專利名稱：具有降低的吸附作用和增加的水解作用的卡氏枯草桿菌蛋白酶變異體的制作方法
技術(shù)領(lǐng)域：
本發(fā)明涉及新的用于各種洗滌組合物的酶變異體和編碼這種酶變異體的DNA序列。
背景技術(shù)：
酶構(gòu)成了最大種類的天然產(chǎn)生的蛋白質(zhì)。每一類酶通常催化(加速反應(yīng)而不被消耗)各種化學(xué)反應(yīng)。稱為蛋白酶的一種酶已知能夠水解(分解化合物成兩個(gè)或更多個(gè)更簡單的化合物，在斷裂的化學(xué)鍵的兩側(cè)接受水分子的H和OH部分)其它蛋白質(zhì)。將天然產(chǎn)生的和蛋白質(zhì)設(shè)計(jì)的蛋白酶作為添加劑加入洗衣洗滌劑制劑中即利用了水解蛋白質(zhì)的這種能力。在衣服上的許多污垢是蛋白質(zhì)的，廣泛特效的蛋白酶可顯著改善這種污垢的除去。
不幸的是，這些蛋白質(zhì)在其天然的，細(xì)菌的環(huán)境中的功效水平通常不能轉(zhuǎn)移至相對(duì)非天然的洗滌環(huán)境。尤其是蛋白酶特性，例如熱穩(wěn)定性、pH穩(wěn)定性、氧化穩(wěn)定性和底物特性對(duì)于酶的天然環(huán)境之外的應(yīng)用未必達(dá)到最佳。
蛋白酶的氨基酸序列決定了蛋白酶的特征。根據(jù)氨基酸序列中改變的位點(diǎn)、性質(zhì)和/或幅度，改變蛋白酶的氨基酸序列可改變酶的特性至各不相同的程度，甚至可失活酶。人們已做了許多嘗試以改變蛋白酶的野生型氨基酸序列，以改善它們的性質(zhì)，從而增加蛋白酶在洗滌環(huán)境中的效率。這些嘗試包括改變氨基酸序列以便在相當(dāng)不同的條件下提高熱穩(wěn)定性和改善氧化穩(wěn)定性。
盡管在現(xiàn)有技術(shù)中描述了各種各樣的嘗試，但人們?nèi)圆粩嘈枰糜谙礈旄鞣N表面的蛋白酶的新有效變異體。發(fā)明目的本發(fā)明的目的在于提供與野生型酶相比較具有改善的水解作用的卡氏(Carlsberg)枯草桿菌蛋白酶酶變異體。
本發(fā)明的另一個(gè)目的在于提供含有這些枯草桿菌蛋白酶變異體的洗滌組合物。
發(fā)明概述本發(fā)明涉及具有野生型卡氏枯草桿菌蛋白酶氨基酸序列的修飾氨基酸序列的卡氏枯草桿菌蛋白酶變異體，野生型氨基酸序列包括第一環(huán)區(qū)域、第二環(huán)區(qū)域、第三環(huán)區(qū)域、第四環(huán)區(qū)域和第五環(huán)區(qū)域；其中，修飾的氨基酸序列在一個(gè)或多個(gè)環(huán)區(qū)域中的特定標(biāo)記的位點(diǎn)上含有與野生型卡氏枯草桿菌蛋白酶不同的氨基酸(即取代)，與野生型卡氏枯草桿菌蛋白酶相比，卡氏枯草桿菌蛋白酶變異體具有對(duì)不溶解底物降低的吸附作用和增加的水解作用。本發(fā)明還涉及編碼這種卡氏枯草桿菌蛋白酶變異體的基因。本發(fā)明還涉及含有這種卡氏枯草桿菌蛋白酶變異體的用于洗滌各種表面的組合物。
發(fā)明詳述I.枯草桿菌蛋白酶變異體本發(fā)明涉及枯草桿菌蛋白酶，尤其是卡氏枯草桿菌蛋白酶，它們通過突變編碼酶的各種核苷酸順序，從而改變酶的氨基酸序列而被修飾。本發(fā)明的修飾枯草桿菌蛋白酶(下文稱為“卡氏枯草桿菌蛋白酶變異體”)與野生型卡氏枯草桿菌蛋白酶相比，具有對(duì)不溶解底物降低的吸附作用和增加的水解作用。本發(fā)明還涉及編碼這種卡氏枯草桿菌蛋白酶變異體的突變基因。
本發(fā)明的枯草桿菌蛋白酶屬于稱為蛋白酶的酶的種類。蛋白酶是斷裂肽鍵的催化劑。一種類型的蛋白酶是絲氨酸蛋白酶。絲氨酸的特征在于在活性位點(diǎn)上存在必需的絲氨酸殘基。
隨著酶濃度的增加溶解底物的酶水解速率增加的現(xiàn)象已經(jīng)作了充分報(bào)導(dǎo)。因而，對(duì)于表面結(jié)合的底物，例如在許多洗滌應(yīng)用中遇到的情況，水解速率隨著表面濃度的增加而增加似乎是可能的，這已由事實(shí)說明。(Brode，P.F.III和D.S.Rauch，LANGMUIR，“枯草芽孢桿菌BPN’。固定化底物上的活性”，卷8，1325-1329頁(1992))。事實(shí)上，當(dāng)酶的表面濃度變化時(shí)，對(duì)于不溶解底物已經(jīng)發(fā)現(xiàn)了表面濃度與速率的直線關(guān)系。(Rubingh，D.N.和M.D.Bauer.“在蛋白質(zhì)溶液界面上的蛋白酶的催化水解”，多聚物溶液，混合和界面(POLYMER SOLUTIONS，BLENDSAND INTERFACES)，by I Noda and d.N.Rubingh，Elsevier編，第464頁(1992))。出乎意料的是，當(dāng)試圖將這種原理用于研究得到較好洗滌性能的變并體蛋白酶時(shí)，我們不能發(fā)現(xiàn)吸附更多的酶得到較好的性能。事實(shí)上，我們驚奇地發(fā)現(xiàn)與之相反的事實(shí)酶與底物的下降的吸附導(dǎo)致增加的底物水解(即，較好的洗滌性能)。
盡管不希望限制于任何理論，但我們相信，當(dāng)將變異體相互比較時(shí)，改善的性能是如下事實(shí)的結(jié)果，即吸附較少的酶也不太緊密地結(jié)合，因而在欲除去不溶解蛋白質(zhì)底物的表面更易移動(dòng)。在可比的酶溶液濃度下，增加的移動(dòng)能力足以勝過通過向表面提供較高濃度酶而給予的任何優(yōu)點(diǎn)。
本發(fā)明描述的突變用于改變(即降低)酶與表面結(jié)合的污垢的吸附。在卡氏枯草桿菌蛋白酶中，某些氨基酸在酶分子上形成外環(huán)。為了討論，這些環(huán)可稱為第一、第二、第三、第四和第五環(huán)區(qū)域。具體地說，位點(diǎn)58-65形成第一環(huán)區(qū)域；位點(diǎn)94-106形成第二環(huán)區(qū)域；位點(diǎn)125-132形成第三環(huán)區(qū)域；位點(diǎn)153-166形成第四環(huán)區(qū)域；位點(diǎn)186-190形成第五環(huán)區(qū)域；和位點(diǎn)199-219形成第六環(huán)區(qū)域(位點(diǎn)編號(hào)類似于野生型枯草桿菌蛋白酶即卡氏枯草桿菌蛋白酶(SEQ ID NO1)的氨基酸序列)。
我們相信，這些環(huán)區(qū)域在酶分子與表面結(jié)合的肽的吸附中起著重要的作用，在一個(gè)或多個(gè)環(huán)區(qū)域中具體的突變對(duì)吸附具有明顯的效果。盡管不希望限制于任何理論，但我們相信出于至少兩個(gè)原因，這些環(huán)區(qū)域?qū)ㄊ峡莶輻U菌蛋白酶分子的吸附是重要的。首先，含有環(huán)區(qū)域的氨基酸能夠密切地與分子暴露的任何表面接觸。其次，卡氏枯草桿菌蛋白酶分子的環(huán)區(qū)域與活性位點(diǎn)的接近和結(jié)合能使得它們?cè)诿概c表面結(jié)合的底物(肽/蛋白質(zhì)污垢)的催化產(chǎn)生的吸附中起作用。
本文使用的術(shù)語“變異體”是指含有不同于野生型氨基酸序列的酶。
本文使用的術(shù)語“突變卡氏枯草桿菌蛋白酶基因”指的是編碼卡氏枯草桿菌蛋白酶變異體的基團(tuán)。
本文使用的術(shù)語“野生型卡氏枯草桿菌蛋白酶”指的是由SEQ IDNO1表示的酶。該卡氏枯草桿菌蛋白酶的氨基酸序列在Smith，E.L.，Delanfe，R.J Evans，W.H.，Landin，M.和Markland，F(xiàn).S.，生物化學(xué)雜志.，卷243，第2184-2191頁(1968)(列為參考文獻(xiàn))中進(jìn)一步描述。
本文使用的術(shù)語“卡氏枯草桿菌蛋白酶野生型氨基酸序列”包括SEQID NO1以及除位點(diǎn)58-65、94-106、125-132、153-166、186-190和199-219之外具有修飾氨基酸序列的SEQ ID NO1。
本文使用的術(shù)語“更親水的氨基酸”指的是參考下表中的親水性值比目標(biāo)氨基酸具有更大親水性值的任何其它氨基酸。如下親水性值表(表1)以增加的親水性值的下降順序列出氨基酸(參見Hopp，T.P.，和Woods，K.R.，“由氨基酸序列判斷蛋白質(zhì)抗原決定簇”，美國國家科學(xué)院院刊，卷78，3824-3828頁，1981，列為參考文獻(xiàn))。
表1氨基酸 親水性值Trp -3.4Phe -2.5Tyr -2.3Leu，Ile-1.8Val -1.5Met -1.3Cys -1.0Ala，His-0.5Thr -0.4Pro，Gly-0.0Gln，Asn 0.2Ser 0.3Arg+，Lys+，Glu-，Asp-3.0表1還說明了帶有電荷(該特征基于約8-9的pH)。正電荷的氨基酸是Arg和Lys，負(fù)電荷氨基酸是Glu和Asp，其余的氨基酸是中性的。在本發(fā)明的優(yōu)選實(shí)施方案中，取代氨基酸是中性的或負(fù)電荷的，更優(yōu)選負(fù)電荷(即Glu或Asp)。
因而，例如說明“用相等或更親水的中性或帶有負(fù)電荷的氨基酸取代的Gln”指的是用Asn(它與Gln的親水性相等)或Ser、Glu或Asp(它們比Gln更親水)取代的Gln。它們每個(gè)均是中性或具有負(fù)電荷，與Gln相比較，它們具有較大的親水性值。同樣，說明“用更親水的中性或帶有負(fù)電荷的氨基酸取代的Pro”指的是用Gln、Asn、Ser、Glu或Asp取代的Pro。
在本發(fā)明的一個(gè)實(shí)施方案中，卡氏枯草桿菌蛋白酶變異體具有卡氏枯草桿菌蛋白酶野生型氨基酸序列的修飾氨基酸序列，其中野生型氨基酸序列含有在第一環(huán)區(qū)域、第二環(huán)區(qū)域、第三環(huán)區(qū)域、第四環(huán)區(qū)域、第五環(huán)區(qū)域或第六環(huán)區(qū)域的一個(gè)或多個(gè)中的一個(gè)或多個(gè)位點(diǎn)上的取代基；從而卡氏枯草桿菌蛋白酶變異體與野生型卡氏枯草桿菌蛋白酶相比較，對(duì)不溶解底物具有降低的吸附作用和增加的水解作用。
在本發(fā)明的優(yōu)選實(shí)施方案中，在一個(gè)或多個(gè)環(huán)區(qū)域中一個(gè)或多個(gè)位點(diǎn)取代氨基酸，參考表1，是中性的或負(fù)電荷的，與在野生型氨基酸序列中目標(biāo)位點(diǎn)的氨基酸相比較，是相等或更親水性的，優(yōu)選是更親水性的。A.第一環(huán)區(qū)域中的取代當(dāng)取代發(fā)生在第一環(huán)區(qū)域中時(shí)，取代發(fā)生在位點(diǎn)58、59、60、61、62、64或65的一個(gè)或多個(gè)上。
當(dāng)取代發(fā)生在位點(diǎn)58時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)59時(shí)，取代氨基酸是Glu。
當(dāng)取代發(fā)生在位點(diǎn)60時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)61時(shí)，取代氨基酸是Asp、Gln、Glu或Ser。
當(dāng)取代發(fā)生在位點(diǎn)62時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)64時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)65時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。B.第二環(huán)區(qū)域的取代當(dāng)取代發(fā)生在第二環(huán)區(qū)域中時(shí)，取代發(fā)生在位點(diǎn)94、95、96、97、98、99、100、101、102、103、104、105或106的一個(gè)或多個(gè)上。
當(dāng)取代發(fā)生在位點(diǎn)94時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Met、Pro、Ser或Thr。
當(dāng)取代發(fā)生在位點(diǎn)95時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Met、Pro、Ser、Thr或Val。
當(dāng)取代發(fā)生在位點(diǎn)96時(shí)，取代氨基酸是Asp、Gln、Glu、Ser。
當(dāng)取代發(fā)生在位點(diǎn)97時(shí)，取代氨基酸是Asp或Glu。
當(dāng)取代發(fā)生在位點(diǎn)98時(shí)，取代氨基酸是Asp或Glu。
當(dāng)取代發(fā)生在位點(diǎn)99時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)100時(shí)，取代氨基酸是Asp或Glu。
當(dāng)取代發(fā)生在位點(diǎn)101時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)102時(shí)，取代氨基酸是Asp或Glu。
當(dāng)取代發(fā)生在位點(diǎn)103時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val。
當(dāng)取代發(fā)生在位點(diǎn)104時(shí)，取代氨基酸是Asp或Glu。
當(dāng)取代發(fā)生在位點(diǎn)105時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)106時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Leu、Met、Pro、Ser、Thr或Val。C.第三環(huán)區(qū)域中的取代當(dāng)取代發(fā)生在第三環(huán)區(qū)域中時(shí)，取代發(fā)生位點(diǎn)125、126、127、128、129、130、131或132的一個(gè)或多個(gè)上。
當(dāng)取代發(fā)生在位點(diǎn)125時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Met、Pro、Ser、Thr或Val。
當(dāng)取代發(fā)生在位點(diǎn)126時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)127時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)128時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、His、Pro、Ser或Thr。
當(dāng)取代發(fā)生在位點(diǎn)129時(shí)，取代氨基酸是Asp或Glu。
當(dāng)取代發(fā)生在位點(diǎn)130時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)131時(shí)，取代氨基酸是Asp或Glu。
當(dāng)取代發(fā)生在位點(diǎn)132時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。D.第四環(huán)區(qū)域中的取代當(dāng)取代發(fā)生在第四環(huán)區(qū)域中時(shí)，取代發(fā)生在位點(diǎn)153、154、155、156、157、158、159、160、161、162、163、164、165或166的一個(gè)或多個(gè)上。
當(dāng)取代發(fā)生在位點(diǎn)153時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)154時(shí)，取代氨基酸是Asp、Gln、Glu或Ser。
當(dāng)取代發(fā)生在位點(diǎn)155時(shí)，取代氨基酸是Asp或Glu。
當(dāng)取代發(fā)生在位點(diǎn)156時(shí)，取代氨基酸是Ash、Asp、Gln、Glu、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)157時(shí)，取代氨基酸是Asp、Gln、Glu、Ser。
當(dāng)取代發(fā)生在位點(diǎn)158時(shí)，取代氨基酸是Asp或Glu。
當(dāng)取代發(fā)生在位點(diǎn)159時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)160時(shí)，取代氨基酸是Asp或Glu。
當(dāng)取代發(fā)生在位點(diǎn)161時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)162時(shí)，取代氨基酸是Asp、Gln、Glu或Ser。
當(dāng)取代發(fā)生在位點(diǎn)163時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)164時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Met、Pro、Ser、Thr或Val。
當(dāng)取代發(fā)生在位點(diǎn)165時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。和當(dāng)取代發(fā)生在位點(diǎn)166時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val。E.第五環(huán)區(qū)域中的取代當(dāng)取代發(fā)生在第五環(huán)區(qū)域中時(shí)，取代發(fā)生在位點(diǎn)186、187、188、189或190一個(gè)或多個(gè)上。
當(dāng)取代發(fā)生在位點(diǎn)186時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、His、Pro、Ser或Thr。
當(dāng)取代發(fā)生在位點(diǎn)187時(shí)，取代氨基酸是Asp或Glu。
當(dāng)取代發(fā)生在位點(diǎn)188時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr、Tyr或Val。
當(dāng)取代發(fā)生在位點(diǎn)189時(shí)，取代氨基酸是Asp或Glu。
當(dāng)取代發(fā)生在位點(diǎn)190時(shí)，取代氨基酸是Asn或Glu。F.第六環(huán)區(qū)域中的取代當(dāng)取代發(fā)生在第六環(huán)區(qū)域中時(shí)，取代發(fā)生在位點(diǎn)199、200、201、202、203、204、205、206、207、208、209、210、211、212、213、214、215、216、217、218或219的一個(gè)或多個(gè)上。
當(dāng)取代發(fā)生在位點(diǎn)199時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、His、Pro、Ser或Thr。
當(dāng)取代發(fā)生在位點(diǎn)200時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly或Ser。
當(dāng)取代發(fā)生在位點(diǎn)201時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)202時(shí)，取代氨基酸是Asp、Asp、Gln、Glu、Gly、His、Pro、Ser或Thr。
當(dāng)取代發(fā)生在位點(diǎn)203時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)204時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Met、Pro、Ser或Thr。
當(dāng)取代發(fā)生在位點(diǎn)205時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val。
當(dāng)取代發(fā)生在位點(diǎn)206時(shí)，取代氨基酸是Asp或Glu。
當(dāng)取代發(fā)生在位點(diǎn)207時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)208時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val。
當(dāng)取代發(fā)生在位點(diǎn)209時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly或Ser。
當(dāng)取代發(fā)生在位點(diǎn)210時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)211時(shí)，取代氨基酸是Asp、Gln、Glu或Ser。
當(dāng)取代發(fā)生在位點(diǎn)212時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)213時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val。
當(dāng)取代發(fā)生在位點(diǎn)214時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、His、Pro、Ser或Thr。
當(dāng)取代發(fā)生在位點(diǎn)215時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)216時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Met、Pro、Ser、Thr或Val。
當(dāng)取代發(fā)生在位點(diǎn)217時(shí)，取代氨基酸是Asp、Gln、Glu或Ser。
當(dāng)取代發(fā)生在位點(diǎn)218時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
當(dāng)取代發(fā)生在位點(diǎn)219時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。
G.酶變異體的制備實(shí)施例1突變體卡氏枯草桿菌蛋白酶基因構(gòu)建含有野生型卡氏枯草桿菌蛋白酶基因的噬菌粒(“TP”)。將質(zhì)粒pUC119的2.8Kbp PvuII限制酶片段(Vieira，J.和Messing，J.，“單鏈的質(zhì)粒DNA的制備”，153酶學(xué)方法3-11(1989))克隆至質(zhì)粒pUB110(Bacillus Genetic Stock Center，Columbus，OH 1E9)的PvuII位點(diǎn)。pUC119-pUB110雜交質(zhì)粒被稱為pJMA601。在pJMA601的BamH1限制位克隆入具有TP的聚合酶鏈反應(yīng)擴(kuò)增的產(chǎn)CP的卡氏枯草桿菌蛋白酶基因。噬菌粒TP被轉(zhuǎn)化入大腸桿菌Ung-菌株CJ236，用VCSM13輔助噬菌體(Kunkel，T.A.，J.D.Roberts和R.A.Zakour，“無需表型選擇的快速和有效的位點(diǎn)專一誘變”，酶學(xué)方法，卷154，367-382頁(1987)；由Yuckenberg，P.D.，F(xiàn).Witney，J.Geisselsoder和J.McClary改進(jìn)，“使用含尿嘧啶的DNA和噬菌粒載體的體外定點(diǎn)誘變”，定向突變，操作指南，M.J.McPherson編，第27-48頁(1991)；均列為本文參考文獻(xiàn))制備單鏈的含尿嘧啶DNA模板。使用Zoller和Smith方法的單引物定點(diǎn)誘變修飾方法(Zoller，M.J.和M.Smith，“使用M13衍生的載體的寡核苷酸定向誘變一種在DNA任何片段中制備點(diǎn)突變的有效和常用方法”，核酸研究，卷10，6487-6500頁(1982)，列為本文參考文獻(xiàn))制備所有突變體(基本上如上述Yuckenberg等人，1991中說明)。寡核苷酸使用Applied BiosystemInc.380B DNA合成儀制備。誘變反應(yīng)產(chǎn)物轉(zhuǎn)化入大腸桿菌菌株MM 294(American Type Culture Collection E.Coli.33625)中。所有突變體由DNA測序證實(shí)，分離的DNA轉(zhuǎn)化入枯草芽孢桿菌表達(dá)菌株BG2036(Yang，M.Y.，E.Ferrari和D.J.Henner，(1984)，“枯草芽孢桿菌的中性蛋白酶基因的克隆和克隆的基因用于產(chǎn)生體外得到的缺失突變”，細(xì)胞學(xué)雜志，卷160，15-21頁)。對(duì)于某些環(huán)突變體，用在相應(yīng)環(huán)中移碼終止的密碼子突變的修飾TP制備尿嘧啶模板。寡核苷酸用來恢復(fù)合適的閱讀框和編碼在位點(diǎn)58、59、60、61、62、64、65、94、95、96、97、98、99、100、101、102、103、104、105、106、125、126、127、128、129、130、131、132、153、154、155、156、157、158、159、160、161、162、163、164、165、166、186、187、188、189、190、199、200、201、202、203、204、205、206、207、208、209、210、211、212、213、214、215、216、217、218和/或219的隨機(jī)取代(在這些密碼子中對(duì)于所有三種堿基所有四種寡核苷酸的等摩爾和/或可變混合物)。校正移碼終止和制備功能酶的突變由它們消化酪蛋白的能力說明。隨機(jī)取代用DNA測序確定。
實(shí)施例2發(fā)酵將含有感興趣的枯草芽孢桿菌突變體的枯草芽孢桿菌細(xì)胞(BG2036)在1升LB-葡萄糖肉湯培養(yǎng)基中的mid-log相中生長，在總體積10升的Biostat ED發(fā)酵器(B.Braun Biotech，Inc.，Allentown，Pennsylvania)中培養(yǎng)。發(fā)酵介質(zhì)含有酵母提取物、淀粉、消泡劑、緩沖劑和痕量礦物質(zhì)(參見發(fā)酵操作指南B.McNeil和L.M.Harvey編，1990)。在發(fā)酵試驗(yàn)過程中，肉湯保持恒定pH7.0。加入氯霉素以進(jìn)行誘變質(zhì)粒的抗菌選擇。細(xì)胞在37℃生長過夜至A600約60，并收獲。
實(shí)施例3提純將發(fā)酵肉湯進(jìn)行如下步驟以得到純的酶。通過離心除去肉湯中的枯草芽孢桿菌細(xì)胞，通過用100K截?cái)嗄こゼ?xì)的粒子而澄清。然后用10K截?cái)嗄ど蠞饪s，進(jìn)行流體透析以降低離子強(qiáng)度，用0.025M MES緩沖劑(2-(N-嗎啉代)乙磺酸)調(diào)節(jié)pH至5.5。通過將酶加入陽離子交換層析柱或親合吸附層析柱，由柱中用氯化鈉或丙二醇梯度洗脫而進(jìn)一步純化(參見，Scopes，R.K.，蛋白質(zhì)純化理論和操作，Springer-Verlag，NewYork(1984)，列為本文參考文獻(xiàn))。
使用pNA試驗(yàn)(DelMar，E.G.，C.Largman，J.W.Brodrick和M.C.Geokas，生物化學(xué)年鑒，卷99，316-320頁(1979)，列為本文參考文獻(xiàn))測定在梯度洗脫過程中收集的級(jí)分的酶濃度。該試驗(yàn)測量當(dāng)酶水解溶解的合成底物，琥珀酰-丙氨酰-丙氨酰-脯氨酰-苯基丙氨酰-對(duì)-硝基苯胺(sAAPF-pNA)時(shí)釋放對(duì)-硝基苯胺的速率。用分光光度計(jì)在410nm測量水解反應(yīng)產(chǎn)生黃色的速率，與活性酶濃度成比例。此外，在280nm的吸附測量用來測定總蛋白質(zhì)濃度?；钚悦?總蛋白質(zhì)比率得到酶的純度，并用于鑒定用作存貯溶液的收集的級(jí)分。
為避免在貯存過程中酶的自解，在由層析柱得到的收集級(jí)分中加入相等重量的丙二醇。在提純過程完成后，存貯酶溶液用SDS-PAGE(十二烷基磺酸鈉聚丙烯酰胺凝膠電泳)校驗(yàn)，用胰蛋白酶抑制劑型II-T由Sigma Chemical Company(St.Louis，Missouri)購買的火雞卵白經(jīng)活性位滴定法測定絕對(duì)酶濃度。測量的轉(zhuǎn)化因子將說明在酶分子中不同位點(diǎn)發(fā)生的改變使得酶變異體與野生型相比較對(duì)溶解底物pNA具有增加的活性。
在使用的制劑中，酶存貯溶液通過Sephadex-G25(Pharmacia，Piscataway，New Jersey)大小排除柱以除去丙二醇和交換緩沖劑。用含有0.01M氯化鈣的0.1M Tris緩沖劑(三(羥基甲基-氨基)甲烷)代替在酶存貯溶液中的MES緩沖劑，用鹽酸調(diào)節(jié)pH至8.6。在恒溫25℃下在Tris緩沖劑中在pH8.6下進(jìn)行所有試驗(yàn)。H酶變異體的特性實(shí)施例4模型表面制備由CPG Inc.(Fairfield，New Jersey)購買的氨基丙基控制孔玻璃(CPG)用作共價(jià)連接由Bachem，Inc.(Torence，California)購買的sAAPD-pNA底物的載體。反應(yīng)在二甲基亞砜中進(jìn)行，用(1-乙基-3-[3-(二甲基氨基)丙基]碳化二亞胺鹽酸鹽)(EDC)作偶合試驗(yàn)。在反應(yīng)完成后(用pNA試驗(yàn)監(jiān)測)，除去過量的溶劑，用二甲基亞砜(DMSO)和雙蒸餾水漂洗CPGsAAPF-pNA。然后在烘箱中氮?dú)獯祾咴诩s70℃下干燥。反應(yīng)方案和固定的底物的制備方法如Brode，P.F.III，和D.S.Rauch，“枯草桿菌蛋白酶BPN’在固定底物上的活性”，LANGMUIR，卷8，1325-1329頁(1992)(列為本文參考文獻(xiàn))中描述進(jìn)行。
CPG表面含有62000±7000pNA分子/μm2。從CPG Inc.報(bào)導(dǎo)的CPG的50.0m2/g的值來看，表面積保持未變。這說明用于在CPG中加入sAAPf-pNA的方法未破壞孔結(jié)構(gòu)(平均直徑486_)。
實(shí)施例5表面水解試驗(yàn)使用CPGsAAPf-pNA，酶變異體的吸附和CPG-結(jié)合的肽的水解可在同一試驗(yàn)中測量。在含有Tris緩沖劑和已脫氣的CPGsAAPF-pNA的燒瓶中加入少量的酶變異體存貯溶液。燒瓶在活塞作用的振蕩器中振蕩90分鐘，在此期間，以不同的時(shí)間間隔停止振蕩器(例如，在吸附水解作用的最初階段，例如最初20分鐘為每2分鐘-在實(shí)驗(yàn)結(jié)束時(shí)為每10分鐘)。使CPGsAAPF-pNA靜置，取樣溶液。如上述Brode等人，1991中所述進(jìn)行吸附和水解的實(shí)驗(yàn)過程和計(jì)算。
測定所有的酶的自溶穩(wěn)定性，在實(shí)驗(yàn)的時(shí)間內(nèi)，應(yīng)未顯示明顯的自溶損失。因此，酶吸附可通過測量溶液消耗測定。最初酶變異體濃度與在每個(gè)間隔的時(shí)間點(diǎn)測量的濃度之間的差值得到吸附的酶變異體的量。由表面水解的pNA的量由在410nm時(shí)試樣的吸附讀數(shù)測定。水解的pNA的總量由取樣的量加上殘余在燒瓶中的量計(jì)算。該值通過減去當(dāng)不存在酶時(shí)在pH8.6下由Tris緩沖劑水解的pNA的量而校準(zhǔn)。根據(jù)酶的效率，基準(zhǔn)水解范圍為總水解的7-29％。
實(shí)施例6溶解底物的動(dòng)力學(xué)分析溶解底物sAAPF-pNA的水解速率通過用DU-70分光光度計(jì)在410nm下測量作為時(shí)間函數(shù)的吸附增量而監(jiān)測。酶濃度保持恒定，并制備成6-10納摩爾的范圍，而對(duì)于每次動(dòng)力學(xué)測定，底物濃度在90-700μM sAAPF-pNA之間變化。在900秒的時(shí)間內(nèi)，每秒收集吸附數(shù)據(jù)點(diǎn)，數(shù)據(jù)傳遞至LOTUSTM記錄紙(Lotus Development Corporation，Cambridge，Massachusetts)。用標(biāo)準(zhǔn)Linewweaver Burk分析法進(jìn)行動(dòng)力學(xué)參數(shù)的分析，其中在試驗(yàn)最初(通常為最初幾分鐘)的數(shù)據(jù)符合于線性回歸曲線以得到v0。以標(biāo)準(zhǔn)倒數(shù)方式標(biāo)繪的v0和s0數(shù)據(jù)得到KM和Kcat。I.卡氏枯草桿菌蛋白酶變異體的實(shí)例降低表面結(jié)合的底物的吸附作用和增加的水解作用的本發(fā)明卡氏枯草桿菌蛋白酶變異體在如下表3-36中說明。在描述的具體突變中，首先給出野生型中出現(xiàn)的原始氨基酸，其次是位點(diǎn)編號(hào)，第三是取代氨基酸。
表2環(huán)1-單突變變異體Thr58AsnThr58AspThr58GlnThr58GluThr58GlyThr58ProThr58SerAsp59GluGly60AsnGly60AspGly60GlnGly60GluGly60ProGly60SerAsn61AspAsn61GlnAsn61GluAsn61SerGly62AsnGly62AspGly62GlnGly62GluGly62ProGly62SerGly64AsnGly64AspGly64GlnGly64GluGly64ProGly64SerThr65AsnThr65AspThr65GlnThr65GluThr65GlyThr65ProThr65Ser表3環(huán)1-雙突變變異體Thr58Gln + Asn61GluAsn61Asp + Thr65Pro
Thr58Gln + Asp59GluThr58Gln + Gly62SerGly62Asn + Thr65ProThr58Gln + Gly60AspThr58Pro + Gly60GluAsn61Asp + Thr65SerAsp59Glu + Gly60GlnAsp59Glu + Gly60ProGly62Asp + Gly64GlnAsn61Ser + Thr65ProThr58Asp + Gly60GlnGly60Ser + Gly62GlnAsp59Glu + Gly62ProGly62Asp + Thr65GlyAsn61Gln + Thr65ProAsn61Gln + Gly64AsnGly60Gln + Gly62GlnThr58Gly + Asn61AspThr58Gln + Gly62GlnGly60Asn + Gly62GlnGly60Asn + Gly62AspThr58Glu + Gly62ProGly64Asp + Thr65ProAsp59Glu + Thr65SerGly60Gln + Thr65GlnAsp59Glu + Gly62SerAsn61Glu + Gly64GlnAsp59Glu + Gly60SerGly60Ser + Gly62SerThr58Gly + Gly64GluThr58Asn + Asn61SerAsp59Glu + Thr65AsnGly60Asn + Asn61SerGly60Glu + Gly62GlnThr58Pro + Gly62AspThr58Ser + Asp59GluThr58Asp + Gly62ProAsn61Asp + Gly62ProThr58Ser + Gly62SerGly60Asn + Gly62SerGly64Pro + Thr65AsnGly64Asp + Thr65GlnGly64Glu + Thr65ProGly64Asn + Thr65ProAsn61Ser + Gly64Asp表4
環(huán)1-三突變變異體Thr58Pro + Gly64Ser + Thr65GlyAsn61Glu + Gly62Gln + Thr65GlnThr58Gln + Gly62Ser + Gly64ProAsp59Glu + Gly62Asn + Thr65ProThr58Pro + Gly60Glu + Gly64AsnThr58Asn + Gly64Ser + Thr65GlyAsp59Glu + Asn61Gln + Gly64SerThr58Asn + Asn61Glu + Thr65ProGly60Pro + Asn61Asp + Thr65ProThr58Glu + Gly62Gln + Gly64AsnThr58Asn + Gly62Glu + Gly64ProThr58Ser + Asp59Glu + Thr65AsnThr58Glu + Gly62Asn + Gly64GlnThr58Pro + Asp59Glu + Gly62SerGly60Asn + Asn61Asp + Gly62AsnGly62Asp + Gly64Ser + Thr65ProAsp59Glu + Gly60Asn + Thr65AsnThr58Gly + Gly60Pro + Thr65GlyThr58Gln + Gly60Glu + Asn61GlnGly60Pro + Asn61Gln + Gly64AspThr58Asn + Gly62Asn + Thr65ProThr58Pro + Asp59Glu + Gly64AsnThr58Pro + Ash61Ser + Gly62AspThr58Asp + Gly50Pro + Gly62AsnThr58Ser + Gly60Pro + Asn61AspGly60Gln + Gly62Gln + Gly64ProAsp59Glu + Asn61Gln + Thr65AsnGly60Asp + Asn61Gln + Thr65ProAsn61Glu + Gly62Glu + Thr65ProThr58Asp + Asp59Glu + Thr65GlnThr58Glu + Asp59Glu + Thr65SerThr58Glu + Asp59Glu + Thr65Gly表5環(huán)1-四突變變異體Asp59Glu + Gly60Pro + Gly62Asn + Gly64GlnThr58Pro + Gly60Pro + Gly62Glu + Thr65GlnThr58Ser + Gly60Pro + Gly64Gln + Thr65ProThr58Pro + Asp59Glu + Asn61Ser + Gly62AsnThr58Glu + Gly60Asn + Gly64Ser + Thr65ProThr58Glu + Gly60Ser + Asn61Gln + Thr65GlnThr58Pro + Asp59Glu + Gly60Asn + Asn61GlnThr58Gly + Gly60Pro + Gly62Glu + Thr65SerGly60Asp + Gly62Ser + Gly64Gln + Thr65GlyThr58Asn + Gly60Pro + Gly62Ser + Gly64Asp
Thr58Pro + Asp59Glu + Gly60Asn + Gly62AsnAsn61Asp + Gly62Gln + Gly64Pro + Thr65AsnThr58Asp + Gly60Pro + Gly62Asn + Gly64SerThr58Asp + Gly60Gln + Gly62Pro + Thr65AsnThr58Asn + Asn61Asp + Gly62Glu + Thr65GlyThr58Ser + Gly60Ser + Asn61Asp + Gly62GluThr58Glu + Asp59Glu + Asn61Gln + Thr65SerThr58Asp + Asp59Glu + Gly62Pro + Thr65GlyGly60Asp + Asn61Asp + Gly62Ser + Thr65GlyGly60Glu + Asn61Glu + Gly62Gln + Gly64ProThr58Pro + Gly60Asp + Asn61Asp + Gly64AsnAsp59Glu + Gly60Asp + Gly64Gln + Thr65SerAsp59Glu + Gly60Asp + Gly64Gln + Thr65ProThr58Pro + Asp59Glu + Gly60Asp + Gly64AsnThr58Ser + Asp59Glu + Gly60Asp + Gly62AsnAsp59Glu + Gly60Glu + Asn61Glu + Gly62SerThr58Glu + Asp59Glu + Gly60Asp + Thr65ProAsp59Glu + Gly60Asp + Gly62Glu + Thr65ProThr58Ser + Asp59Glu + Asn61Asp + Gly62Asp表6環(huán)2-單突變變異體Val 94AlaVal 94AsnVal 94AspVal 94CysVal 94GlnVal 94GluVal 94GlyVal 94HisVal 94MetVal 94ProVal 94SerVal 94ThrLeu 95AlaLeu 95AsnLeu 95AspLeu 95CysLeu 95GlnLeu 95GluLeu 95GlyLeu 95HisLeu 95IleLeu 95MetLeu 95ProLeu 95Ser
Leu 95ThrLeu 95ValAsn 96AspAsn 96GlnAsn 96GluAsn 96SerSer 97AspSer 97GluSer 98AspSer 98GluGly 99AsnGly 99AspGly 99GlnGly 99GluGly 99ProGly 99SerSer100AspSer100GluGly101AsnGly101AspGly101GlnGly101GluGly101ProGly101SerSer102AspSer102GluTyr103AlaTyr103AsnTyr103AspTyr103CysTyr103GlnTyr103GluTyr103GlyTyr103HisTyr103IleTyr103LeuTyr103MetTyr103ProTyr103SerTyr103ThrTyr103ValSer104AspSer104GluGly105AsnGly105AspGly105GlnGly105Glu
Gly105ProGly105SerIle106AlaIle106AsnIle106AspIle106CysIle106GlnIle106GluIle106GlyIle106HisIle106LeuIle106MetIle106ProIle106SerIle106ThrIle106Val表7環(huán)2-雙突變變異體Val 94Gln + Ser100GluAsn 96Ser + Gly 99GlnGly 99Glu + Ile106MetSer 97Asp + Gly105GlnSer102Glu + Tyr103AsnSer100Glu + Ile106GlyAsn 96Asp + Ile106SerGly 99Ser + Ser100GluGly101Gln + Gly105SerAsn 96Asp + Gly 99ProAsn 96Gln + Ser104AspLeu 95Glu + Tyr103ProAsn 96Ser + Gly101AspLeu 95Val + Ser 97GluLeu 95Ala + Gly 99ProVal 94Thr + Gly 99GluLeu 95Gln + Ile106AlaLeu 95Pro + Ile106AlaLeu 95Val + Ser100GluGly101Gln + Tyr103GlyAsn 96Asp + Ile106ProLeu 95Ser + Ser100AspGly 99Pro + Ser100AspVal 94Thr + Leu 95AspSer100Glu + Ile106GlnSer100Asp + Gly101ProLeu 95Cys + Gly105Asp
Ser 98Glu + Gly101SerGly101Asn + Tyr103GlnVal 94Met + Ser102GluGly 99Pro + Gly105AspVal 94Asn + Ser102AspVal 94Cys + Ile106ThrVal 94Gly + Ser104AspVal 94Pro + Gly101ProVal 94Ser + Gly105GluVal 94Met + Ser 98AspVal 94Gln + Ser104AspAsn 96Ser + Ser 98AspSer104Glu + Ile106GlnVal 94Asn + Tyr103ThrTyr103Ala + Ser104AspGly 99Pro + Ser104GluLeu 95Cys + Tyr103LeuVal 94Gly + Gly 99SerGly101Gln + Tyr103SerSer 97Asp + Gly105ProSer 98Asp + Gly105AsnGly 99Gln + Ser100GluGly 99Pro + Ser100GluTyr103Ile + Gly105SerSer100Glu + Gly105ProAsn 96Glu + Tyr103GlyVal 94Met + Gly105AsnVal 94His + Leu 95GluSer100Asp + Tyr103MetLeu 95Met + Gly101GlnGly 99Pro + Gly101AspAsn 96Asp + Gly101AsnVal 94Ala + Gly 99Ser表8環(huán)2-三突變變異體Val 94Gln + Leu 95Thr + Ser100GluSer100Glu + Tyr103Met + Ile106ThrSer 97Asp + Gly101Gln + Ile106AsnLeu 95Gln + Gly105Ser + Ile106GlnLeu 95His + Gly 99Gln + Ser100GluVal 94His + Leu 95Met + Asn 96GlnVal 94Asn + Ser 97Glu + Tyr103ProSer 97Asp + Gly101Ser + Tyr103MetSer 97Glu + Tyr103Met + Ile106GlnVal 94Thr + Gly 99Pro + Gly105Asp
Leu 95Gly + Ser 98Glu + Tyr103CysLeu 95Pro + Gly101Asp + Tyr103GlyLeu 95Val + Gly 99Pro + Ser104AspLeu 95Ile + Gly105Ser + Ile106MetSer 97Glu + Gly101Asn + Ile106ThrAsn 96Ser + Gly101Asp + Tyr103AsnVal 94Pro + Ser 98Glu + Ile106AsnGly 99Pro + Tyr103Glu + Ile106SerAsn 96Asp + Gly 99Asn + Tyr103ProLeu 95Gln + Ser 97Asp + Ile106ValVal 94Pro + Ser 98Glu + Tyr103ValLeu 95Asn + Gly101Pro + Ile106HisVal 94Ala + Leu 95His + Ser100GluVal 94Asn + Gly101Ser + Ser104AspVal 94Cys + Gly105Ser + Ile106ThrLeu 95Gly + Gly101Asp + Ile106CysSer102Glu + Tyr103Ile + Ile106GlyVal 94Pro + Leu 95Asp + Ile106ProVal 94Pro + Leu 95Ile + Ile106CysSer100Asp + Gly101Asp + Ile106AsnLeu 95Glu + Asn 96Glu + Tyr103ProGly101Glu + Ser102Asp + Tyr103LeuVal 94Pro + Gly101Asp + Ser102GluAsn 96Asp + Ser 97Asp + Gly 99ProSer 97Asp + Ser 98Asp + Gly 99GluAsn 96Glu + Ser 97Asp + Gly 99AspGly 99Glu + Ser100Asp + Gly101AspGly 99Glu + Ser100Glu + Gly101GluLeu 95Gln + Ser102Asp + Gly105AspAsn 96Ser + Ser102Glu + Ser104GluSer102Asp + Ser104Glu + Gly105ProSer102Asp + Tyr103Ser + Ser104GluVal 94Cys + Ser 97Glu + Gly 99AspAsn 96Asp + Ser 98Glu + Ile106ThrAsn 96Asp + Ser 98Asp + Ile106CysAsn 96Glu + Ser 98Glu + Gly105AsnSer 98Glu + Ser100Glu + Ile106SerSer 98Asp + Ser100Asp + Gly101AsnGly101Asp + Ser102Asp + Ser104GluLeu 95Glu + Ser 98Asp + Gly101AspVal 94Ser + Ser100Glu + Ser102AspLeu 95Val + Ser 97Glu + Ser100GluAsn 96Gln + Ser 97Asp + Ser100AspAsn 96Ser + Ser 97Asp + Ser100AspVal 94Asp + Gly 99Asn + Ser102GluVal 94Asp + Ser102Glu + Ile106HisLeu 95Asp + Gly 99Glu + Ser102Asp
Val 94Thr + Gly101Asp + Ser104GluGly 99Pro + Gly101Asp + Ser104AspSer 98Asp + Gly101Asp + Tyr103Pro表9環(huán)2-四突變變異體Val 94Met + Leu 95Asn + Ser 98Asp + Gly105SerAsn 96Ser + Gly101Glu + Tyr103Ile + Gly105AsnLeu 95Val + Ser 97Asp + Gly 99Asn + Tyr103IleLeu 95Ile + Asn 96Gln + Gly 99Asn + Tyr103AspVal 94His + Ser100Glu + Tyr103Ile + Ile106MetSer 97Glu + Tyr103Ile + Gly105Gln + Ile106AlaVal 94Thr + Leu 95Ile + Ser 97Asp + Gly99SerVal 94Thr + Leu 95Cys + Gly 99Asp + Ile106ValLeu 95Cys + Ser 98Glu + Gly 99Asn + Ile106MetAsn 96Gln + Ser 98Asp + Gly105Asn + Ile106GlyLeu 95Thr + Ser100Glu + Tyr103Val + Ile106ProVal 94Thr + Leu 95Val + Asn 96Glu + Tyr103AsnVal 94His + Leu 95Asn + Gly 99Gln + Gly105GlnVal 94Ala + Leu 95Thr + Ser 98Asp + Tyr103IleLeu 95Val + Gly101Gln + Tyr103Cys + Ile106GluGly 99Pro + Ser100Glu + Gly101Ser + Tyr103ThrLeu 95Ala + Gly101Asp + Tyr103Ser + Gly105AsnGly 99Asn + Tyr103Asp + Gly105Asn + Ile106ThrAsn 96Asp + Ser 97Asp + Gly101Ser + Tyr103ThrVal 94Ala + Asn 96Glu + Ser 97Asp + Gly105AsnVal 94Cys + Leu 95Gln + Asn 96Glu + Ser 97GluVal 94Gly + Asn 96Gln + Ser104Glu + Gly105GluSer 97Asp + Ser 98Glu + Gly101Asn + Tyr103ValSer 97Glu + Ser 98Asp + Gly 99Gln + Gly101GlnLeu 95Gly + Asn 96Ser + Ser 97Glu + Ser 98GluAsn 96Glu + Ser 97Glu + Ser 98Asp + Gly101ProAsn 96Asp + Ser 97Asp + Ser 98Asp + Tyr103LeuAsn 96Asp + Ser 97Asp + Ser 98Glu + Tyr103GlyLeu 95Glu + Asn 96Glu + Ser100Asp + Ile106AlaSer 97Asp + Gly 99Glu + Tyr103Cys + Gly105AsnVal 94Gly + Asn 96Ser + Ser 97Glu + Gly 99AspAsn 96Ser + Tyr103Leu + Ser104Asp + Ile106GluLeu 95Ala + Asn 96Glu + Gly 99Ser + Ser100AspAsn 96Asp + Ser 97Glu + Ser100Glu + Ile106ValVal 94Cys + Ser 97Asp + Ser 98Asp + Ser100AspSer 97Asp + Ser 98Asp + Ser100Glu + Tyr103ThrAsn 96Glu + Gly 99Glu + Gly101Asp + Tyr103AlaVal 94Asp + Leu 95Ala + Gly101Glu + Ser102GluSer 98Glu + Gly 99Pro + Ser100Asp + Gly101GluVal 94Glu + Asn 96Gln + Gly101Glu + Ile106Thr
Asn 96Ser + Ser102Asp + Tyr103Asn + Ile106AspLeu 95Cys + Ser 98Asp + Gly 99Asp + Gly101AspSer100Glu + Gly101Pro + Ser102Glu + Ile106AlaVal 94Cys + Ser100Glu + Gly101Pro + Ser102GluVal 94Gln + Ser100Asp + Ser102Glu + Tyr103IleSer 97Asp + Ser100Glu + Gly101Glu + Tyr103AsnVal 94Glu + Asn 96Glu + Gly 99Glu + Gly105ProAsn 96Gln + Ser100Glu + Gly101Asp + Ile106GluVal 94Pro + Leu 95Ala + Ser 97Asp + Ser100AspLeu 95Glu + Gly101Asn + Ser102Glu + Tyr103IleVal 94Asn + Asn 96Glu + Gly101Asp + Gly105AspVal 94Asp + Gly 99Asn + Ser100Asp + Ser102AspVal 94Asp + Leu 95Val + Ser102Asp + Tyr103AlaLeu 95Glu + Gly 99Pro + Ser102Asp + Ser104GluLeu 95Ser + Asn 96Gln + Gly101Glu + Ser104AspVal 94Gln + Asn 96Gln + Gly101Glu + Ser104GluGly101Asp + Tyr103Val + Ser104Glu + Ile106ValAsn 96Asp + Ser102Asp + Tyr103Gly + Gly105AspSer100Glu + Ser102Asp + Ser104Asp + Gly105SerVal 94Asp + Ser 97Asp + Ser100Asp + Gly105Gln表10環(huán)3-單突變變異體Leu125AlaLeu125AsnLeu125AspLeu125CysLeu125GlnLeu125GluLeu125GlyLeu125HisLeu125IleLeu125MetLeu125ProLeu125SerLeu125ThrLeu125ValGly126AsnGly126AspGly126GlnGly126GluGly126ProGly126SerGly127AsnGly127AspGly127Gln
Gly127GluGly127ProGly127SerAla128AsnAla128AspAla128GlnAla128GluAla128GlyAla128HisAla128ProAla128SerAla128ThrSer129AspSer129GluGly130AsnGly130AspGly130GlnGly130GluGly130ProGly130SerSer131AspSer131GluThr132AsnThr132AspThr132GlnThr132GluThr132GlyThr132ProThr132Ser表11環(huán)3-雙突變變異體Leu125Gln + Ser129GluGly130Gln + Thr132AsnGly126Asp + Gly127ProLeu125Ile + Ser131GluGly126Gln + Gly130SerGly126Pro + Thr132ProGly126Asp + Thr132SerAla128Gly + Thr132SerLeu125Ala + Gly126GluLeu125Gln + Ser129AspGly126Pro + Ser131AspGly127Ser + Ser129GluGly130Asn + Ser131GluLeu125Glu + Ala128Gln
Gly126Ser + Thr132SerLeu125Ala + Ser129AspGly127Pro + Thr132AspGly126Pro + Gly130SerLeu125Ala + Ala128AspGly127Gln + Ser131AspLeu125Ala + Thr132SerGly127Pro + Thr132GlyGly126Glu + Thr132AsnLeu125Gly + Ser129GluAla128Thr + Gly130AspAla128His + Thr132AspLeu125Gly + Gly126AsnGly126Ser + Ser129GluLeu125Asn + Thr132AsnGly126Glu + Ala128GlySer131Asp + Thr132GlnSer129Glu + Gly130ProGly127Asn + Ser129AspLeu125Glu + Thr132AsnGly126Gln + Gly127GluAla128Thr + Ser131AspLeu125Pro + Ser131GluGly126Pro + Thr132GlyGly126Gln + Ser129AspLeu125Gly + Gly126SerGly127Gln + Ser129AspGly126Gln + Gly130GluGly127Asp + Gly130AsnLeu125Glu + Gly126SerLeu125Ile + Thr132AspGly126Asn + Gly127AspLeu125Cys + Gly126GlnLeu125Glu + Ala128AsnGly126Gln + Ala128AsnGly126Asp + Gly130AsnLeu125Ser + Gly130SerGly126Asn + Ser129AspLeu125Ser + Ser131AspSer129Asp + Thr132AsnGly127Glu + Ala128ThrLeu125Thr + Gly130ProAla128Pro + Thr132AspGly127Ser + Ser131GluGly126Asn + Thr132GluGly130Asp + Thr132Asn
表12環(huán)3-三突變變異體Leu125Asn + Gly126Pro + Ser129GluGly126Asn + Ser129Asp + Gly130ProLeu125Ala + Gly126Ser + Ser131GluGly127Gln + Ala128Thr + Ser129GluLeu125Thr + Ser129Asp + Gly130ProGly126Pro + Ala128Thr + Thr132AspGly126Asn + Gly130Asn + Thr132GlnLeu125Thr + Gly130Gln + Thr132GlnAla128Gly + Ser129Asp + Thr132AsnLeu125Met + Ala128Gln + Ser129AspLsu125Gly + Ala128Asp + Gly130ProLeu125Ile + Gly127Ser + Thr132AspLeu125Val + Gly127Glu + Gly130SerGly127Ser + Gly130Asn + Thr132AsnLeu125Thr + Gly126Ser + Ala128GluLeu125Gln + Gly126Pro + Ser131GluLeu125Glu + Gly130Asn + Thr132AsnGly126Asn + Gly127Pro + Ser129AspGly126Pro + Gly127Pro + Ala128HisGly127Asn + Ala128His + Ser129AspLeu125Thr + Gly130Pro + Thr132SerLeu125Cys + Gly127Gln + Ser129AspGly127Asn + Gly130Glu + Thr132GlyGly127Gln + Gly130Ser + Thr132AspLeu125Thr + Ser129Asp + Thr132ProLeu125Cys + Gly127Glu + Gly130ProAla128Gly + Ser129Glu + Gly130GlnGly127Asp + Gly130Ser + Thr132ProLeu125Pro + Gly126Asn + Gly127AsnGly126Glu + Ala128Ser + Gly130SerLeu125Gly + Gly127Pro + Ser129AspGly126Gln + Ala128Glu + Gly130GlnAla128Gly + Gly130Asn + Thr132AspLeu125Met + Gly126Pro + Gly127GlnLeu125Gln + Ala128Asn + Ser129AspGly127Ser + Ala128Asn + Thr132GlyGly126Gln + Gly127Asp + Gly130SerGly127Asp + Ala128Pro + Thr132ProLeu125His + Ser129Asp + Thr132ProLeu125Ile + Gly126Gln + Thr132GlnGly126Asp + Gly130Ser + Thr132AsnGly127Asn + Ser129Glu + Gly130ProLeu125Pro + Gly127Asp + Gly130AsnLeu125Ile + Ala128His + Ser131Asp
Gly126Pro + Ala128His + Thr132ProLeu125Val + Ala128Gln + Ser129GluLeu125Ser + Gly126Asp + Thr132GlyLeu125Pro + Gly126Asp + Gly127AspAla128Ser + Gly130Asp + Ser131AspGly130Asp + Ser131Glu + Thr132AsnGly126Asn + Gly130Asp + Ser131GluGly126Gln + Gly130Asp + Ser131GluGly130Gln + Ser131Glu + Thr132GluAla128Thr + Ser131Glu + Thr132AspAla128Thr + Ser131Asp + Thr132GluAla128Pro + Ser131Asp + Thr132AspGly127Asp + Ala128Asp + Thr132GlnLeu125Glu + Gly126Glu + Gly130AsnAla128Asp + Ser129Asp + Gly130AsnSer129Asp + Gly130Glu + Thr132Pro表13環(huán)3-四突變變異體Leu125Asn + Ala128Asn + Ser131Glu + Thr132ProLeu125Ala + Gly126Gln + Gly127Pro + Gly1305erLeu125Pro + Gly126Asn + Gly127Gln + Ala128ProLeu125Gly + Gly126Asn + Ser129Glu + Gly130AsnAla128Gly + Gly130Asn + Ser131Asp + Thr132SerLeu125Ile + Gly126Pro + Ala128Gly + Thr132GluLeu125Thr + Gly130Pro + Ser131Asp + Thr132GlyLeu125Asp + Gly126Pro + Gly127Ser + Thr132GlnLeu125Gly + Gly126Pro + Gly127Asp + Thr132ProLeu125Val + Gly127Glu + Gly130Asn + Thr132AsnLeu125Cys + Ala128Pro + Gly130Ser + Ser131GluGly126Gln + Gly127Ser + Ala128Thr + Gly130GluLeu125Gly + Gly127Ser + Gly130Ser + Thr132GlyLeu125Pro + Gly126Ser + Ala128His + Thr132AsnLeu125Val + Gly126Gln + Gly127Pro + Ser129AspLeu125Cys + Gly130Pro + Ser131Asp + Thr132SerLeu125Gly + Gly127Gln + Ser129Glu + Thr132SerLeu125Met + Ala128Ser + Gly130Pro + Thr132GlyLeu125Ala + Gly126Asp + Ala128His + Thr132SerGly127Gln + Gly130Glu + Ser131Asp + Thr132AsnGly126Ser + Gly127Gln + Gly130Glu + Ser131GluGly127Pro + Gly130Asp + Ser131Glu + Thr132ProAla128Gly + Gly130Pro + Ser131Asp + Thr132GluGly126Asn + Ala128Pro + Ser131Asp + Thr132AspGly126Gln + Ala128His + Ser131Asp + Thr132AspLeu125Thr + Gly127Glu + Ala128Asp + Gly130GlnGly126Asn + Ala128Gln + Ser129Glu + Gly130Asp
Ala128Gly + Ser129Glu + Gly130Glu + Thr132GlnGly126Gln + Ser129Asp + Gly130Glu + Thr132AsnGly126Asn + Ala128Thr + Ser129Glu + Gly130GluGly126Ser + Gly127Asn + Ser129Glu + Gly130GluLeu125His + Gly127Glu + Ala128Glu + Ser129AspLeu125Thr + Gly127Glu + Ala128Asp + Ser129GluGly126Pro + Gly127Glu + Ala128Asp + Ser129GluAla128Gly + Ser129Glu + Gly130Asp + Ser131AspGly126Gln + Ser129Asp + Gly130Asp + Ser131AspGly127Ser + Ser129Glu + Gly130Glu + Ser131GluAla128Gly + Ser129Asp + Gly130Asp + Ser131GluGly126Asn + Ser129Asp + Gly130Glu + Ser131AspAla128Asn + Ser129Asp + Gly130Asp + Ser131AspGly127Pro + Ser129Glu + Gly130Asp + Ser131GluGly126Ser + Gly130Asp + Ser131Asp + Thr132GluLeu125Met + Gly127Glu + Ser129Asp + Gly130AspGly126Pro + Gly127Asp + Ser129Glu + Gly130AspLeu125Val + Gly127Glu + Ser129Asp + Gly130AspGly127Asp + Ala128Asn + Ser129Asp + Gly130GluGly127Glu + Ala128Thr + Ser129Glu + Gly130SerGly127Asp + Ala128His + Ser129Glu + Gly130SerGly126Ser + Gly127Glu + Ala128Thr + Ser129GluGly126Pro + Gly127Asp + Ala128Asp + Gly130GluLeu125Gly + Gly126Asp + Gly127Glu + Ser129GluGly126Asp + Gly127Glu + Ala128Ser + Ser129AspGly127Ser + Ser129Asp + Gly130Gln + Ser131AspAla128Gly + Ser129Glu + Gly130Asn + Ser131GluLeu125Val + Ser129Asp + Ser131Asp + Thr132SerLeu125Gly + Ala128Gly + Ser129Glu + Ser131AspLeu125Cys + Gly126Pro + Ser129Glu + Ser131GluGly127Ser + Ala128Asn + Ser129Asp + Ser131AspGly126Asn + Ser129Asp + Ser131Glu + Thr132GlnGly126Pro + Ala128Pro + Ser129Asp + Ser131Glu表14環(huán)4-單突變變異體Gly153AsnGly153AspGly153GlnGly153GluGly153ProGly153SerAsn154AspAsn154GlnAsn154GluAsn154Ser
Ser155AspSer155GluGly156AsnGly156AspGly156GlnGly156GluGly156ProGly156SerAsn157AspAsn157GlnAsn157GluAsn157SerSer158AspSer158GluGly159AsnGly159AspGly159GlnGly159GluGly159ProGly159SerSer160AspSer160GluThr161AsnThr161AspThr161GlnThr161GluThr161GlyThr161ProThr161SerAsn162AspAsn162GlnAsn162GluAsn162SerThr163AsnThr163AspThr163GlnThr163GluThr163GlyThr163ProThr163SerIle164AlaIle164AsnIle164AspIle164CysIle164GlnIle164GluIle164Gly
Ile164HisIle164LeuIle164MetIle164ProIle164SerIle164ThrIle164ValGly165AsnGly165AspGly165GlnGly165GluGly165ProGly165SerTyr166AlaTyr166AsnTyr166AspTyr166CysTyr166GlnTyr166GluTyr166GlyTyr166HisTyr166IleTyr166LeuTyr166MetTyr166ProTyr166SerTyr166ThrTyr166Val表15環(huán)4-雙突變變異體Asn154Ser + Ser155GluGly153Glu + Ile164AlaGly159Glu + Gly165ProAsn154Glu + Gly156SerGly156Pro + Gly159AspGly153Ser + Ser160AspSer155Asp + Gly165SerIle164Gly + Gly165GluGly159Glu + Ile164AsnAsn157Gln + Asn162SerGly156Asn + Thr161GlnAsn162Glu + Tyr166GlnAsn157Glu + Gly165GlnAsn162Glu + Gly165GlnGly153Pro + Ile164Cys
Thr163Glu + Gly165AsnGly153Asp + Ile164AsnGly153Ser + Thr161AspGly156Asp + Tyr166GlnGly165Pro + Tyr1665erGly153Ser + Asn162GluGly159Glu + Tyr166LeuGly156Asn + Thr161GluAsn154Gln + Ser158GluSer160Asp + Thr161AsnSer155Glu + Gly165ProAsn154Ser + Tyr166IleGly153Asp + Thr161AsnSer158Asp + Thr161ProAsn162Glu + Thr163SerGly159Gln + Tyr166CysThr161Gly + Asn162SerGly159Asp + Tyr166AsnSer160Glu + Ile164ThrSer158Asp + Ile164LeuAsn154Ser + Asn157AspAsn154Asp + Thr161GlnGly156Ser + Ser160GluGly156Gln + Tyr166IleGly153Asn + Tyr166AlaAsn154Asp + Ile164SerSer155Asp + Thr163AsnGly156Glu + Gly159GlnGly156Ser + Asn162AspGly153Ser + Asn157GluSer160Glu + Gly165ProGly153Pro + Asn157AspGly153Gln + Ser160AspAsn154Gln + Thr163AsnGly156Ser + Gly159AspGly159Asp + Thr161GlnGly159Ser + Tyr166GluAsn157Asp + Tyr166LeuThr161Asp + Asn162GlnAsn162Gln + Gly165GluGly156Asp + Asn162GlnAsn154Gln + Ile164GlnAsn157Asp + Asn162SerAsn154Asp + Ile164AsnGly159Asp + Gly165Pro表16
環(huán)4-三突變變異體Gly153Gln + Asn154Ser + Ser155GluGly156Asp + Thr161Pro + Tyr166IleAsn157Glu + Gly159Gln + Asn162GlnAsn154Glu + Asn162Gln + Ile164LeuGly153Pro + Gly159Gln + Ile164AsnAsn154Ser + Ile164Met + Tyr166CysGly159Gln + Ser160Asp + Ile164ThrGly156Asn + Thr161Ser + Ile164CysGly156Asn + Ile164His + Tyr166AspAsn157Glu + Asn162Gln + Ile164AlaSer160Asp + Thr163Ser + Ile164GlyAsn154Glu + Thr161Pro + Ile164MetThr163Asn + Ile164Asn + Gly165ProAsn157Glu + Asn162Gln + Ile164ThrAsn162Gln + Ile164Thr + Gly165GluAsn154Gln + Gly159Asp + Tyr166ThrAsn157Gln + Thr161Gly + Thr163ProSer158Glu + Thr163Ser + Gly165SerGly153Pro + Asn157Asp + Thr163AsnGly156Pro + Ser158Asp + Gly165GlnAsn157Gln + Thr163Gly + Gly165ProSer155Glu + Gly159Ser + Tyr166ThrGly153Gln + Asn154Gln + Gly165GluGly153Gln + Asn154Ser + Gly159ProThr161Gly + Asn162Asp + Gly165AsnSer158Glu + Gly159Ser + Gly165GlnSer155Asp + Gly159Asn + Ile164SerAsn157Gln + Thr163Gly + Gly165GluGly153Ser + Gly159Ser + Asn162AspGly153Ser + Ser158Glu + Ile164ThrGly153Asp + Thr161Ser + Gly165AsnThr161Ser + Ile164Ser + Tyr166ProSer155Asp + Gly156Asn + Ile164AsnAsn154Glu + Gly159Gln + Tyr166IleGly153Gln + Gly156Gln + Thr161AspGly159Asn + Thr163Asn + Tyr166SerSer155Asp + Thr163Pro + Ile164GlyGly153Ser + Ser160Asp + Ile164ProAsn162Glu + Ile164Gln + Tyr166GlnGly156Glu + Asn157Glu + Thr161GlyGly159Asp + Ser160Glu + Ile164GlnSer155Asp + Gly156Glu + Thr163AsnSer155Glu + Gly156Asp + Gly165SerSer155Glu + Gly156Asp + Ile164MetSer160Glu + Thr161Asp + Gly165AsnAsn162Asp + Thr163Asp + Gly165Asn
Asn157Asp + Ser158Glu + Thr161GluGly159Asp + Thr161Glu + Thr163ProGly153Asp + Asn154Ser + Gly165AspGly153Glu + Asn162Gln + Gly165AspAsn154Ser + Thr161Glu + Thr163AspSer158Glu + Ser160Asp + Thr163AsnSer158Glu + Ser160Glu + Thr163GlySer158Asp + Ser160Asp + Ile164GlyAsn157Asp + Gly159Glu + Thr163GlnThr163Glu + Gly165Asp + Tyr166ProSer160Asp + Asn162Glu + Gly165AsnAsn154Glu + Gly156Asp + Asn157GluGly159Asp + Ser160Asp + Asn162GluAsn154Glu + Ser155Glu + Asn157Asp表17環(huán)4-四突變變異體Gly153Pro + Gly159Ser + Thr163Pro + Gly165AsnAsn154Gln + Asn162Glu + Ile164Cys + Gly165ProAsn154Ser + Asn157Asp + Thr161Gly + Ile164AsnGly159Glu + Thr161Gln + Thr163Asn + Gly165GlnGly159Ser + Ser160Glu + Thr163Gln + Tyr166IleGly156Ser + Ser160Glu + Thr161Asn + Gly165ProGly153Ser + Gly159Ser + Ser160Asp + Gly165SerGly153Asn + Gly159Glu + Ile164Cys + Gly165ProAsn154Ser + Gly156Glu + Thr161Pro + Tyr166HisGly153Ser + Ser160Glu + Gly165Asn + Tyr166CysAsn157Gln + Thr163Gly + Ile164His + Gly165GlnAsn154Ser + Ser155Glu + Asn162Ser + Ile164HisGly156Pro + Asn157Gln + Ser160Glu + Gly165SerGly153Gln + Ser155Asp + Thr161Gln + Tyr166GlyAsn157Ser + Thr161Gln + Thr163Asp + Ile164ThrGly153Ser + Ser160Asp + Asn162Gln + Tyr166ThrGly153Pro + Ash154Gln + Asn157Gln + Ser160GluGly156Glu + Asn162Gln + Thr163Ser + Gly165GlnGly156Asn + Asn157Glu + Thr161Gly + Thr163SerGly153Ser + Ser158Glu + Gly159Asp + Thr163GlyAsn154Gln + Ser158Asp + Gly159Asp + Thr163AsnSer158Asp + Gly159Asp + Thr163Asn + Tyr166ProGly159Asp + Ser160Glu + Thr163Ser + Gly165SerAsn154Glu + Ser155Asp + Thr163Gln + Tyr165MetGly153Glu + Asn154Asp + Ser155Glu + Gly159AsnSer158Glu + Gly159Glu + Ser160Asp + Gly165AsnSer158Asp + Gly159Asp + Ser160Asp + Asn162GlnAsn157Ser + Ser160Asp + Thr161Glu + Asn162GluGly156Asn + Ser158Asp + Ser160Glu + Thr161Glu
Gly159Glu + Thr161Asp + Thr163Asn + Tyr166ValGly156Pro + Gly159Glu + Thr161Asp + Gly165ProGly153Glu + Gly156Pro + Gly159Gln + Gly165AspGly153Glu + Asn157Gln + Gly159Gln + Gly165AspGly153Asn + Ser155Glu + Thr163Glu + Ile164GlyAsn157Glu + Ser158Asp + Ser160Asp + Thr161AsnAsn154Ser + Asn157Ser + Ser158Asp + Ser160GluSer158Glu + Gly159Gln + Ser160Glu + Thr163GlnSer158Asp + Ser160Glu + Ile164Ala + Tyr166GlnGly155Pro + Asn157Gln + Ser158Glu + Ser160AspGly153Gln + Gly156Pro + Ser158Glu + Ser160AspGly156Ser + Asn157Glu + Gly159Glu + Ile164LeuThr163Asp + Ile164Cys + Gly165Asp + Tyr166CysSer160Asp + Asn162Asp + Ile164Asn + Tyr166ThrAsn157Ser + Ser160Glu + Asn162Asp + Ile164MetSer155Glu + Asn157Asp + Asn162Ser + Gly165ProGly153Ser + Asn154Glu + Ser155Glu + Asn157GluGly153Pro + Asn154Asp + Ser155Asp + Asn157GluGly153Gln + Ser158Glu + Ser160Asp + Asn162GluAsn157Asp + Ser160Glu + Thr161Ser + Asn162GluGly156Asp + Asn157Asp + Thr161Ser + Asn162AspAsn154Gln + Asn157Asp + Ser160Glu + Tyr166SerGly153Gln + Gly156Glu + Ser158Glu + Asn162GluGly156Glu + Gly159Pro + Thr163Asp + Gly165AspSer155Glu + Thr161Ser + Thr163Ser + Gly165AspAsn154Asp + Thr163Gly + Gly165Glu + Tyr166SerSer155Asp + Asn157Glu + Gly159Glu + Gly165GlnAsn162Glu + Ile164Ala + Gly165Asp + Tyr166ThrGly156Gln + Asn162Asp + Gly165Asp + Tyr166AsnSer155Glu + Ser158Asp + Thr161Asn + Tyr166MetSer155Glu + Ser158Glu + Gly159Gln + Ile164Gly表18環(huán)5-單突變變異體Ala186AsnAla186AspAla186GlnAla186GluAla186GlyAla186HisAla186ProAla186SerAla186ThrSer187AspSer187GluPhe188Ala
Phe188AsnPhe188AspPhe188CysPhe188GlnPhe188GluPhe188GlyPhe188HisPhe188IlePhe188LeuPhe188MetPhe188ProPhe188SerPhe188ThrPhe188TyrPhe188ValSer189AspSer189GluSer190AspSer190Glu表19環(huán)5-雙突變變異體Ala186Asp + Phe188GlnAla186Gly + Ser187GluAla186Gln + Ser187AspAla186His + Ser190GluSer187Asp + Phe188CysSer187Asp + Phe188ThrPhe188Gly + Ser190GluAla186Asn + Phe188SerAla186Asp + Phe188IleAla186Ser + Phe188GluAla186Thr + Phe188LeuSer187Glu + Phe188SerSer187Glu + Phe188ProSer187Asp + Phe188GlnAla186Thr + Ser190GluPhe188His + Ser190AspAla186Gln + Ser190AspPhe188Leu + Ser190GluAla186His + Ser190AspAla186Pro + Ser187GluPhe188Ile + Ser190AspAla186Asp + Phe188AsnAla186Ser + Ser187GluSer187Glu + Phe188Leu
Ala186Asn + Phe188IlePhe188Tyr + Ser190GluAla186Pro + Ser190GluAla186Thr + Phe188AspAla186His + Ser187AspAla186Gln + Phe188ThrPhe188Cys + Ser190GluSer187Glu + Phe188ThrSer187Asp + Phe188MetAla186Asn + Ser190AspPhe188Val + Ser190AspAla186Pro + Phe188SerAla186Asp + Phe188SerSer187Glu + Phe188AsnAla186Thr + Ser187AspAla186Thr + Phe188AsnAla186Gly + Phe188LeuPhe188Asn + Ser190GluAla186Thr + Ser187GluAla186Thr + Ser190AspSer187Asp + Phe188AlaAla186Asn + Ser187GluAla186Gln + Phe188TyrPhe188Ile + Ser190GluPhe188Thr + Ser190AspAla186Ser + Ser187AspPhe188Ala + Ser190AspPhe188Gln + Ser190GluAla186Gly + Ser190AspAla186Glu + Phe188ThrAla186Pro + Phe188AlaPhe188Asn + Ser190AspAla186Gly + Phe188ValPhe188Leu + Ser190AspAla186Asn + Phe188MetAla186Gln + Phe188Glu表20環(huán)5-三突變變異體Ala186Pro + Phe188Cys + Ser190GluAla186Gln + Phe188Val + Ser190GluAla186Thr + Phe188Asn + Ser190GluAla186Gly + Phe188Gln + Ser190GluAla186Pro + Phe188Gln + Ser190GluAla186Gln + Phe188Asn + Ser190AspAla186Asn + Ser187Asp + Phe188Leu
Ala186Gln + Ser187Asp + Phe188ProAla186Gln + Ser187Glu + Phe188MetAla186Pro + Phe188Val + Ser190GluAla186His + Phe188Met + Ser190AspAla186Asn + Ser187Glu + Phe188IleAla186Gln + Phe188Ala + Ser190GluAla186Pro + Phe188Thr + Ser190AspAla186Asn + Ser187Glu + Phe188ThrAla186Ser + Phe188Ala + Ser190GluAla186His + Phe188Pro + Ser190AspAla186His + Ser187Asp + Phe188LeuAla186Pro + Ser187Glu + Phe188ValAla186Asn + Ser187Asp + Phe188HisAla186Gly + Ser187Glu + Phe188LeuAla186Pro + Ser187Asp + Phe188HisAla186Gln + Phe188Ile + Ser190AspAla186Pro + Phe188Ile + Ser190AspAla186Gln + Phe188Thr + Ser190AspAla186Gln + Phe188His + Ser190GluAla186Gly + Phe188Ile + Ser190AspAla186Thr + Ser187Glu + Phe188TyrAla186His + Ser187Asp + Phe188SerAla186His + Ser187Glu + Phe188SerAla186His + Phe188Gly + Ser190GluAla186Pro + Ser187Glu + Phe188ProAla186Gln + Ser187Glu + Phe188TyrAla186Asn + Phe188Cys + Ser190AspAla186Gly + Phe188Gly + Ser190AspAla186Gln + Ser187Asp + Phe188ValAla186Pro + Ser187Glu + Phe188GlyAla186Glu + Ser187Glu + Phe188IleAla186Asp + Ser187Glu + Phe188AlaAla186Asp + Ser187Asp + Phe188ThrAla186Glu + Ser187Asp + Phe188ThrAla186Glu + Ser187Glu + Phe188TyrAla186Asp + Ser187Asp + Phe188SerAla186Asp + Ser187Glu + Phe188IleAla186Pro + Ser187Asp + Phe188AspAla186Gln + Ser187Glu + Phe188GluAla186Gln + Ser187Glu + Phe188AspAla186Glu + Ser187Glu + Phe188AspAla186Glu + Ser187Asp + Phe188GluAla186Thr + Phe188Asp + Ser190GluAla186Ser + Phe188Asp + Ser190AspAla186Gly + Phe188Asp + Ser190GluSer187Asp + Phe188Glu + Ser190AspSer187Glu + Phe188Glu + Ser190Asp
Ser187Asp + Phe188Glu + Ser190GluSer187Glu + Phe188Glu + Ser190GluAla186Glu + Phe188Glu + Ser190GluAla186Glu + Phe188Asp + Ser190AspAla186Asp + Ser187Asp + Ser190GluAla186Asp + Ser187Glu + Ser190Asp表21環(huán)5-四突變變異體Ala186Thr + Ser187Glu + Phe188Glu + Ser190AspAla186Gln + Ser187Asp + Phe188Asp + Ser190AspAla186Gly + Ser187Asp + Phe188Glu + Ser190AspAla186Thr + Ser187Asp + Phe188Glu + Ser190GluAla186His + Ser187Glu + Phe188Glu + Ser190AspAla186Gln + Ser187Glu + Phe188Glu + Ser190GluAla186Thr + Ser187Glu + Phe188Asp + Ser190AspAla186Asn + Ser187Glu + Phe188Asp + Ser190AspAla186Ser + Ser187Glu + Phe188Glu + Ser190GluAla186Asn + Ser187Asp + Phe188Asp + Ser190AspAla186Ser + Ser187Asp + Phe188Asp + Ser190GluAla186Asn + Ser187Asp + Phe188Asp + Ser190GluAla186His + Ser187Glu + Phe188Glu + Ser190GluAla186Gln + Ser187Asp + Phe188Glu + Ser190AspAla186Gly + Ser187Asp + Phe188Glu + Ser190GluAla186Pro + Ser187Asp + Phe188Asp + Ser190GluAla186Gly + Ser187Asp + Phe188Asp + Ser190GluAla186Ser + Ser187Glu + Phe188Glu + Ser190AspAla186Thr + Ser187Asp + Phe188Asp + Ser190AspAla186Gly + Ser187Glu + Phe188Asp + Ser190GluAla186Gly + Ser187Asp + Phe188Asp + Ser190AspAla186Asp + Ser187Asp + Phe188Asn + Ser190AspAla186Glu + Ser187Glu + Phe188Tyr + Ser190GluAla186Asp + Ser187Asp + Phe188Ser + Ser190GluAla186Asp + Ser187Glu + Phe188Gln + Ser190GluAla186Glu + Ser187Asp + Phe188Ala + Ser190GluAla186Glu + Ser187Glu + Phe188Gly + Ser190GluAla186Asp + Ser187Asp + Phe188Ala + Ser190AspAla186Glu + Ser187Glu + Phe188Cys + Ser190GluAla186Glu + Ser187Asp + Phe188Ile + Ser190AspAla186Glu + Ser187Glu + Phe188Ala + Ser190GluAla186Glu + Ser187Glu + Phe188His + Ser190AspAla186Asp + Ser187Asp + Phe188Leu + Ser190GluAla186Asp + Ser187Asp + Phe188Gln + Ser190GluAla186Asp + Ser187Glu + Phe188Asn + Ser190AspAla186Asp + Ser187Asp + Phe188Cys + Ser190GluAla186Asp + Ser187Asp + Phe188Thr + Ser190Asp
Ala186Glu + Ser187Glu + Phe188Leu + Ser190AspAla186Glu + Ser187Glu + Phe188Leu + Ser190GluAla186Asp + Ser187Asp + Phe188Pro + Ser190GluAla186Asp + Ser187Glu + Phe188Val + Ser190AspAla185Asp + Ser187Asp + Phe188His + Ser190GluAla186Glu + Ser187Glu + Phe188Pro + Ser190GluAla186Asp + Ser187Glu + Phe188Ser + Ser190GluAla186Glu + Ser187Glu + Phe188Val + Ser190GluAla186Asp + Ser187Glu + Phe188Gln + Ser190AspAla186Asp + Ser187Glu + Phe188Asn + Ser190GluAla186Asp + Ser187Glu + Phe188Gly + Ser190GluAla186Asp + Ser187Asp + Phe188Cys + Ser190AspAla186Asp + Ser187Glu + Phe188Tyr + Ser190GluAla186Glu + Ser187Asp + Phe188Thr + Ser190AspAla186Glu + Ser187Glu + Phe188Thr + Ser190GluAla186Glu + Ser187Glu + Phe188Met + Ser190GluAla186Asp + Ser187Asp + Phe188Gly + Ser190GluAla186Asp + Ser187Glu + Phe188His + Ser190GluAla186Asp + Ser187Glu + Phe188Met + Ser190GluAla186Glu + Ser187Asp + Phe188Ser + Ser190AspAla186Asn + Ser187Glu + Phe188Met + Ser190AspAla186Gln + Ser187Asp + Phe188His + Ser190AspAla186Asn + Ser187Glu + Phe188Ser + Ser190Glu表22環(huán)6-單突變變異體Ala199AsnAla199AspAla199GlnAla199GluAla199GlyAla199HisAla199ProAla199SerAla199ThrPro200AsnPro200AspPro200GlnPro200GluPro200GlyPro200SerGly201AsnGly201AspGly201GlnGly201GluGly201Pro
Gly201SerAla202AsnAla202AspAla202GlnAla202GlnAla202GlyAla202HisAla202ProAla202SerAla202ThrGly203AsnGly203AspGly203GlnGly203GluGly203ProGly203SerVal204AlaVal204AsnVal204AspVal204CysVal204GlnVal204GluVal204GlyVal204HisVal204MetVal204ProVal204SerVal204ThrTyr205AlaTyr205AsnTyr205AspTyr205CysTyr205GlnTyr205GluTyr205GlyTyr205HisTyr205IleTyr205LeuTyr205MetTyr205ProTyr205SerTyr205ThrTyr205ValSer206AspSer206GluThr207AsnThr207Asp
Thr207GlnThr207GluThr207GlyThr207ProThr207SerTyr208AlaTyr208AsnTyr208AspTyr208CysTyr208GlnTyr208GluTyr208GlyTyr208HisTyr208IleTyr208LeuTyr208MetTyr208ProTyr208SerTyr208ThrTyr208ValPro209AsnPro209AspPro209GlnPro209GluPro209GlyPro209SerThr210AsnThr210AspThr210GlnThr210GluThr210GlyThr210ProThr210SerAsn211AspAsn211GlnAsn211GluAsn211SerThr212AsnThr212AspThr212Glnlhr212GluThr212GlyThr212ProThr212SerTyr213AlaTyr213AsnTyr213Asp
Tyr213CysTyr213GlnTyr213GluTyr213GlyTyr213HisTyr213IleTyr213LeuTyr213MetTyr213ProTyr213SerTyr213ThrTyr213ValAla214AsnAla214AspAla214GlnAla214GluAla214GlyAla214HisAla214ProAla214SerAla214ThrThr215AsnThr215AspThr215GlnThr215GluThr215GlyThr215ProThr215SerLeu216AlaLeu216AsnLeu216AspLeu216CysLeu216GlnLeu216GluLeu216GlyLeu216HisLeu216IleLeu216MetLeu216ProLeu216SerLeu216ThrLeu216ValAsn217AspAsn217GlnAsn217GluAsn217SerGly218Asn
Gly218AspGly218GlnGly218GluGly218ProGly218SerThr219AsnThr219AspThr219GlnThr219GluThr219GlyThr219ProThr219Ser表23環(huán)6-雙突變變異體Gly218Gln + Thr219GlnVal204Ala + Thr207GlnTyr213Val + Gly218ProPro209Gly + Tyr213IleThr215Ser + Thr219SerLeu216Ala + Asn217GlnPro209Asp + Thr212SerAla199Asn + Leu216IleAsn217Glu + Thr219AsnTyr208Ile + Thr215AsnAla202Gly + Pro209GluGly201Ser + Thr219GlyThr215Pro + Gly218SerTyr205Leu + Leu216HisVal204Asn + Ala214AsnAsn217Ser + Gly218ProAla202His + Tyr208ValGly201Gln + Tyr213LeuTyr213Cys + Thr219GlyPro200Asn + Thr210ProAla202Asp + Thr219AsnGly201Pro + Val204ProAla214Thr + Thr219GlnThr207Gln + Leu216AspPro200Gln + Tyr213MetVal204Ala + Thr215GluThr212Gln + Thr215AsnLeu216His + Asn217GluAla202Thr + Ala214AspVal204Pro + Thr207GlnGly203Asp + Tyr205Leu
Tyr205His + Asn211AspAsn211Gln + Thr212AsnGly203Pro + Thr219ProPro200Gly + Pro209AspThr210Asp + Gly218ProGly203Asp + Gly218SerTyr205Glu + Gly218ProThr212Gln + Thr219ProAla214Ser + Asn217GluAla202Asn + Leu216SerVal204Thr + Ala214ProPro200Asn + Asr211AspTyr205Gly + Thr219ProAla199His + Asn217SerGly201Ser + Asn211AspGly203Ser + Tyr205AlaAla202Pro + Pro209GluTyr213Pro + Gly218GluPro200Gly + Ala202GluThr210Gly + Asn211AspVal204Asn + Thr215AsnPro200Asn + Tyr208LeuAla202Pro + Ala214ThrTyr205Cys + Leu216HisLeu216Ser + Gly218GlnThr215Glu + Thr219GlnTyr205Leu + Pro209GluAla214Glu + Thr215ProGly203Glu + Gly218GlnTyr205Cys + Thr212ProLeu216Asn + Thr219AsnAla199Ser + Thr219AsnGly201Gln + Tyr213ValGly203Asn + Thr215GlnPro209Glu + Asn211GlnAla199Gly + Asn211AspAla214Glu + Leu216MetVal204Pro + Asn211AspAla202Glu + Gly218GlnTyr208Ile + Gly218GluTyr205Ser + Leu216ThrAsn211Glu + Tyr213GlyThr212Asn + Thr219AsnAsn217Asp + Thr219GlnThr207Asn + Asn217AspGly201Ser + Tyr208GlnThr210Pro + Thr212Gly
Val204Asn + Thr210GlyAla199His + Tyr208ValTyr208Gly + Tyr213ValGly201Pro + Ala214AspGly201Gln + Leu216GlyGly201Gln + Thr207AsnGly203Asn + Gly218GluTyr208Asn + Thr219SerThr207Gly + Leu216GluGly218Glu + Thr219AsnAla199Gln + Thr212GluThr212Asp + Ala214GlyGly203Asn + Ala214SerAsn211Ser + Thr212GlnPro209Asp + Tyr213ThrGly201Gln + Ala202ThrVal204Thr + Tyr208CysGly201Asn + Ala202AsnAla199His + Thr210AspVal204Ser + Thr212AspTyr208Ser + Thr219GlnTyr205Pro + Thr215AspAsn211Gln + Gly218GlnAsn217Asp + Gly218ProAla202Gln + Thr210ProTyr205Asn + Gly218GlnGly201Gln + Thr219SerAla202Asp + Thr207AsnThr212Gly + Asn217GlnThr215Asn + Thr219SerAla214Glu + Thr215GlyThr210Asn + Tyr213GlnAla199Gln + Tyr205GluTyr205His + Thr210AspGly201Asn + Gly203AspGly203Ser + Pro209AspGly201Gln + Thr210GluPro209Asn + Thr215SerThr215Gln + Thr219GlyGly201Gln + Thr215AspTyr208Gly + Tyr213ProGly201Asn + Tyr213AsnAla202Pro + Pro209AsnAla199Thr + Tyr208MetThr210Gln + Gly218AsnThr210Pro + Asn211AspAsn211Asp + Gly218Pro
Gly201Asn + Tyr205GlyAla202Gln + Tyr213GluAla199His + Ala202AspTyr205Ser + Tyr213GluGly201Gln + Pro209AspAla202Ser + Thr219GlnLeu216Met + Thr219AsnVal204Ser + Tyr205SerVal204Thr + Tyr208SerGly203Asn + Ala214ThrThr210Gly + Thr219AsnPro200Gly + Thr210ProAla214Glu + Gly218ProPro200Gly + Thr219GlnAla202Ser + Asn211AspAla199Thr + Val204GlnAla199Gln + Pro200SerPro209Ser + Leu216ThrTyr205Pro + Leu216AlaGly201Pro + Pro209GlyThr207Asn + Thr215GluPro200Ser + Tyr205AlaThr210Gln + Asn217GlnAla199Asn + Gly203GlnGly201Asn + Thr207ProGly203Asp + Tyr208AsnThr207Asn + Asn211AspTyr205Leu + Gly218AspGly203Ser + Thr210ProTyr205Cys + Asn211SerAla214Glu + Asn217SerAla199Pro + Gly201GlnGly201Asn + Tyr213MetTyr205His + Tyr208GlyGly203Glu + Thr210SerThr207Asn + Ala214SerTyr205Val + Thr219SerVal204Asn + Thr207ProThr207Ser + Asn211AspTyr205Leu + Gly218GluAsn211Ser + Thr219ProTyr205Gln + Asn211SerPro200Gly + Val204AsnAla202Glu + Tyr213HisGly201Pro + Gly218GlnAla214Asn + Asn217GlnGly201Asn + Asn211Asp
Pro209Ser + Thr212GlnGly203Gln + Val204GlnGly201Gln + Thr215ProGly201Asn + Ala202SerThr207Gln + Thr212AspVal204Gly + Thr212AspGly201Asn + Thr210AspThr210Asp + Tyr213Cys表24環(huán)6-三突變變異體Pro200Gly + Thr212Ser + Tyr213MetGly201Pro + Pro209Glu + Thr212GlyGly203Asp + Val204Met + Asn217GlnPro200Gly + Thr212Ser + Ala214AspAla199Gly + Tyr213Asp + Gly218SerGly201Asn + Thr207Ser + Thr215GluPro200Ser + Ala214Gly + Asn217GluAla199Pro + Thr207Ser + Thr210ProGly201Asn + Thr210Ser + Asn211GluGly203Asn + Thr207Asn + Asn217GluGly201Asn + Tyr205Pro + Thr207SerPro200Asn + Gly201Gln + Gly203SerPro200Gln + Asn211Glu + Tyr213AlaAla202His + Thr212Asp + Tyr213HisAla199Thr + Gly201Asn + Asn211SerPro200Ser + Gly201Gln + Pro209AsnAla202Gly + Tyr205Pro + Asn211SerGly203Gln + Tyr205Val + Thr210ProGly203Glu + Asn211Gln + Asn217SerPro200Asn + Pro209Glu + Asn211SerGly201Pro + Thr210Ser + Tyr213ThrTyr205Ile + Leu216Asp + Thr219GlyPro200Ser + Val204Met + Pro209AspTyr205Asp + Thr207Gly + Tyr208GlnPro200Ser + Thr210Asp + Thr212ProGly201Gln + Asn211Asp + Tyr213GlyTyr208Val + Thr210Gly + Leu216ValPro209Gly + Thr215Pro + Leu216AlaVal204His + Thr215Glu + Leu216CysAla199Pro + Gly203Ser + Thr210SerGly201Asn + Tyr205Met + Thr210AspPro200Gly + TyT205Cys + Thr212AsnAla199Pro + Pro200Ser + Leu216GluGly201Asn + Tyr205Asp + Tyr213llePro200Ser + Ala202Gly + Thr210Asp
Ala202Asn + Val204Ser + Gly218GluGly203Gln + Val204Cys + Thr207SerVal204Cys + Thr207Asn + Thr212AspGly201Pro + Gly203Ser + Tyr208GlyGly203Asn + Tyr208Cys + Leu216MetThr210Gln + Thr215Asp + Thr219SerPro209Asp + Tyr213Leu + Ala214ThrAla202Asp + Tyr213Pro + Thr215GlnThr215Glu + Gly218Ser + Thr219GlnPro200Gly + Val204His + Thr207SerGly203Ser + Pro209Asp + Thr219SerAla202Asn + Thr207Gln + Ala214HisThr207Ser + Ala214Glu + Leu216SerPro200Asn + Pro209Asn + Thr210ProPro209Gln + Tyr213His + Ala214ThrThr210Asn + Thr215Pro + Leu216GlnGly203Gln + Val204Ser + Thr210GluPro200Asn + Asn211Gln + Gly218SerPro200Gln + Thr207Gln + Leu216GluTyr213Pro + Gly218Glu + Thr219AsnTyr208Cys + Thr210Asp + Thr219SerPro200Gln + Tyr205His + Thr219SerPro209Asp + Thr212Gln + Thr219GlnGly201Asn + Val204Asn + Thr210AspPro200Gly + Tyr208Leu + Thr210AsnPro200Gln + Thr207Gln + Thr219GlnTyr205Val + Pro209Ser + Thr210AspGly203Pro + Val204His + Tyr213GluGly203Asn + Pro209Asn + Tyr213HisAla199Asn + Gly201Asn + Leu216AspPro209Asp + Tyr213His + Gly218SerGly201Ser + Gly203Asp + Asn211SerGly203Asn + Val204Cys + Thr212GlyGly201Pro + Tyr208Met + Leu216GluGly201Ser + Val204Gln + Leu216HisAla199Gly + Ala202Pro + Leu216SerGly203Gln + Tyr205Pro + Thr219GlyTyr205Gly + Tyr213Val + Asn217AspGly201Asn + Ala214His + Thr215GlnPro200Gly + Ala202Gln + Tyr213CysGly203Pro + Tyr208Met + Thr219ProPro200Asn + Tyr205Leu + Gly218GluPro209Gly + Thr212Glu + Tyr213ThrPro200Gln + Thr210Pro + Thr219AsnGly201Ser + Gly218Pro + Thr219AsnPro200Ser + Gly201Pro + Thr219GlyThr207Pro + Tyr213Ala + Leu216Glu
Ala199Ser + Thr207Asn + Thr212AspThr207Gln + Tyr208Ile + Thr210AspTyr213Ser + Thr215Gln + Thr219ProPro200Gly + Tyr208Ile + Ala214SerGly201Asn + Pro209Ser + Thr215AspPro200Ser + Pro209Asp + Tyr213MetTyr208Gly + Thr210Glu + Thr215SerAla202Ser + Ala214Asp + Thr219SerVal204Pro + Pro209Asn + Thr219AsnThr207Asn + Pro209Asn + Gly218SerPro200Gly + Leu216Cys + Gly218SerPro209Gly + Leu216Val + Gly218GluAsn211Gln + Tyr213Thr + Thr215SerTyr213Val + Thr215Glu + Gly218SerTyr213Pro + Gly218Gln + Thr219SerGly201Pro + Ala202Thr + Val204CysAla202Ser + Tyr208Leu + Pro209AspTyr208Thr + Thr210Asn + Ala214ThrTyr205Pro + Pro209Ser + Thr215GlnTyr205Cys + Tyr208Gly + Leu216GluAla199Gly + Gly203Asp + Pro209AsnVal204Gln + Thr207Gly + Gly218GlnAla202His + Gly218Asp + Thr219GlnGly203Asp + Ash211Ser + Leu216MetPro200Ser + Val204Cys + Thr215GluThr207Asn + Leu216Pro + Asn217GluAla199His + Tyr205His + Thr212ProAla199Gln + Gly203Asp + Thr219GlyAla199His + Pro200Gln + Asn217AspVal204Ser + Tyr213Leu + Thr219GlnVal204Ser + Pro209Ser + Leu216GluAla202Gly + Pro209Asn + Thr219AsnThr215Gln + Gly218Asn + Thr219GlnAla199Pro + Ala214Ser + Leu216ValGly203Asn + Thr215Asn + Gly218GluThr207Pro + Tyr208Ala + Thr219AsnAla199Ser + Thr210Ser + Gly218GlnPro209Asn + Thr210Ser + Asn211SerGly201Asn + Val204Cys + Tyr213ValGly201Ser + Val204Ser + Pro209AsnGly201Pro + Thr210Ser + Asn211SerTyr205Gln + Tyr208His + Thr210AspTyr208Ile + Thr215Ser + Gly218GluThr212Glu + Ala214His + Thr215GlnGly201Asn + Val204Gly + Tyr205GlnAla199Thr + Pro209Glu + Thr210SerPro209Gly + Thr210Asn + Gly218Glu
Gly203Pro + Pro209Asn + Thr210AsnGly203Glu + Tyr213Pro + Leu216ThrAla199Pro + Thr212Glu + Ala214ThrThr207Gln + Pro209Asn + Thr219SerAla199Thr + Tyr205Gln + Thr219ProVal204Met + Ala214Gly + Asn217GluGly201Gln + Val204Pro + Thr210GluTyr208Leu + Thr212Asp + Gly218AsnGly201Gln + Tyr205His + Thr210GluGly201Pro + Ala214Gly + Thr219SerGly203Asp + Tyr205Ala + Tyr208SerThr212Pro + Asn217Ser + Gly218AspTyr205Asp + Thr207Gly + Tyr208LeuGly201Pro + Ala202Glu + Leu216SerAla202His + Tyr208Pro + Thr215AspTyr208Thr + Thr212Glu + Thr219SerVal204Met + Pro209Gln + Leu216AspThr207Pro + Thr210Asn + Tyr213GluGly201Ser + Val204Gln + Asn211AspGly203Gln + Tyr205Asn + Tyr213SerAla199Thr + Thr210Asn + Tyr213GluTyr208Ile + Pro209Ser + Thr212ProTyr205Leu + Tyr208Thr + Ala214GluPro200Gln + Val204Gln + Ala214GluGly203Pro + Asn211Asp + Asn217GlnGly201Asn + Gly218Gln + Thr219ProAla199Gln + Gly203Gln + Ala214AspAla202Thr + Tyr205Cys + Tyr213IleGly201Gln + Val204Cys + Asn211GluTyr208Thr + Pro209Gly + Thr215AspThr210Glu + Asn211Gln + Tyr213LeuVal204Cys + Tyr208Pro + Thr219ProAla202Asp + Gly203Asn + Gly218AsnAla199Asn + Val204Gly + Tyr205ProGly203Pro + Val204Met + Tyr208HisPro200Gly + Tyr213Pro + Leu216CysVal204Ala + Ala214His + Asn217GluAla202Gln + Thr207Asn + Ala214GlyGly203Ser + Val204Gly + Tyr213ThrAsn211Asp + Leu216Cys + Thr219AsnPro200Asn + Thr212Gly + Gly218AsnTyr205His + Tyr208Leu + Thr215GlnGly203Ser + Tyr205His + Thr219SerVal204Asn + Thr212Pro + Thr215AspThr212Ser + Thr215Ser + Leu216AlaTyr208Leu + Thr212Glu + Leu216MetThr207Asn + Tyr208Leu + Leu216Asp
Thr210Gly + Thr212Pro + Asn217AspAsn211Ser + Ala214Asn + Leu216GluThr207Ser + Tyr208Thr + Ala214HisAla202Asp + Val204Ser + Tyr208Thr表25環(huán)6-四突變變異體Pro200Ser + Thr207Ser + Tyr213Cys + Gly218GlnAla202Gln + Tyr208Cys + Thr210Pro + Tyr213AsnGly203Asp + Tyr205Ile + Thr215Pro + Thr219SerGly201Gln + Gly203Gln + Val204Asn + Thr207AsnPro200Ser + Gly201Ser + Ala202Ser + Pro209GluGly201Ser + Thr207Pro + Thr215Asp + Thr219ProAla202Gln + Asn211Gln + Tyr213Ser + Thr219AsnGly201Pro + Tyr213Ala + Ala214Pro + Thr215SerAla199Gly + Pro200Gly + Ala202Glu + Gly218ProTyr205Thr + Tyr208Met + Thr210Ser + Asn217GlnGly201Ser + Ala214Ser + Leu216Gly + Gly218AsnPro200Gln + Gly203Gln + Asn211Glu + Thr215ProPro200Ser + Tyr208Leu + Asn211Ser + Gly218AspAla199Asn + Pro209Asn + Thr210Ser + Tyr213MetAla199His + Gly203Ser + Pro209Asp + Thr212GlnTyr205Gly + Thr212Gly + Thr215Asn + Asn217GlnAla199Gln + Tyr208Thr + Thr210Asn + Tyr213GlnGly203Ser + Pro209Asp + Asn211Ser + Ala214ThrPro209Asn + Thr215Ser + Gly218Asp + Thr219GlyAla199Pro + Thr207Gln + Thr210Glu + Thr215ProAla202Asn + Val204Gly + Thr207Gly + Thr212AspVal204Thr + Tyr208Leu + Asn211Ser + Asn217AspPro200Gly + Tyr205Val + Leu216Asp + Thr219GlyPro200Gly + Tyr208Ser + Ala214Ser + Thr219SerAla199Thr + Gly203Glu + Thr207Asn + Tyr213CysPro200Gly + Tyr213Thr + Asn217Gln + Thr219ProGly203Asn + Val204Thr + Thr207Asn + Thr210SerAla202Gly + Thr207Gln + Pro209Gln + Thr210AspAla199His + Gly201Gln + Tyr205Asp + Thr219GlnThr207Ser + Asn211Asp + Tyr213Thr + Leu216SerAla199Ser + Thr210Glu + Asn211Gln + Leu216IleGly201Ser + Gly203Ser + Thr212Pro + Ala214GluPro200Gly + Gly201Gln + Gly203Gln + Leu216HisAla199Pro + Gly201Gln + Gly203Pro + Thr212GluAla199His + Gly201Asn + Val204Thr + Tyr208SerVal204Thr + Tyr208Ile + Ala214Glu + Thr219ProAla199Ser + Tyr205Pro + Thr212Ser + Thr219GlnVal204Met + Leu216Cys + Asn217Ser + Thr219GlyGly201Ser + Tyr205His + Pro209Gln + Thr219Ser
Pro200Gln + Pro209Asn + Asn211Asp + Thr219GlyVal204Met + Thr210Gly + Thr212Asp + Ala214SerPro200Gly + Thr207Asn + Asn211Glu + Leu216HisTyr208Ala + Thr210Asp + Thr215Gln + Thr219GlyGly203Asn + Thr207Gln + Pro209Asp + Thr215GlyAla199Ser + Pro209Glu + Leu216His + Thr219ProGly203Asp + Tyr205His + Tyr213Val + Ala214ThrPro200Asn + Thr212Asn + Leu216His + Gly218ProAla199Ser + Pro209Gln + Thr215Gln + Asn217GluGly203Asp + Thr207Pro + Thr212Gly + Ala214GlnGly201Pro + Gly203Gln + Thr207Pro + Ala214AspVal204Gln + Ala214Asp + Leu216Thr + Gly218SerThr207Asn + Tyr208Pro + Pro209Asp + Tyr213AsnPro209Asp + Asn211Gln + Thr215Gln + Gly218ProVal204Gln + Asn211Ser + Asn217Gln + Gly218ProGly201Gln + Ala202His + Asn217Asp + Gly218GlnVal204Thr + Tyr208Leu + Ala214Pro + Gly218AspGly201Ser + Thr207Pro + Thr212Ser + Thr215ProAla202Gly + Val204Gly + Asn211Glu + Leu216GlyAla199Ser + Gly201Gln + Ala214Ser + Asn217SerAla202Gln + Thr207Pro + Pro209Glu + Tyr213ProAla199Thr + Ala202Ser + Val204Ala + Thr207GlnAla199His + Thr207Ser + Pro209Gln + Ala214GluAla202His + Gly203Asp + Val204Pro + Gly218ProAla202Asn + Gly203Glu + Val204Gln + Thr210GlnPro200Asn + Val204Gly + Tyr205Gln + Thr207SerAla202Glu + Gly203Pro + Thr207Gly + Thr219GlyGly201Asn + Tyr208Val + Gly218Glu + Thr219AsnPro200Ser + Gly201Gln + Tyr205Asp + Thr219GlyPro200Ser + Thr207Gly + Ala214Pro + Thr215AspVal204Ala + Thr207Gln + Asn211Asp + Thr215ProAla202Thr + Val204Asn + Thr207Ser + Pro209AsnAla202Asn + Val204Thr + Thr207Gly + Pro209SerAla199Pro + Thr210Gln + Tyr213Cys + Ala214ThrAla199Pro + Pro200Gln + Ala214Glu + Thr215AsnPro200Ser + Val204Gln + Thr207Ser + Leu216ThrAla199Ser + Thr207Asn + Leu216Gln + Gly218GlnGly201Ser + Tyr208Ala + Ala214Ser + Leu216GlnVal204Thr + Thr207Ser + Thr215Asp + Asn217SerGly201Asn + Thr207Pro + Tyr213Met + Asn217AspVal204Ala + Asn211Gln + Thr215Glu + Gly218SerPro200Gly + Pro209Ser + Thr212Gly + Asn217GluAla199Pro + Tyr205Ile + Thr210Gly + Thr219GlyPro200Gly + Tyr205Ala + Thr212Pro + Tyr213GluGly203Ser + Thr212Ser + Leu216Ala + Asn217AspGly201Pro + Gly203Gln + Pro209Asp + Thr215GlyAla202His + Gly203Ser + Tyr208Cys + Gly218Asn
Ala199Pro + Ala202Gln + Val204Gly + Thr212GlyAla199Asn + Pro200Gly + Gly201Gln + Val204GlnGly201Asn + Gly203Pro + Pro209Ser + Thr215AspGly201Asn + Thr210Ser + Tyr213Asp + Leu216GlnGly203Gln + Val204His + Gly218Gln + Thr219GlyGly201Asn + Tyr213Val + Ala214Glu + Thr219GlyAla202Glu + Thr207Ser + Thr215Ser + Thr219SerGly203Glu + Thr212Asn + Asn217Gln + Thr219SerGly203Ser + Thr210Asn + Thr212Glu + Leu216MetVal204Ala + Asn211Asp + Tyr213Met + Leu216IleThr207Pro + Tyr208Asn + Tyr213Pro + Leu216GlnGly201Gln + Gly203Glu + Tyr213Met + Ala214ThrPro200Gln + Thr212Ser + Tyr213Pro + Thr215AsnPro200Asn + Val204Asn + Asn211Glu + Tyr213GlnPro209Ser + Asn211Gln + Asn217Asp + Thr219AsnPro200Gly + Gly203Asn + Tyr205Val + Tyr208HisAla202Glu + Gly203Pro + Thr212Ser + Gly218AsnAla202Gln + Val204Gly + Pro209Asp + Thr215AsnTyr208Ala + Asn211Glu + Thr215Gln + Thr219SerTyT208Gln + Tyr213Ala + Leu216Thr + Asn217AspAla199Thr + Pro200Gln + Gly201Asn + Asn211AspAla199Thr + Pro200Ser + Gly201Asn + Tyr208GlyVal204Ser + Thr207Gly + Thr212Asp + Gly218GlnPro200Asn + Ala214His + Thr215Gly + Thr219AsnThr207Ser + Tyr208Ser + Leu216Asn + Gly218AspPro200Gly + Gly203Glu + Thr212Gly + Gly218AsnPro200Ser + Gly201Ser + Tyr208Gln + Thr219AsnPro200Asn + Gly201Pro + Asn211Glu + Asn217GlnVal204Met + Thr207Asn + Asn217Gln + Thr219GlyPro200Gly + Gly201Pro + Thr210Glu + Leu216ThrPro200Asn + Gly203Pro + Val204Gln + Thr219GlnAla202Asp + Pro209Asn + Tyr213Cys + Leu216AlaPro200Gln + Gly201Ser + Ala202Glu + Tyr205GlyThr207Pro + Tyr208Asn + Asn211Ser + Tyr213SerAla202Ser + Val204Gly + Thr207Asn + Tyr213GlnGly201Ser + Gly203Gln + Thr212Ser + Leu216AspAla199Thr + Tyr205Met + Tyr208Val + Ala214ThrGly201Gln + Ala202Thr + Val204Cys + Tyr205LeuPro200Gln + Tyr205Cys + Thr210Glu + Thr212AsnPro200Gln + Tyr213Pro + Asn217Glu + Gly218AsnTyr208Pro + Tyr213His + Ala214Glu + Asn217SerPro200Ser + Ala202Gly + Tyr208Val + Thr212GlnPro200Asn + Tyr205Gly + Pro209Gln + Asn211SerTyr205Gly + Asn211Gln + Thr215Gly + Thr219GlyGly201Asn + Gly203Asn + Thr212Asp + Ala214GlyVal204Pro + Pro209Gly + Thr210Glu + Leu216ProTyr205Pro + Tyr213Ala + Gly218Asn + Thr219Pro
Ala199Gln + Val204His + Asn211Asp + Thr215GlyPro209Gln + Thr215Gln + Asn217Asp + Thr219AsnGly201Asn + Asn211Glu + Tyr213Ile + Asn217GlnGly201Ser + Thr207Asn + Tyr213Glu + Ala214GlyAla199Ser + Thr210Asn + Thr215Gln + Gly218ProVal204Asn + Tyr205Thr + Tyr208Met + Thr215ProVal204Gly + Tyr205Val + Thr210Asp + Thr215GlnAla202Ser + Val204Ala + Leu216Ser + Asn217AspAsn211Asp + Thr212Ser + Ala214Asn + Thr215GlyAla199His + Tyr208Leu + Asn217Gln + Thr219AsnAla202Pro + Tyr208Cys + Leu216Ala + Thr219AsnAla199Gly + Pro200Gly + Gly201Asn + Ala202AspPro200Ser + Ala202Gln + Thr210Gln + Leu216AspGly201Gln + Gly203Glu + Val204Gly + Leu216AsnVal204Pro + Tyr208Thr + Pro209Ser + Asn211GluTyr208Thr + Thr215Ser + Leu216Glu + Gly218GlnGly201Asn + Tyr205Ser + Tyr213Asp + Gly218ProAla202Gln + Gly203Gln + Tyr205Ile + Asn217GluThr212Ser + Tyr213Glu + Thr215Ser + Leu216CysGly203Glu + Thr212Pro + Tyr213Leu + Ala214SerPro200Ser + Gly201Gln + Ala202Asp + Thr210AsnVal204Gln + Thr210Asp + Tyr213Ser + Ala214ThrAla199Pro + Gly201Ser + Val204Gly + Asn217GlnPro200Asn + Ala202Ser + Thr207Gln + Tyr208MetPro200Gln + Gly203Pro + Ala214Glu + Gly218ProPro200Ser + Thr207Gly + Gly218Asn + Thr219GlyGly203Ser + Thr207Gln + Thr210Gln + Asn217SerGly201Ser + Thr210Glu + Asn211Glu + Thr219AsnThr207Gly + Thr210Asp + Asn211Asp + Gly218ProPro200Asn + Thr210Glu + Asn211Glu + Thr219ProVal204Ser + Thr210Asp + Asn211Glu + Thr219AsnAla199Ser + Tyr205Thr + Ala214Asp + Thr215AspPro200Asn + Thr215Asp + Leu216Asp + Thr219SerTyr208Asn + Tyr213His + Thr215Asp + Leu216AspGly201Ser + Asn211Glu + Thr212Asp + Ala214HisVal204Thr + Asn211Glu + Thr212Asp + Ala214ProPro200Gly + Thr212Gly + Asn217Glu + Gly218AspThr207Gln + Ala214Gly + Asn217Glu + Gly218GluThr207Asn + Pro209Gly + Asn217Asp + Gly218GluTyr213Thr + Leu216Val + Asn217Glu + Gly218GluAla199Thr + Ala202Asp + Gly203Glu + Gly218ProVal204His + Tyr205Asp + Thr212Gly + Thr215GluPro200Ser + TyT205Glu + Thr215Glu + Leu216CysAla202Glu + Gly203Asp + Asn217Glu + Thr21SProPro209Glu + Asn211Asp + Leu216His + Thr219ProThr207Pro + Tyr208Ser + Pro209Glu + Asn211AspVal204Thr + Pro209Asp + Asn211Asp + Thr212Pro
表26環(huán)6-五取代變異體Pro200Asn + Ala202Asn + Val204Gly + Asn211Asp + Thr219GlyPro200Gly + Gly201Pro + Pro209Glu + Asn211Ser + Thr212GlyPro200Ser + Asn211Gln + Ala214Gly + Asn217Glu + Thr219AsnGly201Asn + Gly203Ser + Thr207Ser + Tyr213Pro + Thr215GlyPro200Ser + Tyr205Pro + Tyr208Leu + Asn211Ser + Gly218AspPro200Gly + Pro209Gly + Thr212Asp + Thr215Asn + Leu216ThrGly201Gln + Ala202Pro + Thr212Asn + Ala214Gln + Thr215GluTyr205Pro + Tyr208Ser + Thr215Asp + Asn217Ser + Thr219GlnPro200Gly + Thr207Gly + Thr212Gln + Ala214Asn + Thr215AsnTyr205Ile + Thr207Pro + Tyr208Ala + Ala214Gln + Leu216AsnGly203Pro + Val204His + Thr207Gln + Tyr213Glu + Thr219SerTyr205Met + Thr207Gly + Pro209Glu + Thr215Ser + Thr219AsnPro200Gly + Thr207Gln + Asn211Asp + Thr215Pro + Gly218SerAla202His + Gly203Pro + Asn211Glu + Tyr213Gly + Thr219GlnAla199Gly + Tyr208Val + Thr210Glu + Tyr213Leu + Thr219GlnAla199His + Pro200Gln + Val204Cys + Thr215Asn + Asn217AspAla199Gly + Pro200Gln + Tyr205Ile + Thr210Gly + Thr215AspTyr208Met + Pro209Asn + Thr210Ser + Asn211Ser + Ala214GluPro200Asn + Gly201Gln + Pro209Asn + Leu216Ala + Gly218GluGly203Gln + Pro209Gly + Thr210Asn + Asn211Ser + Tyr213ValGly201Pro + Thr207Asn + Thr210Gln + Tyr213His + Ala214ThrPro200Gln + Gly201Ser + Val204Cys + Thr210Asn + Asn217GluAla199Pro + Gly203Gln + Tyr205Val + Asn211Asp + Thr215AsnAla202Asn + Val204Ser + Asn211Ser + Tyr213Asn + Leu216AspThr210Glu + Thr212Ser + Ala214Gly + Thr215Gln + Leu216IleAla199Asn + Ala202Asn + Val204Gly + Tyr205Met + Thr215AsnThr210Gln + Thr212Pro + Tyr213Thr + Leu216Gln + Asn217GlnAla199Gln + Pro200Gly + Ala202Pro + Thr212Gln + Thr215SerAla202Gln + Thr207Asn + Thr210Gln + Asn211Glu + Ala214GlyGly203Gln + Thr207Asn + Pro209Gly + Tyr213Asn + Gly218AspVal204Cys + Tyr205Thr + Thr207Asn + Leu216Thr + Thr219GlnGly201Gln + Val204His + Tyr208Ser + Thr212Ser + Thr215GlyPro200Gln + Ala202Asp + Tyr208Gly + Thr212Ser + Leu216ThrAla199Ser + Gly201Ser + Asn211Asp + Ala214Thr + Thr219GlyAla202Gln + Val204Ser + Tyr208Leu + Pro209Asp + Tyr213GlnAla199Ser + Tyr205Ala + Thr210Glu + Thr212Gly + Leu216AlaAla199His + Ala202Glu + Thr210Gly + Asn211Gln + Tyr213ProAla199Ser + Gly203Glu + Tyr213Met + Leu216Ala + Gly218SerGly201Gln + Gly203Ser + Thr207Asn + Gly218Pro + Thr219ProAla199Gly + Pro209Asp + Thr212Ser + Leu216Met + Gly218SerPro200Ser + Tyr205Ala + Ash211Ser + Thr212Glu + Thr215ProPro209Glu + Thr212Gly + Ala214Thr + Leu216Asn + Thr219ProGly203Asn + Tyr205Gln + Pro209Asn + Gly218Asn + Thr219Ser
Gly201Asn + Ala202Gln + Gly203Pro + Thr210Gly + Asn217SerPro200Ser + Gly201Pro + Tyr208Val + Thr210Gly + Thr212AsnAla199His + Gly201Gln + Tyr205Glu + Leu216Met + Gly218AsnTyr205Ser + Asn211Gln + Thr212Asp + Thr215Ser + Thr219GlyAla199Thr + Gly201Gln + Ala202Thr + Gly203Glu + Tyr213ProAla199Gln + Val204Asn + Thr207Asn + Tyr208Gly + Gly218SerAla199His + Gly203Pro + Thr207Gln + Asn211Asp + Ala214AsnAla202Ser + Gly203Pro + Tyr205Ser + Thr210Glu + Leu216AsnThr207Gly + Tyr208Gly + Thr212Asp + Leu216Pro + Thr219AsnAla202Asp + Val204Tnr + Tyr208Ile + Thr212Asn + Gly218SerGly203Gln + Val204Ala + Tyr205Pro + Asn211Gln + Thr215GlnPro200Ser + Gly203Pro + Thr212Pro + Leu216Met + Gly218GluAla199Gly + Gly201Gln + Thr207Ser + Leu216Cys + Asn217GluAla199Thr + Tyr205Met + Tyr208Val + Ala214Thr + Leu216IleAla202Asp + Tyr205Pro + Thr207Ser + Tyr208Thr + Thr210ProAla199Gln + Pro200Gln + Thr207Asn + Leu216Val + Gly218SerGly201Ser + Tyr208Met + Asn211Asp + Thr212Ser + Thr219GlyGly203Ser + Thr207Pro + Asn211Ser + Asn217Asp + Gly218GlnGly201Pro + Gly203Asp + Tyr208Asn + Thr212Ser + Thr215AsnAla199Pro + Pro200Ser + Tyr205Asp + Thr212Pro + Leu216ValTyr208Ala + Tyr213Ile + Ala214Gly + Thr215Pro + Asn217GlnAla199Ser + Pro200Gly + Gly203Asp + Leu216His + Asn217SerGly201Pro + Gly203Ser + Val204Asn + Leu216Ala + Asn217AspVal204Gln + Tyr208His + Asn211Asp + Tyr213His + Leu216ThrPro200Gly + Gly201Asn + Val204Gln + Tyr208Cys + Thr210ProAla202Asn + Tyr205Gln + Pro209Glu + Thr210Gly + Gly218AsnPro200Gly + Thr207Pro + Tyr208Met + Leu216Val + Asn217GluGly201Asn + Tyr205Gln + Thr212Asp + Thr215Pro + Asn217GlnGly203Glu + Tyr208Ile + Asn211Gln + Thr215Ser + Thr219GlyGly201Asn + Ala202Ser + Gly203Gln + Asn211Gln + Ala214AsnPro200Gly + Val204Asn + Pro209Gln + Thr210Pro + Thr219SerTyr208Leu + Pro209Asn + Thr210Gln + Thr215Asn + Gly218AsnAla202Ser + Val204Cys + Thr210Asn + Ala214Glu + Thr215GlyPro200Asn + Gly203Gln + Tyr208Ser + Pro209Gln + Gly218ProAla202Thr + Gly203Asp + Tyr205Leu + Thr212Gly + Tyr213GlnGly203Gln + Val204Pro + Thr210Asp + Gly218Gln + Thr219GlyTyr208His + Thr210Ser + Asn211Asp + Leu216Asn + Asn217GlnGly201Gln + Tyr205His + Thr210Glu + Tyr213lle + Leu216AlaPro200Asn + Gly201Ser + Ala202Glu + Gly203Gln + Thr219SerAla199Gln + Gly201Asn + Ala202Asp + Thr212Asn + Tyr213ThrAla199Pro + Val204Ser + Tyr213His + Asn217Asp + Gly218GlnPro200Asn + Gly201Ser + Ala202Pro + Thr207Ser + Ala214ThrAla199Asn + Gly201Pro + Gly203Asp + Tyr213Cys + Gly218GlnGly201Gln + Gly203Asn + Val204Thr + Thr210Ser + Thr212GlnGly201Asn + Ala202Pro + Thr210Gln + Asn211Gln + Thr215GlnPro200Asn + Gly203Glu + Tyr205Val + Thr207Pro + Tyr208SerAla199Gln + Gly201Ser + Ala202Pro + Thr207Pro + Ash217Glu
Pro200Gln + Tyr205Met + Thr212Pro + Leu216Glu + Thr219SerAla199Pro + Pro200Gln + Ala202Gly + Thr207Gln + Asn217GluTyr208Thr + Asn211Ser + Tyr213Gln + Leu216Gly + Thr219ProGly203Ser + Val204Thr + Tyr208Ala + Tyr213Pro + Thr219AsnGly201Asn + Gly203Pro + Val204Ala + Asn211Ser + Thr219GlyGly203Glu + Thr207Ser + Tyr208Ser + Thr215Asn + Leu216CysGly201Asn + Tyr208Leu + Asn211Glu + Leu216Gly + Asn217SerGly201Pro + Tyr208Val + Thr210Pro + Leu216Gln + Gly218AsnPro200Asn + Ala202Thr + Thr207Ser + Thr212Ser + Ala214AspGly203Glu + Val204Asn + Thr210Pro + Ala214His + Gly218AsnGly201Ser + Val204Met + Ala214Gly + Leu216Ser + Thr219SerPro200Gly + Tyr208Val + Tyr213His + Leu216Ala + Asn217GluThr212Glu + Tyr213Asn + Ala214Thr + Leu216Gln + Gly218ProGly203Pro + Val204Asn + Tyr208Cys + Asn217Glu + Thr219SerPro200Asn + Gly201Pro + Val204Thr + Leu216Asp + Asn217GlnTyr205Met + Tyr208Asn + Pro209Asn + Tyr213Ile + Ala214HisGly201Gln + Gly203Pro + Tyr205Cys + Thr210Gln +Asn217AspAla202Asn + Gly203Asp + Thr207Gly + Asn211Ser + Thr219ProPro200Gly + Gly201Ser + Val204Cys + Thr207Gly + Gly218AspPro200Ser + Gly203Pro + Tyr208Pro + Thr210Asn + Asn211GluPro200Gly + Gly201Gln + Tyr205Leu + Pro209Asn + Ala214HisTyr205Ser + Thr207Gln + Tyr208Ser + Thr212Ser + Thr219AsnAla202Gly + Asn211Asp + Thr212Gln + Ala214Thr + Leu216HisAla202Pro + Val204Ala + Pro209Gln + Asn211Ser + Thr212GluAla202Gly + Thr207Pro + Pro209Gly + Asn217Ser + Gly218AspGly201Gln + Pro209Asn + Thr215Ser + Leu216Thr + Thr219GlyVal204Thr + Tyr208Val + Thr215Asn + Asn217Glu + Gly218AsnTyr205Gln + Tyr213Pro + Thr215Ser + Leu216Gly + Gly218GlnGly203Asn + Thr207Ser + Thr210Asn + Asn211Asp + Leu216MetTyr208His + Pro209Ser + Leu216Gly + Asn217Glu + Gly218GlnThr207Ser + Pro209Gly + Asn211Glu + Thr215Ser + Leu216IleAla199Gly + Ala202Glu + Thr207Pro + Tyr213Val + Ala214GlyAla199His + Pro200Gly + Gly201Ser + Gly218Asp + Thr219ProAla202Glu + Tyr208Ala + Thr212Ser + Tyr213His + Thr219AsnAla199Thr + Val204Thr + Thr207Asn + Thr210Asn + Tyr213GluTyr208Gln + Pro209Gly + Ala214Asn + Thr215Glu + Gly218AsnTyr208Thr + Tyr213Val + Thr215Asp + Leu216Val + Gly218GlnThr207Pro + Pro209Ser + Thr210Glu + Asn217Ser + Thr219ProPro200Ser + Gly201Gln + Tyr205Ile + Pro209Gly + Thr219GlnVal204Met + Thr207Pro + Tyr208Gln + Pro209Asn + Thr215AspVal204Ala + Pro209Asn + Thr210Gly + Tyr213Asp + Ala214ProPro200Ser + Tyr208His + Tyr213Glu + Leu216Cys + Thr219SerGly201Gln + Ala202Asp + Tyr205Ala + Leu216Thr + Asn217GlnAla199Gly + Gly201Asn + Val204Gly + Ala214Asn + Thr219ProPro200Asn + Ala202Asn + Gly203Gln + Tyr208Asn + Asn217AspVal204Gly + Tyr205Asp + Asn211Gln + Tyr213Met + Ala214GlyGly201Asn + Thr207Gln + Pro209Gln + Ala214Asp + Gly218Pro
Pro200Gly + Tyr208Ser + Asn211Asp + Tyr213Ser + Thr219GlyAla202Thr + Val204Pro + Thr210Ser + Asn211Asp + Leu216ThrGly203Glu + Val204Met + Tyr205His + Asn211Gln + Thr219GlnGly201Gln + Ala202Gly + Val204Ser + Thr207Gln + Asn211GluGly203Asn + Tyr208Ala + Pro209Asp + Thr212Asn + Thr215GlnAla199Gln + Pro200Asn + Tyr205Met + Asn211Asp + Thr219GlnVal204Gln + Tyr208Val + Thr210Pro + Asn211Gln + Leu216AspPro200Asn + Val204Pro + Tyr205Gly + Thr210Pro + Ala214GlyGly201Asn + Tyr205Met + Tyr213Cys + Leu216Ile + Thr219AsnAla199Gly + Gly201Asn + Gly203Gln + Val204Thr + Pro209GluAla199His + Ala202Gly + Pro209Asp + Thr210Asp + Gly218SerGly201Gln + Tyr208Ser + Pro209Asp + Thr210Glu + Gly218SerAla199Asn + Pro200Ser + Thr212Asn + Ala214Asp + Thr215GluAla199His + Pro200Ser + Thr215Gly + Asn217Asp + Gly218GluTyr205Thr + Thr207Ser + Asn217Glu + Gly218Asp + Thr219GlyAla202Thr + Tyr205Glu + Tyr208Thr + Thr215Glu + Leu216GlnTyr205Asp + Thr212Gly + Thr215Glu + Asn217Ser + Gly218SerPro200Gly + Ala202Glu + Val204Ala + Tyr208Leu + Asn217AspPro200Gln + Ala202Asp + Tyr205Gly + Tyr213Ile + Asn217AspPro209Asp + Asn211Asp + Tyr213Thr + Leu216Cys + Thr219AsnAla202Asn + Val204His + Pro209Glu + Asn211Glu + Thr219AsnPro200Ser + Ala202Asp + Val204Gln + Leu216Glu + Asn217GluAla199Gly + Pro200Gln + Ala202Gln + Thr210Glu + Thr212AspGly203Asp + Pro209Gly + Tyr213His + Asn217Asp + Gly218SerGly203Asp + Val204Thr + Tyr208Ala + Tyr213Gln + Thr215AspTyr205Pro + Thr207Gly + Pro209Glu + Thr212Asp + Gly218SerAla199His + Ala202Glu + Gly203Gln + Asn211Gln + Gly218AspAla199Asn + Ala202Glu + Thr207Gln + Thr212Gln + Gly218AspAla202Asp + Gly203Asn + Tyr213Val + Leu216Ile + Gly218GluPro200Gly + Ala202Thr + Tyr208Met + Ala214Glu + Leu216AspGly203Glu + Thr207Gln + Thr212Ser + Ala214Glu + Thr215GluVal204His + Tyr205Asp + Tyr208Met + Tyr213Glu + Asn217GlnTyr205Gly + Tyr213Ile + Leu216Glu + Gly218Glu + Thr219GlyVal204Ser + Asn211Gln + Tyr213Thr + Thr215Glu + Asn217GluTyr208Leu + Thr210Gly + Thr212Gln + Tyr213Glu + Thr215GluAla202Glu + Tyr205Glu + Thr210Gly + Asn211Gln + Leu216AspPro200Asn + Thr210Ser + Thr212Asp + Tyr213Glu + Thr215AspAla199Thr + Thr207Ser + Thr210Asp + Thr212Asn + Tyr213AspAla202Glu + Val204Pro + Tyr205Thr + Tyr208His + Thr215GluGly203Glu + Tyr208Ile + Thr210Gln + Tyr213Gly + Gly218AspTyr208Gln + Pro209Glu + Tyr213Thr + Ala214Glu + Thr219GlnTyr205Asp + Thr210Ser + Thr215Gln + Asn217Asp + Gly218GluGly201Asn + Tyr205Glu + Tyr208Leu + Asn217Asp + Gly218Glu表27環(huán)6-六取代變異體Pro200Gly + Gly203Asp + Thr210Gln + Thr212Asn + Ala214His +Leu216AsnPro200Gly + Thr207Asn + Thr210Gln + Thr212Pro + Thr215Asp +Gly218ProPro200Gln + Gly203Ser + Thr207Asn + Tyr208Ile + Pro209Ser +Thr210GluGly203Pro + Tyr205Gly + Tyr208Gly + Asn211Glu + Tyr213Pro +Gly218GlnAla199His + Gly201Gln + Gly203Gln + Thr212Pro + Leu216Ile +Gly218AspVal204Met + Tyr205Thr + Thr207Ser + Tyr213His + Ala214Glu +Thr219AsnAla199Thr + Gly201Ser + Gly203Asp + Thr207Pro + Thr210Gly +Thr212ProPro200Gly + Tyr208Ile + Asn211Glu + Tyr213Ser + Ala214Ser +Thr215GlnAla199Thr + Gly201Gln + Pro209Gln + Thr212Asn + Tyr213His +Thr219GlyAla199Gly + Pro200Gly + Gly201Pro + Val204Cys + Thr207Gly +Thr215GlnAla199Gly + Pro200Ser + Thr207Pro + Leu216Ala + Asn217Glu +Gly218AsnPro200Gln + Val204Met + Asn211Gln + Ala214Gln + Thr215Gln +Leu216AspAla199Ser + Gly203Asn + Thr207Gln + Pro209Asp + Thr215Gly +Leu216HisGly201Pro + Tyr208Ile + Thr210Glu + Ala214Gly + Leu216Val +Thr219SerAla199Gln + Ala202Asn + Pro209Asn + Tyr213Asn + Thr215Gln +Gly218SerPro200Gln + Gly203Pro + Val204Gly + Thr212Pro + Ala214Pro +Thr215GluVal204Met + Tyr208Asn + Tyr213Asp + Ala214His + Thr215Gln +Leu216ThrAla199Gly + Pro200Asn + Ala202Thr + Val204Ser + Gly218Glu +Thr219ProAla199Ser + Gly201Pro + Gly203Asn + Thr207Ser + Thr212Gly +Thr219GlnAla199His + Tyr205Asn + Tyr208Ala + Asn211Gln + Tyr213Asn +Asn217AspTyr205Asp + Thr207Gly + Pro209Asn + Thr210Pro + Tyr213Ser +Leu216SerGly201Gln + Ala202His + Thr207Gly + Pro209Glu + Thr210Ser +Ala214GlyPro200Asn + Val204Gln + Tyr205Cys + Thr207Gln + Thr212Gln +Thr219SerPro200Ser + Tyr205Asp + Thr210Ser + Thr215Gln + Leu216Asn +Asn217GlnAla202His + Tyr205Thr + Tyr208His + Thr212Asp + Tyr213Ile +Gly218GlnGly201Asn + Pro209Glu + Asn211Gln + Thr212Ser + Tyr213Gln +Thr219GlnAla199Pro + Pro200Asn + Tyr205Val + Thr210Glu + Leu216Gly +Gly218AsnAla199Gln + Pro200Gln + Val204Thr + Tyr208Thr + Leu216Met +Asn217AspVal204Met + Thr210Ser + Thr212Asn + Tyr213Cys + Thr215Asn +Thr219ProAla199Gln + Gly201Pro + Ala202His + Thr207Asn + Leu216Val +Gly218SerAla199Thr + Gly203Ser + Tyr205Val + Tyr208Val + Thr210Pro +Ala214GluAla199Asn + Pro200Gln + Tyr205Met + Tyr208Cys + Thr212Glu +Ala214GlnAla202Ser + Thr207Gln + Thr210Glu + Asn211Gln + Tyr213Ile +Leu216AlaAla199Gly + Pro200Gly + Thr207Pro + Tyr213Ser + Leu216Asn +Asn217SerGly201Ser + Ala202Gly + Tyr205Asn + Pro209Gly + Thr210Asn +Asn211GlnGly201Ser + Ala202Gly + Gly203Asp + Thr207Asn + Asn211Gln +Thr215GlyVal204Gln + Tyr205His + Thr207Gln + Tyr208His + Asn211Asp +Leu216ThrAla202Glu + Val204Ser + Tyr205His + Tyr208Ile + Ala214Gln +Leu216GlnAla199Gly + Gly203Glu + Tyr205Val + Tyr213His + Ala214Asn +Asn217SerPro200Ser + Gly201Ser + Tyr205Asn + Thr210Ser + Thr212Glu +Gly218AsnAla199Thr + Gly201Gln + Pro209Ser + Thr210Asp + Thr215Gly +Leu216SerGly201Pro + Tyr205Thr + Thr207Ser + Tyr213His + Gly218Asp +Thr219GlyGly203Glu + Tyr208Ile + Asn211Gln + Ala214Gln + Thr215Ser +Thr219GlyAla199Gly + Gly203Gln + Thr207Gln + Asn211Asp + Thr215Pro +Thr219GlnAla199Pro + Thr207Gly + Pro209Glu + Thr210Ser + Thr212Gly +Leu216ValVal204Asn + Thr207Pro + Asn211Gln + Ala214His + Asn217Glu +Gly218GlnAla202Glu + Gly203Ser + Val204Cys + Thr210Gly + Asn211Gln +Ala214ProAla199Asn + Ala202Gln + Gly203Asn + Thr207Pro + Thr212Pro +Thr215ProAla199Thr + Gly203Gln + Thr207Asn + Tyr208Asn + Tyr213Glu +Thr215GlyPro200Ser + Ala202Asn + Val204Ser + Tyr205Ile + Thr207Gln +Asn211AspPro200Gln + Val204Pro + Tyr208Pro + Pro209Asn + Leu216Glu +Thr219SerPro200Gly + Ala202Gln + Tyr205Thr + Thr210Gln + Asn211Gln +Ala214AspGly203Asn + Pro209Ser + Thr210Glu + Ala214Pro + Gly218Asn +Thr219AsnPro200Ser + Gly201Pro + Ala202Thr + Ala214Gly + Leu216Asp +Thr219SerAla202Ser + Tyr205His + Thr212Gln + Thr215Glu + Asn217Gln +Thr219SerAla202Gly + Thr207Asn + Pro209Gly + Thr210Pro + Tyr213Met +Thr215GlyPro200Gln + Val204Ser + Thr207Ser + Tyr208Cys + Thr212Asn +Gly218GlnAla199Pro + Pro200Gly + Ala202Thr + Thr207Gly + Thr212Glu +Thr219SerAla199His + Gly203Gln + Tyr205Gly + Thr207Ser + Thr210Asn +Thr212SerGly203Glu + Val204Asn + Tyr205Pro + Thr210Pro + Ala214His +Gly218AsnPro200Gln + Val204Ala + Tyr208Cys + Ala214Gln + Leu216Val +Asn217GluGly201Gln + Ala202Asp + Tyr205Gly + Thr210Gly + Leu216Val +Gly218SerPro200Asn + Gly201Ser + Thr207Gln + Thr212Asp + Tyr213Gly +Thr215ProAla202Asn + Gly203Asp + Val204Cys + Thr207Gly + Asn211Ser +Thr219ProAla199Ser + Gly201Ser + Tyr205Ser + Thr207Asn + Tyr213Val +Leu216AlaPro200Gln + Tyr205Ser + Thr207Gln + Tyr208Ser + Thr212Ser +Thr219AsnPro200Asn + Ala202Gly + Asn211Asp + Thr212Gln + Ala214Thr +Leu216HisGly201Pro + Gly203Glu + Val204Met + Pro209Gly + Asn217Ser +Thr219SerPro200Gly + Gly201Pro + Tyr208Asn + Thr212Ser + Tyr213Leu +Ala214AsnAla199Ser + Gly201Pro + Gly203Asn + Asn211Asp + Leu216Cys +Thr219AsnGly203Pro + Val204Gly + Thr207Gln + Leu216Gly + Asn217Ser +Gly218AsnVal204Gln + Thr207Gln + Thr210Gly + Asn211Glu + Ala214Gly +Thr219ProAla202His + Tyr208Asn + Thr210Asn + Leu216Asn + Gly218Pro +Thr219GlyGly201Pro + Gly203Asp + Tyr205Met + Thr207Ser + Tyr213Gln +Asn217GlnGly201Ser + Ala202Ser + Thr207Asn + Tyr208Gly + Asn211Ser +Leu216ProAla202Gln + Gly203Asn + Val204Met + Tyr208Ser + Thr212Gln +Leu216AspAla199Asn + Ala202Thr + Pro209Glu + Thr210Pro + Asn217Ser +Thr219GlyGly203Asn + Tyr208Ser + Asn211Glu + Ala214His + Leu216Met +Gly218ProAla199Ser + Pro200Gln + Val204Asn + Tyr208Asn + Thr215Asp +Thr219GlnGly201Asn + Thr207Asn + Tyr208Gln + Pro209Asn + Asn211Asp +Leu216CysPro200Ser + Gly203Asn + Thr207Gln + Thr212Gln + Asn217Asp +Gly218AsnVal204His + Tyr205Ala + Thr210Asn + Thr215Pro + Leu216Val +Asn217GluAla199Pro + Gly201Asn + Ala202Asp + Val204Thr + Thr210Gly +Thr219SerPro200Asn + Ala202Gly + Gly203Glu + Thr207Gln + Tyr208His +Asn211GlnPro200Gly + Ala202Asn + Gly203Asn + Thr212Asp + Ala214Ser +Leu216ProAla199Gly + Gly201Pro + Thr207Gly + Pro209Asp + Thr215Asn +Gly218SerGly203Asn + Tyr205Thr + Asn211Asp + Ala214Gly + Gly218Asn +Thr219GlyGly201Pro + Val204His + Tyr208Cys + Thr215Glu + Leu216Asn +Gly218ProVal204Ala + Pro209Asn + Thr210Asp + Tyr213Leu + Leu216Ser +Thr219GlyAla202Asp + Tyr205Val + Thr207Asn + Thr210Asn + Tyr213Pro +Ala214AsnGly201Ser + Thr207Gly + Thr212Pro + Thr215Ser + Leu216Asn +Thr219GlyAla199His + Gly203Gln + Tyr205Gln + Tyr208His + Ala214Gln +Thr215AsnVal204Asn + Tyr205Ile + Pro209Glu + Thr210Ser + Tyr213Val +Leu216SerAla199Ser + Pro200Ser + Gly201Asn + Val204Thr + Thr207Pro +Gly218GluAla199Gly + Pro200Gly + Pro209Gln + Thr215Asp + Leu216Cys +Thr219GlyGly201Ser + Gly203Ser + Tyr205Cys + Thr207Gln + Thr212Gly +Ala214ThrPro200Asn + Ala202Gln + Pro209Gly + Thr210Asp + Thr215Pro +Gly218AsnTyr205Pro + Thr207Gln + Thr210Glu + Tyr213Cys + Leu216Pro +Thr219SerGly201Gln + Tyr205Val + Tyr208Val + Thr210Gln + Thr215Glu +Gly218AsnAla199Ser + Tyr208Asn + Asn211Gln + Leu216Ile + Asn217Glu +Gly218ProPro200Asn + Gly201Pro + Val204Thr + Tyr205His + Thr215Glu +Leu216ThrGly203Pro + Val204His + Prn209Gly + Thr210Glu + Thr215Pro +Gly218AsnTyr205Leu + Asn211Gln + Ala214Gln + Leu216Gly + Asn217Asp +Gly218AsnAla199Ser + Gly201Gln + Gly203Pro + Tyr208Thr + Thr215Gly +Thr219AsnPro209Glu + Thr210Ser + Thr212Ser + Tyr213Pro + Ala214Thr +Thr215ProPro200Ser + Ala202Pro + Gly203Asn + Thr207Pro + Thr210Asp +Tyr213ValGly201Asn + Thr207Gly + Pro209Ser + Tyr213Met + Leu216Ile +Gly218AspGly201Pro + Val204His + Thr212Gly + Leu216Asn + Asn217Ser +Thr219GlyThr212Gln + Ala214Glu + Thr215Ser + Asn217Ser + Gly218Ser +Thr219AsnGly203Ser + Tyr205Met + Tyr208Thr + Thr210Asn + Ala214Glu +Leu216IleGly201Gln + Tyr208Gln + Thr210Gly + Thr212Gln + Asn217Ser +Thr219AsnGly201Gln + Tyr205Cys + Thr212Pro + Tyr213Met + Leu216Asn +Gly218GlnAla202Gln + Tyr205Val + Pro209Gln + Thr210Asp + Asn211Ser +Tnr215GlnGly201Pro + Val204Pro + Thr207Asn + Asn211Gln + Asn217Glu +Thr219AsnAla199Ser + Tyr213Ser + Ala214Thr + Leu216Ser + Asn217Ser +Gly218GluPro200Gln + Gly201Ser + Val204Ala + Thr207Gln + Thr210Asn +Ala214GluTyr205Gln + Tyr208Cys + Thr212Gln + Leu216Ser + Gly218Gln +Thr219SerPro200Gly + Gly201Pro + Gly203Pro + Thr207Gln + Thr210Gly +Thr219ProAla199Gln + Pro200Gly + Val204Ser + Thr215Glu + Asn217Ser +Thr219ProAla199Ser + Pro200Gln + Gly201Pro + Val204Pro + Thr210Pro +Thr219GlnAla202Gln + Thr207Gln + Thr210Glu + Ala214Ser + Thr215Pro +Gly218GlnPro200Gln + Ala202Glu + Thr207Pro + Pro209Asn + Thr212Gln +Leu216ValGly201Asn + Asn211Glu + Tyr213Cys + Ala214Thr + Thr215Ser +Leu216HisAla199His + Ala202Gly + Tyr205Ala + Thr207Gly + Asn211Asp +Ala214GlnGly203Ser + Thr207Asn + Pro209Glu + Thr210Asp + Asn217Gln +Thr219GlyGly201Gln + Val204Met + Thr210Asp + Asn211Asp + Leu216Gln +Gly218SerGly203Pro + Tyr205Ile + Tyr208Gln + Asn211Asp + Thr212Glu +Gly218GlnGly203Ser + Tyr205Val + Thr207Asn + Leu216Glu + Asn217Asp +Thr219SerVal204Ala + Tyr205Pro + Pro209Ser + Leu216Ala + Asn217Asp +Gly218GluPro200Asn + Gly201Ser + Tyr208Thr + Thr215Gln + Asn217Asp +Gly218AspAla202Asn + Tyr208Met + Thr210Gln + Asn211Gln + Asn217Asp +Gly218AspAla199Gln + Ala202Asp + Gly203Glu + Thr207Gln + Tyr208Asn +Asn211GlnAla202Asp + Gly203Asp + Thr207Ser + Tyr208Cys + Thr210Asn +Thr215ProAla202Glu + Gly203Asp + Thr207Gly + Tyr208Leu + Leu216Val +Thr219GlnPro200Ser + Gly201Asn + Pro209Asp + Thr210Asp + Asn211Asp +Leu216SerAla199Gln + Val204Gln + Tyr205Asp + Tyr208Leu + Thr210Gln +Thr215GluPro200Gln + Gly203Gln + Tyr205Glu + Thr212Gly + Thr215Glu +Thr219ProPro200Asn + Thr207Ser + Thr210Asp + Asn211Asp + Thr212Glu +Leu216MetAla202Asp + Gly203Asp + Val204Ser + Thr210Ser + Asn217Asp +Gly218ProVal204Ser + Thr207Pro + Tyr213His + Thr215Glu + Leu216Asp +Asn217GluVal204Gly + Asn211Ser + Ala214Gln + Thr215Glu + Leu216Glu +Asn217GluTyr208Ile + Tyr213Ser + Ala214His + Leu216Glu + Asn217Asp +Gly218AspGly203Asp + Thr207Ser + Ala214His + Thr215Asp + Leu216Asp +Thr219SerAla199Ser + Gly201Asn + Thr207Ser + Pro209Glu + Asn211Asp +Thr219SerGly203Glu + Tyr205Glu + Thr210Gly + Tyr213Asn + Ala214Gln +Thr215AspAla199His + Gly203Asp + Val204His + Thr212Ser + Asn217Asp +Gly218ProTyr205Glu + Thr210Gly + Asn211Ser + Ala214Asp + Thr215Ser +Asn217GlnGly201Asn + Tyr205Asp + Asn211Gln + Thr212Ser + Ala214Glu +Gly218SerAla199Asn + Gly201Asn + Gly203Asp + Tyr208Thr + Thr215Asp +Leu216ThrPro200Gly + Tyr208Pro + Pro209Glu + Thr212Asp + Gly218Ser +Thr219SerPro200Asn + Val204Gly + Thr207Asn + Pro209Asp + Thr212Glu +Tyr213AlaAla199Pro + Gly203Asp + Tyr205Glu + Pro209Ser + Ala214His +Thr215AsnPro200Gly + Gly203Glu + Thr207Ser + Thr215Gln + Asn217Asp +Gly218AspAla202Glu + Gly203Glu + Tyr208Gln + Tyr213Thr + Ala214Ser +Gly218AspPro200Gln + Gly203Asp + Tyr213Asn + Ala214Asp + Thr215Asp +Leu216IleAla202Ser + Gly203Ser + Tyr205Glu + Pro209Gly + Thr215Asp +Asn217GluAla202Gln + Thr207Pro + Thr212Asp + Tyr213Pro + Ala214Glu +Thr219GlyPro200Ser + Val204Pro + Thr212Asp + Tyr213Ala + Ala214Asp +Thr219AsnAla199Ser + Ala202Asp + Val204Pro + Pro209Ser + Thr215Asn +Leu216AspAla199Gly + Ala202Asn + Val204Asn + Tyr213Asp + Ala214Glu +Leu216GluAla199Thr + Gly203Gln + Thr210Gln + Thr212Gln + Tyr213Glu +Thr215GluPro200Gln + Gly201Pro + Pro209Asp + Tyr213Glu + Thr215Pro +Gly218AsnVal204Asn + Tyr205Val + Pro209Asp + Thr212Pro + Tyr213Asp +Ala214GlyTyr205Leu + Asn211Glu + Thr212Glu + Tyr213Leu + Ala214Glu +Thr215GlyAla199Ser + Ala202Glu + Tyr213Pro + Ala214Gly + Thr215Asp +Gly218ProPro200Gly + Gly203Asp + Val204His + Thr210Asn + Asn211Gln +Gly218AspGly201Ser + Ala202Gly + Gly203Asp + Ala214Asn + Gly218Glu +Thr219GlyAla199Gln + Pro200Ser + Ala202Ser + Thr210Asp + Asn211Glu +Ala214GluAla202Glu + Thr207Ser + Asn211Gln + Ala214Asp + Leu216Glu +Gly218SerAla199Gln + Pro200Ser + Tyr205Asp + Pro209Ser + Thr215Gly +Asn217GluAla199Ser + Tyr205Asp + Tyr213Met + Ala214Thr + Asn217Asp +Thr219AsnPro200Gln + Ala202Asn + Gly203Gln + Tyr213Glu + Leu216Glu +Thr219AsnAla199Pro + Ala202Gln + Gly203Asn + Thr212Glu + Thr215Asp +Leu216AspAla199Gln + Gly201Gln + Val204Thr + Asn211Asp + Ala214Asp +Thr215AspGly203Gln + Val204Met + Asn211Glu + Thr212Ser + Ala214Asp +Thr219GlyThr207Gly + Tyr208Val + Thr210Gly + Thr215Glu + Leu216Asn +Gly218AspPro200Gly + Val204Asn + Thr207Gly + Thr210Glu + Tyr213Val +Ala214AspGly203Glu + Thr207Asn + Thr210Gly + Thr212Asp + Thr215Asp +Leu216GlnGly203Gln + Val204Cys + Pro209Asn + Thr210Ser + Thr212Asp +Thr215AspGly203Asn + Val204Gly + Tyr205Asp + Ala214Glu + Thr215Pro +Gly218Asp表28環(huán)6-七取代突變變異體Pro200Asn + Ala202Gln + Tyr208Cys + Thr210Pro + Tyr213Asn +Ala214Asn + Thr219SerGly203Pro + Val204Gln + Tyr205Gly + Tyr208Gly + Asn211Glu +Tyr213Pro + Gly218GlnGly201Asn + Gly203Gln + Val204Cys + Thr207Ser + Thr212Asp +Thr215Pro + Thr219GlyGly203Gln + Val204Gln + Tyr205Pro + Thr212Gly + Tyr213Val +Asn217Asp + Thr219GlyGly201Gln + Ala202Gln + Val204Pro + Asn211Asp + Tyr213Ile +Leu216Gly + Gly218ProPro200Gly + Ala202Gln + Val204Pro + Pro209Asn + Tyr213Ala +Asn217Asp + Thr219ProAla199Thr + Gly201Ser + Val204Ser + Thr210Gly + Asn211Asp +Gly218Ser + Thr219SerGly201Gln + Tyr208Val + Pro209Gln + Asn211Ser + Thr215Pro +Asn217Gln + Thr219GlnAla199His + Pro200Gly + Pro209Asn + Thr212Gln + Tyr213Ile +Leu216Gln + Thr219GlyPro200Gln + Thr210Pro + Asn211Ser + Thr212Gln + Leu216Gln +Asn217Asp + Thr219ProPro200Ser + Val204His + Tyr205Ala + Tyr208Ser + Asn211Gln +Ala214His + Thr215AsnAla199Ser + Pro200Ser + Gly201Pro + Tyr205Met + Thr207Pro +Thr210Asp + Thr212SerGly201Gln + Val204Ser + Tyr205Asn + Tyr213Ala + Ala214Glu +Thr215Gln + Leu216ThrAla199Gln + Val204Thr + Thr207Ser + Tyr208Asn + Pro209Asn +Thr212Gln + Gly218ProPro200Asn + Gly203Pro + Thr207Pro + Thr210Gln + Tyr213His +Leu216Ile + Gly218AspAla202Gly + Gly203Gln + Val204Gly + Tyr205Ser + Thr212Gln +Asn217Ser + Thr219GlyPro209Gly + Thr210Gln + Asn211Asp + Tyr213Gln + Ala214Ser +Leu216Asn + Asn217SerAla199His + Val204Pro + Tyr205Glu + Pro209Asn + Tyr213Ala +Leu216Thr + Gly218AsnAla199Gln + Pro200Asn + Ala202Thr + Gly203Pro + Val204Gln +Thr212Ser + Tyr213GlnGly201Asn + Ala202Asn + Gly203Asp + Tyr208Thr + Thr212Gln +Tyr213Thr + Gly218AsnAla199Thr + Gly203Asp + Val204Thr + Thr207Asn + Thr210Asn +Asn211Ser + Tyr213ValPro200Gly + Thr207Pro + Tyr208Met + Pro209Glu + Tyr213His +Thr215Pro + Leu216GlnPro200Ser + Gly201Asn + Ala202Asp + Tyr205Cys + Tyr208Ala +Tyr213His + Thr219AsnAla199Pro + Val204Met + Thr207Gln + Tyr208Val + Asn211Glu +Ala214Ser + Thr219SerGly201Asn + Tyr205Leu + Tyr208Pro + Thr210Pro + Thr212Asn +Tyr213Leu + Ala214AsnPro200Ser + Gly201Pro + Gly203Pro + Val204Asn + Thr207Asn +Ala214Asp + Leu216HisPro200Gly + Gly201Gln + Val204Gln + Thr207Gln + Tyr213His +Ala214Ser + Gly218ProPro200Gln + Val204His + Pro209Ser + Asn211Ser + Thr212Glu +Tyr213Gly + Ala214ProAla199His + Gly201Gln + Ala202Asn + Thr210Asn + Asn211Ser +Ala214Asn + Thr219GlnPro200Gly + Gly201Asn + Val204Gln + Thr212Gly + Thr215Pro +Asn217Ser + Thr219ProAla199Thr + Val204Thr + Tyr205Gly + Tyr208Met + Thr210Asn +Tyr213His + Leu216CysPro200Asn + Ala202Asp + Val204Pro + Tyr205Gly + Thr207Asn +Tyr208Cys + Thr210GlnThr207Ser + Thr210Ser + Thr212Gly + Ala214Thr + Thr215Asn +Asn217Asp + Thr219GlnAla199Ser + Pro200Ser + Gly203Gln + Thr207Ser + Tyr208Ile +Asn211Glu + Leu216MetAla199Gly + Pro200Asn + Gly201Asn + Gly203Ser + Ala214His +Thr215Asp + Leu216ProPro200Gln + Gly201Ser + Ala202Asp + Thr207Asn + Tyr208Pro +Thr215Gln + Leu216CysVal204Thr + Tyr205Pro + Tyr208Thr + Asn211Gln + Thr212Asp +Tyr213Cys + Thr219AsnPro200Gln + Ala202Gly + Gly203Asn + Tyr205Met + Tyr208Val +Tyr213Val + Thr219AsnAla199Gly + Pro200Asn + Ala202Pro + Tyr205Asp + Thr207Gln +Thr212Gln + Leu216CysGly201Pro + Ala202Gly + Val204Thr + Tyr205Pro + Asn211Glu +Ala214Pro + Thr215GlyAla199Gln + Ala202Pro + Val204Met + Tyr205Ile + Tyr208Ile +Asn217Asp + Thr219ProAla199Thr + Gly203Asn + Tyr208Leu + Pro209Gly + Tyr213Ile +Ala214Glu + Leu216GlyGly201Gln + Val204Ala + Tyr208Gly + Thr210Gln + Thr215Gln +Asn217Gln + Gly218AsnPro200Gln + Gly201Asn + Val204Ser + Tyr205Glu + Tyr213Ser +Ala214Gln + Asn217GlnPro200Ser + Gly203Asn + Thr207Gly + Thr210Pro + Tyr213His +Ala214Thr + Thr215ProAla199Asn + Tyr208Ile + Pro209Asp + Thr212Gln + Tyr213Ser +Ala214His + Asn217GlnAla199Asn + Pro200Gln + Ala202Pro + Val204Cys + Tyr208His +Thr215Gln + Thr219AsnAla199Ser + Ala202His + Val204Pro + Thr210Asn + Thr212Pro +Gly218Asp + Thr219AsnGly201Pro + Tyr205Glu + Thr210Gly + Asn211Ser + Tyr213Gln +Ala214Thr + Gly218GlnAla199Pro + Gly203Gln + Thr207Asn + Asn211Asp + Ala214Gln +Thr215Gly + Thr219GlyAla199Gln + Pro200Asn + Thr212Ser + Tyr213Ser + Leu216Ile +Asn217Asp + Thr219ProPro209Asn + Thr212Gln + Ala214Ser + Leu216Gly + Asn217Asp +Gly218Asn + Thr219AsnGly201Ser + Gly203Pro + Thr207Ser + Thr212Ser + Ala214His +Asn217Asp + Gly218GlnAla199Ser + Pro200Gln + Gly201Asn + Tyr205Asn + Ala214Ser +Asn217Asp + Gly218ProAla199His + Ala202Gly + Gly203Glu + Tyr208Gln + Pro209Gln +Leu216Asn + Thr219SerAla199Pro + Thr207Ser + Tyr208Cys + Asn211Gln + Tyr213Gly +Leu216Thr + Thr219GlyPro200Gly + Ala202Thr + Tyr205Cys + Thr212Glu + Leu216Thr +Asn217Ser + Gly218ProAla199Thr + Gly203Asn + Tyr208Ser + Pro209Gly + Asn211Ser +Tyr213Pro + Gly218ProAla199Gly + Tyr205Gln + Pro209Asn + Asn211Glu + Ala214His +Thr215Gln + Thr219AsnAla199Asn + Pro200Asn + Gly203Pro + Tyr208Ile + Pro209Gly +Tyr213Asp + Ala214HisGly203Asn + Val204Thr + Asn211Gln + Thr212Pro + Thr215Asp +Leu216His + Thr219GlnAla199Gln + Gly201Ser + Val204Pro + Tyr205Asn + Pro209Asp +Thr210Gln + Thr219AsnPro200Asn + Gly201Ser + Val204Ser + Tyr205Asp + Thr210Asn +Thr212Pro + Asn217SerPro200Asn + Gly201Asn + Tyr205Asp + Tyr208Pro + Thr215Pro +Asn217Gln + Thr219GlyAla199Gln + Gly201Pro + Tyr205Ile + Tyr208Asn + Thr215Gly +Gly218Asp + Thr219ProAla199Pro + Gly201Gln + Tyr205Asn + Thr207Ser + Thr210Gly +Tyr213Met + Ala214SerPro200Asn + Ala202Glu + Val204Thr + Thr207Asn + Tyr208His +Tyr213Gln + Asn217GlnGly201Ser + Ala202Asp + Val204Pro + Thr207Gly + Pro209Gln +Tyr213Pro + Gly218SerGly201Asn + Ala202Asn + Tyr205Cys + Thr210Gly + Tyr213Gln +Ala214Pro + Thr219GlyPro200Asn + Gly203Ser + Thr207Gly + Thr210Pro + Thr215Asp +Leu216Ser + Gly218SerGly201Gln + Ala202Glu + Tyr208Gly + Thr212Pro + Ala214His +Leu216Thr + Thr219GlnAla199Asn + Pro200Gln + Ala202Asn + Tyr205Pro + Thr207Pro +Tyr208Pro + Pro209AspGly201Pro + Ala202Glu + Pro209Ser + Thr212Ser + Thr215Pro +Leu216Cys + Gly218AsnAla202Thr + Gly203Asp + Thr207Gln + Tyr208Cys + Ala214Pro +Leu216Gln + Asn217SerGly201Ser + Ala202Asn + Gly203Pro + Tyr205Met + Asn211Ser +Leu216Glu + Gly218GlnAla199Gly + Tyr205Gly + Asn211Ser + Thr212Ser + Ala214Gln +Asn217Glu + Thr219AsnGly203Gln + Tyr205Gln + Tyr208Met + Pro209Glu + Thr210Glu +Thr215Pro + Thr219GlnPro200Gly + Tyr205Ile + Thr207Gln + Tyr208Pro + Pro209Asp +Thr210Glu + Leu216ProAla199Pro + Pro200Gly + Gly203Gln + Tyr208Asn + Thr210Asp +Asn211Glu + Leu216ProThr207Asn + Tyr208Gln + Pro209Gly + Ala214Asn + Thr215Glu +Leu216Asp + Gly218AsnAla199Thr + Pro200Gln + Thr210Ser + Ala214Gly + Thr215Asp +Leu216Glu + Gly218GlnAla202Gln + Gly203Ser + Val204Cys + Tyr213Ile + Ala214Pro +Leu216Asp + Asn217AspPro200Gly + Gly203Pro + Tyr205Asp + Tyr208Asn + Thr212Gly +Thr215Asp + Gly218ProAla202Glu + Gly203Glu + Val204Cys + Tyr205Pro + Asn217Glu +Gly218Gln + Thr219ProPro200Gly + Ala202Glu + Val204Cys + Thr207Asn + Leu216Ile +Asn217Glu + Gly218GluAla199Pro + Gly201Pro + Thr207Gln + Tyr208Leu + Thr212Glu +Tyr213Asp + Ala214AspAla199Gly + Gly201Pro + Ala202Pro + Tyr208Ala + Pro209Glu +Asn211Asp + Thr212GlyThr207Pro + Pro209Glu + Asn211Glu + Tyr213Met + Ala214Ser +Thr215Gln + Leu216CysGly201Gln + Gly203Glu + Thr207Gln + Tyr208Met + Thr215Asn +Asn217Asp + Thr219ProGly201Asn + Ala202His + Gly203Pro + Tyr205Asp + Thr212Gln +Ala214Glu + Thr219SerVal204Ser + Thr207Pro + Pro209Glu + Thr212Asp + Tyr213Ser +Leu216Gln + Thr219ProAla199Thr + Thr207Pro + Pro209Glu + Thr212Glu + Tyr213Thr +Leu216Ser + Asn217SerPro200Asn + Tyr205His + Pro209Asp + Thr212Asp + Tyr213Ala +Ala214Pro + Thr219GlyAla199Pro + Pro200Gly + Gly201Ser + Val204His + Pro209Glu +Thr212Asp + Thr219GlnAla199Pro + Ala202Gln + Gly203Asp + Thr207Gly + Tyr208Gln +Leu216Glu + Gly218ProGly203Glu + Val204Ser + Tyr208Val + Tyr213Pro + Ala214Asp +Thr215Glu + Leu216AspGly201Gln + Gly203Glu + Val204Gln + Thr207Ser + Thr215Asp +Leu216Ala + Asn217GluAla202Asn + Val204Gly + Asn211Ser + Ala214Asp + Thr215Glu +Leu216Glu + Asn217GluAla199His + Pro200Ser + Gly201Gln + Ala202Glu + Leu216Ile +Gly218Asp + Thr219GlyAla202Gly + Gly203Glu + Val204Gln + Thr207Gly + Thr215Gln +Asn217Glu + Gly218AspAla199Asn + Gly203Glu + Thr210Gly + Ala214Asn + Thr215Pro +Asn217Glu + Gly218AspAla202Pro + Gly203Asn + Tyr205Glu + Asn211Gln + Thr215Gln +Leu216Glu + Thr219AsnPro200Gly + Val204Thr + Thr207Ser + Tyr208Cys + Thr210Asp +Asn211Glu + Tyr213GluAla202Glu + Gly203Pro + Val204Thr + Thr212Ser + Tyr213Val +Thr215Glu + Asn217AspPro200Gln + Val204Met + Pro209Gly + Thr210Asp + Thr212Asp +Tyr213Asp + Ala214AspAla199His + Pro200Gln + Ala202Gly + Val204Cys + Thr215Glu +Leu216Cys + Asn217AspVal204Gly + Tyr208Leu + Pro209Glu + Thr210Asn + Tyr213Glu +Ala214Asp + Thr215SerPro200Gly + Gly201Gln + Ala202Ser + Pro209Gly + Thr212Glu +Ala214Asp + Leu216AlaGly201Asn + Tyr205His + Tyr208Ser + Tyr213Asp + Ala214Asp +Thr215Ser + Leu216AspGly203Asn + Tyr208Gln + Asn211Gln + Tyr213Asp + Thr215Asp +Asn217Gln + Thr219GlnAla202Ser + Thr207Pro + Tyr208Cys + Tyr213Asp + Ala214Ser +Thr215Asp + Leu216CysAla199Thr + Gly201Pro + Tyr205Ser + Thr207Asn + Tyr208Ile +Asn211Glu + Tyr213GluAla199Pro + Ala202Ser + Gly203Pro + Tyr208Gly + Asn211Asp +Tyr213Glu + Thr215GlnAla199Gln + Pro200Asn + Gly203Asp + Tyr205Glu + Tyr208Asn +Tyr213Met + Ala214AspGly201Asn + Thr207Pro + Thr210Asp + Asn211Glu + Thr212Glu +Ala214Glu + Leu216ProPro200Gln + Gly201Asn + Gly203Asn + Thr207Asn + Asn211Asp +Thr212Glu + Ala214GluThr210Gly + Asn211Asp + Tyr213Asp + Ala214Asp + Asn217Ser +Gly218Pro + Thr219AsnAla199Asn + Gly201Ser + Tyr208Ala + Thr210Glu + Tyr213Glu +Ala214Pro + Leu216HisAla202Asn + Gly203Asp + Tyr205Gln + Thr212Ser + Thr215Glu +Gly218Asp + Thr219ProPro200Gly + Gly203Glu + Thr207Gln + Ala214Gln + Thr215Asp +Leu216Asn + Gly218GluAla199Ser + Gly201Ser + Ala202Glu + Gly203Gln + Thr207Ser +Thr215Asp + Thr219SerAla202Asp + Tyr205Thr + Thr207Asn + Tyr208Ala + Thr215Glu +Leu216Gln + Gly218AsnPro200Asn + Gly203Asp + Val204Pro + Tyr208Cys + Leu216Cys +Asn217Ser + Gly218GluAla199Asn + Pro200Ser + Gly201Asn + Gly203Glu + Val204Gly +Asn211Gln + Gly218GluAla199Thr + Gly201Gln + Ala202Pro + Thr210Asp + Asn211Glu +Ala214Asp + Thr219AsnAla199Gly + Pro200Ser + Tyr205Glu + Thr210Pro + Asn211Asp +Tyr213Asp + Thr215GluPro200Ser + Ala202Glu + Thr207Gln + Asn211Ser + Ala214Glu +Asn217Glu + Gly218AsnAla202Ser + Val204His + Thr207Ser + Thr212Asp + Tyr213Asp +Thr215Asn + Leu216AspGly203Pro + Tyr205Pro + Pro209Asp + Asn211Asp + Ala214Glu +Thr215Asp + Thr219GlyThr207Ser + Pro209Ser + Thr212Glu + Tyr213Glu + Tnr215Glu +Leu216Gln + Asn217AspAla199Ser + Pro200Gly + Gly201Ser + Thr212Gly + Ala214Glu +Asn217Glu + Gly218ProGly201Gln + Tyr208Thr + Ala214Asp + Thr215Pro + Leu216Asn +Asn217Glu + Gly218ProAla199Ser + Tyr205Glu + Thr207Ser + Thr210Asn + Tyr213Glu +Ala214Gln + Asn217AspGly203Glu + Pro209Asp + Thr210Asp + Asn211Glu + Ala214Thr +Thr215Gln + Thr219AsnAla202Pro + Gly203Asp + Tyr205Cys + Thr212Asp + Thr215Pro +Leu216Glu + Asn217AspAla199Asn + Gly201Pro + Ala202Glu + Gly203Glu + Tyr205Glu +Thr207Gly + Thr212AspAla202Asn + Thr207Pro + Asn211Glu + Thr212Asp + Ala214Ser +Thr215Asp + Leu216ValGly201Gln + Ala202Glu + Val204Ser + Tyr205Asp + Tyr213Glu +Thr215Asn + Leu216ThrPro200Gly + Gly203Asn + Tyr205Glu + Thr212Glu + Tyr213Val +Ala214Asn + Leu216GluAla202His + Thr207Gln + Pro209Gln + Thr210Gln + Thr215Glu +Asn217Gln + Gly218GluAla199Gly + Gly201Gln + Pro209Gly + Thr212Gly + Thr215Glu +Asn217Ser + Gly218GluGly203Asp + Val204Met + Thr212Asp + Tyr213Val + Ala214Asp +Asn217Glu + Thr219ProAla202Asp + Gly203Asn + Tyr208Met + Thr212Asp + Asn217Glu +Gly218Glu + Thr219GlyPro200Gln + Val204Asn + Thr207Gly + Thr210Glu + Asn211Gln +Tyr213Val + Ala214AspGly203Gln + Tyr205lle + Tyr208His + Thr210Asp + Ala214Asp +Thr215Pro + Leu216ValGly203Gln + Tyr208Ala + Pro209Gln + Thr210Glu + Asn211Gln +Ala214Asp + Leu216CysGly201Gln + Gly203Glu + Val204Ser + Pro209Asn + Asn211Glu +Tyr213Asp + Thr215GluTyr205Asp + Thr207Pro + Pro209Ser + Thr210Glu + Ala214Glu +Asn217Ser + Thr219ProAla199Asn + Gly201Ser + Ala202Thr + Gly203Asp + Thr210Pro +Ala214Asp +Gly218GluGly203Glu + Thr207Ser + Tyr208Thr + Pro209Gln + Thr212Glu +Ala214Ser + Thr215GluAla199Pro + Gly201Ser + Val204Gln + Tyr205Asp + Thr212Asp +Leu216Ala + Thr219ProPro200Gln + Tyr208Ile + Pro209Glu + Ala214Glu + Leu216Asp +Asn217Ser + Gly218GluAla199Thr + Ala202Glu + Gly203Glu + Thr210Asp + Ala214Ser +Gly218Glu + Thr219GlnAla199Thr + Pro200Ser + Ala202Asp + Gly203Asp + Thr210Gly +Thr212Glu + Gly218GluAla199Pro + Ala202Glu + Tyr208Leu + Thr210Asp + Ala214Ser +Leu216Glu + Gly218AspGly201Asn + Tyr205Ala + Thr207Pro + Thr210Asp + Asn211Asp +Ala214Asp + Asn217GluAla199Ser + Gly203Gln + Tyr205Gln + Pro209Glu + Thr212Glu +Tyr213Asn + Leu216AspAla199Gln + Pro200Asn + Tyr205Glu + Tyr208Leu + Pro209Glu +Thr210Asp + Tyr213GlyPro200Gly + Gly203Glu + Pro209Glu + Thr212Ser + Tyr213Pro +Thr215Asp + Asn217GlnGly201Pro + Tyr205Val + Thr207Ser + Pro209Asp + Ala214Glu +Asn217Glu + Thr219GlnPro200Asn + Ala202Thr + Pro209Gly + Asn211Glu + Thr212Glu +Ala214Asn + Leu216AspGly201Ser + Thr207Ser + Thr212Asp + Ala214Pro + Leu216Asp +Asn217Asp + Gly218ProTyr205Asp + Thr207Asn + Pro209Asp + Asn211Glu + Thr212Pro +Gly218Ser + Thr219ProPro200Gly + Ala202Asn + Gly203Asn + Thr210Ser + Thr212Glu +Tyr213Asp + Asn217GluGly203Pro + Pro209Ser + Thr212Asn + Tyr213Glu + Asn217Glu +Gly218Asp + Thr219GlnAla202Ser + Tyr205Ala + Pro209Glu + Thr210Asp + Leu216Ser +Asn217Asp + Gly218GlnVal204Met + Pro209Asp + Thr210Glu + Ala214Gln + Leu216Gln +Gly218Asp + Thr219ProAla202Glu + Val204Met + Thr207Ser + Pro209Glu + Thr210Glu +Thr212Ser + Gly218ProPro200Ser + Ala202His + Thr210Asp + Asn211Glu + Leu216Gly +Gly218Asp + Thr219GlyPro200Gly + Gly201Asn + Ala202Ser + Thr210Asp + Asn211Glu +Ala214His + Thr215AspPro200Gln + Pro209Gly + Thr210Glu + Asn211Asp + Thr212Gln +Tyr213Met + Leu216AspPro200Gly + Gly203Glu + Tyr208Thr + Thr210Asp + Asn211Asp +Ala214Thr + Leu216AsnGly203Ser + Val204Cys + Thr207Asn + Pro209Ser + Thr210Glu +Thr215Glu + Leu216GluAla199Asn + Pro200Asn + Gly201Pro + Gly203Asn + Asn211Asp +Thr215Asp + Leu216AspAla202Thr + Tyr208Met + Pro209Asn + Asn211Glu + Thr212Glu +Ala214Gly + Gly218GluGly203Gln + Val204Cys + Tyr205Gly + Tyr208Pro + Asn211Glu +Thr212Asp + Gly218GluPro200Gln + Ala202Asp + Tyr205Met + Thr212Asp + Tyr213Glu +Gly218Gln + Thr219GlnAla199Gln + Gly203Pro + Pro209Asn + Thr210Gln + Thr212Asp +Asn217Asp + Gly218AspGly201Asn + Tyr208Asn + Pro209Ser + Thr212Glu + Tyr213Pro +Asn217Asp + Gly218AspAla199Gln + Gly201 Pro + Tyr205Ser + Asn211Glu + Ala214Asn +Asn217Glu + Gly218Glu表29多環(huán)雙突變變異體Leu 95Gly + Gly203ProThr 58Gln + Asn211GluAsp 59Glu + Leu216GlnTyr103Cys + Thr212GlyGly156Glu + Ala202GlnGly126Gln + Thr215ProSer187Glu + Ala214GlyAsn 96Gln + Ile106AlaGly101Asp + Tyr205ProAsn 61Ser + Gly165GlnGly 64Gln + Thr132GlyAsp 59Glu + Tyr205AsnSer100Glu + Tyr205HisAla128Ser + Thr161GlnSer158Asp + Gly203SerAsn 61Asp + Asn162SerThr161Glu + Pro209GlnGly 62Asp + Pro209AsnIle106Leu + Asn211AspTyr103Glu + Gly203ProTyr166Pro + Thr215GluIle106Leu + Pro209GluThr163Pro + Ala202ProThr132Gly + Leu216SerSer104Glu + Phe188Val
Tyr166Asp + Ala186ThrSer160Glu + Thr215SerGly 62Gln + Gly165AspLeu 95Gln + Ala128GluSer 97Glu + Asn211SerLeu 95Asn + Phe188LeuAsn157Gln + Thr212SerLeu125Gln + Gly159AspAsn211Asp + Tyr213AlaSer158Asp + Tyr205CysSer160Glu + Asn162GlnSer187Asp + Tyr213ProSer158Asp + Leu216GlnMet198Ser + Ala202AspGly 99Asn + Ala214GluThr 58Asp + Gly130GlnGly156Glu + Tyr208ProGly105Pro + Leu216IleSer187Asp + Tyr205GlnThr215Ser + Gly218AspGly 60Asn + Ala128GluThr 65Gln + Leu125AsnGly101Gln + Gly218AspAsn211Ser + Thr212AspThr212Gln + Thr215GlyGly203Glu + Tyr205GlyThr132Pro + Thr215SerTyr103Asp + Gly165GlnThr 65Pro + Gly203GluThr 58Glu + Gly 60ProGly153Pro + Leu216ValAsn 96Gln + Pro209GluGly 99Asn + Gly153AsnAla128Asn + Ala202HisGly105Glu + Gly130SerThr212Gly + Gly218GluPhe188Val + Tyr205SerAsn217Glu + Thr219GlyLeu 95Gln + Thr215AspVal 94Gln + Thr212GluAsn 61Ser + Gly203AspSer129Glu + Asn154GlnGly 62Glu + Thr219AsnSer129Glu + Asn157GlnTyr103Ile + Thr215ProIle164Thr + Thr215SerGly101Gln + Tyr208Gln
Asn 61Gln + Thr212AspThr212Asp + Leu216IleAsn211Glu + Leu216HisGly153Glu + Thr163GlnGly203Asp + Thr215AsnAsn 61Ser + Thr132AsnSer158Glu + Gly203ProLeu 95Asn + Thr215AsnThr 58Ser + Ser100AspIle164Met + Tyr205AlaSer158Glu + Tyr205MetPhe188Leu + Thr212GluGly130Pro + Ser190GluGly101Asp + Gly126SerAsn 61Ser + Ala186SerAsn162Gln + Phe188SerThr 58Gly + Thr 65AspSer129Asp + Thr210ProSer129Glu + Gly153AsnAsp 59Glu + Tyr213ThrAsn154Glu + Leu216GlnSer 97Asp + Ala199ProGly203Asn + Gly218AspThr 65Ser + Gly126GlnThr215Gln + Leu216AlaAsn 96Glu + Tyr205HisThr 65Ser + Gly156AspAla128Ser + Thr163GlnThr215Asp + Leu216ValSer129Glu + Leu216AlaAsn211Asp + Tyr213LeuVal 94Ser + Phe188ValSer158Glu + Thr212GlnGly 99Pro + Asn154GlnGly203Asn + Thr215GlnAla202Ser + Tyr205GluTyr205Val + Asn217GluVal 94Gly + Thr215AspSer104Glu + Thr215GlnGly201Pro + Tyr205GluAsn 96Ser + Gly127GluTyr205Thr + Ala214ThrGly 62Ser + Thr215GlyThr 65Ser + Asn211GluGly153Pro + Gly203SerAla128Thr + Leu216GluAsn 96Glu + Leu216Gln
Thr215Asn + Leu216AlaAsp 59Glu + Ala202ProVal 94Thr + Leu216MetSer 97Glu + Thr212GlnGly101Asp + Ala202AsnSer129Asp + Ala202GlyTyr103Ala + Thr132SerLeu 95Met + Leu216AspSer100Asp + Gly101ProSer100Asp + Thr219ProVal 94Asn + Thr215ProTyr103Asn + Ala128AspGly201Asn + Tyr205AspThr 58Glu + Ile106CysThr 65Glu + Tyr103ProVal 94Met + Ser 98GluGly203Glu + Thr210ProPro209Glu + Gly218SerThr161Pro + Gly203SerSer 97Glu + Leu125ProAla128Asp + Ala199HisIle106Gly + Ala214ProAsp 59Glu + Tyr205IleAsn154Asp + Tyr208HisThr210Asp + Leu216ValThr215Asp + Thr219GlnGly 60Glu + Leu125SerSer 97Asp + Gly153AsnTyr166Pro + Tyr205GlySer129Glu + Gly203GlnGly153Gln + Pro209GlnGly203Gln + Thr215GlnAsn 61Glu + Thr212SerThr163Asp + Tyr205HisAsn154Glu + Tyr166ThrAsn 96Ser + Gly105AspPhe188Gly + Asn211SerGly153Glu + Gly218AsnSer100Asp + Thr212ProGly126Pro + Gly156AspLeu125Ser + Thr215GluAsp 59Glu + Thr 65GlyGly156Pro + Gly159AspThr 58Asp + Ala202SerLeu 95Met + Gly 99SerAsn 61Gln + Tyr205GluGly126Asn + Gly159Pro
Leu 95Thr + Gly218AsnThr 65Gln + Leu216AlaGly130Pro + Gly203GluGly 64Gln + Ser 98GluGly126Asn + Gly127GlnThr132Asn + Gly153AsnThr 58Pro + Gly105GluGly130Asp + Ala214HisGly126Glu + Tyr205ProTyr103His + Ser129AspSer 98Asp + Pro209SerAsp 59Glu + Tyr205SerGly 60Gln + Thr215AsnPro209Asn + Asn211AspThr132Asp + Ala202AsnGly218Ser + Thr219GlyThr 58Asp + Gly203GlnIle106Asp + Ala202ThrThr161Asn + Ser190AspSer129Glu + Thr215GlnThr212Asn + Leu216AspAsn 96Gln + Ser158GluGly203Pro + Thr215AspVal 94Asp + Tyr205AsnAla128Pro + Thr215AspGly201Gln + Leu216ProThr161Pro + Leu216CysGly126Asp + Tyr205MetSer 97Asp + Thr212ProThr210Asn + Leu216HisAla186Asn + Tyr205ProSer131Asp + Phe188ThrThr132Ser + Leu216AsnAsn 61Asp + Gly130ProLeu 95Asn + Thr132GlySer100Asp + Thr215AsnTyr166Thr + Asn211GlnThr207Pro + Leu216ThrIle106Ala + Gly165AspTyr205Asn + Thr215AsnAsn 96Gln + Gly218AsnSer160Asp + Phe188GlyThr210Gly + Thr212SerGly 64Ser + Asn 96GlnGly101Asp + Asn154GlnGly 64Asn + Asn157GluAsn157Glu + Gly203Gln
Tyr213Asn + Thr215SerGly101Asn + Ala186AsnSer160Glu + Leu216IleTyr103His + Ser187GluAsn 96Ser + Ala214AsnGly 62Asn + Ser 98AspIle106Ser + Asn157GluThr 65Glu + Thr212GlyGly 62Asn + Thr215ProGly127Ser + Thr210AspGly156Ser + Asn217GluSer 98Asp + Tyr103ThrGly127Asn + Gly165ProGly153Glu + Leu216MetSer129Asp + Gly165ProAsn 61Asp + Thr163GlnGly126Asn + Tyr205MetAsn157Gln + Thr212GlnSer190Glu + Thr215GlySer129Glu + Gly165GlnThr 65Glu + Gly101ProVal 94Gln + Gly165GluTyr103Ile + Serl90GluAsn217Gln + Gly218SerSer 98Glu + Gly 99AsnSer102Asp + Thr212SerThr212Ser + Thr215AspGly130Glu + Pro209AsnAla128Pro + Gly165ProAsn217Gln + Gly218AsnLeu125His + Thr163SerThr212Asp + Tyr213AlaThr 65Pro + Thr210SerGly105Pro + Ala202GlnVal 94Thr + Ser158GluThr132Gly + Tyr205CysGly105Gln + Ala214HisGly127Gln + Tyr205CysGly127Glu + Gly130SerPhe188Tyr + Asn211SerGly 64Asn + Phe188AspThr212Ser + Leu216AspMet198His + Gly203ProTyr208Val + Thr210AspGly165Glu + Tyr205MetAsn 96Gln + Thr132GluSer 98Glu + Gly101Pro
Leu125Cys + Thr215AspLeu125Cys + Gly126SerSer190Asp + Thr215AsnGly 99Gln + Gly153AspGly 60Asp + Thr210SerSer160Asp + Phe188LeuVal204Gln + Thr215GluAsn157Glu + Gly203SerTyr213Glu + Leu216CysAsn 61Gln + Leu216AspLeu125Gly + Ser155AspAla186Pro + Val204GlnIle106Met + Ser160AspLeu125Ile + Leu216SerGly 60Gln + Gly203AsnGly203Asn + Pro209GluGly 60Ser + Gly 62AspAsn154Glu + Tyr213GlnAsn154Gln + Gly203AspAsn154Glu + Thr219GlnThr212Ser + Leu216HisTyr205His + Thr215AspGly 60Asp + Gly126ProGly203Glu + Leu216AlaGly156Ser + Gly203SerSer155Asp + Gly165GlnAla202Gly + Thr212AspAsn157Gln + Tyr213SerGly 99Ser + Gly130SerGly203Ser + Thr219AspGly130Ser + Ala214AsnThr210Gln + Thr212AspAsn211Gln + Thr212GluGly153Asn + Thr212AspGly153Asp + Tyr205SerSer100Glu + Tyr205ThrGly126Asn + Ash154GluSer190Asp + Tyr205ValSer102Glu + Tyr166ThrAsn157Ser + Pro209GlyAsp 59Glu + Tyr205LeuSer131Glu + Thr132AsnGly127Asp + Leu216ThrThr210Asn + Asn217GlnGly203Ser + Gly218AspIle106Gln + Gly203AspThr161Asp + Tyr205Pro
Gly105Pro + Thr212AspTyr205Thr + Leu216CysAsn 96Ser + Ser158GluAsn 61Glu + Tyr205AlaTyr205Ser + Thr212GluThr163Gln + Phe188GluTyr103Ala + Thr212AspGly101Asp + Ala202HisSer155Glu + Gly218AsnVal 94Thr + Thr163SerSer102Glu + Asn217GlnGly159Ser + Ala202GluAsn154Ser + Tyr166AlaAsp 59Glu + Phe188GlyThr163Gln + Gly218SerGly153Asn + Tyr205ValSer129Glu + Leu216ProGly126Ser + Thr212GluGly203Asp + Leu216AsnGly126Glu + Thr212ProVal 94Pro + Ala186GluGly165Gln + Thr215GluSer129Asp + Gly153AsnSer 98Glu + Thr215SerGly203Ser + Leu216ProGly105Asp + Leu216HisIle106Met + Thr210AspLeu216Asn + Asn217GluGly127Gln + Leu216GluVal 94Glu + Ala202GlyThr132Asn + Leu216HisAsp 59Glu + Ala202AsnLeu 95Ile + Tyr205ProThr 65Gln + Thr215ProSer104Glu + Thr215AsnSer160Glu + Gly203ProGly203Asp + Leu216ProAla128Glu + Tyr213MetSer 98Asp + Leu216CysSer158Asp + Thr215GlnVal 94Asp + Thr215AsnGly127Asp + Asn154SerGly126Ser + Gly156GlnLeu125Gly + Thr212AspThr210Glu + Leu216AsnAsn154Ser + Ala202AsnAsn 96Gln + Gly126Asn
Gly 64Ser + Gly127SerAsn211Ser + Thr212GlySer160Asp + Gly218GlnTyr205Ala + Thr210GluThr212Asp + Leu216SerSer158Asp + Thr215Ser表33多環(huán)三突變變異體Leu 95Gly + Gly203Pro + Asn211GluThr 58Gln + Asp 59Glu + Leu216GlnTyr103Cys + Ala202Gln + Thr212GlyGly126Gln + Gly156Glu + Thr215ProAsn 96Gln + Ile106Ala + Tyr205ProAsn 61Ser + Gly101Asp + Gly165GlnGly 64Gln + Thr132Gly + Gly153GluAla128Ser + Ser158Asp + Gly203SerTyr103Glu + Ile106Leu + Gly203ProThr163Pro + Ala202Pro + Leu216SerSer104Glu + Thr132Gly + Phe188ValLeu 95Asn + Phe188Leu + Asn211SerAsn162Gln + Ser187Asp + Tyr213ProThr 58Asp + Gly130Gln + Tyr208ProGly105Pro + Gly156Glu + Leu216IleGly 60Asn + Thr215Ser + Gly218AspThr 65Gln + Leu125Asn + Ala128GluAsn211Ser + Thr212Asp + Gly218GlnGly203Glu + Thr212Gln + Thr215GlyThr132Pro + Tyr205Gly + Thr215SerGly 60Pro + Thr 65Pro + Gly203GluAsn 96Gln + Gly203Asp + Thr215GlyGly 99Asn + Gly153Asn + Pro209GluGly105Glu + Ala128Asn + Ala202HisTyr205Ser + Thr212Gly + Gly218GluGly 99Gln + Asn217Glu + Thr219GlyLeu 95Gln + Ala202Thr + Thr215AspSer129Glu + Asn157Gln + Thr219AsnAsp 59Glu + Tyr103Ile + Thr215ProGly 99Asp + Ile164Thr + Thr215SerAsn 61Gln + Tyr208Gln + Thr212AspGly153Glu + Thr163Gln + Thr215AsnAsn 61Ser + Thr132Asn + Gly203ProLeu 95Asn + Ser158Glu + Thr215AsnThr 58Ser + Ser100Asp + Tyr205AlaSer158Glu + Phe188Leu + Tyr205MetAsn 61Ser + Ala186Ser + Phe188Ser
Asn154Glu + Tyr213Thr + Leu216GlnSer 97Asp + Ala199Pro + Gly203AsnThr 65Ser + Gly126Gln + Gly218AspTyr205His + Thr215Gln + Leu216AlaAla128Ser + Thr163Gln + Thr215AspVal 94Ser + Ser100Asp + Phe188ValGly 99Pro + Asn154Gln + Gly203AsnAsn 96Ser + Thr163Asp + Ala214ThrTyr166Asp + Tyr205Thr + Ala214ThrGly 62Ser + Thr 65Ser + Thr215GlyGly153Pro + Gly203Ser + Thr219GluAsp 59Glu + Ala202Pro + Leu216AlaVal 94Thr + Thr212Gln + Leu216MetSer129Asp + Thr132Ser + Ala202GlyLeu 95Met + Tyr103Ala + Leu216AspSer104Asp + Gly203Asn + Thr215AsnThr 65Glu + Tyr103Prc + Ile106CysVal 94Met + Ser 98Glu + Thr210ProSer 97Glu + Thr161Pro + Gly203SerLeu125Pro + Ala128Asp + Ala199HisAsp 59Glu + Tyr205Ile + Ala214ProSer129Glu + Tyr166Pro + Gly203GlnGly203Gln + Pro209Gln + Thr215GlnAsn 96Ser + Asn154Glu + Tyr166ThrGly105Asp + Phe188Gly + Asn211SerThr 58Asp + Gly156Pro + Ala202SerLeu 95Met + Gly 99Ser + Ser131GluGly126Asn + Gly159Pro + Tyr205GluLeu 95Thr + Leu216Ala + Gly218AsnGly105Glu + Gly130Pro + Leu216ThrThr 58Pro + Thr132Asn + Gly153AsnGly 60Gln + Pro209Asn + Thr215AsnThr132Asp + Ala202Asn + Thr219GlyThr 58Asp + Gly203Gln + Gly218SerSer129Glu + Thr212Asn + Thr215GlnGly203Pro + Tyr205Asn + Thr215AspAla128Pro + Thr215Asp + Leu216ProThr161Pro + Gly201Gln + Leu216CysGly126Asp + Tyr205Met + Thr212ProSer 97Asp + Thr210Asn + Leu216HisAla186Asn + Phe188Thr + Tyr205ProAsn 61Asp + Thr132Ser + Leu216AsnLeu 95Asn + Thr132Gly + Thr215AspIle106Asp + Tyr166Thr + Asn211GlnIle106Ala + Thr207Pro + Leu216ThrTyr205Asn + Thr215Asn + Gly218AsnAsn 96Gln + Ser160Asp + Phe188Gly
Gly 64Ser + Gly101Asp + Asn154GlnThr 58Asn + Gly 64Asn + Asn157GluGly101Asn + Ala186Asn + Thr215SerGly 62Asn + Asn 96Ser + Ala214AsnGly130Asp + Phe188Asn + Gly218ProGly 99Ser + Ala128Glu + Leu216GlyVal 94Ala + Pro209Ser + Thr215AspLeu 95Thr + Gly126Gln + Leu216GlyAsp 59Glu + Gly159Ser + Tyr205ProAla199Gly + Ala202Gln + Gly203GlnSer158Glu + Thr161Gly + Tyr205AsnThr 58Ser + Ser190Glu + Tyr205ProLeu 95Asn + Tyr205Ala + Pro209GluThr 58Asn + Gly153Pro + Thr210ProAla128Gly + Gly203Asn + Tyr205ProVal 94Cys + Asn157Gln + Tyr205GlnSer129Glu + Tyr205Asn + Thr215AsnThr 65Gly + Gly127Asn + Thr212GlnGly 60Asn + Tyr205Thr + Asn217GlnGly126Pro + Ala214Pro + Thr215GlyThr163Gly + Ala202Pro + Thr212ProThr 58Pro + Gly130Ser + Ser160AspGly153Ser + Asn154Glu + Thr212GlyLeu125Gly + Tyr205Thr + Thr210GluTyr103Val + Thr132Gly + Pro200GlnSer100Glu + Gly127Asn + Leu216AlaSer160Glu + Ala186Ser + Tyr205SerGly159Ser + Thr163Glu + Ile164HisGly101Asp + Tyr166Cys + Gly203ProThr 58Asn + Asn 61Glu + Ala214SerGly127Glu + Phe188Met + Ala202GlyAsn162Glu + Gly203Ser + Gly218AsnTyr166Asn + Thr210Glu + Ala214HisIle106Asp + Thr212Ser + Thr215GlnAsn162Glu + Ala202Ser + Thr215AsnTyr103Pro + Ser187Glu + Tyr205ProVal204Thr + Thr212Ser + Thr215GlnSer100Asp + Gly101Asn + Leu216HisLeu 95Cys + Gly203Ser + Leu216ThrAsn 96Ser + Gly153Glu + Tyr205ProIle106Pro + Ala202Ser + Thr212ProSer102Asp + Gly203Ser + Thr215SerLeu 95Cys + Ser104Glu + Phe188CysAsp 59Glu + Ala214Gly + Thr215GlyLeu125Cys + Thr212Gly + Thr215SerLeu125Val + Ala202His + Thr215ProGly101Asp + Gly127Asn + Gly203Gln
Thr 58Pro + Gly105Asn + Gly203GluTyr166Cys + Gly203Ser + Tyr205AspGly126Gln + Ile164Val + Leu216HisGly153Ser + Thr215Pro + Leu216GluSer160Asp + Gly218Gln + Thr219AsnAsn 61Glu + Asn157Ser + Ala214SerGly 62Ser + Ala128Ser + Asn154AspIle106Gly + Tyr205Val + Thr215SerSer160Glu + Asn211Gln + Thr215ProThr 65Gly + Ser100Glu + Thr215ProThr 65Ser + Ser104Glu + Gly203ProAsn 61Ser + Ile106Ser + Thr219ProSer102Asp + Gly153Pro + Leu216HisThr 65Glu + Gly126Gln + Gly203SerSer187Glu + Phe188Ile + Leu216ThrThr132Gln + Gly165Asp + Thr212SerSer129Asp + Asn211Ser + Leu216ThrLeu125Ile + Asn154Ser + Leu216MetThr 65Pro + Gly101Pro + Thr212AspThr 65Gln + Leu125Asn + Leu216MetPro209Gly + Ala214Glu + Thr215ProLeu125Val + Tyr205Ser + Leu216HisLeu 95Ash + Gly156Asp + Thr212GlnLeu125Pro + Thr210Asp + Thr215SerGly105Asn + Thr161Asn + Tyr205ValGly126Ser + Ser160Glu + Gly218AsnVal 94Thr + Thr207Gly + Thr212GluTyr208Gln + Thr212Gln + Gly218AspIle106Asn + Gly159Asn + Phe188AspTyr205Met + Thr212Gln + Tyr213ThrSer131Asp + Thr132Pro + Tyr213AsnGly105Ser + Ala128Thr + Thr215ProLeu 95His + Asn 96Asp + Thr215GlyGly 60Ser + Gly156Asp + Pro209AsnAsn162Asp + Thr163Asn + Ala199GlyTyr103Glu + Leu125His + Thr215SerGly127Glu + Asn157Gln + Tyr205LeuAsp 59Glu + Gly105Pro + Ala186AsnAla128Asp + Tyr213His + Leu216HisThr 65Pro + Ile106His + Ser131GluGly126Gln + Thr161Asp + Ala202SerGly 99Gln + Gly126Asp + Thr212GlyThr 58Asp + Gly159Ser + Thr212SerAsp 59Glu + Ala186Gly + Gly203GlnGly 62Glu + Tyr205Ala + Leu216GlnAsn154Gln + Tyr205Ser + Tyr213MetLeu 95Ser + Ser158Glu + Thr212Gly
Tyr166Asn + Tyr205Glu + Leu216AsnAsp 59Glu + Ala128Asn + Gly153ProGly 99Asp + Gly159Asn + Asn211SerGly165Pro + Thr215Gln + Leu216ThrGly165Asn + Thr215Asn + Leu216AspGly 60Asp + Gly159Asn + Thr212GlnThr 58Glu + Thr163Gly + Thr210GlnGly159Ser + Ala186Gly + Ala199ThrGly203Glu + Asn211Gln + Tyr213GlnGly 99Ser + Asn162Asp + Thr215GlnThr 65Gly + Ser190Glu + Thr219SerVal 94Met + Asn162Ser + Ser190GluAsn 61Gln + Leu 95Ser + Tyr205AspGly126Glu + Gly201Pro + Gly203SerThr 58Gly + Thr212Pro + Gly218GluSer104Asp + Gly105Ser + Gly159ProThr 58Gln + Thr163Gly + Tyr205ProGly 60Ser + Thr 65Pro + Thr163GlyLeu125Met + Ser131Glu + Thr210GlyPhe188Gln + Gly201Ser + Ala202GlyGly203Pro + Leu216Glu + Asn217GlnGly 62Asn + Gly126Pro + Thr210AsnGly 60Asn + Ser104Asp + Gly127GlnSer155Glu + Thr163Pro + Gly203ProLeu 95Met + Thr132Asp + Asn211SerTyr103Ile + Gly159Asp + Ala202GlyGly101Gln + Tyr166Pro + Asn211GlnAsn157Ser + Ala202Ser + Thr215GlyThr132Pro + Ser155Glu + Ala214HisThr132Ser + Thr212Asp + Leu216GlyGly126Asn + Thr212Asp + Gly218GlnTyr205Cys + Thr212Pro + Leu216MetAsn 96Gln + Thr163Gln + Leu216GluAla202Ser + Tyr205Asn + Gly218AspLeu 95Asp + Gly126Asn + Ala202SerSer100Glu + Ile106Gln + Ala202GlnThr 58Ser + Ser100Glu + Ile164LeuGly101Ser + Phe188Asp + Tyr213GlySer104Glu + Gly156Gln + Thr212GlyLeu125Val + Gly203Asn + Tyr205ThrThr163Gln + Thr215Asn + Asn217GluGly 99Pro + Gly165Glu + Ala214GlnMet198Ser + Gly203Glu + Tyr213ValAsn 96Ser + Ser 97Asp + Thr212SerAla202Glu + Gly203Ser + Tyr205GlnGly 64Asn + Asn154Gln + Asn157GluThr 65Ser + Gly130Pro + Tyr205Asp
Gly126Gln + Thr161Gln + Pro209SerAsp 59Glu + Phe188Leu + Leu216AlaSer131Asp + Thr212Pro + Thr215GlnVal204Pro + Gly218Asn + Thr219GluThr 58Ser + Gly203Glu + Thr212ProGly 62Gln + Tyr205Gly + Thr212GluThr 65Gly + Ser129Asp + Met198GlyLeu 95Cys + Asn162Ser + Ala202ThrLeu125Asp + Tyr205Thr + Leu216ThrAsn157Asp + Tyr205Ser + Leu216ProGly156Asn + Gly165Asn + Thr215GluAla128Ser + Thr163Glu + Ala199SerVal 94Ser + Ser187Glu + Thr215GlnVal 94Pro + Thr215Gly + Asn217GluVal 94Asn + Gly130Glu + Gly203SerGly127Gln + Ile164Cys + Thr210GlnSer187Glu + Thr212Gln + Thr215AsnAsn 96Gln + Gly126Gln + Gly203AspGly165Asp + Thr210Ser + Tyr213IleThr 58Gln + Gly153Ser + Asn217GlnGly203Asp + Tyr205Met + Thr210SerAla128Pro + Thr132Glu + Thr215ProSer104Glu + Phe188Ser + Thr212GlySer104Glu + Asn157Ser + Ala214SerIle106Cys + Tyr205Glu + Thr215AsnGly156Gln + Tyr205Pro + Thr219AspAsn157Glu + Tyr213Thr + Leu216IleThr 58Ser + Asn162Ser + Ser190AspGly 62Ser + Thr 65Asp + Tyr205IleLeu125Pro + Gly203Asp + Thr212SerThr 58Gln + Ser 98Asp + Thr212ProSer 97Asp + Leu125Val + Tyr213IleIle106Leu + Phe188Glu + Thr212AsnThr 58Pro + Gly153Pro + Asn162GlnSer190Asp + Thr210Gln + Leu216AlaSer100Glu + Thr215Gly + Leu216MetGly127Glu + Ala128Gln + Thr212ProVal 94Pro + Gly127Ser + Thr215GluThr161Asp + Ala199Asn + Tyr213GlnThr132Pro + Tyr166Leu + Thr212ProGly203Gln + Thr215Asn + Leu216MetGly127Pro + Gly203Asp + Tyr205IleThr163Ser + Tyr205Ala + Pro209GlnVal 94Cys + Thr215Asn + Gly218ProGly126Glu + Gly165Pro + Ala202ThrThr 58Glu + Tyr103Asn + Pro200GlnPhe188Thr + Ala202Asn + Asn217Ser
Gly 60Asp + Leu 95Thr + Tyr205IleSer102Glu + Ala202Ser + Leu216GlnGly 99Asn + Ser102Asp + Gly218GlnIle106Ala + Thr132Pro + Asn157AspGly203Ser + Tyr205Gln + Thr212GluSer 97Asp + Ile106His + Asn211GlnSer100Glu + Gly201Asn + Tyr205HisThr212Gln + Ala214Glu + Thr215GluAsn 61Glu + Gly 62Glu + Gly203ProAsn211Glu + Thr212Asp + Leu216IleGly 99Glu + Ser100Asp + Gly156ProAla202Asp + Gly203Glu + Tyr205ProSer 97Asp + Ser 98Glu + Thr163GlnAla128Thr + Tyr205Asp + Thr215AspSer102Asp + Gly105Asp + Leu216GlyGly127Glu + Gly165Glu + Tyr205ThrGly203Asp + Tyr205Ala + Asn217GluAsp 59Glu + Asn 61Glu + Gly130SerGly 60Glu + Ser 97Glu + Gly 99AsnTyr166Ala + Gly203Glu + Leu216AspSer158Asp + Ser160Glu + Tyr205CysSer129Asp + Tyr166Glu + Thr215GlnSer160Asp + Asn162Glu + Thr210GlySer129Asp + Ser131Asp + Gly218AsnAsn 61Gln + Leu 95Asp + Ser 97AspGly 62Ser + Leu216Asp + Gly218GluAsp 59Glu + Gly165Asn + Pro209GluLeu 95Glu + Gly105Glu + Gly203SerAsn157Glu + Asn162Asp + Thr215GlnTyr103His + Asn162Glu + Ser190AspSer158Asp + Asn162Glu + Thr163ProGly 62Glu + Ser 97Glu + Gly153GlnGly 99Gln + Asn154Asp + Ser187GluSer155Asp + Gly156Asp + Thr219AspGly1535er + Asn154Asp + Gly165AspAsp 59Glu + Asn 96Glu + Tyr205SerAsp 59Glu + Asn 96Asp + Leu216IleSer104Asp + Ser131Asp + Thr210GlySer 97Glu + Gly101Asp + Ala202AsnAsn 96Asp + Ser102Glu + Ile106GlyAsn 61Asp + Leu 95Glu + Val204GlnGly156Asp + Thr161Asp + Gly203SerSer100Asp + Tyr103Asp + Gly126AsnThr 65Asp + Tyr205Asp + Thr215AspThr 58Asp + Ser 97Asp + Thr132GlyThr215Pro + Leu216Asp + Thr219GluSer100Glu + Gly105Glu + Asn162Gln
Ala186Glu + Ala202Glu + Thr215AspAsn 61Asp + Ser100Glu + Asn162SerSer104Asp + Thr132Glu + Tyr205MetGly 99Asn + Thr210Asp + Ala214GluAsn157Asp + Ala186Glu + Thr212SerGly 62Asp + Pro209Asn + Asn211AspGly105Pro + Gly153Glu + Asn162AspAla186Ser + Thr212Asp + Thr215AspGly 99Glu + Ser102Asp + Leu125GlyGly 62Ser + Gly 99Glu + Ser102AspGly126Glu + Ser155Glu + Tyr205ProSer158Glu + Phe188Asp + Ala202SerGly 60Glu + Leu 95Glu + Ile106LeuAla128Asp + Asn154Asp + Thr215GlnAla128Asp + Asn162Glu + Asn217SerAsn157Gln + Ser187Glu + Asn217AspGly 62Asp + Ser 98Asp + Thr212GluGly159Glu + Ser160Asp + Phe188AspGly126Asp + Thr210Glu + Asn211GluSer104Asp + Gly105Asp + Ala128GluAsp 59Glu + Tyr205Glu + Thr215GluGly 60Asp + Tyr205Glu + Thr215AspAsn 61Glu + Pro209Asp + Thr215GluAsn 61Asp + Ser 97Glu + Tyr205AspSer102Asp + Gly105Glu + Pro209AspSer158Asp + Gly203Asp + Thr215GluAsn157Asp + Gly203Asp + Thr215AspGly127Glu + Ser129Glu + Thr215GluAsn 96Glu + Ser100Asp + Thr212GluSer190Glu + Gly203Glu + Tyr205GluSer129Asp + Phe188Glu + Ser190AspSer100Asp + Ser104Glu + Thr212ProSer129Glu + Gly153Glu + Asn157GlnSer 97Glu + Thr212Glu + Ala214GluSer100Asp + Ser102Glu + Ser160AspAsp 59Glu + Ser 97Glu + Asn157AspSer 97Glu + Asn157Glu + Ser160GluGly101Asn + Leu125Glu + Ser129GluGly101Glu + Gly130Glu + Thr161SerAsn154Glu + Ser158Glu + Gly203GlnVal 94Asp + Ser187Asp + Ser190Glu表34多環(huán)四突變變異體Thr 58Gln + Leu 95Gly + Gly203Pro + Asn211GluAsp 59Glu + Tyr103Cys + Thr212Gly + Leu216Gln
Gly126Gln + Gly156Glu + Ala202Gln + Thr215ProAsn 615er + Gly101Asp + Gly165Gln + Tyr205ProSer104Glu + Thr132Gly + Phe188Val + Leu216SerGly127Glu + Thr212Asn + Thr215Ser + Leu216GlnLeu125Thr + Thr210Gln + Thr212Asp + Thr215SerThr132Pro + Gly203Glu + Tyr205Gly + Thr215GlyAsn 96Gln + Gly153Pro + Pro209Glu + Leu216ValGly 99Asn + Ala128Asn + Gly153Asn + Ala202HisPhe188Val + Tyr205Ser + Thr212Gly + Gly218GluTyr103Ile + Ser129Glu + Asn157Gln + Thr215ProLeu 95Asn + Ser158Glu + Gly203Pro + Thr215AsnThr 58Ser + Ser100Asp + Ile164Met + Tyr205AlaThr 65Ser + Gly126Gln + Gly203Asn + Gly218AspAsn 96Glu + Tyr205His + Thr215Gln + Leu216AlaThr 65Ser + Ala128Ser + Gly156Asp + Thr163GlnAsn154Gln + Asn157Glu + Gly203Asn + Thr215GlnGly 62Ser + Tyr205Thr + Ala214Thr + Thr215GlyAsp 59Glu + Asn 61Gln + Ala202Pro + Leu216AlaLeu 95Met + Tyr103Ala + Thr132Ser + Tyr205AspSer 97Glu + Leu125Pro + Thr161Pro + Gly203SerAsp 59Glu + Ile106Gly + Tyr205Ile + Ala214ProSer129Glu + Tyr166Pro + Gly203Gln + Tyr205GlyAsn 61Glu + Gly203Gln + Thr212Ser + Thr215GlnGly153Glu + Phe188Gly + Asn211Ser + Gly218AsnTyr103Val + Gly126Asn + Pro209Asn + Leu216AspAla128Ser + Ser190Glu + Tyr213Ala + Leu216ValGly127Ser + Asn154Ser + Tyr205Leu + Thr212GlnAla128Glu + Asn154Gln + Asn157Gln + Leu216HisSer155Asp + Gly156Ser + Gly203Ser + Leu216AlaGly 99Ser + Gly130Ser + Asn157Gln + Tyr213SerGly130Ser + Gly203Ser + Ala214Asn + Thr219AspAsp 59Glu + Asn157Ser + Tyr166Thr + Pro209GlyGly203Ser + Thr210Asn + Leu216Thr + Asn217GlnTyr103Ala + Thr163Gln + Ala202His + Thr212AspGly 62Glu + Asn154Ser + Tyr166Ala + Phe188GlyAsp 59Glu + Thr163Gln + Tyr205Val + Gly218SerVal 94Pro + Gly153Asn + Gly165Gln + Thr215GluSer 98Glu + Asn154Gln + Gly203Ser + Leu216ProIle106Glu + Thr132Asn + Ala202Gly + Leu216HisAsp 59Glu + Leu 95Ile + Ala202Asn + Tyr205ProThr 65Gln + Asn 96Gln + Ser104Glu + Thr215ProLeu125Gly + Gly126Ser + Asn154Ser + Gly156GlnAsn 96Gln + Gly126Asn + Gly127Ser + Ala202AsnGly 64Ser + Ser160Asp + Asn211Ser + Thr212GlyGly101Pro + Ser158Asp + Thr215Ser + Leu216SerVal 94Glu + Ala202Ser + Tyr205Ser + Thr212GlyGly153Ser + Asn154Glu + Thr161Asn + Thr212Gly
Tyr103Val + Leu125Gly + Thr132Gly + Pro200GlnGly127Pro + Phe188Met + Ala202Gly + Thr215GluAsn162Glu + Gly203Ser + Ala214His + Gly218AsnAsn 96Glu + Gly 99Pro + Thr132Asn + Thr212ProGly101Asn + Tyr205Gly + Tyr213Met + Thr215AspLeu125Pro + Gly126Asn + Ala202Gly + Gly203AspGly 99Pro + Gly127Glu + Tyr205His + Thr212GlnTyr103Pro + Tyr166Ile + Gly203Ser + Thr207GlyThr 58Gly + Gly 62Ser + Gly105Glu + Ala128SerIle106Gly + Tyr205Val + Thr215Ser + Leu216AspAsn 61Ser + Ile106Ser + Ser190Asp + Thr219ProThr 65Glu + Gly126Gln + Gly203Ser + Leu216HisSer187Glu + Phe188Ile + Thr212Ser + Leu216ThrLeu125Ile + Asn154Ser + Asn211Ser + Leu216MetThr 65Gln + Leu125Asn + Pro209Gly + Ala214GluLeu125Val + Tyr205Ser + Thr215Pro + Leu216HisLeu 95Asn + Leu125Pro + Gly156Asp + Thr212GlnGly126Ser + Ser160Glu + Thr161Asn + Gly218AsnLeu 95Gln + Phe188Ile + Ala202His + Thr212AsnSer104Glu + Thr212Gln + Tyr213Thr + Thr215ProAla128Thr + Ser131Asp + Thr132Pro + Tyr213AsnLeu 95His + Ser129Glu + Thr215Gly + Leu216AsnGly105Pro + Ala128Asp + Tyr213His + Leu216HisGly159Ser + Tyr205Ala + Thr212Gly + Leu216AlaGly 62Glu + Asn154Gln + Tyr205Ser + Leu216GlnLeu 95Ser + Ser158Glu + Val204Gln + Thr212GlySer 98Asp + Leu216Ile + Asn217Ser + Gly218AsnLeu125Gly + Ala128Gln + Gly203Glu + Pro209GlnGly105Asn + Tyr166Val + Ala202Asp + Thr215ProSer 97Asp + Ala202Gln + Pro209Gln + Thr215GlyLeu 95Thr + Tyr205Ile + Leu216Val + Thr219SerGly 99Asn + Ser102Asp + Tyr205Ser + Gly218GlnThr 65Pro + Gly165Asn + Ala186Pro + Gly201SerAsn154Asp + Gly203Gln + Tyr205Asn + Thr212AsnIle106His + Ala186Gly + Thr212Asn + Leu216AspGly126Glu + Gly127Gln + Ala128Asn + Ala202AsnLeu 95Ser + Ala128Gly + Thr161Gln + Asn217GluAsp 59Glu + Gly101Pro + Ala199Gly + Gly203SerAsn 61Asp + Val204Thr + Tyr205Cys + Thr215SerGly101Asn + Asn162Asp + Phe188Ala + Tyr205HisAla186Thr + Gly203Asp + Thr215Pro + Asn217SerTyr103Gln + Phe188Met + Thr210Glu + Thr219ProGly 62Asp + Gly126Ser + Leu216Pro + Thr219AsnGly 60Gln + Tyr205Pro + Thr212Ser + Thr215AspIle106Val + Ser187Glu + Thr212Pro + Thr215SerGly159Asp + Asn211Ser + Ala214Gln + Thr219ProAsn 96Ser + Ser160Asp + Ala214Gln + Leu216Gly
Ser104Asp + Gly203Ser + Tyr205Ser + Thr215ProGly153Gln + Phe188Tyr + Thr215Glu + Leu216IleSer129Asp + Thr132Pro + Thr212Gln + Leu216ThrAla128Gln + Ala186Asn + Thr215Asp + Gly218AsnAsn 96Asp + Ala202Pro + Tyr205Gln + Pro209SerThr132Asp + Gly165Pro + Thr210Pro + Leu216CysGly105Pro + Gly130Ser + Thr161Glu + Thr215ProIle106Cys + Asn157Gln + Tyr205His + Thr212GlnLeu 95Gly + Ser100Glu + Gly101Pro + Gly126SerGly 60Ser + Gly105Asp + Thr132Pro + Tyr205ThrThr132Ser + Ile164Val + Gly203Glu + Gly218AsnGly203Gln + Thr212Pro + Thr215Asn + Leu216ProAsn 61Ser + Val 94Gln + Thr132Asp + Val204GlyGly201Ser + Thr212Gly + Thr215Pro + Leu216ProGly153Pro + Ser160Glu + Gly165Gln + Gly218SerTyr166Ile + Tyr213Ser + Ala214Glu + Gly218AsnThr132Ser + Thr163Asp + Tyr166Val + Ala199HisThr132Gln + Asn157Ser + Tyr208Gly + Thr215AspGly203Glu + Tyr205Ile + Tyr208Pro + Thr215SerGly127Ser + Gly165Ser + Tyr213Ile + Leu216HisGly156Glu + Pro209Asn + Asn211Ser + Thr212SerGly165Gln + Phe188Leu + Ala202His + Thr212GluSer102Asp + Gly130Asn + Asn162Ser + Leu216ThrVal 94Cys + Gly156Ser + Tyr205Ala + Leu216AlaGly 99Pro + Tyr103Glu + Phe188His + Leu216CysTyr103Gln + Gly126Gln + Gly156Pro + Tyr166AsnLeu125Pro + Gly153Glu + Asn162Ser + Thr215SerGly 64Ser + Gly 99Asn + Gly203Asp + Thr210GlnGly 99Asn + Ala202Pro + Thr212Asn + Gly218SerAla128Ser + Gly203Gln + Pro209Glu + Gly218SerThr 58Ser + Tyr103Gly + Leu125Gly + Tyr205GlyAsn154Glu + Thr161Ser + Asn162Gln + Gly203GlnGly101Pro + Asn154Ser + Leu216His + Asn217GluThr163Asp + Ala186Ser + Tyr205His + Leu216ProGly159Glu + Gly203Ser + Thr210Gly + Leu216HisLeu125Ser + Thr163Glu + Ala202Pro + Thr210GlnGly153Pro + Ser155Glu + Thr210Asn + Leu216CysGly101Asp + Gly126Gln + Tyr166Ile + Gly203ProSer158Asp + Gly201Asn + Va1204Asn + Pro209SerTyr166Pro + Tyr205Pro + Thr215Asp + Leu216AlaGly 64Asn + Ser160Asp + Tyr205Leu + Thr215GlyGly101Ser + Thr132Gln + Gly159Pro + Thr161SerTyr103Val + Asn154Gln + Gly159Ser + Asn211AspGly127Glu + Asn157Ser + Thr161Asn + Thr212GlyAsn 61Asp + Ile164Asn + Pro209Asn + Leu216ValSer158Asp + Asn162Ser + Tyr205Asn + Thr212GlyLeu125Val + Ala202Thr + Pro209Asn + Thr219Pro
Gly 60Asp + Asn154Gln + Gly203Pro + Tyr205LeuLeu125Gln + Asn162Ser + Gly203Asp + Thr215GlySer102Asp + Tyr103Ile + Tyr205Met + Thr212GlnGly 99Gln + Ala214Gln + Leu216Ala + Gly218GluGly105Asn + Gly156Gln + Gly203Pro + Gly218AspIle106Ser + Thr212Asn + Leu216Glu + Gly218GlnAsn 96Asp + Tyr166Thr + Gly203Ser + Gly218ProSer155Asp + Gly203Gln + Tyr213Thr + Thr215GlyGly101Ser + Thr212Gly + Thr215Asn + Leu216HisThr 65Ser + Leu 95Asn + Thr132Ser + Asn162AspLeu 95Met + Thr132Asn + Gly203Asp + Thr215GlyAsn154Ser + Gly203Glu + Thr212Gly + Leu216AlaGly 60Ser + Ser131Glu + Thr210Ser + Asn217GlnGly 62Asn + Asn162Gln + Gly203Gln + Thr215SerGly 99Ser + Tyr103Ala + Thr210Gln + Thr219SerAsn162Gln + Gly203Glu + Thr215Gln + Thr219GlyAsn162Gln + Gly165Asn + Leu216Ile + Gly218SerThr 65Asn + Gly 99Gln + Thr212Ser + Leu216AspPhe188Val + Ala214Asp + Thr215Asp + Leu216GlnGly 99Glu + Ser100Asp + Gly156Pro + Tyr205GlnVal 94Asn + Tyr103Asp + Ser104Asp + Thr215ProGly 64Gln + Asn154Asp + Ser155Glu + Leu216SerAsn154Glu + Ser155Glu + Gly156Gln + Thr215ProGly153Pro + Asn157Ser + Tyr205Asp + Thr215AspTyr103Leu + Phe188Met + Tyr205Glu + Thr215GluTyr205Ala + Thr210Glu + Thr212Asp + Gly218GlnAla202His + Tyr205Val + Thr210Glu + Thr212AspGly203Asp + Tyr205His + Thr212Ser + Thr215AspGly127Glu + Ser129Asp + Gly153Asn + Thr210ProAla128Asn + Tyr166Ala + Gly203Asp + Leu216AspSer 98Glu + Ser100Glu + Gly203Ser + Asn217GlnAsn 61Asp + Gly 62Pro + Asn 96Asp + Ser 97GluGly159Ser + Thr161Glu + Thr163Glu + Ile164HisSgr160Glu + Asn162Glu + Asn211Gln + Thr215ProAsn 61Asp + Gly 99Asp + Thr161Gly + Thr212ProIle106Val + Ala186Asp + Ala202Glu + Thr215GlnAsp 59Glu + Gly 64Asn + Thr 65Glu + Tyr213SerVal 94Pro + Thr215Asp + Leu216His + Asn217AspAsp 59Glu + Gly127Asn + Ala186Pro + Pro209AspTyr103Glu + Ser131Glu + Tyr213His + Thr215SerSer129Asp + Thr132Gln + Gly165Asp + Leu216ThrTyr205Ile + Thr212Glu + Ala214Glu + Thr219GlnThr 58Gly + Thr212Gln + Tyr213Asp + Thr215AspAsp 59Glu + Gly 62Gln + Ser 97Glu + Tyr205GlyPhe188Asp + Tyr208Gln + Thr212Gln + Gly218AspAla186Asp + Ala202Asp + Gly203Glu + Tyr205ProAsn154Asp + Ser155Asp + Gly165Ser + Ser187Asp
Ser 97Asp + Ser100Glu + Gly201Asn + Tyr205HisSer 97Asp + Ser100Asp + Gly165Ser + Tyr205ThrGly105Ser + Asn157Glu + Ser160Glu + Thr215ProGly156Asp + Thr161Asp + Ser190Asp + Gly203SerThr 65Gly + Ser100Glu + Leu125Glu + Thr215ProGly 62Glu + Ser 97Glu + Gly153Gln + Pro209GlnGly 62Glu + Asn157Gln + Gly203Gln + Thr210GluAsn154Asp + Gly165Glu + Thr212Gly + Gly218ProAsn162Asp + Gly165Asp + Thr212Gln + Leu216CysSer100Glu + Gly126Glu + Ala186Gln + Tyr205AlaAsn154Ser + Ser158Asp + Ser190Glu + Thr215GlnSer129Asp + Thr132Asp + Gly203Gln + Thr215GlnAsn 96Ser + Thr163Asp + Tyr166Asp + Ala214ThrGly 60Glu + Ser 97Glu + Tyr166Asn + Thr210GluIle106Gln + Asn154Glu + Gly203Asp + Gly218AspSer100Glu + Gly105Glu + Asn162Gln + Phe188SerAla186Glu + Ala202Glu + Thr215Asp + Asn217GlnAsp 59Glu + Gly 99Asp + Ile164Thr + Thr215SerSer129Glu + Asn154Glu + Asn162Ser + Gly165AspSer104Glu + Thr132Glu + Tyr205Met + Thr215ProGly 62Asp + Ile106Leu + Pro209Asn + Asn211AspAsp 59Glu + Ser 98Asp + Gly156Asn + Gly218AsnAsn 96Asp + Ser100Asp + Ser104Glu + Thr132AsnAla128Asp + Asn154Asp + Tyr213Met + Thr215GlnSer131Glu + Gly153Asp + Gly165Asp + Tyr205GlyGly101Asp + Gly105Asp + Gly126Asn + Ser131GluSer129Asp + Asn157Glu + Thr163Glu + Thr212ProGly 99Asn + Ala128Asp + Asn162Glu + Asn217SerSer 98Asp + Ser100Asp + Ser104Glu + Thr212ProAsn 96Gln + Ile106Ala + Gly156Glu + Ser160AspGly203Asp + Pro209Glu + Thr215Asp + Leu216ProAsn 96Asp + Ser 97Glu + Ser104Asp + Ala214SerGly159Asn + Gly165Glu + Leu216Cys + Thr219AspGly153Asp + Ser187Asp + Gly203Asn + Leu216SerThr 65Glu + Tyr103Pro + Thr215Asp + Asn217AspGly 62Asp + Ala202Glu + Thr215Asp + Leu216SerThr 65Asp + Gly101Gln + Gly130Ser + Tyr205AspAsn 61Ser + Thr 65Asp + Thr132Asn + Tyr205AspGly 99Pro + Ser187Glu + Gly203Glu + Thr215AspThr 65Ser + Gly126Asp + Thr210Glu + Asn211GluAsn 96Asp + Thr161Asp + Asn162Glu + Ala202AsnGly101Glu + Ile106Met + Ala214Asp + Thr215GluAsn 61Asp + Ala186Asp + Ser187Glu + Leu216ProSer104Asp + Tyr205Val + Thr215Asp + Leu216AspLeu 95Asp + Ala202His + Thr215Glu + Leu216GluSer129Asp + Gly130Asp + Thr210Gln + Thr212AspIle106Glu + Asn162Ser + Asn211Asp + Thr212Glu
Thr 58Asp + Asp 59Glu + Ile106Glu + Phe188SerSer104Asp + Gly105Asp + Ala128Glu + Thr215GlyGly 99Asp + Ser100Glu + Thr161Glu + Pro209GlnGly 99Asp + Ser100Glu + Ser190Asp + Tyr205HisAsp 59Glu + Ala202Asp + Gly203Glu + Leu216CysAsp 59Glu + Gly 60Glu + Ala128His + Thr215AspGly126Asn + Asn154Glu + Ser155Asp + Gly203AspSer 97Glu + Ser 98Asp + Ser131Glu + Asn162SerSer 97Glu + Ser 98Glu + Ser104Asp + Asn217GlnGly156Ser + Ser158Glu + Tyr205Asp + Thr215AspAsn 61Glu + Ser 97Glu + Ala128His + Tyr205AspGly 64Asn + Thr 65Glu + Phe188Tyr + Thr215GluTyr103Glu + Ile106Glu + Gly203Pro + Gly218GluGly 99Glu + Ser102Asp + Leu125Gly + Ser131GluSer102Glu + Asn154Glu + Ser190Asp + Tyr205ValGly127Glu + Gly165Glu + Tyr205Thr + Thr215GluVal 94Glu + Gly105Asp + Gly165Pro + Thr212GluAsn 96Glu + Ser 98Asp + Tyr103Leu + Ser155AspAsp 59Glu + Asn 61Glu + Gly105Glu + Gly130SerAsp 59Glu + Asn 61Asp + Thr 65Gly + Tyr205GluLeu 95Gln + Tyr103Asp + Gly203Asp + Thr215AspAsn157Asp + Gly203Asp + Thr215Asp + Thr219GlnSer 98Glu + Gly203Glu + Thr215Glu + Gly218SerSer131Asp + Asn154Gln + Gly203Asp + Thr215GluGly101Asn + Ile106Glu + Gly159Asp + Thr161AspGly127Glu + Ser129Glu + Gly130Ser + Thr215GluGly153Gln + Ser155Asp + Pro209Glu + Thr212AspGly203Glu + Thr210Asp + Leu216Asp + Gly218ProVal 94Ser + Ser 98Glu + Ser100Asp + Phe188AspAsn 96Asp + Ser100Asp + Gly153Ser + Asn162GluGly 62Glu + Leu 95Glu + Ile106Leu + Thr212GluSer160Asp + Asn162Asp + Ala202Glu + Thr215GlnAsn 61Glu + Ser 98Asp + Gly126Glu + Leu216AsnSer100Asp + Ser104Asp + Gly105Asn + Tyr205CysAsn 61Asp + Gly 99Glu + Tyr103Thr + Ser131AspVal 94Glu + Asn157Asp + Thr163Glu + Tyr205AlaGly127Asp + Ser131Glu + Thr132Asn + Ser190AspGly 60Glu + Gly156Ser + Asn211Asp + Thr212GlnGly126Asp + Gly156Glu + Ser158Glu + Tyr205MetAsn157Asp + Tyr205Val + Thr215Asp + Asn217GluAsp 59Glu + Gly203Glu + Tyr205Asn + Pro209AspGly105Glu + Ser131Asp + Gly156Asn + Leu216ProAsp 59Glu + Ser 97Glu + Asn157Asp + Phe188HisSer 97Asp + Ser100Asp + Ala202Pro + Thr212GluVal 94Gly + Ser104Glu + Asn157Asp + Ser160AspAsn 61Glu + Asn162Glu + Ser190Asp + Thr215GlyAsn 81Asp + Asn 96Glu + Gly127Ser + Gly130Asp
Thr 65Glu + Gly165Glu + Phe188Val + Ser190GluTyr103Pro + Asn154Glu + Ser158Glu + Tyr205ProSer131Glu + Gly156Glu + Tyr166Asp + Ala186ThrAsn 61Glu + Thr 65Asp + Tyr103Pro + Ser131AspGly130Glu + Asn154Asp + Ser187Asp + Tyr208HisSer131Glu + Thr163Pro + Gly203Glu + Gly218GluAsp 59Glu + Asn162Asp + Thr163Gln + Thr210GluLeu125Asp + Gly165Asp + Gly203Asp + Tyr213GlnGly101Asp + Asn162Asp + Gly165Glu + Leu216AsnSer155Glu + Ser158Asp + Tyr205Glu + Leu216ProGly 60Asp + Tyr205Pro + Thr212Asp + Thr215GluSer160Glu + Thr163Asp + Tyr205His + Thr215AspThr 58Pro + Asn 61Glu + Asn154Glu + Asn157AspAsp 59Glu + Gly 99Ser + Gly153Asn + Thr212AspSer 97Glu + Gly101Asp + Thr132Glu + Ala202AsnThr 58Glu + Asn 61Asp + Ala186Gly + Ser187AspThr 58Glu + Asn 61Glu + Ile106Cys + Thr132AspSer158Glu + Gly203Glu + Asn211Gln + Ala214AspSer129Asp + Gly203Glu + Ala214Glu + Thr215SerSer104Asp + Ser155Glu + Asn157Gln + Asn162GluThr 65Glu + Ser160Glu + Tyr205Cys + Thr210AspAsn 61Asp + Leu 95Glu + Tyr205Glu + Leu216AsnSer100Glu + Gly127Glu + Thr215Asp + Leu216ProSer102Glu + Gly127Glu + Ala128Pro + Asn162GluAsn 96Asp + Gly105Glu + Asn162Asp + Thr212GlnSer 98Glu + Gly127Asp + Ser131Glu + Ala202SerAsp 59Glu + Leu 95Asp + Gly159Asp + Tyr205ValThr 65Glu + Ser187Glu + Thr215Gln + Gly218GluGly156Glu + Gly159Glu + Tyr205Met + Thr215GluGly 64Gln + Gly130Glu + Thr132Gly + Gly153GluAsp 59Glu + Gly 99Glu + Ser129Glu + Tyr205AsnGly126Glu + Ala186Glu + Gly203Asp + Thr212ProThr 58Asp + Tyr103Val + Ser104Glu + Thr132AspSer104Glu + Thr132Glu + Ala202Glu + Asn217GlnSer 98Glu + Thr132Gly + Ser187Glu + Thr219GluGly 62Asp + Ser100Asp + Gly105Asn + Tyr205ValSer 97Asp + Ala186Asn + Thr212Glu + Thr215AspSer187Asp + Ala202Gly + Thr212Glu + Thr215GluAsp 59Glu + Ser 98Glu + Gly153Ser + Asn154GluGly126Glu + Thr132Asp + Ser155Asp + Ala202GlyAsp 59Glu + Gly 62Pro + Ser158Glu + Phe188GluSer104Asp + Gly130Asp + Tyr205Pro + Thr212GluThr 65Pro + Ser102Asp + Gly126Asp + Ala202AspGly 62Asp + Ser129Glu + Thr215Asp + Leu216ValAla128Asp + Asn162Asp + Ala202Ser + Tyr205GluSer 97Glu + Tnr132Gly + Gly153Asp + Ser187AspSer102Asp + Leu125Asp + Gly156Asp + Gly218Asn
Asn 61Gln + Ser160Glu + Ser190Asp + Gly203GluAsn 96Asp + Tyr166Asp + Ser190Glu + Leu216CysThr 65Asp + Asn162Glu + Tyr205Asp + Thr215ProThr 65Asp + Ser 97Glu + Gly105Glu + Leu216MetVal 94Asp + Ser 98Asp + Ser158Asp + Thr215GlnThr 65Glu + Gly101Asp + Thr215Glu + Gly218AsnThr 58Asp + Gly156Ser + Asn211Asp + Thr215GluGly 99Gln + Gly126Gln + Asn154Asp + Thr161AspGly156Asn + Ala186Asp + Thr212Glu + Leu216AspSer 97Asp + Ile106Gln + Leu125Asp + Thr215SerSer100Glu + Ser104Glu + Tyr166Ala + Ser187AspSer 98Asp + Ser102Glu + Thr215Glu + Leu216GlnSer129Asp + Thr132Gln + Ser155Glu + Ser160AspSer 97Glu + Thr163Asp + Ser187Glu + Thr219SerLeu125Cys + Ser129Glu + Ser155Glu + Thr215GlyAsn 61Asp + Gly126Glu + Ala214Asp + Thr219GlnThr 58Asn + Gly 64Pro + Thr 65Glu + Val 94AspTyr205Leu + Tyr213Asp + Thr215Gly + Asn217AspAsp 59Glu + Ala202Gln + Thr212Asp + Asn217AspAsp 59Glu + Ser187Asp + Pro209Asn + Thr212AspThr 58Asn + Leu 95Glu + Phe188Asp + Gly203AspSer187Glu + Tyr205Ile + Thr212Glu + Leu216GluSer129Glu + Gly156Gln + Thr212Asp + Leu216AspGly101Glu + Ser129Glu + Asn157Glu + Tyr205ThrGly 60Glu + Leu125Ser + Gly153Glu + Ala202AspAsn 61Glu + Phe188Glu + Tyr205Ala + Thr212GluAsn 61Glu + Gly153Asn + Gly159Asp + Thr212AspThr 65Gly + Tyr205Glu + Asn211Glu + Leu216CysThr 65Glu + Asn 96Glu + Gly156Asp + Leu216Cys表35多環(huán)五突變變異體Tyr103Cys + Gly156Glu + Ala202Gln + Thr212Gly + Leu216GlnAsn 61Ser + Gly101Asp + Thr132Gly + Gly165Gln + Tyr205ProGly 60Asn + Asn162Gln + Thr212Pro + Tyr213Gln + Asn217AspGly 60Ser + Leu 95His + Ser100Glu + Gly156Pro + Thr215GlyLeu 95Ile + Thr163Gln + Tyr205Glu + Tyr213Ala + Thr219AsnGly159Glu + Phe188Tyr + Gly203Pro + Tyr205Gly + Ala214ThrGly126Pro + Gly127Gln + Tyr208Asn + Thr212Gln + Leu216AlaLeu 95Gly + Asn162Ser + Gly165Glu + Pro209Gly + Thr210GlyThr 58Ser + Leu 95Asn + Ser100Asp + Tyr205Ala + Thr215AsnAsn 61Ser + Ser100Glu + Asn162Gln + Phe188Ser + Gly203SerTyr103Ser + Thr161Pro + Phe188Gln + Gly201Gln + Leu215ProSer 97Asp + Gly 99Pro + Tyr205Pro + Thr210Asn + Leu216HisLeu 95Met + Ser187Asp + Tyr205Val + Thr212Gly + Leu216AsnIle106Asp + Tyr166Thr + Thr207Pro + Asn211Gln + Leu216Thr
Asn 96Gln + Ser160Asp + Phe188Gly + Thr215Asn + Gly218AsnThr 58Pro + Ser 98Asp + Gly130Pro + Tyr205Val + Tyr208ProVal 94Ala + Gly 99Ser + Ala128Glu + Phe188Asn + Leu216GlyLeu 95Thr + Gly126Gln + Pro209Ser + Thr215Asp + Leu216GlySer 98Glu + Asn154Gln + Gly203Ser + Thr215Ser + Leu216ProThr 58Asp + Leu125Pro + Ala202Thr + Gly203Pro + Tyr205ValAla128Asp + Gly153Gln + Ala186His + Ala199Gln + Gly203AsnAsp 59Glu + Gly159Gln + Ala186His + Leu216Ser + Gly218SerAla186His + Phe188Gln + Ser190Glu + Pro200Ser + Tyr205ValVal 94Gly + Ser 98Asp + Tyr166Pro + Ala202Gly + Tyr205HisAsn 61Gln + Tyr103Met + Asn157Asp + Thr161Ser + Gly203AsnThr 65Gln + Gly203Gln + Pro209Ser + Thr215Gln + Asn217AspGly126Ser + Ser160Asp + Thr163Gln + Gly203Ser + Tyr205GlyTyr103Gln + Gly156Pro + Tyr166Asn + Phe188Tyr + Thr215GluVal 94Met + Gly 99Asn + Thr212Asn + Tyr213Gly + Thr215GluAla128Ser + Ala202Pro + Gly203Gln + Pro209Glu + Gly218SerThr 58Ser + Tyr103Gly + Leu125Gly + Tyr205Gly + Gly218SerLeu125Pro + Asn154Glu + Thr161Ser + Asn162Gln + Gly203GlnGly 60Asp + Gly 62Pro + Asn 96Ser + Ala186Gly + Thr210GlnAsn 61Gln + Thr 65Ser + Leu 95Met + Tyr103Gly + Ser104AspAsn 61Ser + Gly155Gln + Gly156Gln + Ala202Gly + Asn211GlnSer102Asp + Gly126Gln + Ala202Asn + Tyr205Leu + Thr212GlyGly 60Asn + Ser104Asp + Gly105Asn + Thr163Ser + Tyr205AlaVal 94Asn + Gly101Asn + Leu125Asn + Gly130Asp + Leu216CysGly159Pro + Gly165Pro + Ala186Thr + Gly201Pro + Leu216SerLeu 95Cys + Gly130Gln + Thr212Ser + Ala214Asn + Leu216ValGly 60Glu + Thr161Gly + Gly203Gln + Tyr205Ile + Thr212GlyGly153Ser + Gly159Asn + Ala186Pro + Gly203Ser + Thr219AsnAsp 59Glu + Gly 62Gln + Leu125Ala + Thr215Asn + Asn217GlnVal 94Gly + Leu125Ser + Gly203Glu + Thr215Gln + Leu216AsnAla186Gln + Ser187Glu + Gly201Gln + Ala202Pro + Gly203SerAsn157Asp + Gly203Gln + Thr212Pro + Ala214Gln + Leu216AlaThr 65Ser + Ser 98Asp + Thr161Ser + Gly203Ser + Tyr205CysGly 99Asp + Gly165Ser + Met198Cys + Pro209Gly + Thr215GlyGly153Pro + Tyr208Ile + Thr212Ser + Ala214His + Gly218GluSer 98Asp + Ile106Ala + Tyr166Leu + Ala202Gln + Ala214ThrGly 99Gln + Ser129Glu + Thr161Asn + Phe188Cys + Tyr205ThrGly 60Asn + Gly159Gln + Ser160Glu + Thr163Pro + Ala186GlyAsn 96Asp + Gly 99Pro + Val204Ala + Tyr205Val + Asn217GlnGly 60Ser + Phe188Ile + Thr212Asn + Thr215Ser + Leu216AspAla128Gly + Thr132Gln + Gly165Gln + Met198Thr + Gly203GluAsn 96Asp + Asn157Ser + Gly201Asn + Thr212Gly + Thr215GlyGly 99Asp + Phe188Val + Gly203Asn + Thr207Asn + Leu216ProAla128Pro + Thr132Pro + Thr163Pro + Phe188Thr + Thr212GluVal 94His + Gly105Glu + Ala186Pro + Thr210Gln + Thr212GlyGly101Pro + Thr132Asp + Asn162Ser + Thr212Gln + Leu216ProGly 60Asn + Tyr103Asn + Gly105Gln + Ile106Ala + Leu216Gln
Gly126Pro + Gly203Pro + Tyr205Val + Thr215Asn + Leu216SerThr 58Glu + Thr161Gly + Tyr205Asn + Tyr208Leu + Leu216ThrGly101Asn + Gly159Pro + Asn162Ser + Ala202Gly + Tyr205GlnTyr103Ile + Gly105Pro + Ala202Gln + Thr210Gln + Leu216ProGly 60Asp + Gly153Ser + Thr163Gly + Thr212Pro + Leu216MetGly101Pro + Ser104Glu + Gly105Pro + Gly153Ser + Tyr205ThrAsn162Asp + Phe188His + Ala199Gly + Thr212Pro + Leu216ProAla186Gly + Ala202Pro + Tyr205Met + Thr210Gln + Leu216ValGly 62Ser + Tyr103Met + Asn154Glu + Thr212Gly + Leu216MetThr 65Ser + Gly101Pro + Tyr103Leu + Thr161Asp + Thr219SerGly105Ser + Ile106His + Ala186Thr + Met198Pro + Thr212ProThr163Gly + Gly203Asp + Thr212Asn + Leu216His + Asn217GlnGly126Ser + Thr161Gln + Gly203Pro + Thr210Pro + Leu216ValThr 65Gln + Ser131Asp + Ala186Gly + Thr215Ser + Leu216HisThr132Gln + Asn162Ser + Gly165Asp + Tyr205Gly + Tyr208CysGly127Gln + Gly153Glu + Gly203Pro + Thr215Gly + Thr219GlyIle106Met + Leu125Cys + Gly126Ser + Ser131Glu + Gly156AsnSer 97Glu + Thr132Gly + Pro200Gly + Tyr205Ala + Asn211SerThr 65Asp + Tyr166Gln + Gly203Asn + Thr210Ser + Thr212GlnGly 99Ser + Tyr166Ser + Ser187Glu + Tyr205Gln + Leu216SerGly126Ser + Gly130Asn + Thr161Asp + Phe188Val + Thr207GlnLeu 95Pro + Pro200Ser + Tyr205Leu + Thr210Glu + Leu216CysTyr103Glu + Gly127Gln + Tyr166Thr + Ala202His + Thr210SerThr 58Gly + Ala202Pro + Pro209Gln + Thr212Ser + Ala214GluGly 62Gln + Tyr103Thr + Thr163Gly + Ser187Asp + Gly203GlnGly101Gln + Leu125Glu + Thr132Ser + Gly203Ser + Thr212GlnAsn 96Ser + Ala128His + Ala186Gln + Tyr205His + Thr215SerGly101Ser + Gly165Asp + Thr210Ser + Thr212Gly + Thr215GlyGly156Gln + Ala202Gly + Ala214His + Thr215Pro + Leu216ThrVal 94Cys + Gly 99Gln + Gly126Asn + Gly159Pro + Thr219ProLeu 95Pro + Gly105Pro + Thr161Asp + Gly203Pro + Leu216AsnGly 60Glu + Asn 61Gln + Thr 65Gln + Thr210Gly + Leu216GlnLeu125Gly + Ser187Asp + Phe188Gln + Ala202Asn + Thr212AsnPhe188Ile + Gly203Glu + Thr210Gly + Thr212Gln + Leu216ProAsp 59Glu + Leu 95Ile + Ile106Leu + Gly203Gln + Tyr213IleGly 99Gln + Ala186Pro + Tyr205Gly + Thr215Glu + Gly218GlnSer 98Glu + Thr161Pro + Ala202Gln + Tyr205Leu + Thr207GlnSer100Glu + Gly153Asn + Thr163Gln + Thr212Pro + Leu216GlnLeu 95Ser + Ile106Asp + Tyr205Val + Leu216Pro + Thr219AsnTyr166Val + Ser190Asp + Thr207Gly + Thr212Ser + Leu216AsnAsp 59Glu + Gly105Gln + Thr132Gln + Ala186Ser + Tyr205IleGly 60Pro + Tyr205Ile + Asn211Gln + Leu216Ile + Asn217GlnGly 99Asn + Leu125T r + Asn157Gln + Asn162Glu + Thr212GlnIle106Ala + Asn162Glu + Tyr166His + Asn211Ser + Thr215GlnAsn154Asp + Phe188Gly + Gly203Pro + Tyr205Ala + Pro209SerLeu 95Asn + Gly105Glu + Tyr166Ala + Tyr205Leu + Thr212SerVal 94Ala + Ser131Glu + Thr132Glu + Phe188His + Thr212Gly
Gly 64Gln + Gly159Asp + Ser160Glu + Thr215Pro + Leu216AlaThr 58Glu + Asp 59Glu + Pro209Gln + Thr212Asn + Thr215GlnAsn 96Asp + Ser 97Glu + Tyr205Thr + Ala214Thr + Leu216SerAsp 59Glu + Gly 60Glu + Gly159Asn + Thr210Pro + Gly218AsnAsp 59Glu + Gly 60Asp + Leu 95Ser + Gly203Pro + Tyr205SerTyr103Ile + Asn154Ser + Tyr205Glu + Ala214His + Thr215AspGly105Asn + Gly127Gln + Ala202Glu + Gly203Ser + Asn217GluAsn 61Asp + Thr 65Gln + Ser 97Asp + Tyr205Ala + Leu216SerAsn 61Glu + Ser 97Glu + Gly105Gln + Asn162Ser + Thr212SerAsn 61Asp + Leu 95Ser + Ser 97Glu + Val204Gln + Thr212GlyTyr103His + Asn154Glu + Phe188Ile + Ser190Glu + Thr212GlnVal 94Pro + Asn 96Asp + Ser 98Glu + Tyr103Pro + Thr212SerVal 94Met + Ser158Asp + Thr161Glu + Gly203Ser + Gly218SerGly203Asp + Tyr213Pro + Thr215Asn + Asn217Asp + Gly218AspSer160Glu + Asn162Glu + Tyr205Val + Asn211Gln + Thr215ProAsn 61Asp + Ser 98Glu + Tyr103Cys + Gly203Ser + Tyr213AlaGly101Asn + Tyr103Pro + Ile106Asn + Asn157Glu + Thr161AspGly130Asn + Ala186Glu + Ala202Glu + Thr215Pro + Thr219AsnGly126Asp + Ala128Glu + Thr132Ser + Met198Cys + Gly218SerVal 94Pro + Leu125Gly + Thr215Asp + Leu216His + Asn217AspGly 62Ser + Ile106Ala + Ala128Thr + Thr215Asp + Asn217GluGly105Pro + Ser129Glu + Thr132Asn + Gly165Glu + Asn211GlnAsp 59Glu + Asn 96Asp + Ser 97Glu + Phe188His + Thr210ProAsn 61Gln + Gly153Glu + Thr163Asp + Gly203Gln + Leu216SerAsp 59Glu + Thr 65Gly + Leu 95Gly + Ser 97Asp + Tyr166GlyAsn157Asp + Ser160Asp + Asn162Glu + Thr210Gly + Thr212SerSer 97Glu + Ser100Asp + Gly101Ser + Thr207Gln + Thr212AsnGly 64Ser + Gly203Ser + Thr212Asp + Ala214Asp + Thr215AspGly 60Asn + Thr132Asn + Ser155Glu + Gly156Glu + Asn162AspGly 99Ser + Tyr103Ile + Gly130Gln + Phe188Glu + Thr219GluGly156Asp + Thr161Asp + Ser190Asp + Gly203Ser + Ala214GlnGly101Asp + Ser104Asp + Gly127Ser + Thr210Gln + Thr212GlnGly130Gln + Ser155Asp + Tyr213Met + Gly218Asp + Thr219GluGly 99Ser + Asn154Ser + Gly203Asp + Leu216Asn + Gly218AspSer102Glu + Gly130Asp + Ser131Asp + Ala202His + Thr210AsnGly153Pro + Gly159Asp + Asn162Asp + Ala186Gln + Ala202GlnAsn 96Asp + Ser100Glu + Gly105Asp + Val204Thr + Thr215ProAsn162Asp + Gly165Asp + Thr212Gln + Thr215Ser + Leu216CysSer155Asp + Gly165Ser + Ser187Glu + Tyr205His + Thr212GlySer100Glu + Gly126Glu + Ala186Pro + Tyr205Asn + Tyr213GlnGly 62Pro + Ser104Glu + Ser131Asp + Thr212Gln + Tyr213ThrGly203Asp + Tyr205Asn + Asn211Gln + Thr212Gly + Ala214AspIle106Cys + Asn154Glu + Ala186Glu + Gly203Ser + Leu216AlaThr 58Pro + Asn 61Asp + Val 94Glu + Asn 96Asp + Gly165ProLeu 95Glu + Ser 97Glu + Ser102G1u + Thr161Ser + Tyr205LeuThr 58Gly + Ala128Pro + Gly156Asp + Thr161Asp + Gly203AsnGly 60Ser + Gly126Pro + Thr161Glu + Ser190Glu + Thr212Gly
Gly 60Ser + Ser102Glu + Gly105Asp + Ser131Glu + Gly203AsnAsn154Gln + Ala186Ser + Tyr213Glu + Leu216Glu + Thr219ProGly101Asp + Ile106Cys + Gly126Asp + Met198Gln + Thr219ProLeu 95Gln + Asn154Asp + Ser190Asp + Thr212Gly + Asn217GluThr 58Gln + Asn 61Glu + Ser100Asp + Tyr205Ala + Thr215GlyAsn 61Glu + Gly 99Gln + Ser100Glu + Tyr103Met + Leu125SerAla128Ser + Gly130Ser + Gly159Asp + Thr163Glu + Tyr20SCysAsp 59Glu + Gly 99Asp + Tyr103Ile + Ile164Thr + Thr215SerLeu 95Ile + Ser102Asp + Ser131Asp + Ile164Val + Leu216IleAsn 96Gln + Thr212Pro + Ala214Asp + Leu216Glu + Thr219GluIle106Glu + Ser131Asp + Ala202Gly + Tyr205Asn + Thr215SerIle106Glu + Gly130Ser + Ser131Asp + Phe188Ala + Tyr205ProThr 65Gln + Asn154Asp + Thr215Glu + Leu216His + Asn217GluAsn 61Gln + Gly 99Gln + Asn154Glu + Ser158Asp + Ser187GluGly101Ser + Gly126Asn + Thr212Glu + Thr215Asp + Leu216GlnAsp 59Glu + Thr 65Pro + Ser 98Asp + Tyr103Pro + Leu125IleGly153Ser + Ser155Asp + Gly165Asp + Ser187Asp + Asn217SerSer100Asp + Ser102Glu + Gly130Glu + Gly156Pro + Tyr205IleGly105Ser + Gly153Glu + Thr215Asn + Leu216Asp + Asn217GluThr 58Asp + Asn 96Glu + Ala186Gly + Gly203Pro + Thr219GlyAsn154Asp + Ser160Asp + Gly165Ser + Ser190Asp + Ala202SerAsp 59Glu + Ser 98Glu + Tyr103Asn + Thr132Ser + Pro209GluAla128Asp + Asn154Asp + Tyr166Ala + Pro200Gly + Tyr205MetGly101Gln + Asn157Gln + Gly203Asp + Thr215Gly + Thr219AspAsn 61Gln + Gly126Asp + Asn162Glu + Ser190Asp + Thr215GlyThr 58Gln + Asp 59Glu + Leu 95Gly + Gly203Pro + Asn211GluIle106Asn + Leu125Val + Ser129Glu + Thr163Asp + Ala214GlyGly126Gln + Gly127Asp + Ala128Ser + Ser155Asp + Ser158GluSer190Glu + Ala202Glu + Thr212Asn + Thr215Gly + Asn217AspSer155Glu + Gly156Glu + Ala202Asp + Thr215Gly + Thr219GlnSer160Glu + Ser190Asp + Gly203Gln + Tyr205Asn + Thr212AsnGly156Asn + Tyr166Ala + Gly203Glu + Thr212Glu + Leu216AspGly 99Gln + Ser102Glu + Gly126Asp + Asn154Glu + Tyr205AsnThr 65Pro + Gly127Glu + Ser131Glu + Ser155Asp + Tyr205CysAsn 96Gln + Thr161Gly + Ser190Glu + Thr210Glu + Asn211AspGly130Asn + Ser187Glu + Pro209Gln + Ala214Glu + Thr215GluGly101Glu + Ile106Met + Thr210Ser + Ala214Asp + Thr215GluSer104Asp + Thr210Gly + Thr212Gln + Ala214Glu + Thr215GluAsn 96Gln + Ser158Asp + Gly159Asp + Thr163Asn + Leu216GluAsn 96Gln + Ser158Glu + Gly203Pro + Thr215Asp + Leu216AspGly127Asn + Gly130Gln + Ser160Glu + Thr215Asp + Leu216AspGly 60Asn + Asn 61Glu + Gly 62Glu + Asn157Asp + Tyr205SerGly105Asn + Ala128Glu + Ser129Glu + Gly153Pro + Ala202AspThr 58Asp + Tyr103Met + Gly153Ser + Asn157Glu + Ser158GluGly 64Ser + Gly 99Asn + Ser129Asp + Gly130Asp + Asn157AspAsn154Gln + Ala202Asp + Asn211Glu + Thr212Asp + Leu216IleGly101Asp + Ser102Glu + Gly159Glu + Ile164Cys + Asn217Gln
Gly 62Pro + Gly101Asp + Ser102Glu + Ser190Glu + Thr215AsnThr 58Glu + Asp 59Glu + Gly130Glu + Ala214Gln + Leu216ValAsn 61Ser + Ser102Glu + Ser160Glu + Thr161Asp + Leu216ValSer102Asp + Ser160Glu + Thr161Asp + Tyr205Ser + Asn211GlnGly 62Pro + Ser104Glu + Gly105Asp + Leu125Met + Ala214AspVal 94Cys + Gly105Asp + Asn162Glu + Thr163Glu + Leu216IleSer 97Asp + Ser 98Asp + Ala202Asp + Tyr205His + Leu216ThrGly 62Glu + Asn 96Glu + Gly105Asp + Tyr166Ser + Pro209GlyVal 94Asp + Ala128Asp + Thr132Asn + Gly165Asp + Thr212SerSer102Glu + Gly105Glu + Asn154Asp + Gly156Asn + Leu216GlyTyr103Glu + Gly130Gln + Ile164Leu + Thr210Glu + Thr212AspAsp 59Glu + Asn157Ser + Tyr166Met + Gly203Asp + Asn217AspGly 60Gln + Gly130Asn + Ser155Glu + Thr215Glu + Asn217AspVal 94Thr + Gly 99Asp + Gly153Pro + Tyr205Glu + Ala214GluGly101Asn + Ser131Asp + Asn154Gln + Gly203Asp + Thr2l5GluGly101Pro + Gly127Pro + Asn157Glu + Gly203Asp + Thr215AspThr 65Gln + Gly153Glu + Gly165Glu + Gly203Asp + Leu216CysSer 98Asp + Ser100Glu + Ala128Gly + Gly159Asp + Gly165ProSer 98Asp + Ser100Asp + Ser187Asp + Gly203Asn + Thr212SerSer 98Glu + Ser100Glu + Ser155Asp + Asn162Ser + Thr210AsnAsn 96Asp + Ser100Asp + Gly127Asn + Tyr205Cys + Thr219AspAsn 96Asp + Ser100Glu + Gly159Asp + Tyr205Gln + Ala214ProLeu 95Gln + Gly101Pro + Ser102Asp + Gly153Asp + Tyr166GluAsn 96Ser + Gly159Asp + Ser190Asp + Tyr205Ile + Ala214GlyThr 65Glu + Ser 97Glu + Gly101Glu + Ala186Pro + Pro200GlyGly 99Asn + Gly126Asp + Ser158Glu + Ser160Glu + Tyr205GlnGly 62Glu + Leu 95Glu + Ile106Leu + Thr212Glu + Ala214SerAsn 61Ser + Gly 99Pro + Ser104Asp + Gly203Asp + Tyr205AspGly101Ser + Tyr103Ala + Gly203Gln + Tyr205Asp + Gly218AspSer160Asp + Asn162Asp + Phe188Pro + Thr212Glu + Leu216SerIle106Ser + Ser160Glu + Asn162Glu + Gly203Asp + Leu216ValVal 94Asp + Gly126Asp + Gly130Pro + Thr212Ser + Leu216AsnThr 65Pro + Ser155Asp + Asn157Asp + Ala186Thr + Leu216AspSer100Glu + Gly105Pro + Ala128Gly + Asn157Asp + Thr163GluGly126Glu + Ala186Asp + Ala202Asp + Gly203Gln + Leu216AsnThr132Gly + Ser160Asp + Ala186Glu + Ala202Asp + Tyr205GlnTyr103Pro + Ser129Glu + Ala186Glu + Ala202Glu + Thr215AsnLeu125Pro + Ser129Asp + Gly153Glu + Asn162Ser + Leu216IleAsp 59Glu + Thr 65Glu + Gly105Ser + Ser131Asp + Tyr213SerGly126Asp + Gly156Glu + Ser158Glu + Ile164Met + Tyr205MetAsn 96Glu + Tyr103Gly + Leu125Glu + Asn154Glu + Thr163GlnThr 58Glu + Ala128Ser + Gly156Asp + Thr163Asp + Thr210GlnGly 99Asp + Leu125Thr + Gly156Glu + Ser158Asp + Gly203ProGly101Glu + Gly156Asp + Ser158Asp + Ala202Gln + Pro209AsnSer131Glu + Gly156Asp + Ser158Glu + Phe188Leu + Gly203AsnGly 62Ser + Ser129Glu + Asn157Gln + Gly165Asp + Thr215GluSer 97Asp + Ser129Glu + Gly165Asp + Thr215Asn + Asn217Ser
Ala186Asp + Tyr205Cys + Thr212Asp + Ala214Asp + Thr215SerVal 94His + Ser158Glu + Thr212Asp + Ala214Glu + Thr215GlnLeu125Glu + Gly159Pro + Ala186His + Thr212Asp + Ala214GluAsn 61Asp + Tyr103Gln + Gly105Gln + Leu125Asp + Pro209AspGly 99Gln + Gly159Ser + Pro209Asp + Thr215Glu + Leu216AlaSer100Asp + Ser102Asp + Gly105Asn + Gly203Asp + Asn211GlnSer 97Glu + Ser100Glu + Gly165Glu + Phe188Val + Leu216AsnAsn 96Ser + Leu125Asp + Ser129Glu + Tyr205Gln + Thr215ProAsn 96Glu + Asn157Asp + Gly165Gln + Ser187Asp + Tyr205ValLeu 95Asp + Ser102Asp + Gly203Glu + Val204Ser + Leu216SerGly 62Glu + Thr 65Gln + Asn157Asp + Asn162Asp + Thr215GlnSer131Glu + Gly165Asp + Leu216Cys + Asn217Gln + Thr219AspVal 94Asp + Ser187Asp + Ser190Glu + Tyr205Asn + Thr215ProAsp 59Glu + Asn 96Gln + Ser187Glu + Ser190Asp + Thr215AsnSer158Asp + Asn162Asp + Gly203Ser + Leu216Glu + Asn217SerGly126Pro + Ser158Asp + Asn162Asp + Thr212Asp + Leu216ThrSer102Asp + Ser158Asp + Thr163Asp + Met198Gly + Thr207ProGly 60Glu + Leu125Val + Ser155Glu + Gly165Glu + Val204AsnSer131Glu + Asn154Glu + Ser187Glu + Tyr213Val + Asn217GlnGly 62Asp + Ala128His + Asn154Glu + Ser187Glu + Gly203SerAsn 61Gln + Gly 62Pro + Gly105Asp + Gly203Glu + Gly218AspSer102Asp + Gly203Glu + Ala214Gly + Thr215Pro + Gly218AspAsp 59Glu + Asn157Asp + Thr163Pro + Pro209Gly + Thr210AspLeu125Glu + Asn162Ser + Gly165Glu + Thr212Asp + Thr215GlnIle106Asn + Ser155Glu + Tyr166Val + Ser187Asp + Thr212AspGly105Pro + Ser155Glu + Ser187Asp + Tyr205Pro + Thr215AspAsn 96Glu + Ser155Glu + Ser158Asp + Thr161Pro + Thr215ProThr 58Asp + Thr 65Asp + Tyr103Val + Thr132Asp + Phe188GlyAsp 59Glu + Asn 96Glu + Ser190Glu + Gly203Ser + Ala214SerSer155Glu + Ile164Ser + Ala202Asp + Tyr205Asp + Leu216ProTyr103Ala + Ser104Asp + Ser131Asp + Ser160Asp + Tyr213LeuThr 58Glu + Asn 61Asp + Ala186Gly + Ser187Asp + Tyr205AlaAsn 96Glu + Ser102Glu + Gly153Asp + Gly203Pro + Asn217GlnAsn 96Glu + Ser102Asp + Gly105Pro + Asn162Glu + Tyr213ValSer129Asp + Gly153Asn + Gly203Glu + Ala214Glu + Thr215GlnSer100Glu + Gly127Glu + Gly203Glu + Tyr205Ala + Tyr213ThrSer158Asp + Ser190Asp + Gly203Asp + Thr212Pro + Thr215GlyLeu 95Ile + Thr132Gly + Ser187Asp + Tyr213Asp + Leu216GluGly 60Glu + Gly 62Pro + Ala128Glu + Ser155Glu + Ala202SerThr 58Asp + Ser 97Asp + Gly153Ser + Gly159Glu + Ala214ThrAsn 61Asp + Ser100Glu + Gly126Asn + Ser129Asp + Thr219SerAsp 59Glu + Leu 95Asp + Phe188Glu + Pro209Ser + Gly218SerTyr103Glu + Ser187Glu + Thr212Ser + Thr215Gln + Gly218AspSer104Glu + Ser190Asp + Tyr205Asn + Tyr208Val + Gly218AspAla128Asp + Ser131Glu + Thr161Gln + Tyr205Glu + Leu216MetSer102Asp + Ala128Glu + Ile164Gly + Gly203Asn + Thr212SerGly 62Asp + Gly105Glu + Tyr208Ala + Thr210Ser + Thr212Asp
Ser 97Glu + Ser102Glu + Ile106Ser + Asn157Ser + Leu216GlyGly 62Asn + Leu 95Met + Ser 97Asp + Ser102Asp + Asn217SerSer102Glu + Leu125Cys + Ser131Asp + Ala202Ser + Asn217GluSer102Asp + Ser131Glu + Thr163Asp + Thr207Gly + Thr215AsnSer 97Asp + Ile106Thr + Ala186Asp + Gly203Asp + Thr215GlyThr 58Asp + Gly127Asp + Ala128Thr + Ser155Asp + Gly203SerVal 94Gly + Gly127Asp + Ser155Asp + Gly156Gln + Tyr205GluIle106Glu + Gly126Asp + Gly127Asn + Gly156Glu + Ala202AsnSer 97Asp + Gly203Pro + Asn211Ser + Thr215Asp + Gly218GluGly 99Asp + Ser104Asp + Ile106Val + Thr132Asp + Thr215AsnGly 62Glu + Ser100Glu + Gly127Pro + Tyr205Ile + Thr212GlnGly 99Glu + Gly126Asp + Ala186Glu + Gly203Ser + Asn217SerThr132Asp + Tyr205Glu + Pro209Gln + Tnr212Glu + Asn217SerSer 97Asp + Gly127Gln + Ala186Asn + Thr212Glu + Thr215AspSer102Asp + Asn154Gln + Thr212Asp + Thr215Asp + Leu216GlyLeu125Gln + Ser129Asp + Asn162Gln + Thr212Glu + Thr215AspAsp 59Glu + Ser 98Glu + Leu125Gly + Ala128Asn + Ser129AspAsp 59Glu + Ser 98Asp + Gly156Asn + Ala202Asp + Gly218AsnAsn 61Ser + Asn157Glu + Gly203Asp + Tyr205His + Thr219AspIle106Asp + Gly130Asp + Tyr166Ile + Ala186Ser + Thr212AspAsp 59Glu + Gly156Asp + Thr161Asn + Leu216Ala + Gly218AspThr 58Asp + Asn 96Glu + Ser131Asp + Phe188Val + Thr215GlyLeu 95Val + Ser155Glu + Tyr166Ala + Ala186Glu + Thr210AspSer 98Asp + Leu125Cys + Ala128Asp + Thr161Gly + Asn162GluAsn 96Gln + Ile106Ala + Gly156Glu + Ser160Asp + Thr210GluAsp 59Glu + Tyr166Glu + Tyr205Asn + Thr210Gly + Asn211GluAsp 59Glu + Gly153Glu + Pro209Ser + Asn211Asp + Thr212ProAsp 59Glu + Tyr103Gly + Gly153Glu + Ser187Glu + Asn211GlnAsp 59Glu + Thr163Ser + Ser187Glu + Val204Thr + Asn217GluSer104Glu + Ile106Ala + Gly156Asp + Gly165Asp + Tyr205AsnGly126Glu + Ser131Asp + Asn154Ser + Tyr205Ser + Thr215AspAsp 59Glu + Gly101Gln + Thr212Glu + Ala214Pro + Leu216AspAsn154Ser + Ile164Cys + Thr212Asn + Ala214Asp + Gly218GluThr 65Glu + Ile106Asp + Gly127Asn + Thr215Asp + Leu216HisGly 60Gln + Gly 99Asp + Gly127Glu + Tyr205Glu + Thr215AsnThr 58Gln + Gly 60Glu + Ile106Val + Tyr205Glu + Asn211GluGly130Pro + Gly159Glu + Phe188Leu + Ser190Glu + Thr212GluAsn 61Glu + Gly127Asp + Ser190Glu + Tyr205Ala + Leu216SerSer187Glu + Pro209Asp + Thr212Asn + Ala214Ser + Thr215AspSer155Glu + Tyr166Glu + Gly203Glu + Val204Pro + Thr215SerSer 98Asp + Ser102Glu + Ala202Ser + Thr215Glu + Leu216GlnVal 94Asn + Gly101Asn + Leu125Glu + Ser129Glu + Thr212AspSer129Asp + Ser155Asp + Ala202His + Gly203Asn + Asn211SerAsn 96Asp + Tyr103His + Gly105Asn + Ala202Glu + Ala214GluLeu 95His + Gly 99Gln + Ser190Glu + Ala202Asp + Thr210AsnGly101Asp + Ala128Glu + Thr132Glu + Gly156Asn + Tyr166IleGly105Asp + Leu125Asp + Ala202Pro + Gly203Glu + Leu216Gln
Gly105Asp + Leu125Glu + Gly203Glu + Thr212Gly + Leu216CysGly 62Glu + Val 94Ala + Ser129Asp + Tyr205Asp + Thr212SerAsn 61Asp + G1y101Glu + Tyr205Glu + Thr215Pro + Leu216HisSer104Asp + Leu125Glu + Ser190Glu + Asn211Gln + Leu216AsnGly105Asn + Gly127Asp + Ala128Gly + Ser158Asp + Thr219GluSer 97Asp + Ser155Glu + Ser160Asp + Asn211Gln + Thr215SerAsp 59Glu + Gly 64Gln + Gly130Glu + Gly153Glu + Tyr205AsnAsn157Asp + Tyr205Leu + Thr212Glu + Thr215Gly + Leu216GluThr 58Glu + Gly101Glu + Gly105Pro + Gly130Ser + Tyr166AspSer 97Glu + Gly159Asp + Tyr166Ala + Ser187Glu + Leu216ValGly101Asp + Ser131Glu + Thr212Gln + Thr215Asp + Leu216IleGly153Asn + Asn157Asp + Tyr213Leu + Asn217Ser + Thr219AspGly153Asp + Ser158Glu + Tyr205Gly + Leu216Val + Gly218SerGly 99Ser + Ser187Asp + Gly203Glu + Pro209Asp + Asn211GlnLeu 95Ala + Ser102Asp + Ser187Glu + Gly203Asp + Thr219GlnGly 62Asp + Ala128Thr + Ser187Glu + Gly203Glu + Asn211SerGly101Asp + Ala128Thr + Asn154Gln + Phe188Asp + Leu216Glu表36多環(huán)六突變變異體Tyr103Cys + Gly126Gln + Gly156Glu + Ala202Gln + Thr212Gly +Thr215ProAsn 61Gln + Gly101Asp + Ile106Ala + Asn162Ser + Gly165Gln +Tyr205ProGly126Pro + Ala128His + Ser187Glu + Gly203Ser + Tyr208Asn +Leu216AlaThr 65Ser + Gly126Gln + Gly203Asn + Thr215Gln + Leu216Ala +Gly218AspIle106Ala + Gly165Asp + Tyr166Thr + Thr207Pro + Asn211Gln +Leu216ThrSer158Glu + Thr161Gly + Ala199Gly + Ala202Gln + Gly203Gln +Tyr205AsnThr 58Asn + Leu 95Asn + Gly153Pro + Tyr205Ala + Pro209Glu +Thr210ProGly126Glu + Ala128Gly + Asn157Gln + Gly165Asn + Gly203Asn +Tyr205ProGly 64Ser +Asn 96Gln + Gly126Asn + Gly127Ser + Asn211Ser +Thr212GlyGly126Asn + Ser158Asp + Gly203Pro + Tyr205Met + Thr215Gly +Thr219GlnLeu125Val + Gly156Asp + Tyr205Ser + Thr212Gln + Thr215Pro +Leu216HisGly105Asn + Gly126Ser + Ser160Glu + Thr161Asn + Tyr205Val +Gly218AsnGly105Ser + Ala128Thr + Thr132Pro + Ser160Glu + Tyr213Asn +Thr215ProAsp 59Glu + Gly105Pro + Asn157Gln + Ala186Asn + Tyr213His +Leu216HisGly 99Gln + Gly126Gln + Asn154Asp + Gly159Ser + Tyr205Ala +Thr212GlyThr 58Gln + Ser104Asp + Gly105Ser + Gly159Pro + Thr163Gly +Tyr205ProLeu125Met + Ser131Glu + Phe188Gln + Gly201Ser + Ala202Gly +Thr210GlyGly 60Asn + Gly 62Asn + Ser104Asp + Gly126Pro + Gly127Gln +Thr210AsnGly101Gln + Asn157Ser + Tyr166Pro + Ala202Ser + Asn211Gln +Thr215GlySer 97Aslp + Gly130Pro + Thr163Pro + Val204Thr + Tyr205Cys +Ala214ProTyr103Ala + Ala128His + Gly153Gln + Asn154Ser + Ser187Glu +Gly203SerAla128His + Tyr166Pro + Thr207Ser + Thr212Pro + Thr215Pro +Gly218GluAsn157Gln + Ser187Asp + Phe188Ala + Gly203Pro + Tyr205Ser +Thr212SerVal 94Pro + Tyr103Gly + Gly105Gln + Gly126Ser + Ala214Asp +Thr215GlyLeu 95Asn + Gly203Pro + Tyr205Met + Thr212Asn + Thr215Gln +Leu216HisLeu 95His + Tyr166Cys + Ala202Gly + Thr207Gln + Pro209Gly +Leu216HisGly 62Ser + Asn 96Gln + Thr132Gln + Gly159Ser + Gly165Pro +Thr219GluThr132Gln + Asn157Ser + Tyr166Val + Ala199His + Tyr208Gly +Thr215AspGly127Ser + Gly165Ser + Gly203Glu + Tyr205Ile + Tyr208Pro +Thr215SerGly156Glu + Pro209Asn + Asn211Ser + Thr212Ser + Tyr213Ile +Leu216HisTyr103Gln + Gly126Gln + Ala128Gln + Gly156Pro + Tyr166Asn +Ala202ThrThr 58Ser + Tyr103Gly + Leu125Gly + Thr161Ser + Asn162Gln +Tyr205GlyGly 60Ser + Gly101Gln + Gly127Asp + Gly203Pro + Pro209Gly +Thr215GlnGly105Gln + Leu125Asn + Ser187Glu + Pro200Gly + Ala214Ser +Leu216ThrAsn 61Glu + Leu 95Ile + Asn 96Ser + Tyr166Asn + Thr210Ser +Gly218SerGly 60Asn + Ser104Asp + Gly105Asn + Thr161Ser + Thr163Ser +Tyr205AlaVal 94Gly + Asn162Asp + Gly165Asn + Asn211Gln + Leu216Ile +Gly218SerSer 97Glu + Tyr103Met + Gly153Pro + Pro200Ser + Gly203Ser +Leu216MetThr 65Gln + Tyr103Leu + Ala128Asp + Ile164His + Tyr205Thr +Leu216SerGly127Glu + Phe188Gln + Gly203Gln + Pro209Asn + Leu216Met +Gly218ProGly 62Glu + Gly156Asn + Asn162Gln + Gly201Ser + Thr210Pro +Thr212AsnGly 60Pro + Tyr103Asn + Gly127Asn + Thr161Gly + Thr215Asp +Gly218GlnGly 60Asn +Gly159Gln + Ser160Glu + Thr163Pro + Ala186Gly +Pro209GlnAla202Thr + Gly203Ser + Tyr205Ala + Pro209Glu + Thr210Gln +Thr215GlnVal 94Asn + Ile106Asp + Gly126Pro + Tyr205Cys + Thr215Ser +Leu216CysVal 94Gln + Gly105Pro + Gly159Asp + Ala202Ser + Thr210Ser +Thr215SerAsn 61Ser + Asn154Asp + Ala186Thr + Pro209Ser + Tyr213Ile +Ala214GlnThr 58Pro + Gly127Ser + Gly159Ser + Ala186Glu + Thr215Pro +Thr219AsnAla128Gly + Ser129Glu + Thr132Gln + Gly165Gln + Met198Thr +Thr219GlnVal 94Gln + Asn157Asp + Tyr166Cys + Tyr205Ser + Thr207Pro +Thr215AsnAsn 96Gln + Thr132Glu + Ile164Ser + Thr210Pro + Thr212Gln +Leu216MetGly101Asn + Tyr103Ile + Gly159Pro + Asn162Ser + Ala202Gly +Thr210GlnGly 60Asn + Val 94Gln + Ser 97Glu + Gly203Gln + Tyr205Pro +Leu216MetTyr103Gln + Gly130Pro + Ser155Glu + Gly159Asn + Tyr205Pro +Thr212SerAsn157Gln + Gly203Asp + Pro209Ser + Thr215Ser + Leu216Cys +Thr219SerGly101Pro + Ile106Val + Gly126Glu + Ala128His + Asn157Ser +Thr219ProGly 64Pro + Gly203Ser + Tyr205Pro + Pro209Asp + Thr210Asp +Ala214GlnThr 65Gln + Gly203Pro + Pro209Gln + Thr212Pro + Gly218Glu +Thr219AspAsn 61Ser + Ser104Asp + Gly105Asp + Asn162Gln + Ala186Ser +Phe188SerTyr103Asp + Ser104Asp + Gly153Ser + Asn162Ser + Ala202Gly +Tyr208GlnAsn154Asp + Ser155Glu + Asn162Ser + Gly203Asn + Pro209Gly +Leu216AsnSer 97Glu + Ser 98Glu + Asn157Gln + Ala202Asn + Tyr205Asn +Leu216MetThr 65Gln + Gly 99Ser + Thr132Ser + Asn154Asp + Ser155Asp +Ser190AspAsn 96Asp + Gly 99Asp + Gly165Pro + Gly203Asn + Thr215Gln +Leu216ThrAsp 59Glu + Asn 61Asp + Val 94Asn + Gly101Ser + Asn217Gln +Thr219SerGly101Asn + Ile106Gln + Gly130Gln + Gly159Asp + Thr161Asp +Phe188SerSer 98Glu + Gly 99Ser + Ser100Glu + Tyr103Ala + Gly203Ser +Asn217GlnThr 65Gly + Val 94Met + Asn157Asp + Ser190Glu + Leu216Ser +Thr219SerGly130Asn + Ser160Glu + Asn162Glu + Ala186Asn + Gly203Pro +Tyr205GlnAsn 61Glu + Leu 95Pro + Ser 98Glu + Leu125Cys + Phe188Cys +Gly203ProSer129Asp + Ser131Asp + Gly203Pro + Tyr205Asn + Thr212Ser +Thr215GlnGly101Asp + Tyr103Asp + Leu125Thr + Thr161Asn + Thr212Gly +Leu216AsnGly105Pro + Ser129Glu + Thr132Asn + Gly165Glu + Ala202Asn +Asn211GlnAla186Pro + Phe188Met + Ala202Glu + Gly203Ser + Tyr213Val +Leu216GluIle106His + Gly130Ser + Tyr205Ser + Thr210Gly + Asn211Asp +Tyr213AspThr 65Asn + Gly203Gln + Pro209Gln + Thr212Asp + Tyr213Glu +Thr215AspGly 62Asn + Asn 96Ser + Tyr103His + Asn162Glu + Ser190Asp +Ala214AsnIle106Asn + Gly156Glu + Gly159Asn + Phe188Asp + Tyr208Gln +Gly218AspGly 60Pro + Gly 62Gln + Ala202Asp + Thr215Gly + Leu216Thr +Thr219GluGly153Asp + Gly156Pro + Ser187Glu + Phe188Asp + Gly203Gln +Ala214HisAla186Glu + Ala202Glu + Tyr205Ala + Tyr213Val + Thr215Asp +Asn217GluAsp 59Glu + Asn157Gln + Ala186Thr + Gly203Gln + Pro209Glu +Thr212AspAsp 59Glu +Asn 96Glu + Gly 99Glu + Gly101Glu + Tyr103His +Tyr205SerGly159Ser + Ser160Glu + Thr163Glu + Ile164His + Ala186Ser +Tyr205SerThr 58Asp + Gly 62Asp + Thr132Ser + Asn154Gln + Gly203Pro +Tyr205HisAsn154Ser + Ser158Asp + Ser190Glu + Thr212Gly + Thr215Gln +Gly218ProThr 58Asp +Asn 96Ser + Ser 97Asp + Ile106Pro + Thr132Gly +Thr212ProSer155Asp + Ser160Asp + Asn162Glu + Phe188Gly + Thr210Gly +Thr212SerAsp 59Glu + Gly 99Asp + Ser100Asp + Tyr103Ile + Ile164Thr +Thr215SerGly127Gln + Ser187Glu + Gly203Gln + Thr212Gln + Thr215Asn +Gly218GluGly 60Glu + Ser 97Glu + Gly 99Asn + Tyr166Asn + Thr210Glu +Ala214HisLeu 95Ser + Tyr103Met + Leu125Glu + Gly165Glu + Ser190Glu +Gly218ProAsn 96Asp + Ser 97Glu + Ser100Asp + Ser104Glu + Thr132Asn +Leu216HisLeu 95Ile + Ser104Glu + Ala128Glu + Ser129Glu + Ser131Asp +Asn157GlnAsp 59Glu + Asn 61Asp + Ser100Glu + Gly203Asn + Tyr205Ile +Thr212GlnThr 58Asn + Thr132Ser + Asn157Ser + Thr163Glu + Phe188Glu +Tyr205ProSer129Glu + Asn154Glu + Gly156Asn + Asn162Ser + Gly165Asp +Leu216CysGly 60Glu + Gly153Asn + Gly159Ser + Tyr205Asp + Ala214Asp +Thr215GluVal 94Glu + Ser102Asp + Gly105Asp + Ala128Asp + Thr215Gly +Leu216GlyGly126Asp + Gly156Glu + Ile164Met + Phe188Leu + Ser190Glu +Tyr205AlaGly 62Gln + Ser155Asp + Ser190Asp + Leu216Glu + Asn217Ser +Thr219GluVal 94Cys + Gly203Asp + Val204Asn + Tyr205Ile + Thr212Glu +Thr215AspAsn 61Ser + Val 94Gln + Leu 95Gln + Gly203Asp + Thr212Glu +Thr215AspAsn 61Gln + Gly 99Gln + Gly153Gln + Asn154Glu + Ser158Asp +Ser187GluGly 99Glu + Gly101Gln + Gly126Asp + Phe188Met + Tyr205Ser +Thr212ProGly101Asn + Tyr205Gly + Thr212Asp + Tyr213Met + Thr215Asp +Teu216IleGly130Pro + Asn154Glu + Ser160Glu + Ser187Asp + Ser190Glu +Tyr205LeuLeu125Pro + Asn154Glu + Thr161Ser + Ala202Glu + Gly203Gln +Leu216GluGly101Asp + Ile106Ala + Gly153Asp + Ser190Asp + Leu216Asn +Thr219AspGly 60Gln + Tyr103Gly + Tyr205Leu + Thr212Glu + Thr215Glu +Asn217AspGly 62Gln + Asn154Asp + Ser155Asp + Ser160Asp + Gly165Glu +Tyr205MetGly 62Glu + Asn154Gln + Tyr205Ala + Tyr213Met + Thr215Asp +Leu216GlnAsn154Ser + Asn157Asp + Ala202Asp + Gly203Glu + Tyr205Asp +Thr219GlyLeu 95Gln + Gly126Glu + Asn154Ser + Gly159Asn + Ser190Asp +Gly218AspAsn 61Ser + Thr 65Asp + Thr132Asn + Gly203Asp + Tyr205Asp +Thr215AsnAsn 61Glu + Leu 95Asp + Ser 97Glu + Thr132Asp + Thr161Pro +Thr163SerGly126Ser + Ser131Glu + Gly203Glu + Tyr205Ala + Leu216Asp +Gly218GluGly 60Glu + Ser100Asp + Gly101Asp + Ile106His + Gly130Gln +Ala214HisGly 62Pro + Gly101Asp + Ser102Glu + Gly126Glu + Ser190Glu +Thr215AsnAsn 61Asp + Gly 99Asp + Ser100Glu + Thr161Glu + Pro209Gln +Ala214AsnThr 58Gln + Asp 59Glu + Leu 95Gly + Gly203Pro + Asn211Glu +Leu216GlnAsn 96Gln + Gly156Asp + Gly165Glu + Tyr205Asn + Thr215Asn +Gly218AsnGly 60Pro + Ser102Glu + Tyr103Glu + Thr163Glu + Tyr166Asp +Gly203SerGly127Gln + Gly153Gln + Asn157Glu + Ser160Glu + Thr163Glu +Asn217GluAsn 61Gln + Ser102Asp + Gly126Asp + Ser155Glu + Ser190Asp +Ala202SerLeu125Pro + Gly156Asp + Phe188Glu + Tyr205Cys + Thr212Gly +Leu216AspThr 58Gly + Asn 61Glu + Gly126Ser + Thr212Pro + Leu216Asp +Gly218GluVal 94Ser + Gly156Asp + Thr161Ser + Thr212Gly + Thr215Asp +Leu216GluVal 94Gly + Gly101Glu + Gly159Asp + Ser160Asp + Ala186Pro +Leu216ProAsn 61Asp + Gly 62Glu + Leu125Gln + Asn154Asp + Gly159Asn +Thr212GlnAsn 96Gln + Gly101Asp + Ser102Glu + Gly159Glu + Ile164Cys +Asn217GlnThr 58Asp + Asp 59Glu + Ser190Glu + Gly203Gln + Tyr205Cys +Pro209SerGly 62Pro + Gly105Glu + Ile106Glu + Asn154Asp + Leu216Gln +Thr219AsnVal 94Ser + Ala128Asn + Ala202Glu + Gly203Asp + Tyr205Cys +Thr212AspAsn 61Glu + Tyr103Glu + Ser104Glu + Gly203Asn + Thr212Asn +Thr215ProThr 65Ser + Ser 97Asp + Ser 98Glu + Ser104Glu + Thr163Gln +Gly203ProGly126Asp + Ala128Pro + Gly156Asp + Ser160Glu + Thr161Asp +Gly203AsnThr 65Gly + Ser100Glu + Gly101Asn + Leu125Glu + Ser129Glu +Thr215ProAsn 61Glu + Gly159Ser + Ala186Gly + Pro209Asp + Asn211Gln +Thr215GluVal 94Ser + Ser100Glu + Gly101Gln + Gly126Asp + Ser131Asp +Gly203SerAsn 61Asp + Thr 65Gln + Ser 97Asp + Gly203Glu + Tyr205Ala +Leu216ValGly156Ser + Ser158Glu + Ala186Ser + Tyr205Asp + Thr212Asp +Thr215AspAsp 59Glu + Gly 60Glu + Tyr205Ala + Tyr213Glu + Thr215Glu +Leu216CysThr132Ser + Thr163Asp + Ser190Glu + Tyr205Gly + Pro209Asp +Thr215AsnGly 62Glu +Asn 96Glu + Ser 98Glu + Ser155Asp + Tyr166Ala +Leu216ValSer160Asp + Tyr205Ala + Thr210Glu + Thr212Asp + Leu216Ser +Gly218Glnlhr 65Pro + Leu 95Glu + Gly 99Glu + Ile106His + Leu125Gly +Ser131GluGly 60Glu + Thr 65Gly + Gly 99Ser + Gly203Glu + Tyr213Gln +Thr215GluGly 62Asp + Asn 96Gln + Gly101Glu + Leu125Cys + Gly127Ser +Pro209AspSer 98Asp + Ser100Asp + Ala202Gln + Thr210Glu + Thr212Gly +Thr215AsnSer 98Glu + Ser100Asp + Ala128Gln + Gly130Gln + Thr210Gln +Asn217AspLeu 95Pro + Gly130Asn + Ser155Glu + Phe188Glu + Tyr205Asn +Thr215GluAsp 59Glu + Val 94Glu + Ser 98Asp + Ser104Glu + Gly126Ser +Tyr213GlySer100Asp + Ser102Asp + Gly127Asn + Tyr205Asp + Thr215Asp +Leu216HisGly127Asp + Ala186Gly + Ser190Glu + Tyr205Val + Thr212Gln +Leu216ThrLeu 95Glu + Ser 97Glu + Ser102Glu + Gly159Asp + Tyr205Leu +Leu216AlaGly 60Glu +Asn 61Glu + Leu125Asp + Gly165Glu + Phe188His +Gly203AsnSer102Glu + Ser104Asp + Ala202Thr + Gly203Ser + Thr212Glu +Ala214GluGly 62Glu + Leu 95Glu + Ile106Leu + Tyr166Ile + Gly203Ser + Thr212GluAsn154Ser + Ala202Asn + Gly203Asp + Tyr205Glu + Thr210Glu +Leu216AsnAsn 61Glu + Ser 98Asp + Gly 99Ser + Gly101Glu + Asn154Asp +Gly203GlnSer160Asp + Asn162Asp + Ala202Glu + Tyr208Thr + Asn211Ser +Thr215GlnGly101Asp + Ser160Asp + Asn162Glu + Thr163Ser + Tyr205Asn +Thr212GlnVal 94Asn + Ser160Glu + Asn162Glu + Asn211Gln + Thr212Asp +Thr215ProThr 58Asn + Ser 97Glu + Ser155Asp + Asn157Asp + Thr212Ser +Leu216CysSer104Glu + Leu125Thr + Ala186Thr + Tyr205Glu + Pro209Asn +Leu216AspAsp 59Glu + Gly 64Gln + Ser 97Asp + Ser100Asp + Ser160Asp +Tyr213ValGly101Pro + Gly130Asp + Asn154Ser + Thr215Glu + Leu216His +Asn217GluAla186Asp + Gly203Asn + Tyr205Cys + Thr212Asp + Ala214Asp +Thr215SerGly 99Asp + Ile106Gly + Ala128Asn + Asn154Gln + Thr212Asp +Ala214GluThr 65Gln + Asn 96Ser + Ser100Asp + Gly126Pro + Ala202Asp +Leu216AspThr 65Ser + Ser129Asp + Gly153Asn + Tyr205Leu + Tyr213Asp +Thr215AspAsn 61Ser + Ser104Asp + Ile106Glu + Ala128Asp + Thr163Asp +Thr212GlnGly105Glu + Ser131Asp + Gly156Asn + Tyr205Ala + Thr215Pro +Leu216ProGly 62Asp + Ser100Asp + Ala128Asn + Tyr166Ala + Gly203Asp +Leu216AspGly 60Asn + Asn 61Glu + Gly 62Asp + Ser187Glu + Ala202Glu +Tyr205SerSer 97Glu + Ser100Asp + Leu125Asp + Gly203Pro + Thr207Gln +Thr215AspAsn 96Ser + Ser 98Asp + Ser102Glu + Tyr205Gln + Thr212Gln +Thr215GlyGly 64Ser + Ser 98Glu + Ser100Glu + Ser155Asp + Ser187Asp +Ala199GlnSer104Asp + Leu125His + Ser129Asp + Ser131Asp + Ala202Glu +Gly218AsnSer155Glu + Thr163Glu + Ala202Glu + Tyr205Glu + Thr207Gly +Gly218SerAsn 61Ser + Thr132Gln + Ser155Glu + Gly159Asp + Thr212Pro +Thr215GlnSer100Glu + Gly127Glu + Gly130Asn + Gly203Asp + Thr215Asp +Leu216ThrAsn162Asp + Phe188His + Ser190Asp + Pro209Glu + Thr212Pro +Leu216ProSer129Glu + Gly130Asp + Gly153Asn + Asn162Ser + Asn211Glu +Ala214GluGly 60Asp + Thr 65Asn + Ser 98Asp + Gly159Asn + Leu216Asp +Gly218GluGly 62Pro + Ser131Glu + Gly165Glu + Tyr205Ser + Leu216Thr +Thr219AspGly 62Glu + Asn 96Asp + Ser100Asp + Thr132Glu + Thr215Pro +Gly218ProThr 58Asn + Gly105Gln + Ser187Asp + Ser190Glu + Thr212Glu +Leu216ProGly127Glu + Ser129Glu + Gly153Ser + Asn154Glu + Thr212Gly +Thr215GluThr 65Pro + Ser102Asp + Ser104Glu + Gly126Asp + Asn162Ser +Ala202AspThr 58Asn + Gly 64Pro + Thr 65Glu + Val 94Asp + Tyr205Asp +Thr212AspSer102Glu + Ser104Asp + Gly203Pro + Thr207Pro + Thr212Asp +Thr215AspGly 62Asp + Ser104Glu + Gly159Gln + Thr210As1 + Thr212Pro +Leu216ProGly105Gln + Gly153Ser + Gly156Asp + Ala186Glu + Ala214Glu +Thr215ProThr 58Gln + Leu 95Pro + Ser155Glu + Gly165Glu + Gly203Pro +Leu216GluGly 99Glu + Tyr103Pro + Asn154Glu + Ser158Glu + Ser187Glu +Tyr205ProAsn 96Asp + Ser 97Asp + Leu125Val + Tyr166Asp + Ser190Glu +Leu216CysLeu 95Thr + Asn154Asp + Ser187Asp + Phe188Asn + Thr212Asp +Thr215GlySer131Glu + Asn154Glu + Ser187Glu + Tyr205Gln + Tyr213Val +Asn217GlnGly 62Asp + Ser129Glu + Gly203Asp + Tyr213Leu + Thr215Asp +Leu216ValGly101Asp + Ala128Asp + Ser155Asp + Ser187Asp + Thr207Gly +Thr212GlnSer131Asp + Asn162Ser + Gly203Asp + Thr212Ser + Thr215Asn +Gly218GluGly101Gln + Gly130Pro + Gly159Glu + Ser190Glu + Asn211Glu +Thr212GluTyr103Asn + Ile106Asp + Tyr166Ala + Ala186Glu + Leu216Pro +Thr219GluAsn 96Glu + Ile106Glu + Ala128Ser + Ser158Asp + Ser160Asp +Ala214AsnThr 58Asp + Val 94His + Ser155Glu + Ser158Asp + Gly159Asn +Tyr213ProAsp 59Glu + Val 94Pro + Asn 96Asp + Tyr103Pro + Gly159Glu +Leu216ValSer 97Asp + Asn154Glu + Asn157Ser + Thr163Glu + Ala202Ser +Asn217GluLeu 95Gly + Gly127Asp + Ser131Glu + Asn154Ser + Thr161Asn +Asn162AspGly 60Asp + Leu125Gly + Gly126Ser + Gly156Gln + Thr212Asp +Thr215GluSer100Glu + Ala128Glu + Gly165Pro + Phe188Tyr + Gly203Ser +Tyr205GlyLeu 95Gly + Asn157Asp + Asn162Ser + Gly165Glu + Pro209Gly +Thr210GlySer104Glu + Ser131Glu + Thr132Gly + Gly156Glu + Phe188Val +Leu216SerGly 99Glu + Ser100Asp + Ala128Glu + Pro209Gly + Thr212Asn +Ala214GluThr 58Asn + Asn 96Glu + Ser102Asp + Tyr103Met + Ser155Asp +Gly156GlnLeu 95His + Gly126Glu + Gly165Gln + Ser190Glu + Tyr213Asp +Thr215AspGly105Glu + Leu125Cys + Ser129Glu + Gly165Glu + Thr212Ser +Thr219GluSer 98Asp + Ser102Asp + Gly105Asp + Phe188Leu + Tyr205Glu +Thr212ProLeu 95Val + Gly 99Asp + Tyr103Val + Gly105Asp + Tyr205Asp +Thr215AspGly159Glu + Gly165Asp + Thr215Ser + Leu216Cys + Asn217Glu +Thr219AspTyr103Ile + Ser129Asp + Gly159Asp + Phe188Glu + Ser190Asp +Ala202GlyAsp 59Glu + Asn 96Asp + Gly105Glu + Asn162Asp + Thr163Gln +Thr212GlnGly 99Pro + Gly101Glu + Ser104Glu + Gly153Glu + Asn162Asp +Thr215GlnAsn 61Glu + Gly126Asp + Gly159Gln + Asn162Glu + Ser190Asp +Thr215GlyGly 99Asp + Ser158Asp + Asn162Asp + Thr163Asn + Ser187Glu +Ala199GlyAsn154Glu + Ser155Glu + Gly156Gln + Thr210Asp + Leu216Asp +Gly218ProGly126Asn + Ser129Asp + Asn162Glu + Ala186Ser + Ala202Glu +Asn217AspSer100Asp + Gly101Asp + Ala128Thr + Asn154Gln + Phe188Asp +Leu216GluAsn 61Ser + Ile106Ser + Ser158Glu + Ser190Asp + Tyr205Asp +Thr219ProThr 58Ser + Val 94Ala + Ser 98Glu + Ser158Glu + Ser190Glu +Tyr205ProSer100Glu + Gly127Glu + Asn154Glu + Gly203Glu + Tyr205Ala +Tyr213ThrGly101Glu + Leu125Gly + Ser155Asp + Tyr205Val + Leu216Pro +Thr219AspSer 97Asp + Asn157Gln + Gly165Ser + Tyr205Ser + Leu216Asp +Thr219AspSer 97Glu + Ser 98Asp + Gly105Glu + Gly165Gln + Ser190Glu +Thr215SerGly 60Gln + Thr 65Gln + Val 94Asp + Leu125Glu + Ser131Asp +Ala202GlyTyr166Glu + Ser187Asp + Ser190Glu + Gly203Pro + Tyr205Gly +Leu216AspThr 58Pro + Val 94Glu + Ser 97Asp + Gly165Glu + Tyr205Ser +Tyr213HisGly 62Asp + Val 94Ala + Ser129Glu + Gly203Gln + Tyr205Val +Thr212GluTyr103Asn + Gly130Glu + Ser131Asp + Ser155Glu + Phe188Met +Thr212AspSer155Asp + Gly203Asn + Pro209Asp + Thr212Asp + Thr215Ser +Asn217GluAsn 96Glu + Ser160Glu + Asn162Ser + Asn211Gln + Ala214Glu +Thr215AspAsn 96Glu + Ser158Asp + Gly159Asp + Thr163Asn + Thr210Pro +Leu216GluThr 58Asn + Ser129Asp + Gly130Glu + Gly156Glu + Thr161Ser +Tyr205AspGly153Gln + Ser158Asp + Ala202Asp + Asn211Glu + Thr212Asp +Leu216HisSer104Glu + Leu125Cys + Gly156Pro + Ala202Glu + Gly203Asp +Pro209GluThr 65Asp + Ser100Glu + Gly105Asp + Gly127Ser + Gly203Ser +Tyr208MetSer 97Asp + Ser 98Asp + Ser158Asp + Thr212Gln + Thr215Pro +Leu216GluGly 62Gln + Ser 97Glu + Ser 98Asp + Gly126Glu + Gly130Pro +Thr212GluGly 60Asp + Asn157Glu + Tyr205Glu + Tyr213Thr + Thr215Asp +Leu216IleGly 62Glu + Ser102Asp + Gly127Glu + Ser155Asp + Tyr213Pro +Ala214GlnThr 58Pro + Asn 96Asp + Gly127Glu + Ser155Glu + Asn162Ser +Thr212SerGly 62Gln + Ser100Glu + Gly130Glu + Asn154Gln + Gly203Glu +Asn217GluAsp 59Glu + Gly 62Pro + Phe188Glu + Ala202Ser + Tyr205Glu +Pro209AspAsn 61Glu + Ser 97Glu + Ala128His + Asn162Gln + Tyr166Glu +Tyr205AspIle106Pro + Gly126Pro + Gly153Glu + Ser158Glu + Gly203Glu +Thr215GluSer102Glu + Tyr103Pro + Gly105Ser + Gly130Glu + Ser158Glu +Ala199ProAsp 59Glu + Asn 61Ser + Ser158Asp + Thr161Asn + Ser190Glu +Asn211AspAsn 61Gln + Ser104Asp + Gly127Glu + Asn157Glu + Asn162Glu +Tyr205AlaVal 94Asp + Gly105Glu + Gly165Asp + Ala186Pro + Ala202Asp +Gly203GlnAsn 61Glu + Val 94Gln + Ser102Asp + Leu125Asp + Gly159Gln +Gly165SerIle106Asp + Ser131Glu + Asn162Glu + Ala202Ser + Thr212Ser +Thr215AsnIle106Asp + Ser131Asp + Gly159Asp + Gly203Pro + Leu216Ala +Thr219GlnAsn154Gln + Gly159Asp + Ser190Asp + Ala214Glu + Leu216Pro +Gly218AspAsn 61Asp + Ser158Asp + Gly203Asp + Thr212Gln + Ala214Gly +Thr215GluGly 62Asp + Thr 65Glu + Leu 95Met + Ser104Asp + Thr132Gln +Asn162AspAsn 96Asp + Ser 98Asp + Gly127Asn + Ser129Asp + Tyr205Asn +Thr215AspSer 97Asp + Gly126Pro + Thr161Ser + Ala186Glu + Phe188Glu +Thr212GluGly 60Asn + Ser 98Glu + Gly159Asn + Gly203Glu + Thr212Asp +Asn217GluSer131Asp + Ser190Asp + Gly203Asp + Tyr205His + Thr215Asp +Leu216GlnAsn 61Glu + Gly105Gln + Ser131Asp + Met198Gly + Pro209Gly +Thr210GluThr132Pro + Gly153Asp + Asn162Glu + Gly203Asn + Pro209Gln +Leu216GluSer100Glu + Gly105Pro + Asn157Asp + Gly203Glu + Asn211Ser +Leu216GluThr 58Asn + Ser 98Glu + Gly156Glu + Gly203Asp + Ala214Gln +Leu216AspAsp 59Glu + Gly 99Gln + Ser100Asp + Gly101Pro + Ser102Glu +Gly203GluSer 98Asp + Gly126Gln + Thr161Asn + Ala202Asn + Thr212Glu +Thr215GluSer160Asp + Thr161Asn + Thr212Glu + Thr215Glu + Leu216Val +Thr219SerSer 98Glu + Ser100Asp + Ser160Asp + Phe188Asp + Val204Pro +Asn211GlnLeu 95Gly + Ser 98Glu + Ser100Glu + Gly101Asn + Gly203Asp +Thr210GluAsn 96Glu + Ser100Asp + Asn157Asp + Thr163Pro + Ala202Pro +Thr212GluAsp 59Glu + Gly 99Asp + Ser104Glu + Gly127Glu + Gly203Asn +Tyr205GlyGly 62Glu + Thr132Gln + Gly156Glu + Thr163Asp + Ala186Pro +Ala202AspSer 98Asp + Ser158Glu + Ser160Asp + Phe188Ile + Pro209Asn +Gly218GluThr 58Asn + Gly 62Glu + Gly 64Asn + Asn157Glu + Ser190Glu +Gly203GluAsn 61Asp + Gly 99Pro + Ser104Asp + Gly203Asp + Tyr205Asp +Thr212ProGly 60Asp + Ile106Pro + Gly127Glu + Ala202Asp + Leu216Ile +Gly218AspAsn 61Asp + Ser129Glu + Ser131Asp + Thr161Asn + Ala214Gln +Thr215GluAsn 61Asp + Ser 98Asp + Ile106Ser + Ser131Asp + Ser160Glu +Ile164HisSer100Glu + Ser160Glu + Tyr205Asp + Thr210Pro + Thr212Ser +Leu216GluLeu 95Pro + Gly105Asp + Thr161Glu + Tyr205Glu + Leu216Asp +Thr219GlySer 98Asp + Ser102Glu + Leu125Glu + Ala128Pro + Tyr213Gly +Leu216AspLeu 95Val + Ser155Glu + Tyr166Ala + Ala186Glu + Thr210Asp +Leu216MetThr 58Asn + Gly 99Glu + Gly105Glu + Gly127Glu + Ser190Asp +Tyr205LeuAsp 59Glu + Gly 99Glu + Leu125Ser + Thr212Glu + Leu216Asp +Gly218SerThr 58Gln + Gly130Glu + Thr132Glu + Gly203Asn + Thr212Glu +Asn217AspSer 97Glu + Gly127Gln + Ser129Asp + Gly156Asp + Ser158Glu +Asn211SerSer102Glu + Gly126Glu + Gly153Ser + Ser190Glu + Ala202Thr +Asn211AspTyr103Glu + Leu125His + Ser131Glu + Gly203Asp + Asn211Asp +Thr215SerSer129Asp + Gly130Gln + Gly165Asp + Gly203Asp + Thr212Glu +Leu216ValGly127Asn + Ser131Asp + Gly159Glu + Tyr205Gln + Thr212Asp +Ala214GluSer 97Glu + Ile106Glu + Tyr166Gly + Ala202Asp + Thr215Ser +Leu216AspLeu 95Glu + Leu125His + Gly126Asp + Tyr205Gln + Thr215Asp +Thr219AsnAsp 59Glu + Ser 97Glu + Ala128Asn + Thr132Glu + Asn157Asp +Tyr205GlySer 98Asp + Ser102Asp + Ala128His + Ser187Glu + Thr215Pro +Gly218AspThr 58Pro + Gly130Ser + Gly156Asp + Ser160Asp + Thr212Ser +Thr215GluThr 58Ser + Gly 60Glu + Thr 65Asp + Ser100Glu + Gly159Glu +Leu216HisSer 97Glu + Ser100Glu + Asn154Glu + Gly203Pro + Thr212Asn +Thr215GluSer 97Asp + Ser100Asp + Ile106Val + Ser155Asp + Gly203Pro +Thr215AspSer 97Glu + Asn157Glu + Ser160Asp + Thr210Gln + Thr212Asn +Asn217AspAsp 59Glu + Leu 95Cys + Ala128Asn + Thr132Gly + Gly153Asp +Ser187AspThr 65Asp + Leu125Asp + Ser190Glu + Gly203Asn + Asn211Asp +Thr219SerAsp 59Glu + Asn 96Asp + Leu125Cys + Thr215Asp + Leu216Ile +Thr219AspSer 98Glu + Gly105Asp + Leu125His + Asn162Asp + Ser190Glu +Ala202SerGly 60Ser + Thr 65Glu + Ser158Glu + Thr163Gly + Thr215Pro +Leu216AspVal 94Glu + Asn 96Ser + Gly101Pro + Ser102Asp + Ser155Glu +Thr212AspSer100Glu + Gly127Asn + Ser129Asp + Thr132Glu + Asn162Glu +Leu216AlaSer 98Glu + Ser102Asp + Ala202His + Thr212Pro + Thr215Asp +Gly218AspAsn 61Asp + Asn 96Asp + Gly165Glu + Tyr166Ser + Gly203Asp +Asn211GlnAsn 61Asp + Asn 96Glu + Gly127Ser + Gly130Asp + Gly203Glu +Thr212ProSer 97Glu + Gly101Asp + Ser129Glu + Ser158Glu + Thr161Ser +Ala214GlnGly 62Glu + Ser 97Asp + Ser129Asp + Ala199Pro + Leu216Gln +Gly218AspSer129Glu + Asn157Gln + Ser187Asp + Ser190Glu + Gly203Asn +Tyr205AspGly156Asn + Asn162Gln + Ser187Glu + Gly203Glu + Thr215Gln +Thr219GlyAla128Ser + Ser187Glu + Gly203Glu + Tyr205Leu + Thr210Ser +Leu216ThrThr 58Gly + Thr 65Asp + Ser100Glu + Gly105Glu + Ala128Asp +Leu216ThrAsp 59Glu + Thr 65Gly + Gly156Asp + Tyr205Glu + Leu216Cys +Asn217AspThr 58Pro + Asn 96Asp + Ser160Asp + Ser190Asp + Ala202Asn +Thr212AsnSer160Glu + Ser190Asp + Tyr205Asp + Thr207Pro + Thr212Gln +Leu216MetAsn 61Asp + Leu 95Glu + Tyr166Asn + Tyr205Glu + Asn211Glu +Leu216AsnSer 98Glu + Ser158Glu + Thr163Glu + Tyr166Leu + Gly203Glu +Tyr205ThrAsp 59Glu + Ser104Glu + Ser158Asp + Thr163Glu + Phe188His +Thr212ProSer158Asp + Thr163Glu + Ala202Asp + Tyr205Cys + Thr212Asp +Leu216ProTyr166Glu + Ser190Glu + Met198Ser + Gly203Asn + Asn211Asp +Tyr213ProAsn 61Ser + Gly 64Ser + Thr 65Asp + Ser129Glu + Tyr205Glu +Tyr213GlyAla128Pro + Thr132Glu + Gly153Ser + Asn154Asp + Gly165Asp +Gly203GluAsp 59Glu + Thr132Glu + Thr163Pro + Tyr166Ile + Thr210Asp +Leu216AspThr 58Asp + Asn 96Gln + Ser104Asp + Gly126Gln + Gly203Asp +Pro209GluGly 64Asn + Thr 65Glu + Gly126Glu + Tyr166Ala + Phe188Tyr +Thr215GluAsp 59Glu + Asn 96Asp + Gly105Asn + Gly127Asp + Ser158Glu +Thr215SerSer104Asp + Ser131Asp + Gly159Glu + Phe188Asp + Thr210Gly +Thr212GlnVal 94Met + Ser104Asp + Gly126Ser + Ser131Glu + Thr210Glu +Thr215AspThr 58Asp + Ser 97Asp + Gly105Asp + Ser187Glu + Met198Thr +Tyr205ValVal 94Asp + Gly 99Asp + Gly126Asn + Thr163Glu + Gly203Glu +Leu216CysThr 58Glu + Gly 62Glu + Gly105Glu + Gly156Glu + Phe188Thr +Thr212GlyThr 58Gln + Gly159Asp + Ser190Glu + Gly203Asp + Tyr205Ile +Gly218SerThr 58Pro + Ser131Glu + Tyr166Thr + Tyr205Asp + Thr2155er +Gly218AspGly 60Asp + Thr161Asp + Gly165Glu + Pro209Gln + Thr215Gly +Thr219GlyAsp 59Glu + Ser102Asp + Thr161Asp + Asn162Gln + Ser190Glu +Ala202SerLeu 95Cys + Ser100Glu + Ser104Glu + Phe188Cys + Tyr205Met +Leu216AspSer100Asp + Ser104Asp + Gly105Asn + Tyr205Cys + Tyr208Asn +Pro209GluGly 64Ser + Thr 65Gln + Ser100Glu + Ser104Glu + Ser160Asp +Thr207AsnThr 65Gly + Gly 99Asp + Ala128Glu + Ser155Glu + Gly203Gln +Thr215AspGly 99Asp + Ala128Asp + Ser155Asp + Ala202Glu + Thr210Asn +Leu216ValThr 58Asp + Ser 97Asp + Thr132Asp + Thr215Gln + Leu216Asn +Gly218GluThr 58Asp + Ser 97Glu + Gly127Asp + Tyr166Ala + Ala186Ser +Asn217GluThr 65Asp + Thr132Glu + Asn154Asp + Thr212Glu + Leu216Gly +Asn217SerGly 99Gln + Ser131Glu + Thr161Asn + Tyr205Thr + Pro209Asp +Thr215GluTyr103His + Ser131Glu + Gly156Asp + Gly159Glu + Thr212Glu +Leu216MetThr 58Gln + Ser 98Asp + Ser102Glu + Ala202His + Thr212Pro +Thr215AspGly127Glu + Asn162Glu + Phe188Met + Ala202Gly + Gly203Ser +Gly218AsnII洗滌組合物在本發(fā)明的另一實(shí)施方案中，有效量的一種或多種酶變異體包括在用于洗滌各種需要除去蛋白質(zhì)污垢的表面的組合物中。這種洗滌組合物包括用于洗滌硬表面的非限制形式洗滌組合物(例如，液體或顆粒)；用于洗滌織物的非限制形式的洗滌組合物(顆粒、液體或條狀制劑)；餐具洗滌組合物(非限制形式)；非限制形式的口腔用洗滌組合物(例如，潔牙劑、牙膏和漱口劑制劑)；非限制形式的假牙洗滌組合物(例如，液體、片劑)和非限制形式的接觸透鏡洗滌組合物(例如，液體、片劑)。
除了本文上述的卡氏枯草桿菌蛋白酶變異體之外，洗滌組合物還包括與蛋白酶相容的一種或多種洗滌組合物材料。本文使用的術(shù)語“洗滌組合物材料”指的是選擇用于所需的洗滌組合物的具體類型和產(chǎn)物形式(例如，液體、顆粒、條狀、噴霧劑、棒狀、膏狀、凝膠)的任何液體、固體或氣態(tài)材料，該材料還應(yīng)與組合物中使用的卡氏枯草桿菌蛋白酶變異體相容。通過考慮所洗滌的表面材料、在使用過程中用于洗滌條件下所需的組合物的形式(例如通過洗滌劑使用)很容易地具體選擇洗滌組合物材料。本文中使用的術(shù)語“相容”指的是洗滌組合物材料不降低卡氏枯草桿菌蛋白酶變異體的蛋白水解活性至在正常使用場合根據(jù)需要蛋白酶失效的程度。具體的洗滌組合物材料在下文詳細(xì)舉例說明。
本文中使用的“有效量的酶變異體”指的是為獲得在具體洗滌組合物中所需的酶活性而必需的酶變異體的量。這種有效量易于被本領(lǐng)域的技術(shù)人員確定，其基于許多因素，例如所用的具體的酶變異體、洗滌應(yīng)用、洗滌組合物的具體組成，和是否需要液體或無水(例如顆粒、條狀)組合物等等。洗滌組合物優(yōu)選含有約0.0001％-約10％一種或多種本發(fā)明的酶變異體，更優(yōu)選約0.001％-約1％，還優(yōu)選約0.01％-約0.1％。其中使用酶變異體的各種洗滌組合物的若干實(shí)例在下文中進(jìn)一步地討論。本文使用的所有份數(shù)、百分?jǐn)?shù)和比率均為按重量計(jì)，除非另有說明。
本文中使用的“非織物洗滌組合物”包括硬表面洗滌組合物、餐具洗滌組合物、口腔洗滌組合物、假牙洗滌組合物和接觸透鏡洗滌組合物。A.用于硬表面、餐具和織物的洗滌組合物本發(fā)明的酶變異體可用于其中需要高泡和良好不溶底物去除的各種洗滌組合物中。因此，酶變異體可與各種常規(guī)成分一起使用以提供全配制的硬表面洗滌劑。餐具洗滌組合物、織物洗滌組合物等等。這種組合物可以是液體、顆粒、條狀形式等等。這種組合物可配制成含有多達(dá)按表面活性劑重量計(jì)30％-60％的現(xiàn)代“濃縮”洗滌劑。
本發(fā)明的洗滌組合物可選擇性地，并優(yōu)選地含有各種陰離子、非離子、兩性離子等等的表面活性劑。這些表面活性劑通常以組合物的約5％-約35％的含量存在。
用于本發(fā)明的表面活性劑的非限制性實(shí)例包括常規(guī)的C11-C18烷基苯磺酸鹽和伯和無規(guī)烷基硫酸鹽、式為CH3(CH2)x(CHOSO3c-M+)CH3和CH3(CH2)y(CHOSO3-M+)CH2CH3的C10-C18仲(2，3)烷基硫酸鹽，其中x和(y+1)是至少約7，優(yōu)選至少約9的整數(shù)，M是水溶性陽離子，尤其是鈉、C10-C18烷基烷氧基硫酸鹽(尤其是EO1-5乙氧基硫酸鹽)、C10-C18烷基烷氧基羧酸鹽(尤其是EO1-5乙氧基羧酸鹽)、C10-C18烷基聚糖苷和它們相應(yīng)的硫酸化聚糖苷，和C12-C18α-磺化脂肪酸酯、C12-C18烷基和烷基苯酚烷氧基化物(尤其是乙氧基化物和混合的乙氧基化物/丙氧基化物)、C12-C18甜菜堿和磺基甜菜堿(“sultaines”)、C10-C18胺氧化物等等。本發(fā)明優(yōu)選烷基烷氧基硫酸鹽(AES)和烷基烷氧基羧酸鹽(AEC)。(根據(jù)配制者的需要，使用這種表面活性劑與上述的胺氧化物和/或甜菜堿或磺基甜菜堿表面活性劑的混合物也是優(yōu)選的)。其它常規(guī)使用的表面活性劑在標(biāo)準(zhǔn)文獻(xiàn)中列出。尤其有用的表面活性劑包括在1993年3月16日頒布的Connor等人的US5194639中的C10-C18N-甲基葡糖酰胺，列為本文參考文獻(xiàn)。
用于洗滌組合物中的各種各樣的其它成分可包括在本發(fā)明的組合物中，其包括其它活性成分、載體、增溶劑、加工助劑、染料或顏料、用于液體配方的溶劑等等。如果需要附加地增加泡沫，在組合物中，通常以約1％-約10％的含量加入發(fā)泡劑，例如C10-C16烷醇酰胺。C10-C14單乙醇和二乙醇酰胺說明了這種發(fā)泡劑的典型種類。與上述的高泡輔助表面活性劑，例如胺氧化物、甜菜堿和磺基甜菜堿一起使用這類發(fā)泡劑也是有利的。如果需要，溶解的鎂鹽，例如氯化鎂、硫酸鎂等等可以通常約0.1％-約2％的含量加入以提供附加的起泡。
本發(fā)明的液體洗滌組合物可含有作為載體的水和其它溶劑。低分子量的伯或仲醇，例如甲醇、乙醇、丙醇和異丙醇是適宜的。單羥基醇對(duì)于表面活性劑的增溶是優(yōu)選的，但多元醇，例如含有約2-約6個(gè)碳原子和約2-約6個(gè)羥基的多元醇(例如，1，3-丙二醇、乙二醇、甘油和1，2-丙二醇)也可以使用。組合物可含有約5％-約90％，典型地約10％-約50％的這種載體。
本發(fā)明的洗滌組合物將優(yōu)選配制成在使用過程以含水洗滌操作，洗滌水將具有約6.8-約11.0的pH，最終的產(chǎn)物因此通常在該范圍內(nèi)配制?？刂苝H在所需要的范圍內(nèi)的技術(shù)包括使用緩沖劑、堿、酸等等，是本領(lǐng)域技術(shù)人員已知的。
在配制本發(fā)明的硬表面洗滌組合物和織物洗滌組合物時(shí)，配制者可以按重量計(jì)約5％-約50％的含量使用各種助劑。典型的助劑包括1-10微米沸石、多羧酸鹽，例如檸檬酸鹽和氧聯(lián)二琥珀酸鹽、層狀硅酸鹽、磷酸鹽等等。其它常規(guī)的助劑在標(biāo)準(zhǔn)配方中列出。
同樣，配制者可在組合物中，通常以按重量計(jì)約0.001％-1％的含量使用各種其它的酶，例如纖維素酶、脂酶、淀粉酶和蛋白酶。在洗衣洗滌劑領(lǐng)域中已知各種洗滌和織物護(hù)理酶。
各種漂白化合物，例如過碳酸鹽、過硼酸鹽等等可通常以按重量計(jì)約1％-約15％的含量用于該組合物中。如果需要，該組合物還可以含有漂白活性劑，例如四乙?；叶贰⑷甚Ｑ趸交撬猁}等等，它們也是現(xiàn)有技術(shù)中已知的。使用含量通常為按重量計(jì)約1％-約10％。
各種污垢解脫劑，尤其是陰離子低聚酯類型、各種螯合劑，尤其是氨基膦酸鹽和乙二胺二琥珀酸鹽、各種粘土去污劑，尤其是乙氧基化四亞乙五胺、各種分散劑，尤其是聚丙烯酸酯和聚天冬氨酸酯、各種增白劑，尤其是陰離子增白劑、各種泡沫抑制劑，尤其硅烷和仲醇、各種織物柔軟劑，尤其是綠土等等部可以按重量計(jì)約1％-約35％的含量用于組合物。在標(biāo)準(zhǔn)配方和公開的所得中有此常規(guī)物質(zhì)的詳細(xì)描述。
在洗滌組合物中也可使用酶穩(wěn)定劑。酶穩(wěn)定劑包括丙二醇(優(yōu)選約1％-約10％)、甲酸鈉(優(yōu)選約0.1％-約1％)和甲酸鈣(優(yōu)選約0.1％-約1％)。1.硬表面洗滌組合物本文使用的“硬表面洗滌組合物”指的是用于洗滌硬表面，例如地板、墻壁、浴室瓷磚等等的液體和顆粒洗滌組合物。本發(fā)明的硬表面洗滌組合物含有有效量的一種或多種本發(fā)明的酶變異體，按組合物活性酶重量計(jì)優(yōu)選為約0.001％-約10％，更優(yōu)選約0.01％-約5％，還優(yōu)選約0.05％-約1％。除了含有一種或多種酶變異體之外，硬表面洗滌組合物通常含有表面活性劑和水溶性螯合劑。然而，在某些特殊的產(chǎn)物，例如噴霧玻璃清潔劑中，有時(shí)是不使用表面活性劑的，因?yàn)樗鼈儠?huì)產(chǎn)生殘留在玻璃表面的膜狀/條紋殘余物。
當(dāng)存在時(shí)，表面活性劑組分可含有少至組合物的0.1％，但通常組合物將含有約0.25％-約10％，更優(yōu)選約1％-約5％的表面活性劑。
組合物通常含有約0.5％-約50％的洗滌劑助劑，優(yōu)選為約1％-約10％。
pH值優(yōu)選在約8-12的范圍內(nèi)。如果需要進(jìn)行調(diào)節(jié)，可使用常規(guī)的pH調(diào)節(jié)劑，例如氫氧化鈉、碳酸鈉或鹽酸。
在組合物中可包括溶劑。有用的溶劑包括，但不限制于，乙二醇醚，例如二乙二醇單己基醚、二乙二醇單丁基醚、乙二醇單丁基醚、乙二醇單己基醚、丙二醇單丁基醚、二丙二醇單丁基醚，和二醇，例如2，2，4-三甲基-1，3-戊二醇和2-乙基-1，3-己二醇。當(dāng)使用時(shí)，溶劑通常以約0.5％-約15％，優(yōu)選約3％-約11％的含量存在。
此外，當(dāng)組合物以“完全強(qiáng)度”應(yīng)用于表面后表面不進(jìn)行漂洗時(shí)，在本發(fā)明的組合物中可使用高揮發(fā)性溶劑，例如異丙醇或乙醇，以有助于組合物從表面快速蒸發(fā)。當(dāng)使用時(shí)，揮發(fā)性溶劑通常以約2％-約12％的含量存在于組合物中。
本發(fā)明的硬表面洗滌組合物的實(shí)施方案通過如下實(shí)施例說明。
實(shí)施例7-12液體硬表面洗滌組合物實(shí)施 例組分 7 8 9 101112Thr58Asn0.05 0.50 0.02 0.03 0.10 0.03Thr58Gln+Asn61Glu - - - - 0.20 0.02Na2DIDA*EDTA**- - 2.90 2.90 - -檸檬酸鈉- - - - 2.90 2.90C12烷基苯磺酸鈉1.95 - 1.95 - 1.95 -C12烷基硫酸鈉 - 2.20 - 2.20 -2.20C12(乙氧基)***硫酸鈉 - 2.20 - 2.20 -2.20C12二甲基胺氧化物 - 0.50 - 0.50 -0.50枯烯磺酸鈉 1.30 - 1.30 - 1.30 -己基卡必醇***6.30 6.30 6.30 6.30 6.30 6.30水****平衡至100％*N-二乙二醇-N，N-亞氨基二乙酸二鈉**乙二胺二乙酸四鈉***二乙二醇單己基醚****所有配方調(diào)節(jié)到pH7
在實(shí)施例7-10中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體代替Thr58Asn，得到基本上類似的結(jié)果。
在實(shí)施例11-12中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體的任何組合代替Thr58Asn和Thr58Gln+Asn6lGlu，得到基本上類似的結(jié)果。
實(shí)施例13-18用于清潔硬表面和除去家庭霉變的噴霧組合物實(shí)施例組分1314151617 18Val94Ala 0.50 0.05 0.60 0.30 0.20 0.30Leu125Asn+Gly126Pro+Ser129Glu - - - - 0.30 0.10辛基硫酸鈉 2.00 2.00 2.00 2.00 2.00 2.00十二烷基硫酸鈉 4.00 4.00 4.00 4.00 4.00 4.00氫氧化鈉 0.80 0.80 0.80 0.80 0.80 0.80硅酸鹽(鈉) 0.04 0.04 0.04 0.04 0.04 0.04香料 0.35 0.35 0.35 0.35 0.35 0.35水平衡至100％產(chǎn)物pH為約7。
在實(shí)施例13-16中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體代替Val94Ala，得到基本上類似的結(jié)果。
在實(shí)施例17-18中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體的任何組合代替Val94Ala和Leu125Asn+Gly126Pro+Ser129Glu，得到基本上類似的結(jié)果。2.餐具洗滌組合物在本發(fā)明的另一實(shí)施方案中，餐具洗滌組合物含有一種或多種本發(fā)明的酶變異體。本文使用的“餐具洗滌組合物”指的是用于洗滌餐具的組合物的所有形式，其包括，但不限制于，顆粒和液體形式。本發(fā)明的餐具洗滌組合物的實(shí)施方案由如下實(shí)施例說明。
實(shí)施例19-24餐具洗滌組合物實(shí)施例組分 19 2021222324Gly153Asn0.05 0.50 0.02 0.40 0.10 0.03Ala186asp+Phe188Gln - - - -0.40 0.02C12-C14N-甲基葡糖酰胺 0.90 0.90 0.90 0.90 0.90 0.90C12乙氧基(1)硫酸鹽 12.00 12.00 12.00 12.00 12.00 12.002-甲基十一烷酸 4.50 4.50 4.50 4.50 4.50 4.50C12乙氧基(2)羧酸鹽 4.50 4.50 4.50 4.50 4.50 4.50C12醇乙氧基化物(4) 3.00 3.00 3.00 3.00 3.00 3.00C12胺氧化物 3.00 3.00 3.00 3.00 3.00 3.00枯烯磺酸鈉 2.00 2.00 2.00 2.00 2.00 2.00乙醇 4.00 4.00 4.00 4.00 4.00 4.00Mg++(作為氯化鎂)0.20 0.20 0.20 0.20 0.20 0.20Ca++(作為氯化鈣)0.40 0.40 0.40 0.40 0.40 0.40水 平衡至100％產(chǎn)物pH調(diào)節(jié)至7。
在實(shí)施例19-22中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體代替Gly153Asn，得到基本上類似的結(jié)果。
在實(shí)施例23-24中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體的任何組合代替Gly153Asn和Ala186asp+Phe188Gln，得到基本上類似的結(jié)果。3.織物洗滌組合物在本發(fā)明的另一實(shí)施方案中，織物洗滌組合物含有一種或多種本發(fā)明的酶變異體。本文使用的“織物洗滌組合物”指的是用于洗滌織物的組合物的所有形式，其包括，但不限制于，顆粒、液體和條狀形式。優(yōu)選的織物洗滌組合物是液體形式。a.顆?？椢锵礈旖M合物本發(fā)明的顆粒織物洗滌組合物含有有效量的一種或多種本發(fā)明的酶變異體，按組合物活性酶重量計(jì)優(yōu)選為約0.001％-約10％，更優(yōu)選約0.005％-約5％，還優(yōu)選約0.01％-約1％。除了含有一種或多種酶變異體之外，顆?？椢锵礈旖M合物通常含有至少一種表面活性劑、一種或多種助劑和，在某些情況下，漂白劑。
本發(fā)明的顆?？椢锵礈旖M合物的實(shí)施方案由如下實(shí)施例說明。
實(shí)施例25-28顆?？椢锵礈旖M合物實(shí)施例組分 25262728Pro200Asn 0.10 0.20 0.03 0.05Gly203Asp+Tyr205Ala+Tyr208Ser- - 0.02 0.05C13直鏈烷基苯磺酸鹽22.00 22.00 22.00 22.00磷酸鹽(作為三聚磷酸鈉) 23.00 23.00 23.00 23.00碳酸鈉 23.00 23.00 23.00 23.00硅酸鈉 14.00 14.00 14.00 14.00沸石8.20 8.20 8.20 8.20螯合劑(二亞乙三胺五乙酸)0.40 0.40 0.40 0.40硫酸鈉 5.50 5.50 5.50 5.50水平衡至100％在實(shí)施例25-26中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體代替Pro200Asn，得到基本上類似的結(jié)果。
在實(shí)施例27-28中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體的任何組合代替Pro200Asn和Gly203Asp+Tyr205Ala+Tyr208Ser，得到基本上類似的結(jié)果。
實(shí)施例29-32顆?？椢锵礈旖M合物實(shí)施例組分29 30 3132Pro200Asn+Ala202Asn+Vla204 0.10 0.20 0.03 0.05Gly+Asn211Asp+Thr219GlyLeu95Gly+Gly203Pro - -0.02 0.05C12烷基苯磺酸鹽12.00 12.00 12.00 12.00沸石A(1-10微米) 26.00 26.00 26.00 26.002-丁基辛酸 4.00 4.00 4.00 4.00C12-C14仲(2，3)烷基硫酸鈉 5.00 5.00 5.00 5.00檸檬酸鈉 5.00 5.00 5.00 5.00熒光增白劑 0.10 0.10 0.10 0.10硫酸鈉 17.00 17.00 17.00 17.00水和少量組分平衡至100％在實(shí)施例29-30中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體代替Pro200Asn+Ala202Asn+Vla204Gly+Asn211Asp+Thr219Gly，得到基本上類似的結(jié)果。
在實(shí)施例31-32中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體的任何組合代替Pro200Asn+Ala202Asn+Vla204Gly+Asn211Asp+Thr219Gly和Leu95Gly+Gly203Pro，得到基本上類似的結(jié)果。
實(shí)施例33-36顆粒織物洗滌組合物實(shí)施例組分33343536Thr58Gln+Asp59Glu+Leu216Gln 0.10 0.20 0.03 0.05Pro200Asn+Ala202Gln+Tyr208Cys+Thr210 -- 0.02 0.05Pro+Tyr213Asn+Ala214Asn+Thr219SerC13直鏈烷基苯磺酸鹽22.00 22.00 22.00 22.00磷酸鹽(作為三聚磷酸鈉) 23.00 23.00 23.00 23.00碳酸鈉 23.00 23.00 23.00 23.00硅酸鈉 14.00 14.00 14.00 14.00沸石8.20 8.20 8.20 8.20螯合劑(二亞乙三胺五乙酸)0.40 0.40 0.40 0.40硫酸鈉 5.50 5.50 5.50 5.50水 平衡至100％在實(shí)施例33-34中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體代替Thr58Gln+Asp59Glu+Leu216Gln，得到基本上類似的結(jié)果。
在實(shí)施例35-36中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體的任何組合代替Thr58Gln+Asp59Glu+Leu216Gln和Pro200Asn+Ala202Gln+Tyr208Cys+Thr210Pro+Tyr213Asn+Ala214Asn+Thr219Ser，得到基本上類似的結(jié)果。
實(shí)施例37-40顆?？椢锵礈旖M合物實(shí)施例組分37383940Asp59Glu+Gly60Pro+ 0.10 0.20 0.03 0.05Gly62Asn+Gly64GlLeu125Ala - - 0.02 0.05C12烷基苯磺酸鹽 12.00 12.00 12.00 12.00沸石(1-10微米) 26.00 26.00 26.00 26.002-丁基辛酸 4.00 4.00 4.00 4.00C12-C14仲(2，3)烷基硫酸鈉5.00 5.00 5.00 5.00檸檬酸鈉 5.00 5.00 5.00 5.00熒光增白劑 0.10 0.10 0.10 0.10硫酸鈉 17.00 17.00 17.00 17.00水和少量組分 平衡至100％在實(shí)施例37-38中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體代替Asp59Glu+Gly60Pro+Gly62Asn+Gly64Gl，得到基本上類似的結(jié)果。
在實(shí)施例39-40中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體的任何組合代替Asp59Glu+Gly60Pro+Gly62Asn+G1y64Gl和Leu125Ala，得到基本上類似的結(jié)果。
實(shí)施例41-42顆?？椢锵礈旖M合物實(shí)施例組分 4142直鏈烷基苯磺酸鹽 11.4 10.70動(dòng)物脂烷基硫酸鹽 1.80 2.40C14-15烷基硫酸鹽 3.003.10C14-15醇7倍乙氧基化4.004.00動(dòng)物脂醇11倍乙氧基化 1.801.80分散劑 0.070.1硅氧烷液體 0.800.80檸檬酸三鈉 14.00 15.00檸檬酸 3.002.50沸石 32.50 32.10馬來酸丙烯酸共聚物 5.005.00二亞乙三胺五亞甲基膦酸 1.000.20Gly127Glu+Asn162Glu+Phe218Met+ 0.300.30Ala202Gly+Gly203Ser+Gly218Asn脂酶 0.360.40淀粉酶 0.300.30硅酸鈉 2.002.50硫酸鈉 3.505.20聚乙烯吡咯烷酮 0.300.50過硼酸鹽 0.5 1苯酚磺酸鹽 0.1 0.2過氧化酶 0.1 0.1少量組分 至100 至100實(shí)施例43-44顆粒織物洗滌組合物實(shí)施例組分4344直鏈C12烷基苯磺酸鈉6.5 8.0硫酸鈉 15.0 18.0沸石A 26.0 22.0次氮基三乙酸鈉 5.0 5.0聚乙烯吡咯烷酮 0.5 0.7四乙?；叶? 3.0 3.0硼酸 4.0 -過硼酸鹽 0.5 1苯酚磺酸鹽 0.1 0.2Gly218Gln+Thr219Gln0.4 0.4填料(例如，硅酸鹽；碳酸鹽；香料；水) 至100至100實(shí)施例45壓型顆?？椢锵礈旖M合物組分 重量％烷基硫酸鹽 8.0烷基乙氧基硫酸鹽2.0C25和C45醇3和7倍乙氧基化的混合物 6.0多羥基脂肪酸酰胺2.5沸石17.0層狀硅酸鹽/檸檬酸鹽 16.0碳酸鹽 7.0馬來酸丙烯酸共聚物 5.0污垢解脫聚合物 0.4羧甲基纖維素0.4聚(4-乙烯基吡啶)-N-氧化物 0.1乙烯基咪唑和乙烯基吡咯烷酮共聚物0.1PEG2000 0.2Pro209Gln 0.5脂酶0.2纖維素酶0.2四乙?；叶? 6.0過碳酸鹽22.0乙二胺二琥珀酸 0.3泡沫抑制劑 3.54，4’-雙(2-嗎啉基-4-苯胺基-s-三嗪-6-基氨基)芪-2，2’- 0.25二磺酸二鈉4，4’-雙(2-磺基苯乙烯基)聯(lián)苯二鈉 0.05水、香料和少量組分 至100
實(shí)施例46顆粒織物洗滌組合物組分 重量％直鏈烷基苯磺酸鹽 7.6C16-C18烷基硫酸鹽 1.3C14-15醇7倍乙氧基化物4.0椰子烷基二甲基羥基乙基銨氯化物 1.4分散劑 0.07硅氧烷液體 0.8檸檬酸三鈉 5.0沸石4A 15.0馬來酸丙烯酸共聚物 4.0二亞乙三胺五亞甲基膦酸 0.4過硼酸鹽 15.0四乙?；叶? 5.0綠土 10.0聚(環(huán)氧乙烷)(MW300000) 0.3Thr58Pro+Gly64Ser+Thr65Gly 0.4脂酶 0.2淀粉酶 0.3纖維素酶 0.2硅酸鈉 3.0碳酸鈉 10.0羧甲基纖維素 0.2增白劑 0.2水、香料和少量組分 至100
實(shí)施例47顆?？椢锵礈旖M合物組分 重量％直鏈烷基苯磺酸鹽 6.92動(dòng)物脂烷基硫酸鹽 2.05C14-15醇7倍乙氧基化物4.4C12-15烷基乙氧基硫酸鹽-3倍乙氧基化 0.16沸石 20.2檸檬酸鹽 5.5碳酸鹽 15.4硅酸鹽 3.4馬來酸丙烯酸共聚物 4.0羧甲基纖維素酶 0.31污垢解脫聚合物 0.30Val94Gln+Ser100Glu 0.2脂酶 0.36纖維素酶 0.13過硼酸鹽四水合物 11.64過硼酸鹽單水合物 8.7四乙?；叶? 5.0二亞乙三胺五甲基膦酸 0.38硫酸鎂 0.40增白劑 0.19香料、硅氧烷、泡沫抑制劑 0.85少量組分 至100b.液體織物洗滌組合物本發(fā)明的液體織物洗滌組合物含有有效量的一種或多種本發(fā)明的酶變異體，按組合物活性酶重量計(jì)優(yōu)選為約0.005％-約5％，更優(yōu)選為約0.01％-約1％。該液體織物洗滌組合物通常還含有一種陰離子表面活性劑、脂肪酸、水溶性洗滌劑助劑和水。
本發(fā)明的液體織物洗滌組合物的實(shí)施方案由如下實(shí)施例說明。
實(shí)施例48-52液體織物洗滌組合物實(shí)施例組分 48 49 505152Leu125Ala+Gly126Glu 0.05 0.03 0.30 0.03 0.10Gly153Glu+Ile164Ala - - - 0.01 0.20C12-C14烷基硫酸鈉 20.00 20.00 20.00 20.00 20.002-丁基辛酸 5.00 5.00 5.00 5.00 5.00檸檬酸鈉 1.00 1.00 1.00 1.00 1.00C10醇乙氧基化物(3) 13.00 13.00 13.00 13.00 13.00單乙醇胺 2.50 2.50 2.50 2.50 2.50水/丙二醇/乙醇(100∶1∶1)平衡至100％在實(shí)施例48-50中，用表3-36所列的卡氏枯草桿菌蛋白酶變異體代替Leu125Ala+Gly126Glu，得到基本上類似的結(jié)果。
在實(shí)施例51-52中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體的任何組合代替Leu125Ala+Gly126Glu和Gly153Glu+Ile164Ala，得到基本上類似的結(jié)果。
實(shí)施例53-57液體織物洗滌組合物實(shí)施例組分 53 54 55 56 57Phe188Ser0.05 0.03 0.30 0.03 0.10Ala186Gln+Ser187Glu - - - 0.01 0.20+Phe188AspC12-C14烷基硫酸鈉 20.00 20.00 20.00 20.00 20.002-丁基辛酸 5.00 5.00 5.00 5.00 5.00檸檬酸鈉 1.00 1.00 1.00 1.00 1.00C10醇乙氧基化物(3) 13.00 13.00 13.00 13.00 13.00單乙醇胺 2.50 2.50 2.50 2.50 2.50水/丙二醇/乙醇(100∶1∶1) 平衡至100％在實(shí)施例53-55中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體代替Phe188Ser，得到基本上類似的結(jié)果。
在實(shí)施例56-57中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體的任何組合代替Phe188Ser和Ala186Gln+Ser187Glu+Phe188Asp，得到基本上類似的結(jié)果。
實(shí)施例58-59顆粒織物洗滌組合物實(shí)施例組分 5859C12-14烯基琥珀酸 3.0 8.0檸檬酸單水合物 10.0 15.0C12-15烷基硫酸鈉 8.0 8.0C12-15醇2倍乙氧基化硫酸鈉 -3.0C12-15醇7倍乙氧基化物 -8.0C12-15醇5倍乙氧基化物 8.0 -二亞乙三胺五(亞甲基膦酸) 0.2 -油酸 1.8 -乙醇 4.0 4.0丙二醇 2.0 2.0Thr58Ser+Ser100Asp+Ile164Met+Tyr205Ala 0.2 0.2聚乙烯吡咯烷酮 1.0 2.0泡沫抑制劑 0.15 0.15氫氧化鈉 至pH 7.5過硼酸鹽 0.5 1苯酚磺酸鹽 0.1 0.2過氧化酶 0.4 0.1水和少量組分 至10 0份在實(shí)施例58和59中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體代替Thr58Ser+Ser100Asp+Ile164Met+Tyr205Ala，得到基本上類似的結(jié)果。
實(shí)施例60-62液體織物洗滌組合物實(shí)施例組分 60 61 62檸檬酸 7.103.003.00脂肪酸 2.00 - 2.00乙醇1.933.203.20硼酸2.223.503.50單乙醇胺0.711.091.091，2-丙二醇 7.898.008.00枯烯磺酸鈉 1.803.003.00甲酸鈉 0.080.080.08氫氧化鈉6.703.803.80含硅消泡劑 1.161.181.18Asn61Glu0.0145 - -Tyr103Cys+Gly156Glu+Ala202Gln - 0.0145 -+Thr212Gly+Leu216GlnGly105Asn+Gly126Ser+Ser160Glu -- 0.0145+Thr161Asn+Tyr205Val+Gly218Asn脂酶0.200 0.200 0.200纖維素酶- 7.507.50污垢解脫聚合物 0.290.150.15消泡劑 0.060.085 0.085增白劑360.095- -增白劑3 - 0.050.05C12烷基苯磺酸 9.86 - -C12-15烷基多乙氧基化(2.5)硫酸鹽 13.80 18.00 18.00C12葡糖酰胺- 5.005.00C12-13烷基聚乙氧基化物(9) 2.00 2.002.00水、香料和少量組分 平衡至 100％c.條狀織物洗滌組合物本發(fā)明的適合于手洗臟織物的條狀織物洗滌組合物含有有效量的一種或多種本發(fā)明的酶變異體，按組合物活性酶重量計(jì)優(yōu)選為約0.001％-約10％，更優(yōu)選為約0.01％-約1％。
本發(fā)明的條狀織物洗滌組合物的實(shí)施方案由如下實(shí)施例說明。
實(shí)施例63-66條狀織物洗滌組合物實(shí)施例組分 63646566Thr207Glu 0.3 - 0.1 0.02Leu125Ile+Ser131Glu- - 0.4 0.03C12-C16烷基硫酸鈉20.0 20.0 20.0 20.00C12-C14N-甲基葡糖酰胺5.0 5.0 5.0 5.00C11-C13烷基苯磺酸鈉 10.0 10.0 10.0 10.00碳酸鈉 25.0 25.0 25.0 25.00焦磷酸鈉 7.0 7.0 7.0 7.00三聚磷酸鈉 7.0 7.0 7.0 7.00沸石A(0.1-10μ)5.0 5.0 5.0 5.00羧甲基纖維素 0.2 0.2 0.2 0.20聚丙烯酸酯(MW1400) 0.2 0.2 0.2 0.20椰子烷基單乙醇胺 5.0 5.0 5.0 5.00增白劑、香料 0.2 0.2 0.2 0.20硫酸鈣 1.0 1.0 1.0 1.00硫酸鎂 1.0 1.0 1.0 1.00水 4.0 4.0 4.0 4.00填料*平衡至 100％*可以選自常規(guī)材料，例如碳酸鈣、滑石、粘土、硅酸鹽等等。
在實(shí)施例63-64中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體代替Thr207Glu，得到基本上類似的結(jié)果。
在實(shí)施例65-66中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體的任何組合代替Thr207Glu和Leu125Ile+Ser131Glu，得到基本上類似的結(jié)果。
實(shí)施例67-70條狀織物洗滌組合物實(shí)施例組分 67 68 6970Pr0200Ser+Ala214Gly+Asn217Glu 0.3 -0.1 0.02Gly101Asp+Ala128Asp+Ser155Asp+- 0.3 0.4 0.03Ser187Asp+Thr207Gly+Thr212GlnC12-C16烷基硫酸鈉 20.020.0 20.0 20.0C12-C14N-甲基葡糖酰胺 5.0 5.0 5.05.00C11-C13烷基苯磺酸鈉 10.010.0 10.0 10.00碳酸鈉25.025.0 25.0 25.00焦磷酸鈉 7.0 7.0 7.07.00三聚磷酸鈉7.0 7.0 7.07.00沸石A(0.1-10μ) 5.0 5.0 5.05.00羧甲基纖維素 0.2 0.2 0.20.20聚丙烯酸酯(MW1400)0.2 0.2 0.20.20椰子烷基單乙醇胺 5.0 5.0 5.05.00增白劑、香料 0.2 0.2 0.20.20硫酸鈣1.0 1.0 1.01.00硫酸鎂1.0 1.0 1.01.00水4.0 4.0 4.04.00填料*平衡至100％*可以選自常規(guī)材料，例如碳酸鈣、滑石、粘土、硅酸鹽等等。
在實(shí)施例67中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體代替Pro200Ser+Ala214Gly+Asn217Glu，得到基本上類似的結(jié)果。
在實(shí)施例68中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體代替Pro200Ser+Ala214Gly+Asn217Glu和Gly101Asp+Ala128Asp+Ser155Asp+Ser187Asp+Thr207Gly+Thr212Gln，得到基本上類似的結(jié)果。
在實(shí)施例69-70中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體的任何組合代替Va1203Glu和Gly100Glu+Ilel07Ser，得到基本上類似的結(jié)果。B.其它洗滌組合物除了上述硬表面洗滌、餐具洗滌和織物洗滌組合物之外，本發(fā)明的一種或多種酶變異體可加入各種需要水解不溶解底物的其它洗滌組合物中。該附加的洗滌組合物包括，但不限制于，口腔洗滌組合物、假牙洗滌組合物和接觸透鏡洗滌組合物。1.口腔洗滌組合物在本發(fā)明的另一實(shí)施方案中，在組合物中包含一種或多種可藥用量的本發(fā)明的酶變異體以從牙齒或假牙中除去蛋白質(zhì)污垢。本文使用的“口腔洗滌組合物”指的是潔齒粉、牙膏、牙凝膠、牙粉、漱口劑、口腔噴霧劑、口腔凝膠、咀嚼膠、錠劑、香囊、片劑、生物凝膠、預(yù)防膏、假牙處理溶液等等。按組合物的重量計(jì)，洗滌組合物優(yōu)選含有約0.0001-約20％，更優(yōu)選約0.001％-約10％，還優(yōu)選約0.01％-約5％的一種或多種本發(fā)明的酶變異體和可藥用的載體。本文使用的“可藥用的”指的是該術(shù)語描述的藥物、藥劑或惰性成分適合于與人體和低級(jí)動(dòng)物的組織接觸，而沒有不適宜的毒性、不適性、不穩(wěn)定性、刺激性、過敏反應(yīng)等等，與有益效果/風(fēng)險(xiǎn)比率相當(dāng)。
口腔洗滌組合物的可藥用的口腔洗滌載體組分按組合物重量計(jì)通常占約50％-約99.99％，優(yōu)選約65％-約99.99％，更優(yōu)選約65％-約99％。
可包括在本發(fā)明的口腔洗滌組合物中的可藥用的載體組分和選擇性的組分是本領(lǐng)域技術(shù)人員已知的。用于口腔洗滌組合物的各種組合物類型、載體組分和選擇性組分在1992年3月17日頒布的Seibel的US5096700；1991年7月2日頒布的Sampathkumar的US5028414和1991年7月2日頒布的Benedict、Bush和Sunberg的US5028415中，列為本文參考文獻(xiàn)。
本發(fā)明的口腔洗滌組合物的實(shí)施方案在如下實(shí)施例中說明。
實(shí)施例71-74潔牙粉組合物實(shí)施例組分7172 73 74Thy212Gln 2.000 3.500 1.500 2.000山梨糖醇(70％水溶液) 35.000 35.000 35.000 35.000PEG-6*1.000 1.000 1.000 1.000氧化硅牙用磨料**20.000 20.000 20.000 20.000氟化鈉0.243 0.243 0.243 0.243二氧化鈦 0.500 0.500 0.500 0.500糖精鈉0.286 0.286 0.286 0.286烷基硫酸鈉(27.9％水溶液) 4.000 4.000 4.000 4.000香料 1.040 1.040 1.040 1.040羧乙烯基聚合物***0.300 0.300 0.300 0.300Carrageenan****0.800 0.800 0.800 0.800水平衡至 100％*PEG-6＝分子量600的聚乙二醇。**由J.M.Huber提供的沉淀的氧化硅Zeodent119***由B.F.Goodrich Chemical Company提供的Carbopol****由Hercules Chemical Company提供的lota Carrageenan。
在實(shí)施例71-74中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體代替Thy212Gln，得到基本上類似的結(jié)果。
實(shí)施例75-78漱口劑組合物實(shí)施例組分 75767778Val204Asn+Thr215Asn 3.00 7.50 1.00 5.00SDA40醇 8.00 8.00 8.00 8.00香料0.08 0.08 0.08 0.08乳化劑 0.08 0.08 0.08 0.08氟化鈉 0.05 0.05 0.05 0.05甘油 10.0010.0010.0010.00增甜劑 0.02 0.02 0.02 0.02苯甲酸 0.05 0.05 0.05 0.05氫氧化鈉 0.20 0.20 0.20 0.20染料 0.04 0.04 0.04 0.04水 平衡至100％在實(shí)施例75-78中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體代替Val204Asn+Thr215Asn，得到基本上類似的結(jié)果。
實(shí)施例79-82錠劑組合物實(shí)施例組分79808182Thr65Asp+Thr132Glu+Asn154Asp+ 0.01 0.03 0.10 0.02Thr212Glu+Leu216Gly+Asn217Ser山梨糖醇17.50 17.50 17.50 17.50甘露糖醇17.50 17.50 17.50 17.50淀粉13.60 13.60 13.60 13.60增甜劑 1.20 1.20 1.20 1.20香料11.70 11.70 11.70 11.70色素0.10 0.10 0.10 0.10玉米糖漿 平衡至100％在實(shí)施例79-82中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體代替Thr65Asp+Thr132Glu+Asn154Asp+Thr212Glu+Leu216Gly+Asn217Ser，得到基本上類似的結(jié)果。
實(shí)施例83-86口香糖組合物實(shí)施例組分83 848586Tyr103Ala 0.03 0.02 0.10 0.05山梨糖醇結(jié)晶 38.44 38.40 38.40 38.40Palija-T膠基料*20.0020.0020.00 20.00山梨糖醇(70％水溶液) 22.0022.0022.00 22.00甘露糖醇 10.0010.0010.00 10.00甘油 7.56 7.56 7.56 7.56香料 1.00 1.00 1.00 1.00*L A.Dreyfus Company提供。
在實(shí)施例83-86中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體代替Tyr103A1a，得到基本上類似的結(jié)果。2.假牙洗滌組合物在本發(fā)明的另一實(shí)施方案中，用于在口腔外洗滌假牙的假牙洗滌組合物含有一種或多種本發(fā)明的酶變異體。該假牙洗滌組合物含有有效量的一種或多種酶變異體，優(yōu)選按組合物重量計(jì)，約0.0001-約50％，更優(yōu)選約0.001％-約35％，還優(yōu)選約0.01％-約20％的一種或多種本發(fā)明的酶變異體，和假牙洗滌載體。各種假牙洗滌組合物配方，如泡騰片等等是現(xiàn)有技術(shù)中已知的(參見例如Young的US5055305，列為本文參考文獻(xiàn))，通常適合于加入一種或多種酶變異體以便從假牙除去蛋白質(zhì)污垢。
本發(fā)明的假牙洗滌組合物的實(shí)施方案由如下實(shí)施例說明。
實(shí)施例87-90雙層泡騰假牙洗滌片實(shí)施例組分87888990酸性層Leu216Pro 1.0 1.5 0.01 0.05酒石酸 24.0 24.0 24.00 24.00碳酸鈉 4.0 4.0 4.00 4.00氨基磺酸10.0 10.0 10.00 10.00PEG200004.0 4.0 4.00 4.00碳酸氫鈉24.5 24.5 24.50 24.50過硫酸鉀15.0 15.0 15.00 15.00酸性焦磷酸鈉7.0 7.0 7.00 7.00熱解氧化硅 2.0 2.0 2.00 2.00TAED*7.07.07.007.00蓖麻酰基磺基琥珀酸鹽0.50.50.500.50香料1.01.01.001.00堿性層過硼酸鈉單水合物32.0 32.0 32.00 32.00碳酸氫鈉19.0 19.0 19.00 19.00EDTA3.03.03.003.00三聚磷酸鈉 12.0 12.0 12.00 12.00PEG200002.02.02.002.0過硫酸鉀26.0 26.0 26.00 26.00碳酸鈉 2.02.02.002.00熱解氧化硅 2.02.02.002.00染料/香料 2.02.02.002.00*四乙?；叶吩趯?shí)施例87-90中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體代替Leu216Pro，得到基本上類似的結(jié)果。2.接觸透鏡洗滌組合物在本發(fā)明的另一實(shí)施方案中，接觸透鏡洗滌組合物含有一種或多種本發(fā)明的酶變異體。該接觸透鏡洗滌組合物含有有效量的一種或多種酶變異體，優(yōu)選按組合物重量計(jì)，約0.01-約50％，更優(yōu)選約0.01％-約20％，還優(yōu)選約1％-約5％的一種或多種本發(fā)明的酶變異體，和接觸透鏡洗滌載體。各種接觸透鏡洗滌組合物配方，例如片劑、液體等等在現(xiàn)有技術(shù)中是已知的(參見例如1989年9月5日授權(quán)的Davies，Meaken和Rees，US4863627；1988年5月24日再次授權(quán)的Huth，Lam和Kirai，US Re32672；有1986年9月2日授權(quán)的Schafer，US4609493；1987年9月1日授權(quán)的Ogunbiyi和Smith，US4690793；1986年9月30日授權(quán)的Ogunbiyi，Riedhammer和Smiht，US4614549；和1981年8月25日頒布的Ogata，US4285738；均列為本文參考文獻(xiàn))，通常適合于加入本發(fā)明的一種或多種酶變異體以便從接觸透鏡除去蛋白質(zhì)污垢。
本發(fā)明的接觸透鏡洗滌組合物的實(shí)施方案由如下實(shí)施例說明。
實(shí)施例91-94酶解接觸透鏡洗滌溶液實(shí)施例組分 91929394Ala199Thr+Val204Gln 0.01 0.5 0.1 2.0葡萄糖 50.00 50.0 50.0 50.0非離子表面活性劑(聚氧乙烯- 2.00 2.0 2.0 2.0聚氧丙烯共聚物)陰離子表面活性劑(聚氧乙烯- 1.00 1.0 1.0 1.0烷基苯基醚硫酸酯鈉)氯化鈉 1.00 1.0 1.0 1.0硼砂0.30 0.3 0.3 0.3水 平衡至100％在實(shí)施例91-94中，用表3-36中所列的卡氏枯草桿菌蛋白酶變異體代替Ala199Thr+Val204Gln，得到基本上類似的結(jié)果。
盡管描述了本發(fā)明的具體實(shí)施方案，但顯然本領(lǐng)域的技術(shù)人員可作出本發(fā)明的各種改變和改進(jìn)而不會(huì)違背本發(fā)明的精神和范圍。在所附的權(quán)利要求中，本發(fā)明的范圍覆蓋了所有這些改進(jìn)。
序列表(1)一般信息(i)申請(qǐng)人BRODE，PHILIPF。
BARNETT，BOBBYL。
RUBINGH，DONNN。
(ii)發(fā)明項(xiàng)目SUBTILISIN CARLSBERG VARIANTS HANINGDECREASED ASDORPTION AND INCREASEDHYDROLYSIS(iii)序列數(shù)1(iv)通信地址(A)收信人THE PROCTER & GAMBLE COMPANY(B)街11810 East Miami River Road(C)城市Ross(D)州OH(E)國家USA(F)郵區(qū)45061(v)計(jì)算機(jī)可讀形式(A)介質(zhì)類型軟盤(B)計(jì)算機(jī)IBM PC兼容(C)操作系統(tǒng)PC-DOS/MS-DOS(D)軟件Patentln Release#1.0，Version#1.03(vi)當(dāng)前申請(qǐng)資料(A)申請(qǐng)?zhí)?B)申請(qǐng)日(C)分類(viii)代理人/事務(wù)所信息(A)姓名CORSTANJE，BRAHMJ。
(B)登記號(hào)34，804(C)參考/存檔號(hào)5602(ix)電訊信息(A)電話(513)627-2858(B)電傳(513)627-0260(2)SEQ ID NO1的信息(i)序列特征(A)長度274氨基酸(B)類型氨基酸(C)鏈型單(D)拓?fù)鋵W(xué)線性(xi)序列描述SEQID NO1(xi) SEQUENCE DESCRIPTIONSEQ ID NO1Ala Gln Thr Val Pro Tyr Gly Ile Pro Leu Ile Lys Ala Asp Lys Val15 10 15Gln Ala Gln Gly Phe Lys Gly Ala Asn Val Lys Val Ala Val Leu Asp20 25 30Thr Gly Ile Gln Ala Ser His Pro Asp Leu Asn Val Val Gly Gly Ala35 40 45Ser Phe Val Ala Gly Glu Ala Tyr Asn Thr Asp Gly Asn Gly His Gly5055 60Thr His Val Ala Gly Thr Val Ala Ala Leu Asp Asn Thr Thr Gly Val65 70 7580Leu Gly Val Ala Pro Ser Val Ser Leu Tyr Ala Val Lys Val Leu Asn8590 95Ser Ser Gly Ser Gly Ser Tyr Ser Gly Ile Val Ser Gly Ile Glu Trp100 105 110Ala Thr Thr Asn Gly Met Asp Val Ile Asn Met Ser Leu Gly Gly Ala115120 125Ser Gly Ser Thr Ala Met Lys Gln Ala Val Asp Asn Ala Tyr Ala Arg130 135140Gly Val Val Val Val Ala Ala Ala Gly Asn Ser Gly Asn Ser Gly Ser145 150 155 160Thr Asn Thr Ile Gly Tyr Pro Ala Lys Tyr Asp Ser Val Ile Ala Val165 170175Gly Ala Val Asp Ser Asn Ser Asn Arg Ala Ser Phe Ser Ser Val Gly180 185 190Ala Glu Leu Glu Val Met Ala Pro Gly Ala Gly Val Tyr Ser Thr Tyr195200 205Pro Thr Asn Thr Tyr Ala Thr Leu Asn Gly Thr Ser Met Ala Ser Pro210215220His Val Ala Gly Ala Ala Ala Leu Ile Leu Ser Lys His Pro Asn Leu225230 235240Ser Ala Ser Gln Val Arg Asn Arg Leu Ser Ser Thr Ala Thr Tyr Leu245250 255Gly Ser Ser Phe Tyr Tyr Gly Lys Gly Leu Ile Asn Val Glu Ala Ala260 265 270Ala Gln
權(quán)利要求
1.具有卡氏枯草桿菌蛋白酶野生型氨基酸序列的修飾氨基酸序列的卡氏枯草桿菌蛋白酶變異體，野生型氨基酸序列包括第一環(huán)區(qū)域、第二環(huán)區(qū)域、第三環(huán)區(qū)域、第四環(huán)區(qū)域、第五環(huán)區(qū)域和第六環(huán)區(qū)域；其中修飾的氨基酸序列在一個(gè)或多個(gè)環(huán)區(qū)域內(nèi)的一個(gè)或多個(gè)位點(diǎn)上含有取代基；其中A.當(dāng)取代發(fā)生在第一環(huán)區(qū)域中時(shí)，取代發(fā)生在位點(diǎn)58、59、60、61、62、64或65的一個(gè)或多個(gè)上；其中a.當(dāng)取代發(fā)生在位點(diǎn)58時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；b.當(dāng)取代發(fā)生在位點(diǎn)59時(shí)，取代氨基酸是Glu；c.當(dāng)取代發(fā)生在位點(diǎn)60時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；d.當(dāng)取代發(fā)生在位點(diǎn)61時(shí)，取代氨基酸是Asp、Gln、Glu或Ser；e.當(dāng)取代發(fā)生在位點(diǎn)62時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；f.當(dāng)取代發(fā)生在位點(diǎn)64時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；和g.當(dāng)取代發(fā)生在位點(diǎn)65時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；B.當(dāng)取代發(fā)生在第二環(huán)區(qū)域中時(shí)，取代發(fā)生在位點(diǎn)94、95、96、97、98、99、100、101、102、103、104、105或106的一個(gè)或多個(gè)上；其中a.當(dāng)取代發(fā)生在位點(diǎn)94時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Met、Pro、Ser或Thr；b.當(dāng)取代發(fā)生在位點(diǎn)95時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Met、Pro、Ser、Thr或Val；c.當(dāng)取代發(fā)生在位點(diǎn)96時(shí)，取代氨基酸是Asp、Gln、Glu、Ser；d.當(dāng)取代發(fā)生在位點(diǎn)97時(shí)，取代氨基酸是Asp或Glu；e.當(dāng)取代發(fā)生在位點(diǎn)98時(shí)，取代氨基酸是Asp或Glu；f.當(dāng)取代發(fā)生在位點(diǎn)99時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；g.當(dāng)取代發(fā)生在位點(diǎn)100時(shí)，取代氨基酸是Asp或Glu；h.當(dāng)取代發(fā)生在位點(diǎn)101時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；i.當(dāng)取代發(fā)生在位點(diǎn)102時(shí)，取代氨基酸是Asp或Glu；j.當(dāng)取代發(fā)生在位點(diǎn)103時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val；k.當(dāng)取代發(fā)生在位點(diǎn)104時(shí)，取代氨基酸是Asp或Glu；l.當(dāng)取代發(fā)生在位點(diǎn)105時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；和m.當(dāng)取代發(fā)生在位點(diǎn)106時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Leu、Met、Pro、Ser、Thr或Val；C.當(dāng)取代發(fā)生在第三環(huán)區(qū)域中時(shí)，取代發(fā)生位點(diǎn)125、126、127、128、129、130、131或132的一個(gè)或多個(gè)上；其中a.當(dāng)取代發(fā)生在位點(diǎn)125時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Met、Pro、Ser、Thr或Val；b.當(dāng)取代發(fā)生在位點(diǎn)126時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；c.當(dāng)取代發(fā)生在位點(diǎn)127時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；d.當(dāng)取代發(fā)生在位點(diǎn)128時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、His、Pro、Ser或Thr；e.當(dāng)取代發(fā)生在位點(diǎn)129時(shí)，取代氨基酸是Asp或Glu；f.當(dāng)取代發(fā)生在位點(diǎn)130時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；g.當(dāng)取代發(fā)生在位點(diǎn)131時(shí)，取代氨基酸是Asp或Glu；和h.當(dāng)取代發(fā)生在位點(diǎn)132時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；D.當(dāng)取代發(fā)生在第四環(huán)區(qū)域中時(shí)，取代發(fā)生在位點(diǎn)153、154、155、156、157、158、159、160、161、162、163、164、165或166的一個(gè)或多個(gè)上；其中a.當(dāng)取代發(fā)生在位點(diǎn)153時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；b.當(dāng)取代發(fā)生在位點(diǎn)154時(shí)，取代氨基酸是Asp、Gln、Glu或Ser；c.當(dāng)取代發(fā)生在位點(diǎn)155時(shí)，取代氨基酸是Asp或Glu；d.當(dāng)取代發(fā)生在位點(diǎn)156時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；e.當(dāng)取代發(fā)生在位點(diǎn)157時(shí)，取代氨基酸是Asp、Gln、Glu、Ser；f.當(dāng)取代發(fā)生在位點(diǎn)158時(shí)，取代氨基酸是Asp或Glu；g.當(dāng)取代發(fā)生在位點(diǎn)159時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；h.當(dāng)取代發(fā)生在位點(diǎn)160時(shí)，取代氨基酸是Asp或Glu；i.當(dāng)取代發(fā)生在位點(diǎn)161時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；j.當(dāng)取代發(fā)生在位點(diǎn)162時(shí)，取代氨基酸是Asp、Gln、Glu或Ser；k.當(dāng)取代發(fā)生在位點(diǎn)163時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；l.當(dāng)取代發(fā)生在位點(diǎn)164時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Met、Pro、Ser、Thr或Val；m.當(dāng)取代發(fā)生在位點(diǎn)165時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；和n.當(dāng)取代發(fā)生在位點(diǎn)166時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val；和E.當(dāng)取代發(fā)生在第五環(huán)區(qū)域中時(shí)，取代發(fā)生在位點(diǎn)186、187、188、189或190一個(gè)或多個(gè)上；其中a.當(dāng)取代發(fā)生在位點(diǎn)186時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、His、Pro、Ser或Thr；b.當(dāng)取代發(fā)生在位點(diǎn)187時(shí)，取代氨基酸是Asp或Glu；c.當(dāng)取代發(fā)生在位點(diǎn)188時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr、Tyr或Val；d.當(dāng)取代發(fā)生在位點(diǎn)189時(shí)，取代氨基酸是Asp或Glu；e.當(dāng)取代發(fā)生在位點(diǎn)190時(shí)，取代氨基酸是Asn或Glu；和F.當(dāng)取代發(fā)生在第六環(huán)區(qū)域中時(shí)，取代發(fā)生在位點(diǎn)199、200、201、202、203、204、205、206、207、208、209、210、211、212、213、214、215、216、217、218或219的一個(gè)或多個(gè)上；其中a.當(dāng)取代發(fā)生在位點(diǎn)199時(shí)，取代氨基酸是Asn、Asp、gln、Glu、Gly、His、Pro、Ser或Thr；b.當(dāng)取代發(fā)生在位點(diǎn)200時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly或Ser；c.當(dāng)取代發(fā)生在位點(diǎn)201時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；d.當(dāng)取代發(fā)生在位點(diǎn)202時(shí)，取代氨基酸是Asp、Asp、Gln、Glu、Gly、His、Pro、Ser或Thr；e.當(dāng)取代發(fā)生在位點(diǎn)203時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；f.當(dāng)取代發(fā)生在位點(diǎn)204時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Met、Pro、Ser或Thr；g.當(dāng)取代發(fā)生在位點(diǎn)205時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val；h.當(dāng)取代發(fā)生在位點(diǎn)206時(shí)，取代氨基酸是Asp或Glu；i.當(dāng)取代發(fā)生在位點(diǎn)207時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；j.當(dāng)取代發(fā)生在位點(diǎn)208時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val；k.當(dāng)取代發(fā)生在位點(diǎn)209時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly或Ser；l.當(dāng)取代發(fā)生在位點(diǎn)210時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；m.當(dāng)取代發(fā)生在位點(diǎn)211時(shí)，取代氨基酸是Asp、Gln、Glu或Ser；n.當(dāng)取代發(fā)生在位點(diǎn)212時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；o.當(dāng)取代發(fā)生在位點(diǎn)213時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val；p.當(dāng)取代發(fā)生在位點(diǎn)214時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、His、Pro、Ser或Thr；q.當(dāng)取代發(fā)生在位點(diǎn)215時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；r.當(dāng)取代發(fā)生在位點(diǎn)216時(shí)，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Met、Pro、Ser、Thr或Val；s.當(dāng)取代發(fā)生在位點(diǎn)217時(shí)，取代氨基酸是Asp、Gln、Glu或Ser；t.當(dāng)取代發(fā)生在位點(diǎn)218時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；和u.當(dāng)取代發(fā)生在位點(diǎn)219時(shí)，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；從而，與野生型卡氏枯草桿菌蛋白酶相比較，卡氏枯草桿菌蛋白酶變異體對(duì)不溶解底物具有降低的吸附作用和增加的水解作用。
2.權(quán)利要求1的卡氏枯草桿菌蛋白酶變異體，其中一個(gè)或多個(gè)取代發(fā)生在第一環(huán)區(qū)域。
3.權(quán)利要求1的卡氏枯草桿菌蛋白酶變異體，其中一個(gè)或多個(gè)取代發(fā)生在第二環(huán)區(qū)域。
4.權(quán)利要求1的卡氏枯草桿菌蛋白酶變異體，其中一個(gè)或多個(gè)取代發(fā)生在第三環(huán)區(qū)域。
5.權(quán)利要求1的卡氏枯草桿菌蛋白酶變異體，其中一個(gè)或多個(gè)取代發(fā)生在第四環(huán)區(qū)域。
6.權(quán)利要求1的卡氏枯草桿菌蛋白酶變異體，其中一個(gè)或多個(gè)取代發(fā)生在第五環(huán)區(qū)域。
7.權(quán)利要求1的卡氏枯草桿菌蛋白酶變異體，其中一個(gè)或多個(gè)取代發(fā)生在第六環(huán)區(qū)域。
8.一種選自硬表面洗滌組合物、餐具洗滌組合物、口腔洗滌組合物、假牙洗滌組合物、接觸透鏡洗滌組合物和織物洗滌組合物的洗滌組合物，其特征在于其中洗滌組合物含有權(quán)利要求1-7的任何一項(xiàng)的卡氏枯草桿菌蛋白酶變異體和洗滌組合物載體；洗滌組合物優(yōu)選是硬表面洗滌組合物或洗滌組合物是織物洗滌組合物；組合物優(yōu)選含有按組合物重量計(jì)至少約5％的表面活性劑和至少約5％助劑；組合物還優(yōu)選含有選自溶劑、緩沖劑、酶、污垢釋放劑、粘土去污劑、分散劑、增白劑、泡沫抑制劑、織物柔軟劑、增泡劑、酶穩(wěn)定劑、漂白劑、染料、香料和它們的混合物的洗滌組合物材料。
9.編碼權(quán)利要求1-7的任何一項(xiàng)的卡氏枯草桿菌蛋白酶變異體的突變體卡氏枯草桿菌蛋白酶基因。
全文摘要
本發(fā)明涉及具有野生型卡氏枯草桿菌蛋白酶氨基酸序列的修飾氨基酸序列的卡氏枯草桿菌蛋白酶變異體,野生型氨基酸序列包括第一環(huán)區(qū)域、第二環(huán)區(qū)域、第三環(huán)區(qū)域、第四環(huán)區(qū)域、第五環(huán)區(qū)域和第六環(huán)區(qū)域;其中,修飾的氨基酸序列在一個(gè)或多個(gè)環(huán)區(qū)域中的特定標(biāo)記的位點(diǎn)上含有與野生型卡氏枯草桿菌蛋白酶不同的氨基酸(即取代),從而與野生型卡氏枯草桿菌蛋白酶相比,卡氏枯草桿菌蛋白酶變異體具有對(duì)不溶解底物降低的吸附作用和增加的水解作用。本發(fā)明還涉及編碼這種卡氏枯草桿菌蛋白酶變異體的基因。本發(fā)明還涉及含有這種卡氏枯草桿菌蛋白酶變異體的用于洗滌各種表面的組合物。
文檔編號(hào)C11D3/00GK1183119SQ96193612
公開日1998年5月27日 申請(qǐng)日期1996年2月14日 優(yōu)先權(quán)日1995年3月8日
發(fā)明者P·F·布羅德三世, B·L·巴爾內(nèi)特, D·N·魯賓 申請(qǐng)人:普羅格特-甘布爾公司