CN1183119A - 具有降低的吸附作用和增加的水解作用的卡氏枯草杆菌蛋白酶变异体 - Google Patents
具有降低的吸附作用和增加的水解作用的卡氏枯草杆菌蛋白酶变异体 Download PDFInfo
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Abstract
本发明涉及具有野生型卡氏枯草杆菌蛋白酶氨基酸序列的修饰氨基酸序列的卡氏枯草杆菌蛋白酶变异体,野生型氨基酸序列包括第一环区域、第二环区域、第三环区域、第四环区域、第五环区域和第六环区域;其中,修饰的氨基酸序列在一个或多个环区域中的特定标记的位点上含有与野生型卡氏枯草杆菌蛋白酶不同的氨基酸(即取代),从而与野生型卡氏枯草杆菌蛋白酶相比,卡氏枯草杆菌蛋白酶变异体具有对不溶解底物降低的吸附作用和增加的水解作用。本发明还涉及编码这种卡氏枯草杆菌蛋白酶变异体的基因。本发明还涉及含有这种卡氏枯草杆菌蛋白酶变异体的用于洗涤各种表面的组合物。
Description
技术领域
本发明涉及新的用于各种洗涤组合物的酶变异体和编码这种酶变异体的DNA序列。
发明背景
酶构成了最大种类的天然产生的蛋白质。每一类酶通常催化(加速反应而不被消耗)各种化学反应。称为蛋白酶的一种酶已知能够水解(分解化合物成两个或更多个更简单的化合物，在断裂的化学键的两侧接受水分子的H和OH部分)其它蛋白质。将天然产生的和蛋白质设计的蛋白酶作为添加剂加入洗衣洗涤剂制剂中即利用了水解蛋白质的这种能力。在衣服上的许多污垢是蛋白质的，广泛特效的蛋白酶可显著改善这种污垢的除去。
不幸的是，这些蛋白质在其天然的，细菌的环境中的功效水平通常不能转移至相对非天然的洗涤环境。尤其是蛋白酶特性，例如热稳定性、pH稳定性、氧化稳定性和底物特性对于酶的天然环境之外的应用未必达到最佳。
蛋白酶的氨基酸序列决定了蛋白酶的特征。根据氨基酸序列中改变的位点、性质和/或幅度，改变蛋白酶的氨基酸序列可改变酶的特性至各不相同的程度，甚至可失活酶。人们已做了许多尝试以改变蛋白酶的野生型氨基酸序列，以改善它们的性质，从而增加蛋白酶在洗涤环境中的效率。这些尝试包括改变氨基酸序列以便在相当不同的条件下提高热稳定性和改善氧化稳定性。
尽管在现有技术中描述了各种各样的尝试，但人们仍不断需要用于洗涤各种表面的蛋白酶的新有效变异体。发明目的
本发明的目的在于提供与野生型酶相比较具有改善的水解作用的卡氏(Carlsberg)枯草杆菌蛋白酶酶变异体。
本发明的另一个目的在于提供含有这些枯草杆菌蛋白酶变异体的洗涤组合物。
发明概述
本发明涉及具有野生型卡氏枯草杆菌蛋白酶氨基酸序列的修饰氨基酸序列的卡氏枯草杆菌蛋白酶变异体，野生型氨基酸序列包括第一环区域、第二环区域、第三环区域、第四环区域和第五环区域；其中，修饰的氨基酸序列在一个或多个环区域中的特定标记的位点上含有与野生型卡氏枯草杆菌蛋白酶不同的氨基酸(即取代)，与野生型卡氏枯草杆菌蛋白酶相比，卡氏枯草杆菌蛋白酶变异体具有对不溶解底物降低的吸附作用和增加的水解作用。本发明还涉及编码这种卡氏枯草杆菌蛋白酶变异体的基因。本发明还涉及含有这种卡氏枯草杆菌蛋白酶变异体的用于洗涤各种表面的组合物。
发明详述I.枯草杆菌蛋白酶变异体
本发明涉及枯草杆菌蛋白酶，尤其是卡氏枯草杆菌蛋白酶，它们通过突变编码酶的各种核苷酸顺序，从而改变酶的氨基酸序列而被修饰。本发明的修饰枯草杆菌蛋白酶(下文称为“卡氏枯草杆菌蛋白酶变异体”)与野生型卡氏枯草杆菌蛋白酶相比，具有对不溶解底物降低的吸附作用和增加的水解作用。本发明还涉及编码这种卡氏枯草杆菌蛋白酶变异体的突变基因。
本发明的枯草杆菌蛋白酶属于称为蛋白酶的酶的种类。蛋白酶是断裂肽键的催化剂。一种类型的蛋白酶是丝氨酸蛋白酶。丝氨酸的特征在于在活性位点上存在必需的丝氨酸残基。
随着酶浓度的增加溶解底物的酶水解速率增加的现象已经作了充分报导。因而，对于表面结合的底物，例如在许多洗涤应用中遇到的情况，水解速率随着表面浓度的增加而增加似乎是可能的，这已由事实说明。(Brode，P.F.III和D.S.Rauch，LANGMUIR，“枯草芽孢杆菌BPN’。固定化底物上的活性”，卷8，1325-1329页(1992))。事实上，当酶的表面浓度变化时，对于不溶解底物已经发现了表面浓度与速率的直线关系。(Rubingh，D.N.和M.D.Bauer.“在蛋白质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编，第464页(1992))。出乎意料的是，当试图将这种原理用于研究得到较好洗涤性能的变并体蛋白酶时，我们不能发现吸附更多的酶得到较好的性能。事实上，我们惊奇地发现与之相反的事实：酶与底物的下降的吸附导致增加的底物水解(即，较好的洗涤性能)。
尽管不希望限制于任何理论，但我们相信，当将变异体相互比较时，改善的性能是如下事实的结果，即吸附较少的酶也不太紧密地结合，因而在欲除去不溶解蛋白质底物的表面更易移动。在可比的酶溶液浓度下，增加的移动能力足以胜过通过向表面提供较高浓度酶而给予的任何优点。
本发明描述的突变用于改变(即降低)酶与表面结合的污垢的吸附。在卡氏枯草杆菌蛋白酶中，某些氨基酸在酶分子上形成外环。为了讨论，这些环可称为第一、第二、第三、第四和第五环区域。具体地说，位点58-65形成第一环区域；位点94-106形成第二环区域；位点125-132形成第三环区域；位点153-166形成第四环区域；位点186-190形成第五环区域；和位点199-219形成第六环区域(位点编号类似于野生型枯草杆菌蛋白酶即卡氏枯草杆菌蛋白酶(SEQ ID NO：1)的氨基酸序列)。
我们相信，这些环区域在酶分子与表面结合的肽的吸附中起着重要的作用，在一个或多个环区域中具体的突变对吸附具有明显的效果。尽管不希望限制于任何理论，但我们相信出于至少两个原因，这些环区域对卡氏枯草杆菌蛋白酶分子的吸附是重要的。首先，含有环区域的氨基酸能够密切地与分子暴露的任何表面接触。其次，卡氏枯草杆菌蛋白酶分子的环区域与活性位点的接近和结合能使得它们在酶与表面结合的底物(肽/蛋白质污垢)的催化产生的吸附中起作用。
本文使用的术语“变异体”是指含有不同于野生型氨基酸序列的酶。
本文使用的术语“突变卡氏枯草杆菌蛋白酶基因”指的是编码卡氏枯草杆菌蛋白酶变异体的基团。
本文使用的术语“野生型卡氏枯草杆菌蛋白酶”指的是由SEQ IDNO：1表示的酶。该卡氏枯草杆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.S.，生物化学杂志.，卷243，第2184-2191页(1968)(列为参考文献)中进一步描述。
本文使用的术语“卡氏枯草杆菌蛋白酶野生型氨基酸序列”包括SEQID NO：1以及除位点58-65、94-106、125-132、153-166、186-190和199-219之外具有修饰氨基酸序列的SEQ ID NO：1。
本文使用的术语“更亲水的氨基酸”指的是参考下表中的亲水性值比目标氨基酸具有更大亲水性值的任何其它氨基酸。如下亲水性值表(表1)以增加的亲水性值的下降顺序列出氨基酸(参见Hopp，T.P.，和Woods，K.R.，“由氨基酸序列判断蛋白质抗原决定簇”，美国国家科学院院刊，卷78，3824-3828页，1981，列为参考文献)。
表1氨基酸 亲水性值
Trp -3.4
Phe -2.5
Tyr -2.3
Leu，Ile -1.8
Val -1.5
Met -1.3
Cys -1.0
Ala，His -0.5
Thr -0.4
Pro，Gly -0.0
Gln，Asn 0.2
Ser 0.3Arg+，Lys+，Glu-，Asp- 3.0
表1还说明了带有电荷(该特征基于约8-9的pH)。正电荷的氨基酸是Arg和Lys，负电荷氨基酸是Glu和Asp，其余的氨基酸是中性的。在本发明的优选实施方案中，取代氨基酸是中性的或负电荷的，更优选负电荷(即Glu或Asp)。
因而，例如说明“用相等或更亲水的中性或带有负电荷的氨基酸取代的Gln”指的是用Asn(它与Gln的亲水性相等)或Ser、Glu或Asp(它们比Gln更亲水)取代的Gln。它们每个均是中性或具有负电荷，与Gln相比较，它们具有较大的亲水性值。同样，说明“用更亲水的中性或带有负电荷的氨基酸取代的Pro”指的是用Gln、Asn、Ser、Glu或Asp取代的Pro。
在本发明的一个实施方案中，卡氏枯草杆菌蛋白酶变异体具有卡氏枯草杆菌蛋白酶野生型氨基酸序列的修饰氨基酸序列，其中野生型氨基酸序列含有在第一环区域、第二环区域、第三环区域、第四环区域、第五环区域或第六环区域的一个或多个中的一个或多个位点上的取代基；从而卡氏枯草杆菌蛋白酶变异体与野生型卡氏枯草杆菌蛋白酶相比较，对不溶解底物具有降低的吸附作用和增加的水解作用。
在本发明的优选实施方案中，在一个或多个环区域中一个或多个位点取代氨基酸，参考表1，是中性的或负电荷的，与在野生型氨基酸序列中目标位点的氨基酸相比较，是相等或更亲水性的，优选是更亲水性的。A.第一环区域中的取代
当取代发生在第一环区域中时，取代发生在位点58、59、60、61、62、64或65的一个或多个上。
当取代发生在位点58时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。
当取代发生在位点59时，取代氨基酸是Glu。
当取代发生在位点60时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
当取代发生在位点61时，取代氨基酸是Asp、Gln、Glu或Ser。
当取代发生在位点62时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
当取代发生在位点64时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
当取代发生在位点65时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。B.第二环区域的取代
当取代发生在第二环区域中时，取代发生在位点94、95、96、97、98、99、100、101、102、103、104、105或106的一个或多个上。
当取代发生在位点94时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Met、Pro、Ser或Thr。
当取代发生在位点95时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Met、Pro、Ser、Thr或Val。
当取代发生在位点96时，取代氨基酸是Asp、Gln、Glu、Ser。
当取代发生在位点97时，取代氨基酸是Asp或Glu。
当取代发生在位点98时，取代氨基酸是Asp或Glu。
当取代发生在位点99时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
当取代发生在位点100时，取代氨基酸是Asp或Glu。
当取代发生在位点101时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
当取代发生在位点102时，取代氨基酸是Asp或Glu。
当取代发生在位点103时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val。
当取代发生在位点104时，取代氨基酸是Asp或Glu。
当取代发生在位点105时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
当取代发生在位点106时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Leu、Met、Pro、Ser、Thr或Val。C.第三环区域中的取代
当取代发生在第三环区域中时，取代发生位点125、126、127、128、129、130、131或132的一个或多个上。
当取代发生在位点125时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Met、Pro、Ser、Thr或Val。
当取代发生在位点126时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
当取代发生在位点127时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
当取代发生在位点128时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、His、Pro、Ser或Thr。
当取代发生在位点129时，取代氨基酸是Asp或Glu。
当取代发生在位点130时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
当取代发生在位点131时，取代氨基酸是Asp或Glu。
当取代发生在位点132时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。D.第四环区域中的取代
当取代发生在第四环区域中时，取代发生在位点153、154、155、156、157、158、159、160、161、162、163、164、165或166的一个或多个上。
当取代发生在位点153时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
当取代发生在位点154时，取代氨基酸是Asp、Gln、Glu或Ser。
当取代发生在位点155时，取代氨基酸是Asp或Glu。
当取代发生在位点156时，取代氨基酸是Ash、Asp、Gln、Glu、Pro或Ser。
当取代发生在位点157时，取代氨基酸是Asp、Gln、Glu、Ser。
当取代发生在位点158时，取代氨基酸是Asp或Glu。
当取代发生在位点159时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
当取代发生在位点160时，取代氨基酸是Asp或Glu。
当取代发生在位点161时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。
当取代发生在位点162时，取代氨基酸是Asp、Gln、Glu或Ser。
当取代发生在位点163时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。
当取代发生在位点164时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Met、Pro、Ser、Thr或Val。
当取代发生在位点165时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。和
当取代发生在位点166时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val。E.第五环区域中的取代
当取代发生在第五环区域中时，取代发生在位点186、187、188、189或190一个或多个上。
当取代发生在位点186时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、His、Pro、Ser或Thr。
当取代发生在位点187时，取代氨基酸是Asp或Glu。
当取代发生在位点188时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr、Tyr或Val。
当取代发生在位点189时，取代氨基酸是Asp或Glu。
当取代发生在位点190时，取代氨基酸是Asn或Glu。F.第六环区域中的取代
当取代发生在第六环区域中时，取代发生在位点199、200、201、202、203、204、205、206、207、208、209、210、211、212、213、214、215、216、217、218或219的一个或多个上。
当取代发生在位点199时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、His、Pro、Ser或Thr。
当取代发生在位点200时，取代氨基酸是Asn、Asp、Gln、Glu、Gly或Ser。
当取代发生在位点201时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
当取代发生在位点202时，取代氨基酸是Asp、Asp、Gln、Glu、Gly、His、Pro、Ser或Thr。
当取代发生在位点203时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
当取代发生在位点204时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Met、Pro、Ser或Thr。
当取代发生在位点205时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val。
当取代发生在位点206时，取代氨基酸是Asp或Glu。
当取代发生在位点207时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。
当取代发生在位点208时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val。
当取代发生在位点209时，取代氨基酸是Asn、Asp、Gln、Glu、Gly或Ser。
当取代发生在位点210时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。
当取代发生在位点211时，取代氨基酸是Asp、Gln、Glu或Ser。
当取代发生在位点212时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。
当取代发生在位点213时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val。
当取代发生在位点214时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、His、Pro、Ser或Thr。
当取代发生在位点215时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。
当取代发生在位点216时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Met、Pro、Ser、Thr或Val。
当取代发生在位点217时，取代氨基酸是Asp、Gln、Glu或Ser。
当取代发生在位点218时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser。
当取代发生在位点219时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser。
G.酶变异体的制备
实施例1
突变体卡氏枯草杆菌蛋白酶基因
构建含有野生型卡氏枯草杆菌蛋白酶基因的噬菌粒(“TP”)。将质粒pUC119的2.8Kbp PvuII限制酶片段(Vieira，J.和Messing，J.，“单链的质粒DNA的制备”，153酶学方法3-11(1989))克隆至质粒pUB110(Bacillus Genetic Stock Center，Columbus，OH 1E9)的PvuII位点。pUC119-pUB110杂交质粒被称为pJMA601。在pJMA601的BamH1限制位克隆入具有TP的聚合酶链反应扩增的产CP的卡氏枯草杆菌蛋白酶基因。噬菌粒TP被转化入大肠杆菌Ung-菌株CJ236，用VCSM13辅助噬菌体(Kunkel，T.A.，J.D.Roberts和R.A.Zakour，“无需表型选择的快速和有效的位点专一诱变”，酶学方法，卷154，367-382页(1987)；由Yuckenberg，P.D.，F.Witney，J.Geisselsoder和J.McClary改进，“使用含尿嘧啶的DNA和噬菌粒载体的体外定点诱变”，定向突变，操作指南，M.J.McPherson编，第27-48页(1991)；均列为本文参考文献)制备单链的含尿嘧啶DNA模板。使用Zoller和Smith方法的单引物定点诱变修饰方法(Zoller，M.J.和M.Smith，“使用M13衍生的载体的寡核苷酸定向诱变：一种在DNA任何片段中制备点突变的有效和常用方法”，核酸研究，卷10，6487-6500页(1982)，列为本文参考文献)制备所有突变体(基本上如上述Yuckenberg等人，1991中说明)。寡核苷酸使用Applied BiosystemInc.380B DNA合成仪制备。诱变反应产物转化入大肠杆菌菌株MM 294(American Type Culture Collection E.Coli.33625)中。所有突变体由DNA测序证实，分离的DNA转化入枯草芽孢杆菌表达菌株BG2036(Yang，M.Y.，E.Ferrari和D.J.Henner，(1984)，“枯草芽孢杆菌的中性蛋白酶基因的克隆和克隆的基因用于产生体外得到的缺失突变”，细胞学杂志，卷160，15-21页)。对于某些环突变体，用在相应环中移码终止的密码子突变的修饰TP制备尿嘧啶模板。寡核苷酸用来恢复合适的阅读框和编码在位点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的随机取代(在这些密码子中对于所有三种碱基所有四种寡核苷酸的等摩尔和/或可变混合物)。校正移码终止和制备功能酶的突变由它们消化酪蛋白的能力说明。随机取代用DNA测序确定。
实施例2
发酵
将含有感兴趣的枯草芽孢杆菌突变体的枯草芽孢杆菌细胞(BG2036)在1升LB-葡萄糖肉汤培养基中的mid-log相中生长，在总体积10升的Biostat ED发酵器(B.Braun Biotech，Inc.，Allentown，Pennsylvania)中培养。发酵介质含有酵母提取物、淀粉、消泡剂、缓冲剂和痕量矿物质(参见发酵：操作指南B.McNeil和L.M.Harvey编，1990)。在发酵试验过程中，肉汤保持恒定pH7.0。加入氯霉素以进行诱变质粒的抗菌选择。细胞在37℃生长过夜至A600约60，并收获。
实施例3
提纯
将发酵肉汤进行如下步骤以得到纯的酶。通过离心除去肉汤中的枯草芽孢杆菌细胞，通过用100K截断膜除去细的粒子而澄清。然后用10K截断膜上浓缩，进行流体透析以降低离子强度，用0.025M MES缓冲剂(2-(N-吗啉代)乙磺酸)调节pH至5.5。通过将酶加入阳离子交换层析柱或亲合吸附层析柱，由柱中用氯化钠或丙二醇梯度洗脱而进一步纯化(参见，Scopes，R.K.，蛋白质纯化理论和操作，Springer-Verlag，NewYork(1984)，列为本文参考文献)。
使用pNA试验(DelMar，E.G.，C.Largman，J.W.Brodrick和M.C.Geokas，生物化学年鉴，卷99，316-320页(1979)，列为本文参考文献)测定在梯度洗脱过程中收集的级分的酶浓度。该试验测量当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-对-硝基苯胺(sAAPF-pNA)时释放对-硝基苯胺的速率。用分光光度计在410nm测量水解反应产生黄色的速率，与活性酶浓度成比例。此外，在280nm的吸附测量用来测定总蛋白质浓度。活性酶/总蛋白质比率得到酶的纯度，并用于鉴定用作存贮溶液的收集的级分。
为避免在贮存过程中酶的自解，在由层析柱得到的收集级分中加入相等重量的丙二醇。在提纯过程完成后，存贮酶溶液用SDS-PAGE(十二烷基磺酸钠聚丙烯酰胺凝胶电泳)校验，用胰蛋白酶抑制剂型II-T：由Sigma Chemical Company(St.Louis，Missouri)购买的火鸡卵白经活性位滴定法测定绝对酶浓度。测量的转化因子将说明在酶分子中不同位点发生的改变使得酶变异体与野生型相比较对溶解底物pNA具有增加的活性。
在使用的制剂中，酶存贮溶液通过Sephadex-G25(Pharmacia，Piscataway，New Jersey)大小排除柱以除去丙二醇和交换缓冲剂。用含有0.01M氯化钙的0.1M Tris缓冲剂(三(羟基甲基-氨基)甲烷)代替在酶存贮溶液中的MES缓冲剂，用盐酸调节pH至8.6。在恒温25℃下在Tris缓冲剂中在pH8.6下进行所有试验。H酶变异体的特性
实施例4
模型表面制备
由CPG Inc.(Fairfield，New Jersey)购买的氨基丙基控制孔玻璃(CPG)用作共价连接由Bachem，Inc.(Torence，California)购买的sAAPD-pNA底物的载体。反应在二甲基亚砜中进行，用(1-乙基-3-[3-(二甲基氨基)丙基]碳化二亚胺盐酸盐)(EDC)作偶合试验。在反应完成后(用pNA试验监测)，除去过量的溶剂，用二甲基亚砜(DMSO)和双蒸馏水漂洗CPG：sAAPF-pNA。然后在烘箱中氮气吹扫在约70℃下干燥。反应方案和固定的底物的制备方法如Brode，P.F.III，和D.S.Rauch，“枯草杆菌蛋白酶BPN’：在固定底物上的活性”，LANGMUIR，卷8，1325-1329页(1992)(列为本文参考文献)中描述进行。
CPG表面含有62000±7000pNA分子/μm2。从CPG Inc.报导的CPG的50.0m2/g的值来看，表面积保持未变。这说明用于在CPG中加入sAAPf-pNA的方法未破坏孔结构(平均直径486_)。
实施例5
表面水解试验
使用CPG：sAAPf-pNA，酶变异体的吸附和CPG-结合的肽的水解可在同一试验中测量。在含有Tris缓冲剂和已脱气的CPG：sAAPF-pNA的烧瓶中加入少量的酶变异体存贮溶液。烧瓶在活塞作用的振荡器中振荡90分钟，在此期间，以不同的时间间隔停止振荡器(例如，在吸附水解作用的最初阶段，例如最初20分钟为每2分钟-在实验结束时为每10分钟)。使CPG：sAAPF-pNA静置，取样溶液。如上述Brode等人，1991中所述进行吸附和水解的实验过程和计算。
测定所有的酶的自溶稳定性，在实验的时间内，应未显示明显的自溶损失。因此，酶吸附可通过测量溶液消耗测定。最初酶变异体浓度与在每个间隔的时间点测量的浓度之间的差值得到吸附的酶变异体的量。由表面水解的pNA的量由在410nm时试样的吸附读数测定。水解的pNA的总量由取样的量加上残余在烧瓶中的量计算。该值通过减去当不存在酶时在pH8.6下由Tris缓冲剂水解的pNA的量而校准。根据酶的效率，基准水解范围为总水解的7-29％。
实施例6
溶解底物的动力学分析
溶解底物sAAPF-pNA的水解速率通过用DU-70分光光度计在410nm下测量作为时间函数的吸附增量而监测。酶浓度保持恒定，并制备成6-10纳摩尔的范围，而对于每次动力学测定，底物浓度在90-700μM sAAPF-pNA之间变化。在900秒的时间内，每秒收集吸附数据点，数据传递至LOTUSTM记录纸(Lotus Development Corporation，Cambridge，Massachusetts)。用标准Linewweaver Burk分析法进行动力学参数的分析，其中在试验最初(通常为最初几分钟)的数据符合于线性回归曲线以得到v0。以标准倒数方式标绘的v0和s0数据得到KM和Kcat。I.卡氏枯草杆菌蛋白酶变异体的实例
降低表面结合的底物的吸附作用和增加的水解作用的本发明卡氏枯草杆菌蛋白酶变异体在如下表3-36中说明。在描述的具体突变中，首先给出野生型中出现的原始氨基酸，其次是位点编号，第三是取代氨基酸。
表2
环1-单突变变异体
Thr58Asn
Thr58Asp
Thr58Gln
Thr58Glu
Thr58Gly
Thr58Pro
Thr58Ser
Asp59Glu
Gly60Asn
Gly60Asp
Gly60Gln
Gly60Glu
Gly60Pro
Gly60Ser
Asn61Asp
Asn61Gln
Asn61Glu
Asn61Ser
Gly62Asn
Gly62Asp
Gly62Gln
Gly62Glu
Gly62Pro
Gly62Ser
Gly64Asn
Gly64Asp
Gly64Gln
Gly64Glu
Gly64Pro
Gly64Ser
Thr65Asn
Thr65Asp
Thr65Gln
Thr65Glu
Thr65Gly
Thr65Pro
Thr65Ser
表3
环1-双突变变异体
Thr58Gln + Asn61Glu
Asn61Asp + Thr65Pro
Thr58Gln + Asp59Glu
Thr58Gln + Gly62Ser
Gly62Asn + Thr65Pro
Thr58Gln + Gly60Asp
Thr58Pro + Gly60Glu
Asn61Asp + Thr65Ser
Asp59Glu + Gly60Gln
Asp59Glu + Gly60Pro
Gly62Asp + Gly64Gln
Asn61Ser + Thr65Pro
Thr58Asp + Gly60Gln
Gly60Ser + Gly62Gln
Asp59Glu + Gly62Pro
Gly62Asp + Thr65Gly
Asn61Gln + Thr65Pro
Asn61Gln + Gly64Asn
Gly60Gln + Gly62Gln
Thr58Gly + Asn61Asp
Thr58Gln + Gly62Gln
Gly60Asn + Gly62Gln
Gly60Asn + Gly62Asp
Thr58Glu + Gly62Pro
Gly64Asp + Thr65Pro
Asp59Glu + Thr65Ser
Gly60Gln + Thr65Gln
Asp59Glu + Gly62Ser
Asn61Glu + Gly64Gln
Asp59Glu + Gly60Ser
Gly60Ser + Gly62Ser
Thr58Gly + Gly64Glu
Thr58Asn + Asn61Ser
Asp59Glu + Thr65Asn
Gly60Asn + Asn61Ser
Gly60Glu + Gly62Gln
Thr58Pro + Gly62Asp
Thr58Ser + Asp59Glu
Thr58Asp + Gly62Pro
Asn61Asp + Gly62Pro
Thr58Ser + Gly62Ser
Gly60Asn + Gly62Ser
Gly64Pro + Thr65Asn
Gly64Asp + Thr65Gln
Gly64Glu + Thr65Pro
Gly64Asn + Thr65Pro
Asn61Ser + Gly64Asp
表4
环1-三突变变异体
Thr58Pro + Gly64Ser + Thr65Gly
Asn61Glu + Gly62Gln + Thr65Gln
Thr58Gln + Gly62Ser + Gly64Pro
Asp59Glu + Gly62Asn + Thr65Pro
Thr58Pro + Gly60Glu + Gly64Asn
Thr58Asn + Gly64Ser + Thr65Gly
Asp59Glu + Asn61Gln + Gly64Ser
Thr58Asn + Asn61Glu + Thr65Pro
Gly60Pro + Asn61Asp + Thr65Pro
Thr58Glu + Gly62Gln + Gly64Asn
Thr58Asn + Gly62Glu + Gly64Pro
Thr58Ser + Asp59Glu + Thr65Asn
Thr58Glu + Gly62Asn + Gly64Gln
Thr58Pro + Asp59Glu + Gly62Ser
Gly60Asn + Asn61Asp + Gly62Asn
Gly62Asp + Gly64Ser + Thr65Pro
Asp59Glu + Gly60Asn + Thr65Asn
Thr58Gly + Gly60Pro + Thr65Gly
Thr58Gln + Gly60Glu + Asn61Gln
Gly60Pro + Asn61Gln + Gly64Asp
Thr58Asn + Gly62Asn + Thr65Pro
Thr58Pro + Asp59Glu + Gly64Asn
Thr58Pro + Ash61Ser + Gly62Asp
Thr58Asp + Gly50Pro + Gly62Asn
Thr58Ser + Gly60Pro + Asn61Asp
Gly60Gln + Gly62Gln + Gly64Pro
Asp59Glu + Asn61Gln + Thr65Asn
Gly60Asp + Asn61Gln + Thr65Pro
Asn61Glu + Gly62Glu + Thr65Pro
Thr58Asp + Asp59Glu + Thr65Gln
Thr58Glu + Asp59Glu + Thr65Ser
Thr58Glu + Asp59Glu + Thr65Gly
表5
环1-四突变变异体
Asp59Glu + Gly60Pro + Gly62Asn + Gly64Gln
Thr58Pro + Gly60Pro + Gly62Glu + Thr65Gln
Thr58Ser + Gly60Pro + Gly64Gln + Thr65Pro
Thr58Pro + Asp59Glu + Asn61Ser + Gly62Asn
Thr58Glu + Gly60Asn + Gly64Ser + Thr65Pro
Thr58Glu + Gly60Ser + Asn61Gln + Thr65Gln
Thr58Pro + Asp59Glu + Gly60Asn + Asn61Gln
Thr58Gly + Gly60Pro + Gly62Glu + Thr65Ser
Gly60Asp + Gly62Ser + Gly64Gln + Thr65Gly
Thr58Asn + Gly60Pro + Gly62Ser + Gly64Asp
Thr58Pro + Asp59Glu + Gly60Asn + Gly62Asn
Asn61Asp + Gly62Gln + Gly64Pro + Thr65Asn
Thr58Asp + Gly60Pro + Gly62Asn + Gly64Ser
Thr58Asp + Gly60Gln + Gly62Pro + Thr65Asn
Thr58Asn + Asn61Asp + Gly62Glu + Thr65Gly
Thr58Ser + Gly60Ser + Asn61Asp + Gly62Glu
Thr58Glu + Asp59Glu + Asn61Gln + Thr65Ser
Thr58Asp + Asp59Glu + Gly62Pro + Thr65Gly
Gly60Asp + Asn61Asp + Gly62Ser + Thr65Gly
Gly60Glu + Asn61Glu + Gly62Gln + Gly64Pro
Thr58Pro + Gly60Asp + Asn61Asp + Gly64Asn
Asp59Glu + Gly60Asp + Gly64Gln + Thr65Ser
Asp59Glu + Gly60Asp + Gly64Gln + Thr65Pro
Thr58Pro + Asp59Glu + Gly60Asp + Gly64Asn
Thr58Ser + Asp59Glu + Gly60Asp + Gly62Asn
Asp59Glu + Gly60Glu + Asn61Glu + Gly62Ser
Thr58Glu + Asp59Glu + Gly60Asp + Thr65Pro
Asp59Glu + Gly60Asp + Gly62Glu + Thr65Pro
Thr58Ser + Asp59Glu + Asn61Asp + Gly62Asp
表6
环2-单突变变异体
Val 94Ala
Val 94Asn
Val 94Asp
Val 94Cys
Val 94Gln
Val 94Glu
Val 94Gly
Val 94His
Val 94Met
Val 94Pro
Val 94Ser
Val 94Thr
Leu 95Ala
Leu 95Asn
Leu 95Asp
Leu 95Cys
Leu 95Gln
Leu 95Glu
Leu 95Gly
Leu 95His
Leu 95Ile
Leu 95Met
Leu 95Pro
Leu 95Ser
Leu 95Thr
Leu 95Val
Asn 96Asp
Asn 96Gln
Asn 96Glu
Asn 96Ser
Ser 97Asp
Ser 97Glu
Ser 98Asp
Ser 98Glu
Gly 99Asn
Gly 99Asp
Gly 99Gln
Gly 99Glu
Gly 99Pro
Gly 99Ser
Ser100Asp
Ser100Glu
Gly101Asn
Gly101Asp
Gly101Gln
Gly101Glu
Gly101Pro
Gly101Ser
Ser102Asp
Ser102Glu
Tyr103Ala
Tyr103Asn
Tyr103Asp
Tyr103Cys
Tyr103Gln
Tyr103Glu
Tyr103Gly
Tyr103His
Tyr103Ile
Tyr103Leu
Tyr103Met
Tyr103Pro
Tyr103Ser
Tyr103Thr
Tyr103Val
Ser104Asp
Ser104Glu
Gly105Asn
Gly105Asp
Gly105Gln
Gly105Glu
Gly105Pro
Gly105Ser
Ile106Ala
Ile106Asn
Ile106Asp
Ile106Cys
Ile106Gln
Ile106Glu
Ile106Gly
Ile106His
Ile106Leu
Ile106Met
Ile106Pro
Ile106Ser
Ile106Thr
Ile106Val
表7
环2-双突变变异体
Val 94Gln + Ser100Glu
Asn 96Ser + Gly 99Gln
Gly 99Glu + Ile106Met
Ser 97Asp + Gly105Gln
Ser102Glu + Tyr103Asn
Ser100Glu + Ile106Gly
Asn 96Asp + Ile106Ser
Gly 99Ser + Ser100Glu
Gly101Gln + Gly105Ser
Asn 96Asp + Gly 99Pro
Asn 96Gln + Ser104Asp
Leu 95Glu + Tyr103Pro
Asn 96Ser + Gly101Asp
Leu 95Val + Ser 97Glu
Leu 95Ala + Gly 99Pro
Val 94Thr + Gly 99Glu
Leu 95Gln + Ile106Ala
Leu 95Pro + Ile106Ala
Leu 95Val + Ser100Glu
Gly101Gln + Tyr103Gly
Asn 96Asp + Ile106Pro
Leu 95Ser + Ser100Asp
Gly 99Pro + Ser100Asp
Val 94Thr + Leu 95Asp
Ser100Glu + Ile106Gln
Ser100Asp + Gly101Pro
Leu 95Cys + Gly105Asp
Ser 98Glu + Gly101Ser
Gly101Asn + Tyr103Gln
Val 94Met + Ser102Glu
Gly 99Pro + Gly105Asp
Val 94Asn + Ser102Asp
Val 94Cys + Ile106Thr
Val 94Gly + Ser104Asp
Val 94Pro + Gly101Pro
Val 94Ser + Gly105Glu
Val 94Met + Ser 98Asp
Val 94Gln + Ser104Asp
Asn 96Ser + Ser 98Asp
Ser104Glu + Ile106Gln
Val 94Asn + Tyr103Thr
Tyr103Ala + Ser104Asp
Gly 99Pro + Ser104Glu
Leu 95Cys + Tyr103Leu
Val 94Gly + Gly 99Ser
Gly101Gln + Tyr103Ser
Ser 97Asp + Gly105Pro
Ser 98Asp + Gly105Asn
Gly 99Gln + Ser100Glu
Gly 99Pro + Ser100Glu
Tyr103Ile + Gly105Ser
Ser100Glu + Gly105Pro
Asn 96Glu + Tyr103Gly
Val 94Met + Gly105Asn
Val 94His + Leu 95Glu
Ser100Asp + Tyr103Met
Leu 95Met + Gly101Gln
Gly 99Pro + Gly101Asp
Asn 96Asp + Gly101Asn
Val 94Ala + Gly 99Ser
表8
环2-三突变变异体
Val 94Gln + Leu 95Thr + Ser100Glu
Ser100Glu + Tyr103Met + Ile106Thr
Ser 97Asp + Gly101Gln + Ile106Asn
Leu 95Gln + Gly105Ser + Ile106Gln
Leu 95His + Gly 99Gln + Ser100Glu
Val 94His + Leu 95Met + Asn 96Gln
Val 94Asn + Ser 97Glu + Tyr103Pro
Ser 97Asp + Gly101Ser + Tyr103Met
Ser 97Glu + Tyr103Met + Ile106Gln
Val 94Thr + Gly 99Pro + Gly105Asp
Leu 95Gly + Ser 98Glu + Tyr103Cys
Leu 95Pro + Gly101Asp + Tyr103Gly
Leu 95Val + Gly 99Pro + Ser104Asp
Leu 95Ile + Gly105Ser + Ile106Met
Ser 97Glu + Gly101Asn + Ile106Thr
Asn 96Ser + Gly101Asp + Tyr103Asn
Val 94Pro + Ser 98Glu + Ile106Asn
Gly 99Pro + Tyr103Glu + Ile106Ser
Asn 96Asp + Gly 99Asn + Tyr103Pro
Leu 95Gln + Ser 97Asp + Ile106Val
Val 94Pro + Ser 98Glu + Tyr103Val
Leu 95Asn + Gly101Pro + Ile106His
Val 94Ala + Leu 95His + Ser100Glu
Val 94Asn + Gly101Ser + Ser104Asp
Val 94Cys + Gly105Ser + Ile106Thr
Leu 95Gly + Gly101Asp + Ile106Cys
Ser102Glu + Tyr103Ile + Ile106Gly
Val 94Pro + Leu 95Asp + Ile106Pro
Val 94Pro + Leu 95Ile + Ile106Cys
Ser100Asp + Gly101Asp + Ile106Asn
Leu 95Glu + Asn 96Glu + Tyr103Pro
Gly101Glu + Ser102Asp + Tyr103Leu
Val 94Pro + Gly101Asp + Ser102Glu
Asn 96Asp + Ser 97Asp + Gly 99Pro
Ser 97Asp + Ser 98Asp + Gly 99Glu
Asn 96Glu + Ser 97Asp + Gly 99Asp
Gly 99Glu + Ser100Asp + Gly101Asp
Gly 99Glu + Ser100Glu + Gly101Glu
Leu 95Gln + Ser102Asp + Gly105Asp
Asn 96Ser + Ser102Glu + Ser104Glu
Ser102Asp + Ser104Glu + Gly105Pro
Ser102Asp + Tyr103Ser + Ser104Glu
Val 94Cys + Ser 97Glu + Gly 99Asp
Asn 96Asp + Ser 98Glu + Ile106Thr
Asn 96Asp + Ser 98Asp + Ile106Cys
Asn 96Glu + Ser 98Glu + Gly105Asn
Ser 98Glu + Ser100Glu + Ile106Ser
Ser 98Asp + Ser100Asp + Gly101Asn
Gly101Asp + Ser102Asp + Ser104Glu
Leu 95Glu + Ser 98Asp + Gly101Asp
Val 94Ser + Ser100Glu + Ser102Asp
Leu 95Val + Ser 97Glu + Ser100Glu
Asn 96Gln + Ser 97Asp + Ser100Asp
Asn 96Ser + Ser 97Asp + Ser100Asp
Val 94Asp + Gly 99Asn + Ser102Glu
Val 94Asp + Ser102Glu + Ile106His
Leu 95Asp + Gly 99Glu + Ser102Asp
Val 94Thr + Gly101Asp + Ser104Glu
Gly 99Pro + Gly101Asp + Ser104Asp
Ser 98Asp + Gly101Asp + Tyr103Pro
表9
环2-四突变变异体
Val 94Met + Leu 95Asn + Ser 98Asp + Gly105Ser
Asn 96Ser + Gly101Glu + Tyr103Ile + Gly105Asn
Leu 95Val + Ser 97Asp + Gly 99Asn + Tyr103Ile
Leu 95Ile + Asn 96Gln + Gly 99Asn + Tyr103Asp
Val 94His + Ser100Glu + Tyr103Ile + Ile106Met
Ser 97Glu + Tyr103Ile + Gly105Gln + Ile106Ala
Val 94Thr + Leu 95Ile + Ser 97Asp + Gly99Ser
Val 94Thr + Leu 95Cys + Gly 99Asp + Ile106Val
Leu 95Cys + Ser 98Glu + Gly 99Asn + Ile106Met
Asn 96Gln + Ser 98Asp + Gly105Asn + Ile106Gly
Leu 95Thr + Ser100Glu + Tyr103Val + Ile106Pro
Val 94Thr + Leu 95Val + Asn 96Glu + Tyr103Asn
Val 94His + Leu 95Asn + Gly 99Gln + Gly105Gln
Val 94Ala + Leu 95Thr + Ser 98Asp + Tyr103Ile
Leu 95Val + Gly101Gln + Tyr103Cys + Ile106Glu
Gly 99Pro + Ser100Glu + Gly101Ser + Tyr103Thr
Leu 95Ala + Gly101Asp + Tyr103Ser + Gly105Asn
Gly 99Asn + Tyr103Asp + Gly105Asn + Ile106Thr
Asn 96Asp + Ser 97Asp + Gly101Ser + Tyr103Thr
Val 94Ala + Asn 96Glu + Ser 97Asp + Gly105Asn
Val 94Cys + Leu 95Gln + Asn 96Glu + Ser 97Glu
Val 94Gly + Asn 96Gln + Ser104Glu + Gly105Glu
Ser 97Asp + Ser 98Glu + Gly101Asn + Tyr103Val
Ser 97Glu + Ser 98Asp + Gly 99Gln + Gly101Gln
Leu 95Gly + Asn 96Ser + Ser 97Glu + Ser 98Glu
Asn 96Glu + Ser 97Glu + Ser 98Asp + Gly101Pro
Asn 96Asp + Ser 97Asp + Ser 98Asp + Tyr103Leu
Asn 96Asp + Ser 97Asp + Ser 98Glu + Tyr103Gly
Leu 95Glu + Asn 96Glu + Ser100Asp + Ile106Ala
Ser 97Asp + Gly 99Glu + Tyr103Cys + Gly105Asn
Val 94Gly + Asn 96Ser + Ser 97Glu + Gly 99Asp
Asn 96Ser + Tyr103Leu + Ser104Asp + Ile106Glu
Leu 95Ala + Asn 96Glu + Gly 99Ser + Ser100Asp
Asn 96Asp + Ser 97Glu + Ser100Glu + Ile106Val
Val 94Cys + Ser 97Asp + Ser 98Asp + Ser100Asp
Ser 97Asp + Ser 98Asp + Ser100Glu + Tyr103Thr
Asn 96Glu + Gly 99Glu + Gly101Asp + Tyr103Ala
Val 94Asp + Leu 95Ala + Gly101Glu + Ser102Glu
Ser 98Glu + Gly 99Pro + Ser100Asp + Gly101Glu
Val 94Glu + Asn 96Gln + Gly101Glu + Ile106Thr
Asn 96Ser + Ser102Asp + Tyr103Asn + Ile106Asp
Leu 95Cys + Ser 98Asp + Gly 99Asp + Gly101Asp
Ser100Glu + Gly101Pro + Ser102Glu + Ile106Ala
Val 94Cys + Ser100Glu + Gly101Pro + Ser102Glu
Val 94Gln + Ser100Asp + Ser102Glu + Tyr103Ile
Ser 97Asp + Ser100Glu + Gly101Glu + Tyr103Asn
Val 94Glu + Asn 96Glu + Gly 99Glu + Gly105Pro
Asn 96Gln + Ser100Glu + Gly101Asp + Ile106Glu
Val 94Pro + Leu 95Ala + Ser 97Asp + Ser100Asp
Leu 95Glu + Gly101Asn + Ser102Glu + Tyr103Ile
Val 94Asn + Asn 96Glu + Gly101Asp + Gly105Asp
Val 94Asp + Gly 99Asn + Ser100Asp + Ser102Asp
Val 94Asp + Leu 95Val + Ser102Asp + Tyr103Ala
Leu 95Glu + Gly 99Pro + Ser102Asp + Ser104Glu
Leu 95Ser + Asn 96Gln + Gly101Glu + Ser104Asp
Val 94Gln + Asn 96Gln + Gly101Glu + Ser104Glu
Gly101Asp + Tyr103Val + Ser104Glu + Ile106Val
Asn 96Asp + Ser102Asp + Tyr103Gly + Gly105Asp
Ser100Glu + Ser102Asp + Ser104Asp + Gly105Ser
Val 94Asp + Ser 97Asp + Ser100Asp + Gly105Gln
表10
环3-单突变变异体
Leu125Ala
Leu125Asn
Leu125Asp
Leu125Cys
Leu125Gln
Leu125Glu
Leu125Gly
Leu125His
Leu125Ile
Leu125Met
Leu125Pro
Leu125Ser
Leu125Thr
Leu125Val
Gly126Asn
Gly126Asp
Gly126Gln
Gly126Glu
Gly126Pro
Gly126Ser
Gly127Asn
Gly127Asp
Gly127Gln
Gly127Glu
Gly127Pro
Gly127Ser
Ala128Asn
Ala128Asp
Ala128Gln
Ala128Glu
Ala128Gly
Ala128His
Ala128Pro
Ala128Ser
Ala128Thr
Ser129Asp
Ser129Glu
Gly130Asn
Gly130Asp
Gly130Gln
Gly130Glu
Gly130Pro
Gly130Ser
Ser131Asp
Ser131Glu
Thr132Asn
Thr132Asp
Thr132Gln
Thr132Glu
Thr132Gly
Thr132Pro
Thr132Ser
表11
环3-双突变变异体
Leu125Gln + Ser129Glu
Gly130Gln + Thr132Asn
Gly126Asp + Gly127Pro
Leu125Ile + Ser131Glu
Gly126Gln + Gly130Ser
Gly126Pro + Thr132Pro
Gly126Asp + Thr132Ser
Ala128Gly + Thr132Ser
Leu125Ala + Gly126Glu
Leu125Gln + Ser129Asp
Gly126Pro + Ser131Asp
Gly127Ser + Ser129Glu
Gly130Asn + Ser131Glu
Leu125Glu + Ala128Gln
Gly126Ser + Thr132Ser
Leu125Ala + Ser129Asp
Gly127Pro + Thr132Asp
Gly126Pro + Gly130Ser
Leu125Ala + Ala128Asp
Gly127Gln + Ser131Asp
Leu125Ala + Thr132Ser
Gly127Pro + Thr132Gly
Gly126Glu + Thr132Asn
Leu125Gly + Ser129Glu
Ala128Thr + Gly130Asp
Ala128His + Thr132Asp
Leu125Gly + Gly126Asn
Gly126Ser + Ser129Glu
Leu125Asn + Thr132Asn
Gly126Glu + Ala128Gly
Ser131Asp + Thr132Gln
Ser129Glu + Gly130Pro
Gly127Asn + Ser129Asp
Leu125Glu + Thr132Asn
Gly126Gln + Gly127Glu
Ala128Thr + Ser131Asp
Leu125Pro + Ser131Glu
Gly126Pro + Thr132Gly
Gly126Gln + Ser129Asp
Leu125Gly + Gly126Ser
Gly127Gln + Ser129Asp
Gly126Gln + Gly130Glu
Gly127Asp + Gly130Asn
Leu125Glu + Gly126Ser
Leu125Ile + Thr132Asp
Gly126Asn + Gly127Asp
Leu125Cys + Gly126Gln
Leu125Glu + Ala128Asn
Gly126Gln + Ala128Asn
Gly126Asp + Gly130Asn
Leu125Ser + Gly130Ser
Gly126Asn + Ser129Asp
Leu125Ser + Ser131Asp
Ser129Asp + Thr132Asn
Gly127Glu + Ala128Thr
Leu125Thr + Gly130Pro
Ala128Pro + Thr132Asp
Gly127Ser + Ser131Glu
Gly126Asn + Thr132Glu
Gly130Asp + Thr132Asn
表12
环3-三突变变异体
Leu125Asn + Gly126Pro + Ser129Glu
Gly126Asn + Ser129Asp + Gly130Pro
Leu125Ala + Gly126Ser + Ser131Glu
Gly127Gln + Ala128Thr + Ser129Glu
Leu125Thr + Ser129Asp + Gly130Pro
Gly126Pro + Ala128Thr + Thr132Asp
Gly126Asn + Gly130Asn + Thr132Gln
Leu125Thr + Gly130Gln + Thr132Gln
Ala128Gly + Ser129Asp + Thr132Asn
Leu125Met + Ala128Gln + Ser129Asp
Lsu125Gly + Ala128Asp + Gly130Pro
Leu125Ile + Gly127Ser + Thr132Asp
Leu125Val + Gly127Glu + Gly130Ser
Gly127Ser + Gly130Asn + Thr132Asn
Leu125Thr + Gly126Ser + Ala128Glu
Leu125Gln + Gly126Pro + Ser131Glu
Leu125Glu + Gly130Asn + Thr132Asn
Gly126Asn + Gly127Pro + Ser129Asp
Gly126Pro + Gly127Pro + Ala128His
Gly127Asn + Ala128His + Ser129Asp
Leu125Thr + Gly130Pro + Thr132Ser
Leu125Cys + Gly127Gln + Ser129Asp
Gly127Asn + Gly130Glu + Thr132Gly
Gly127Gln + Gly130Ser + Thr132Asp
Leu125Thr + Ser129Asp + Thr132Pro
Leu125Cys + Gly127Glu + Gly130Pro
Ala128Gly + Ser129Glu + Gly130Gln
Gly127Asp + Gly130Ser + Thr132Pro
Leu125Pro + Gly126Asn + Gly127Asn
Gly126Glu + Ala128Ser + Gly130Ser
Leu125Gly + Gly127Pro + Ser129Asp
Gly126Gln + Ala128Glu + Gly130Gln
Ala128Gly + Gly130Asn + Thr132Asp
Leu125Met + Gly126Pro + Gly127Gln
Leu125Gln + Ala128Asn + Ser129Asp
Gly127Ser + Ala128Asn + Thr132Gly
Gly126Gln + Gly127Asp + Gly130Ser
Gly127Asp + Ala128Pro + Thr132Pro
Leu125His + Ser129Asp + Thr132Pro
Leu125Ile + Gly126Gln + Thr132Gln
Gly126Asp + Gly130Ser + Thr132Asn
Gly127Asn + Ser129Glu + Gly130Pro
Leu125Pro + Gly127Asp + Gly130Asn
Leu125Ile + Ala128His + Ser131Asp
Gly126Pro + Ala128His + Thr132Pro
Leu125Val + Ala128Gln + Ser129Glu
Leu125Ser + Gly126Asp + Thr132Gly
Leu125Pro + Gly126Asp + Gly127Asp
Ala128Ser + Gly130Asp + Ser131Asp
Gly130Asp + Ser131Glu + Thr132Asn
Gly126Asn + Gly130Asp + Ser131Glu
Gly126Gln + Gly130Asp + Ser131Glu
Gly130Gln + Ser131Glu + Thr132Glu
Ala128Thr + Ser131Glu + Thr132Asp
Ala128Thr + Ser131Asp + Thr132Glu
Ala128Pro + Ser131Asp + Thr132Asp
Gly127Asp + Ala128Asp + Thr132Gln
Leu125Glu + Gly126Glu + Gly130Asn
Ala128Asp + Ser129Asp + Gly130Asn
Ser129Asp + Gly130Glu + Thr132Pro
表13
环3-四突变变异体
Leu125Asn + Ala128Asn + Ser131Glu + Thr132Pro
Leu125Ala + Gly126Gln + Gly127Pro + Gly1305er
Leu125Pro + Gly126Asn + Gly127Gln + Ala128Pro
Leu125Gly + Gly126Asn + Ser129Glu + Gly130Asn
Ala128Gly + Gly130Asn + Ser131Asp + Thr132Ser
Leu125Ile + Gly126Pro + Ala128Gly + Thr132Glu
Leu125Thr + Gly130Pro + Ser131Asp + Thr132Gly
Leu125Asp + Gly126Pro + Gly127Ser + Thr132Gln
Leu125Gly + Gly126Pro + Gly127Asp + Thr132Pro
Leu125Val + Gly127Glu + Gly130Asn + Thr132Asn
Leu125Cys + Ala128Pro + Gly130Ser + Ser131Glu
Gly126Gln + Gly127Ser + Ala128Thr + Gly130Glu
Leu125Gly + Gly127Ser + Gly130Ser + Thr132Gly
Leu125Pro + Gly126Ser + Ala128His + Thr132Asn
Leu125Val + Gly126Gln + Gly127Pro + Ser129Asp
Leu125Cys + Gly130Pro + Ser131Asp + Thr132Ser
Leu125Gly + Gly127Gln + Ser129Glu + Thr132Ser
Leu125Met + Ala128Ser + Gly130Pro + Thr132Gly
Leu125Ala + Gly126Asp + Ala128His + Thr132Ser
Gly127Gln + Gly130Glu + Ser131Asp + Thr132Asn
Gly126Ser + Gly127Gln + Gly130Glu + Ser131Glu
Gly127Pro + Gly130Asp + Ser131Glu + Thr132Pro
Ala128Gly + Gly130Pro + Ser131Asp + Thr132Glu
Gly126Asn + Ala128Pro + Ser131Asp + Thr132Asp
Gly126Gln + Ala128His + Ser131Asp + Thr132Asp
Leu125Thr + Gly127Glu + Ala128Asp + Gly130Gln
Gly126Asn + Ala128Gln + Ser129Glu + Gly130Asp
Ala128Gly + Ser129Glu + Gly130Glu + Thr132Gln
Gly126Gln + Ser129Asp + Gly130Glu + Thr132Asn
Gly126Asn + Ala128Thr + Ser129Glu + Gly130Glu
Gly126Ser + Gly127Asn + Ser129Glu + Gly130Glu
Leu125His + Gly127Glu + Ala128Glu + Ser129Asp
Leu125Thr + Gly127Glu + Ala128Asp + Ser129Glu
Gly126Pro + Gly127Glu + Ala128Asp + Ser129Glu
Ala128Gly + Ser129Glu + Gly130Asp + Ser131Asp
Gly126Gln + Ser129Asp + Gly130Asp + Ser131Asp
Gly127Ser + Ser129Glu + Gly130Glu + Ser131Glu
Ala128Gly + Ser129Asp + Gly130Asp + Ser131Glu
Gly126Asn + Ser129Asp + Gly130Glu + Ser131Asp
Ala128Asn + Ser129Asp + Gly130Asp + Ser131Asp
Gly127Pro + Ser129Glu + Gly130Asp + Ser131Glu
Gly126Ser + Gly130Asp + Ser131Asp + Thr132Glu
Leu125Met + Gly127Glu + Ser129Asp + Gly130Asp
Gly126Pro + Gly127Asp + Ser129Glu + Gly130Asp
Leu125Val + Gly127Glu + Ser129Asp + Gly130Asp
Gly127Asp + Ala128Asn + Ser129Asp + Gly130Glu
Gly127Glu + Ala128Thr + Ser129Glu + Gly130Ser
Gly127Asp + Ala128His + Ser129Glu + Gly130Ser
Gly126Ser + Gly127Glu + Ala128Thr + Ser129Glu
Gly126Pro + Gly127Asp + Ala128Asp + Gly130Glu
Leu125Gly + Gly126Asp + Gly127Glu + Ser129Glu
Gly126Asp + Gly127Glu + Ala128Ser + Ser129Asp
Gly127Ser + Ser129Asp + Gly130Gln + Ser131Asp
Ala128Gly + Ser129Glu + Gly130Asn + Ser131Glu
Leu125Val + Ser129Asp + Ser131Asp + Thr132Ser
Leu125Gly + Ala128Gly + Ser129Glu + Ser131Asp
Leu125Cys + Gly126Pro + Ser129Glu + Ser131Glu
Gly127Ser + Ala128Asn + Ser129Asp + Ser131Asp
Gly126Asn + Ser129Asp + Ser131Glu + Thr132Gln
Gly126Pro + Ala128Pro + Ser129Asp + Ser131Glu
表14
环4-单突变变异体
Gly153Asn
Gly153Asp
Gly153Gln
Gly153Glu
Gly153Pro
Gly153Ser
Asn154Asp
Asn154Gln
Asn154Glu
Asn154Ser
Ser155Asp
Ser155Glu
Gly156Asn
Gly156Asp
Gly156Gln
Gly156Glu
Gly156Pro
Gly156Ser
Asn157Asp
Asn157Gln
Asn157Glu
Asn157Ser
Ser158Asp
Ser158Glu
Gly159Asn
Gly159Asp
Gly159Gln
Gly159Glu
Gly159Pro
Gly159Ser
Ser160Asp
Ser160Glu
Thr161Asn
Thr161Asp
Thr161Gln
Thr161Glu
Thr161Gly
Thr161Pro
Thr161Ser
Asn162Asp
Asn162Gln
Asn162Glu
Asn162Ser
Thr163Asn
Thr163Asp
Thr163Gln
Thr163Glu
Thr163Gly
Thr163Pro
Thr163Ser
Ile164Ala
Ile164Asn
Ile164Asp
Ile164Cys
Ile164Gln
Ile164Glu
Ile164Gly
Ile164His
Ile164Leu
Ile164Met
Ile164Pro
Ile164Ser
Ile164Thr
Ile164Val
Gly165Asn
Gly165Asp
Gly165Gln
Gly165Glu
Gly165Pro
Gly165Ser
Tyr166Ala
Tyr166Asn
Tyr166Asp
Tyr166Cys
Tyr166Gln
Tyr166Glu
Tyr166Gly
Tyr166His
Tyr166Ile
Tyr166Leu
Tyr166Met
Tyr166Pro
Tyr166Ser
Tyr166Thr
Tyr166Val
表15
环4-双突变变异体
Asn154Ser + Ser155Glu
Gly153Glu + Ile164Ala
Gly159Glu + Gly165Pro
Asn154Glu + Gly156Ser
Gly156Pro + Gly159Asp
Gly153Ser + Ser160Asp
Ser155Asp + Gly165Ser
Ile164Gly + Gly165Glu
Gly159Glu + Ile164Asn
Asn157Gln + Asn162Ser
Gly156Asn + Thr161Gln
Asn162Glu + Tyr166Gln
Asn157Glu + Gly165Gln
Asn162Glu + Gly165Gln
Gly153Pro + Ile164Cys
Thr163Glu + Gly165Asn
Gly153Asp + Ile164Asn
Gly153Ser + Thr161Asp
Gly156Asp + Tyr166Gln
Gly165Pro + Tyr1665er
Gly153Ser + Asn162Glu
Gly159Glu + Tyr166Leu
Gly156Asn + Thr161Glu
Asn154Gln + Ser158Glu
Ser160Asp + Thr161Asn
Ser155Glu + Gly165Pro
Asn154Ser + Tyr166Ile
Gly153Asp + Thr161Asn
Ser158Asp + Thr161Pro
Asn162Glu + Thr163Ser
Gly159Gln + Tyr166Cys
Thr161Gly + Asn162Ser
Gly159Asp + Tyr166Asn
Ser160Glu + Ile164Thr
Ser158Asp + Ile164Leu
Asn154Ser + Asn157Asp
Asn154Asp + Thr161Gln
Gly156Ser + Ser160Glu
Gly156Gln + Tyr166Ile
Gly153Asn + Tyr166Ala
Asn154Asp + Ile164Ser
Ser155Asp + Thr163Asn
Gly156Glu + Gly159Gln
Gly156Ser + Asn162Asp
Gly153Ser + Asn157Glu
Ser160Glu + Gly165Pro
Gly153Pro + Asn157Asp
Gly153Gln + Ser160Asp
Asn154Gln + Thr163Asn
Gly156Ser + Gly159Asp
Gly159Asp + Thr161Gln
Gly159Ser + Tyr166Glu
Asn157Asp + Tyr166Leu
Thr161Asp + Asn162Gln
Asn162Gln + Gly165Glu
Gly156Asp + Asn162Gln
Asn154Gln + Ile164Gln
Asn157Asp + Asn162Ser
Asn154Asp + Ile164Asn
Gly159Asp + Gly165Pro
表16
环4-三突变变异体
Gly153Gln + Asn154Ser + Ser155Glu
Gly156Asp + Thr161Pro + Tyr166Ile
Asn157Glu + Gly159Gln + Asn162Gln
Asn154Glu + Asn162Gln + Ile164Leu
Gly153Pro + Gly159Gln + Ile164Asn
Asn154Ser + Ile164Met + Tyr166Cys
Gly159Gln + Ser160Asp + Ile164Thr
Gly156Asn + Thr161Ser + Ile164Cys
Gly156Asn + Ile164His + Tyr166Asp
Asn157Glu + Asn162Gln + Ile164Ala
Ser160Asp + Thr163Ser + Ile164Gly
Asn154Glu + Thr161Pro + Ile164Met
Thr163Asn + Ile164Asn + Gly165Pro
Asn157Glu + Asn162Gln + Ile164Thr
Asn162Gln + Ile164Thr + Gly165Glu
Asn154Gln + Gly159Asp + Tyr166Thr
Asn157Gln + Thr161Gly + Thr163Pro
Ser158Glu + Thr163Ser + Gly165Ser
Gly153Pro + Asn157Asp + Thr163Asn
Gly156Pro + Ser158Asp + Gly165Gln
Asn157Gln + Thr163Gly + Gly165Pro
Ser155Glu + Gly159Ser + Tyr166Thr
Gly153Gln + Asn154Gln + Gly165Glu
Gly153Gln + Asn154Ser + Gly159Pro
Thr161Gly + Asn162Asp + Gly165Asn
Ser158Glu + Gly159Ser + Gly165Gln
Ser155Asp + Gly159Asn + Ile164Ser
Asn157Gln + Thr163Gly + Gly165Glu
Gly153Ser + Gly159Ser + Asn162Asp
Gly153Ser + Ser158Glu + Ile164Thr
Gly153Asp + Thr161Ser + Gly165Asn
Thr161Ser + Ile164Ser + Tyr166Pro
Ser155Asp + Gly156Asn + Ile164Asn
Asn154Glu + Gly159Gln + Tyr166Ile
Gly153Gln + Gly156Gln + Thr161Asp
Gly159Asn + Thr163Asn + Tyr166Ser
Ser155Asp + Thr163Pro + Ile164Gly
Gly153Ser + Ser160Asp + Ile164Pro
Asn162Glu + Ile164Gln + Tyr166Gln
Gly156Glu + Asn157Glu + Thr161Gly
Gly159Asp + Ser160Glu + Ile164Gln
Ser155Asp + Gly156Glu + Thr163Asn
Ser155Glu + Gly156Asp + Gly165Ser
Ser155Glu + Gly156Asp + Ile164Met
Ser160Glu + Thr161Asp + Gly165Asn
Asn162Asp + Thr163Asp + Gly165Asn
Asn157Asp + Ser158Glu + Thr161Glu
Gly159Asp + Thr161Glu + Thr163Pro
Gly153Asp + Asn154Ser + Gly165Asp
Gly153Glu + Asn162Gln + Gly165Asp
Asn154Ser + Thr161Glu + Thr163Asp
Ser158Glu + Ser160Asp + Thr163Asn
Ser158Glu + Ser160Glu + Thr163Gly
Ser158Asp + Ser160Asp + Ile164Gly
Asn157Asp + Gly159Glu + Thr163Gln
Thr163Glu + Gly165Asp + Tyr166Pro
Ser160Asp + Asn162Glu + Gly165Asn
Asn154Glu + Gly156Asp + Asn157Glu
Gly159Asp + Ser160Asp + Asn162Glu
Asn154Glu + Ser155Glu + Asn157Asp
表17
环4-四突变变异体
Gly153Pro + Gly159Ser + Thr163Pro + Gly165Asn
Asn154Gln + Asn162Glu + Ile164Cys + Gly165Pro
Asn154Ser + Asn157Asp + Thr161Gly + Ile164Asn
Gly159Glu + Thr161Gln + Thr163Asn + Gly165Gln
Gly159Ser + Ser160Glu + Thr163Gln + Tyr166Ile
Gly156Ser + Ser160Glu + Thr161Asn + Gly165Pro
Gly153Ser + Gly159Ser + Ser160Asp + Gly165Ser
Gly153Asn + Gly159Glu + Ile164Cys + Gly165Pro
Asn154Ser + Gly156Glu + Thr161Pro + Tyr166His
Gly153Ser + Ser160Glu + Gly165Asn + Tyr166Cys
Asn157Gln + Thr163Gly + Ile164His + Gly165Gln
Asn154Ser + Ser155Glu + Asn162Ser + Ile164His
Gly156Pro + Asn157Gln + Ser160Glu + Gly165Ser
Gly153Gln + Ser155Asp + Thr161Gln + Tyr166Gly
Asn157Ser + Thr161Gln + Thr163Asp + Ile164Thr
Gly153Ser + Ser160Asp + Asn162Gln + Tyr166Thr
Gly153Pro + Ash154Gln + Asn157Gln + Ser160Glu
Gly156Glu + Asn162Gln + Thr163Ser + Gly165Gln
Gly156Asn + Asn157Glu + Thr161Gly + Thr163Ser
Gly153Ser + Ser158Glu + Gly159Asp + Thr163Gly
Asn154Gln + Ser158Asp + Gly159Asp + Thr163Asn
Ser158Asp + Gly159Asp + Thr163Asn + Tyr166Pro
Gly159Asp + Ser160Glu + Thr163Ser + Gly165Ser
Asn154Glu + Ser155Asp + Thr163Gln + Tyr165Met
Gly153Glu + Asn154Asp + Ser155Glu + Gly159Asn
Ser158Glu + Gly159Glu + Ser160Asp + Gly165Asn
Ser158Asp + Gly159Asp + Ser160Asp + Asn162Gln
Asn157Ser + Ser160Asp + Thr161Glu + Asn162Glu
Gly156Asn + Ser158Asp + Ser160Glu + Thr161Glu
Gly159Glu + Thr161Asp + Thr163Asn + Tyr166Val
Gly156Pro + Gly159Glu + Thr161Asp + Gly165Pro
Gly153Glu + Gly156Pro + Gly159Gln + Gly165Asp
Gly153Glu + Asn157Gln + Gly159Gln + Gly165Asp
Gly153Asn + Ser155Glu + Thr163Glu + Ile164Gly
Asn157Glu + Ser158Asp + Ser160Asp + Thr161Asn
Asn154Ser + Asn157Ser + Ser158Asp + Ser160Glu
Ser158Glu + Gly159Gln + Ser160Glu + Thr163Gln
Ser158Asp + Ser160Glu + Ile164Ala + Tyr166Gln
Gly155Pro + Asn157Gln + Ser158Glu + Ser160Asp
Gly153Gln + Gly156Pro + Ser158Glu + Ser160Asp
Gly156Ser + Asn157Glu + Gly159Glu + Ile164Leu
Thr163Asp + Ile164Cys + Gly165Asp + Tyr166Cys
Ser160Asp + Asn162Asp + Ile164Asn + Tyr166Thr
Asn157Ser + Ser160Glu + Asn162Asp + Ile164Met
Ser155Glu + Asn157Asp + Asn162Ser + Gly165Pro
Gly153Ser + Asn154Glu + Ser155Glu + Asn157Glu
Gly153Pro + Asn154Asp + Ser155Asp + Asn157Glu
Gly153Gln + Ser158Glu + Ser160Asp + Asn162Glu
Asn157Asp + Ser160Glu + Thr161Ser + Asn162Glu
Gly156Asp + Asn157Asp + Thr161Ser + Asn162Asp
Asn154Gln + Asn157Asp + Ser160Glu + Tyr166Ser
Gly153Gln + Gly156Glu + Ser158Glu + Asn162Glu
Gly156Glu + Gly159Pro + Thr163Asp + Gly165Asp
Ser155Glu + Thr161Ser + Thr163Ser + Gly165Asp
Asn154Asp + Thr163Gly + Gly165Glu + Tyr166Ser
Ser155Asp + Asn157Glu + Gly159Glu + Gly165Gln
Asn162Glu + Ile164Ala + Gly165Asp + Tyr166Thr
Gly156Gln + Asn162Asp + Gly165Asp + Tyr166Asn
Ser155Glu + Ser158Asp + Thr161Asn + Tyr166Met
Ser155Glu + Ser158Glu + Gly159Gln + Ile164Gly
表18
环5-单突变变异体
Ala186Asn
Ala186Asp
Ala186Gln
Ala186Glu
Ala186Gly
Ala186His
Ala186Pro
Ala186Ser
Ala186Thr
Ser187Asp
Ser187Glu
Phe188Ala
Phe188Asn
Phe188Asp
Phe188Cys
Phe188Gln
Phe188Glu
Phe188Gly
Phe188His
Phe188Ile
Phe188Leu
Phe188Met
Phe188Pro
Phe188Ser
Phe188Thr
Phe188Tyr
Phe188Val
Ser189Asp
Ser189Glu
Ser190Asp
Ser190Glu
表19
环5-双突变变异体
Ala186Asp + Phe188Gln
Ala186Gly + Ser187Glu
Ala186Gln + Ser187Asp
Ala186His + Ser190Glu
Ser187Asp + Phe188Cys
Ser187Asp + Phe188Thr
Phe188Gly + Ser190Glu
Ala186Asn + Phe188Ser
Ala186Asp + Phe188Ile
Ala186Ser + Phe188Glu
Ala186Thr + Phe188Leu
Ser187Glu + Phe188Ser
Ser187Glu + Phe188Pro
Ser187Asp + Phe188Gln
Ala186Thr + Ser190Glu
Phe188His + Ser190Asp
Ala186Gln + Ser190Asp
Phe188Leu + Ser190Glu
Ala186His + Ser190Asp
Ala186Pro + Ser187Glu
Phe188Ile + Ser190Asp
Ala186Asp + Phe188Asn
Ala186Ser + Ser187Glu
Ser187Glu + Phe188Leu
Ala186Asn + Phe188Ile
Phe188Tyr + Ser190Glu
Ala186Pro + Ser190Glu
Ala186Thr + Phe188Asp
Ala186His + Ser187Asp
Ala186Gln + Phe188Thr
Phe188Cys + Ser190Glu
Ser187Glu + Phe188Thr
Ser187Asp + Phe188Met
Ala186Asn + Ser190Asp
Phe188Val + Ser190Asp
Ala186Pro + Phe188Ser
Ala186Asp + Phe188Ser
Ser187Glu + Phe188Asn
Ala186Thr + Ser187Asp
Ala186Thr + Phe188Asn
Ala186Gly + Phe188Leu
Phe188Asn + Ser190Glu
Ala186Thr + Ser187Glu
Ala186Thr + Ser190Asp
Ser187Asp + Phe188Ala
Ala186Asn + Ser187Glu
Ala186Gln + Phe188Tyr
Phe188Ile + Ser190Glu
Phe188Thr + Ser190Asp
Ala186Ser + Ser187Asp
Phe188Ala + Ser190Asp
Phe188Gln + Ser190Glu
Ala186Gly + Ser190Asp
Ala186Glu + Phe188Thr
Ala186Pro + Phe188Ala
Phe188Asn + Ser190Asp
Ala186Gly + Phe188Val
Phe188Leu + Ser190Asp
Ala186Asn + Phe188Met
Ala186Gln + Phe188Glu
表20
环5-三突变变异体
Ala186Pro + Phe188Cys + Ser190Glu
Ala186Gln + Phe188Val + Ser190Glu
Ala186Thr + Phe188Asn + Ser190Glu
Ala186Gly + Phe188Gln + Ser190Glu
Ala186Pro + Phe188Gln + Ser190Glu
Ala186Gln + Phe188Asn + Ser190Asp
Ala186Asn + Ser187Asp + Phe188Leu
Ala186Gln + Ser187Asp + Phe188Pro
Ala186Gln + Ser187Glu + Phe188Met
Ala186Pro + Phe188Val + Ser190Glu
Ala186His + Phe188Met + Ser190Asp
Ala186Asn + Ser187Glu + Phe188Ile
Ala186Gln + Phe188Ala + Ser190Glu
Ala186Pro + Phe188Thr + Ser190Asp
Ala186Asn + Ser187Glu + Phe188Thr
Ala186Ser + Phe188Ala + Ser190Glu
Ala186His + Phe188Pro + Ser190Asp
Ala186His + Ser187Asp + Phe188Leu
Ala186Pro + Ser187Glu + Phe188Val
Ala186Asn + Ser187Asp + Phe188His
Ala186Gly + Ser187Glu + Phe188Leu
Ala186Pro + Ser187Asp + Phe188His
Ala186Gln + Phe188Ile + Ser190Asp
Ala186Pro + Phe188Ile + Ser190Asp
Ala186Gln + Phe188Thr + Ser190Asp
Ala186Gln + Phe188His + Ser190Glu
Ala186Gly + Phe188Ile + Ser190Asp
Ala186Thr + Ser187Glu + Phe188Tyr
Ala186His + Ser187Asp + Phe188Ser
Ala186His + Ser187Glu + Phe188Ser
Ala186His + Phe188Gly + Ser190Glu
Ala186Pro + Ser187Glu + Phe188Pro
Ala186Gln + Ser187Glu + Phe188Tyr
Ala186Asn + Phe188Cys + Ser190Asp
Ala186Gly + Phe188Gly + Ser190Asp
Ala186Gln + Ser187Asp + Phe188Val
Ala186Pro + Ser187Glu + Phe188Gly
Ala186Glu + Ser187Glu + Phe188Ile
Ala186Asp + Ser187Glu + Phe188Ala
Ala186Asp + Ser187Asp + Phe188Thr
Ala186Glu + Ser187Asp + Phe188Thr
Ala186Glu + Ser187Glu + Phe188Tyr
Ala186Asp + Ser187Asp + Phe188Ser
Ala186Asp + Ser187Glu + Phe188Ile
Ala186Pro + Ser187Asp + Phe188Asp
Ala186Gln + Ser187Glu + Phe188Glu
Ala186Gln + Ser187Glu + Phe188Asp
Ala186Glu + Ser187Glu + Phe188Asp
Ala186Glu + Ser187Asp + Phe188Glu
Ala186Thr + Phe188Asp + Ser190Glu
Ala186Ser + Phe188Asp + Ser190Asp
Ala186Gly + Phe188Asp + Ser190Glu
Ser187Asp + Phe188Glu + Ser190Asp
Ser187Glu + Phe188Glu + Ser190Asp
Ser187Asp + Phe188Glu + Ser190Glu
Ser187Glu + Phe188Glu + Ser190Glu
Ala186Glu + Phe188Glu + Ser190Glu
Ala186Glu + Phe188Asp + Ser190Asp
Ala186Asp + Ser187Asp + Ser190Glu
Ala186Asp + Ser187Glu + Ser190Asp
表21
环5-四突变变异体
Ala186Thr + Ser187Glu + Phe188Glu + Ser190Asp
Ala186Gln + Ser187Asp + Phe188Asp + Ser190Asp
Ala186Gly + Ser187Asp + Phe188Glu + Ser190Asp
Ala186Thr + Ser187Asp + Phe188Glu + Ser190Glu
Ala186His + Ser187Glu + Phe188Glu + Ser190Asp
Ala186Gln + Ser187Glu + Phe188Glu + Ser190Glu
Ala186Thr + Ser187Glu + Phe188Asp + Ser190Asp
Ala186Asn + Ser187Glu + Phe188Asp + Ser190Asp
Ala186Ser + Ser187Glu + Phe188Glu + Ser190Glu
Ala186Asn + Ser187Asp + Phe188Asp + Ser190Asp
Ala186Ser + Ser187Asp + Phe188Asp + Ser190Glu
Ala186Asn + Ser187Asp + Phe188Asp + Ser190Glu
Ala186His + Ser187Glu + Phe188Glu + Ser190Glu
Ala186Gln + Ser187Asp + Phe188Glu + Ser190Asp
Ala186Gly + Ser187Asp + Phe188Glu + Ser190Glu
Ala186Pro + Ser187Asp + Phe188Asp + Ser190Glu
Ala186Gly + Ser187Asp + Phe188Asp + Ser190Glu
Ala186Ser + Ser187Glu + Phe188Glu + Ser190Asp
Ala186Thr + Ser187Asp + Phe188Asp + Ser190Asp
Ala186Gly + Ser187Glu + Phe188Asp + Ser190Glu
Ala186Gly + Ser187Asp + Phe188Asp + Ser190Asp
Ala186Asp + Ser187Asp + Phe188Asn + Ser190Asp
Ala186Glu + Ser187Glu + Phe188Tyr + Ser190Glu
Ala186Asp + Ser187Asp + Phe188Ser + Ser190Glu
Ala186Asp + Ser187Glu + Phe188Gln + Ser190Glu
Ala186Glu + Ser187Asp + Phe188Ala + Ser190Glu
Ala186Glu + Ser187Glu + Phe188Gly + Ser190Glu
Ala186Asp + Ser187Asp + Phe188Ala + Ser190Asp
Ala186Glu + Ser187Glu + Phe188Cys + Ser190Glu
Ala186Glu + Ser187Asp + Phe188Ile + Ser190Asp
Ala186Glu + Ser187Glu + Phe188Ala + Ser190Glu
Ala186Glu + Ser187Glu + Phe188His + Ser190Asp
Ala186Asp + Ser187Asp + Phe188Leu + Ser190Glu
Ala186Asp + Ser187Asp + Phe188Gln + Ser190Glu
Ala186Asp + Ser187Glu + Phe188Asn + Ser190Asp
Ala186Asp + Ser187Asp + Phe188Cys + Ser190Glu
Ala186Asp + Ser187Asp + Phe188Thr + Ser190Asp
Ala186Glu + Ser187Glu + Phe188Leu + Ser190Asp
Ala186Glu + Ser187Glu + Phe188Leu + Ser190Glu
Ala186Asp + Ser187Asp + Phe188Pro + Ser190Glu
Ala186Asp + Ser187Glu + Phe188Val + Ser190Asp
Ala185Asp + Ser187Asp + Phe188His + Ser190Glu
Ala186Glu + Ser187Glu + Phe188Pro + Ser190Glu
Ala186Asp + Ser187Glu + Phe188Ser + Ser190Glu
Ala186Glu + Ser187Glu + Phe188Val + Ser190Glu
Ala186Asp + Ser187Glu + Phe188Gln + Ser190Asp
Ala186Asp + Ser187Glu + Phe188Asn + Ser190Glu
Ala186Asp + Ser187Glu + Phe188Gly + Ser190Glu
Ala186Asp + Ser187Asp + Phe188Cys + Ser190Asp
Ala186Asp + Ser187Glu + Phe188Tyr + Ser190Glu
Ala186Glu + Ser187Asp + Phe188Thr + Ser190Asp
Ala186Glu + Ser187Glu + Phe188Thr + Ser190Glu
Ala186Glu + Ser187Glu + Phe188Met + Ser190Glu
Ala186Asp + Ser187Asp + Phe188Gly + Ser190Glu
Ala186Asp + Ser187Glu + Phe188His + Ser190Glu
Ala186Asp + Ser187Glu + Phe188Met + Ser190Glu
Ala186Glu + Ser187Asp + Phe188Ser + Ser190Asp
Ala186Asn + Ser187Glu + Phe188Met + Ser190Asp
Ala186Gln + Ser187Asp + Phe188His + Ser190Asp
Ala186Asn + Ser187Glu + Phe188Ser + Ser190Glu
表22
环6-单突变变异体
Ala199Asn
Ala199Asp
Ala199Gln
Ala199Glu
Ala199Gly
Ala199His
Ala199Pro
Ala199Ser
Ala199Thr
Pro200Asn
Pro200Asp
Pro200Gln
Pro200Glu
Pro200Gly
Pro200Ser
Gly201Asn
Gly201Asp
Gly201Gln
Gly201Glu
Gly201Pro
Gly201Ser
Ala202Asn
Ala202Asp
Ala202Gln
Ala202Gln
Ala202Gly
Ala202His
Ala202Pro
Ala202Ser
Ala202Thr
Gly203Asn
Gly203Asp
Gly203Gln
Gly203Glu
Gly203Pro
Gly203Ser
Val204Ala
Val204Asn
Val204Asp
Val204Cys
Val204Gln
Val204Glu
Val204Gly
Val204His
Val204Met
Val204Pro
Val204Ser
Val204Thr
Tyr205Ala
Tyr205Asn
Tyr205Asp
Tyr205Cys
Tyr205Gln
Tyr205Glu
Tyr205Gly
Tyr205His
Tyr205Ile
Tyr205Leu
Tyr205Met
Tyr205Pro
Tyr205Ser
Tyr205Thr
Tyr205Val
Ser206Asp
Ser206Glu
Thr207Asn
Thr207Asp
Thr207Gln
Thr207Glu
Thr207Gly
Thr207Pro
Thr207Ser
Tyr208Ala
Tyr208Asn
Tyr208Asp
Tyr208Cys
Tyr208Gln
Tyr208Glu
Tyr208Gly
Tyr208His
Tyr208Ile
Tyr208Leu
Tyr208Met
Tyr208Pro
Tyr208Ser
Tyr208Thr
Tyr208Val
Pro209Asn
Pro209Asp
Pro209Gln
Pro209Glu
Pro209Gly
Pro209Ser
Thr210Asn
Thr210Asp
Thr210Gln
Thr210Glu
Thr210Gly
Thr210Pro
Thr210Ser
Asn211Asp
Asn211Gln
Asn211Glu
Asn211Ser
Thr212Asn
Thr212Asp
Thr212Gln
lhr212Glu
Thr212Gly
Thr212Pro
Thr212Ser
Tyr213Ala
Tyr213Asn
Tyr213Asp
Tyr213Cys
Tyr213Gln
Tyr213Glu
Tyr213Gly
Tyr213His
Tyr213Ile
Tyr213Leu
Tyr213Met
Tyr213Pro
Tyr213Ser
Tyr213Thr
Tyr213Val
Ala214Asn
Ala214Asp
Ala214Gln
Ala214Glu
Ala214Gly
Ala214His
Ala214Pro
Ala214Ser
Ala214Thr
Thr215Asn
Thr215Asp
Thr215Gln
Thr215Glu
Thr215Gly
Thr215Pro
Thr215Ser
Leu216Ala
Leu216Asn
Leu216Asp
Leu216Cys
Leu216Gln
Leu216Glu
Leu216Gly
Leu216His
Leu216Ile
Leu216Met
Leu216Pro
Leu216Ser
Leu216Thr
Leu216Val
Asn217Asp
Asn217Gln
Asn217Glu
Asn217Ser
Gly218Asn
Gly218Asp
Gly218Gln
Gly218Glu
Gly218Pro
Gly218Ser
Thr219Asn
Thr219Asp
Thr219Gln
Thr219Glu
Thr219Gly
Thr219Pro
Thr219Ser
表23
环6-双突变变异体
Gly218Gln + Thr219Gln
Val204Ala + Thr207Gln
Tyr213Val + Gly218Pro
Pro209Gly + Tyr213Ile
Thr215Ser + Thr219Ser
Leu216Ala + Asn217Gln
Pro209Asp + Thr212Ser
Ala199Asn + Leu216Ile
Asn217Glu + Thr219Asn
Tyr208Ile + Thr215Asn
Ala202Gly + Pro209Glu
Gly201Ser + Thr219Gly
Thr215Pro + Gly218Ser
Tyr205Leu + Leu216His
Val204Asn + Ala214Asn
Asn217Ser + Gly218Pro
Ala202His + Tyr208Val
Gly201Gln + Tyr213Leu
Tyr213Cys + Thr219Gly
Pro200Asn + Thr210Pro
Ala202Asp + Thr219Asn
Gly201Pro + Val204Pro
Ala214Thr + Thr219Gln
Thr207Gln + Leu216Asp
Pro200Gln + Tyr213Met
Val204Ala + Thr215Glu
Thr212Gln + Thr215Asn
Leu216His + Asn217Glu
Ala202Thr + Ala214Asp
Val204Pro + Thr207Gln
Gly203Asp + Tyr205Leu
Tyr205His + Asn211Asp
Asn211Gln + Thr212Asn
Gly203Pro + Thr219Pro
Pro200Gly + Pro209Asp
Thr210Asp + Gly218Pro
Gly203Asp + Gly218Ser
Tyr205Glu + Gly218Pro
Thr212Gln + Thr219Pro
Ala214Ser + Asn217Glu
Ala202Asn + Leu216Ser
Val204Thr + Ala214Pro
Pro200Asn + Asr211Asp
Tyr205Gly + Thr219Pro
Ala199His + Asn217Ser
Gly201Ser + Asn211Asp
Gly203Ser + Tyr205Ala
Ala202Pro + Pro209Glu
Tyr213Pro + Gly218Glu
Pro200Gly + Ala202Glu
Thr210Gly + Asn211Asp
Val204Asn + Thr215Asn
Pro200Asn + Tyr208Leu
Ala202Pro + Ala214Thr
Tyr205Cys + Leu216His
Leu216Ser + Gly218Gln
Thr215Glu + Thr219Gln
Tyr205Leu + Pro209Glu
Ala214Glu + Thr215Pro
Gly203Glu + Gly218Gln
Tyr205Cys + Thr212Pro
Leu216Asn + Thr219Asn
Ala199Ser + Thr219Asn
Gly201Gln + Tyr213Val
Gly203Asn + Thr215Gln
Pro209Glu + Asn211Gln
Ala199Gly + Asn211Asp
Ala214Glu + Leu216Met
Val204Pro + Asn211Asp
Ala202Glu + Gly218Gln
Tyr208Ile + Gly218Glu
Tyr205Ser + Leu216Thr
Asn211Glu + Tyr213Gly
Thr212Asn + Thr219Asn
Asn217Asp + Thr219Gln
Thr207Asn + Asn217Asp
Gly201Ser + Tyr208Gln
Thr210Pro + Thr212Gly
Val204Asn + Thr210Gly
Ala199His + Tyr208Val
Tyr208Gly + Tyr213Val
Gly201Pro + Ala214Asp
Gly201Gln + Leu216Gly
Gly201Gln + Thr207Asn
Gly203Asn + Gly218Glu
Tyr208Asn + Thr219Ser
Thr207Gly + Leu216Glu
Gly218Glu + Thr219Asn
Ala199Gln + Thr212Glu
Thr212Asp + Ala214Gly
Gly203Asn + Ala214Ser
Asn211Ser + Thr212Gln
Pro209Asp + Tyr213Thr
Gly201Gln + Ala202Thr
Val204Thr + Tyr208Cys
Gly201Asn + Ala202Asn
Ala199His + Thr210Asp
Val204Ser + Thr212Asp
Tyr208Ser + Thr219Gln
Tyr205Pro + Thr215Asp
Asn211Gln + Gly218Gln
Asn217Asp + Gly218Pro
Ala202Gln + Thr210Pro
Tyr205Asn + Gly218Gln
Gly201Gln + Thr219Ser
Ala202Asp + Thr207Asn
Thr212Gly + Asn217Gln
Thr215Asn + Thr219Ser
Ala214Glu + Thr215Gly
Thr210Asn + Tyr213Gln
Ala199Gln + Tyr205Glu
Tyr205His + Thr210Asp
Gly201Asn + Gly203Asp
Gly203Ser + Pro209Asp
Gly201Gln + Thr210Glu
Pro209Asn + Thr215Ser
Thr215Gln + Thr219Gly
Gly201Gln + Thr215Asp
Tyr208Gly + Tyr213Pro
Gly201Asn + Tyr213Asn
Ala202Pro + Pro209Asn
Ala199Thr + Tyr208Met
Thr210Gln + Gly218Asn
Thr210Pro + Asn211Asp
Asn211Asp + Gly218Pro
Gly201Asn + Tyr205Gly
Ala202Gln + Tyr213Glu
Ala199His + Ala202Asp
Tyr205Ser + Tyr213Glu
Gly201Gln + Pro209Asp
Ala202Ser + Thr219Gln
Leu216Met + Thr219Asn
Val204Ser + Tyr205Ser
Val204Thr + Tyr208Ser
Gly203Asn + Ala214Thr
Thr210Gly + Thr219Asn
Pro200Gly + Thr210Pro
Ala214Glu + Gly218Pro
Pro200Gly + Thr219Gln
Ala202Ser + Asn211Asp
Ala199Thr + Val204Gln
Ala199Gln + Pro200Ser
Pro209Ser + Leu216Thr
Tyr205Pro + Leu216Ala
Gly201Pro + Pro209Gly
Thr207Asn + Thr215Glu
Pro200Ser + Tyr205Ala
Thr210Gln + Asn217Gln
Ala199Asn + Gly203Gln
Gly201Asn + Thr207Pro
Gly203Asp + Tyr208Asn
Thr207Asn + Asn211Asp
Tyr205Leu + Gly218Asp
Gly203Ser + Thr210Pro
Tyr205Cys + Asn211Ser
Ala214Glu + Asn217Ser
Ala199Pro + Gly201Gln
Gly201Asn + Tyr213Met
Tyr205His + Tyr208Gly
Gly203Glu + Thr210Ser
Thr207Asn + Ala214Ser
Tyr205Val + Thr219Ser
Val204Asn + Thr207Pro
Thr207Ser + Asn211Asp
Tyr205Leu + Gly218Glu
Asn211Ser + Thr219Pro
Tyr205Gln + Asn211Ser
Pro200Gly + Val204Asn
Ala202Glu + Tyr213His
Gly201Pro + Gly218Gln
Ala214Asn + Asn217Gln
Gly201Asn + Asn211Asp
Pro209Ser + Thr212Gln
Gly203Gln + Val204Gln
Gly201Gln + Thr215Pro
Gly201Asn + Ala202Ser
Thr207Gln + Thr212Asp
Val204Gly + Thr212Asp
Gly201Asn + Thr210Asp
Thr210Asp + Tyr213Cys
表24
环6-三突变变异体
Pro200Gly + Thr212Ser + Tyr213Met
Gly201Pro + Pro209Glu + Thr212Gly
Gly203Asp + Val204Met + Asn217Gln
Pro200Gly + Thr212Ser + Ala214Asp
Ala199Gly + Tyr213Asp + Gly218Ser
Gly201Asn + Thr207Ser + Thr215Glu
Pro200Ser + Ala214Gly + Asn217Glu
Ala199Pro + Thr207Ser + Thr210Pro
Gly201Asn + Thr210Ser + Asn211Glu
Gly203Asn + Thr207Asn + Asn217Glu
Gly201Asn + Tyr205Pro + Thr207Ser
Pro200Asn + Gly201Gln + Gly203Ser
Pro200Gln + Asn211Glu + Tyr213Ala
Ala202His + Thr212Asp + Tyr213His
Ala199Thr + Gly201Asn + Asn211Ser
Pro200Ser + Gly201Gln + Pro209Asn
Ala202Gly + Tyr205Pro + Asn211Ser
Gly203Gln + Tyr205Val + Thr210Pro
Gly203Glu + Asn211Gln + Asn217Ser
Pro200Asn + Pro209Glu + Asn211Ser
Gly201Pro + Thr210Ser + Tyr213Thr
Tyr205Ile + Leu216Asp + Thr219Gly
Pro200Ser + Val204Met + Pro209Asp
Tyr205Asp + Thr207Gly + Tyr208Gln
Pro200Ser + Thr210Asp + Thr212Pro
Gly201Gln + Asn211Asp + Tyr213Gly
Tyr208Val + Thr210Gly + Leu216Val
Pro209Gly + Thr215Pro + Leu216Ala
Val204His + Thr215Glu + Leu216Cys
Ala199Pro + Gly203Ser + Thr210Ser
Gly201Asn + Tyr205Met + Thr210Asp
Pro200Gly + TyT205Cys + Thr212Asn
Ala199Pro + Pro200Ser + Leu216Glu
Gly201Asn + Tyr205Asp + Tyr213lle
Pro200Ser + Ala202Gly + Thr210Asp
Ala202Asn + Val204Ser + Gly218Glu
Gly203Gln + Val204Cys + Thr207Ser
Val204Cys + Thr207Asn + Thr212Asp
Gly201Pro + Gly203Ser + Tyr208Gly
Gly203Asn + Tyr208Cys + Leu216Met
Thr210Gln + Thr215Asp + Thr219Ser
Pro209Asp + Tyr213Leu + Ala214Thr
Ala202Asp + Tyr213Pro + Thr215Gln
Thr215Glu + Gly218Ser + Thr219Gln
Pro200Gly + Val204His + Thr207Ser
Gly203Ser + Pro209Asp + Thr219Ser
Ala202Asn + Thr207Gln + Ala214His
Thr207Ser + Ala214Glu + Leu216Ser
Pro200Asn + Pro209Asn + Thr210Pro
Pro209Gln + Tyr213His + Ala214Thr
Thr210Asn + Thr215Pro + Leu216Gln
Gly203Gln + Val204Ser + Thr210Glu
Pro200Asn + Asn211Gln + Gly218Ser
Pro200Gln + Thr207Gln + Leu216Glu
Tyr213Pro + Gly218Glu + Thr219Asn
Tyr208Cys + Thr210Asp + Thr219Ser
Pro200Gln + Tyr205His + Thr219Ser
Pro209Asp + Thr212Gln + Thr219Gln
Gly201Asn + Val204Asn + Thr210Asp
Pro200Gly + Tyr208Leu + Thr210Asn
Pro200Gln + Thr207Gln + Thr219Gln
Tyr205Val + Pro209Ser + Thr210Asp
Gly203Pro + Val204His + Tyr213Glu
Gly203Asn + Pro209Asn + Tyr213His
Ala199Asn + Gly201Asn + Leu216Asp
Pro209Asp + Tyr213His + Gly218Ser
Gly201Ser + Gly203Asp + Asn211Ser
Gly203Asn + Val204Cys + Thr212Gly
Gly201Pro + Tyr208Met + Leu216Glu
Gly201Ser + Val204Gln + Leu216His
Ala199Gly + Ala202Pro + Leu216Ser
Gly203Gln + Tyr205Pro + Thr219Gly
Tyr205Gly + Tyr213Val + Asn217Asp
Gly201Asn + Ala214His + Thr215Gln
Pro200Gly + Ala202Gln + Tyr213Cys
Gly203Pro + Tyr208Met + Thr219Pro
Pro200Asn + Tyr205Leu + Gly218Glu
Pro209Gly + Thr212Glu + Tyr213Thr
Pro200Gln + Thr210Pro + Thr219Asn
Gly201Ser + Gly218Pro + Thr219Asn
Pro200Ser + Gly201Pro + Thr219Gly
Thr207Pro + Tyr213Ala + Leu216Glu
Ala199Ser + Thr207Asn + Thr212Asp
Thr207Gln + Tyr208Ile + Thr210Asp
Tyr213Ser + Thr215Gln + Thr219Pro
Pro200Gly + Tyr208Ile + Ala214Ser
Gly201Asn + Pro209Ser + Thr215Asp
Pro200Ser + Pro209Asp + Tyr213Met
Tyr208Gly + Thr210Glu + Thr215Ser
Ala202Ser + Ala214Asp + Thr219Ser
Val204Pro + Pro209Asn + Thr219Asn
Thr207Asn + Pro209Asn + Gly218Ser
Pro200Gly + Leu216Cys + Gly218Ser
Pro209Gly + Leu216Val + Gly218Glu
Asn211Gln + Tyr213Thr + Thr215Ser
Tyr213Val + Thr215Glu + Gly218Ser
Tyr213Pro + Gly218Gln + Thr219Ser
Gly201Pro + Ala202Thr + Val204Cys
Ala202Ser + Tyr208Leu + Pro209Asp
Tyr208Thr + Thr210Asn + Ala214Thr
Tyr205Pro + Pro209Ser + Thr215Gln
Tyr205Cys + Tyr208Gly + Leu216Glu
Ala199Gly + Gly203Asp + Pro209Asn
Val204Gln + Thr207Gly + Gly218Gln
Ala202His + Gly218Asp + Thr219Gln
Gly203Asp + Ash211Ser + Leu216Met
Pro200Ser + Val204Cys + Thr215Glu
Thr207Asn + Leu216Pro + Asn217Glu
Ala199His + Tyr205His + Thr212Pro
Ala199Gln + Gly203Asp + Thr219Gly
Ala199His + Pro200Gln + Asn217Asp
Val204Ser + Tyr213Leu + Thr219Gln
Val204Ser + Pro209Ser + Leu216Glu
Ala202Gly + Pro209Asn + Thr219Asn
Thr215Gln + Gly218Asn + Thr219Gln
Ala199Pro + Ala214Ser + Leu216Val
Gly203Asn + Thr215Asn + Gly218Glu
Thr207Pro + Tyr208Ala + Thr219Asn
Ala199Ser + Thr210Ser + Gly218Gln
Pro209Asn + Thr210Ser + Asn211Ser
Gly201Asn + Val204Cys + Tyr213Val
Gly201Ser + Val204Ser + Pro209Asn
Gly201Pro + Thr210Ser + Asn211Ser
Tyr205Gln + Tyr208His + Thr210Asp
Tyr208Ile + Thr215Ser + Gly218Glu
Thr212Glu + Ala214His + Thr215Gln
Gly201Asn + Val204Gly + Tyr205Gln
Ala199Thr + Pro209Glu + Thr210Ser
Pro209Gly + Thr210Asn + Gly218Glu
Gly203Pro + Pro209Asn + Thr210Asn
Gly203Glu + Tyr213Pro + Leu216Thr
Ala199Pro + Thr212Glu + Ala214Thr
Thr207Gln + Pro209Asn + Thr219Ser
Ala199Thr + Tyr205Gln + Thr219Pro
Val204Met + Ala214Gly + Asn217Glu
Gly201Gln + Val204Pro + Thr210Glu
Tyr208Leu + Thr212Asp + Gly218Asn
Gly201Gln + Tyr205His + Thr210Glu
Gly201Pro + Ala214Gly + Thr219Ser
Gly203Asp + Tyr205Ala + Tyr208Ser
Thr212Pro + Asn217Ser + Gly218Asp
Tyr205Asp + Thr207Gly + Tyr208Leu
Gly201Pro + Ala202Glu + Leu216Ser
Ala202His + Tyr208Pro + Thr215Asp
Tyr208Thr + Thr212Glu + Thr219Ser
Val204Met + Pro209Gln + Leu216Asp
Thr207Pro + Thr210Asn + Tyr213Glu
Gly201Ser + Val204Gln + Asn211Asp
Gly203Gln + Tyr205Asn + Tyr213Ser
Ala199Thr + Thr210Asn + Tyr213Glu
Tyr208Ile + Pro209Ser + Thr212Pro
Tyr205Leu + Tyr208Thr + Ala214Glu
Pro200Gln + Val204Gln + Ala214Glu
Gly203Pro + Asn211Asp + Asn217Gln
Gly201Asn + Gly218Gln + Thr219Pro
Ala199Gln + Gly203Gln + Ala214Asp
Ala202Thr + Tyr205Cys + Tyr213Ile
Gly201Gln + Val204Cys + Asn211Glu
Tyr208Thr + Pro209Gly + Thr215Asp
Thr210Glu + Asn211Gln + Tyr213Leu
Val204Cys + Tyr208Pro + Thr219Pro
Ala202Asp + Gly203Asn + Gly218Asn
Ala199Asn + Val204Gly + Tyr205Pro
Gly203Pro + Val204Met + Tyr208His
Pro200Gly + Tyr213Pro + Leu216Cys
Val204Ala + Ala214His + Asn217Glu
Ala202Gln + Thr207Asn + Ala214Gly
Gly203Ser + Val204Gly + Tyr213Thr
Asn211Asp + Leu216Cys + Thr219Asn
Pro200Asn + Thr212Gly + Gly218Asn
Tyr205His + Tyr208Leu + Thr215Gln
Gly203Ser + Tyr205His + Thr219Ser
Val204Asn + Thr212Pro + Thr215Asp
Thr212Ser + Thr215Ser + Leu216Ala
Tyr208Leu + Thr212Glu + Leu216Met
Thr207Asn + Tyr208Leu + Leu216Asp
Thr210Gly + Thr212Pro + Asn217Asp
Asn211Ser + Ala214Asn + Leu216Glu
Thr207Ser + Tyr208Thr + Ala214His
Ala202Asp + Val204Ser + Tyr208Thr
表25
环6-四突变变异体
Pro200Ser + Thr207Ser + Tyr213Cys + Gly218Gln
Ala202Gln + Tyr208Cys + Thr210Pro + Tyr213Asn
Gly203Asp + Tyr205Ile + Thr215Pro + Thr219Ser
Gly201Gln + Gly203Gln + Val204Asn + Thr207Asn
Pro200Ser + Gly201Ser + Ala202Ser + Pro209Glu
Gly201Ser + Thr207Pro + Thr215Asp + Thr219Pro
Ala202Gln + Asn211Gln + Tyr213Ser + Thr219Asn
Gly201Pro + Tyr213Ala + Ala214Pro + Thr215Ser
Ala199Gly + Pro200Gly + Ala202Glu + Gly218Pro
Tyr205Thr + Tyr208Met + Thr210Ser + Asn217Gln
Gly201Ser + Ala214Ser + Leu216Gly + Gly218Asn
Pro200Gln + Gly203Gln + Asn211Glu + Thr215Pro
Pro200Ser + Tyr208Leu + Asn211Ser + Gly218Asp
Ala199Asn + Pro209Asn + Thr210Ser + Tyr213Met
Ala199His + Gly203Ser + Pro209Asp + Thr212Gln
Tyr205Gly + Thr212Gly + Thr215Asn + Asn217Gln
Ala199Gln + Tyr208Thr + Thr210Asn + Tyr213Gln
Gly203Ser + Pro209Asp + Asn211Ser + Ala214Thr
Pro209Asn + Thr215Ser + Gly218Asp + Thr219Gly
Ala199Pro + Thr207Gln + Thr210Glu + Thr215Pro
Ala202Asn + Val204Gly + Thr207Gly + Thr212Asp
Val204Thr + Tyr208Leu + Asn211Ser + Asn217Asp
Pro200Gly + Tyr205Val + Leu216Asp + Thr219Gly
Pro200Gly + Tyr208Ser + Ala214Ser + Thr219Ser
Ala199Thr + Gly203Glu + Thr207Asn + Tyr213Cys
Pro200Gly + Tyr213Thr + Asn217Gln + Thr219Pro
Gly203Asn + Val204Thr + Thr207Asn + Thr210Ser
Ala202Gly + Thr207Gln + Pro209Gln + Thr210Asp
Ala199His + Gly201Gln + Tyr205Asp + Thr219Gln
Thr207Ser + Asn211Asp + Tyr213Thr + Leu216Ser
Ala199Ser + Thr210Glu + Asn211Gln + Leu216Ile
Gly201Ser + Gly203Ser + Thr212Pro + Ala214Glu
Pro200Gly + Gly201Gln + Gly203Gln + Leu216His
Ala199Pro + Gly201Gln + Gly203Pro + Thr212Glu
Ala199His + Gly201Asn + Val204Thr + Tyr208Ser
Val204Thr + Tyr208Ile + Ala214Glu + Thr219Pro
Ala199Ser + Tyr205Pro + Thr212Ser + Thr219Gln
Val204Met + Leu216Cys + Asn217Ser + Thr219Gly
Gly201Ser + Tyr205His + Pro209Gln + Thr219Ser
Pro200Gln + Pro209Asn + Asn211Asp + Thr219Gly
Val204Met + Thr210Gly + Thr212Asp + Ala214Ser
Pro200Gly + Thr207Asn + Asn211Glu + Leu216His
Tyr208Ala + Thr210Asp + Thr215Gln + Thr219Gly
Gly203Asn + Thr207Gln + Pro209Asp + Thr215Gly
Ala199Ser + Pro209Glu + Leu216His + Thr219Pro
Gly203Asp + Tyr205His + Tyr213Val + Ala214Thr
Pro200Asn + Thr212Asn + Leu216His + Gly218Pro
Ala199Ser + Pro209Gln + Thr215Gln + Asn217Glu
Gly203Asp + Thr207Pro + Thr212Gly + Ala214Gln
Gly201Pro + Gly203Gln + Thr207Pro + Ala214Asp
Val204Gln + Ala214Asp + Leu216Thr + Gly218Ser
Thr207Asn + Tyr208Pro + Pro209Asp + Tyr213Asn
Pro209Asp + Asn211Gln + Thr215Gln + Gly218Pro
Val204Gln + Asn211Ser + Asn217Gln + Gly218Pro
Gly201Gln + Ala202His + Asn217Asp + Gly218Gln
Val204Thr + Tyr208Leu + Ala214Pro + Gly218Asp
Gly201Ser + Thr207Pro + Thr212Ser + Thr215Pro
Ala202Gly + Val204Gly + Asn211Glu + Leu216Gly
Ala199Ser + Gly201Gln + Ala214Ser + Asn217Ser
Ala202Gln + Thr207Pro + Pro209Glu + Tyr213Pro
Ala199Thr + Ala202Ser + Val204Ala + Thr207Gln
Ala199His + Thr207Ser + Pro209Gln + Ala214Glu
Ala202His + Gly203Asp + Val204Pro + Gly218Pro
Ala202Asn + Gly203Glu + Val204Gln + Thr210Gln
Pro200Asn + Val204Gly + Tyr205Gln + Thr207Ser
Ala202Glu + Gly203Pro + Thr207Gly + Thr219Gly
Gly201Asn + Tyr208Val + Gly218Glu + Thr219Asn
Pro200Ser + Gly201Gln + Tyr205Asp + Thr219Gly
Pro200Ser + Thr207Gly + Ala214Pro + Thr215Asp
Val204Ala + Thr207Gln + Asn211Asp + Thr215Pro
Ala202Thr + Val204Asn + Thr207Ser + Pro209Asn
Ala202Asn + Val204Thr + Thr207Gly + Pro209Ser
Ala199Pro + Thr210Gln + Tyr213Cys + Ala214Thr
Ala199Pro + Pro200Gln + Ala214Glu + Thr215Asn
Pro200Ser + Val204Gln + Thr207Ser + Leu216Thr
Ala199Ser + Thr207Asn + Leu216Gln + Gly218Gln
Gly201Ser + Tyr208Ala + Ala214Ser + Leu216Gln
Val204Thr + Thr207Ser + Thr215Asp + Asn217Ser
Gly201Asn + Thr207Pro + Tyr213Met + Asn217Asp
Val204Ala + Asn211Gln + Thr215Glu + Gly218Ser
Pro200Gly + Pro209Ser + Thr212Gly + Asn217Glu
Ala199Pro + Tyr205Ile + Thr210Gly + Thr219Gly
Pro200Gly + Tyr205Ala + Thr212Pro + Tyr213Glu
Gly203Ser + Thr212Ser + Leu216Ala + Asn217Asp
Gly201Pro + Gly203Gln + Pro209Asp + Thr215Gly
Ala202His + Gly203Ser + Tyr208Cys + Gly218Asn
Ala199Pro + Ala202Gln + Val204Gly + Thr212Gly
Ala199Asn + Pro200Gly + Gly201Gln + Val204Gln
Gly201Asn + Gly203Pro + Pro209Ser + Thr215Asp
Gly201Asn + Thr210Ser + Tyr213Asp + Leu216Gln
Gly203Gln + Val204His + Gly218Gln + Thr219Gly
Gly201Asn + Tyr213Val + Ala214Glu + Thr219Gly
Ala202Glu + Thr207Ser + Thr215Ser + Thr219Ser
Gly203Glu + Thr212Asn + Asn217Gln + Thr219Ser
Gly203Ser + Thr210Asn + Thr212Glu + Leu216Met
Val204Ala + Asn211Asp + Tyr213Met + Leu216Ile
Thr207Pro + Tyr208Asn + Tyr213Pro + Leu216Gln
Gly201Gln + Gly203Glu + Tyr213Met + Ala214Thr
Pro200Gln + Thr212Ser + Tyr213Pro + Thr215Asn
Pro200Asn + Val204Asn + Asn211Glu + Tyr213Gln
Pro209Ser + Asn211Gln + Asn217Asp + Thr219Asn
Pro200Gly + Gly203Asn + Tyr205Val + Tyr208His
Ala202Glu + Gly203Pro + Thr212Ser + Gly218Asn
Ala202Gln + Val204Gly + Pro209Asp + Thr215Asn
Tyr208Ala + Asn211Glu + Thr215Gln + Thr219Ser
TyT208Gln + Tyr213Ala + Leu216Thr + Asn217Asp
Ala199Thr + Pro200Gln + Gly201Asn + Asn211Asp
Ala199Thr + Pro200Ser + Gly201Asn + Tyr208Gly
Val204Ser + Thr207Gly + Thr212Asp + Gly218Gln
Pro200Asn + Ala214His + Thr215Gly + Thr219Asn
Thr207Ser + Tyr208Ser + Leu216Asn + Gly218Asp
Pro200Gly + Gly203Glu + Thr212Gly + Gly218Asn
Pro200Ser + Gly201Ser + Tyr208Gln + Thr219Asn
Pro200Asn + Gly201Pro + Asn211Glu + Asn217Gln
Val204Met + Thr207Asn + Asn217Gln + Thr219Gly
Pro200Gly + Gly201Pro + Thr210Glu + Leu216Thr
Pro200Asn + Gly203Pro + Val204Gln + Thr219Gln
Ala202Asp + Pro209Asn + Tyr213Cys + Leu216Ala
Pro200Gln + Gly201Ser + Ala202Glu + Tyr205Gly
Thr207Pro + Tyr208Asn + Asn211Ser + Tyr213Ser
Ala202Ser + Val204Gly + Thr207Asn + Tyr213Gln
Gly201Ser + Gly203Gln + Thr212Ser + Leu216Asp
Ala199Thr + Tyr205Met + Tyr208Val + Ala214Thr
Gly201Gln + Ala202Thr + Val204Cys + Tyr205Leu
Pro200Gln + Tyr205Cys + Thr210Glu + Thr212Asn
Pro200Gln + Tyr213Pro + Asn217Glu + Gly218Asn
Tyr208Pro + Tyr213His + Ala214Glu + Asn217Ser
Pro200Ser + Ala202Gly + Tyr208Val + Thr212Gln
Pro200Asn + Tyr205Gly + Pro209Gln + Asn211Ser
Tyr205Gly + Asn211Gln + Thr215Gly + Thr219Gly
Gly201Asn + Gly203Asn + Thr212Asp + Ala214Gly
Val204Pro + Pro209Gly + Thr210Glu + Leu216Pro
Tyr205Pro + Tyr213Ala + Gly218Asn + Thr219Pro
Ala199Gln + Val204His + Asn211Asp + Thr215Gly
Pro209Gln + Thr215Gln + Asn217Asp + Thr219Asn
Gly201Asn + Asn211Glu + Tyr213Ile + Asn217Gln
Gly201Ser + Thr207Asn + Tyr213Glu + Ala214Gly
Ala199Ser + Thr210Asn + Thr215Gln + Gly218Pro
Val204Asn + Tyr205Thr + Tyr208Met + Thr215Pro
Val204Gly + Tyr205Val + Thr210Asp + Thr215Gln
Ala202Ser + Val204Ala + Leu216Ser + Asn217Asp
Asn211Asp + Thr212Ser + Ala214Asn + Thr215Gly
Ala199His + Tyr208Leu + Asn217Gln + Thr219Asn
Ala202Pro + Tyr208Cys + Leu216Ala + Thr219Asn
Ala199Gly + Pro200Gly + Gly201Asn + Ala202Asp
Pro200Ser + Ala202Gln + Thr210Gln + Leu216Asp
Gly201Gln + Gly203Glu + Val204Gly + Leu216Asn
Val204Pro + Tyr208Thr + Pro209Ser + Asn211Glu
Tyr208Thr + Thr215Ser + Leu216Glu + Gly218Gln
Gly201Asn + Tyr205Ser + Tyr213Asp + Gly218Pro
Ala202Gln + Gly203Gln + Tyr205Ile + Asn217Glu
Thr212Ser + Tyr213Glu + Thr215Ser + Leu216Cys
Gly203Glu + Thr212Pro + Tyr213Leu + Ala214Ser
Pro200Ser + Gly201Gln + Ala202Asp + Thr210Asn
Val204Gln + Thr210Asp + Tyr213Ser + Ala214Thr
Ala199Pro + Gly201Ser + Val204Gly + Asn217Gln
Pro200Asn + Ala202Ser + Thr207Gln + Tyr208Met
Pro200Gln + Gly203Pro + Ala214Glu + Gly218Pro
Pro200Ser + Thr207Gly + Gly218Asn + Thr219Gly
Gly203Ser + Thr207Gln + Thr210Gln + Asn217Ser
Gly201Ser + Thr210Glu + Asn211Glu + Thr219Asn
Thr207Gly + Thr210Asp + Asn211Asp + Gly218Pro
Pro200Asn + Thr210Glu + Asn211Glu + Thr219Pro
Val204Ser + Thr210Asp + Asn211Glu + Thr219Asn
Ala199Ser + Tyr205Thr + Ala214Asp + Thr215Asp
Pro200Asn + Thr215Asp + Leu216Asp + Thr219Ser
Tyr208Asn + Tyr213His + Thr215Asp + Leu216Asp
Gly201Ser + Asn211Glu + Thr212Asp + Ala214His
Val204Thr + Asn211Glu + Thr212Asp + Ala214Pro
Pro200Gly + Thr212Gly + Asn217Glu + Gly218Asp
Thr207Gln + Ala214Gly + Asn217Glu + Gly218Glu
Thr207Asn + Pro209Gly + Asn217Asp + Gly218Glu
Tyr213Thr + Leu216Val + Asn217Glu + Gly218Glu
Ala199Thr + Ala202Asp + Gly203Glu + Gly218Pro
Val204His + Tyr205Asp + Thr212Gly + Thr215Glu
Pro200Ser + TyT205Glu + Thr215Glu + Leu216Cys
Ala202Glu + Gly203Asp + Asn217Glu + Thr21SPro
Pro209Glu + Asn211Asp + Leu216His + Thr219Pro
Thr207Pro + Tyr208Ser + Pro209Glu + Asn211Asp
Val204Thr + Pro209Asp + Asn211Asp + Thr212Pro
表26
环6-五取代变异体
Pro200Asn + Ala202Asn + Val204Gly + Asn211Asp + Thr219Gly
Pro200Gly + Gly201Pro + Pro209Glu + Asn211Ser + Thr212Gly
Pro200Ser + Asn211Gln + Ala214Gly + Asn217Glu + Thr219Asn
Gly201Asn + Gly203Ser + Thr207Ser + Tyr213Pro + Thr215Gly
Pro200Ser + Tyr205Pro + Tyr208Leu + Asn211Ser + Gly218Asp
Pro200Gly + Pro209Gly + Thr212Asp + Thr215Asn + Leu216Thr
Gly201Gln + Ala202Pro + Thr212Asn + Ala214Gln + Thr215Glu
Tyr205Pro + Tyr208Ser + Thr215Asp + Asn217Ser + Thr219Gln
Pro200Gly + Thr207Gly + Thr212Gln + Ala214Asn + Thr215Asn
Tyr205Ile + Thr207Pro + Tyr208Ala + Ala214Gln + Leu216Asn
Gly203Pro + Val204His + Thr207Gln + Tyr213Glu + Thr219Ser
Tyr205Met + Thr207Gly + Pro209Glu + Thr215Ser + Thr219Asn
Pro200Gly + Thr207Gln + Asn211Asp + Thr215Pro + Gly218Ser
Ala202His + Gly203Pro + Asn211Glu + Tyr213Gly + Thr219Gln
Ala199Gly + Tyr208Val + Thr210Glu + Tyr213Leu + Thr219Gln
Ala199His + Pro200Gln + Val204Cys + Thr215Asn + Asn217Asp
Ala199Gly + Pro200Gln + Tyr205Ile + Thr210Gly + Thr215Asp
Tyr208Met + Pro209Asn + Thr210Ser + Asn211Ser + Ala214Glu
Pro200Asn + Gly201Gln + Pro209Asn + Leu216Ala + Gly218Glu
Gly203Gln + Pro209Gly + Thr210Asn + Asn211Ser + Tyr213Val
Gly201Pro + Thr207Asn + Thr210Gln + Tyr213His + Ala214Thr
Pro200Gln + Gly201Ser + Val204Cys + Thr210Asn + Asn217Glu
Ala199Pro + Gly203Gln + Tyr205Val + Asn211Asp + Thr215Asn
Ala202Asn + Val204Ser + Asn211Ser + Tyr213Asn + Leu216Asp
Thr210Glu + Thr212Ser + Ala214Gly + Thr215Gln + Leu216Ile
Ala199Asn + Ala202Asn + Val204Gly + Tyr205Met + Thr215Asn
Thr210Gln + Thr212Pro + Tyr213Thr + Leu216Gln + Asn217Gln
Ala199Gln + Pro200Gly + Ala202Pro + Thr212Gln + Thr215Ser
Ala202Gln + Thr207Asn + Thr210Gln + Asn211Glu + Ala214Gly
Gly203Gln + Thr207Asn + Pro209Gly + Tyr213Asn + Gly218Asp
Val204Cys + Tyr205Thr + Thr207Asn + Leu216Thr + Thr219Gln
Gly201Gln + Val204His + Tyr208Ser + Thr212Ser + Thr215Gly
Pro200Gln + Ala202Asp + Tyr208Gly + Thr212Ser + Leu216Thr
Ala199Ser + Gly201Ser + Asn211Asp + Ala214Thr + Thr219Gly
Ala202Gln + Val204Ser + Tyr208Leu + Pro209Asp + Tyr213Gln
Ala199Ser + Tyr205Ala + Thr210Glu + Thr212Gly + Leu216Ala
Ala199His + Ala202Glu + Thr210Gly + Asn211Gln + Tyr213Pro
Ala199Ser + Gly203Glu + Tyr213Met + Leu216Ala + Gly218Ser
Gly201Gln + Gly203Ser + Thr207Asn + Gly218Pro + Thr219Pro
Ala199Gly + Pro209Asp + Thr212Ser + Leu216Met + Gly218Ser
Pro200Ser + Tyr205Ala + Ash211Ser + Thr212Glu + Thr215Pro
Pro209Glu + Thr212Gly + Ala214Thr + Leu216Asn + Thr219Pro
Gly203Asn + Tyr205Gln + Pro209Asn + Gly218Asn + Thr219Ser
Gly201Asn + Ala202Gln + Gly203Pro + Thr210Gly + Asn217Ser
Pro200Ser + Gly201Pro + Tyr208Val + Thr210Gly + Thr212Asn
Ala199His + Gly201Gln + Tyr205Glu + Leu216Met + Gly218Asn
Tyr205Ser + Asn211Gln + Thr212Asp + Thr215Ser + Thr219Gly
Ala199Thr + Gly201Gln + Ala202Thr + Gly203Glu + Tyr213Pro
Ala199Gln + Val204Asn + Thr207Asn + Tyr208Gly + Gly218Ser
Ala199His + Gly203Pro + Thr207Gln + Asn211Asp + Ala214Asn
Ala202Ser + Gly203Pro + Tyr205Ser + Thr210Glu + Leu216Asn
Thr207Gly + Tyr208Gly + Thr212Asp + Leu216Pro + Thr219Asn
Ala202Asp + Val204Tnr + Tyr208Ile + Thr212Asn + Gly218Ser
Gly203Gln + Val204Ala + Tyr205Pro + Asn211Gln + Thr215Gln
Pro200Ser + Gly203Pro + Thr212Pro + Leu216Met + Gly218Glu
Ala199Gly + Gly201Gln + Thr207Ser + Leu216Cys + Asn217Glu
Ala199Thr + Tyr205Met + Tyr208Val + Ala214Thr + Leu216Ile
Ala202Asp + Tyr205Pro + Thr207Ser + Tyr208Thr + Thr210Pro
Ala199Gln + Pro200Gln + Thr207Asn + Leu216Val + Gly218Ser
Gly201Ser + Tyr208Met + Asn211Asp + Thr212Ser + Thr219Gly
Gly203Ser + Thr207Pro + Asn211Ser + Asn217Asp + Gly218Gln
Gly201Pro + Gly203Asp + Tyr208Asn + Thr212Ser + Thr215Asn
Ala199Pro + Pro200Ser + Tyr205Asp + Thr212Pro + Leu216Val
Tyr208Ala + Tyr213Ile + Ala214Gly + Thr215Pro + Asn217Gln
Ala199Ser + Pro200Gly + Gly203Asp + Leu216His + Asn217Ser
Gly201Pro + Gly203Ser + Val204Asn + Leu216Ala + Asn217Asp
Val204Gln + Tyr208His + Asn211Asp + Tyr213His + Leu216Thr
Pro200Gly + Gly201Asn + Val204Gln + Tyr208Cys + Thr210Pro
Ala202Asn + Tyr205Gln + Pro209Glu + Thr210Gly + Gly218Asn
Pro200Gly + Thr207Pro + Tyr208Met + Leu216Val + Asn217Glu
Gly201Asn + Tyr205Gln + Thr212Asp + Thr215Pro + Asn217Gln
Gly203Glu + Tyr208Ile + Asn211Gln + Thr215Ser + Thr219Gly
Gly201Asn + Ala202Ser + Gly203Gln + Asn211Gln + Ala214Asn
Pro200Gly + Val204Asn + Pro209Gln + Thr210Pro + Thr219Ser
Tyr208Leu + Pro209Asn + Thr210Gln + Thr215Asn + Gly218Asn
Ala202Ser + Val204Cys + Thr210Asn + Ala214Glu + Thr215Gly
Pro200Asn + Gly203Gln + Tyr208Ser + Pro209Gln + Gly218Pro
Ala202Thr + Gly203Asp + Tyr205Leu + Thr212Gly + Tyr213Gln
Gly203Gln + Val204Pro + Thr210Asp + Gly218Gln + Thr219Gly
Tyr208His + Thr210Ser + Asn211Asp + Leu216Asn + Asn217Gln
Gly201Gln + Tyr205His + Thr210Glu + Tyr213lle + Leu216Ala
Pro200Asn + Gly201Ser + Ala202Glu + Gly203Gln + Thr219Ser
Ala199Gln + Gly201Asn + Ala202Asp + Thr212Asn + Tyr213Thr
Ala199Pro + Val204Ser + Tyr213His + Asn217Asp + Gly218Gln
Pro200Asn + Gly201Ser + Ala202Pro + Thr207Ser + Ala214Thr
Ala199Asn + Gly201Pro + Gly203Asp + Tyr213Cys + Gly218Gln
Gly201Gln + Gly203Asn + Val204Thr + Thr210Ser + Thr212Gln
Gly201Asn + Ala202Pro + Thr210Gln + Asn211Gln + Thr215Gln
Pro200Asn + Gly203Glu + Tyr205Val + Thr207Pro + Tyr208Ser
Ala199Gln + Gly201Ser + Ala202Pro + Thr207Pro + Ash217Glu
Pro200Gln + Tyr205Met + Thr212Pro + Leu216Glu + Thr219Ser
Ala199Pro + Pro200Gln + Ala202Gly + Thr207Gln + Asn217Glu
Tyr208Thr + Asn211Ser + Tyr213Gln + Leu216Gly + Thr219Pro
Gly203Ser + Val204Thr + Tyr208Ala + Tyr213Pro + Thr219Asn
Gly201Asn + Gly203Pro + Val204Ala + Asn211Ser + Thr219Gly
Gly203Glu + Thr207Ser + Tyr208Ser + Thr215Asn + Leu216Cys
Gly201Asn + Tyr208Leu + Asn211Glu + Leu216Gly + Asn217Ser
Gly201Pro + Tyr208Val + Thr210Pro + Leu216Gln + Gly218Asn
Pro200Asn + Ala202Thr + Thr207Ser + Thr212Ser + Ala214Asp
Gly203Glu + Val204Asn + Thr210Pro + Ala214His + Gly218Asn
Gly201Ser + Val204Met + Ala214Gly + Leu216Ser + Thr219Ser
Pro200Gly + Tyr208Val + Tyr213His + Leu216Ala + Asn217Glu
Thr212Glu + Tyr213Asn + Ala214Thr + Leu216Gln + Gly218Pro
Gly203Pro + Val204Asn + Tyr208Cys + Asn217Glu + Thr219Ser
Pro200Asn + Gly201Pro + Val204Thr + Leu216Asp + Asn217Gln
Tyr205Met + Tyr208Asn + Pro209Asn + Tyr213Ile + Ala214His
Gly201Gln + Gly203Pro + Tyr205Cys + Thr210Gln +Asn217Asp
Ala202Asn + Gly203Asp + Thr207Gly + Asn211Ser + Thr219Pro
Pro200Gly + Gly201Ser + Val204Cys + Thr207Gly + Gly218Asp
Pro200Ser + Gly203Pro + Tyr208Pro + Thr210Asn + Asn211Glu
Pro200Gly + Gly201Gln + Tyr205Leu + Pro209Asn + Ala214His
Tyr205Ser + Thr207Gln + Tyr208Ser + Thr212Ser + Thr219Asn
Ala202Gly + Asn211Asp + Thr212Gln + Ala214Thr + Leu216His
Ala202Pro + Val204Ala + Pro209Gln + Asn211Ser + Thr212Glu
Ala202Gly + Thr207Pro + Pro209Gly + Asn217Ser + Gly218Asp
Gly201Gln + Pro209Asn + Thr215Ser + Leu216Thr + Thr219Gly
Val204Thr + Tyr208Val + Thr215Asn + Asn217Glu + Gly218Asn
Tyr205Gln + Tyr213Pro + Thr215Ser + Leu216Gly + Gly218Gln
Gly203Asn + Thr207Ser + Thr210Asn + Asn211Asp + Leu216Met
Tyr208His + Pro209Ser + Leu216Gly + Asn217Glu + Gly218Gln
Thr207Ser + Pro209Gly + Asn211Glu + Thr215Ser + Leu216Ile
Ala199Gly + Ala202Glu + Thr207Pro + Tyr213Val + Ala214Gly
Ala199His + Pro200Gly + Gly201Ser + Gly218Asp + Thr219Pro
Ala202Glu + Tyr208Ala + Thr212Ser + Tyr213His + Thr219Asn
Ala199Thr + Val204Thr + Thr207Asn + Thr210Asn + Tyr213Glu
Tyr208Gln + Pro209Gly + Ala214Asn + Thr215Glu + Gly218Asn
Tyr208Thr + Tyr213Val + Thr215Asp + Leu216Val + Gly218Gln
Thr207Pro + Pro209Ser + Thr210Glu + Asn217Ser + Thr219Pro
Pro200Ser + Gly201Gln + Tyr205Ile + Pro209Gly + Thr219Gln
Val204Met + Thr207Pro + Tyr208Gln + Pro209Asn + Thr215Asp
Val204Ala + Pro209Asn + Thr210Gly + Tyr213Asp + Ala214Pro
Pro200Ser + Tyr208His + Tyr213Glu + Leu216Cys + Thr219Ser
Gly201Gln + Ala202Asp + Tyr205Ala + Leu216Thr + Asn217Gln
Ala199Gly + Gly201Asn + Val204Gly + Ala214Asn + Thr219Pro
Pro200Asn + Ala202Asn + Gly203Gln + Tyr208Asn + Asn217Asp
Val204Gly + Tyr205Asp + Asn211Gln + Tyr213Met + Ala214Gly
Gly201Asn + Thr207Gln + Pro209Gln + Ala214Asp + Gly218Pro
Pro200Gly + Tyr208Ser + Asn211Asp + Tyr213Ser + Thr219Gly
Ala202Thr + Val204Pro + Thr210Ser + Asn211Asp + Leu216Thr
Gly203Glu + Val204Met + Tyr205His + Asn211Gln + Thr219Gln
Gly201Gln + Ala202Gly + Val204Ser + Thr207Gln + Asn211Glu
Gly203Asn + Tyr208Ala + Pro209Asp + Thr212Asn + Thr215Gln
Ala199Gln + Pro200Asn + Tyr205Met + Asn211Asp + Thr219Gln
Val204Gln + Tyr208Val + Thr210Pro + Asn211Gln + Leu216Asp
Pro200Asn + Val204Pro + Tyr205Gly + Thr210Pro + Ala214Gly
Gly201Asn + Tyr205Met + Tyr213Cys + Leu216Ile + Thr219Asn
Ala199Gly + Gly201Asn + Gly203Gln + Val204Thr + Pro209Glu
Ala199His + Ala202Gly + Pro209Asp + Thr210Asp + Gly218Ser
Gly201Gln + Tyr208Ser + Pro209Asp + Thr210Glu + Gly218Ser
Ala199Asn + Pro200Ser + Thr212Asn + Ala214Asp + Thr215Glu
Ala199His + Pro200Ser + Thr215Gly + Asn217Asp + Gly218Glu
Tyr205Thr + Thr207Ser + Asn217Glu + Gly218Asp + Thr219Gly
Ala202Thr + Tyr205Glu + Tyr208Thr + Thr215Glu + Leu216Gln
Tyr205Asp + Thr212Gly + Thr215Glu + Asn217Ser + Gly218Ser
Pro200Gly + Ala202Glu + Val204Ala + Tyr208Leu + Asn217Asp
Pro200Gln + Ala202Asp + Tyr205Gly + Tyr213Ile + Asn217Asp
Pro209Asp + Asn211Asp + Tyr213Thr + Leu216Cys + Thr219Asn
Ala202Asn + Val204His + Pro209Glu + Asn211Glu + Thr219Asn
Pro200Ser + Ala202Asp + Val204Gln + Leu216Glu + Asn217Glu
Ala199Gly + Pro200Gln + Ala202Gln + Thr210Glu + Thr212Asp
Gly203Asp + Pro209Gly + Tyr213His + Asn217Asp + Gly218Ser
Gly203Asp + Val204Thr + Tyr208Ala + Tyr213Gln + Thr215Asp
Tyr205Pro + Thr207Gly + Pro209Glu + Thr212Asp + Gly218Ser
Ala199His + Ala202Glu + Gly203Gln + Asn211Gln + Gly218Asp
Ala199Asn + Ala202Glu + Thr207Gln + Thr212Gln + Gly218Asp
Ala202Asp + Gly203Asn + Tyr213Val + Leu216Ile + Gly218Glu
Pro200Gly + Ala202Thr + Tyr208Met + Ala214Glu + Leu216Asp
Gly203Glu + Thr207Gln + Thr212Ser + Ala214Glu + Thr215Glu
Val204His + Tyr205Asp + Tyr208Met + Tyr213Glu + Asn217Gln
Tyr205Gly + Tyr213Ile + Leu216Glu + Gly218Glu + Thr219Gly
Val204Ser + Asn211Gln + Tyr213Thr + Thr215Glu + Asn217Glu
Tyr208Leu + Thr210Gly + Thr212Gln + Tyr213Glu + Thr215Glu
Ala202Glu + Tyr205Glu + Thr210Gly + Asn211Gln + Leu216Asp
Pro200Asn + Thr210Ser + Thr212Asp + Tyr213Glu + Thr215Asp
Ala199Thr + Thr207Ser + Thr210Asp + Thr212Asn + Tyr213Asp
Ala202Glu + Val204Pro + Tyr205Thr + Tyr208His + Thr215Glu
Gly203Glu + Tyr208Ile + Thr210Gln + Tyr213Gly + Gly218Asp
Tyr208Gln + Pro209Glu + Tyr213Thr + Ala214Glu + Thr219Gln
Tyr205Asp + Thr210Ser + Thr215Gln + Asn217Asp + Gly218Glu
Gly201Asn + Tyr205Glu + Tyr208Leu + Asn217Asp + Gly218Glu
表27
环6-六取代变异体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环6-七取代突变变异体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
多环双突变变异体
Leu 95Gly + Gly203Pro
Thr 58Gln + Asn211Glu
Asp 59Glu + Leu216Gln
Tyr103Cys + Thr212Gly
Gly156Glu + Ala202Gln
Gly126Gln + Thr215Pro
Ser187Glu + Ala214Gly
Asn 96Gln + Ile106Ala
Gly101Asp + Tyr205Pro
Asn 61Ser + Gly165Gln
Gly 64Gln + Thr132Gly
Asp 59Glu + Tyr205Asn
Ser100Glu + Tyr205His
Ala128Ser + Thr161Gln
Ser158Asp + Gly203Ser
Asn 61Asp + Asn162Ser
Thr161Glu + Pro209Gln
Gly 62Asp + Pro209Asn
Ile106Leu + Asn211Asp
Tyr103Glu + Gly203Pro
Tyr166Pro + Thr215Glu
Ile106Leu + Pro209Glu
Thr163Pro + Ala202Pro
Thr132Gly + Leu216Ser
Ser104Glu + Phe188Val
Tyr166Asp + Ala186Thr
Ser160Glu + Thr215Ser
Gly 62Gln + Gly165Asp
Leu 95Gln + Ala128Glu
Ser 97Glu + Asn211Ser
Leu 95Asn + Phe188Leu
Asn157Gln + Thr212Ser
Leu125Gln + Gly159Asp
Asn211Asp + Tyr213Ala
Ser158Asp + Tyr205Cys
Ser160Glu + Asn162Gln
Ser187Asp + Tyr213Pro
Ser158Asp + Leu216Gln
Met198Ser + Ala202Asp
Gly 99Asn + Ala214Glu
Thr 58Asp + Gly130Gln
Gly156Glu + Tyr208Pro
Gly105Pro + Leu216Ile
Ser187Asp + Tyr205Gln
Thr215Ser + Gly218Asp
Gly 60Asn + Ala128Glu
Thr 65Gln + Leu125Asn
Gly101Gln + Gly218Asp
Asn211Ser + Thr212Asp
Thr212Gln + Thr215Gly
Gly203Glu + Tyr205Gly
Thr132Pro + Thr215Ser
Tyr103Asp + Gly165Gln
Thr 65Pro + Gly203Glu
Thr 58Glu + Gly 60Pro
Gly153Pro + Leu216Val
Asn 96Gln + Pro209Glu
Gly 99Asn + Gly153Asn
Ala128Asn + Ala202His
Gly105Glu + Gly130Ser
Thr212Gly + Gly218Glu
Phe188Val + Tyr205Ser
Asn217Glu + Thr219Gly
Leu 95Gln + Thr215Asp
Val 94Gln + Thr212Glu
Asn 61Ser + Gly203Asp
Ser129Glu + Asn154Gln
Gly 62Glu + Thr219Asn
Ser129Glu + Asn157Gln
Tyr103Ile + Thr215Pro
Ile164Thr + Thr215Ser
Gly101Gln + Tyr208Gln
Asn 61Gln + Thr212Asp
Thr212Asp + Leu216Ile
Asn211Glu + Leu216His
Gly153Glu + Thr163Gln
Gly203Asp + Thr215Asn
Asn 61Ser + Thr132Asn
Ser158Glu + Gly203Pro
Leu 95Asn + Thr215Asn
Thr 58Ser + Ser100Asp
Ile164Met + Tyr205Ala
Ser158Glu + Tyr205Met
Phe188Leu + Thr212Glu
Gly130Pro + Ser190Glu
Gly101Asp + Gly126Ser
Asn 61Ser + Ala186Ser
Asn162Gln + Phe188Ser
Thr 58Gly + Thr 65Asp
Ser129Asp + Thr210Pro
Ser129Glu + Gly153Asn
Asp 59Glu + Tyr213Thr
Asn154Glu + Leu216Gln
Ser 97Asp + Ala199Pro
Gly203Asn + Gly218Asp
Thr 65Ser + Gly126Gln
Thr215Gln + Leu216Ala
Asn 96Glu + Tyr205His
Thr 65Ser + Gly156Asp
Ala128Ser + Thr163Gln
Thr215Asp + Leu216Val
Ser129Glu + Leu216Ala
Asn211Asp + Tyr213Leu
Val 94Ser + Phe188Val
Ser158Glu + Thr212Gln
Gly 99Pro + Asn154Gln
Gly203Asn + Thr215Gln
Ala202Ser + Tyr205Glu
Tyr205Val + Asn217Glu
Val 94Gly + Thr215Asp
Ser104Glu + Thr215Gln
Gly201Pro + Tyr205Glu
Asn 96Ser + Gly127Glu
Tyr205Thr + Ala214Thr
Gly 62Ser + Thr215Gly
Thr 65Ser + Asn211Glu
Gly153Pro + Gly203Ser
Ala128Thr + Leu216Glu
Asn 96Glu + Leu216Gln
Thr215Asn + Leu216Ala
Asp 59Glu + Ala202Pro
Val 94Thr + Leu216Met
Ser 97Glu + Thr212Gln
Gly101Asp + Ala202Asn
Ser129Asp + Ala202Gly
Tyr103Ala + Thr132Ser
Leu 95Met + Leu216Asp
Ser100Asp + Gly101Pro
Ser100Asp + Thr219Pro
Val 94Asn + Thr215Pro
Tyr103Asn + Ala128Asp
Gly201Asn + Tyr205Asp
Thr 58Glu + Ile106Cys
Thr 65Glu + Tyr103Pro
Val 94Met + Ser 98Glu
Gly203Glu + Thr210Pro
Pro209Glu + Gly218Ser
Thr161Pro + Gly203Ser
Ser 97Glu + Leu125Pro
Ala128Asp + Ala199His
Ile106Gly + Ala214Pro
Asp 59Glu + Tyr205Ile
Asn154Asp + Tyr208His
Thr210Asp + Leu216Val
Thr215Asp + Thr219Gln
Gly 60Glu + Leu125Ser
Ser 97Asp + Gly153Asn
Tyr166Pro + Tyr205Gly
Ser129Glu + Gly203Gln
Gly153Gln + Pro209Gln
Gly203Gln + Thr215Gln
Asn 61Glu + Thr212Ser
Thr163Asp + Tyr205His
Asn154Glu + Tyr166Thr
Asn 96Ser + Gly105Asp
Phe188Gly + Asn211Ser
Gly153Glu + Gly218Asn
Ser100Asp + Thr212Pro
Gly126Pro + Gly156Asp
Leu125Ser + Thr215Glu
Asp 59Glu + Thr 65Gly
Gly156Pro + Gly159Asp
Thr 58Asp + Ala202Ser
Leu 95Met + Gly 99Ser
Asn 61Gln + Tyr205Glu
Gly126Asn + Gly159Pro
Leu 95Thr + Gly218Asn
Thr 65Gln + Leu216Ala
Gly130Pro + Gly203Glu
Gly 64Gln + Ser 98Glu
Gly126Asn + Gly127Gln
Thr132Asn + Gly153Asn
Thr 58Pro + Gly105Glu
Gly130Asp + Ala214His
Gly126Glu + Tyr205Pro
Tyr103His + Ser129Asp
Ser 98Asp + Pro209Ser
Asp 59Glu + Tyr205Ser
Gly 60Gln + Thr215Asn
Pro209Asn + Asn211Asp
Thr132Asp + Ala202Asn
Gly218Ser + Thr219Gly
Thr 58Asp + Gly203Gln
Ile106Asp + Ala202Thr
Thr161Asn + Ser190Asp
Ser129Glu + Thr215Gln
Thr212Asn + Leu216Asp
Asn 96Gln + Ser158Glu
Gly203Pro + Thr215Asp
Val 94Asp + Tyr205Asn
Ala128Pro + Thr215Asp
Gly201Gln + Leu216Pro
Thr161Pro + Leu216Cys
Gly126Asp + Tyr205Met
Ser 97Asp + Thr212Pro
Thr210Asn + Leu216His
Ala186Asn + Tyr205Pro
Ser131Asp + Phe188Thr
Thr132Ser + Leu216Asn
Asn 61Asp + Gly130Pro
Leu 95Asn + Thr132Gly
Ser100Asp + Thr215Asn
Tyr166Thr + Asn211Gln
Thr207Pro + Leu216Thr
Ile106Ala + Gly165Asp
Tyr205Asn + Thr215Asn
Asn 96Gln + Gly218Asn
Ser160Asp + Phe188Gly
Thr210Gly + Thr212Ser
Gly 64Ser + Asn 96Gln
Gly101Asp + Asn154Gln
Gly 64Asn + Asn157Glu
Asn157Glu + Gly203Gln
Tyr213Asn + Thr215Ser
Gly101Asn + Ala186Asn
Ser160Glu + Leu216Ile
Tyr103His + Ser187Glu
Asn 96Ser + Ala214Asn
Gly 62Asn + Ser 98Asp
Ile106Ser + Asn157Glu
Thr 65Glu + Thr212Gly
Gly 62Asn + Thr215Pro
Gly127Ser + Thr210Asp
Gly156Ser + Asn217Glu
Ser 98Asp + Tyr103Thr
Gly127Asn + Gly165Pro
Gly153Glu + Leu216Met
Ser129Asp + Gly165Pro
Asn 61Asp + Thr163Gln
Gly126Asn + Tyr205Met
Asn157Gln + Thr212Gln
Ser190Glu + Thr215Gly
Ser129Glu + Gly165Gln
Thr 65Glu + Gly101Pro
Val 94Gln + Gly165Glu
Tyr103Ile + Serl90Glu
Asn217Gln + Gly218Ser
Ser 98Glu + Gly 99Asn
Ser102Asp + Thr212Ser
Thr212Ser + Thr215Asp
Gly130Glu + Pro209Asn
Ala128Pro + Gly165Pro
Asn217Gln + Gly218Asn
Leu125His + Thr163Ser
Thr212Asp + Tyr213Ala
Thr 65Pro + Thr210Ser
Gly105Pro + Ala202Gln
Val 94Thr + Ser158Glu
Thr132Gly + Tyr205Cys
Gly105Gln + Ala214His
Gly127Gln + Tyr205Cys
Gly127Glu + Gly130Ser
Phe188Tyr + Asn211Ser
Gly 64Asn + Phe188Asp
Thr212Ser + Leu216Asp
Met198His + Gly203Pro
Tyr208Val + Thr210Asp
Gly165Glu + Tyr205Met
Asn 96Gln + Thr132Glu
Ser 98Glu + Gly101Pro
Leu125Cys + Thr215Asp
Leu125Cys + Gly126Ser
Ser190Asp + Thr215Asn
Gly 99Gln + Gly153Asp
Gly 60Asp + Thr210Ser
Ser160Asp + Phe188Leu
Val204Gln + Thr215Glu
Asn157Glu + Gly203Ser
Tyr213Glu + Leu216Cys
Asn 61Gln + Leu216Asp
Leu125Gly + Ser155Asp
Ala186Pro + Val204Gln
Ile106Met + Ser160Asp
Leu125Ile + Leu216Ser
Gly 60Gln + Gly203Asn
Gly203Asn + Pro209Glu
Gly 60Ser + Gly 62Asp
Asn154Glu + Tyr213Gln
Asn154Gln + Gly203Asp
Asn154Glu + Thr219Gln
Thr212Ser + Leu216His
Tyr205His + Thr215Asp
Gly 60Asp + Gly126Pro
Gly203Glu + Leu216Ala
Gly156Ser + Gly203Ser
Ser155Asp + Gly165Gln
Ala202Gly + Thr212Asp
Asn157Gln + Tyr213Ser
Gly 99Ser + Gly130Ser
Gly203Ser + Thr219Asp
Gly130Ser + Ala214Asn
Thr210Gln + Thr212Asp
Asn211Gln + Thr212Glu
Gly153Asn + Thr212Asp
Gly153Asp + Tyr205Ser
Ser100Glu + Tyr205Thr
Gly126Asn + Ash154Glu
Ser190Asp + Tyr205Val
Ser102Glu + Tyr166Thr
Asn157Ser + Pro209Gly
Asp 59Glu + Tyr205Leu
Ser131Glu + Thr132Asn
Gly127Asp + Leu216Thr
Thr210Asn + Asn217Gln
Gly203Ser + Gly218Asp
Ile106Gln + Gly203Asp
Thr161Asp + Tyr205Pro
Gly105Pro + Thr212Asp
Tyr205Thr + Leu216Cys
Asn 96Ser + Ser158Glu
Asn 61Glu + Tyr205Ala
Tyr205Ser + Thr212Glu
Thr163Gln + Phe188Glu
Tyr103Ala + Thr212Asp
Gly101Asp + Ala202His
Ser155Glu + Gly218Asn
Val 94Thr + Thr163Ser
Ser102Glu + Asn217Gln
Gly159Ser + Ala202Glu
Asn154Ser + Tyr166Ala
Asp 59Glu + Phe188Gly
Thr163Gln + Gly218Ser
Gly153Asn + Tyr205Val
Ser129Glu + Leu216Pro
Gly126Ser + Thr212Glu
Gly203Asp + Leu216Asn
Gly126Glu + Thr212Pro
Val 94Pro + Ala186Glu
Gly165Gln + Thr215Glu
Ser129Asp + Gly153Asn
Ser 98Glu + Thr215Ser
Gly203Ser + Leu216Pro
Gly105Asp + Leu216His
Ile106Met + Thr210Asp
Leu216Asn + Asn217Glu
Gly127Gln + Leu216Glu
Val 94Glu + Ala202Gly
Thr132Asn + Leu216His
Asp 59Glu + Ala202Asn
Leu 95Ile + Tyr205Pro
Thr 65Gln + Thr215Pro
Ser104Glu + Thr215Asn
Ser160Glu + Gly203Pro
Gly203Asp + Leu216Pro
Ala128Glu + Tyr213Met
Ser 98Asp + Leu216Cys
Ser158Asp + Thr215Gln
Val 94Asp + Thr215Asn
Gly127Asp + Asn154Ser
Gly126Ser + Gly156Gln
Leu125Gly + Thr212Asp
Thr210Glu + Leu216Asn
Asn154Ser + Ala202Asn
Asn 96Gln + Gly126Asn
Gly 64Ser + Gly127Ser
Asn211Ser + Thr212Gly
Ser160Asp + Gly218Gln
Tyr205Ala + Thr210Glu
Thr212Asp + Leu216Ser
Ser158Asp + Thr215Ser
表33
多环三突变变异体
Leu 95Gly + Gly203Pro + Asn211Glu
Thr 58Gln + Asp 59Glu + Leu216Gln
Tyr103Cys + Ala202Gln + Thr212Gly
Gly126Gln + Gly156Glu + Thr215Pro
Asn 96Gln + Ile106Ala + Tyr205Pro
Asn 61Ser + Gly101Asp + Gly165Gln
Gly 64Gln + Thr132Gly + Gly153Glu
Ala128Ser + Ser158Asp + Gly203Ser
Tyr103Glu + Ile106Leu + Gly203Pro
Thr163Pro + Ala202Pro + Leu216Ser
Ser104Glu + Thr132Gly + Phe188Val
Leu 95Asn + Phe188Leu + Asn211Ser
Asn162Gln + Ser187Asp + Tyr213Pro
Thr 58Asp + Gly130Gln + Tyr208Pro
Gly105Pro + Gly156Glu + Leu216Ile
Gly 60Asn + Thr215Ser + Gly218Asp
Thr 65Gln + Leu125Asn + Ala128Glu
Asn211Ser + Thr212Asp + Gly218Gln
Gly203Glu + Thr212Gln + Thr215Gly
Thr132Pro + Tyr205Gly + Thr215Ser
Gly 60Pro + Thr 65Pro + Gly203Glu
Asn 96Gln + Gly203Asp + Thr215Gly
Gly 99Asn + Gly153Asn + Pro209Glu
Gly105Glu + Ala128Asn + Ala202His
Tyr205Ser + Thr212Gly + Gly218Glu
Gly 99Gln + Asn217Glu + Thr219Gly
Leu 95Gln + Ala202Thr + Thr215Asp
Ser129Glu + Asn157Gln + Thr219Asn
Asp 59Glu + Tyr103Ile + Thr215Pro
Gly 99Asp + Ile164Thr + Thr215Ser
Asn 61Gln + Tyr208Gln + Thr212Asp
Gly153Glu + Thr163Gln + Thr215Asn
Asn 61Ser + Thr132Asn + Gly203Pro
Leu 95Asn + Ser158Glu + Thr215Asn
Thr 58Ser + Ser100Asp + Tyr205Ala
Ser158Glu + Phe188Leu + Tyr205Met
Asn 61Ser + Ala186Ser + Phe188Ser
Asn154Glu + Tyr213Thr + Leu216Gln
Ser 97Asp + Ala199Pro + Gly203Asn
Thr 65Ser + Gly126Gln + Gly218Asp
Tyr205His + Thr215Gln + Leu216Ala
Ala128Ser + Thr163Gln + Thr215Asp
Val 94Ser + Ser100Asp + Phe188Val
Gly 99Pro + Asn154Gln + Gly203Asn
Asn 96Ser + Thr163Asp + Ala214Thr
Tyr166Asp + Tyr205Thr + Ala214Thr
Gly 62Ser + Thr 65Ser + Thr215Gly
Gly153Pro + Gly203Ser + Thr219Glu
Asp 59Glu + Ala202Pro + Leu216Ala
Val 94Thr + Thr212Gln + Leu216Met
Ser129Asp + Thr132Ser + Ala202Gly
Leu 95Met + Tyr103Ala + Leu216Asp
Ser104Asp + Gly203Asn + Thr215Asn
Thr 65Glu + Tyr103Prc + Ile106Cys
Val 94Met + Ser 98Glu + Thr210Pro
Ser 97Glu + Thr161Pro + Gly203Ser
Leu125Pro + Ala128Asp + Ala199His
Asp 59Glu + Tyr205Ile + Ala214Pro
Ser129Glu + Tyr166Pro + Gly203Gln
Gly203Gln + Pro209Gln + Thr215Gln
Asn 96Ser + Asn154Glu + Tyr166Thr
Gly105Asp + Phe188Gly + Asn211Ser
Thr 58Asp + Gly156Pro + Ala202Ser
Leu 95Met + Gly 99Ser + Ser131Glu
Gly126Asn + Gly159Pro + Tyr205Glu
Leu 95Thr + Leu216Ala + Gly218Asn
Gly105Glu + Gly130Pro + Leu216Thr
Thr 58Pro + Thr132Asn + Gly153Asn
Gly 60Gln + Pro209Asn + Thr215Asn
Thr132Asp + Ala202Asn + Thr219Gly
Thr 58Asp + Gly203Gln + Gly218Ser
Ser129Glu + Thr212Asn + Thr215Gln
Gly203Pro + Tyr205Asn + Thr215Asp
Ala128Pro + Thr215Asp + Leu216Pro
Thr161Pro + Gly201Gln + Leu216Cys
Gly126Asp + Tyr205Met + Thr212Pro
Ser 97Asp + Thr210Asn + Leu216His
Ala186Asn + Phe188Thr + Tyr205Pro
Asn 61Asp + Thr132Ser + Leu216Asn
Leu 95Asn + Thr132Gly + Thr215Asp
Ile106Asp + Tyr166Thr + Asn211Gln
Ile106Ala + Thr207Pro + Leu216Thr
Tyr205Asn + Thr215Asn + Gly218Asn
Asn 96Gln + Ser160Asp + Phe188Gly
Gly 64Ser + Gly101Asp + Asn154Gln
Thr 58Asn + Gly 64Asn + Asn157Glu
Gly101Asn + Ala186Asn + Thr215Ser
Gly 62Asn + Asn 96Ser + Ala214Asn
Gly130Asp + Phe188Asn + Gly218Pro
Gly 99Ser + Ala128Glu + Leu216Gly
Val 94Ala + Pro209Ser + Thr215Asp
Leu 95Thr + Gly126Gln + Leu216Gly
Asp 59Glu + Gly159Ser + Tyr205Pro
Ala199Gly + Ala202Gln + Gly203Gln
Ser158Glu + Thr161Gly + Tyr205Asn
Thr 58Ser + Ser190Glu + Tyr205Pro
Leu 95Asn + Tyr205Ala + Pro209Glu
Thr 58Asn + Gly153Pro + Thr210Pro
Ala128Gly + Gly203Asn + Tyr205Pro
Val 94Cys + Asn157Gln + Tyr205Gln
Ser129Glu + Tyr205Asn + Thr215Asn
Thr 65Gly + Gly127Asn + Thr212Gln
Gly 60Asn + Tyr205Thr + Asn217Gln
Gly126Pro + Ala214Pro + Thr215Gly
Thr163Gly + Ala202Pro + Thr212Pro
Thr 58Pro + Gly130Ser + Ser160Asp
Gly153Ser + Asn154Glu + Thr212Gly
Leu125Gly + Tyr205Thr + Thr210Glu
Tyr103Val + Thr132Gly + Pro200Gln
Ser100Glu + Gly127Asn + Leu216Ala
Ser160Glu + Ala186Ser + Tyr205Ser
Gly159Ser + Thr163Glu + Ile164His
Gly101Asp + Tyr166Cys + Gly203Pro
Thr 58Asn + Asn 61Glu + Ala214Ser
Gly127Glu + Phe188Met + Ala202Gly
Asn162Glu + Gly203Ser + Gly218Asn
Tyr166Asn + Thr210Glu + Ala214His
Ile106Asp + Thr212Ser + Thr215Gln
Asn162Glu + Ala202Ser + Thr215Asn
Tyr103Pro + Ser187Glu + Tyr205Pro
Val204Thr + Thr212Ser + Thr215Gln
Ser100Asp + Gly101Asn + Leu216His
Leu 95Cys + Gly203Ser + Leu216Thr
Asn 96Ser + Gly153Glu + Tyr205Pro
Ile106Pro + Ala202Ser + Thr212Pro
Ser102Asp + Gly203Ser + Thr215Ser
Leu 95Cys + Ser104Glu + Phe188Cys
Asp 59Glu + Ala214Gly + Thr215Gly
Leu125Cys + Thr212Gly + Thr215Ser
Leu125Val + Ala202His + Thr215Pro
Gly101Asp + Gly127Asn + Gly203Gln
Thr 58Pro + Gly105Asn + Gly203Glu
Tyr166Cys + Gly203Ser + Tyr205Asp
Gly126Gln + Ile164Val + Leu216His
Gly153Ser + Thr215Pro + Leu216Glu
Ser160Asp + Gly218Gln + Thr219Asn
Asn 61Glu + Asn157Ser + Ala214Ser
Gly 62Ser + Ala128Ser + Asn154Asp
Ile106Gly + Tyr205Val + Thr215Ser
Ser160Glu + Asn211Gln + Thr215Pro
Thr 65Gly + Ser100Glu + Thr215Pro
Thr 65Ser + Ser104Glu + Gly203Pro
Asn 61Ser + Ile106Ser + Thr219Pro
Ser102Asp + Gly153Pro + Leu216His
Thr 65Glu + Gly126Gln + Gly203Ser
Ser187Glu + Phe188Ile + Leu216Thr
Thr132Gln + Gly165Asp + Thr212Ser
Ser129Asp + Asn211Ser + Leu216Thr
Leu125Ile + Asn154Ser + Leu216Met
Thr 65Pro + Gly101Pro + Thr212Asp
Thr 65Gln + Leu125Asn + Leu216Met
Pro209Gly + Ala214Glu + Thr215Pro
Leu125Val + Tyr205Ser + Leu216His
Leu 95Ash + Gly156Asp + Thr212Gln
Leu125Pro + Thr210Asp + Thr215Ser
Gly105Asn + Thr161Asn + Tyr205Val
Gly126Ser + Ser160Glu + Gly218Asn
Val 94Thr + Thr207Gly + Thr212Glu
Tyr208Gln + Thr212Gln + Gly218Asp
Ile106Asn + Gly159Asn + Phe188Asp
Tyr205Met + Thr212Gln + Tyr213Thr
Ser131Asp + Thr132Pro + Tyr213Asn
Gly105Ser + Ala128Thr + Thr215Pro
Leu 95His + Asn 96Asp + Thr215Gly
Gly 60Ser + Gly156Asp + Pro209Asn
Asn162Asp + Thr163Asn + Ala199Gly
Tyr103Glu + Leu125His + Thr215Ser
Gly127Glu + Asn157Gln + Tyr205Leu
Asp 59Glu + Gly105Pro + Ala186Asn
Ala128Asp + Tyr213His + Leu216His
Thr 65Pro + Ile106His + Ser131Glu
Gly126Gln + Thr161Asp + Ala202Ser
Gly 99Gln + Gly126Asp + Thr212Gly
Thr 58Asp + Gly159Ser + Thr212Ser
Asp 59Glu + Ala186Gly + Gly203Gln
Gly 62Glu + Tyr205Ala + Leu216Gln
Asn154Gln + Tyr205Ser + Tyr213Met
Leu 95Ser + Ser158Glu + Thr212Gly
Tyr166Asn + Tyr205Glu + Leu216Asn
Asp 59Glu + Ala128Asn + Gly153Pro
Gly 99Asp + Gly159Asn + Asn211Ser
Gly165Pro + Thr215Gln + Leu216Thr
Gly165Asn + Thr215Asn + Leu216Asp
Gly 60Asp + Gly159Asn + Thr212Gln
Thr 58Glu + Thr163Gly + Thr210Gln
Gly159Ser + Ala186Gly + Ala199Thr
Gly203Glu + Asn211Gln + Tyr213Gln
Gly 99Ser + Asn162Asp + Thr215Gln
Thr 65Gly + Ser190Glu + Thr219Ser
Val 94Met + Asn162Ser + Ser190Glu
Asn 61Gln + Leu 95Ser + Tyr205Asp
Gly126Glu + Gly201Pro + Gly203Ser
Thr 58Gly + Thr212Pro + Gly218Glu
Ser104Asp + Gly105Ser + Gly159Pro
Thr 58Gln + Thr163Gly + Tyr205Pro
Gly 60Ser + Thr 65Pro + Thr163Gly
Leu125Met + Ser131Glu + Thr210Gly
Phe188Gln + Gly201Ser + Ala202Gly
Gly203Pro + Leu216Glu + Asn217Gln
Gly 62Asn + Gly126Pro + Thr210Asn
Gly 60Asn + Ser104Asp + Gly127Gln
Ser155Glu + Thr163Pro + Gly203Pro
Leu 95Met + Thr132Asp + Asn211Ser
Tyr103Ile + Gly159Asp + Ala202Gly
Gly101Gln + Tyr166Pro + Asn211Gln
Asn157Ser + Ala202Ser + Thr215Gly
Thr132Pro + Ser155Glu + Ala214His
Thr132Ser + Thr212Asp + Leu216Gly
Gly126Asn + Thr212Asp + Gly218Gln
Tyr205Cys + Thr212Pro + Leu216Met
Asn 96Gln + Thr163Gln + Leu216Glu
Ala202Ser + Tyr205Asn + Gly218Asp
Leu 95Asp + Gly126Asn + Ala202Ser
Ser100Glu + Ile106Gln + Ala202Gln
Thr 58Ser + Ser100Glu + Ile164Leu
Gly101Ser + Phe188Asp + Tyr213Gly
Ser104Glu + Gly156Gln + Thr212Gly
Leu125Val + Gly203Asn + Tyr205Thr
Thr163Gln + Thr215Asn + Asn217Glu
Gly 99Pro + Gly165Glu + Ala214Gln
Met198Ser + Gly203Glu + Tyr213Val
Asn 96Ser + Ser 97Asp + Thr212Ser
Ala202Glu + Gly203Ser + Tyr205Gln
Gly 64Asn + Asn154Gln + Asn157Glu
Thr 65Ser + Gly130Pro + Tyr205Asp
Gly126Gln + Thr161Gln + Pro209Ser
Asp 59Glu + Phe188Leu + Leu216Ala
Ser131Asp + Thr212Pro + Thr215Gln
Val204Pro + Gly218Asn + Thr219Glu
Thr 58Ser + Gly203Glu + Thr212Pro
Gly 62Gln + Tyr205Gly + Thr212Glu
Thr 65Gly + Ser129Asp + Met198Gly
Leu 95Cys + Asn162Ser + Ala202Thr
Leu125Asp + Tyr205Thr + Leu216Thr
Asn157Asp + Tyr205Ser + Leu216Pro
Gly156Asn + Gly165Asn + Thr215Glu
Ala128Ser + Thr163Glu + Ala199Ser
Val 94Ser + Ser187Glu + Thr215Gln
Val 94Pro + Thr215Gly + Asn217Glu
Val 94Asn + Gly130Glu + Gly203Ser
Gly127Gln + Ile164Cys + Thr210Gln
Ser187Glu + Thr212Gln + Thr215Asn
Asn 96Gln + Gly126Gln + Gly203Asp
Gly165Asp + Thr210Ser + Tyr213Ile
Thr 58Gln + Gly153Ser + Asn217Gln
Gly203Asp + Tyr205Met + Thr210Ser
Ala128Pro + Thr132Glu + Thr215Pro
Ser104Glu + Phe188Ser + Thr212Gly
Ser104Glu + Asn157Ser + Ala214Ser
Ile106Cys + Tyr205Glu + Thr215Asn
Gly156Gln + Tyr205Pro + Thr219Asp
Asn157Glu + Tyr213Thr + Leu216Ile
Thr 58Ser + Asn162Ser + Ser190Asp
Gly 62Ser + Thr 65Asp + Tyr205Ile
Leu125Pro + Gly203Asp + Thr212Ser
Thr 58Gln + Ser 98Asp + Thr212Pro
Ser 97Asp + Leu125Val + Tyr213Ile
Ile106Leu + Phe188Glu + Thr212Asn
Thr 58Pro + Gly153Pro + Asn162Gln
Ser190Asp + Thr210Gln + Leu216Ala
Ser100Glu + Thr215Gly + Leu216Met
Gly127Glu + Ala128Gln + Thr212Pro
Val 94Pro + Gly127Ser + Thr215Glu
Thr161Asp + Ala199Asn + Tyr213Gln
Thr132Pro + Tyr166Leu + Thr212Pro
Gly203Gln + Thr215Asn + Leu216Met
Gly127Pro + Gly203Asp + Tyr205Ile
Thr163Ser + Tyr205Ala + Pro209Gln
Val 94Cys + Thr215Asn + Gly218Pro
Gly126Glu + Gly165Pro + Ala202Thr
Thr 58Glu + Tyr103Asn + Pro200Gln
Phe188Thr + Ala202Asn + Asn217Ser
Gly 60Asp + Leu 95Thr + Tyr205Ile
Ser102Glu + Ala202Ser + Leu216Gln
Gly 99Asn + Ser102Asp + Gly218Gln
Ile106Ala + Thr132Pro + Asn157Asp
Gly203Ser + Tyr205Gln + Thr212Glu
Ser 97Asp + Ile106His + Asn211Gln
Ser100Glu + Gly201Asn + Tyr205His
Thr212Gln + Ala214Glu + Thr215Glu
Asn 61Glu + Gly 62Glu + Gly203Pro
Asn211Glu + Thr212Asp + Leu216Ile
Gly 99Glu + Ser100Asp + Gly156Pro
Ala202Asp + Gly203Glu + Tyr205Pro
Ser 97Asp + Ser 98Glu + Thr163Gln
Ala128Thr + Tyr205Asp + Thr215Asp
Ser102Asp + Gly105Asp + Leu216Gly
Gly127Glu + Gly165Glu + Tyr205Thr
Gly203Asp + Tyr205Ala + Asn217Glu
Asp 59Glu + Asn 61Glu + Gly130Ser
Gly 60Glu + Ser 97Glu + Gly 99Asn
Tyr166Ala + Gly203Glu + Leu216Asp
Ser158Asp + Ser160Glu + Tyr205Cys
Ser129Asp + Tyr166Glu + Thr215Gln
Ser160Asp + Asn162Glu + Thr210Gly
Ser129Asp + Ser131Asp + Gly218Asn
Asn 61Gln + Leu 95Asp + Ser 97Asp
Gly 62Ser + Leu216Asp + Gly218Glu
Asp 59Glu + Gly165Asn + Pro209Glu
Leu 95Glu + Gly105Glu + Gly203Ser
Asn157Glu + Asn162Asp + Thr215Gln
Tyr103His + Asn162Glu + Ser190Asp
Ser158Asp + Asn162Glu + Thr163Pro
Gly 62Glu + Ser 97Glu + Gly153Gln
Gly 99Gln + Asn154Asp + Ser187Glu
Ser155Asp + Gly156Asp + Thr219Asp
Gly1535er + Asn154Asp + Gly165Asp
Asp 59Glu + Asn 96Glu + Tyr205Ser
Asp 59Glu + Asn 96Asp + Leu216Ile
Ser104Asp + Ser131Asp + Thr210Gly
Ser 97Glu + Gly101Asp + Ala202Asn
Asn 96Asp + Ser102Glu + Ile106Gly
Asn 61Asp + Leu 95Glu + Val204Gln
Gly156Asp + Thr161Asp + Gly203Ser
Ser100Asp + Tyr103Asp + Gly126Asn
Thr 65Asp + Tyr205Asp + Thr215Asp
Thr 58Asp + Ser 97Asp + Thr132Gly
Thr215Pro + Leu216Asp + Thr219Glu
Ser100Glu + Gly105Glu + Asn162Gln
Ala186Glu + Ala202Glu + Thr215Asp
Asn 61Asp + Ser100Glu + Asn162Ser
Ser104Asp + Thr132Glu + Tyr205Met
Gly 99Asn + Thr210Asp + Ala214Glu
Asn157Asp + Ala186Glu + Thr212Ser
Gly 62Asp + Pro209Asn + Asn211Asp
Gly105Pro + Gly153Glu + Asn162Asp
Ala186Ser + Thr212Asp + Thr215Asp
Gly 99Glu + Ser102Asp + Leu125Gly
Gly 62Ser + Gly 99Glu + Ser102Asp
Gly126Glu + Ser155Glu + Tyr205Pro
Ser158Glu + Phe188Asp + Ala202Ser
Gly 60Glu + Leu 95Glu + Ile106Leu
Ala128Asp + Asn154Asp + Thr215Gln
Ala128Asp + Asn162Glu + Asn217Ser
Asn157Gln + Ser187Glu + Asn217Asp
Gly 62Asp + Ser 98Asp + Thr212Glu
Gly159Glu + Ser160Asp + Phe188Asp
Gly126Asp + Thr210Glu + Asn211Glu
Ser104Asp + Gly105Asp + Ala128Glu
Asp 59Glu + Tyr205Glu + Thr215Glu
Gly 60Asp + Tyr205Glu + Thr215Asp
Asn 61Glu + Pro209Asp + Thr215Glu
Asn 61Asp + Ser 97Glu + Tyr205Asp
Ser102Asp + Gly105Glu + Pro209Asp
Ser158Asp + Gly203Asp + Thr215Glu
Asn157Asp + Gly203Asp + Thr215Asp
Gly127Glu + Ser129Glu + Thr215Glu
Asn 96Glu + Ser100Asp + Thr212Glu
Ser190Glu + Gly203Glu + Tyr205Glu
Ser129Asp + Phe188Glu + Ser190Asp
Ser100Asp + Ser104Glu + Thr212Pro
Ser129Glu + Gly153Glu + Asn157Gln
Ser 97Glu + Thr212Glu + Ala214Glu
Ser100Asp + Ser102Glu + Ser160Asp
Asp 59Glu + Ser 97Glu + Asn157Asp
Ser 97Glu + Asn157Glu + Ser160Glu
Gly101Asn + Leu125Glu + Ser129Glu
Gly101Glu + Gly130Glu + Thr161Ser
Asn154Glu + Ser158Glu + Gly203Gln
Val 94Asp + Ser187Asp + Ser190Glu
表34
多环四突变变异体
Thr 58Gln + Leu 95Gly + Gly203Pro + Asn211Glu
Asp 59Glu + Tyr103Cys + Thr212Gly + Leu216Gln
Gly126Gln + Gly156Glu + Ala202Gln + Thr215Pro
Asn 615er + Gly101Asp + Gly165Gln + Tyr205Pro
Ser104Glu + Thr132Gly + Phe188Val + Leu216Ser
Gly127Glu + Thr212Asn + Thr215Ser + Leu216Gln
Leu125Thr + Thr210Gln + Thr212Asp + Thr215Ser
Thr132Pro + Gly203Glu + Tyr205Gly + Thr215Gly
Asn 96Gln + Gly153Pro + Pro209Glu + Leu216Val
Gly 99Asn + Ala128Asn + Gly153Asn + Ala202His
Phe188Val + Tyr205Ser + Thr212Gly + Gly218Glu
Tyr103Ile + Ser129Glu + Asn157Gln + Thr215Pro
Leu 95Asn + Ser158Glu + Gly203Pro + Thr215Asn
Thr 58Ser + Ser100Asp + Ile164Met + Tyr205Ala
Thr 65Ser + Gly126Gln + Gly203Asn + Gly218Asp
Asn 96Glu + Tyr205His + Thr215Gln + Leu216Ala
Thr 65Ser + Ala128Ser + Gly156Asp + Thr163Gln
Asn154Gln + Asn157Glu + Gly203Asn + Thr215Gln
Gly 62Ser + Tyr205Thr + Ala214Thr + Thr215Gly
Asp 59Glu + Asn 61Gln + Ala202Pro + Leu216Ala
Leu 95Met + Tyr103Ala + Thr132Ser + Tyr205Asp
Ser 97Glu + Leu125Pro + Thr161Pro + Gly203Ser
Asp 59Glu + Ile106Gly + Tyr205Ile + Ala214Pro
Ser129Glu + Tyr166Pro + Gly203Gln + Tyr205Gly
Asn 61Glu + Gly203Gln + Thr212Ser + Thr215Gln
Gly153Glu + Phe188Gly + Asn211Ser + Gly218Asn
Tyr103Val + Gly126Asn + Pro209Asn + Leu216Asp
Ala128Ser + Ser190Glu + Tyr213Ala + Leu216Val
Gly127Ser + Asn154Ser + Tyr205Leu + Thr212Gln
Ala128Glu + Asn154Gln + Asn157Gln + Leu216His
Ser155Asp + Gly156Ser + Gly203Ser + Leu216Ala
Gly 99Ser + Gly130Ser + Asn157Gln + Tyr213Ser
Gly130Ser + Gly203Ser + Ala214Asn + Thr219Asp
Asp 59Glu + Asn157Ser + Tyr166Thr + Pro209Gly
Gly203Ser + Thr210Asn + Leu216Thr + Asn217Gln
Tyr103Ala + Thr163Gln + Ala202His + Thr212Asp
Gly 62Glu + Asn154Ser + Tyr166Ala + Phe188Gly
Asp 59Glu + Thr163Gln + Tyr205Val + Gly218Ser
Val 94Pro + Gly153Asn + Gly165Gln + Thr215Glu
Ser 98Glu + Asn154Gln + Gly203Ser + Leu216Pro
Ile106Glu + Thr132Asn + Ala202Gly + Leu216His
Asp 59Glu + Leu 95Ile + Ala202Asn + Tyr205Pro
Thr 65Gln + Asn 96Gln + Ser104Glu + Thr215Pro
Leu125Gly + Gly126Ser + Asn154Ser + Gly156Gln
Asn 96Gln + Gly126Asn + Gly127Ser + Ala202Asn
Gly 64Ser + Ser160Asp + Asn211Ser + Thr212Gly
Gly101Pro + Ser158Asp + Thr215Ser + Leu216Ser
Val 94Glu + Ala202Ser + Tyr205Ser + Thr212Gly
Gly153Ser + Asn154Glu + Thr161Asn + Thr212Gly
Tyr103Val + Leu125Gly + Thr132Gly + Pro200Gln
Gly127Pro + Phe188Met + Ala202Gly + Thr215Glu
Asn162Glu + Gly203Ser + Ala214His + Gly218Asn
Asn 96Glu + Gly 99Pro + Thr132Asn + Thr212Pro
Gly101Asn + Tyr205Gly + Tyr213Met + Thr215Asp
Leu125Pro + Gly126Asn + Ala202Gly + Gly203Asp
Gly 99Pro + Gly127Glu + Tyr205His + Thr212Gln
Tyr103Pro + Tyr166Ile + Gly203Ser + Thr207Gly
Thr 58Gly + Gly 62Ser + Gly105Glu + Ala128Ser
Ile106Gly + Tyr205Val + Thr215Ser + Leu216Asp
Asn 61Ser + Ile106Ser + Ser190Asp + Thr219Pro
Thr 65Glu + Gly126Gln + Gly203Ser + Leu216His
Ser187Glu + Phe188Ile + Thr212Ser + Leu216Thr
Leu125Ile + Asn154Ser + Asn211Ser + Leu216Met
Thr 65Gln + Leu125Asn + Pro209Gly + Ala214Glu
Leu125Val + Tyr205Ser + Thr215Pro + Leu216His
Leu 95Asn + Leu125Pro + Gly156Asp + Thr212Gln
Gly126Ser + Ser160Glu + Thr161Asn + Gly218Asn
Leu 95Gln + Phe188Ile + Ala202His + Thr212Asn
Ser104Glu + Thr212Gln + Tyr213Thr + Thr215Pro
Ala128Thr + Ser131Asp + Thr132Pro + Tyr213Asn
Leu 95His + Ser129Glu + Thr215Gly + Leu216Asn
Gly105Pro + Ala128Asp + Tyr213His + Leu216His
Gly159Ser + Tyr205Ala + Thr212Gly + Leu216Ala
Gly 62Glu + Asn154Gln + Tyr205Ser + Leu216Gln
Leu 95Ser + Ser158Glu + Val204Gln + Thr212Gly
Ser 98Asp + Leu216Ile + Asn217Ser + Gly218Asn
Leu125Gly + Ala128Gln + Gly203Glu + Pro209Gln
Gly105Asn + Tyr166Val + Ala202Asp + Thr215Pro
Ser 97Asp + Ala202Gln + Pro209Gln + Thr215Gly
Leu 95Thr + Tyr205Ile + Leu216Val + Thr219Ser
Gly 99Asn + Ser102Asp + Tyr205Ser + Gly218Gln
Thr 65Pro + Gly165Asn + Ala186Pro + Gly201Ser
Asn154Asp + Gly203Gln + Tyr205Asn + Thr212Asn
Ile106His + Ala186Gly + Thr212Asn + Leu216Asp
Gly126Glu + Gly127Gln + Ala128Asn + Ala202Asn
Leu 95Ser + Ala128Gly + Thr161Gln + Asn217Glu
Asp 59Glu + Gly101Pro + Ala199Gly + Gly203Ser
Asn 61Asp + Val204Thr + Tyr205Cys + Thr215Ser
Gly101Asn + Asn162Asp + Phe188Ala + Tyr205His
Ala186Thr + Gly203Asp + Thr215Pro + Asn217Ser
Tyr103Gln + Phe188Met + Thr210Glu + Thr219Pro
Gly 62Asp + Gly126Ser + Leu216Pro + Thr219Asn
Gly 60Gln + Tyr205Pro + Thr212Ser + Thr215Asp
Ile106Val + Ser187Glu + Thr212Pro + Thr215Ser
Gly159Asp + Asn211Ser + Ala214Gln + Thr219Pro
Asn 96Ser + Ser160Asp + Ala214Gln + Leu216Gly
Ser104Asp + Gly203Ser + Tyr205Ser + Thr215Pro
Gly153Gln + Phe188Tyr + Thr215Glu + Leu216Ile
Ser129Asp + Thr132Pro + Thr212Gln + Leu216Thr
Ala128Gln + Ala186Asn + Thr215Asp + Gly218Asn
Asn 96Asp + Ala202Pro + Tyr205Gln + Pro209Ser
Thr132Asp + Gly165Pro + Thr210Pro + Leu216Cys
Gly105Pro + Gly130Ser + Thr161Glu + Thr215Pro
Ile106Cys + Asn157Gln + Tyr205His + Thr212Gln
Leu 95Gly + Ser100Glu + Gly101Pro + Gly126Ser
Gly 60Ser + Gly105Asp + Thr132Pro + Tyr205Thr
Thr132Ser + Ile164Val + Gly203Glu + Gly218Asn
Gly203Gln + Thr212Pro + Thr215Asn + Leu216Pro
Asn 61Ser + Val 94Gln + Thr132Asp + Val204Gly
Gly201Ser + Thr212Gly + Thr215Pro + Leu216Pro
Gly153Pro + Ser160Glu + Gly165Gln + Gly218Ser
Tyr166Ile + Tyr213Ser + Ala214Glu + Gly218Asn
Thr132Ser + Thr163Asp + Tyr166Val + Ala199His
Thr132Gln + Asn157Ser + Tyr208Gly + Thr215Asp
Gly203Glu + Tyr205Ile + Tyr208Pro + Thr215Ser
Gly127Ser + Gly165Ser + Tyr213Ile + Leu216His
Gly156Glu + Pro209Asn + Asn211Ser + Thr212Ser
Gly165Gln + Phe188Leu + Ala202His + Thr212Glu
Ser102Asp + Gly130Asn + Asn162Ser + Leu216Thr
Val 94Cys + Gly156Ser + Tyr205Ala + Leu216Ala
Gly 99Pro + Tyr103Glu + Phe188His + Leu216Cys
Tyr103Gln + Gly126Gln + Gly156Pro + Tyr166Asn
Leu125Pro + Gly153Glu + Asn162Ser + Thr215Ser
Gly 64Ser + Gly 99Asn + Gly203Asp + Thr210Gln
Gly 99Asn + Ala202Pro + Thr212Asn + Gly218Ser
Ala128Ser + Gly203Gln + Pro209Glu + Gly218Ser
Thr 58Ser + Tyr103Gly + Leu125Gly + Tyr205Gly
Asn154Glu + Thr161Ser + Asn162Gln + Gly203Gln
Gly101Pro + Asn154Ser + Leu216His + Asn217Glu
Thr163Asp + Ala186Ser + Tyr205His + Leu216Pro
Gly159Glu + Gly203Ser + Thr210Gly + Leu216His
Leu125Ser + Thr163Glu + Ala202Pro + Thr210Gln
Gly153Pro + Ser155Glu + Thr210Asn + Leu216Cys
Gly101Asp + Gly126Gln + Tyr166Ile + Gly203Pro
Ser158Asp + Gly201Asn + Va1204Asn + Pro209Ser
Tyr166Pro + Tyr205Pro + Thr215Asp + Leu216Ala
Gly 64Asn + Ser160Asp + Tyr205Leu + Thr215Gly
Gly101Ser + Thr132Gln + Gly159Pro + Thr161Ser
Tyr103Val + Asn154Gln + Gly159Ser + Asn211Asp
Gly127Glu + Asn157Ser + Thr161Asn + Thr212Gly
Asn 61Asp + Ile164Asn + Pro209Asn + Leu216Val
Ser158Asp + Asn162Ser + Tyr205Asn + Thr212Gly
Leu125Val + Ala202Thr + Pro209Asn + Thr219Pro
Gly 60Asp + Asn154Gln + Gly203Pro + Tyr205Leu
Leu125Gln + Asn162Ser + Gly203Asp + Thr215Gly
Ser102Asp + Tyr103Ile + Tyr205Met + Thr212Gln
Gly 99Gln + Ala214Gln + Leu216Ala + Gly218Glu
Gly105Asn + Gly156Gln + Gly203Pro + Gly218Asp
Ile106Ser + Thr212Asn + Leu216Glu + Gly218Gln
Asn 96Asp + Tyr166Thr + Gly203Ser + Gly218Pro
Ser155Asp + Gly203Gln + Tyr213Thr + Thr215Gly
Gly101Ser + Thr212Gly + Thr215Asn + Leu216His
Thr 65Ser + Leu 95Asn + Thr132Ser + Asn162Asp
Leu 95Met + Thr132Asn + Gly203Asp + Thr215Gly
Asn154Ser + Gly203Glu + Thr212Gly + Leu216Ala
Gly 60Ser + Ser131Glu + Thr210Ser + Asn217Gln
Gly 62Asn + Asn162Gln + Gly203Gln + Thr215Ser
Gly 99Ser + Tyr103Ala + Thr210Gln + Thr219Ser
Asn162Gln + Gly203Glu + Thr215Gln + Thr219Gly
Asn162Gln + Gly165Asn + Leu216Ile + Gly218Ser
Thr 65Asn + Gly 99Gln + Thr212Ser + Leu216Asp
Phe188Val + Ala214Asp + Thr215Asp + Leu216Gln
Gly 99Glu + Ser100Asp + Gly156Pro + Tyr205Gln
Val 94Asn + Tyr103Asp + Ser104Asp + Thr215Pro
Gly 64Gln + Asn154Asp + Ser155Glu + Leu216Ser
Asn154Glu + Ser155Glu + Gly156Gln + Thr215Pro
Gly153Pro + Asn157Ser + Tyr205Asp + Thr215Asp
Tyr103Leu + Phe188Met + Tyr205Glu + Thr215Glu
Tyr205Ala + Thr210Glu + Thr212Asp + Gly218Gln
Ala202His + Tyr205Val + Thr210Glu + Thr212Asp
Gly203Asp + Tyr205His + Thr212Ser + Thr215Asp
Gly127Glu + Ser129Asp + Gly153Asn + Thr210Pro
Ala128Asn + Tyr166Ala + Gly203Asp + Leu216Asp
Ser 98Glu + Ser100Glu + Gly203Ser + Asn217Gln
Asn 61Asp + Gly 62Pro + Asn 96Asp + Ser 97Glu
Gly159Ser + Thr161Glu + Thr163Glu + Ile164His
Sgr160Glu + Asn162Glu + Asn211Gln + Thr215Pro
Asn 61Asp + Gly 99Asp + Thr161Gly + Thr212Pro
Ile106Val + Ala186Asp + Ala202Glu + Thr215Gln
Asp 59Glu + Gly 64Asn + Thr 65Glu + Tyr213Ser
Val 94Pro + Thr215Asp + Leu216His + Asn217Asp
Asp 59Glu + Gly127Asn + Ala186Pro + Pro209Asp
Tyr103Glu + Ser131Glu + Tyr213His + Thr215Ser
Ser129Asp + Thr132Gln + Gly165Asp + Leu216Thr
Tyr205Ile + Thr212Glu + Ala214Glu + Thr219Gln
Thr 58Gly + Thr212Gln + Tyr213Asp + Thr215Asp
Asp 59Glu + Gly 62Gln + Ser 97Glu + Tyr205Gly
Phe188Asp + Tyr208Gln + Thr212Gln + Gly218Asp
Ala186Asp + Ala202Asp + Gly203Glu + Tyr205Pro
Asn154Asp + Ser155Asp + Gly165Ser + Ser187Asp
Ser 97Asp + Ser100Glu + Gly201Asn + Tyr205His
Ser 97Asp + Ser100Asp + Gly165Ser + Tyr205Thr
Gly105Ser + Asn157Glu + Ser160Glu + Thr215Pro
Gly156Asp + Thr161Asp + Ser190Asp + Gly203Ser
Thr 65Gly + Ser100Glu + Leu125Glu + Thr215Pro
Gly 62Glu + Ser 97Glu + Gly153Gln + Pro209Gln
Gly 62Glu + Asn157Gln + Gly203Gln + Thr210Glu
Asn154Asp + Gly165Glu + Thr212Gly + Gly218Pro
Asn162Asp + Gly165Asp + Thr212Gln + Leu216Cys
Ser100Glu + Gly126Glu + Ala186Gln + Tyr205Ala
Asn154Ser + Ser158Asp + Ser190Glu + Thr215Gln
Ser129Asp + Thr132Asp + Gly203Gln + Thr215Gln
Asn 96Ser + Thr163Asp + Tyr166Asp + Ala214Thr
Gly 60Glu + Ser 97Glu + Tyr166Asn + Thr210Glu
Ile106Gln + Asn154Glu + Gly203Asp + Gly218Asp
Ser100Glu + Gly105Glu + Asn162Gln + Phe188Ser
Ala186Glu + Ala202Glu + Thr215Asp + Asn217Gln
Asp 59Glu + Gly 99Asp + Ile164Thr + Thr215Ser
Ser129Glu + Asn154Glu + Asn162Ser + Gly165Asp
Ser104Glu + Thr132Glu + Tyr205Met + Thr215Pro
Gly 62Asp + Ile106Leu + Pro209Asn + Asn211Asp
Asp 59Glu + Ser 98Asp + Gly156Asn + Gly218Asn
Asn 96Asp + Ser100Asp + Ser104Glu + Thr132Asn
Ala128Asp + Asn154Asp + Tyr213Met + Thr215Gln
Ser131Glu + Gly153Asp + Gly165Asp + Tyr205Gly
Gly101Asp + Gly105Asp + Gly126Asn + Ser131Glu
Ser129Asp + Asn157Glu + Thr163Glu + Thr212Pro
Gly 99Asn + Ala128Asp + Asn162Glu + Asn217Ser
Ser 98Asp + Ser100Asp + Ser104Glu + Thr212Pro
Asn 96Gln + Ile106Ala + Gly156Glu + Ser160Asp
Gly203Asp + Pro209Glu + Thr215Asp + Leu216Pro
Asn 96Asp + Ser 97Glu + Ser104Asp + Ala214Ser
Gly159Asn + Gly165Glu + Leu216Cys + Thr219Asp
Gly153Asp + Ser187Asp + Gly203Asn + Leu216Ser
Thr 65Glu + Tyr103Pro + Thr215Asp + Asn217Asp
Gly 62Asp + Ala202Glu + Thr215Asp + Leu216Ser
Thr 65Asp + Gly101Gln + Gly130Ser + Tyr205Asp
Asn 61Ser + Thr 65Asp + Thr132Asn + Tyr205Asp
Gly 99Pro + Ser187Glu + Gly203Glu + Thr215Asp
Thr 65Ser + Gly126Asp + Thr210Glu + Asn211Glu
Asn 96Asp + Thr161Asp + Asn162Glu + Ala202Asn
Gly101Glu + Ile106Met + Ala214Asp + Thr215Glu
Asn 61Asp + Ala186Asp + Ser187Glu + Leu216Pro
Ser104Asp + Tyr205Val + Thr215Asp + Leu216Asp
Leu 95Asp + Ala202His + Thr215Glu + Leu216Glu
Ser129Asp + Gly130Asp + Thr210Gln + Thr212Asp
Ile106Glu + Asn162Ser + Asn211Asp + Thr212Glu
Thr 58Asp + Asp 59Glu + Ile106Glu + Phe188Ser
Ser104Asp + Gly105Asp + Ala128Glu + Thr215Gly
Gly 99Asp + Ser100Glu + Thr161Glu + Pro209Gln
Gly 99Asp + Ser100Glu + Ser190Asp + Tyr205His
Asp 59Glu + Ala202Asp + Gly203Glu + Leu216Cys
Asp 59Glu + Gly 60Glu + Ala128His + Thr215Asp
Gly126Asn + Asn154Glu + Ser155Asp + Gly203Asp
Ser 97Glu + Ser 98Asp + Ser131Glu + Asn162Ser
Ser 97Glu + Ser 98Glu + Ser104Asp + Asn217Gln
Gly156Ser + Ser158Glu + Tyr205Asp + Thr215Asp
Asn 61Glu + Ser 97Glu + Ala128His + Tyr205Asp
Gly 64Asn + Thr 65Glu + Phe188Tyr + Thr215Glu
Tyr103Glu + Ile106Glu + Gly203Pro + Gly218Glu
Gly 99Glu + Ser102Asp + Leu125Gly + Ser131Glu
Ser102Glu + Asn154Glu + Ser190Asp + Tyr205Val
Gly127Glu + Gly165Glu + Tyr205Thr + Thr215Glu
Val 94Glu + Gly105Asp + Gly165Pro + Thr212Glu
Asn 96Glu + Ser 98Asp + Tyr103Leu + Ser155Asp
Asp 59Glu + Asn 61Glu + Gly105Glu + Gly130Ser
Asp 59Glu + Asn 61Asp + Thr 65Gly + Tyr205Glu
Leu 95Gln + Tyr103Asp + Gly203Asp + Thr215Asp
Asn157Asp + Gly203Asp + Thr215Asp + Thr219Gln
Ser 98Glu + Gly203Glu + Thr215Glu + Gly218Ser
Ser131Asp + Asn154Gln + Gly203Asp + Thr215Glu
Gly101Asn + Ile106Glu + Gly159Asp + Thr161Asp
Gly127Glu + Ser129Glu + Gly130Ser + Thr215Glu
Gly153Gln + Ser155Asp + Pro209Glu + Thr212Asp
Gly203Glu + Thr210Asp + Leu216Asp + Gly218Pro
Val 94Ser + Ser 98Glu + Ser100Asp + Phe188Asp
Asn 96Asp + Ser100Asp + Gly153Ser + Asn162Glu
Gly 62Glu + Leu 95Glu + Ile106Leu + Thr212Glu
Ser160Asp + Asn162Asp + Ala202Glu + Thr215Gln
Asn 61Glu + Ser 98Asp + Gly126Glu + Leu216Asn
Ser100Asp + Ser104Asp + Gly105Asn + Tyr205Cys
Asn 61Asp + Gly 99Glu + Tyr103Thr + Ser131Asp
Val 94Glu + Asn157Asp + Thr163Glu + Tyr205Ala
Gly127Asp + Ser131Glu + Thr132Asn + Ser190Asp
Gly 60Glu + Gly156Ser + Asn211Asp + Thr212Gln
Gly126Asp + Gly156Glu + Ser158Glu + Tyr205Met
Asn157Asp + Tyr205Val + Thr215Asp + Asn217Glu
Asp 59Glu + Gly203Glu + Tyr205Asn + Pro209Asp
Gly105Glu + Ser131Asp + Gly156Asn + Leu216Pro
Asp 59Glu + Ser 97Glu + Asn157Asp + Phe188His
Ser 97Asp + Ser100Asp + Ala202Pro + Thr212Glu
Val 94Gly + Ser104Glu + Asn157Asp + Ser160Asp
Asn 61Glu + Asn162Glu + Ser190Asp + Thr215Gly
Asn 81Asp + Asn 96Glu + Gly127Ser + Gly130Asp
Thr 65Glu + Gly165Glu + Phe188Val + Ser190Glu
Tyr103Pro + Asn154Glu + Ser158Glu + Tyr205Pro
Ser131Glu + Gly156Glu + Tyr166Asp + Ala186Thr
Asn 61Glu + Thr 65Asp + Tyr103Pro + Ser131Asp
Gly130Glu + Asn154Asp + Ser187Asp + Tyr208His
Ser131Glu + Thr163Pro + Gly203Glu + Gly218Glu
Asp 59Glu + Asn162Asp + Thr163Gln + Thr210Glu
Leu125Asp + Gly165Asp + Gly203Asp + Tyr213Gln
Gly101Asp + Asn162Asp + Gly165Glu + Leu216Asn
Ser155Glu + Ser158Asp + Tyr205Glu + Leu216Pro
Gly 60Asp + Tyr205Pro + Thr212Asp + Thr215Glu
Ser160Glu + Thr163Asp + Tyr205His + Thr215Asp
Thr 58Pro + Asn 61Glu + Asn154Glu + Asn157Asp
Asp 59Glu + Gly 99Ser + Gly153Asn + Thr212Asp
Ser 97Glu + Gly101Asp + Thr132Glu + Ala202Asn
Thr 58Glu + Asn 61Asp + Ala186Gly + Ser187Asp
Thr 58Glu + Asn 61Glu + Ile106Cys + Thr132Asp
Ser158Glu + Gly203Glu + Asn211Gln + Ala214Asp
Ser129Asp + Gly203Glu + Ala214Glu + Thr215Ser
Ser104Asp + Ser155Glu + Asn157Gln + Asn162Glu
Thr 65Glu + Ser160Glu + Tyr205Cys + Thr210Asp
Asn 61Asp + Leu 95Glu + Tyr205Glu + Leu216Asn
Ser100Glu + Gly127Glu + Thr215Asp + Leu216Pro
Ser102Glu + Gly127Glu + Ala128Pro + Asn162Glu
Asn 96Asp + Gly105Glu + Asn162Asp + Thr212Gln
Ser 98Glu + Gly127Asp + Ser131Glu + Ala202Ser
Asp 59Glu + Leu 95Asp + Gly159Asp + Tyr205Val
Thr 65Glu + Ser187Glu + Thr215Gln + Gly218Glu
Gly156Glu + Gly159Glu + Tyr205Met + Thr215Glu
Gly 64Gln + Gly130Glu + Thr132Gly + Gly153Glu
Asp 59Glu + Gly 99Glu + Ser129Glu + Tyr205Asn
Gly126Glu + Ala186Glu + Gly203Asp + Thr212Pro
Thr 58Asp + Tyr103Val + Ser104Glu + Thr132Asp
Ser104Glu + Thr132Glu + Ala202Glu + Asn217Gln
Ser 98Glu + Thr132Gly + Ser187Glu + Thr219Glu
Gly 62Asp + Ser100Asp + Gly105Asn + Tyr205Val
Ser 97Asp + Ala186Asn + Thr212Glu + Thr215Asp
Ser187Asp + Ala202Gly + Thr212Glu + Thr215Glu
Asp 59Glu + Ser 98Glu + Gly153Ser + Asn154Glu
Gly126Glu + Thr132Asp + Ser155Asp + Ala202Gly
Asp 59Glu + Gly 62Pro + Ser158Glu + Phe188Glu
Ser104Asp + Gly130Asp + Tyr205Pro + Thr212Glu
Thr 65Pro + Ser102Asp + Gly126Asp + Ala202Asp
Gly 62Asp + Ser129Glu + Thr215Asp + Leu216Val
Ala128Asp + Asn162Asp + Ala202Ser + Tyr205Glu
Ser 97Glu + Tnr132Gly + Gly153Asp + Ser187Asp
Ser102Asp + Leu125Asp + Gly156Asp + Gly218Asn
Asn 61Gln + Ser160Glu + Ser190Asp + Gly203Glu
Asn 96Asp + Tyr166Asp + Ser190Glu + Leu216Cys
Thr 65Asp + Asn162Glu + Tyr205Asp + Thr215Pro
Thr 65Asp + Ser 97Glu + Gly105Glu + Leu216Met
Val 94Asp + Ser 98Asp + Ser158Asp + Thr215Gln
Thr 65Glu + Gly101Asp + Thr215Glu + Gly218Asn
Thr 58Asp + Gly156Ser + Asn211Asp + Thr215Glu
Gly 99Gln + Gly126Gln + Asn154Asp + Thr161Asp
Gly156Asn + Ala186Asp + Thr212Glu + Leu216Asp
Ser 97Asp + Ile106Gln + Leu125Asp + Thr215Ser
Ser100Glu + Ser104Glu + Tyr166Ala + Ser187Asp
Ser 98Asp + Ser102Glu + Thr215Glu + Leu216Gln
Ser129Asp + Thr132Gln + Ser155Glu + Ser160Asp
Ser 97Glu + Thr163Asp + Ser187Glu + Thr219Ser
Leu125Cys + Ser129Glu + Ser155Glu + Thr215Gly
Asn 61Asp + Gly126Glu + Ala214Asp + Thr219Gln
Thr 58Asn + Gly 64Pro + Thr 65Glu + Val 94Asp
Tyr205Leu + Tyr213Asp + Thr215Gly + Asn217Asp
Asp 59Glu + Ala202Gln + Thr212Asp + Asn217Asp
Asp 59Glu + Ser187Asp + Pro209Asn + Thr212Asp
Thr 58Asn + Leu 95Glu + Phe188Asp + Gly203Asp
Ser187Glu + Tyr205Ile + Thr212Glu + Leu216Glu
Ser129Glu + Gly156Gln + Thr212Asp + Leu216Asp
Gly101Glu + Ser129Glu + Asn157Glu + Tyr205Thr
Gly 60Glu + Leu125Ser + Gly153Glu + Ala202Asp
Asn 61Glu + Phe188Glu + Tyr205Ala + Thr212Glu
Asn 61Glu + Gly153Asn + Gly159Asp + Thr212Asp
Thr 65Gly + Tyr205Glu + Asn211Glu + Leu216Cys
Thr 65Glu + Asn 96Glu + Gly156Asp + Leu216Cys
表35
多环五突变变异体
Tyr103Cys + Gly156Glu + Ala202Gln + Thr212Gly + Leu216Gln
Asn 61Ser + Gly101Asp + Thr132Gly + Gly165Gln + Tyr205Pro
Gly 60Asn + Asn162Gln + Thr212Pro + Tyr213Gln + Asn217Asp
Gly 60Ser + Leu 95His + Ser100Glu + Gly156Pro + Thr215Gly
Leu 95Ile + Thr163Gln + Tyr205Glu + Tyr213Ala + Thr219Asn
Gly159Glu + Phe188Tyr + Gly203Pro + Tyr205Gly + Ala214Thr
Gly126Pro + Gly127Gln + Tyr208Asn + Thr212Gln + Leu216Ala
Leu 95Gly + Asn162Ser + Gly165Glu + Pro209Gly + Thr210Gly
Thr 58Ser + Leu 95Asn + Ser100Asp + Tyr205Ala + Thr215Asn
Asn 61Ser + Ser100Glu + Asn162Gln + Phe188Ser + Gly203Ser
Tyr103Ser + Thr161Pro + Phe188Gln + Gly201Gln + Leu215Pro
Ser 97Asp + Gly 99Pro + Tyr205Pro + Thr210Asn + Leu216His
Leu 95Met + Ser187Asp + Tyr205Val + Thr212Gly + Leu216Asn
Ile106Asp + Tyr166Thr + Thr207Pro + Asn211Gln + Leu216Thr
Asn 96Gln + Ser160Asp + Phe188Gly + Thr215Asn + Gly218Asn
Thr 58Pro + Ser 98Asp + Gly130Pro + Tyr205Val + Tyr208Pro
Val 94Ala + Gly 99Ser + Ala128Glu + Phe188Asn + Leu216Gly
Leu 95Thr + Gly126Gln + Pro209Ser + Thr215Asp + Leu216Gly
Ser 98Glu + Asn154Gln + Gly203Ser + Thr215Ser + Leu216Pro
Thr 58Asp + Leu125Pro + Ala202Thr + Gly203Pro + Tyr205Val
Ala128Asp + Gly153Gln + Ala186His + Ala199Gln + Gly203Asn
Asp 59Glu + Gly159Gln + Ala186His + Leu216Ser + Gly218Ser
Ala186His + Phe188Gln + Ser190Glu + Pro200Ser + Tyr205Val
Val 94Gly + Ser 98Asp + Tyr166Pro + Ala202Gly + Tyr205His
Asn 61Gln + Tyr103Met + Asn157Asp + Thr161Ser + Gly203Asn
Thr 65Gln + Gly203Gln + Pro209Ser + Thr215Gln + Asn217Asp
Gly126Ser + Ser160Asp + Thr163Gln + Gly203Ser + Tyr205Gly
Tyr103Gln + Gly156Pro + Tyr166Asn + Phe188Tyr + Thr215Glu
Val 94Met + Gly 99Asn + Thr212Asn + Tyr213Gly + Thr215Glu
Ala128Ser + Ala202Pro + Gly203Gln + Pro209Glu + Gly218Ser
Thr 58Ser + Tyr103Gly + Leu125Gly + Tyr205Gly + Gly218Ser
Leu125Pro + Asn154Glu + Thr161Ser + Asn162Gln + Gly203Gln
Gly 60Asp + Gly 62Pro + Asn 96Ser + Ala186Gly + Thr210Gln
Asn 61Gln + Thr 65Ser + Leu 95Met + Tyr103Gly + Ser104Asp
Asn 61Ser + Gly155Gln + Gly156Gln + Ala202Gly + Asn211Gln
Ser102Asp + Gly126Gln + Ala202Asn + Tyr205Leu + Thr212Gly
Gly 60Asn + Ser104Asp + Gly105Asn + Thr163Ser + Tyr205Ala
Val 94Asn + Gly101Asn + Leu125Asn + Gly130Asp + Leu216Cys
Gly159Pro + Gly165Pro + Ala186Thr + Gly201Pro + Leu216Ser
Leu 95Cys + Gly130Gln + Thr212Ser + Ala214Asn + Leu216Val
Gly 60Glu + Thr161Gly + Gly203Gln + Tyr205Ile + Thr212Gly
Gly153Ser + Gly159Asn + Ala186Pro + Gly203Ser + Thr219Asn
Asp 59Glu + Gly 62Gln + Leu125Ala + Thr215Asn + Asn217Gln
Val 94Gly + Leu125Ser + Gly203Glu + Thr215Gln + Leu216Asn
Ala186Gln + Ser187Glu + Gly201Gln + Ala202Pro + Gly203Ser
Asn157Asp + Gly203Gln + Thr212Pro + Ala214Gln + Leu216Ala
Thr 65Ser + Ser 98Asp + Thr161Ser + Gly203Ser + Tyr205Cys
Gly 99Asp + Gly165Ser + Met198Cys + Pro209Gly + Thr215Gly
Gly153Pro + Tyr208Ile + Thr212Ser + Ala214His + Gly218Glu
Ser 98Asp + Ile106Ala + Tyr166Leu + Ala202Gln + Ala214Thr
Gly 99Gln + Ser129Glu + Thr161Asn + Phe188Cys + Tyr205Thr
Gly 60Asn + Gly159Gln + Ser160Glu + Thr163Pro + Ala186Gly
Asn 96Asp + Gly 99Pro + Val204Ala + Tyr205Val + Asn217Gln
Gly 60Ser + Phe188Ile + Thr212Asn + Thr215Ser + Leu216Asp
Ala128Gly + Thr132Gln + Gly165Gln + Met198Thr + Gly203Glu
Asn 96Asp + Asn157Ser + Gly201Asn + Thr212Gly + Thr215Gly
Gly 99Asp + Phe188Val + Gly203Asn + Thr207Asn + Leu216Pro
Ala128Pro + Thr132Pro + Thr163Pro + Phe188Thr + Thr212Glu
Val 94His + Gly105Glu + Ala186Pro + Thr210Gln + Thr212Gly
Gly101Pro + Thr132Asp + Asn162Ser + Thr212Gln + Leu216Pro
Gly 60Asn + Tyr103Asn + Gly105Gln + Ile106Ala + Leu216Gln
Gly126Pro + Gly203Pro + Tyr205Val + Thr215Asn + Leu216Ser
Thr 58Glu + Thr161Gly + Tyr205Asn + Tyr208Leu + Leu216Thr
Gly101Asn + Gly159Pro + Asn162Ser + Ala202Gly + Tyr205Gln
Tyr103Ile + Gly105Pro + Ala202Gln + Thr210Gln + Leu216Pro
Gly 60Asp + Gly153Ser + Thr163Gly + Thr212Pro + Leu216Met
Gly101Pro + Ser104Glu + Gly105Pro + Gly153Ser + Tyr205Thr
Asn162Asp + Phe188His + Ala199Gly + Thr212Pro + Leu216Pro
Ala186Gly + Ala202Pro + Tyr205Met + Thr210Gln + Leu216Val
Gly 62Ser + Tyr103Met + Asn154Glu + Thr212Gly + Leu216Met
Thr 65Ser + Gly101Pro + Tyr103Leu + Thr161Asp + Thr219Ser
Gly105Ser + Ile106His + Ala186Thr + Met198Pro + Thr212Pro
Thr163Gly + Gly203Asp + Thr212Asn + Leu216His + Asn217Gln
Gly126Ser + Thr161Gln + Gly203Pro + Thr210Pro + Leu216Val
Thr 65Gln + Ser131Asp + Ala186Gly + Thr215Ser + Leu216His
Thr132Gln + Asn162Ser + Gly165Asp + Tyr205Gly + Tyr208Cys
Gly127Gln + Gly153Glu + Gly203Pro + Thr215Gly + Thr219Gly
Ile106Met + Leu125Cys + Gly126Ser + Ser131Glu + Gly156Asn
Ser 97Glu + Thr132Gly + Pro200Gly + Tyr205Ala + Asn211Ser
Thr 65Asp + Tyr166Gln + Gly203Asn + Thr210Ser + Thr212Gln
Gly 99Ser + Tyr166Ser + Ser187Glu + Tyr205Gln + Leu216Ser
Gly126Ser + Gly130Asn + Thr161Asp + Phe188Val + Thr207Gln
Leu 95Pro + Pro200Ser + Tyr205Leu + Thr210Glu + Leu216Cys
Tyr103Glu + Gly127Gln + Tyr166Thr + Ala202His + Thr210Ser
Thr 58Gly + Ala202Pro + Pro209Gln + Thr212Ser + Ala214Glu
Gly 62Gln + Tyr103Thr + Thr163Gly + Ser187Asp + Gly203Gln
Gly101Gln + Leu125Glu + Thr132Ser + Gly203Ser + Thr212Gln
Asn 96Ser + Ala128His + Ala186Gln + Tyr205His + Thr215Ser
Gly101Ser + Gly165Asp + Thr210Ser + Thr212Gly + Thr215Gly
Gly156Gln + Ala202Gly + Ala214His + Thr215Pro + Leu216Thr
Val 94Cys + Gly 99Gln + Gly126Asn + Gly159Pro + Thr219Pro
Leu 95Pro + Gly105Pro + Thr161Asp + Gly203Pro + Leu216Asn
Gly 60Glu + Asn 61Gln + Thr 65Gln + Thr210Gly + Leu216Gln
Leu125Gly + Ser187Asp + Phe188Gln + Ala202Asn + Thr212Asn
Phe188Ile + Gly203Glu + Thr210Gly + Thr212Gln + Leu216Pro
Asp 59Glu + Leu 95Ile + Ile106Leu + Gly203Gln + Tyr213Ile
Gly 99Gln + Ala186Pro + Tyr205Gly + Thr215Glu + Gly218Gln
Ser 98Glu + Thr161Pro + Ala202Gln + Tyr205Leu + Thr207Gln
Ser100Glu + Gly153Asn + Thr163Gln + Thr212Pro + Leu216Gln
Leu 95Ser + Ile106Asp + Tyr205Val + Leu216Pro + Thr219Asn
Tyr166Val + Ser190Asp + Thr207Gly + Thr212Ser + Leu216Asn
Asp 59Glu + Gly105Gln + Thr132Gln + Ala186Ser + Tyr205Ile
Gly 60Pro + Tyr205Ile + Asn211Gln + Leu216Ile + Asn217Gln
Gly 99Asn + Leu125T r + Asn157Gln + Asn162Glu + Thr212Gln
Ile106Ala + Asn162Glu + Tyr166His + Asn211Ser + Thr215Gln
Asn154Asp + Phe188Gly + Gly203Pro + Tyr205Ala + Pro209Ser
Leu 95Asn + Gly105Glu + Tyr166Ala + Tyr205Leu + Thr212Ser
Val 94Ala + Ser131Glu + Thr132Glu + Phe188His + Thr212Gly
Gly 64Gln + Gly159Asp + Ser160Glu + Thr215Pro + Leu216Ala
Thr 58Glu + Asp 59Glu + Pro209Gln + Thr212Asn + Thr215Gln
Asn 96Asp + Ser 97Glu + Tyr205Thr + Ala214Thr + Leu216Ser
Asp 59Glu + Gly 60Glu + Gly159Asn + Thr210Pro + Gly218Asn
Asp 59Glu + Gly 60Asp + Leu 95Ser + Gly203Pro + Tyr205Ser
Tyr103Ile + Asn154Ser + Tyr205Glu + Ala214His + Thr215Asp
Gly105Asn + Gly127Gln + Ala202Glu + Gly203Ser + Asn217Glu
Asn 61Asp + Thr 65Gln + Ser 97Asp + Tyr205Ala + Leu216Ser
Asn 61Glu + Ser 97Glu + Gly105Gln + Asn162Ser + Thr212Ser
Asn 61Asp + Leu 95Ser + Ser 97Glu + Val204Gln + Thr212Gly
Tyr103His + Asn154Glu + Phe188Ile + Ser190Glu + Thr212Gln
Val 94Pro + Asn 96Asp + Ser 98Glu + Tyr103Pro + Thr212Ser
Val 94Met + Ser158Asp + Thr161Glu + Gly203Ser + Gly218Ser
Gly203Asp + Tyr213Pro + Thr215Asn + Asn217Asp + Gly218Asp
Ser160Glu + Asn162Glu + Tyr205Val + Asn211Gln + Thr215Pro
Asn 61Asp + Ser 98Glu + Tyr103Cys + Gly203Ser + Tyr213Ala
Gly101Asn + Tyr103Pro + Ile106Asn + Asn157Glu + Thr161Asp
Gly130Asn + Ala186Glu + Ala202Glu + Thr215Pro + Thr219Asn
Gly126Asp + Ala128Glu + Thr132Ser + Met198Cys + Gly218Ser
Val 94Pro + Leu125Gly + Thr215Asp + Leu216His + Asn217Asp
Gly 62Ser + Ile106Ala + Ala128Thr + Thr215Asp + Asn217Glu
Gly105Pro + Ser129Glu + Thr132Asn + Gly165Glu + Asn211Gln
Asp 59Glu + Asn 96Asp + Ser 97Glu + Phe188His + Thr210Pro
Asn 61Gln + Gly153Glu + Thr163Asp + Gly203Gln + Leu216Ser
Asp 59Glu + Thr 65Gly + Leu 95Gly + Ser 97Asp + Tyr166Gly
Asn157Asp + Ser160Asp + Asn162Glu + Thr210Gly + Thr212Ser
Ser 97Glu + Ser100Asp + Gly101Ser + Thr207Gln + Thr212Asn
Gly 64Ser + Gly203Ser + Thr212Asp + Ala214Asp + Thr215Asp
Gly 60Asn + Thr132Asn + Ser155Glu + Gly156Glu + Asn162Asp
Gly 99Ser + Tyr103Ile + Gly130Gln + Phe188Glu + Thr219Glu
Gly156Asp + Thr161Asp + Ser190Asp + Gly203Ser + Ala214Gln
Gly101Asp + Ser104Asp + Gly127Ser + Thr210Gln + Thr212Gln
Gly130Gln + Ser155Asp + Tyr213Met + Gly218Asp + Thr219Glu
Gly 99Ser + Asn154Ser + Gly203Asp + Leu216Asn + Gly218Asp
Ser102Glu + Gly130Asp + Ser131Asp + Ala202His + Thr210Asn
Gly153Pro + Gly159Asp + Asn162Asp + Ala186Gln + Ala202Gln
Asn 96Asp + Ser100Glu + Gly105Asp + Val204Thr + Thr215Pro
Asn162Asp + Gly165Asp + Thr212Gln + Thr215Ser + Leu216Cys
Ser155Asp + Gly165Ser + Ser187Glu + Tyr205His + Thr212Gly
Ser100Glu + Gly126Glu + Ala186Pro + Tyr205Asn + Tyr213Gln
Gly 62Pro + Ser104Glu + Ser131Asp + Thr212Gln + Tyr213Thr
Gly203Asp + Tyr205Asn + Asn211Gln + Thr212Gly + Ala214Asp
Ile106Cys + Asn154Glu + Ala186Glu + Gly203Ser + Leu216Ala
Thr 58Pro + Asn 61Asp + Val 94Glu + Asn 96Asp + Gly165Pro
Leu 95Glu + Ser 97Glu + Ser102G1u + Thr161Ser + Tyr205Leu
Thr 58Gly + Ala128Pro + Gly156Asp + Thr161Asp + Gly203Asn
Gly 60Ser + Gly126Pro + Thr161Glu + Ser190Glu + Thr212Gly
Gly 60Ser + Ser102Glu + Gly105Asp + Ser131Glu + Gly203Asn
Asn154Gln + Ala186Ser + Tyr213Glu + Leu216Glu + Thr219Pro
Gly101Asp + Ile106Cys + Gly126Asp + Met198Gln + Thr219Pro
Leu 95Gln + Asn154Asp + Ser190Asp + Thr212Gly + Asn217Glu
Thr 58Gln + Asn 61Glu + Ser100Asp + Tyr205Ala + Thr215Gly
Asn 61Glu + Gly 99Gln + Ser100Glu + Tyr103Met + Leu125Ser
Ala128Ser + Gly130Ser + Gly159Asp + Thr163Glu + Tyr20SCys
Asp 59Glu + Gly 99Asp + Tyr103Ile + Ile164Thr + Thr215Ser
Leu 95Ile + Ser102Asp + Ser131Asp + Ile164Val + Leu216Ile
Asn 96Gln + Thr212Pro + Ala214Asp + Leu216Glu + Thr219Glu
Ile106Glu + Ser131Asp + Ala202Gly + Tyr205Asn + Thr215Ser
Ile106Glu + Gly130Ser + Ser131Asp + Phe188Ala + Tyr205Pro
Thr 65Gln + Asn154Asp + Thr215Glu + Leu216His + Asn217Glu
Asn 61Gln + Gly 99Gln + Asn154Glu + Ser158Asp + Ser187Glu
Gly101Ser + Gly126Asn + Thr212Glu + Thr215Asp + Leu216Gln
Asp 59Glu + Thr 65Pro + Ser 98Asp + Tyr103Pro + Leu125Ile
Gly153Ser + Ser155Asp + Gly165Asp + Ser187Asp + Asn217Ser
Ser100Asp + Ser102Glu + Gly130Glu + Gly156Pro + Tyr205Ile
Gly105Ser + Gly153Glu + Thr215Asn + Leu216Asp + Asn217Glu
Thr 58Asp + Asn 96Glu + Ala186Gly + Gly203Pro + Thr219Gly
Asn154Asp + Ser160Asp + Gly165Ser + Ser190Asp + Ala202Ser
Asp 59Glu + Ser 98Glu + Tyr103Asn + Thr132Ser + Pro209Glu
Ala128Asp + Asn154Asp + Tyr166Ala + Pro200Gly + Tyr205Met
Gly101Gln + Asn157Gln + Gly203Asp + Thr215Gly + Thr219Asp
Asn 61Gln + Gly126Asp + Asn162Glu + Ser190Asp + Thr215Gly
Thr 58Gln + Asp 59Glu + Leu 95Gly + Gly203Pro + Asn211Glu
Ile106Asn + Leu125Val + Ser129Glu + Thr163Asp + Ala214Gly
Gly126Gln + Gly127Asp + Ala128Ser + Ser155Asp + Ser158Glu
Ser190Glu + Ala202Glu + Thr212Asn + Thr215Gly + Asn217Asp
Ser155Glu + Gly156Glu + Ala202Asp + Thr215Gly + Thr219Gln
Ser160Glu + Ser190Asp + Gly203Gln + Tyr205Asn + Thr212Asn
Gly156Asn + Tyr166Ala + Gly203Glu + Thr212Glu + Leu216Asp
Gly 99Gln + Ser102Glu + Gly126Asp + Asn154Glu + Tyr205Asn
Thr 65Pro + Gly127Glu + Ser131Glu + Ser155Asp + Tyr205Cys
Asn 96Gln + Thr161Gly + Ser190Glu + Thr210Glu + Asn211Asp
Gly130Asn + Ser187Glu + Pro209Gln + Ala214Glu + Thr215Glu
Gly101Glu + Ile106Met + Thr210Ser + Ala214Asp + Thr215Glu
Ser104Asp + Thr210Gly + Thr212Gln + Ala214Glu + Thr215Glu
Asn 96Gln + Ser158Asp + Gly159Asp + Thr163Asn + Leu216Glu
Asn 96Gln + Ser158Glu + Gly203Pro + Thr215Asp + Leu216Asp
Gly127Asn + Gly130Gln + Ser160Glu + Thr215Asp + Leu216Asp
Gly 60Asn + Asn 61Glu + Gly 62Glu + Asn157Asp + Tyr205Ser
Gly105Asn + Ala128Glu + Ser129Glu + Gly153Pro + Ala202Asp
Thr 58Asp + Tyr103Met + Gly153Ser + Asn157Glu + Ser158Glu
Gly 64Ser + Gly 99Asn + Ser129Asp + Gly130Asp + Asn157Asp
Asn154Gln + Ala202Asp + Asn211Glu + Thr212Asp + Leu216Ile
Gly101Asp + Ser102Glu + Gly159Glu + Ile164Cys + Asn217Gln
Gly 62Pro + Gly101Asp + Ser102Glu + Ser190Glu + Thr215Asn
Thr 58Glu + Asp 59Glu + Gly130Glu + Ala214Gln + Leu216Val
Asn 61Ser + Ser102Glu + Ser160Glu + Thr161Asp + Leu216Val
Ser102Asp + Ser160Glu + Thr161Asp + Tyr205Ser + Asn211Gln
Gly 62Pro + Ser104Glu + Gly105Asp + Leu125Met + Ala214Asp
Val 94Cys + Gly105Asp + Asn162Glu + Thr163Glu + Leu216Ile
Ser 97Asp + Ser 98Asp + Ala202Asp + Tyr205His + Leu216Thr
Gly 62Glu + Asn 96Glu + Gly105Asp + Tyr166Ser + Pro209Gly
Val 94Asp + Ala128Asp + Thr132Asn + Gly165Asp + Thr212Ser
Ser102Glu + Gly105Glu + Asn154Asp + Gly156Asn + Leu216Gly
Tyr103Glu + Gly130Gln + Ile164Leu + Thr210Glu + Thr212Asp
Asp 59Glu + Asn157Ser + Tyr166Met + Gly203Asp + Asn217Asp
Gly 60Gln + Gly130Asn + Ser155Glu + Thr215Glu + Asn217Asp
Val 94Thr + Gly 99Asp + Gly153Pro + Tyr205Glu + Ala214Glu
Gly101Asn + Ser131Asp + Asn154Gln + Gly203Asp + Thr2l5Glu
Gly101Pro + Gly127Pro + Asn157Glu + Gly203Asp + Thr215Asp
Thr 65Gln + Gly153Glu + Gly165Glu + Gly203Asp + Leu216Cys
Ser 98Asp + Ser100Glu + Ala128Gly + Gly159Asp + Gly165Pro
Ser 98Asp + Ser100Asp + Ser187Asp + Gly203Asn + Thr212Ser
Ser 98Glu + Ser100Glu + Ser155Asp + Asn162Ser + Thr210Asn
Asn 96Asp + Ser100Asp + Gly127Asn + Tyr205Cys + Thr219Asp
Asn 96Asp + Ser100Glu + Gly159Asp + Tyr205Gln + Ala214Pro
Leu 95Gln + Gly101Pro + Ser102Asp + Gly153Asp + Tyr166Glu
Asn 96Ser + Gly159Asp + Ser190Asp + Tyr205Ile + Ala214Gly
Thr 65Glu + Ser 97Glu + Gly101Glu + Ala186Pro + Pro200Gly
Gly 99Asn + Gly126Asp + Ser158Glu + Ser160Glu + Tyr205Gln
Gly 62Glu + Leu 95Glu + Ile106Leu + Thr212Glu + Ala214Ser
Asn 61Ser + Gly 99Pro + Ser104Asp + Gly203Asp + Tyr205Asp
Gly101Ser + Tyr103Ala + Gly203Gln + Tyr205Asp + Gly218Asp
Ser160Asp + Asn162Asp + Phe188Pro + Thr212Glu + Leu216Ser
Ile106Ser + Ser160Glu + Asn162Glu + Gly203Asp + Leu216Val
Val 94Asp + Gly126Asp + Gly130Pro + Thr212Ser + Leu216Asn
Thr 65Pro + Ser155Asp + Asn157Asp + Ala186Thr + Leu216Asp
Ser100Glu + Gly105Pro + Ala128Gly + Asn157Asp + Thr163Glu
Gly126Glu + Ala186Asp + Ala202Asp + Gly203Gln + Leu216Asn
Thr132Gly + Ser160Asp + Ala186Glu + Ala202Asp + Tyr205Gln
Tyr103Pro + Ser129Glu + Ala186Glu + Ala202Glu + Thr215Asn
Leu125Pro + Ser129Asp + Gly153Glu + Asn162Ser + Leu216Ile
Asp 59Glu + Thr 65Glu + Gly105Ser + Ser131Asp + Tyr213Ser
Gly126Asp + Gly156Glu + Ser158Glu + Ile164Met + Tyr205Met
Asn 96Glu + Tyr103Gly + Leu125Glu + Asn154Glu + Thr163Gln
Thr 58Glu + Ala128Ser + Gly156Asp + Thr163Asp + Thr210Gln
Gly 99Asp + Leu125Thr + Gly156Glu + Ser158Asp + Gly203Pro
Gly101Glu + Gly156Asp + Ser158Asp + Ala202Gln + Pro209Asn
Ser131Glu + Gly156Asp + Ser158Glu + Phe188Leu + Gly203Asn
Gly 62Ser + Ser129Glu + Asn157Gln + Gly165Asp + Thr215Glu
Ser 97Asp + Ser129Glu + Gly165Asp + Thr215Asn + Asn217Ser
Ala186Asp + Tyr205Cys + Thr212Asp + Ala214Asp + Thr215Ser
Val 94His + Ser158Glu + Thr212Asp + Ala214Glu + Thr215Gln
Leu125Glu + Gly159Pro + Ala186His + Thr212Asp + Ala214Glu
Asn 61Asp + Tyr103Gln + Gly105Gln + Leu125Asp + Pro209Asp
Gly 99Gln + Gly159Ser + Pro209Asp + Thr215Glu + Leu216Ala
Ser100Asp + Ser102Asp + Gly105Asn + Gly203Asp + Asn211Gln
Ser 97Glu + Ser100Glu + Gly165Glu + Phe188Val + Leu216Asn
Asn 96Ser + Leu125Asp + Ser129Glu + Tyr205Gln + Thr215Pro
Asn 96Glu + Asn157Asp + Gly165Gln + Ser187Asp + Tyr205Val
Leu 95Asp + Ser102Asp + Gly203Glu + Val204Ser + Leu216Ser
Gly 62Glu + Thr 65Gln + Asn157Asp + Asn162Asp + Thr215Gln
Ser131Glu + Gly165Asp + Leu216Cys + Asn217Gln + Thr219Asp
Val 94Asp + Ser187Asp + Ser190Glu + Tyr205Asn + Thr215Pro
Asp 59Glu + Asn 96Gln + Ser187Glu + Ser190Asp + Thr215Asn
Ser158Asp + Asn162Asp + Gly203Ser + Leu216Glu + Asn217Ser
Gly126Pro + Ser158Asp + Asn162Asp + Thr212Asp + Leu216Thr
Ser102Asp + Ser158Asp + Thr163Asp + Met198Gly + Thr207Pro
Gly 60Glu + Leu125Val + Ser155Glu + Gly165Glu + Val204Asn
Ser131Glu + Asn154Glu + Ser187Glu + Tyr213Val + Asn217Gln
Gly 62Asp + Ala128His + Asn154Glu + Ser187Glu + Gly203Ser
Asn 61Gln + Gly 62Pro + Gly105Asp + Gly203Glu + Gly218Asp
Ser102Asp + Gly203Glu + Ala214Gly + Thr215Pro + Gly218Asp
Asp 59Glu + Asn157Asp + Thr163Pro + Pro209Gly + Thr210Asp
Leu125Glu + Asn162Ser + Gly165Glu + Thr212Asp + Thr215Gln
Ile106Asn + Ser155Glu + Tyr166Val + Ser187Asp + Thr212Asp
Gly105Pro + Ser155Glu + Ser187Asp + Tyr205Pro + Thr215Asp
Asn 96Glu + Ser155Glu + Ser158Asp + Thr161Pro + Thr215Pro
Thr 58Asp + Thr 65Asp + Tyr103Val + Thr132Asp + Phe188Gly
Asp 59Glu + Asn 96Glu + Ser190Glu + Gly203Ser + Ala214Ser
Ser155Glu + Ile164Ser + Ala202Asp + Tyr205Asp + Leu216Pro
Tyr103Ala + Ser104Asp + Ser131Asp + Ser160Asp + Tyr213Leu
Thr 58Glu + Asn 61Asp + Ala186Gly + Ser187Asp + Tyr205Ala
Asn 96Glu + Ser102Glu + Gly153Asp + Gly203Pro + Asn217Gln
Asn 96Glu + Ser102Asp + Gly105Pro + Asn162Glu + Tyr213Val
Ser129Asp + Gly153Asn + Gly203Glu + Ala214Glu + Thr215Gln
Ser100Glu + Gly127Glu + Gly203Glu + Tyr205Ala + Tyr213Thr
Ser158Asp + Ser190Asp + Gly203Asp + Thr212Pro + Thr215Gly
Leu 95Ile + Thr132Gly + Ser187Asp + Tyr213Asp + Leu216Glu
Gly 60Glu + Gly 62Pro + Ala128Glu + Ser155Glu + Ala202Ser
Thr 58Asp + Ser 97Asp + Gly153Ser + Gly159Glu + Ala214Thr
Asn 61Asp + Ser100Glu + Gly126Asn + Ser129Asp + Thr219Ser
Asp 59Glu + Leu 95Asp + Phe188Glu + Pro209Ser + Gly218Ser
Tyr103Glu + Ser187Glu + Thr212Ser + Thr215Gln + Gly218Asp
Ser104Glu + Ser190Asp + Tyr205Asn + Tyr208Val + Gly218Asp
Ala128Asp + Ser131Glu + Thr161Gln + Tyr205Glu + Leu216Met
Ser102Asp + Ala128Glu + Ile164Gly + Gly203Asn + Thr212Ser
Gly 62Asp + Gly105Glu + Tyr208Ala + Thr210Ser + Thr212Asp
Ser 97Glu + Ser102Glu + Ile106Ser + Asn157Ser + Leu216Gly
Gly 62Asn + Leu 95Met + Ser 97Asp + Ser102Asp + Asn217Ser
Ser102Glu + Leu125Cys + Ser131Asp + Ala202Ser + Asn217Glu
Ser102Asp + Ser131Glu + Thr163Asp + Thr207Gly + Thr215Asn
Ser 97Asp + Ile106Thr + Ala186Asp + Gly203Asp + Thr215Gly
Thr 58Asp + Gly127Asp + Ala128Thr + Ser155Asp + Gly203Ser
Val 94Gly + Gly127Asp + Ser155Asp + Gly156Gln + Tyr205Glu
Ile106Glu + Gly126Asp + Gly127Asn + Gly156Glu + Ala202Asn
Ser 97Asp + Gly203Pro + Asn211Ser + Thr215Asp + Gly218Glu
Gly 99Asp + Ser104Asp + Ile106Val + Thr132Asp + Thr215Asn
Gly 62Glu + Ser100Glu + Gly127Pro + Tyr205Ile + Thr212Gln
Gly 99Glu + Gly126Asp + Ala186Glu + Gly203Ser + Asn217Ser
Thr132Asp + Tyr205Glu + Pro209Gln + Tnr212Glu + Asn217Ser
Ser 97Asp + Gly127Gln + Ala186Asn + Thr212Glu + Thr215Asp
Ser102Asp + Asn154Gln + Thr212Asp + Thr215Asp + Leu216Gly
Leu125Gln + Ser129Asp + Asn162Gln + Thr212Glu + Thr215Asp
Asp 59Glu + Ser 98Glu + Leu125Gly + Ala128Asn + Ser129Asp
Asp 59Glu + Ser 98Asp + Gly156Asn + Ala202Asp + Gly218Asn
Asn 61Ser + Asn157Glu + Gly203Asp + Tyr205His + Thr219Asp
Ile106Asp + Gly130Asp + Tyr166Ile + Ala186Ser + Thr212Asp
Asp 59Glu + Gly156Asp + Thr161Asn + Leu216Ala + Gly218Asp
Thr 58Asp + Asn 96Glu + Ser131Asp + Phe188Val + Thr215Gly
Leu 95Val + Ser155Glu + Tyr166Ala + Ala186Glu + Thr210Asp
Ser 98Asp + Leu125Cys + Ala128Asp + Thr161Gly + Asn162Glu
Asn 96Gln + Ile106Ala + Gly156Glu + Ser160Asp + Thr210Glu
Asp 59Glu + Tyr166Glu + Tyr205Asn + Thr210Gly + Asn211Glu
Asp 59Glu + Gly153Glu + Pro209Ser + Asn211Asp + Thr212Pro
Asp 59Glu + Tyr103Gly + Gly153Glu + Ser187Glu + Asn211Gln
Asp 59Glu + Thr163Ser + Ser187Glu + Val204Thr + Asn217Glu
Ser104Glu + Ile106Ala + Gly156Asp + Gly165Asp + Tyr205Asn
Gly126Glu + Ser131Asp + Asn154Ser + Tyr205Ser + Thr215Asp
Asp 59Glu + Gly101Gln + Thr212Glu + Ala214Pro + Leu216Asp
Asn154Ser + Ile164Cys + Thr212Asn + Ala214Asp + Gly218Glu
Thr 65Glu + Ile106Asp + Gly127Asn + Thr215Asp + Leu216His
Gly 60Gln + Gly 99Asp + Gly127Glu + Tyr205Glu + Thr215Asn
Thr 58Gln + Gly 60Glu + Ile106Val + Tyr205Glu + Asn211Glu
Gly130Pro + Gly159Glu + Phe188Leu + Ser190Glu + Thr212Glu
Asn 61Glu + Gly127Asp + Ser190Glu + Tyr205Ala + Leu216Ser
Ser187Glu + Pro209Asp + Thr212Asn + Ala214Ser + Thr215Asp
Ser155Glu + Tyr166Glu + Gly203Glu + Val204Pro + Thr215Ser
Ser 98Asp + Ser102Glu + Ala202Ser + Thr215Glu + Leu216Gln
Val 94Asn + Gly101Asn + Leu125Glu + Ser129Glu + Thr212Asp
Ser129Asp + Ser155Asp + Ala202His + Gly203Asn + Asn211Ser
Asn 96Asp + Tyr103His + Gly105Asn + Ala202Glu + Ala214Glu
Leu 95His + Gly 99Gln + Ser190Glu + Ala202Asp + Thr210Asn
Gly101Asp + Ala128Glu + Thr132Glu + Gly156Asn + Tyr166Ile
Gly105Asp + Leu125Asp + Ala202Pro + Gly203Glu + Leu216Gln
Gly105Asp + Leu125Glu + Gly203Glu + Thr212Gly + Leu216Cys
Gly 62Glu + Val 94Ala + Ser129Asp + Tyr205Asp + Thr212Ser
Asn 61Asp + G1y101Glu + Tyr205Glu + Thr215Pro + Leu216His
Ser104Asp + Leu125Glu + Ser190Glu + Asn211Gln + Leu216Asn
Gly105Asn + Gly127Asp + Ala128Gly + Ser158Asp + Thr219Glu
Ser 97Asp + Ser155Glu + Ser160Asp + Asn211Gln + Thr215Ser
Asp 59Glu + Gly 64Gln + Gly130Glu + Gly153Glu + Tyr205Asn
Asn157Asp + Tyr205Leu + Thr212Glu + Thr215Gly + Leu216Glu
Thr 58Glu + Gly101Glu + Gly105Pro + Gly130Ser + Tyr166Asp
Ser 97Glu + Gly159Asp + Tyr166Ala + Ser187Glu + Leu216Val
Gly101Asp + Ser131Glu + Thr212Gln + Thr215Asp + Leu216Ile
Gly153Asn + Asn157Asp + Tyr213Leu + Asn217Ser + Thr219Asp
Gly153Asp + Ser158Glu + Tyr205Gly + Leu216Val + Gly218Ser
Gly 99Ser + Ser187Asp + Gly203Glu + Pro209Asp + Asn211Gln
Leu 95Ala + Ser102Asp + Ser187Glu + Gly203Asp + Thr219Gln
Gly 62Asp + Ala128Thr + Ser187Glu + Gly203Glu + Asn211Ser
Gly101Asp + Ala128Thr + Asn154Gln + Phe188Asp + Leu216Glu
表36
多环六突变变异体
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洗涤组合物
在本发明的另一实施方案中，有效量的一种或多种酶变异体包括在用于洗涤各种需要除去蛋白质污垢的表面的组合物中。这种洗涤组合物包括用于洗涤硬表面的非限制形式洗涤组合物(例如，液体或颗粒)；用于洗涤织物的非限制形式的洗涤组合物(颗粒、液体或条状制剂)；餐具洗涤组合物(非限制形式)；非限制形式的口腔用洗涤组合物(例如，洁牙剂、牙膏和漱口剂制剂)；非限制形式的假牙洗涤组合物(例如，液体、片剂)：和非限制形式的接触透镜洗涤组合物(例如，液体、片剂)。
除了本文上述的卡氏枯草杆菌蛋白酶变异体之外，洗涤组合物还包括与蛋白酶相容的一种或多种洗涤组合物材料。本文使用的术语“洗涤组合物材料”指的是选择用于所需的洗涤组合物的具体类型和产物形式(例如，液体、颗粒、条状、喷雾剂、棒状、膏状、凝胶)的任何液体、固体或气态材料，该材料还应与组合物中使用的卡氏枯草杆菌蛋白酶变异体相容。通过考虑所洗涤的表面材料、在使用过程中用于洗涤条件下所需的组合物的形式(例如通过洗涤剂使用)很容易地具体选择洗涤组合物材料。本文中使用的术语“相容”指的是洗涤组合物材料不降低卡氏枯草杆菌蛋白酶变异体的蛋白水解活性至在正常使用场合根据需要蛋白酶失效的程度。具体的洗涤组合物材料在下文详细举例说明。
本文中使用的“有效量的酶变异体”指的是为获得在具体洗涤组合物中所需的酶活性而必需的酶变异体的量。这种有效量易于被本领域的技术人员确定，其基于许多因素，例如所用的具体的酶变异体、洗涤应用、洗涤组合物的具体组成，和是否需要液体或无水(例如颗粒、条状)组合物等等。洗涤组合物优选含有约0.0001％-约10％一种或多种本发明的酶变异体，更优选约0.001％-约1％，还优选约0.01％-约0.1％。其中使用酶变异体的各种洗涤组合物的若干实例在下文中进一步地讨论。本文使用的所有份数、百分数和比率均为按重量计，除非另有说明。
本文中使用的“非织物洗涤组合物”包括硬表面洗涤组合物、餐具洗涤组合物、口腔洗涤组合物、假牙洗涤组合物和接触透镜洗涤组合物。A.用于硬表面、餐具和织物的洗涤组合物
本发明的酶变异体可用于其中需要高泡和良好不溶底物去除的各种洗涤组合物中。因此，酶变异体可与各种常规成分一起使用以提供全配制的硬表面洗涤剂。餐具洗涤组合物、织物洗涤组合物等等。这种组合物可以是液体、颗粒、条状形式等等。这种组合物可配制成含有多达按表面活性剂重量计30％-60％的现代“浓缩”洗涤剂。
本发明的洗涤组合物可选择性地，并优选地含有各种阴离子、非离子、两性离子等等的表面活性剂。这些表面活性剂通常以组合物的约5％-约35％的含量存在。
用于本发明的表面活性剂的非限制性实例包括常规的C11-C18烷基苯磺酸盐和伯和无规烷基硫酸盐、式为CH3(CH2)x(CHOSO3c-M+)CH3和CH3(CH2)y(CHOSO3 -M+)CH2CH3的C10-C18仲(2，3)烷基硫酸盐，其中x和(y+1)是至少约7，优选至少约9的整数，M是水溶性阳离子，尤其是钠、C10-C18烷基烷氧基硫酸盐(尤其是EO1-5乙氧基硫酸盐)、C10-C18烷基烷氧基羧酸盐(尤其是EO1-5乙氧基羧酸盐)、C10-C18烷基聚糖苷和它们相应的硫酸化聚糖苷，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甜菜碱和磺基甜菜碱(“sultaines”)、C10-C18胺氧化物等等。本发明优选烷基烷氧基硫酸盐(AES)和烷基烷氧基羧酸盐(AEC)。(根据配制者的需要，使用这种表面活性剂与上述的胺氧化物和/或甜菜碱或磺基甜菜碱表面活性剂的混合物也是优选的)。其它常规使用的表面活性剂在标准文献中列出。尤其有用的表面活性剂包括在1993年3月16日颁布的Connor等人的US5194639中的C10-C18 N-甲基葡糖酰胺，列为本文参考文献。
用于洗涤组合物中的各种各样的其它成分可包括在本发明的组合物中，其包括其它活性成分、载体、增溶剂、加工助剂、染料或颜料、用于液体配方的溶剂等等。如果需要附加地增加泡沫，在组合物中，通常以约1％-约10％的含量加入发泡剂，例如C10-C16烷醇酰胺。C10-C14单乙醇和二乙醇酰胺说明了这种发泡剂的典型种类。与上述的高泡辅助表面活性剂，例如胺氧化物、甜菜碱和磺基甜菜碱一起使用这类发泡剂也是有利的。如果需要，溶解的镁盐，例如氯化镁、硫酸镁等等可以通常约0.1％-约2％的含量加入以提供附加的起泡。
本发明的液体洗涤组合物可含有作为载体的水和其它溶剂。低分子量的伯或仲醇，例如甲醇、乙醇、丙醇和异丙醇是适宜的。单羟基醇对于表面活性剂的增溶是优选的，但多元醇，例如含有约2-约6个碳原子和约2-约6个羟基的多元醇(例如，1，3-丙二醇、乙二醇、甘油和1，2-丙二醇)也可以使用。组合物可含有约5％-约90％，典型地约10％-约50％的这种载体。
本发明的洗涤组合物将优选配制成在使用过程以含水洗涤操作，洗涤水将具有约6.8-约11.0的pH，最终的产物因此通常在该范围内配制。控制pH在所需要的范围内的技术包括使用缓冲剂、碱、酸等等，是本领域技术人员已知的。
在配制本发明的硬表面洗涤组合物和织物洗涤组合物时，配制者可以按重量计约5％-约50％的含量使用各种助剂。典型的助剂包括1-10微米沸石、多羧酸盐，例如柠檬酸盐和氧联二琥珀酸盐、层状硅酸盐、磷酸盐等等。其它常规的助剂在标准配方中列出。
同样，配制者可在组合物中，通常以按重量计约0.001％-1％的含量使用各种其它的酶，例如纤维素酶、脂酶、淀粉酶和蛋白酶。在洗衣洗涤剂领域中已知各种洗涤和织物护理酶。
各种漂白化合物，例如过碳酸盐、过硼酸盐等等可通常以按重量计约1％-约15％的含量用于该组合物中。如果需要，该组合物还可以含有漂白活性剂，例如四乙酰基乙二胺、壬酰氧基苯磺酸盐等等，它们也是现有技术中已知的。使用含量通常为按重量计约1％-约10％。
各种污垢解脱剂，尤其是阴离子低聚酯类型、各种螯合剂，尤其是氨基膦酸盐和乙二胺二琥珀酸盐、各种粘土去污剂，尤其是乙氧基化四亚乙五胺、各种分散剂，尤其是聚丙烯酸酯和聚天冬氨酸酯、各种增白剂，尤其是阴离子增白剂、各种泡沫抑制剂，尤其硅烷和仲醇、各种织物柔软剂，尤其是绿土等等部可以按重量计约1％-约35％的含量用于组合物。在标准配方和公开的所得中有此常规物质的详细描述。
在洗涤组合物中也可使用酶稳定剂。酶稳定剂包括丙二醇(优选约1％-约10％)、甲酸钠(优选约0.1％-约1％)和甲酸钙(优选约0.1％-约1％)。1.硬表面洗涤组合物
本文使用的“硬表面洗涤组合物”指的是用于洗涤硬表面，例如地板、墙壁、浴室瓷砖等等的液体和颗粒洗涤组合物。本发明的硬表面洗涤组合物含有有效量的一种或多种本发明的酶变异体，按组合物活性酶重量计优选为约0.001％-约10％，更优选约0.01％-约5％，还优选约0.05％-约1％。除了含有一种或多种酶变异体之外，硬表面洗涤组合物通常含有表面活性剂和水溶性螯合剂。然而，在某些特殊的产物，例如喷雾玻璃清洁剂中，有时是不使用表面活性剂的，因为它们会产生残留在玻璃表面的膜状/条纹残余物。
当存在时，表面活性剂组分可含有少至组合物的0.1％，但通常组合物将含有约0.25％-约10％，更优选约1％-约5％的表面活性剂。
组合物通常含有约0.5％-约50％的洗涤剂助剂，优选为约1％-约10％。
pH值优选在约8-12的范围内。如果需要进行调节，可使用常规的pH调节剂，例如氢氧化钠、碳酸钠或盐酸。
在组合物中可包括溶剂。有用的溶剂包括，但不限制于，乙二醇醚，例如二乙二醇单己基醚、二乙二醇单丁基醚、乙二醇单丁基醚、乙二醇单己基醚、丙二醇单丁基醚、二丙二醇单丁基醚，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。当使用时，溶剂通常以约0.5％-约15％，优选约3％-约11％的含量存在。
此外，当组合物以“完全强度”应用于表面后表面不进行漂洗时，在本发明的组合物中可使用高挥发性溶剂，例如异丙醇或乙醇，以有助于组合物从表面快速蒸发。当使用时，挥发性溶剂通常以约2％-约12％的含量存在于组合物中。
本发明的硬表面洗涤组合物的实施方案通过如下实施例说明。
实施例7-12
液体硬表面洗涤组合物
实施 例组分 7 8 9 10 11 12Thr58Asn 0.05 0.50 0.02 0.03 0.10 0.03Thr58Gln+Asn61Glu - - - - 0.20 0.02Na2DIDA*EDTA** - - 2.90 2.90 - -柠檬酸钠 - - - - 2.90 2.90C12烷基苯磺酸钠 1.95 - 1.95 - 1.95 -C12烷基硫酸钠 - 2.20 - 2.20 - 2.20C12(乙氧基)***硫酸钠 - 2.20 - 2.20 - 2.20C12二甲基胺氧化物 - 0.50 - 0.50 - 0.50枯烯磺酸钠 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-亚氨基二乙酸二钠**乙二胺二乙酸四钠***二乙二醇单己基醚****所有配方调节到pH7
在实施例7-10中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体代替Thr58Asn，得到基本上类似的结果。
在实施例11-12中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体的任何组合代替Thr58Asn和Thr58Gln+Asn6lGlu，得到基本上类似的结果。
实施例13-18
用于清洁硬表面和除去家庭霉变的喷雾组合物
实施例组分 13 14 15 16 17 18Val94Ala 0.50 0.05 0.60 0.30 0.20 0.30Leu125Asn+Gly126Pro+Ser129Glu - - - - 0.30 0.10辛基硫酸钠 2.00 2.00 2.00 2.00 2.00 2.00十二烷基硫酸钠 4.00 4.00 4.00 4.00 4.00 4.00氢氧化钠 0.80 0.80 0.80 0.80 0.80 0.80硅酸盐(钠) 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％产物pH为约7。
在实施例13-16中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体代替Val94Ala，得到基本上类似的结果。
在实施例17-18中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体的任何组合代替Val94Ala和Leu125Asn+Gly126Pro+Ser129Glu，得到基本上类似的结果。2.餐具洗涤组合物
在本发明的另一实施方案中，餐具洗涤组合物含有一种或多种本发明的酶变异体。本文使用的“餐具洗涤组合物”指的是用于洗涤餐具的组合物的所有形式，其包括，但不限制于，颗粒和液体形式。本发明的餐具洗涤组合物的实施方案由如下实施例说明。
实施例19-24
餐具洗涤组合物
实施例组分 19 20 21 22 23 24Gly153Asn 0.05 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)硫酸盐 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)羧酸盐 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枯烯磺酸钠 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++(作为氯化镁) 0.20 0.20 0.20 0.20 0.20 0.20Ca++(作为氯化钙) 0.40 0.40 0.40 0.40 0.40 0.40水 平衡至100％产物pH调节至7。
在实施例19-22中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体代替Gly153Asn，得到基本上类似的结果。
在实施例23-24中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体的任何组合代替Gly153Asn和Ala186asp+Phe188Gln，得到基本上类似的结果。3.织物洗涤组合物
在本发明的另一实施方案中，织物洗涤组合物含有一种或多种本发明的酶变异体。本文使用的“织物洗涤组合物”指的是用于洗涤织物的组合物的所有形式，其包括，但不限制于，颗粒、液体和条状形式。优选的织物洗涤组合物是液体形式。a.颗粒织物洗涤组合物
本发明的颗粒织物洗涤组合物含有有效量的一种或多种本发明的酶变异体，按组合物活性酶重量计优选为约0.001％-约10％，更优选约0.005％-约5％，还优选约0.01％-约1％。除了含有一种或多种酶变异体之外，颗粒织物洗涤组合物通常含有至少一种表面活性剂、一种或多种助剂和，在某些情况下，漂白剂。
本发明的颗粒织物洗涤组合物的实施方案由如下实施例说明。
实施例25-28
颗粒织物洗涤组合物
实施例组分 25 26 27 28Pro200Asn 0.10 0.20 0.03 0.05Gly203Asp+Tyr205Ala+Tyr208Ser - - 0.02 0.05C13直链烷基苯磺酸盐 22.00 22.00 22.00 22.00磷酸盐(作为三聚磷酸钠) 23.00 23.00 23.00 23.00碳酸钠 23.00 23.00 23.00 23.00硅酸钠 14.00 14.00 14.00 14.00沸石 8.20 8.20 8.20 8.20螯合剂(二亚乙三胺五乙酸) 0.40 0.40 0.40 0.40硫酸钠 5.50 5.50 5.50 5.50水 平衡至100％
在实施例25-26中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体代替Pro200Asn，得到基本上类似的结果。
在实施例27-28中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体的任何组合代替Pro200Asn和Gly203Asp+Tyr205Ala+Tyr208Ser，得到基本上类似的结果。
实施例29-32
颗粒织物洗涤组合物
实施例组分 29 30 31 32Pro200Asn+Ala202Asn+Vla204 0.10 0.20 0.03 0.05Gly+Asn211Asp+Thr219GlyLeu95Gly+Gly203Pro - - 0.02 0.05C12烷基苯磺酸盐 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)烷基硫酸钠 5.00 5.00 5.00 5.00柠檬酸钠 5.00 5.00 5.00 5.00荧光增白剂 0.10 0.10 0.10 0.10硫酸钠 17.00 17.00 17.00 17.00水和少量组分 平衡至100％
在实施例29-30中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体代替Pro200Asn+Ala202Asn+Vla204Gly+Asn211Asp+Thr219Gly，得到基本上类似的结果。
在实施例31-32中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体的任何组合代替Pro200Asn+Ala202Asn+Vla204Gly+Asn211Asp+Thr219Gly和Leu95Gly+Gly203Pro，得到基本上类似的结果。
实施例33-36
颗粒织物洗涤组合物
实施例组分 33 34 35 36Thr58Gln+Asp59Glu+Leu216Gln 0.10 0.20 0.03 0.05Pro200Asn+Ala202Gln+Tyr208Cys+Thr210 - - 0.02 0.05Pro+Tyr213Asn+Ala214Asn+Thr219SerC13直链烷基苯磺酸盐 22.00 22.00 22.00 22.00磷酸盐(作为三聚磷酸钠) 23.00 23.00 23.00 23.00碳酸钠 23.00 23.00 23.00 23.00硅酸钠 14.00 14.00 14.00 14.00沸石 8.20 8.20 8.20 8.20螯合剂(二亚乙三胺五乙酸) 0.40 0.40 0.40 0.40硫酸钠 5.50 5.50 5.50 5.50水 平衡至100％
在实施例33-34中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体代替Thr58Gln+Asp59Glu+Leu216Gln，得到基本上类似的结果。
在实施例35-36中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体的任何组合代替Thr58Gln+Asp59Glu+Leu216Gln和Pro200Asn+Ala202Gln+Tyr208Cys+Thr210Pro+Tyr213Asn+Ala214Asn+Thr219Ser，得到基本上类似的结果。
实施例37-40
颗粒织物洗涤组合物
实施例组分 37 38 39 40Asp59Glu+Gly60Pro+ 0.10 0.20 0.03 0.05Gly62Asn+Gly64GlLeu125Ala - - 0.02 0.05C12烷基苯磺酸盐 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)烷基硫酸钠 5.00 5.00 5.00 5.00柠檬酸钠 5.00 5.00 5.00 5.00荧光增白剂 0.10 0.10 0.10 0.10硫酸钠 17.00 17.00 17.00 17.00水和少量组分 平衡至100％
在实施例37-38中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体代替Asp59Glu+Gly60Pro+Gly62Asn+Gly64Gl，得到基本上类似的结果。
在实施例39-40中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体的任何组合代替Asp59Glu+Gly60Pro+Gly62Asn+G1y64Gl和Leu125Ala，得到基本上类似的结果。
实施例41-42
颗粒织物洗涤组合物
实施例组分 41 42直链烷基苯磺酸盐 11.4 10.70动物脂烷基硫酸盐 1.80 2.40C14-15烷基硫酸盐 3.00 3.10C14-15醇7倍乙氧基化 4.00 4.00动物脂醇11倍乙氧基化 1.80 1.80分散剂 0.07 0.1硅氧烷液体 0.80 0.80柠檬酸三钠 14.00 15.00柠檬酸 3.00 2.50沸石 32.50 32.10马来酸丙烯酸共聚物 5.00 5.00二亚乙三胺五亚甲基膦酸 1.00 0.20Gly127Glu+Asn162Glu+Phe218Met+ 0.30 0.30Ala202Gly+Gly203Ser+Gly218Asn脂酶 0.36 0.40淀粉酶 0.30 0.30硅酸钠 2.00 2.50硫酸钠 3.50 5.20聚乙烯吡咯烷酮 0.30 0.50过硼酸盐 0.5 1苯酚磺酸盐 0.1 0.2过氧化酶 0.1 0.1少量组分 至100 至100
实施例43-44
颗粒织物洗涤组合物
实施例组分 43 44直链C12烷基苯磺酸钠 6.5 8.0硫酸钠 15.0 18.0沸石A 26.0 22.0次氮基三乙酸钠 5.0 5.0聚乙烯吡咯烷酮 0.5 0.7四乙酰基乙二胺 3.0 3.0硼酸 4.0 -过硼酸盐 0.5 1苯酚磺酸盐 0.1 0.2Gly218Gln+Thr219Gln 0.4 0.4填料(例如，硅酸盐；碳酸盐；香料；水) 至100 至100
实施例45
压型颗粒织物洗涤组合物组分 重量％烷基硫酸盐 8.0烷基乙氧基硫酸盐 2.0C25和C45醇3和7倍乙氧基化的混合物 6.0多羟基脂肪酸酰胺 2.5沸石 17.0层状硅酸盐/柠檬酸盐 16.0碳酸盐 7.0马来酸丙烯酸共聚物 5.0污垢解脱聚合物 0.4羧甲基纤维素 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纤维素酶 0.2四乙酰基乙二胺 6.0过碳酸盐 22.0乙二胺二琥珀酸 0.3泡沫抑制剂 3.54，4’-双(2-吗啉基-4-苯胺基-s-三嗪-6-基氨基)芪-2，2’- 0.25二磺酸二钠4，4’-双(2-磺基苯乙烯基)联苯二钠 0.05水、香料和少量组分 至100
实施例46
颗粒织物洗涤组合物组分 重量％直链烷基苯磺酸盐 7.6C16-C18烷基硫酸盐 1.3C14-15醇7倍乙氧基化物 4.0椰子烷基二甲基羟基乙基铵氯化物 1.4分散剂 0.07硅氧烷液体 0.8柠檬酸三钠 5.0沸石4A 15.0马来酸丙烯酸共聚物 4.0二亚乙三胺五亚甲基膦酸 0.4过硼酸盐 15.0四乙酰基乙二胺 5.0绿土 10.0聚(环氧乙烷)(MW300000) 0.3Thr58Pro+Gly64Ser+Thr65Gly 0.4脂酶 0.2淀粉酶 0.3纤维素酶 0.2硅酸钠 3.0碳酸钠 10.0羧甲基纤维素 0.2增白剂 0.2水、香料和少量组分 至100
实施例47
颗粒织物洗涤组合物组分 重量％直链烷基苯磺酸盐 6.92动物脂烷基硫酸盐 2.05C14-15醇7倍乙氧基化物 4.4C12-15烷基乙氧基硫酸盐-3倍乙氧基化 0.16沸石 20.2柠檬酸盐 5.5碳酸盐 15.4硅酸盐 3.4马来酸丙烯酸共聚物 4.0羧甲基纤维素酶 0.31污垢解脱聚合物 0.30Val94Gln+Ser100Glu 0.2脂酶 0.36纤维素酶 0.13过硼酸盐四水合物 11.64过硼酸盐单水合物 8.7四乙酰基乙二胺 5.0二亚乙三胺五甲基膦酸 0.38硫酸镁 0.40增白剂 0.19香料、硅氧烷、泡沫抑制剂 0.85少量组分 至100b.液体织物洗涤组合物
本发明的液体织物洗涤组合物含有有效量的一种或多种本发明的酶变异体，按组合物活性酶重量计优选为约0.005％-约5％，更优选为约0.01％-约1％。该液体织物洗涤组合物通常还含有一种阴离子表面活性剂、脂肪酸、水溶性洗涤剂助剂和水。
本发明的液体织物洗涤组合物的实施方案由如下实施例说明。
实施例48-52
液体织物洗涤组合物
实施例组分 48 49 50 51 52Leu125Ala+Gly126Glu 0.05 0.03 0.30 0.03 0.10Gly153Glu+Ile164Ala - - - 0.01 0.20C12-C14烷基硫酸钠 20.00 20.00 20.00 20.00 20.002-丁基辛酸 5.00 5.00 5.00 5.00 5.00柠檬酸钠 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单乙醇胺 2.50 2.50 2.50 2.50 2.50水/丙二醇/乙醇(100∶1∶1) 平衡至100％
在实施例48-50中，用表3-36所列的卡氏枯草杆菌蛋白酶变异体代替Leu125Ala+Gly126Glu，得到基本上类似的结果。
在实施例51-52中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体的任何组合代替Leu125Ala+Gly126Glu和Gly153Glu+Ile164Ala，得到基本上类似的结果。
实施例53-57
液体织物洗涤组合物
实施例组分 53 54 55 56 57Phe188Ser 0.05 0.03 0.30 0.03 0.10Ala186Gln+Ser187Glu - - - 0.01 0.20+Phe188AspC12-C14烷基硫酸钠 20.00 20.00 20.00 20.00 20.002-丁基辛酸 5.00 5.00 5.00 5.00 5.00柠檬酸钠 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单乙醇胺 2.50 2.50 2.50 2.50 2.50水/丙二醇/乙醇(100∶1∶1) 平衡至100％
在实施例53-55中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体代替Phe188Ser，得到基本上类似的结果。
在实施例56-57中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体的任何组合代替Phe188Ser和Ala186Gln+Ser187Glu+Phe188Asp，得到基本上类似的结果。
实施例58-59
颗粒织物洗涤组合物
实施例组分 58 59C12-14烯基琥珀酸 3.0 8.0柠檬酸单水合物 10.0 15.0C12-15烷基硫酸钠 8.0 8.0C12-15醇2倍乙氧基化硫酸钠 - 3.0C12-15醇7倍乙氧基化物 - 8.0C12-15醇5倍乙氧基化物 8.0 -二亚乙三胺五(亚甲基膦酸) 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泡沫抑制剂 0.15 0.15氢氧化钠 至pH 7.5过硼酸盐 0.5 1苯酚磺酸盐 0.1 0.2过氧化酶 0.4 0.1水和少量组分 至10 0份
在实施例58和59中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体代替Thr58Ser+Ser100Asp+Ile164Met+Tyr205Ala，得到基本上类似的结果。
实施例60-62
液体织物洗涤组合物
实施例组分 60 61 62柠檬酸 7.10 3.00 3.00脂肪酸 2.00 - 2.00乙醇 1.93 3.20 3.20硼酸 2.22 3.50 3.50单乙醇胺 0.71 1.09 1.091，2-丙二醇 7.89 8.00 8.00枯烯磺酸钠 1.80 3.00 3.00甲酸钠 0.08 0.08 0.08氢氧化钠 6.70 3.80 3.80含硅消泡剂 1.16 1.18 1.18Asn61Glu 0.0145 - -Tyr103Cys+Gly156Glu+Ala202Gln - 0.0145 -+Thr212Gly+Leu216GlnGly105Asn+Gly126Ser+Ser160Glu - - 0.0145+Thr161Asn+Tyr205Val+Gly218Asn脂酶 0.200 0.200 0.200纤维素酶 - 7.50 7.50污垢解脱聚合物 0.29 0.15 0.15消泡剂 0.06 0.085 0.085增白剂36 0.095 - -增白剂3 - 0.05 0.05C12烷基苯磺酸 9.86 - -C12-15烷基多乙氧基化(2.5)硫酸盐 13.80 18.00 18.00C12葡糖酰胺 - 5.00 5.00C12-13烷基聚乙氧基化物(9) 2.00 2.00 2.00水、香料和少量组分 平衡至 100％c.条状织物洗涤组合物
本发明的适合于手洗脏织物的条状织物洗涤组合物含有有效量的一种或多种本发明的酶变异体，按组合物活性酶重量计优选为约0.001％-约10％，更优选为约0.01％-约1％。
本发明的条状织物洗涤组合物的实施方案由如下实施例说明。
实施例63-66
条状织物洗涤组合物
实施例组分 63 64 65 66Thr207Glu 0.3 - 0.1 0.02Leu125Ile+Ser131Glu - - 0.4 0.03C12-C16烷基硫酸钠 20.0 20.0 20.0 20.00C12-C14N-甲基葡糖酰胺 5.0 5.0 5.0 5.00C11-C13烷基苯磺酸钠 10.0 10.0 10.0 10.00碳酸钠 25.0 25.0 25.0 25.00焦磷酸钠 7.0 7.0 7.0 7.00三聚磷酸钠 7.0 7.0 7.0 7.00沸石A(0.1-10μ) 5.0 5.0 5.0 5.00羧甲基纤维素 0.2 0.2 0.2 0.20聚丙烯酸酯(MW1400) 0.2 0.2 0.2 0.20椰子烷基单乙醇胺 5.0 5.0 5.0 5.00增白剂、香料 0.2 0.2 0.2 0.20硫酸钙 1.0 1.0 1.0 1.00硫酸镁 1.0 1.0 1.0 1.00水 4.0 4.0 4.0 4.00填料* 平衡至 100％*可以选自常规材料，例如碳酸钙、滑石、粘土、硅酸盐等等。
在实施例63-64中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体代替Thr207Glu，得到基本上类似的结果。
在实施例65-66中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体的任何组合代替Thr207Glu和Leu125Ile+Ser131Glu，得到基本上类似的结果。
实施例67-70
条状织物洗涤组合物
实施例组分 67 68 69 70Pr0200Ser+Ala214Gly+Asn217Glu 0.3 - 0.1 0.02Gly101Asp+Ala128Asp+Ser155Asp+ - 0.3 0.4 0.03Ser187Asp+Thr207Gly+Thr212GlnC12-C16烷基硫酸钠 20.0 20.0 20.0 20.0C12-C14N-甲基葡糖酰胺 5.0 5.0 5.0 5.00C11-C13烷基苯磺酸钠 10.0 10.0 10.0 10.00碳酸钠 25.0 25.0 25.0 25.00焦磷酸钠 7.0 7.0 7.0 7.00三聚磷酸钠 7.0 7.0 7.0 7.00沸石A(0.1-10μ) 5.0 5.0 5.0 5.00羧甲基纤维素 0.2 0.2 0.2 0.20聚丙烯酸酯(MW1400) 0.2 0.2 0.2 0.20椰子烷基单乙醇胺 5.0 5.0 5.0 5.00增白剂、香料 0.2 0.2 0.2 0.20硫酸钙 1.0 1.0 1.0 1.00硫酸镁 1.0 1.0 1.0 1.00水 4.0 4.0 4.0 4.00填料* 平衡至100％*可以选自常规材料，例如碳酸钙、滑石、粘土、硅酸盐等等。
在实施例67中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体代替Pro200Ser+Ala214Gly+Asn217Glu，得到基本上类似的结果。
在实施例68中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体代替Pro200Ser+Ala214Gly+Asn217Glu和Gly101Asp+Ala128Asp+Ser155Asp+Ser187Asp+Thr207Gly+Thr212Gln，得到基本上类似的结果。
在实施例69-70中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体的任何组合代替Va1203Glu和Gly100Glu+Ilel07Ser，得到基本上类似的结果。B.其它洗涤组合物
除了上述硬表面洗涤、餐具洗涤和织物洗涤组合物之外，本发明的一种或多种酶变异体可加入各种需要水解不溶解底物的其它洗涤组合物中。该附加的洗涤组合物包括，但不限制于，口腔洗涤组合物、假牙洗涤组合物和接触透镜洗涤组合物。1.口腔洗涤组合物
在本发明的另一实施方案中，在组合物中包含一种或多种可药用量的本发明的酶变异体以从牙齿或假牙中除去蛋白质污垢。本文使用的“口腔洗涤组合物”指的是洁齿粉、牙膏、牙凝胶、牙粉、漱口剂、口腔喷雾剂、口腔凝胶、咀嚼胶、锭剂、香囊、片剂、生物凝胶、预防膏、假牙处理溶液等等。按组合物的重量计，洗涤组合物优选含有约0.0001-约20％，更优选约0.001％-约10％，还优选约0.01％-约5％的一种或多种本发明的酶变异体和可药用的载体。本文使用的“可药用的”指的是该术语描述的药物、药剂或惰性成分适合于与人体和低级动物的组织接触，而没有不适宜的毒性、不适性、不稳定性、刺激性、过敏反应等等，与有益效果/风险比率相当。
口腔洗涤组合物的可药用的口腔洗涤载体组分按组合物重量计通常占约50％-约99.99％，优选约65％-约99.99％，更优选约65％-约99％。
可包括在本发明的口腔洗涤组合物中的可药用的载体组分和选择性的组分是本领域技术人员已知的。用于口腔洗涤组合物的各种组合物类型、载体组分和选择性组分在1992年3月17日颁布的Seibel的US5096700；1991年7月2日颁布的Sampathkumar的US5028414和1991年7月2日颁布的Benedict、Bush和Sunberg的US5028415中，列为本文参考文献。
本发明的口腔洗涤组合物的实施方案在如下实施例中说明。
实施例71-74
洁牙粉组合物
实施例组分 71 72 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氟化钠 0.243 0.243 0.243 0.243二氧化钛 0.500 0.500 0.500 0.500糖精钠 0.286 0.286 0.286 0.286烷基硫酸钠(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。
在实施例71-74中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体代替Thy212Gln，得到基本上类似的结果。
实施例75-78
漱口剂组合物
实施例组分 75 76 77 78Val204Asn+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乳化剂 0.08 0.08 0.08 0.08氟化钠 0.05 0.05 0.05 0.05甘油 10.00 10.00 10.00 10.00增甜剂 0.02 0.02 0.02 0.02苯甲酸 0.05 0.05 0.05 0.05氢氧化钠 0.20 0.20 0.20 0.20染料 0.04 0.04 0.04 0.04水 平衡至 100％
在实施例75-78中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体代替Val204Asn+Thr215Asn，得到基本上类似的结果。
实施例79-82
锭剂组合物
实施例组分 79 80 81 82Thr65Asp+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增甜剂 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玉米糖浆 平衡至100％
在实施例79-82中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体代替Thr65Asp+Thr132Glu+Asn154Asp+Thr212Glu+Leu216Gly+Asn217Ser，得到基本上类似的结果。
实施例83-86
口香糖组合物
实施例组分 83 84 85 86Tyr103Ala 0.03 0.02 0.10 0.05山梨糖醇结晶 38.44 38.40 38.40 38.40Palija-T胶基料* 20.00 20.00 20.00 20.00山梨糖醇(70％水溶液) 22.00 22.00 22.00 22.00甘露糖醇 10.00 10.00 10.00 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。
在实施例83-86中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体代替Tyr103A1a，得到基本上类似的结果。2.假牙洗涤组合物
在本发明的另一实施方案中，用于在口腔外洗涤假牙的假牙洗涤组合物含有一种或多种本发明的酶变异体。该假牙洗涤组合物含有有效量的一种或多种酶变异体，优选按组合物重量计，约0.0001-约50％，更优选约0.001％-约35％，还优选约0.01％-约20％的一种或多种本发明的酶变异体，和假牙洗涤载体。各种假牙洗涤组合物配方，如泡腾片等等是现有技术中已知的(参见例如Young的US5055305，列为本文参考文献)，通常适合于加入一种或多种酶变异体以便从假牙除去蛋白质污垢。
本发明的假牙洗涤组合物的实施方案由如下实施例说明。
实施例87-90
双层泡腾假牙洗涤片
实施例组分 87 88 89 90酸性层Leu216Pro 1.0 1.5 0.01 0.05酒石酸 24.0 24.0 24.00 24.00碳酸钠 4.0 4.0 4.00 4.00氨基磺酸 10.0 10.0 10.00 10.00PEG20000 4.0 4.0 4.00 4.00碳酸氢钠 24.5 24.5 24.50 24.50过硫酸钾 15.0 15.0 15.00 15.00酸性焦磷酸钠 7.0 7.0 7.00 7.00热解氧化硅 2.0 2.0 2.00 2.00TAED* 7.0 7.0 7.00 7.00蓖麻酰基磺基琥珀酸盐 0.5 0.5 0.50 0.50香料 1.0 1.0 1.00 1.00碱性层过硼酸钠单水合物 32.0 32.0 32.00 32.00碳酸氢钠 19.0 19.0 19.00 19.00EDTA 3.0 3.0 3.00 3.00三聚磷酸钠 12.0 12.0 12.00 12.00PEG20000 2.0 2.0 2.00 2.0过硫酸钾 26.0 26.0 26.00 26.00碳酸钠 2.0 2.0 2.00 2.00热解氧化硅 2.0 2.0 2.00 2.00染料/香料 2.0 2.0 2.00 2.00*四乙酰基乙二胺
在实施例87-90中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体代替Leu216Pro，得到基本上类似的结果。2.接触透镜洗涤组合物
在本发明的另一实施方案中，接触透镜洗涤组合物含有一种或多种本发明的酶变异体。该接触透镜洗涤组合物含有有效量的一种或多种酶变异体，优选按组合物重量计，约0.01-约50％，更优选约0.01％-约20％，还优选约1％-约5％的一种或多种本发明的酶变异体，和接触透镜洗涤载体。各种接触透镜洗涤组合物配方，例如片剂、液体等等在现有技术中是已知的(参见例如1989年9月5日授权的Davies，Meaken和Rees，US4863627；1988年5月24日再次授权的Huth，Lam和Kirai，US Re32672；有1986年9月2日授权的Schafer，US4609493；1987年9月1日授权的Ogunbiyi和Smith，US4690793；1986年9月30日授权的Ogunbiyi，Riedhammer和Smiht，US4614549；和1981年8月25日颁布的Ogata，US4285738；均列为本文参考文献)，通常适合于加入本发明的一种或多种酶变异体以便从接触透镜除去蛋白质污垢。
本发明的接触透镜洗涤组合物的实施方案由如下实施例说明。
实施例91-94
酶解接触透镜洗涤溶液
实施例组分 91 92 93 94Ala199Thr+Val204Gln 0.01 0.5 0.1 2.0葡萄糖 50.00 50.0 50.0 50.0非离子表面活性剂(聚氧乙烯- 2.00 2.0 2.0 2.0聚氧丙烯共聚物)阴离子表面活性剂(聚氧乙烯- 1.00 1.0 1.0 1.0烷基苯基醚硫酸酯钠)氯化钠 1.00 1.0 1.0 1.0硼砂 0.30 0.3 0.3 0.3水 平衡至100％
在实施例91-94中，用表3-36中所列的卡氏枯草杆菌蛋白酶变异体代替Ala199Thr+Val204Gln，得到基本上类似的结果。
尽管描述了本发明的具体实施方案，但显然本领域的技术人员可作出本发明的各种改变和改进而不会违背本发明的精神和范围。在所附的权利要求中，本发明的范围覆盖了所有这些改进。
序列表(1)一般信息：
(i)申请人：BRODE，PHILIPF。
BARNETT，BOBBYL。
RUBINGH，DONNN。
(ii)发明项目：SUBTILISIN CARLSBERG VARIANTS HANING
DECREASED ASDORPTION AND INCREASED
HYDROLYSIS(iii)序列数：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)国家：USA
(F)邮区：45061(v)计算机可读形式：
(A)介质类型：软盘
(B)计算机：IBM PC兼容
(C)操作系统：PC-DOS/MS-DOS
(D)软件：Patentln Release#1.0，Version#1.03(vi)当前申请资料：
(A)申请号：
(B)申请日：
(C)分类：(viii)代理人/事务所信息：
(A)姓名：CORSTANJE，BRAHMJ。
(B)登记号：34，804
(C)参考/存档号：5602(ix)电讯信息：
(A)电话：(513)627-2858
(B)电传：(513)627-0260(2)SEQ ID NO：1的信息：(i)序列特征：
(A)长度：274氨基酸
(B)类型：氨基酸
(C)链型：单
(D)拓扑学：线性
(xi)序列描述：SEQID NO：1：(xi) SEQUENCE DESCRIPTION：SEQ ID NO：1：Ala Gln Thr Val Pro Tyr Gly Ile Pro Leu Ile Lys Ala Asp Lys Val1 5 10 15Gln Ala Gln Gly Phe Lys Gly Ala Asn Val Lys Val Ala Val Leu Asp
20 25 30Thr Gly Ile Gln Ala Ser His Pro Asp Leu Asn Val Val Gly Gly Ala
35 40 45Ser Phe Val Ala Gly Glu Ala Tyr Asn Thr Asp Gly Asn Gly His Gly50 55 60Thr His Val Ala Gly Thr Val Ala Ala Leu Asp Asn Thr Thr Gly Val65 70 75 80Leu Gly Val Ala Pro Ser Val Ser Leu Tyr Ala Val Lys Val Leu Asn
85 90 95Ser Ser Gly Ser Gly Ser Tyr Ser Gly Ile Val Ser Gly Ile Glu Trp
100 105 110Ala Thr Thr Asn Gly Met Asp Val Ile Asn Met Ser Leu Gly Gly Ala
115 120 125Ser Gly Ser Thr Ala Met Lys Gln Ala Val Asp Asn Ala Tyr Ala Arg130 135 140Gly 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 Val
165 170 175Gly Ala Val Asp Ser Asn Ser Asn Arg Ala Ser Phe Ser Ser Val Gly
180 185 190Ala Glu Leu Glu Val Met Ala Pro Gly Ala Gly Val Tyr Ser Thr Tyr
195 200 205Pro Thr Asn Thr Tyr Ala Thr Leu Asn Gly Thr Ser Met Ala Ser Pro210 215 220His Val Ala Gly Ala Ala Ala Leu Ile Leu Ser Lys His Pro Asn Leu225 230 235 240Ser Ala Ser Gln Val Arg Asn Arg Leu Ser Ser Thr Ala Thr Tyr Leu
245 250 255Gly Ser Ser Phe Tyr Tyr Gly Lys Gly Leu Ile Asn Val Glu Ala Ala
260 265 270Ala Gln
Claims (9)
1.具有卡氏枯草杆菌蛋白酶野生型氨基酸序列的修饰氨基酸序列的卡氏枯草杆菌蛋白酶变异体，野生型氨基酸序列包括第一环区域、第二环区域、第三环区域、第四环区域、第五环区域和第六环区域；其中修饰的氨基酸序列在一个或多个环区域内的一个或多个位点上含有取代基；其中
A.当取代发生在第一环区域中时，取代发生在位点58、59、60、61、62、64或65的一个或多个上；其中
a.当取代发生在位点58时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；
b.当取代发生在位点59时，取代氨基酸是Glu；
c.当取代发生在位点60时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；
d.当取代发生在位点61时，取代氨基酸是Asp、Gln、Glu或Ser；
e.当取代发生在位点62时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；
f.当取代发生在位点64时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；和
g.当取代发生在位点65时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；
B.当取代发生在第二环区域中时，取代发生在位点94、95、96、97、98、99、100、101、102、103、104、105或106的一个或多个上；其中
a.当取代发生在位点94时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Met、Pro、Ser或Thr；
b.当取代发生在位点95时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Met、Pro、Ser、Thr或Val；
c.当取代发生在位点96时，取代氨基酸是Asp、Gln、Glu、Ser；
d.当取代发生在位点97时，取代氨基酸是Asp或Glu；
e.当取代发生在位点98时，取代氨基酸是Asp或Glu；
f.当取代发生在位点99时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；
g.当取代发生在位点100时，取代氨基酸是Asp或Glu；
h.当取代发生在位点101时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；
i.当取代发生在位点102时，取代氨基酸是Asp或Glu；
j.当取代发生在位点103时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val；
k.当取代发生在位点104时，取代氨基酸是Asp或Glu；
l.当取代发生在位点105时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；和
m.当取代发生在位点106时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Leu、Met、Pro、Ser、Thr或Val；
C.当取代发生在第三环区域中时，取代发生位点125、126、127、128、129、130、131或132的一个或多个上；其中
a.当取代发生在位点125时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Met、Pro、Ser、Thr或Val；
b.当取代发生在位点126时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；
c.当取代发生在位点127时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；
d.当取代发生在位点128时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、His、Pro、Ser或Thr；
e.当取代发生在位点129时，取代氨基酸是Asp或Glu；
f.当取代发生在位点130时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；
g.当取代发生在位点131时，取代氨基酸是Asp或Glu；和
h.当取代发生在位点132时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；
D.当取代发生在第四环区域中时，取代发生在位点153、154、155、156、157、158、159、160、161、162、163、164、165或166的一个或多个上；其中
a.当取代发生在位点153时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；
b.当取代发生在位点154时，取代氨基酸是Asp、Gln、Glu或Ser；
c.当取代发生在位点155时，取代氨基酸是Asp或Glu；
d.当取代发生在位点156时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；
e.当取代发生在位点157时，取代氨基酸是Asp、Gln、Glu、Ser；
f.当取代发生在位点158时，取代氨基酸是Asp或Glu；
g.当取代发生在位点159时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；
h.当取代发生在位点160时，取代氨基酸是Asp或Glu；
i.当取代发生在位点161时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；
j.当取代发生在位点162时，取代氨基酸是Asp、Gln、Glu或Ser；
k.当取代发生在位点163时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；
l.当取代发生在位点164时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Met、Pro、Ser、Thr或Val；
m.当取代发生在位点165时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；和
n.当取代发生在位点166时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val；和
E.当取代发生在第五环区域中时，取代发生在位点186、187、188、189或190一个或多个上；其中
a.当取代发生在位点186时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、His、Pro、Ser或Thr；
b.当取代发生在位点187时，取代氨基酸是Asp或Glu；
c.当取代发生在位点188时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr、Tyr或Val；
d.当取代发生在位点189时，取代氨基酸是Asp或Glu；
e.当取代发生在位点190时，取代氨基酸是Asn或Glu；和
F.当取代发生在第六环区域中时，取代发生在位点199、200、201、202、203、204、205、206、207、208、209、210、211、212、213、214、215、216、217、218或219的一个或多个上；其中
a.当取代发生在位点199时，取代氨基酸是Asn、Asp、gln、Glu、Gly、His、Pro、Ser或Thr；
b.当取代发生在位点200时，取代氨基酸是Asn、Asp、Gln、Glu、Gly或Ser；
c.当取代发生在位点201时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；
d.当取代发生在位点202时，取代氨基酸是Asp、Asp、Gln、Glu、Gly、His、Pro、Ser或Thr；
e.当取代发生在位点203时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；
f.当取代发生在位点204时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Met、Pro、Ser或Thr；
g.当取代发生在位点205时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val；
h.当取代发生在位点206时，取代氨基酸是Asp或Glu；
i.当取代发生在位点207时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；
j.当取代发生在位点208时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val；
k.当取代发生在位点209时，取代氨基酸是Asn、Asp、Gln、Glu、Gly或Ser；
l.当取代发生在位点210时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；
m.当取代发生在位点211时，取代氨基酸是Asp、Gln、Glu或Ser；
n.当取代发生在位点212时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；
o.当取代发生在位点213时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Leu、Met、Pro、Ser、Thr或Val；
p.当取代发生在位点214时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、His、Pro、Ser或Thr；
q.当取代发生在位点215时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；
r.当取代发生在位点216时，取代氨基酸是Ala、Asn、Asp、Cys、Gln、Glu、Gly、His、Ile、Met、Pro、Ser、Thr或Val；
s.当取代发生在位点217时，取代氨基酸是Asp、Gln、Glu或Ser；
t.当取代发生在位点218时，取代氨基酸是Asn、Asp、Gln、Glu、Pro或Ser；和
u.当取代发生在位点219时，取代氨基酸是Asn、Asp、Gln、Glu、Gly、Pro或Ser；
从而，与野生型卡氏枯草杆菌蛋白酶相比较，卡氏枯草杆菌蛋白酶变异体对不溶解底物具有降低的吸附作用和增加的水解作用。
2.权利要求1的卡氏枯草杆菌蛋白酶变异体，其中一个或多个取代发生在第一环区域。
3.权利要求1的卡氏枯草杆菌蛋白酶变异体，其中一个或多个取代发生在第二环区域。
4.权利要求1的卡氏枯草杆菌蛋白酶变异体，其中一个或多个取代发生在第三环区域。
5.权利要求1的卡氏枯草杆菌蛋白酶变异体，其中一个或多个取代发生在第四环区域。
6.权利要求1的卡氏枯草杆菌蛋白酶变异体，其中一个或多个取代发生在第五环区域。
7.权利要求1的卡氏枯草杆菌蛋白酶变异体，其中一个或多个取代发生在第六环区域。
8.一种选自硬表面洗涤组合物、餐具洗涤组合物、口腔洗涤组合物、假牙洗涤组合物、接触透镜洗涤组合物和织物洗涤组合物的洗涤组合物，其特征在于其中洗涤组合物含有权利要求1-7的任何一项的卡氏枯草杆菌蛋白酶变异体和洗涤组合物载体；洗涤组合物优选是硬表面洗涤组合物或洗涤组合物是织物洗涤组合物；组合物优选含有按组合物重量计至少约5％的表面活性剂和至少约5％助剂；组合物还优选含有选自溶剂、缓冲剂、酶、污垢释放剂、粘土去污剂、分散剂、增白剂、泡沫抑制剂、织物柔软剂、增泡剂、酶稳定剂、漂白剂、染料、香料和它们的混合物的洗涤组合物材料。
9.编码权利要求1-7的任何一项的卡氏枯草杆菌蛋白酶变异体的突变体卡氏枯草杆菌蛋白酶基因。
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| US08/400,698 | 1995-03-08 | ||
| US08/400,698 US6455295B1 (en) | 1995-03-08 | 1995-03-08 | Subtilisin Carlsberg variants having decreased adsorption and increased hydrolysis |
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| Publication Number | Publication Date |
|---|---|
| CN1183119A true CN1183119A (zh) | 1998-05-27 |
Family
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| Application Number | Title | Priority Date | Filing Date |
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| CN96193612A Pending CN1183119A (zh) | 1995-03-08 | 1996-02-14 | 具有降低的吸附作用和增加的水解作用的卡氏枯草杆菌蛋白酶变异体 |
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| EP (1) | EP0813596A2 (zh) |
| JP (1) | JPH11501812A (zh) |
| CN (1) | CN1183119A (zh) |
| AU (1) | AU4868896A (zh) |
| BR (1) | BR9607356A (zh) |
| CA (1) | CA2214348C (zh) |
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| MA (1) | MA23818A1 (zh) |
| MX (1) | MX9706851A (zh) |
| WO (1) | WO1996028566A2 (zh) |
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-
1995
- 1995-03-08 US US08/400,698 patent/US6455295B1/en not_active Expired - Fee Related
-
1996
- 1996-02-14 CZ CZ972793A patent/CZ279397A3/cs unknown
- 1996-02-14 BR BR9607356A patent/BR9607356A/pt not_active Application Discontinuation
- 1996-02-14 HU HU9800765A patent/HUP9800765A3/hu unknown
- 1996-02-14 MX MX9706851A patent/MX9706851A/es unknown
- 1996-02-14 JP JP8527598A patent/JPH11501812A/ja active Pending
- 1996-02-14 WO PCT/US1996/002168 patent/WO1996028566A2/en not_active Application Discontinuation
- 1996-02-14 CN CN96193612A patent/CN1183119A/zh active Pending
- 1996-02-14 AU AU48688/96A patent/AU4868896A/en not_active Abandoned
- 1996-02-14 CA CA002214348A patent/CA2214348C/en not_active Expired - Fee Related
- 1996-02-14 EP EP96904645A patent/EP0813596A2/en not_active Withdrawn
- 1996-02-26 IL IL11726696A patent/IL117266A0/xx unknown
- 1996-02-29 ZA ZA961631A patent/ZA961631B/xx unknown
- 1996-03-07 MA MA24175A patent/MA23818A1/fr unknown
Also Published As
| Publication number | Publication date |
|---|---|
| HUP9800765A2 (hu) | 1998-07-28 |
| WO1996028566A2 (en) | 1996-09-19 |
| EP0813596A2 (en) | 1997-12-29 |
| US6455295B1 (en) | 2002-09-24 |
| BR9607356A (pt) | 1999-06-15 |
| ZA961631B (en) | 1996-09-06 |
| IL117266A0 (en) | 1996-06-18 |
| HUP9800765A3 (en) | 2001-01-29 |
| MA23818A1 (fr) | 1996-10-01 |
| WO1996028566A3 (en) | 1997-07-10 |
| AU4868896A (en) | 1996-10-02 |
| CZ279397A3 (en) | 1997-11-12 |
| JPH11501812A (ja) | 1999-02-16 |
| CA2214348A1 (en) | 1996-09-19 |
| CA2214348C (en) | 2001-10-30 |
| MX9706851A (es) | 1997-11-29 |
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