CN105555951A - 包含脂解酶变体的组合物和方法 - Google Patents

包含脂解酶变体的组合物和方法 Download PDF

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CN105555951A
CN105555951A CN201480051103.9A CN201480051103A CN105555951A CN 105555951 A CN105555951 A CN 105555951A CN 201480051103 A CN201480051103 A CN 201480051103A CN 105555951 A CN105555951 A CN 105555951A
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lipolytic enzyme
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T·P·格雷卡尔
A·J·保罗斯
D·A·埃斯特尔
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Abstract

本发明提供了脂解酶变体。具体地讲，本发明提供了与亲本脂解酶相比具有两个、三个或更多个修饰，且具有至少一种改善的性质的脂解酶变体。此外，本发明提供了包含本发明脂解酶变体的组合物。本发明还提供了使用包含本发明脂解酶变体的组合物进行清洁的方法。

Description

包含脂解酶变体的组合物和方法
相关专利申请的交叉引用
本申请要求于2013年7月19日提交的美国临时申请USSN61/856,524的优先权权益，该申请全文以引用方式并入本文。
背景技术
包括脂肪酶和角质酶在内的脂解酶已被应用于洗涤剂清洁组合物中以去除油性污渍。脂解酶发挥作用的一个机制是将甘油三酯水解而产生脂肪酸。然而，这些酶常常受存在于清洁组合物中的表面活性剂和其他组分的抑制，从而干扰其去除油性污渍的能力。因此，需要具有改进功能的脂解酶，这些酶能够在清洁组合物的苛刻环境中起作用。
发明内容
本发明提供了改善的脂解酶，尤其是可用于洗涤剂组合物的酶。具体地讲，本发明提供了与亲本脂解酶相比具有两个或更多个修饰(例如置换)的脂解酶变体，该脂解酶变体具有改善的脂解活性，例如改善的丁酸对硝基苯酯、辛酸对硝基苯酯或棕榈酸对硝基苯酯的水解，增加的pNPP/pNPB或pNPO/pNPB比活性比值，增加的pNPB/pNPP比活性比值，增加的热稳定性，增加的洗涤剂稳定性，增加的LAS稳定性，增加的织物粘附性，降低的织物粘附性或增加的清洁性能。这一改善的活性可提高变体酶在洗涤循环中的有效性。本发明提供了特别适用于并可用于多种清洁应用的变体脂解酶，包括但不限于变体脂肪酶的脂解酶。本发明还提供了使用本发明的脂解酶变体进行清洁的方法。
在一些实施方案中，本发明是脂解酶变体或其活性片段，该脂解酶变体或其活性片段包含对亲本脂解酶的至少两、三、四、五、六、七、八、九或十或更多个氨基酸修饰，其中第一氨基酸修饰位于所述脂解酶变体中选自以下的位置：第4、11、18、23、24、27、29、33、45、48、49、51、56、58、73、75、77、90、91、94、111、130、137、154、156、163、187、189、227、233、252、256和264位，其中该变体的氨基酸位置是根据与SEQIDNO:1中示出的疏棉状嗜热丝孢菌(Thermomyceslanuginosa)脂肪酶的氨基酸序列的对应性进行编号的。
在一些实施方案中，本发明是脂解酶变体或其活性片段，其中该变体或其活性片段包含表2-2中的氨基酸修饰，其中该变体的氨基酸位置是根据与SEQIDNO:1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。
在一些上述实施方案中，该变体或活性片段具有脂解活性。在一些上述实施方案中，与亲本脂解酶相比，该变体或活性片段水解丁酸对硝基苯酯、辛酸对硝基苯酯或棕榈酸对硝基苯酯的性能指数(pI)大于1.0，在一些情况下，所述性能指数是使用实施例1的丁酸对硝基苯酯、辛酸对硝基苯酯或棕榈酸对硝基苯酯测定法测得的。在一些上述实施方案中，该变体或活性片段具有增加的pNPP/pNPB或pNPO/pNPB比活性比值、增加的pNPB/pNPP比活性比值、增加的热稳定性、增加的洗涤剂稳定性、增加的LAS稳定性、增加的织物粘附性、降低的织物粘附性或增加的清洁性能。
在一些实施方案中，本发明是包含至少一种上文所列的脂解酶变体的组合物。该组合物可以是清洁组合物或清洁添加剂。在一些实施方案中，本发明还包括附加的酶，所述附加的酶选自蛋白酶、半纤维素酶、纤维素酶、过氧化物酶、脂解酶、金属脂解酶(metallolipolyticenzyme)、木聚糖酶、脂肪酶、磷脂酶、酯酶、过水解酶、角质酶、果胶酶、果胶酸裂解酶、甘露聚糖酶、角蛋白酶、还原酶、氧化酶、酚氧化酶、脂氧合酶、木质素酶、支链淀粉酶、鞣酸酶、戊聚糖酶、麦拉宁酶(malanase)、β-葡聚糖酶、阿拉伯糖苷酶、透明质酸酶、软骨素酶、漆酶和淀粉酶。
在一个实施方案中，本发明是水解脂肪酸酯或甘油三酯的方法，该方法包括使脂肪酸酯或甘油三酯与上文所列的脂解酶变体接触。在一个实施方案中，本发明是清洁方法，该方法包括使表面或物品与包含至少一种上文所列的脂解酶变体的清洁组合物接触。
发明详述
本发明提供了改善的脂解酶，尤其是可用于洗涤剂组合物的酶。具体地讲，本发明提供了与亲本脂解酶相比具有两个或更多个修饰(例如置换)的脂解酶变体，该脂解酶变体具有改善的脂解活性，例如改善的脂肪酸酯或甘油三酯的水解，或例如改善的丁酸对硝基苯酯、辛酸对硝基苯酯或棕榈酸对硝基苯酯的水解。本发明提供了特别适用于并可用于多种清洁应用的变体脂解酶，包括但不限于变体脂肪酶的脂解酶。本发明包括了包含至少一种本文所述的变体脂解酶(例如，变体脂肪酶)的组合物。一些这种组合物包含洗涤剂组合物。本发明提供了嗜热丝孢菌属(Thermomyces)物种变体脂解酶和包含一种或多种这种变体脂肪酶的组合物。本发明的脂解酶变体可以与可用于洗涤剂组合物中的其他酶组合使用。本发明还提供了使用本发明的脂解酶变体进行清洁的方法。
本发明包括了脂解酶的酶变体，该酶变体具有两个或更多个对亲本脂解酶的修饰。亲本脂解酶可以是野生型酶，也可以是所述变体脂解酶所源自的任何起始参考脂解酶。
另外，本发明在可用于洗涤剂组合物的脂解酶中的2、3或更多个氨基酸位置处提供了修饰(例如置换)，其中在所述位置的有利修饰的结果是与亲本脂解酶(例如，SEQIDNO:1)或与参考脂肪酶(例如，SEQIDNO:2)相比，脂解活性的性能指数(pI)提高。就对亲本脂解酶的组合修饰而言，可认为这些氨基酸位置是有用的位置。
除非本文另有定义，否则本文使用的所有技术和科学术语的含义与本发明所属技术领域的普通技术人员一般理解的含义相同。尽管与本文所述方法和材料类似或等同的许多方法和材料也可以用于本发明的实践，但是本文描述了一些方法和材料。因此，对于下文即将定义的术语，在说明书整体上可以获得更加全面的描述。藉此将本文提及的所有专利、专利申请、文章和出版物，包括上下文中的，都明确以引用的方式并入本文。
此外，如本文所用，单数术语“一个”、“一种”、“该”和“所述”包括复数涵义，除非上下文中清楚地另有指明。除非另有指明，否则核酸以5′至3′方向从左至右书写；氨基酸序列以氨基至羧基方向从左至右书写。应当理解，本发明不限于所描述的具体方法、方案和试剂，因为这些方面可根据本领域技术人员使用它们的背景而有所变化。
预期在整个本说明书中给定的每一最大数值限值包括每一较低数值限值，就如同此类较低数值限值在本文中明确写出一样。在整个本说明书中给定的每一最小数值限值包括每一较高数值限值，就如同此类较高数值限值在本文中明确写出一样。在整个本说明书中给出的每一数值范围将包括落在此种较宽数值范围内的每一较窄数值范围，就如同此类较窄数值范围在本文中明确写出一样。
“蛋白质”或“多肽”包含氨基酸残基的聚合序列。术语“蛋白质”和“多肽”在本文中互换使用。遵照国际纯化学及应用化学联合会-国际生物化学联合会的生物化学命名委员会(IUPAC-IUBJointCommissiononBiochemicalNomenclature，JCBN))所定义的氨基酸的单字母和三字母代码在整个本公开中使用。还应理解，由于遗传密码的简并性，多肽可由不止一种核苷酸序列编码。突变按以下方式命名：亲本氨基酸的单字母代码后跟数字，然后是变体氨基酸的单字母代码。例如，第87位处的甘氨酸(G)突变为丝氨酸(S)可表示为“G087S”或“G87S”。多重突变可通过在突变之间插入“-”、“+”或“，”来表示。例如，在第87位和第90位处发生的突变可表示为“G087S-A090Y”或“G87S-A90Y”或者“G87S+A90Y”或“G087S+A090Y”。
术语“源自”和“获得自”不仅指由所考虑的生物株产生的脂解酶或可由所考虑的生物株产生的脂解酶，也指由分离自该株的DNA序列编码且在含有该DNA序列的宿主生物体中产生的脂解酶。另外该术语指由合成和/或cDNA起源的DNA序列编码的并且具有所考虑的脂解酶的鉴别特性的脂解酶。例如，“源自疏棉状嗜热丝孢菌的脂解酶”是指由疏棉状嗜热丝孢菌天然产生的具有脂解活性的那些酶，以及类似于疏棉状嗜热丝孢菌来源产生的那些酶的脂解酶(但其使用基因工程技术，由用编码该脂解酶的核酸转化的非疏棉状嗜热丝孢菌生物产生)。
如本文所用，“％同一性或百分比同一性”是指序列相似性。百分比同一性可用本领域已知的标准技术来测定(参见例如，Smith和Waterman，Adv.Appl.Math.2:482[1981]；Needleman和Wunsch，J.Mol.Biol.48:443[1970]；Pearson和Lipman，Proc.Natl.Acad.Sci.USA85:2444[1988]；在威斯康星遗传学软件包(WisconsinGeneticsSoftwarePackage)中的软件程序诸如GAP、BESTFIT、FASTA和TFASTA(遗传计算机公司(GeneticsComputerGroup，Madison，WI)；以及Devereux等人，Nucl.AcidRes.12:387-395[1984])。可用算法的一个例子是PILEUP。PILEUP利用渐进式成对序列比对法，由一组相关的序列产生多序列比对。其还能够绘制显示聚类关系(用于产生比对)的树形图。PILEUP使用Feng和Doolittle的简化渐进式比对方法(参见，Feng和Doolittle，J.Mol.Evol.35:351-360)。该方法类似于Higgins和Sharp所描述的方法(参见，Higgins和Sharp，CABIOS5:151-153)。可用的PILEUP参数包括默认空位权重3.00、默认空位长度权重0.10和加权末端空位。其他可用的算法是Altschul等人描述的BLAST算法(参见，Altschul等人，J.Mol.Biol.215:403-410[1990]；以及Karlin和Altschul，Proc.Natl.Acad.Sci.USA90:5873-5787[1993])。BLAST程序利用多个搜索参数，其中大部分参数设定为默认值。
NCBIBLAST算法可找出就生物学相似性而言最相关的序列，但不推荐将该算法用于少于20个残基的查询序列(AltschulSF等人，(1997)NucleicAcidsRes.25:3389-3402；以及SchafferAA等人，(2001)NucleicAcidsRes.29:2994-3005)。用于核酸序列搜索的示例性默认BLAST参数为：
以下是用于氨基酸序列搜索的参数：
氨基酸序列的同一性百分比值(％)通过用匹配的相同残基数除以“参考”序列的残基总数(包括由程序为进行最优/最大比对所创建的任何空位)来确定。如果某一序列与SEQIDNO:A具有90％的同一性，那么SEQIDNO:A为“参考”序列。BLAST算法将“参考”序列称作“查询”序列。
CLUSTALW算法是序列比对算法的另一个例子。参见Thompson等人，(1994)NucleicAcidsRes.22:4673-4680。对于CLUSTALW算法的默认参数为：
在CLUSTAL算法中，两端发生的缺失也包括在内。例如，500个氨基酸的多肽的任一端(或多肽内)出现5个氨基酸缺失的变体将具有相对于“参考”多肽为99％的序列同一性百分比(495/500相同残基×100)。此类变体将被与该多肽具有“至少99％的序列同一性”的变体所涵盖。
如果所关注的多肽包含与参考多肽的氨基酸序列具有至少约60％、至少约65％、至少约70％、至少约75％、至少约80％、至少约85％、至少约90％、至少约91％、至少约92％、至少约93％、至少约94％、至少约95％、至少约96％、至少约97％、至少约98％、至少约99％或至少约99.5％的序列同一性的氨基酸序列，则可称所关注的多肽与参考多肽“基本上相同”。两条这类多肽之间的百分比同一性可通过观察两条经最佳比对的多肽序列来手动确定，或通过使用软件程序或算法(如BLAST、ALIGN、CLUSTAL)采用标准参数来确定。两条多肽基本上相同的一个指示是第一多肽与第二多肽是免疫交叉反应性的。通常，因保守氨基酸置换而不同的多肽是免疫交叉反应性的。因而，例如在某条多肽与第二多肽仅因保守氨基酸置换或一个或多个保守氨基酸置换而不同的情况中，这两条多肽是基本上相同的。
如果所关注的核酸包含与参考核酸的核苷酸序列具有至少约60％、至少约65％、至少约70％、至少约75％、至少约80％、至少约85％、至少约90％、至少约91％、至少约92％、至少约93％、至少约94％、至少约95％、至少约96％、至少约97％、至少约98％、至少约99％或至少约99.5％的序列同一性的核苷酸序列，则可称所关注的核酸与参考核酸“基本上相同”。两条这类核酸之间的百分比同一性可通过观察两条经最佳比对的核酸序列来手动确定，或通过使用软件程序或算法(如BLAST、ALIGN、CLUSTAL)采用标准参数来确定。两条核酸序列基本上相同的一个指示是这两条核酸分子在严格条件(例如，在中等严格性至高严格性的范围内)下彼此杂交。
当将核酸或多核苷酸从其他组分(包括但不限于，例如，其他蛋白质、核酸、细胞等)部分地或完全地分离时，该核酸或多核苷酸是“分离的”。同样，当将多肽、蛋白质或肽从其他组分(包括但不限于，例如，其他蛋白质、核酸、细胞等)部分地或完全地分离时，该多肽、蛋白质或肽是“分离的”。以摩尔计，在组合物中，分离的物质的丰度比其他物质更高。例如，分离的物质可占所有存在的大分子物质的至少约50％、约70％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、约99％或约100％(以摩尔计)。优选地，所关注的物质纯化为基本上均一的(即通过常规检测方法在组合物中检测不到污染物)。纯度和均一性可以使用本领域熟知的多种技术测定，例如蛋白质或核酸样品的琼脂糖或聚丙烯酰胺凝胶电泳，再在染色后目测观察。如果需要，可采用高分辨率技术，例如高效液相色谱法(HPLC)或类似的方法来纯化材料。
应用于核酸或多肽的术语“纯化的”一般表示核酸或多肽基本上不含其他组分，如本领域熟知的分析技术所测定(如，纯化的多肽或多核苷酸在电泳凝胶、色谱洗脱物和/或经密度梯度离心的介质中形成离散的条带)。例如，在电泳凝胶中基本上产生一条带的核酸或多肽是“纯化的”。纯化的核酸或多肽的纯度为至少约50％，通常其纯度为至少约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、约99％、约99.5％、约99.6％、约99.7％、约99.8％或更大(如，以摩尔计的重量百分比)。在相关的意义上，本发明提供使组合物富集一种或多种本发明的分子，例如一种或多种本发明的多肽或多核苷酸的方法。在应用纯化或富集技术后某分子的浓度有实质性增加时，则组合物富集了该分子。基本上纯的本发明的多肽或多核苷酸(如，分别为基本上纯的本发明变体脂解酶或编码本发明变体脂解酶的多核苷酸)通常占特定组合物中的所有大分子物质重量(以摩尔计)的至少约55％、约60％、约70％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、约99％、约99.5％或更高。
在本文中，给定氨基酸序列中氨基酸残基的位置通常利用SEQIDNO:1中示出的疏棉状嗜热丝孢菌脂肪酶TLL氨基酸序列的对应氨基酸残基的位置的编号方式进行编号。SEQIDNO:1的疏棉状嗜热丝孢菌脂肪酶TLL氨基酸序列因而充当参考亲本序列。可利用本文所述的比对算法将给定氨基酸序列(例如，本文所述的变体脂解酶氨基酸序列)与TLL序列(SEQIDNO:1)比对，该给定氨基酸序列中与TLL序列中的氨基酸残基比对(优选最佳比对)的氨基酸残基可通过参考脂肪酶TLL序列中对应的氨基酸残基便利地进行编号。
本发明的脂解酶
如本文所用，脂解酶包括具有降解脂质的能力(例如，降解甘油三酯或磷脂的能力)的酶、多肽或蛋白质。脂解酶可为例如脂肪酶、磷脂酶、酯酶、聚酯酶或角质酶。脂解酶可为具有αβ水解酶折叠的脂解酶。这些酶通常具有丝氨酸、天冬氨酸和组氨酸残基的催化三联体。αβ水解酶包括脂肪酶和角质酶。角质酶显示极少的(若有的话)界面激活，而脂肪酶在脂质-水界面存在的情况下往往发生构象变化(Longhi和Cambillau(1999)BiochimicaetBiophysicaActa1441:185-96)。脂解酶的活性片段是脂解酶中保持脂质降解能力的部分。活性片段保留该催化三联体。如本文所用，可根据任何本领域已知的工序测定脂解活性(参见例如，Gupta等人，Biotechnol.Appl.Biochem.，37:63-71，2003；美国专利5,990,069；以及国际专利公布WO96/18729A1)。
在一些实施方案中，本发明的脂解酶是α/β水解酶。在一些实施方案中，本发明的脂解酶是脂肪酶。在一些实施方案中，本发明的脂解酶是角质酶。在一些实施方案中，本发明的脂解酶是聚酯酶。
脂解酶的生产性位置(productivepositions)
本发明在可用于洗涤剂组合物的脂解酶中的2、3、4、5、6、7、8、9、10或更多个氨基酸位置处提供修饰(例如置换)，其中有利修饰的结果是与亲本脂解酶相比，脂解活性的性能指数(pI)提高，例如改善的脂肪酸酯或甘油三酯的水解，或例如改善的丁酸对硝基苯酯、辛酸对硝基苯酯或棕榈酸对硝基苯酯的水解，增加的pNPP/pNPB或pNPO/pNPB比活性比值，增加的pNPB/pNPP比活性比值，增加的热稳定性，增加的洗涤剂稳定性，增加的LAS稳定性，增加的织物粘附性，降低的织物粘附性或增加的清洁性能。就对亲本脂解酶的组合修饰而言，可认为这些氨基酸位置是有用的位置。
被认为有用位置的脂解酶氨基酸位置可以具有适用于洗涤剂组合物的不同修饰。修饰可包括在特定位置处的插入、缺失或置换。在一个实施方案中，修饰方式为置换。
在一些实施方案中，本发明是脂解酶变体或其活性片段，该脂解酶变体或其活性片段包含对亲本脂解酶的至少两、三、四、五、六、七、八、九或十或更多个氨基酸修饰，其中第一氨基酸修饰位于所述脂解酶变体中选自以下的位置：第4、11、18、23、24、27、29、33、45、48、49、51、56、58、73、75、77、90、91、94、111、130、137、154、156、163、187、189、227、233、252、256和264位，其中该变体的氨基酸位置是根据与SEQIDNO：1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。在一些实施方案中，本发明是脂解酶变体或其活性片段，其中所述对亲本脂解酶的至少两、三、四、五、六、七、八、九或十或更多个氨基酸修饰位于所述脂解酶变体中选自以下的位置：第4、11、18、23、24、27、29、33、45、48、49、51、56、58、73、75、77、90、91、94、111、130、137、154、156、163、187、189、227、233、252、256和264位，其中该变体的氨基酸位置是根据与SEQIDNO：1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。
在一些实施方案中，本发明的脂解酶变体或其活性片段可具有两个或更多个修饰，其中所述修饰为A018K/D027S、A018K/E045F、A018K/L075D，A018K/L075Q，A018K/T189D，D027E/G163P，D027N/E056K，D027N/N233Q，D027S/N033D，D027S/P256T，D111A/L264R，D130A/L264R，D130A/T189Q，D130A/V187N，D130A/V187T，E045F/N073R，E045F/T189D，F051T/L075R，G023K/D130A，G023K/L264R，G023Q/L075Q，K024A/L075Q，K024A/V154I，L075Q/D130A，L075Q/G156W，L075R/L264R，L227M/L264R，N011K/D027S，N033D/E045F，N073R/L075D，N094R/G156W，P029E/N033D，S058M/L264R，V187T/L264R，A018K/A049V/L075Q，A018K/A049V/V187T，A018K/D027S/E045F，A018K/D027S/N073R，A018K/D027S/T189D，A018K/E045F/N073R，A018K/E045F/T189D，A018K/E045F/V187T，A018K/G023K/G156W，A018K/G023K/L075Q，A018K/G023Q/L075Q，A018K/K024A/V154I，A018K/L075D/T189D，A018K/L075Q/G156W，A018K/L075Q/V187T，A018K/N033D/L075D，A018K/N033D/T189D，A018K/N073R/L075D，A018K/P029E/T189D，A018K/V154I/G156W，A049V/L075Q/T189D，A049V/V187T/T189D，D027E/S058M/G163P，D027N/N233Q/P256T，D027N/S058M/P256T，D027Q/N233Q/P256T，D027Q/S058M/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/N073R/L075D，D027S/N233Q/P256T，D027S/S058M/P256T，D111A/D130A/L264R，D130A/T189Q/L264R，D130A/V154I/G156W，D130A/V187N/L264R，D130A/V187T/L264R，E045F/L075D/T189D，E045F/N073R/L075D，E056K/D130A/L264R，E056K/D130A/T189Q，F051T/I252Q/L264R，G023K/D130A/L264R，G023K/D130A/V187T，G023K/E056K/V187T，G023K/L075R/L264R，G023K/V187N/L264R，G023K/V187T/L264R，G023Q/A049V/T189D，G023Q/D111A/L264R，G023Q/D130A/G156W，G023Q/E045F/V077I，G023Q/E056K/L075R，G023Q/K024A/G156W，G023Q/L075Q/V077I，G023Q/L075Q/V187T，G163P/L227M/L264R，I090F/N233Q/P256T，K024A/D130A/V154I，K024A/L075Q/G156W，K024A/L075Q/V077I，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/D130A/V187T，L075Q/G156W/T189D，L075Q/G156W/V187N，L075Q/G156W/V187T，L075Q/N094R/V154I，L075Q/V077I/T189D，L075Q/V154I/V187T，L075Q/V187N/L264R，L075Q/V187T/L264R，L075Q/V187T/T189D，L075R/D130A/L264R，L075R/D130A/V187T，L075R/V187N/L264R，L075R/V187T/L264R，N011K/D027S/S058M，N011K/N233Q/P256T，N033D/E045F/L075D，N033D/E045F/N073R，N033D/N073R/T189D，N073R/L075D/T189D，N094R/D130A/V187T，P029E/E045F/L075D，P029E/L075D/T189D，P029E/N033D/E045F，P029E/N033D/L075D，P029E/N073R/L075D，Q004D/D027S/P256T，Q004D/N233Q/P256T，Q004D/S058M/N233Q，S058M/L227M/L264R，V077I/D130A/V154I，V077I/V187A/T189D，V187N/T189Q/L264R，A018K/A049V/L075Q/T189D，A018K/A049V/L075Q/V187T，A018K/D027S/E045F/N073R，A018K/D027S/E045F/T189D，A018K/D027S/L075D/T189D，A018K/D027S/N033D/L075D，A018K/D027S/P029E/T189D，A018K/D111A/G156W/T189Q，A018K/D130A/G156W/V187T，A018K/E045F/A049V/G156W，A018K/E045F/A049V/V187T，A018K/E045F/L075D/T189D，A018K/E045F/L075Q/T189D，A018K/E045F/L075Q/V187T，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/T189D，A018K/G023Q/A049V/V187T，A018K/G023Q/E045F/T189D，A018K/G023Q/E045F/V187T，A018K/G023Q/G156W/V187T，A018K/G023Q/L075Q/T189D，A018K/G023Q/L075Q/V077I，A018K/G023Q/L075Q/V187T，A018K/G023Q/L075R/D130A，A018K/G023Q/V077I/G156W，A018K/G023Q/V077I/V187T，A018K/G023Q/V187T/T189D，A018K/K024A/D130A/G156W，A018K/L075Q/G156W/T189D，A018K/L075Q/G156W/V187T，A018K/L075Q/N094R/D111A，A018K/L075Q/N094R/D130A，A018K/L075Q/N094R/V187Q，A018K/L075Q/V077I/N094R，A018K/L075Q/V187T/T189D，A018K/N033D/L075D/T189D，A018K/N073R/L075D/T189D，A018K/P029E/N033D/L075D，A018K/P029E/N033D/T189D，A018K/P029E/N073R/L075D，A018K/V154I/G156W/V187T，A049V/L075Q/V077I/T189D，A049V/L075Q/V187T/T189D，D027E/D048Q/G163P/L264R，D027E/D130A/N233Q/L264R，D027E/D137Q/G163P/L227M，D027E/D137Q/L227M/L264R，D027E/G163P/L227M/L264R，D027E/L227M/N233Q/L264R，D027E/S058M/G163P/L264R，D027N/E056K/N233Q/P256T，D027N/E056K/S058M/P256T，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027Q/S058M/I090F/P256T，D027S/D130A/I252Q/L264R，D027S/E045F/L075D/T189D，D027S/E045F/N073R/L075D，D027S/E045F/N073R/T189D，D027S/E056K/D111A/V154I，D027S/E056K/I090F/P256T，D027S/N033D/E045F/N073R，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/L075D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/L075D，D027S/P029E/E045F/N073R，D027S/P029E/E045F/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D027S/P029E/N033D/N073R，D027S/P029E/N033D/T189D，D048Q/D130A/G163P/L264R，D048Q/D137Q/G163P/L227M，D048Q/G163P/N233Q/L264R，D111A/D130A/V154I/G156W，D111A/D130A/V154I/L264R，D111A/D130A/V187T/L264R，D130A/G163P/L227M/L264R，D130A/G163P/N233Q/L264R，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，E056K/D130A/V187N/L264R，E056K/D130A/V187T/L264R，E056K/L075Q/V187N/L264R，F051T/D130A/I252Q/L264R，F051T/L075G/I252Q/L264R，F051T/L075Q/I252Q/L264R，G023K/D027Q/F051T/L075Q，G023K/D027S/L075Q/L264R，G023K/D111A/D130A/L264R，G023K/D130A/G156W/T189Q，G023K/D130A/V187N/L264R，G023K/D130A/V187T/L264R，G023K/D130A/V187T/T189Q，G023K/E056K/D130A/L264R，G023K/E056K/D130A/V187N，G023K/E056K/L075R/L264R，G023K/E056K/V187T/L264R，G023K/K024A/V154I/V187N，G023K/L075Q/D130A/V187N，G023K/L075Q/G156W/L264R，G023K/L075Q/G156W/V187N，G023K/L075R/D130A/L264R，G023K/L075R/V187N/L264R，G023K/L075R/V187T/L264R，G023Q/A049V/L075Q/T189D，G023Q/A049V/L075Q/V077I，G023Q/A049V/V077I/G156W，G023Q/D027S/D111A/G156W，G023Q/E045F/A049V/T189D，G023Q/E045F/A049V/V187T，G023Q/E045F/L075Q/G156W，G023Q/E045F/L075Q/V187T，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/K024A/D130A/G156W，G023Q/K024A/L075R/G156W，G023Q/K024A/L075R/V154I，G023Q/K024A/V077I/G156W，G023Q/L075Q/D130A/L264R，G023Q/L075Q/G156W/V187N，G023Q/L075Q/G156W/V187T，G023Q/L075Q/V187T/L264R，G023Q/V154I/G156W/V187N，G091Q/V187T/I252Q/L264R，K024A/D130A/V154I/V187T，K024A/L075Q/D130A/G156W，K024A/L075Q/D130A/V154I，K024A/L075Q/V187T/T189Q，K024A/L075R/G156W/V187N，L075G/D130A/V187T/I252Q，L075G/G091Q/V187N/L264R，L075G/V187H/I252Q/L264R，L075Q/D130A/G156W/V187N，L075Q/D130A/V154I/G156W，L075Q/G156W/V187T/L264R，L075Q/V077I/D130A/V187Q，L075Q/V077I/G156W/V187N，L075Q/V077I/N094R/G156W，L075Q/V077I/V154I/V187Q，L075Q/V077I/V187T/T189D，L075Q/V187N/T189Q/L264R，L075R/D111A/V154I/G156W，L075R/D130A/V154I/L264R，L075R/D130A/V187T/L264R，N011K/D027N/E056K/S058M，N011K/D027N/S058M/P256T，N011K/D027Q/S058M/P256T，N011K/D027S/E056K/P256T，N011K/D027S/N233Q/P256T，N011K/D027S/S058M/N233Q，N011K/E056K/N233Q/P256T，N011K/E056K/S058M/P256T，N011K/G023K/L075Q/D111A，N011K/S058M/N233Q/P256T，N033D/E045F/N073R/T189A，P029E/E045F/L075D/T189D，P029E/E045F/N073R/L075D，P029E/E045F/N073R/T189D，P029E/N033D/E045F/L075D，P029E/N033D/L075D/T189D，P029E/N073R/L075D/T189D，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/D027S/I090F/P256T，Q004D/N011K/D027N/P256T，Q004D/N011K/D027Q/N233Q，Q004D/N011K/D027S/P256T，Q004D/N011K/E056K/S058M，S058M/D137Q/G163P/N233Q，S058M/G163P/L227M/L264R，S058M/G163P/N233Q/L264R，S058M/L075Q/G091Q/I252Q，S058M/L075Q/I252Q/L264R，A018K/A049V/L075Q/V187T/T189D，A018K/D027S/E045F/L075D/T189D，A018K/D027S/E045F/N073R/D137V，A018K/D027S/N033D/E045F/T189D，A018K/D027S/N033D/L075D/T189D，A018K/D027S/N033D/N073R/T189D，A018K/D027S/N073R/L075D/T189D，A018K/D027S/P029E/E045F/T189D，A018K/D027S/P029E/N033D/L075D，A018K/D027S/P029E/N073R/L075D，A018K/D130A/G156W/V187N/L264R，A018K/D130A/G156W/V187T/L264R，A018K/E045F/A049V/L075Q/G156W，A018K/E045F/A049V/L075Q/T189D，A018K/E045F/A049V/L075Q/V187T，A018K/E045F/N073R/L075D/T189D，A018K/G023K/D130A/V154I/G156W，A018K/G023K/K024A/D130A/G156W，A018K/G023K/K024A/D130A/V154I，A018K/G023K/K024A/L075R/N094R，A018K/G023K/L075Q/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/A049V/L075Q/G156W，A018K/G023Q/A049V/L075Q/V077I，A018K/G023Q/A049V/L075Q/V187T，A018K/G023Q/A049V/V077I/V187T，A018K/G023Q/E045F/A049V/L075Q，A018K/G023Q/E045F/A049V/V187T，A018K/G023Q/E045F/L075Q/V187T，A018K/G023Q/E045F/V077I/T189D，A018K/G023Q/K024A/D130A/V154I，A018K/G023Q/K024A/D130A/V187Q，A018K/G023Q/K024A/L075Q/G156W，A018K/G023Q/L075Q/G156W/V187T，A018K/G023Q/L075Q/V187T/T189D，A018K/G023Q/V077I/D130A/G156W，A018K/G023Q/V077I/V187T/T189D，A018K/K024A/L075Q/D130A/L264R，A018K/K024A/L075Q/V154I/G156W，A018K/K024A/L075Q/V154I/T189D，A018K/K024A/L075R/D130A/V154I，A018K/K024A/L075R/N094R/D130A，A018K/K024A/N094R/D130A/V187N，A018K/K024A/N094R/G156W/V187T，A018K/L075Q/D111A/D130A/V187T，A018K/L075Q/G156W/V187T/T189D，A018K/L075Q/N094R/D130A/V187N，A018K/L075Q/N094R/G156W/V187N，A018K/L075Q/V077I/N094R/G156W，A018K/L075Q/V187T/T189Q/L264R，A018K/N033D/E045F/L075D/T189D，A018K/P029E/E045F/N073R/L075D，A018K/P029E/N033D/N073R/L075D，A018K/V077I/G156W/V187T/T189D，D027E/D048Q/D137Q/L227M/L264R，D027E/D048Q/G163P/N233Q/L264R，D027E/D048Q/L227M/N233Q/L264R，D027E/D048Q/S058M/G163P/N233Q，D027E/D130A/D137Q/G163P/L264R，D027E/D130A/G163P/L227M/L264R，D027E/D130A/G163P/N233Q/L264R，D027E/D137Q/G163P/L227M/L264R，D027E/D137Q/G163P/N233Q/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L264R，D027E/S058M/D137Q/G163P/N233Q，D027E/S058M/G163P/L227M/L264R，D027N/E056K/S058M/N233Q/P256T，D027N/S058M/I090F/N233Q/P256T，D027Q/E056K/I090F/N233Q/P256T，D027Q/E056K/S058M/N233Q/P256T，D027Q/F051T/L075Q/D130A/L264R，D027S/E056K/D111A/V187N/L264R，D027S/E056K/S058M/N233Q/P256T，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D027S/L075Q/D111A/D130A/V187N，D027S/L075R/V154I/T189Q/L264R，D027S/N033D/E045F/N073R/L075D，D027S/N033D/N073R/L075D/T189D，D027S/P029E/E045F/N073R/L075D，D027S/P029E/E045F/N073R/T189D，D027S/P029E/N033D/E045F/N073R，D027S/P029E/N033D/N073R/L075D，D027S/P029E/N073R/L075D/T189D，D048Q/D130A/D137Q/G163P/L264R，D048Q/D130A/G163P/L227M/L264R，D048Q/D137Q/G163P/L227M/L264R，D048Q/D137Q/G163P/N233Q/L264R，D048Q/S058M/D130A/N233Q/L264R，D048Q/S058M/D137Q/G163P/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/D137Q/N233Q/L264R，D048Q/S058M/G163P/L227M/L264R，D048Q/S058M/G163P/N233Q/L264R，D130A/D137Q/G163P/L227M/L264R，D130A/D137Q/G163P/N233Q/L264R，D130A/G163P/L227M/N233Q/L264R，D130A/V154I/G156W/V187N/T189Q，E045F/A049V/L075Q/V187T/T189D，E056K/D130A/V187H/I252Q/L264R，E056K/L075G/V187N/I252Q/L264R，E056K/L075Q/G156W/T189Q/L264R，E056K/L075R/D130A/V187N/L264R，E056K/L075R/D130A/V187T/L264R，E056K/N094R/G156W/V187N/L264R，E056K/S058M/I090F/N233Q/P256T，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/G091Q/D130A/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/L264R，F051T/L075Q/V187N/I252Q/L264R，F051T/L075R/V187N/I252Q/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/D027S/E056K/V187T/L264R，G023K/D111A/V15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1A/V154I/V187T/L264R，A018K/G023K/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/G023K/L075R/V077I/N094R/D130A/G156W/V187Q，A018K/G023Q/E045F/L075Q/V077I/G156W/V187T/T189D，A018K/G023Q/K024A/L075Q/V077I/D130A/G156W/V187Q，A018K/G023Q/K024A/L075R/D130A/V154I/G156W/V187N，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，A018K/G023Q/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/K024A/L075Q/V077I/N094R/D130A/V154I/G156W，A018K/K024A/L075R/D130A/V154I/G156W/V187T/L264R，A018K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/E056K/L075G/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/D027S/F051T/E056K/L075R/D130A/V187T/L264R，G023K/D027S/F051T/S058M/L075Q/V187H/I252Q/L264R，G023K/E056K/L075Q/D130A/V154I/V187N/T189Q/L264R，G023K/F051T/E056K/L075R/G091Q/D130A/V187T/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187T/L264R，G023K/K024A/L075R/N094R/V154I/G156W/V187T/T189Q，G023K/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，G023K/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/E056K/S058M/L075R/G091Q/V187N/I252Q，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/E056K/L075Q/D111A/V154I/V187N/T189Q/L264R，G023Q/E056K/L075Q/N094R/D111A/G156W/T189Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/K024A/L075Q/V077I/D130A/V154I/G156W/V187N，G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/V187N，G023Q/L075Q/D111A/D130A/V154I/G156W/V187N/L264R，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，G023Q/L075Q/N094R/D130A/V154I/G156W/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，K024A/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，N011K/G023K/D027S/L075Q/D111A/D130A/V154I/T189Q，N011K/G023K/D027S/L075R/V154I/G156W/V187N/T189Q，N011K/G023K/E056K/L075Q/D130A/V154I/T189Q/L264R，N011K/G023K/L075Q/D111A/G156W/V187N/T189Q/L264R，N011K/G023Q/D027S/L075Q/N094R/V154I/G156W/T189Q，N011K/G023Q/D027S/N094R/V154I/G156W/T189Q/L264R，N011K/G023Q/L075Q/D111A/D130A/V154I/V187N/L264R，N011K/G023Q/L075Q/N094R/V154I/G156W/T189Q/L264R，A018K/G023K/K024A/L075Q/N094R/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/V154I/G156W/V187N，A018K/G023K/K024A/L075Q/N094R/D130A/V187T/T189Q/L264K，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，A018K/G023Q/D027S/L075Q/V077I/R108K/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/G156W/V187H，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/P029E/S058M/V077I/R108K/H135F/G156W，A018K/G023Q/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W/V187N，A018K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V187N/T189Q/L264R，G023K/D027S/F051T/E056K/S058M/L075Q/G091Q/V187N/L264R，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/D027S/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023K/L075Q/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/F051T/E056K/L075G/G091Q/V187N/I252Q/L264R，G023Q/D027S/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，G023Q/D027S/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/G023K/D027S/E056K/L075R/D111A/G156W/V187N/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，N011K/G023Q/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/E045F/A049V/S058M/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/S058M/L075Q/V077I/R108K/H135F/G156W/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023K/D027S/F051T/E056K/S058M/L075G/G091Q/D130A/I252Q/L264R，G023Q/D027S/F051T/E056K/L075R/G091Q/D130A/V187H/I252Q/L264R，G023Q/D027S/F051T/E056K/S058M/L075R/G091Q/D130A/V187H/I252Q，G023Q/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023K/L075R/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，N011K/G023Q/D027S/E056K/L075R/D130A/V154I/G156W/T189Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，N011K/G023Q/L075Q/N094R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T，N011K/G023Q/D027S/L075Q/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/H135F/V187T/T189D，或A018K/G023Q/D027S/P029E/N033D/E045F/A049V/S058M/N073S/K074S/L075Q/V077I/N101D/R108K/H135F/D137V/G156W/V187T/T189D，
其中该变体的氨基酸位置是根据与SEQIDNO：1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。在一些实施方案中，该变体或活性片段具有脂解活性，例如相对于亲本脂解酶，水解丁酸对硝基苯酯、辛酸对硝基苯酯或棕榈酸对硝基苯酯的性能指数(pI)在pH6或8时大于1.0。
在上述实施方案的任何一个中，本发明提供了本发明的变体脂解酶，相对于亲本脂解酶(例如，野生型脂解酶，如野生型脂肪酶)或参考酶(例如，SEQIDNO：2)，该变体脂解酶表现出下列一种或多种性质：改善的脂解活性，例如改善的丁酸对硝基苯酯、辛酸对硝基苯酯或棕榈酸对硝基苯酯的水解，增加的pNPP/pNPB或pNPO/pNPB比活性比值，增加的pNPB/pNPP比活性比值，增加的热稳定性，增加的洗涤剂稳定性，增加的LAS稳定性，增加的织物粘附性，降低的织物粘附性，增加的清洁性能，改善的手洗性能，改善的手工或人工盘碟洗涤性能，改善的自动盘碟洗涤性能，改善的衣物洗涤性能和/或改善的稳定性。
就对亲本脂解酶的组合修饰而言，可认为这些氨基酸位置是有用的位置。因此，本发明包括在上述任何位置处具有一个或多个修饰的脂解酶。
本发明的多肽
本发明提供了新型多肽，可统称为“本发明的多肽”。本发明的多肽包括分离、重组、基本上纯的、或非天然存在的变体脂解酶多肽，包括例如具有酶活性(例如，脂解活性)的变体脂解酶多肽。在一些实施方案中，本发明的多肽可用于清洁应用，而且可掺入清洁组合物，这些清洁组合物可用于清洁需要清洁的物品或表面(例如，物品的表面)的方法中。
在一些实施方案中，该脂解酶变体可为源自嗜热丝孢菌属的亲本脂解酶的变体。已发现嗜热丝孢菌属中存在多种脂解酶，这些酶彼此具有高度的同一性，而且与源自疏棉状嗜热丝孢菌的脂解酶(TLL)(如SEQIDNO：1中所示)也具有高度的同一性。在其他实施方案中，脂解酶变体可以是例如疣孢菌属(Verrucosispora)、糖单孢菌属(Saccharomonospora)、链霉菌属(Streptomyces)、小单孢菌属(Micromonospora)、链孢囊菌属(Streptosporangium)、拟无枝酸菌(Amycolatopsis)、纤维单胞菌属(Cellulomonas)、放线束丝菌属(Actinosynnema)、克里布所菌属(Kribbella)、高温单孢菌属(Thermomonospora)、异常球菌属(Deinococcus)、动球菌属(Kineococcus)、拟诺卡氏菌属(Nocardiopsis)、弗兰克氏菌属(Frankia)、琼斯氏菌属(Jonesia)、假单胞菌属(Pseudomonas)、食酸菌属(Acidovorax)或类诺卡氏菌科(Nocardioidaceae)的亲本脂解酶的变体。
源自疏棉状嗜热丝孢菌的脂解酶(TLL)的序列如下所示：
EVSQDLFNQFNLFAQYSAAAYCGKNNDAPAGTNITCTGNACPEVEKADATFLYSFEDSGVGDVTGFLALDNTNKLIVLSFRGSRSIENWIGNLNFDLKEINDICSGCRGHDGFTSSWRSVADTLRQKVEDAVREHPDYRVVFTGHSLGGALATVAGADLRGNGYDIDVFSYGAPRVGNRAFAEFLTVQTGGTLYRITHTNDIVPRLPPREFGYSHSSPEYWIKSGTLVPVTRNDIVKIEGIDATGGNNQPNIPDIPAHLWYFGLIGTCL(SEQIDNO：1).
在一些实施方案中，脂解酶变体可为与源自嗜热丝孢菌属的脂解酶具有50％、60％、70％、75％、80％、85％、90％、95％、96％、97％、98％、99％或100％同一性的变体。在一些实施方案中，脂解酶变体可为与源自疏棉状嗜热丝孢菌的脂解酶具有50％、60％、70％、75％、80％、85％、90％、95％、96％、97％、98％、99％或100％同一性的变体，所述脂解酶具有以SEQIDNO：1示出的序列。
在一个具体的实施方案中，本发明是源自嗜热丝孢菌属的酶。在一个具体的实施方案中，本发明是源自脂解酶的酶，该脂解酶来源于疏棉状嗜热丝孢菌并且具有以SEQIDNO：1示出的序列。
本发明描述了与克隆自疏棉状嗜热丝孢菌的脂肪酶(TLL)相关的组合物和方法。这些组合物和方法部分地基于克隆表达的TLL在洗涤剂组合物存在时具有羧酸酯水解酶活性(作用于羧酸酯)这一观测结果。甚至在存在蛋白酶的情况下，TLL在洗涤剂组合物中也展示出良好的稳定性。由于TLL具备这些特征，所以特别适用于多种清洁应用，在这些清洁应用中，TLL在存在洗涤剂组合物中的表面活性剂与其他组分的情况下可使脂质水解。
虽然TLL对多种天然和合成底物表现出活性，但该酶表现出对C4-C16底物的偏好。这种特异性使TLL特别适用于水解短链甘油三酯和进行涉及短链脂肪酸的酯交换反应。
在上述实施方案的任何一个中，相对于亲本脂解酶，本发明的变体脂解酶可具有对C4-C16底物的改善的脂解活性。在一些实施方案中，本发明的变体具有增加的pNPP/pNPB或pNPO/pNPB比活性比值。在一些实施方案中，本发明的变体具有增加的pNPB/pNPP比活性比值。在上述实施方案的任何一个中，相对于亲本脂解酶(例如，SEQIDNO：1)，本发明的变体脂解酶水解丁酸对硝基苯酯、辛酸对硝基苯酯或棕榈酸对硝基苯酯的性能指数(pI)可大于1.0。这可在pH6或8下测定。在上述实施方案的任何一个中，相对于亲本脂解酶(例如SEQIDNO：1)，本发明变体脂解酶的洗涤剂稳定性的性能指数(pI)可大于1.0。在上述实施方案的任何一个中，相对于亲本脂解酶(例如SEQIDNO：1)，本发明变体脂解酶的LAS稳定性的性能指数(pI)可大于1.0。在上述实施方案的任何一个中，相对于亲本脂解酶(例如SEQIDNO：1)，本发明变体脂解酶的织物粘附性的性能指数(pI)可大于1.0。在上述实施方案的任何一个中，相对于亲本脂解酶(例如SEQIDNO：1)，本发明变体脂解酶的织物粘附性的性能指数(pI)可小于1.0。在上述实施方案的任何一个中，相对于亲本脂解酶(例如SEQIDNO：1)，本发明变体脂解酶的清洁性能的性能指数(pI)可大于1.0。这可使用CS-61微布样测定法，利用半剂量+辅剂或全剂量测得。在上述实施方案的任何一个中，相对于参考脂解酶(例如SEQIDNO：2)，本发明变体脂解酶的如下方面的性能指数(pI)可大于1.0：丁酸对硝基苯酯、辛酸对硝基苯酯或棕榈酸对硝基苯酯的水解，洗涤剂稳定性，LAS稳定性，织物粘附性或清洁性能。在上述实施方案的任何一个中，相对于参考脂解酶(例如SEQIDNO：2)，本发明变体脂解酶的织物粘附性的性能指数(pI)可小于1.0。
参考脂解酶(源自疏棉状嗜热丝孢菌(TLL)的变体)的序列示于下：
EVSQDLFNQFNLFAQYSAAAYCGKNNDAPAGTNITCTGNACPEVEKADATFLYSFEDSGVGDVTGFLALDNTNKLIVLSFRGSRSIENWIGNLNFDLKEINDICSGCRGHDGFTSSWRSVADTLRQKVEDAVREHPDYRVVFTGHSLGGALATVAGADLRGNGYDIDVFSYGAPRVGNRAFAEFLTVQTGGTLYRITHTNDIVPRLPPREFGYSHSSPEYWIKSGTLVPVRRRDIVKIEGIDATGGNNQPNIPDIPAHLWYFGLIGTCL(SEQIDNO：2).
在上述实施方案的一些中，本发明组合物和方法提供了变体TLL多肽。亲本TLL多肽疏棉状嗜热丝孢菌脂肪酶在WO1994025577中有所描述。成熟TLL多肽具有SEQIDNO：1的氨基酸序列。类似的、基本上相同的TLL多肽可存在于自然界中，例如，存在于疏棉状嗜热丝孢菌的其他菌株或分离株中。这些和其他重组TLL多肽涵盖在本发明的组合物和方法中。
在上述实施方案的任何一个中，本发明包括具有脂解活性的分离、重组、基本上纯的或非天然存在的变体脂解酶，该多肽包含与本文所提供的亲本脂解酶具有至少约85％、至少约86％、至少约87％、至少约88％、至少约89％、至少约90％、至少约91％、至少约92％、至少约93％、至少约94％、至少约95％、至少约96％、至少约97％、至少约98％、至少约99％、至少约99.5％、或100％的序列同一性的多肽序列。
在一些实施方案中，该变体多肽是与例示的TLL多肽具有特定程度的氨基酸序列同源性的变体，例如，与SEQIDNO：1的氨基酸序列具有至少70％、至少75％、至少80％、至少85％、至少90％、至少91％、至少92％、至少93％、至少94％、至少95％、至少96％、至少97％、至少98％、或甚至至少99％的序列同源性。同源性可通过氨基酸序列比对，例如，使用如本文所述的程序如BLAST、ALIGN、或CLUSTAL来确定。
本发明也提供分离、重组、基本上纯的、或非天然存在的序列，该序列编码具有脂解活性的变体脂解酶，所述变体脂解酶(例如，变体脂肪酶)包含与SEQIDNO：1的氨基酸序列相差不超过50个、不超过40个、不超过30个、不超过35个、不超过25个、不超过20个、不超过19个、不超过18个、不超过17个、不超过16个、不超过15个、不超过14个、不超过13个、不超过12个、不超过11个、不超过10个、不超过9个、不超过8个、不超过7个、不超过6个、不超过5个、不超过4个、不超过3个、或不超过2个氨基酸残基的氨基酸序列，其中变体脂肪酶的氨基酸位置是根据SEQIDNO：1中所示疏棉状嗜热丝孢菌脂肪酶TLL的氨基酸序列中对应氨基酸位置的编号方式进行编号的，这通过比对变体脂解酶的氨基酸序列与疏棉状嗜热丝孢菌脂肪酶TLL的氨基酸序列来确定。
在一些实施方案中，本发明涉及具有脂肪酶活性的分离的多肽，该多肽由多核苷酸编码，该多核苷酸优选在非常低严格性条件下，更优选低严格性条件下，更优选中等严格性条件下，更优选中高等严格性条件下，甚至更优选高严格性条件下，以及最优选非常高严格性条件下与SEQIDNO：1的成熟多肽编码序列、或其全长互补链杂交(J.Sambrook、E.F.Fritsch和T.Maniatis，1989年，“MolecularCloning，ALaboratoryManual”，第二版，ColdSpringHarbor，NewYork)。
如上所述，本发明的变体脂解酶多肽具有酶活性(例如，脂解活性)，因而可用于清洁应用，清洁应用包括但不限于清洁盘碟物品、餐具物品、织物、以及具有硬质表面的物品(例如，桌子、桌面、墙壁、家具物品、地板、天花板等的硬质表面)的方法。包含一种或多种本发明的变体脂解酶多肽的示例性清洁组合物在下文中描述。本发明的变体脂解酶多肽的酶活性(例如，脂解酶活性)可使用本领域普通技术人员熟知的工序容易地测定。本发明的变体脂解酶在去除污渍(例如，脂质污渍)、清洁硬质表面、或清洁衣物、盘碟或餐具物品方面的性能可用本领域所熟知的工序容易地测定。
可对本发明的多肽进行多种改变，例如一个或多个氨基酸的插入、缺失和/或置换(保守的或非保守的置换)，包括其中这类改变不会实质上改变该多肽的酶活性的情况。类似地，也可对本发明的核酸进行多种改变，例如在一个或多个密码子中对一个或多个核酸进行一个或多个置换以使得特定的密码子编码相同或不同的氨基酸，从而导致沉默变异(例如，核苷酸序列中的突变导致氨基酸序列中的沉默突变，如当所编码的氨基酸未因该核酸突变而变更时)或非沉默变异；序列中一个或多个核酸(或密码子)的一个或多个缺失；序列中一个或多个核酸(或密码子)的一个或多个添加或插入；和/或序列中一个或多个核酸(或密码子)的剪切或序列中一个或多个核酸(或密码子)的一个或多个截短。与初始核酸序列编码的变体脂解酶相比，核酸序列中的许多此类改变可能不会实质上改变所得的编码的变体脂解酶的酶活性。还可对本发明的核酸进行修饰以包括一个或多个使得能在表达系统(例如，细菌表达系统)中进行最优表达的密码子，同时如果需要，所述一个或多个密码子仍编码相同氨基酸。
在一些实施方案中，本发明提供了包含具有所需酶活性(例如，脂解酶活性或清洁性能活性)的变体脂解酶多肽的一类多肽，其包含具有本文所述的氨基酸置换的序列并且其还包含一个或多个另外的氨基酸置换，例如保守和非保守置换，其中所述多肽表现出、维持或大致维持所需的酶活性(例如，在变体脂解酶的清洁活性或性能中所反映出的脂解酶活性或脂肪酶活性)。根据本发明的氨基酸置换可包括但不限于一个或多个非保守置换和/或一个或多个保守氨基酸置换。保守氨基酸残基置换通常涉及将一功能类别的氨基酸残基中的成员替换为属于同一功能类别的残基(在计算功能同源性百分数时相同氨基酸残基被视为在功能上是同源的或保守的)。保守氨基酸置换通常涉及用功能相似的氨基酸置换氨基酸序列中的氨基酸。例如，丙氨酸、甘氨酸、丝氨酸和苏氨酸是功能相似的，因而可充当彼此的保守氨基酸置换。天冬氨酸和谷氨酸可充当彼此的保守置换。天冬酰胺和谷氨酰胺可充当彼此的保守置换。精氨酸、赖氨酸和组氨酸可充当彼此的保守置换。异亮氨酸、亮氨酸、甲硫氨酸和缬氨酸可充当彼此的保守置换。苯丙氨酸、酪氨酸和色氨酸可充当彼此的保守置换。
可以设想其他的保守氨基酸置换组。例如，可通过相似的功能或化学结构或组成对氨基酸进行分组(例如，酸性、碱性、脂族、芳族、含硫氨基酸)。例如，脂族组可包括：甘氨酸(G)、丙氨酸(A)、缬氨酸(V)、亮氨酸(L)、异亮氨酸(I)。含有可以被视为彼此的保守置换的氨基酸的其他组包括：芳族氨基酸：苯丙氨酸(F)、酪氨酸(Y)、色氨酸(W)；含硫氨基酸：甲硫氨酸(M)、半胱氨酸(C)；碱性氨基酸：精氨酸(R)、赖氨酸(K)、组氨酸(H)；酸性氨基酸：天冬氨酸(D)、谷氨酸(E)；非极性不带电荷残基：半胱氨酸(C)、甲硫氨酸(M)和脯氨酸(P)；亲水性不带电荷残基：丝氨酸(S)、苏氨酸(T)、天冬酰胺(N)和谷氨酰胺(Q)。另外的氨基酸分组是本领域技术人员所熟知的并在各种标准教科书中有描述。本文的多肽序列的列表，结合上文的置换组，提供了所有经保守置换的多肽序列的明确列表。
在上文描述的氨基酸残基类别中存在更保守的置换，其也可以是合适的或者其作为另一种选择可以是合适的。更保守置换的保守组包括：缬氨酸-亮氨酸-异亮氨酸、苯丙氨酸-酪氨酸、赖氨酸-精氨酸、丙氨酸-缬氨酸和天冬酰胺-谷氨酰胺。因此，例如在一些实施方案中，本发明提供分离或重组的具有脂解活性的变体脂解酶多肽(例如，变体脂肪酶)，所述变体脂解酶多肽包含与SEQIDNO：1的氨基酸序列具有至少约90％、约95％、约96％、约97％、约98％、约99％或约99.5％的序列同一性的氨基酸序列。本发明的变体脂解酶中一个氨基酸保守置换为另一个氨基酸预计不会显著改变该变体脂解酶的酶活性或清洁性能活性。所得脂解酶的酶活性或清洁性能活性可容易地用标准测定法和本文所述的测定法进行测定。
本发明的多肽序列(例如，本发明的变体脂解酶)的保守置换变异包括用保守选择的相同保守置换组的氨基酸置换该多肽序列的少部分氨基酸，有时低于约25％、约20％、约15％、约14％、约13％、约12％、约11％、约10％、约9％、约8％、约7％或约6％的氨基酸，或低于约5％、约4％、约3％、约2％或约1％的氨基酸。
本发明的核酸
本发明提供编码本发明多肽的、分离的、非天然存在的、或重组的核酸(在本文中也称为“多核苷酸”)，其可统称为“本发明的核酸”或“本发明的多核苷酸”。本发明的核酸(包括下文描述的全部核酸在内)可用于本发明多肽的重组生产(例如，表达)，通常是通过表达包含编码所关注的多肽或其片段的序列的质粒表达载体来进行。如上文所述，多肽包括变体脂解酶多肽，包括具有酶活性(例如，脂解活性)的变体脂肪酶多肽，其可用于清洁应用和清洁组合物，以供清洁需要进行清洁的物品或表面(例如，物品的表面)。
在一些实施方案中，本发明提供分离、重组、基本上纯的或非天然存在的核酸，该核酸包含编码上述题为“本发明的多肽”的章节中以及本文其他地方所述的任何本发明多肽(包括任何融合蛋白等)的核苷酸序列。本发明还提供分离、重组、基本上纯的或非天然存在的核酸，该核酸包含编码上述和本文其他地方所述的任何本发明多肽中两种或更多种的组合的核苷酸序列。
本发明的核酸可通过使用任何合适的合成、操纵和/或分离技术或这些技术的组合来产生。例如，本发明的多核苷酸可用标准的核酸合成技术，如本领域技术人员所熟知的固相合成技术来产生。还可通过本领域已知的任何合适的方法促进(或者实现)本发明核酸的合成，所述方法包括但不限于使用经典的亚磷酰胺方法[参见参见例如Beaucage等人，TetrahedronLetters22：1859-69(1981)]的化学合成；或Matthes等人所描述的方法[Matthes等人，EMBOJ.3：801-805(1984)]，如通常在自动合成方法中实施的。本发明的核酸也可使用自动DNA合成仪来产生。可以从多种商业来源订购定制的核酸(例如，米德兰标准试剂公司(MidlandCertifiedReagentCompany)、大美国基因公司(GreatAmericanGeneCompany)、欧佩沦技术公司(OperonTechnologiesInc.)和DNA2.0公司)。用于合成核酸的其他技术和相关原理在本领域中是已知的(参见例如Itakura等人，Ann.Rev.Biochem.53：323(1984)；以及Itakura等人，Science198：1056(1984))。
制备本发明的经修饰变体脂解酶的方法
多种适于产生编码本发明变体脂解酶的本发明经修饰的多核苷酸的方法是本领域已知的，包括但不限于例如点饱和诱变、扫描诱变、插入诱变、缺失诱变、随机诱变、定点诱变和定向进化，以及多种其他重组方法。制备经修饰的多核苷酸和蛋白(例如，变体脂解酶)的方法包括DNA改组方法、基于基因的非同源重组的方法，例如ITCHY(参见Ostermeier等人，7：2139-44(1999))、SCRACHY(参见Lutz等人，98：11248-53(2001))、SHIPREC(参见Sieber等人，19：456-60(2001))、和NRR(参见Bittker等人，20：1024-9(2001)；Bittker等人，101：7011-6(2004))，以及依赖于使用寡核苷酸来插入随机和定向突变、缺失和/或插入的方法(参见Ness等人，20：1251-5(2002)；Coco等人，20：1246-50(2002)；Zha等人，4：34-9(2003)；Glaser等人，149：3903-13(1992))。
产生本发明的变体脂解酶的载体、细胞和方法
本发明提供分离的或重组的载体，该载体包含至少一种本文所述的本发明多核苷酸(例如，编码本文所述的本发明变体脂解酶的多核苷酸)；分离的或重组的包含至少一种本发明的核酸或多核苷酸的表达载体或表达盒；分离的、基本上纯的或重组的包含至少一种本发明的核酸或多核苷酸的DNA构建体；分离的或重组的包含至少一种本发明多核苷酸的细胞；包含细胞的细胞培养物，该细胞包含至少一种本发明的多核苷酸；包含至少一种本发明的核酸或多核苷酸的细胞培养物；以及包含一种或多种此类载体、核酸、表达载体、表达盒、DNA构建体、细胞、细胞培养物、或它们的任何组合或混合物的组合物。
在一些实施方案中，本发明提供包含至少一种本发明载体(例如，表达载体或DNA构建体)的重组细胞，该载体包含至少一种本发明的核酸或多核苷酸。一些这类重组细胞用所述至少一种载体转化或转染。此类细胞通常称为宿主细胞。一些此类细胞包含细菌细胞，包括但不限于芽孢杆菌属(bacillus)细胞，例如枯草芽孢杆菌(B.subtilis)细胞。一些此类细胞包含真菌细胞，包括但不限于木霉属(Trichoderma)细胞，例如里氏木霉(T.reesei)细胞。本发明还提供包含至少一种本发明的变体脂解酶的重组细胞(例如，重组宿主细胞)。
在一些实施方案中，本发明提供包含本发明的核酸或多核苷酸的载体。在一些实施方案中，该载体为表达载体或表达盒，其中编码本发明变体脂解酶的本发明多核苷酸序列有效连接至有效基因表达所需的一个或多个核酸片段(例如，有效连接至编码本发明变体脂解酶的本发明多核苷酸的启动子)。载体可包括转录终止子和/或选择基因，例如抗生素抗性基因，该抗性基因使得能通过在含有抗微生物剂的培养基中生长而对质粒感染的宿主细胞进行连续的培养维持。
表达载体可得自质粒或病毒DNA，或在可供选择的实施方案中，包含这二者的元件。示例性载体包括但不限于pXX、pC194、pJH101、pE194、pHP13(参见Harwood和Cutting编辑，第3章，MolecularBiologicalMethodsforBacillus，JohnWiley&Sons；合适的枯草芽孢杆菌复制质粒包括列于第92页的那些质粒；另参见Perego，“IntegrationalVectorsforGeneticManipulationsinBacillussubtilis”，载于Sonenshein等人编辑的BacillussubtilisandOtherGram-PositiveBacteria：Biochemistry，PhysiologyandMolecularGenetics，AmericanSocietyforMicrobiology，Washington，D.C.，第615-624页)。
为了在细胞中表达和产生所关注的蛋白质(例如，变体脂解酶)，将包含至少一个拷贝(优选包含多个拷贝)的编码经修饰脂解酶的多核苷酸的至少一个表达载体在适于表达该脂解酶的条件下转化进细胞中。在本发明的一些实施方案中，编码变体脂解酶的多核苷酸序列(以及载体中包含的其他序列)整合进宿主细胞的基因组中，而在其他实施方案中，包含编码变体脂解酶的多核苷酸序列的质粒载体在该细胞内保持为自主的染色体外元件。本发明提供染色体外核酸元件以及整合进宿主细胞基因组中的输入核苷酸序列二者。本文所述的载体可以用于产生本发明的变体脂解酶。在一些实施方案中，编码变体脂解酶的多核苷酸构建体存在于整合载体上，这使得编码变体脂解酶的多核苷酸整合到细菌染色体中以及任选地扩增。整合位点的示例是本领域的技术人员所熟知的。在一些实施方案中，编码本发明变体脂解酶的多核苷酸的转录通过启动子来实现，该启动子对所选择的前体脂解酶而言是野生型启动子。在一些其他实施方案中，启动子对于前体脂解酶而言是异源的，但在宿主细胞中有功能。具体地讲，适用于细菌宿主细胞的启动子例子包括但不限于，例如amyE、amyQ、amyL、pstS、sacB、pSPAC、pAprE、pVeg、pHpaII启动子；嗜热脂肪芽孢杆菌(B.stearothermophilus)产麦芽糖淀粉酶基因、疏棉状嗜热丝孢菌(BAN)淀粉酶基因、枯草芽孢杆菌碱性脂解酶基因、克劳氏芽孢杆菌(B.clausii)碱性脂解酶基因、短小芽孢杆菌(B.pumilis)木糖苷酶基因、苏云金芽孢杆菌(B.thuringiensis)cryIIIA、以及地衣芽孢杆菌(B.licheniformis)α-淀粉酶基因的启动子。另外的启动子包括但不限于A4启动子以及噬菌体λPR或PL启动子，和大肠杆菌lac、trp或tac启动子。
本发明的变体脂解酶可在任何合适的革兰氏阳性微生物(包括细菌和真菌)的宿主细胞中产生。例如，在一些实施方案中，变体脂解酶在真菌和/或细菌来源的宿主细胞中产生。在一些实施方案中，宿主细胞为芽孢杆菌属物种、链霉菌属物种、埃希杆菌属(Escherichia)物种、曲霉菌属(Aspergillus)物种、木霉属物种、假单胞菌属物种、棒状杆菌属(Corynebacterium)物种、酵母属(Saccharomyces)物种、或毕赤酵母属(Pichia)物种。在一些实施方案中，变体脂解酶由芽孢杆菌属物种宿主细胞产生。用于产生本发明变体脂解酶的芽孢杆菌属物种宿主细胞的示例包括但不限于地衣芽孢杆菌、迟缓芽孢杆菌(B.lentus)、枯草芽孢杆菌、解淀粉芽孢杆菌(B.amyloliquefaciens)、迟缓芽孢杆菌、短芽孢杆菌(B.brevis)、嗜热脂肪芽孢杆菌、嗜碱芽孢杆菌(B.alkalophilus)、凝结芽孢杆菌(B.coagulans)、环状芽孢杆菌(B.circulans)、短小芽孢杆菌、苏云金芽孢杆菌、克劳氏芽孢杆菌、巨大芽孢杆菌(B.megaterium)、以及芽孢杆菌属内的其他生物体。在一些实施方案中，枯草芽孢杆菌宿主细胞用于产生变体脂解酶。美国专利5,264,366和4,760,025(RE34,606)描述了可用于产生本发明的变体脂解酶的不同芽孢杆菌属宿主菌株，但也可使用其他合适的菌株。
可用于产生本发明变体脂解酶的多种工业细菌菌株包括非重组(即，野生型)芽孢杆菌属物种菌株以及天然存在的菌株和/或重组菌株的变体。在一些实施方案中，宿主菌株为重组菌株，其中编码所关注的多肽的多核苷酸已引入该宿主中。在一些实施方案中，宿主菌株为枯草芽孢杆菌宿主菌株，尤其是重组枯草芽孢杆菌宿主菌株。许多枯草芽孢杆菌菌株是已知的，包括但不限于例如1A6(ATCC39085)、168(1A01)、SB19、W23、Ts85、B637、PB1753到PB1758、PB3360、JH642、1A243(ATCC39,087)、ATCC21332、ATCC6051、MI113、DE100(ATCC39,094)、GX4931、PBT110和PEP211菌株(参见例如Hoch等人，Genetics73：215-228[1973]；另参见美国专利4,450,235和4,302,544以及EP0134048，这些文献全文以引用方式并入本文)。使用枯草芽孢杆菌作为表达宿主细胞是本领域熟知的(参见例如Palva等人，Gene19：81-87[1982]；Fahnestock和Fischer，J.Bacteriol.，165：796-804[1986]；以及Wang等人，Gene69：39-47[1988])。
在一些实施方案中，芽孢杆菌宿主细胞是在如下基因中的至少一者中包括突变或缺失的芽孢杆菌属物种：degU、degS、degR和degQ。优选地，突变在degU基因中，并且更优选地突变是degU(Hy)32(参见例如Msadek等人，J.Bacteriol.172：824-834[1990]；以及Olmos等人，Mol.Gen.Genet.253：562-567[1997])。一个合适的宿主菌株是携带degU32(Hy)突变的枯草芽孢杆菌。在一些实施方案中，芽孢杆菌宿主包含如下基因中的突变或缺失：scoC4(参见例如Caldwell等人，J.Bacteriol.183：7329-7340[2001])；spoIIE(参见例如Arigoni等人，Mol.Microbiol.31：1407-1415[1999])；和/或oppA或opp操纵子的其他基因(参见例如Perego等人，Mol.Microbiol.5：173-185[1991])。实际上，设想到，opp操纵子中可以与oppA基因突变导致相同的表型的任何突变，均可用于本发明的改变的芽孢杆菌菌株的一些实施方案。在一些实施方案中，这些突变单独地出现，而在其他实施方案中，存在突变的组合。在一些实施方案中，可用于产生本发明的变体脂解酶的改变的芽孢杆菌宿主细胞菌株是已在上述基因的一者或多者中包含突变的芽孢杆菌宿主菌株。此外，可以使用包含内源脂解酶基因的突变和/或缺失的芽孢杆菌属物种宿主细胞。在一些实施方案中，芽孢杆菌宿主细胞包含aprE和nprE基因的缺失。在其他实施方案中，芽孢杆菌属物种宿主细胞包含5个脂解酶基因的缺失，而在其他实施方案中，芽孢杆菌属物种宿主细胞包含9个脂解酶基因的缺失(参见例如美国专利申请公布2005/0202535，该专利以引用方式并入本文)。
可以使用任何合适的本领域已知的方法用至少一条编码至少一种本发明变体脂解酶的核酸转化宿主细胞。无论是将核酸整合进载体中还是在不存在质粒DNA的情况下使用核酸，通常将其引入微生物中，在一些实施方案中，优选引入大肠杆菌细胞或感受态芽孢杆菌细胞中。利用质粒DNA构建体或载体并将这类质粒DNA构建体或载体转化进芽孢杆菌细胞或大肠杆菌细胞中来将核酸(如，DNA)引入这类细胞中的方法是众所周知的。在一些实施方案中，随后从大肠杆菌细胞分离该质粒并且转化进芽孢杆菌细胞中。然而，使用例如大肠杆菌的中间微生物不是必需的，在一些实施方案中，将DNA构建体或载体直接引入芽孢杆菌宿主中。
本领域的技术人员十分了解适于将本发明的核酸或多核苷酸序列引入芽孢杆菌细胞中的方法(参见例如Ferrari等人，“Genetics”载于Harwood等人[编辑]的Bacillus，PlenumPublishingCorp.，第57-72页[1989]；Saunders等人，J.Bacteriol.157：718-726[1984]；Hoch等人，J.Bacteriol.93：1925-1937[1967]；Mann等人，CurrentMicrobiol.13：131-135[1986]；Holubova，FoliaMicrobiol.30：97[1985]；Chang等人，Mol.Gen.Genet.168：11-115[1979]；Vorobjeva等人，FEMSMicrobiol.Lett.7：261-263[1980]；Smith等人，Appl.Env.Microbiol.51：634[1986]；Fisher等人，Arch.Microbiol.139：213-217[1981]；以及McDonald，J.Gen.Microbiol.130：203[1984])。实际上，诸如转化(包括原生质体转化)、以及congression、转导和原生质体融合之类的方法是众所周知的并且适用于本发明。使用转化方法将包含编码本发明变体脂解酶的核酸的DNA构建体或载体引入宿主细胞中。本领域已知的转化芽孢杆菌细胞的方法包括例如质粒标记拯救转化的方法，质粒标记拯救转化涉及携带部分同源的内生质粒的感受态细胞摄入供体质粒(参见Contente等人，Plasmid2：555-571[1979]；Haima等人，Mol.Gen.Genet.223：185-191[1990]；Weinrauch等人，J.Bacteriol.154：1077-1087[1983]；以及Weinrauch等人，J.Bacteriol.169：1205-1211[1987])。在该方法中，输入的供体质粒与内生的“辅助”质粒的同源区在模拟染色体转化的过程中发生重组。
除了通常使用的方法，在一些实施方案中，将宿主细胞直接用包含编码本发明变体脂解酶的核酸的DNA构建体或载体转化(即，引入宿主细胞之前，未使用中间细胞来扩增该DNA构建体或载体，或未以别的方式处理该DNA构建体或载体)。将本发明的DNA构建体或载体引入宿主细胞包括在本领域已知的那些在不插入质粒或载体的情况下将核酸序列(例如，DNA序列)引入宿主细胞中的物理和化学方法。此类方法包括但不限于氯化钙沉淀法、电穿孔法、裸DNA法、脂质体法等等。在另外的实施方案中，将DNA构建体或载体与质粒共同转化，而不插入质粒中。在另外的实施方案中，通过本领域已知的方法从已改变的芽孢杆菌菌株缺失掉选择标记(参见Stahl等人，J.Bacteriol.158：411-418[1984]；以及Palmeros等人，Gene247：255-264[2000])。
在一些实施方案中，将本发明的转化的细胞在常规营养培养基中培养。合适的具体培养条件，例如温度、pH等是本领域技术人员已知的，并且在科学文献中很好地进行了描述。在一些实施方案中，本发明提供包含至少一种本发明的变体脂解酶或至少一种本发明的核酸的培养物(例如，细胞培养物)。还提供的是包含至少一种本发明的核酸、载体或DNA构建体的组合物。
在一些实施方案中，将用至少一条编码至少一种本发明变体脂解酶的多核苷酸序列转化的宿主细胞在允许本发明脂解酶表达的条件下在合适的营养培养基中培养，此后从该培养物回收所得的脂解酶。用于培养细胞的培养基包含任何适于宿主细胞生长的常规培养基，例如含有适当补充剂的基本培养基或复合培养基。合适的培养基得自商业供应商或可根据公布的配方(参见例如，美国典型培养物保藏中心(AmericanTypeCultureCollection)的目录中公布的配方)制备。在一些实施方案中，通过常规的工序从培养基回收由细胞产生的脂解酶，包括但不限于例如，通过离心或过滤从培养基分离出宿主细胞、借助于盐(例如，硫酸铵)沉淀出上清液或滤液的蛋白质组分、进行色谱纯化(如离子交换色谱、凝胶过滤色谱、亲和色谱等)。任何适于回收或纯化变体脂解酶的方法均可用于本发明。
在一些实施方案中，由重组宿主细胞产生的变体脂解酶分泌进培养基中。编码利于纯化的结构域的核酸序列可用于促进可溶性蛋白质的纯化。包含编码变体脂解酶的多核苷酸序列的载体或DNA构建体，还可以包含编码利于纯化的结构域的核酸序列以促进该变体脂解酶的纯化(参见例如Kroll等人，DNACellBiol.12：441-53[1993])。此类利于纯化的结构域包括但不限于，例如金属螯合肽，如允许在固定化金属上纯化的组氨酸-色氨酸模块(参见Porath，ProteinExpr.Purif.3：263-281[1992])、允许在固定化免疫球蛋白上纯化的蛋白A结构域，以及用于FLAGS延伸/亲和纯化系统的结构域(例如，得自ImmunexCorp.，Seattle，WA的蛋白A结构域)。在纯化结构域和异源蛋白质之间包括可剪切的接头序列如XA因子或肠激酶(例如，得自美国加利福尼亚州圣地亚哥市的英杰公司(Invitrogen，SanDiego，CA)的序列)也可以用于促进纯化。
用于检测和测量酶(如本发明的变体脂解酶)的酶活性的测定法是熟知的。各种用于检测和测量脂解酶(例如，本发明的变体脂解酶)活性的测定法也是本领域的普通技术人员已知的。如本文所用，可以根据本领域中已知的任何工序测定脂解活性。例如，诸如三酰基甘油脂解的凝胶扩散测定法、使用恒定pH方法测量脂肪酸释放量的滴定测量、使用分光光度法测量从对硝基苯酯释放的对硝基苯酚量、以及ELISA测定法等测定法可用于确定脂肪酶活性和/或特异性(参见例如Gupta等人，Biotechnol.Appl.Biochem，37：63-71，2003)。其他测定法可见于例如美国专利5,990,069；以及国际公布WO96/18729A1。
多种方法可用于测定成熟脂解酶(例如，本发明的成熟变体脂解酶)在宿主细胞中的产生水平。这类方法包括但不限于例如利用脂解酶特异性的多克隆或单克隆抗体的方法。示例性方法包括但不限于酶联免疫吸附测定法(ELISA)、放射性免疫测定法(RIA)、荧光免疫测定法(FIA)、以及荧光激活细胞分选法(FACS)。这些和其他测定法是本领域中所熟知的(参见例如Maddox等人，J.Exp.Med.158：1211[1983])。
在一些其他实施方案中，本发明提供用于制备或产生本发明的成熟变体脂解酶的方法。成熟变体脂解酶不包含信号肽或前肽序列。一些方法包括在重组细菌宿主细胞，例如芽孢杆菌属物种细胞(如，枯草芽孢杆菌细胞)中制备或产生本发明的变体脂解酶。在一些实施方案中，本发明提供产生本发明的变体脂解酶的方法，该方法包括在有利于产生变体脂解酶的条件下培养包含重组表达载体的重组宿主细胞，该重组表达载体包含编码本发明的变体脂解酶的核酸。一些这类方法还包括从培养物回收变体脂解酶。
在一些实施方案中，本发明提供产生本发明的变体脂解酶的方法，该方法包括：(a)将包含编码本发明变体脂解酶的核酸的重组表达载体引入细胞群(例如，细菌细胞，如枯草芽孢杆菌细胞)；以及(b)在有利于产生由该表达载体编码的变体脂解酶的条件下在培养基中培养该细胞。一些这类方法还包括：(c)从所述细胞或从所述培养基分离所述变体脂解酶。
织物和家居护理产品
在一些实施方案中，本发明的变体脂解酶可用于包含辅助材料和变体脂解酶的组合物中，其中该组合物为织物和家居护理产品。合适组合物的示例在实施例1中有所描述。
在一些实施方案中，含有至少一种变体脂解酶的织物和家居护理产品组合物包含以下成分中的一种或多种(基于组合物总重量计)：约0.0005重量％至约0.5重量％、约0.001重量％至约0.1重量％、或甚至约0.002重量％至约0.05重量％的所述脂解酶变体；以及以下一种或多种：约0.00003重量％至约0.1重量％的织物调色剂、约0.001重量％至约5重量％的香料囊、约0.001重量％至约1重量％的冷水可溶性增白剂、约0.00003重量％至约0.1重量％的漂白催化剂、约0.00003重量％至约0.1重量％的细菌清洁纤维素酶、和/或约0.05重量％至约20重量％的格尔贝特(Guerbet)非离子表面活性剂。
如本文所用，脂解酶(例如，本发明的变体脂解酶)的“洗涤性能”是指该脂解酶对洗涤的贡献，即与未向组合物中添加该变体脂解酶的洗涤剂相比，其为洗涤剂提供额外的清洁性能。对洗涤性能在相关洗涤条件下进行比较。在一些测试系统中，其他相关因素(如洗涤剂组成、泡沫浓度、水硬度、洗涤力学、时间、pH和/或温度)可以通过模拟某细分市场中家居应用(例如，手工或人工盘碟洗涤、自动盘碟洗涤、盘碟清洁、餐具清洁、织物清洁等)的典型条件进行控制。
在一些实施方案中，织物和家居护理产品组合物是颗粒状或粉末衣物洗涤剂。
在一些实施方案中，织物和家居护理产品组合物是液体衣物洗涤剂或盘碟洗涤剂。
意在使织物和家居护理产品以任何合适的形式(包括流体或固体)提供。织物和家居护理产品可以是单位剂量小袋的形式，尤其是液体的形式时，并且通常该织物和家居护理产品至少部分、或甚至完全被水溶性小袋封闭。此外，在包含至少一种变体脂解酶的织物和家居护理产品的一些实施方案中，织物和家居护理产品可以具有上面详述的参数和/或特性的任何组合。
清洁组合物
清洁组合物和清洁制剂包括适于对任何物体、物品和/或表面进行清洁、漂白、消毒和/或灭菌的任何组合物。此类组合物和制剂包括但不限于例如：液体和/或固体组合物，包括清洁或洗涤剂组合物(例如，液态、片状、凝胶状、条棒状、颗粒状和/或固态衣物清洁或洗涤剂组合物及精细织物洗涤剂组合物)；硬质表面清洁组合物和制剂，例如用于玻璃、木材、陶瓷和金属台面与窗户；地毯清洁剂；烘箱清洁剂；织物清新剂；织物软化剂；以及纺织物、衣物助洗剂清洁或洗涤剂组合物、衣物添加剂清洁组合物和衣物预洗剂清洁组合物；盘碟洗涤组合物，包括手动或人工盘碟洗涤组合物(例如，“手动”或“人工”盘碟洗涤剂)和自动盘碟洗涤组合物(例如，“自动盘碟洗涤剂”)。
除非另外指明，否则如本文所用的清洁组合物或清洁制剂包括：颗粒或粉末形式的多功能或重垢型洗涤剂，尤其是清洁洗涤剂；液态、颗粒状、凝胶状、固态、片状、或糊状形式的多功能洗涤剂，尤其是所谓的重垢型液体(HDL)洗涤剂或重垢型粉末洗涤剂(HDD)类型；液体精细织物洗涤剂；手工或人工盘碟洗涤剂，包括那些高泡类型；手工或人工盘碟洗涤剂、自动盘碟洗涤剂、或盘碟或餐具洗涤剂，包括用于家居和公共场所用途的多种片状、粉末状、固态、颗粒状、液态、凝胶状和漂洗助剂类型；液体清洁和消毒剂，包括抗菌手洗类型、清洁棒、漱口水、假牙清洁剂、汽车清洗剂、地毯清洗剂、浴室清洁剂；人和其他动物的洗发剂和/或毛发清洗剂；沐浴露和泡沫浴剂和金属清洁剂；以及洗涤助剂，例如漂白添加剂和“去污棒”或预处理类型。在一些实施方案中，颗粒状组合物为“致密”(compact)形式；在一些实施方案中，液体组合物为“浓缩”形式。
如本文所用，术语“洗涤剂组合物”或“洗涤剂制剂”用于指旨在用于洗涤介质中用来清洁染污的或脏的物体(包括特定的织物和/或非织物物体或物品)的组合物。本发明的这类组合物不限于任何特定的洗涤剂组合物或制剂。实际上，在一些实施方案中，本发明的洗涤剂包含至少一种本发明的变体脂解酶，此外还包含一种或多种表面活性剂、转移酶、水解酶、氧化还原酶、助洗剂(例如，助洗剂盐)、漂白剂、漂白活化剂、上蓝剂、荧光染料、结块抑制剂、掩蔽剂、酶活化剂、抗氧化剂和/或增溶剂。在某些情况下，助洗剂盐是硅酸盐和磷酸盐的混合物，优选硅酸盐(例如，偏硅酸钠)比磷酸盐(例如，三聚磷酸钠)多。一些本发明的组合物(例如但不限于清洁组合物或洗涤剂组合物)不含有任何磷酸盐(例如，磷酸盐或磷酸盐助洗剂)。
除非另外指明，否则本文提供的所有组分或组合物水平都是指所述组分或组合物的活性水平，而且不包括市售来源中可能存在的杂质(例如残余溶剂或副产物)。酶组分重量基于总活性蛋白计。除非另外指明，否则所有百分比和比率都按重量计算。除非另外指明，否则所有百分比和比率都基于总组合物计算。在例示的洗涤剂组合物中，酶水平是以纯酶占总组合物的重量比表示，并且除非另外指明，否则洗涤剂成分是以占总组合物的重量比表示。
如本文所指出，在一些实施方案中，本发明的清洁组合物还包含辅助材料，包括但不限于：表面活性剂、助洗剂、漂白剂、漂白活化剂、漂白催化剂、其他酶、酶稳定系统、螯合剂、荧光增白剂、去垢聚合物(soilreleasepolymers)、染料转移剂、分散剂、抑泡剂、染料、香料、着色剂、填料盐、水溶助长剂、光活化剂、荧光剂、织物调理剂、可水解性表面活性剂、防腐剂、抗氧化剂、抗收缩剂、抗皱剂、杀菌剂、杀真菌剂、色粒、银护理剂(silvercare)、抗晦暗剂和/或抗腐蚀剂、碱度源、增溶剂、载体、加工助剂、颜料和pH控制剂(参见例如，美国专利6,610,642、6,605,458、5,705,464、5,710,115、5,698,504、5,695,679、5,686,014和5,646,101，所有这些专利均以引用方式并入本文)。特定清洁组合物材料的实施方案在下文中详细举例说明。在清洁辅助材料与本发明的变体脂解酶在清洁组合物中不相容的实施方案中，则使用保持清洁辅助材料与所述脂解酶分离(即，相互不接触)直到这两种组分的组合恰当时的合适方法。这种分离方法包括本领域已知的任何合适方法(例如，囊形片(gelcap)、包封法、片剂法、物理分离法等)。
有利地，本发明的清洁组合物用于例如衣物洗涤应用、硬质表面清洁、盘碟洗涤应用，以及化妆应用，如假牙、牙齿、毛发和皮肤。此外，归因于在低温溶液中效力增加的独特优势，本发明的酶理想地适合于衣物洗涤应用。此外，本发明的酶可以用于颗粒状和液体组合物。
本发明的变体脂解酶也可以用于清洁添加剂产品。在一些实施方案中，用于低温溶液清洁应用。在一些实施方案中，本发明提供了包含本发明的至少一种酶的清洁添加剂产品，当需要额外漂白效力时，该酶非常适于添加在洗涤过程中。这种情况包括但不限于低温溶液清洁应用。在一些实施方案中，添加剂产品为其最简单形式，即一种或多种脂解酶。在一些实施方案中，添加剂被包装成用于添加至清洁过程的剂型。在一些实施方案中，添加剂被包装成用于添加到采用过氧源并且需要增加的漂白效力的清洁过程中的剂型。任何合适的单一剂量单位形式都可用于本发明，包括但不限于：小丸、小片、囊形片，或其他单一剂量单位例如预先测量的粉末或液体。在一些实施方案中，此类组合物包括填料或载体材料以增大其体积。合适的填料或载体材料包括但不限于多种硫酸盐、碳酸盐和硅酸盐，以及滑石、粘土等。合适的液体组合物填料或载体材料包括但不限于水或低分子量伯醇和仲醇(包括多元醇和二醇)。此类醇的示例包括但不限于甲醇、乙醇、丙醇和异丙醇。在一些实施方案中，组合物含有约5％至约90％的这类材料。酸性填料可用于降低pH。或者，在一些实施方案中，清洁添加剂包含辅助成分，如下文所更全面描述的。
本发明清洁组合物和清洁添加剂需要有效量的单独的或与其他脂解酶和/或附加的酶进行组合的至少一种本文所提供的变体脂解酶。所需的酶水平通过添加一种或多种本发明的变体脂解酶来实现。通常，本发明清洁组合物包含至少约0.0001重量％、约0.0001重量％至约10重量％、约0.001重量％至约1重量％或甚至约0.01重量％至约0.1重量％的至少一种本发明变体脂解酶。
本文的清洁组合物通常配制为使得在水性清洁操作中使用期间，洗涤水具有约5.0至约11.5的pH，或约6.0至8.0的pH，或甚至约7.5至约10.5的pH。液体产品制剂通常配制成具有约3.0至约9.0或甚至约3至约8的净pH。颗粒状衣物洗涤用产品通常配制成具有约6至约11或甚至约8至约10的pH。用于将pH控制在推荐的使用水平的技术包括使用缓冲液、碱、酸等，并且这些技术是本领域技术人员所熟知的。
合适的“低pH清洁组合物”通常具有约3至约8的净pH，并且通常不含在此类pH环境中水解的表面活性剂。此类表面活性剂包括烷基硫酸钠表面活性剂，该表面活性剂包含至少一个环氧乙烷部分或甚至约1至约16摩尔的环氧乙烷。这类清洁组合物通常包含足量的pH调节剂，如氢氧化钠、单乙醇胺或盐酸，从而得到具有约3至约8的净pH的这类清洁组合物。此类组合物通常包含至少一种酸稳定性酶。在一些实施方案中，所述组合物为液体，而在其他实施方案中，其为固体。此类液体组合物的pH通常是以净pH度量。此类固体组合物的pH是以所述组合物的10％固形物溶液度量，其中溶剂是蒸馏水。在这些实施方案中，除非另外指明，否则所有pH测量值均在20℃下取得。
在一些实施方案中，当颗粒组合物或液体中采用本发明的变体脂解酶时，期望变体脂解酶为封装颗粒的形式，以在存储过程中保护变体脂解酶免受该颗粒组合物的其他组分的影响。此外，封装还是控制变体脂解酶在清洁过程期间的利用度的方式。在一些实施方案中，封装可增强变体脂解酶和/或附加酶的性能。就这一点而言，本发明的变体脂解酶可以使用本领域已知的任何合适的封装材料封装。在一些实施方案中，封装材料通常封装本发明的变体脂解酶的催化剂的至少一部分。通常，封装材料具有水溶性和/或水分散性。在一些实施方案中，封装材料具有0℃或更高的玻璃化转变温度(Tg)。在WO97/11151中对玻璃化转变温度进行了更详细的描述。封装材料通常选自碳水化合物、天然的或合成的树胶、甲壳质、脱乙酰壳多糖、纤维素和纤维素衍生物、硅酸盐、磷酸盐、硼酸盐、聚乙烯醇、聚乙二醇、石蜡、以及它们的组合。当封装材料为碳水化合物时，其通常选自单糖、低聚糖、多糖、以及它们的组合。在一些典型的实施方案中，封装材料为淀粉(参见例如，EP0922499、US4,977,252、US5,354,559和US5,935,826)。在一些实施方案中，封装材料是由塑料例如热塑性塑料、丙烯腈、甲基丙烯腈、聚丙烯腈、聚甲基丙烯腈以及它们的混合物制成的微球体；适用的市售微球体包括但不限于以(瑞典斯托克韦斯文肯市(Stockviksverken，Sweden))和PM6545、PM6550、PM7220、PM7228、和(宾夕法尼亚州瓦利福奇市的PQ公司(PQCorp.，ValleyForge，PA))供应的那些微球体。
如本文所述，本发明的变体脂解酶尤其可用于清洁行业，包括但不限于衣物和盘碟洗涤剂。这些应用将酶置于多种环境胁迫下。本发明的变体脂解酶由于在多种条件下的稳定性而提供了优于许多当前所用酶的优点。
实际上，参与洗涤的脂解酶会暴露于多种洗涤条件，包括变化的洗涤剂配方、洗涤水体积、洗涤水温度和洗涤时间长度。此外，不同地理区域中使用的洗涤剂配方在洗涤水中存在不同浓度的其相关组分。例如，欧洲的洗涤剂通常在洗涤水中具有约2000至9000ppm的洗涤剂组分，而日本的洗涤剂通常在洗涤水中具有大约500至1500ppm的洗涤剂组分。在北美，尤其是在美国，洗涤剂通常在洗涤水中具有约975ppm的洗涤剂组分。
低洗涤剂浓度系统包括在洗涤水中存在少于约800ppm的洗涤剂组分的洗涤剂。日本洗涤剂通常被认为是低洗涤剂浓度系统，因为其在洗涤水中具有大约667ppm的洗涤剂组分。
中等洗涤剂浓度系统包括在洗涤水中存在介于约800ppm和约2000ppm之间的洗涤剂组分的洗涤剂。北美洗涤剂一般被认为是中等洗涤剂浓度系统，因为其在洗涤水中具有大约975ppm的洗涤剂组分。巴西通常在洗涤水中具有大约1500ppm的洗涤剂组分。
高洗涤剂浓度系统包括在洗涤水中存在超过约2000ppm的洗涤剂组分的洗涤剂。欧洲洗涤剂一般被认为是高洗涤剂浓度系统，因为其在洗涤水中具有大约4500至5000ppm的洗涤剂组分。
拉丁美洲的洗涤剂一般是高泡磷酸盐助洗剂洗涤剂，而且拉丁美洲使用的洗涤剂的范围可在中等洗涤剂浓度至高洗涤剂浓度之间，因为其在洗涤水中具有1500ppm至6000ppm的洗涤剂组分。如上所述，巴西通常在洗涤水中具有大约1500ppm的洗涤剂组分。然而，其他高泡磷酸盐助洗剂洗涤剂地理位置(不限于其他拉丁美洲国家)也可具有高洗涤剂浓度系统，即，在洗涤水中存在最高达约6000ppm的洗涤剂组分。
按照上文所述，很明显在全世界范围内，典型洗涤溶液中洗涤剂组合物的浓度在低于约800ppm(“低洗涤剂浓度地理位置”；例如在日本约667ppm)至介于约800ppm至约2000ppm之间(“中等洗涤剂浓度地理位置”；例如在美国约975ppm，以及在巴西约1500ppm)再到高于约2000ppm(“高洗涤剂浓度地理位置”；例如在欧洲约4500ppm至约5000ppm，以及在高泡磷酸盐助洗剂地理位置约6000ppm)的洗涤剂组合物间变动。
典型的洗涤溶液的浓度凭经验确定。例如，在美国，典型洗衣机容纳约64.4L体积的洗涤溶液。因此，为了在洗涤溶液内获得约975ppm的洗涤剂浓度，必须将约62.79g洗涤剂组合物添加至64.4L洗涤溶液中。该量是由消费者使用随洗涤剂一起提供的量杯计量加入洗涤水中的典型量。
再举个例子，不同地理位置使用不同的洗涤温度。日本的洗涤水温度通常低于欧洲所用的洗涤水温度。例如，北美和日本的洗涤水温度通常介于约10℃至约30℃之间(例如，约20℃)，而欧洲的洗涤水温度通常介于约30℃至约60℃之间(例如，约40℃)。然而，为了节省能源，许多消费者转向使用冷水洗涤。此外，在一些另外的区域中，通常将冷水用于衣物洗涤以及盘碟洗涤应用。在一些实施方案中，本发明的“冷水洗涤”利用适于在以下温度进行洗涤的“冷水洗涤剂”：约10℃至约40℃，或约20℃至约30℃，或约15℃至约25℃，以及在约15℃至约35℃范围内的所有其他温度组合、和在10℃至40℃内的所有温度范围。
再举个例子，不同地理位置通常具有不同的水硬度。水硬度通常以每加仑中混合的Ca2+/Mg2+的格令数来描述。硬度是水中钙(Ca2+)和镁(Mg2+)的量的量度。在美国，大多数水都比较硬，但硬度有波动。中等硬(60-120ppm)至硬(121-181ppm)水具有60至181ppm(ppm换算成每美制加仑格令数是ppm数除以17.1等于每加仑格令数)的硬度矿物质。
水 每加仑格令数 ppm
软 小于1.0 小于17
略硬 1.0至3.5 17至60
中等硬 3.5至7.0 60至120
硬 7.0至10.5 120至180
非常硬 大于10.5 大于180
欧洲的水硬度通常是每加仑中混合Ca2+/Mg2+高于约10.5(例如约10.5至约20.0)格令(例如每加仑的混合Ca2+/Mg2+为约15格令)。北美的水硬度通常高于日本的水硬度，但低于欧洲的水硬度。例如，北美的水硬度可介于约3至约10格令、约3至约8格令之间或为约6格令。日本的水硬度通常低于北美的水硬度，通常小于约4，例如每加仑混合的Ca2+/Mg2+为约3格令。
因此，在一些实施方案中，本发明提供的变体脂解酶在至少一组洗涤条件(例如水温、水硬度和/或洗涤剂浓度)下显示出惊人的洗涤性能。在一些实施方案中，本发明的变体脂解酶与其他脂肪酶脂解酶的洗涤性能相当。在一些实施方案中，与目前市售的脂肪酶脂解酶相比，本发明的变体脂解酶表现出增强的洗涤性能。因此，在本发明的一些实施方案中，本文提供的变体脂解酶表现出增强的氧化稳定性、增强的热稳定性、增强的在各种条件下的清洁能力和/或增强的螯合剂稳定性。此外，本发明的变体脂解酶(单独或与助洗剂和稳定剂组合)用于不包含洗涤剂的清洁组合物。
在本发明的一些实施方案中，清洁组合物包含以组合物的重量计水平为约0.00001％至约10％的至少一种本发明变体脂解酶，以及以组合物的重量计余量(例如约99.999％至约90.0％)包含清洁辅助材料。在本发明的一些其他实施方案中，本发明的清洁组合物包含以组合物的重量计水平为约0.0001％至约10％、约0.001％至约5％、约0.001％至约2％、约0.005％至约0.5％的至少一种变体脂解酶，清洁组合物的余量(例如，约99.9999重量％至约90.0重量％、约99.999重量％至约98重量％、约99.995重量％至约99.5重量％)包含清洁辅助材料。
在一些实施方案中，本发明的清洁组合物包含如上文所述的变体脂解酶作为主要的酶组分，例如在单组分组合物中。在一些实施方案中，本发明的清洁组合物包含一种或多种另外的洗涤剂酶，该酶提供清洁性能和/或织物护理和/或盘碟洗涤益处。合适的酶的示例包括但不限于：蛋白酶、过水解酶、半纤维素酶、纤维素酶、过氧化物酶、脂解酶、木聚糖酶、脂肪酶、磷脂酶、酯酶、角质酶、果胶酶、果胶酸裂解酶、甘露聚糖酶、角蛋白酶、还原酶、氧化酶、酚氧化酶、脂氧合酶、木质素酶、支链淀粉酶、鞣酸酶、戊聚糖酶、麦拉宁酶、β-葡聚糖酶、阿拉伯糖苷酶、透明质酸酶、软骨素酶、漆酶和淀粉酶以及它们的任何组合或混合物。在一些实施方案中，使用包含常规应用性酶的酶组合(即“混合物”)，如使用脂解酶、脂肪酶、角质酶和/或纤维素酶与淀粉酶组合。
例如，本发明的脂解酶变体可与蛋白酶组合使用。合适的蛋白水解酶包括源于动物、植物或微生物的蛋白水解酶。在一些实施方案中，使用微生物蛋白水解酶。在一些实施方案中，蛋白水解酶优选为碱性微生物蛋白水解酶或胰蛋白酶样蛋白水解酶。碱性脂解酶的示例包括脂肪酶，尤其是源自芽孢杆菌属的那些(例如，迟缓芽孢杆菌、解淀粉芽孢杆菌的Carlsberg、309、147和168)。另外的示例包括在美国专利RE34,606、5,955,340、5,700,676、6,312,936、和6,482,628中所述的那些突变蛋白水解酶，这些专利均以引用方式并入本文。另外的蛋白酶例子包括但不限于胰蛋白酶(例如源于猪或牛的胰蛋白酶)和WO89/06270中所述的镰孢属(Fusarium)蛋白酶。在一些实施方案中，可用于本发明的市售蛋白酶包括但不限于 OXP、PURAMAXTM、EXCELLASETM和PURAFASTTM(Genencor)； DURAZYMTM、 和(诺维信公司(Novozymes))；BLAPTM和BLAPTM变体(汉高股份两合公司(HenkelKommanditgesellschaftaufAktien，Duesseldorf，Germany))和KAP(嗜碱芽孢杆菌脂肪酶；日本东京市的花王公司(KaoCorp.，Tokyo，Japan))。多种蛋白水解酶在WO95/23221、WO92/21760、美国专利公布2008/0090747和美国专利5,801,039、5,340,735、5,500,364、5,855,625、USRE34,606、5,955,340、5,700,676、6,312,936和6,482,628、以及多种其他专利中有所描述。在一些另外的实施方案中，金属蛋白酶应用于本发明，包括但不限于WO07/044993中所述的中性金属蛋白酶。
在本发明的一些实施方案中，本发明的清洁组合物还包含以组合物的重量计蛋白酶水平为约0.00001％至约10％的蛋白酶，和以组合物的重量计余量的清洁辅助材料。在本发明的一些其他实施方案中，本发明的清洁组合物还包含以组合物的重量计蛋白酶水平为约0.0001％至约10％、约0.001％至约5％、约0.001％至约2％、约0.005％至约0.5％的蛋白酶。
在一些实施方案中，本发明的脂解酶变体可与淀粉酶组合使用。在本发明的一些实施方案中，任何合适的淀粉酶均用于本发明。在一些实施方案中，还使用适用于碱性溶液的任何淀粉酶(例如，α和/或β淀粉酶)。合适的淀粉酶包括但不限于来源于细菌或真菌的淀粉酶。一些实施方案包括经化学或遗传修饰的突变体。用于本发明的淀粉酶，包括但不限于从地衣芽孢杆菌获得的α淀粉酶(参见例如，GB1,296,839)。用于本发明的市售淀粉酶，包括但不限于STAINZYMESTAINZYMEBANTM(诺维信公司)，以及POWERASETM、和P(杰能科公司(Genencor))。
在本发明的一些实施方案中，本发明的清洁组合物还包含以组合物的重量计淀粉酶水平为约0.00001％至约10％的附加淀粉酶，和以组合物的重量计余量的清洁辅助材料。在本发明的一些其他实施方案中，本发明的清洁组合物还包含以组合物的重量计淀粉酶水平为约0.0001％至约10％、约0.001％至约5％、约0.001％至约2％、约0.005％至约0.5％的淀粉酶。
在一些另外的实施方案中，任何合适的纤维素酶均用于本发明的清洁组合物中。合适的纤维素酶包括但不限于来源于细菌或真菌的纤维素酶。一些实施方案包括经化学或遗传修饰的突变体。合适的纤维素酶包括但不限于特异腐质霉(Humicolainsolens)纤维素酶(参见例如，美国专利4,435,307)。特别合适的纤维素酶是具有颜色护理有益效果的纤维素酶(参见例如，EP0495257)。用于本发明的市售纤维素酶，包括但不限于(诺维信公司)和KAC-500(B)TM(花王公司)。在一些实施方案中，纤维素酶作为成熟野生型纤维素酶或变体纤维素酶的部分或片段掺入，其中N末端的一部分缺失(参见例如，美国专利5,874,276)。在一些实施方案中，本发明的清洁组合物还包含以组合物的重量计纤维素酶水平为约0.00001％至约10％的额外纤维素酶，以组合物的重量计余量的清洁辅助材料。在本发明的一些其他实施方案中，本发明的清洁组合物还包含以组合物的重量计纤维素酶水平为约0.0001％至约10％、约0.001％至约5％、约0.001％至约2％、约0.005％至约0.5％的纤维素酶。
适用于洗涤剂组合物中的任何甘露聚糖酶也可以用于本发明中。合适的甘露聚糖酶包括但不限于来源于细菌或真菌的甘露聚糖酶。一些实施方案包括经化学或遗传修饰的突变体。已知多种可以用于本发明的甘露聚糖酶(参见例如，美国专利6,566,114，美国专利6,602,842和美国专利6,440,991，这些专利均以引用方式并入本文)。在一些实施方案中，本发明的清洁组合物还包含以组合物的重量计甘露聚糖酶水平为约0.00001％至约10％的额外甘露聚糖酶，以组合物的重量计余量的清洁辅助材料。在本发明的一些实施方案中，本发明的清洁组合物还包含以组合物的重量计甘露聚糖酶水平为约0.0001％至约10％、约0.001％至约5％、约0.001％至约2％、约0.005％至约0.5％的甘露聚糖酶。
在一些实施方案中，在本发明的组合物中组合使用过氧化物酶与过氧化氢或其来源(例如，过碳酸盐、过硼酸盐或过硫酸盐)。在一些可供选择的实施方案中，组合使用氧化酶与氧。这两种类型的酶，优选连同增强剂一起，用于“溶液漂白”(即，当多个织物一起在洗涤液中洗涤时防止纺织品染料由染色的织物转移到另一织物)(参见例如，WO94/12621和WO95/01426)。合适的过氧化物酶/氧化酶包括但不限于来源于植物、细菌或真菌的过氧化物酶/氧化酶。一些实施方案包括经化学或遗传修饰的突变体。在一些实施方案中，本发明的清洁组合物还包含以组合物的重量计过氧化物酶和/或氧化酶水平为约0.00001％至约10％的额外过氧化物酶和/或氧化酶，以组合物的重量计余量的清洁辅助材料。在本发明的一些其他实施方案中，本发明的清洁组合物还包含以组合物的重量计过氧化物酶和/或氧化酶水平为约0.0001％至约10％、约0.001％至约5％、约0.001％至约2％、约0.005％至约0.5％的过氧化物酶和/或氧化酶。
在一些实施方案中，还使用附加的酶，包括但不限于过水解酶(参见例如，WO05/056782)。此外，在一些实施方案中，本文涵盖上述酶的混合物，尤其是一种或多种附加的脂解酶、淀粉酶、蛋白酶、甘露聚糖酶和/或至少一种纤维素酶。实际上，预期这些酶的各种混合物将应用于本发明。还预期，变体脂解酶与一种或多种附加的酶的变动水平可以独立地变动到约10％，该清洁组合物的余量为清洁辅助材料。通过考虑待清洁表面、物品或织物以及使用(例如，通过使用洗涤剂)期间的清洁条件所需的组合物形式，清洁辅助材料的具体选择变得容易。
合适的清洁辅助材料的示例包括但不限于表面活性剂、助洗剂、漂白剂、漂白活化剂、漂白催化剂、其他酶、酶稳定系统、螯合剂、荧光增白剂、去垢聚合物、染料转移剂、染料转移抑制剂、催化材料、过氧化氢、过氧化氢源、预先形成的过酸、聚合物分散剂、粘土污垢去除剂、结构增塑剂、分散剂、抑泡剂、染料、香料、着色剂、填料盐、水溶助长剂、光活化剂、荧光剂、织物调理剂、织物软化剂、载体、水溶助长剂、加工助剂、溶剂、颜料、可水解性表面活性剂、防腐剂、抗氧化剂、抗收缩剂、抗皱剂、杀菌剂、杀真菌剂、色粒、银护理剂、抗晦暗剂和/或抗腐蚀剂、碱度源、增溶剂、载体、加工助剂、颜料和pH控制剂(参见例如，美国专利6,610,642、6,605,458、5,705,464、5,710,115、5,698,504、5,695,679、5,686,014和5,646,101，所有这些专利均以引用方式并入本文)。特定清洁组合物材料的实施方案在下文中详细举例说明。在清洁辅助材料与本发明的变体脂解酶在清洁组合物中不相容的实施方案中，则使用保持清洁辅助材料与所述脂解酶分离(即，相互不接触)直到这两种组分适于组合在一起的合适方法。这种分离方法包括本领域已知的任何合适方法(例如，囊形片、包封法、片剂法、物理分离法等)。
在一些实施方案中，适用于清洁需要去除脂质污渍的多种表面的组合物中包含有效量的一种或多种本文提供的变体脂解酶。此类清洁组合物包括用于(例如)清洁硬质表面、织物和盘碟等应用的清洁组合物。实际上，在一些实施方案中，本发明提供织物清洁组合物，而在其他实施方案中，本发明提供非织物清洁组合物。预期本发明涵盖任何形式(即，液态、颗粒状、条棒状、半固态、凝胶状、乳液状、片状、胶囊状等)的洗涤剂组合物。
下文将以举例的方式更详细地描述应用本发明的变体脂解酶的几种清洁组合物。在本发明的清洁组合物配制成适用于洗衣机洗涤方法中的组合物的一些实施方案中，本发明组合物优选包含至少一种表面活性剂和至少一种助洗剂化合物，以及一种或多种清洁辅助材料，所述清洁辅助材料优选选自有机聚合化合物、漂白剂、附加的酶、抑泡剂、分散剂、钙皂分散剂、悬污剂和抗再沉积剂以及腐蚀抑制剂。在一些实施方案中，衣物洗涤组合物还含有软化剂(即，作为附加的清洁辅助材料)。本发明组合物还用于固体或液体形式的洗涤添加剂产品。此类添加剂产品旨在补充和/或增强常规洗涤剂组合物的性能，并可以在清洁过程的任何阶段添加。在一些实施方案中，在20℃测量时，本文中衣物洗涤剂组合物的密度在约400至约1200g/L的范围内，而在其他实施方案中，该组合物的密度在约500至约950g/L的范围内。
在配制为用于人工盘碟洗涤方法中的组合物的实施方案中，本发明组合物优选包含至少一种表面活性剂，以及优选至少一种附加的选自如下的清洁辅助材料：有机聚合化合物、泡沫增强剂、II族金属离子、溶剂、水溶助长剂和附加的酶。
在一些实施方案中，各种清洁组合物(例如)美国专利6,605,458中提供的组合物与本发明的变体脂解酶一起使用。因此，在一些实施方案中，包含至少一种本发明变体脂解酶的组合物是致密型颗粒状织物清洁组合物，而在其他实施方案中，所述组合物是用于洗涤有色织物的颗粒状织物清洁组合物，在另外的实施方案中，所述组合物是通过洗涤能力来提供软化的颗粒状织物清洁组合物，在其他实施方案中，所述组合物是重垢型液体织物清洁组合物。在一些实施方案中，包含至少一种本发明变体脂解酶的组合物是织物清洁组合物，例如美国专利6,610,642和6,376,450中所述的那些。此外，本发明的变体脂解酶可以用于在欧洲或日本洗涤条件下尤其有用的颗粒状衣物洗涤剂组合物中(参见例如，美国专利6,610,642)。
在一些可供选择的实施方案中，本发明提供包含本文提供的至少一种变体脂解酶的硬质表面清洁组合物。因此，在一些实施方案中，包含至少一种本发明变体脂解酶的组合物是硬质表面清洁组合物，例如美国专利6,610,642、6,376,450和6,376,450中所述的那些组合物。
在另一些实施方案中，本发明提供包含本文提供的至少一种变体脂解酶的盘碟洗涤组合物。因此，在一些实施方案中，包含至少一种本发明变体脂解酶的组合物是硬质表面清洁组合物，例如美国专利6,610,642和6,376,450中所述的那些。在一些另外的实施方案中，本发明提供包含本文提供的至少一种变体脂解酶的盘碟洗涤组合物。在一些另外的实施方案中，包含至少一种本发明变体脂解酶的组合物包括口腔护理组合物，例如美国专利6,376,450和6,376,450中所述的那些组合物。在上述美国专利6,376,450、6,605,458、6,605,458和6,610,642中所包含的化合物和清洁辅助材料的配制和描述可用于本文提供的变体脂解酶。
本发明的清洁组合物可以被配制成任何合适的形式并通过配制者所选择的任何方法制备，所述方法的非限制性例子在美国专利5,879,584、5,691,297、5,574,005、5,569,645、5,565,422、5,516,448、5,489,392和5,486,303中有所描述，所有这些专利均以引用方式并入本文。当需要低pH清洁组合物时，可以通过添加诸如单乙醇胺之类的材料或诸如HCl之类的酸性材料来调整这种组合物的pH。
尽管不是实现本发明目的所必需的，但下文示例的辅助材料的非限制性列表适用于本发明清洁组合物。在一些实施方案中，掺入这些辅助材料，例如以帮助或增强清洁性能以便处理待清洁的基底，或改变清洁组合物的美观性，如利用香料、着色剂、染料等情形就是如此。应当理解，这些辅助材料是本发明的变体脂解酶的补充。这些附加组分的精确性质以及其掺入水平将取决于组合物的物理形式和打算使用组合物的清洁操作的性质。合适的辅助材料包括但不限于：表面活性剂、助洗剂、螯合剂、染料转移抑制剂、沉积助剂、分散剂、附加的酶和酶稳定剂、催化材料、漂白活化剂、漂白增强剂、过氧化氢、过氧化氢源、预先形成的过酸、聚合物分散剂、粘土污垢去除剂/抗再沉积剂、增亮剂、抑泡剂、染料、香料、结构增塑剂、织物软化剂、载体、水溶助长剂、加工助剂和/或颜料。除下文的公开内容外，此类其他辅助材料及其使用量的合适例子见于美国专利5,576,282、6,306,812和6,326,348(以引用的方式并入)中。上述辅助成分可以构成本发明清洁组合物的余量。
在一些实施方案中，根据本发明的清洁组合物包含至少一种表面活性剂和/或表面活性剂体系，其中表面活性剂选自：非离子表面活性剂、阴离子表面活性剂、阳离子表面活性剂、两性表面活性剂、两性离子表面活性剂、半极性非离子表面活性剂以及它们的混合物。在一些低pH清洁组合物实施方案(例如，具有约3至约5净pH的组合物)中，组合物通常不含烷基乙氧基化硫酸盐，因为据信这种表面活性剂可能会被所述组合物的酸性内容物水解。在一些实施方案中，以清洁组合物的重量计，表面活性剂以约0.1％至约60％的水平存在，而在另外的实施方案中，所述水平为约1％至约50％，而在其他另外的实施方案中，所述水平为约5％至约40％。
在一些实施方案中，本发明的清洁组合物包含一种或多种洗涤剂助洗剂或助洗剂体系。在一些掺有至少一种助洗剂的实施方案中，以清洁组合物的重量计，清洁组合物包含至少约1％，约3％至约60％，或甚至约5％至约40％的助洗剂。助洗剂包括但不限于：聚磷酸的碱金属盐、铵盐和链烷醇铵盐；碱金属硅酸盐；碱土金属和碱金属碳酸盐；硅铝酸盐；聚羧酸盐化合物；醚羟基聚羧酸盐；马来酸酐与乙烯或乙烯基甲基醚的共聚物；1，3，5-三羟基苯-2，4，6-三磺酸；和羧甲基氧基琥珀酸；聚乙酸(如乙二胺四乙酸和次氮基三乙酸)的多种碱金属盐、铵盐和取代的铵盐；以及聚羧酸盐，如苯六甲酸、琥珀酸、柠檬酸、氧联二琥珀酸、聚马来酸、苯1，3，5-三甲酸、羧甲基氧基琥珀酸以及它们的可溶性盐。实际上，预期任何合适的助洗剂都可以应用于本发明的各种实施方案。
在一些实施方案中，助洗剂形成水溶性硬度离子络合物(例如，多价螯合助洗剂)，如柠檬酸盐和聚磷酸盐(例如，三聚磷酸钠和六水合三聚磷酸钠、三聚磷酸钾以及三聚磷酸钠与三聚磷酸钾的混合物等)。预期任何合适的助洗剂将应用于本发明，包括本领域已知的那些助洗剂(参见例如，EP2100949)。
在一些实施方案中，本发明的清洁组合物含有至少一种螯合剂。合适的螯合剂包括但不限于铜、铁和/或锰螯合剂以及它们的混合物。在使用至少一种螯合剂的实施方案中，以主题清洁组合物的重量计，本发明的清洁组合物包含约0.1％至约15％或甚至约3.0％至约10％的螯合剂。
在一些另外的实施方案中，本文中提供的清洁组合物含有至少一种沉积助剂。合适的沉积助剂包括但不限于：聚乙二醇；聚丙二醇；聚羧酸盐；去垢聚合物，如聚对苯二甲酸；粘土，如高岭土、蒙脱土、绿坡缕石(atapulgite)、伊利石、膨润土、多水高岭土；以及它们的混合物。
如本文所指出，在一些实施方案中，抗再沉积剂可用于本发明的一些实施方案中。在一些实施方案中，使用非离子表面活性剂。例如，在自动盘碟洗涤实施方案中，非离子表面活性剂可用于表面改性目的，尤其用于膜片化(sheeting)，以避免成膜和起斑以及用于改善光泽度。这些非离子表面活性剂还可以用于防止污垢再沉积。在一些实施方案中，抗再沉积剂是本领域已知的非离子表面活性剂(参见例如，EP2100949)。
在一些实施方案中，本发明的清洁组合物包含一种或多种染料转移抑制剂。合适的聚合物染料转移抑制剂包括但不限于：聚乙烯吡咯烷酮聚合物、聚胺N-氧化物聚合物、N-乙烯基吡咯烷酮与N-乙烯基咪唑的共聚物、聚乙烯唑烷酮和聚乙烯咪唑或它们的混合物。在使用至少一种染料转移抑制剂的实施方案中，以清洁组合物的重量计，本发明的清洁组合物包含约0.0001％至约10％、约0.01％至约5％或甚至约0.1％至约3％的染料转移抑制剂。
在一些实施方案中，本发明的组合物中包含硅酸盐。在一些此类实施方案中，使用硅酸钠(例如，二硅酸钠、偏硅酸钠和结晶页硅酸盐(phyllosilicate))。在一些实施方案中，硅酸盐以约1％至约20％的水平存在。在一些实施方案中，以组合物的重量计，硅酸盐以约5％至约15％的水平存在。
在一些另外的实施方案中，本发明的清洁组合物还包含分散剂。合适的水溶性有机材料包括但不限于均聚酸或共聚酸或它们的盐，其中多元羧酸包含至少两个彼此相隔不超过两个碳原子的羧基。
在一些另外的实施方案中，通过任何合适的技术使清洁组合物中使用的酶稳定。在一些实施方案中，本文所用的酶因成品组合物中存在水溶性的钙和/或镁离子源而稳定，其中所述水溶性离子源为酶提供这类离子。在一些实施方案中，酶稳定剂包括寡糖、多糖和无机二价金属盐(包括碱土金属盐，如钙盐)。预期用于稳定酶的各种技术将用于本发明。例如，在一些实施方案中，本文所采用的酶因成品组合物中存在为酶提供锌(II)、钙(II)和/或镁(II)离子的这类离子的水溶性来源，以及其他金属离子(例如，钡(II)、钪(II)、铁(II)、锰(II)、铝(III)、锡(II)、钴(II)、铜(II)、镍(II)和氧钒(IV))而稳定。氯化物和硫酸盐也可用于本发明的一些实施方案中。合适的寡糖和多糖(例如，糊精)的示例是本领域已知的(参见例如，WO07/145964)。在一些实施方案中，还可使用可逆性脂解酶抑制剂，如含硼化合物(例如，硼酸盐、4-甲酰基苯基硼酸)和/或必要时，使用三肽醛以进一步提高稳定性。
在一些实施方案中，漂白剂、漂白活化剂和/或漂白催化剂存在于本发明的组合物中。在一些实施方案中，本发明的清洁组合物包含无机和/或有机漂白化合物。无机漂白剂包括但不限于过氧化氢合物盐(例如，过硼酸盐、过碳酸盐、过磷酸盐、过硫酸盐和过硅酸盐)。在一些实施方案中，无机过氧化氢合物盐是碱金属盐。在一些实施方案中，包括的无机过氧化氢合物盐为结晶固体形式，无额外保护，但在一些其他实施方案中，所述盐带涂层。本领域已知的任何合适的盐均可以用于本发明(参见例如，EP2100949)。
在一些实施方案中，漂白活化剂用于本发明的组合物中。漂白活化剂通常为有机过酸前体，这些有机过酸前体在60℃和低于60℃温度下进行的清洁过程中会增强漂白作用。适用于本文中的漂白活化剂包括在过水解条件下产生脂族过氧羧酸和/或任选地取代的过苯甲酸的化合物，所述脂族过氧羧酸优选具有约1至约10个碳原子，尤其约2至约4个碳原子。附加的漂白活化剂是本领域已知的并且可以用于本发明中(参见例如，EP2100949)。
此外，在一些实施方案中以及如本文中进一步描述的，本发明的清洁组合物还包含至少一种漂白催化剂。在一些实施方案中，使用三氮杂环壬烷锰和相关络合物，以及钴、铜、锰和铁络合物。其他漂白催化剂可以用于本发明中(参见例如，US4,246,612、5,227,084、4,810410、WO99/06521和EP2100949)。
在一些实施方案中，本发明的清洁组合物含有一种或多种催化性金属络合物。在一些实施方案中，使用含金属的漂白催化剂。在一些实施方案中，金属漂白催化剂包含催化剂体系，所述催化剂体系包含具有确定的漂白催化活性的过渡金属阳离子(例如，铜、铁、钛、钌、钨、钼或锰阳离子)、具有极低或无漂白催化活性的辅助性金属阳离子(例如，锌或铝阳离子)，以及对催化性和辅助性金属阳离子具有确定的稳定性常数的多价螯合剂，尤其乙二胺四乙酸、乙二胺四(亚甲基膦酸)以及它们的水溶性盐(参见例如，美国专利4,430,243)。在一些实施方案中，借助锰化合物催化本发明的清洁组合物。此类化合物及其使用量是本领域熟知的(参见例如，美国专利5,576,282)。在另外的实施方案中，钴漂白催化剂用于本发明的清洁组合物中。各种钴漂白催化剂是本领域已知的(参见例如，美国专利5,597,936和5,595,967)，并且易于通过已知方法制备。
在一些另外的实施方案中，本发明的清洁组合物包括大多环刚性配体(MRL)的过渡金属络合物。作为一种实践(但绝非限制)，在一些实施方案中，调整本发明提供的组合物和清洁方法以便在水性洗涤介质中提供约至少一亿分之一的活性MRL物质，而在一些实施方案中，在洗涤液中提供约0.005ppm至约25ppm，更优选约0.05ppm至约10ppm，并且最优选约0.1ppm至约5ppm的MRL。
在一些实施方案中，本发明过渡金属漂白催化剂中的过渡金属包括但不限于锰、铁和铬。MRL还包括但不限于，交叉桥接的特定超刚性配体(例如，5，12-二乙基-1，5，8，12-四氮杂双环[6.6.2]十六烷)。合适的过渡金属MRL易于通过已知工序制备(参见例如，WO2000/32601和美国专利6,225,464)。
在一些实施方案中，本发明的清洁组合物包含金属护理剂。金属护理剂用于防止和/或减少包括铝、不锈钢和非铁金属(例如，银和铜)在内的金属的锈污、腐蚀和/或氧化。合适的金属护理剂包括EP2100949、WO9426860和WO94/26859中所述的金属护理剂。在一些实施方案中，金属护理剂是锌盐。在一些另外的实施方案中，本发明的清洁组合物包含约0.1重量％至约5重量％的一种或多种金属护理剂。
如上文所指出，本发明的清洁组合物可以被配制成任何合适的形式并通过配制者所选择的任何方法制备，所述方法的非限制性例子在美国专利5,879,584、5,691,297、5,574,005、5,569,645、5,516,448、5,489,392和5,486,303中有所描述，所有这些专利均以引用方式并入本文。在需要低pH清洁组合物的一些实施方案中，通过添加诸如HCl之类的酸性材料来调整这种组合物的pH。
本文所公开的清洁组合物可以用于原位清洁(例如，表面、物品、盘碟或织物)。通常，使所述位置的至少一部分与本发明清洁组合物的实施方案(纯的形式或稀释在洗涤液中的形式)接触，随后任选洗涤和/或冲洗所述位置。出于本发明的目的，“洗涤”包括但不限于擦洗和机械搅动。在一些实施方案中，清洁组合物在溶液中通常以约500ppm至约15,000ppm的浓度使用。当洗涤溶剂是水时，水温通常在约5℃至约90℃的范围内，并且当所述位置包括织物时，水与织物的质量比通常为约1∶1至约30∶1。
用于清洁的组合物
本发明的组合物和方法的一个方面是包括脂解酶作为组分的清洁组合物。在用于手洗、衣物洗涤、盘碟洗涤和其他硬质表面清洁的洗涤剂组合物中可使用脂解酶多肽作为组分。
在某些实施方案中，以洗涤剂中常规使用的脂解酶浓度或接近的浓度将脂解酶掺入到洗涤剂中。例如，可以对应于每升洗涤/盘碟洗涤液体0.00001至1mg(以纯酶蛋白计)脂解酶的量加入脂解酶多肽。本文提供了示例性的配方，例示如下：
脂解酶多肽可以是洗涤剂组合物中仅有的酶组分；或者，洗涤剂组合物除脂解酶多肽外，还包含其他酶(包括其他淀粉分解酶在内)。因此，它可以无尘颗粒、稳定的液体，或受保护的酶的形式包括在洗涤剂组合物中。例如，可如美国专利4,106,991和4,661,452中所公开的制备无尘颗粒，并且可任选以本领域已知的方法进行涂覆。蜡质涂覆材料的示例是具有平均摩尔质量1,000至20,000的聚(环氧乙烷)产品(聚乙二醇，PEG)；具有16至50的环氧乙烷单元的乙氧基化壬基酚；乙氧基化脂肪醇，其中醇含12至20个碳原子并存在15至80个环氧乙烷单元；脂肪醇；脂肪酸；单甘油脂肪酸酯和二甘油脂肪酸酯和三甘油脂肪酸酯。适合于应用流化床技术的成膜包衣材料的示例列于例如GB1483591中。例如，可根据已建立的方法，通过添加多元醇例如丙二醇、糖或糖醇、乳酸或硼酸，使液体酶制品稳定化。其他的酶稳定剂是本领域已知的。可按照在例如EP238216中所公开的方法制备保护的酶。多元醇早已被公认为蛋白质的稳定剂，以及提高蛋白质的溶解度。
所述洗涤剂组合物可为任何可用的形式，例如，粉末状、颗粒状、糊状或液态。液体洗涤剂可为含水的，其通常包含最高至约70％的水和0％至约30％的有机溶剂。它也可为仅含约30％水的致密凝胶类型的形式。
所述洗涤剂组合物包含一种或多种表面活性剂，其中的每种表面活性剂可为阴离子型、非离子型、阳离子型或两性离子型。洗涤剂一般含0％至约50％的阴离子型表面活性剂，例如直链烷基苯磺酸盐(LAS)、α-烯烃磺酸盐(AOS)、烷基硫酸盐(脂肪醇硫酸盐)(AS)、醇乙氧基硫酸盐(AEOS或AES)、仲烷基磺酸盐(SAS)、α-磺基脂肪酸甲酯、烷基或烯基琥珀酸，或者皂。洗涤剂组合物还可含0％至约40％的非离子型表面活性剂，例如醇乙氧基化物(AEO或AE)、羧基化醇乙氧基化物、壬基酚乙氧基化物、烷基多糖苷、烷基二甲基胺氧化物、乙氧基化脂肪酸单乙醇胺、脂肪酸单乙醇胺或多羟基烷基脂肪酸酰胺(例如WO92/06154中所述)。
所述洗涤剂组合物可还包含任意组合的一种或多种其他酶，诸如蛋白酶、其它淀粉分解酶、角质酶、脂肪酶、纤维素酶、果胶酸裂解酶、过水解酶、木聚糖酶、过氧化物酶、和/或漆酶。
洗涤剂组合物可包含约1％至约65％的洗涤剂助洗剂或络合剂，例如沸石、二磷酸盐、三磷酸盐、膦酸盐、柠檬酸盐、次氮基三乙酸(NTA)、乙二胺四乙酸(EDTA)、二亚乙基三胺五乙酸(DTMPA)、烷基或烯基琥珀酸、可溶性硅酸盐或层状硅酸盐(如，赫希司特出品的SKS-6)。洗涤剂也可以无助洗(unbuilt)，也就是说，基本上不含洗涤剂助洗剂。酶可用于与酶的稳定性相容的任何组合物中。一般来讲，酶可通过已知的封装形式来免于受到有害成分的影响，例如，通过制粒或隔离在水凝胶中。可以在多种组合物中使用酶，尤其是脂解酶，这些酶有无淀粉结合结构域均可；这些组合物包括用来衣物洗涤、洗涤盘碟、清洁表面的组合物，和以淀粉或生物质为原料用来生产乙醇的组合物。
洗涤剂可包含一种或多种聚合物。例子包括羧甲基纤维素(CMC)、聚乙烯吡咯烷酮(PVP)、聚乙二醇(PEG)、聚乙烯醇(PVA)、聚羧酸酯(如聚丙烯酸酯)、马来酸/丙烯酸共聚物、和甲基丙烯酸月桂酯/丙烯酸共聚物。
洗涤剂组合物可包含漂白体系，该漂白体系可含有H2O2源如过硼酸盐或过碳酸盐，这种物质可与形成过酸的漂白活化剂如四乙酰乙二胺(TAED)或壬酰氧基苯磺酸盐(NOBS)组合起效。或者，漂白体系可含有过氧酸(如酰胺、二酰亚胺或砜型的过氧酸)。漂白体系也可以是酶漂白体系，例如过水解酶，如国际PCT专利申请WO2005/056783中描述的过水解酶。
可用常规的稳定剂稳定洗涤剂组合物中的酶，这些稳定剂是例如多元醇，如丙二醇或甘油；糖或糖醇；乳酸；硼酸或硼酸衍生物，如芳香族硼酸酯；可按例如专利WO92/19709和WO92/19708中描述的方法配制组合物。
洗涤剂还可包含其他的常规洗涤剂成分，例如织物调理剂，包括粘土、泡沫促进剂、抑泡剂、耐腐蚀剂、污垢悬浮剂、抗污垢再沉积剂、染料、杀菌剂、晦暗抑制剂、荧光增白剂或香料。
pH(在使用浓度的水溶液中测得)通常为中性或碱性，例如pH为约7.0至约11.0。
可包含本发明的α-脂解酶的洗涤剂组合物的具体形式描述如下。为了便于使用，这些组合物中的许多可以以单位剂量形式提供。单位剂量制剂和包装描述在例如US20090209445A1，US20100081598A1，US7001878B2，EP1504994B1，WO2001085888A2，WO2003089562A1，WO2009098659A1，WO2009098660A1，WO2009112992A1，WO2009124160A1，WO2009152031A1，WO2010059483A1，WO2010088112A1，WO2010090915A1，WO2010135238A1，WO2011094687A1，WO2011094690A1，WO2011127102A1，WO2011163428A1，WO2008000567A1，WO2006045391A1，WO2006007911A1，WO2012027404A1，EP1740690B1，WO2012059336A1，US6730646B1，WO2008087426A1，WO2010116139A1，和WO2012104613A1。
重垢液体(HDL)衣物洗涤剂组合物
示例性的HDL衣物洗涤剂组合物包含去污表面活性剂(10重量％-40重量％)，包括阴离子去污表面活性剂(选自直链或支链或无规链的、取代或未取代的烷基硫酸盐、烷基磺酸盐、烷基烷氧基化硫酸盐、烷基磷酸盐、烷基膦酸盐、烷基羧酸盐、和/或它们的混合物)，和任选的非离子表面活性剂(选自直链或支链或无规链的、取代或未取代的烷基烷氧基化醇，例如C8-C18烷基乙氧基化醇和/或C6-C12烷基酚烷氧基化物)，其中阴离子去污表面活性剂(具有从6.0至9的亲水指数(HIc))与非离子去污表面活性剂的重量比大于1∶1。合适的去污表面活性剂也包括阳离子去污表面活性剂(选自烷基吡啶化合物、烷基季铵化合物、烷基季化合物、烷基叔锍化合物、和/或它们的混合物)；两性离子和/或兼性去污表面活性剂(选自链烷醇胺磺基甜菜碱)；两性表面活性剂；半极性非离子表面活性剂以及它们的混合物。
所述组合物可任选包含表面活性增强聚合物，其由以下物质组成：两亲性的烷氧基化油脂清洁聚合物(选自具有亲水性和疏水性支链的烷氧基化聚合物，诸如在0.05重量％至10重量％范围内的烷氧基化聚亚烷基亚胺)和/或无规接枝聚合物(通常包含亲水性主链，其包含选自下列的单体：不饱和的C1-C6羧酸、醚、醇、醛、酮、酯、糖单元、烷氧基单元、马来酸酐、饱和多元醇诸如甘油、以及它们的混合物；以及疏水侧链，其选自：C4-C25烷基基团、聚丙烯、聚丁烯、饱和C1-C6单羧酸的乙烯基酯、丙烯酸或甲基丙烯酸的C1-C6烷基酯、以及它们的混合物)。
所述组合物可包含另外的聚合物诸如去垢聚合物(包括阴离子封端的聚酯(例如SRP1)，无规或嵌段构型的、包含选自糖类、二元羧酸、多元醇以及它们的组合的至少一种单体单元的聚合物，基于对苯二甲酸乙二酯的聚合物以及其无规或嵌段构型的共聚物(例如Repel-o-texSF、SF-2和SRP6、TexcareSRA100、SRA300、SRN100、SRN170、SRN240、SRN300和SRN325、MarloquestSL))；抗再沉积聚合物(0.1重量％至10重量％，包括羧酸酯聚合物，诸如包含选自丙烯酸、马来酸(或马来酸酐)、延胡索酸、衣康酸、乌头酸、中康酸、柠康酸、亚甲基丙二酸和它们的任何混合物的至少一种单体的聚合物，乙烯基吡咯烷酮均聚物，和/或聚乙二醇，分子量在500至100,000Da的范围内)；纤维素聚合物(包括选自烷基纤维素、烷基烷氧基烷基纤维素、羧烷基纤维素、烷基羧烷基纤维素的纤维素聚合物，其例子包括羧甲基纤维素、甲基纤维素、甲基羟乙基纤维素、甲基羧甲基纤维素以及它们的混合物)和聚合羧酸盐(诸如马来酸/丙烯酸无规共聚物或聚丙烯酸均聚物)。
所述组合物可还包含饱和或不饱和的脂肪酸，例如，饱和或不饱和的C12-C24脂肪酸(0重量％至10重量％)；沉积助剂(其示例包括多糖，例如，纤维素聚合物，聚二烯丙基二甲基铵卤化物(DADMAC)，和无规或嵌段构造的DADMAC与乙烯基吡咯烷酮、丙烯酰胺、咪唑、咪唑卤化物以及它们的混合物的共聚物，阳离子瓜尔胶，阳离子纤维素诸如阳离子羟乙基纤维素等，阳离子淀粉，阳离子聚丙烯酰胺，以及它们的混合物。
所述组合物可还包括染料转移抑制剂，其示例包括锰酞菁，过氧化物酶，聚乙烯吡咯烷酮聚合物，多胺N-氧化物聚合物，N-乙烯基吡咯烷酮和N-乙烯基咪唑的共聚物，聚乙烯基唑烷酮和聚乙烯基咪唑和/或它们的混合物；螯合剂，其示例包括乙二胺四乙酸(EDTA)、二亚乙基三胺五亚甲基膦酸(DTPMP)、羟基乙烷二膦酸(HEDP)、乙二胺-N，N′-二琥珀酸(EDDS)、甲基甘氨酸二乙酸(MGDA)、二亚乙基三胺五乙酸(DTPA)、丙二胺四乙酸(PDTA)、2-羟基吡啶-N-氧化物(HPNO)、或甲基甘氨酸二乙酸(MGDA)、谷氨酸N，N-二乙酸(N，N-二羧甲基谷氨酸的四钠盐)(GLDA)、次氮基三乙酸(NTA)、4，5-二羟基间苯二磺酸、柠檬酸及其任何盐、N-羟乙基乙二胺三乙酸(HEDTA)、三亚乙基四胺六乙酸(TTHA)、N-羟乙基亚氨基二乙酸(HEIDA)、二羟乙基甘氨酸(DHEG)、乙二胺四丙酸(EDTP)，以及它们的衍生物。
所述组合物还可包含酶(大体约0.01重量％的活性酶至0.03重量％的活性酶)，其选自蛋白酶、淀粉酶、脂肪酶、纤维素酶、胆碱氧化酶、过氧化物酶/氧化酶、果胶酸裂解酶、甘露聚糖酶、角质酶、漆酶、磷脂酶、溶血磷脂酶、酰基转移酶、过氧水解酶、芳基酯酶以及它们的任何混合物。所述组合物可包括酶稳定剂(其示例包括多元醇诸如丙二醇或甘油，糖或糖醇，乳酸，可逆的蛋白酶抑制剂，硼酸或硼酸衍生物，例如，芳族硼酸酯、或苯基硼酸衍生物诸如4-甲酰基苯基硼酸)。
所述组合物任选包含硅氧烷或基于脂肪酸的抑泡剂；调色染料、钙离子和镁离子、可视信号成分、消泡剂(0.001重量％至约4.0重量％)和/或结构剂/增稠剂(0.01重量％至5重量％，选自甘油二酯和甘油三酯、乙二醇双硬脂酸酯、微晶纤维素、基于纤维素的材料、微纤纤维素、生物聚合物、黄原胶、结冷胶以及它们的混合物)。
所述组合物可为任何液体形式，例如液体或凝胶形式，或它们的任何组合。所述组合物可为任何单位剂型，例如小袋。
重垢型干燥/固体(HDD)衣物洗涤剂组合物
示例性的HDD衣物洗涤剂组合物包含去污表面活性剂，包括阴离子去污表面活性剂(例如，直链、支链或无规链的，取代或未取代的烷基硫酸盐、烷基磺酸盐、烷基烷氧基化硫酸盐、烷基磷酸盐、烷基膦酸盐、烷基羧酸盐和/或这些物质的混合物)、非离子型去污表面活性剂(例如，直链、支链或无规链的，取代或未取代的C8至C18烷基乙氧基化物和/或C6至C12烷基酚烷氧基化物)、阳离子型去污表面活性剂(例如，烷基吡啶化合物，烷基季铵化合物，烷基季化合物，烷基叔锍化合物，以及它们的混合物)、两性离子和/或兼性去污表面活性剂(例如，链烷醇胺磺基甜菜碱)、两性表面活性剂、半极性非离子表面活性剂以及这些表面活性剂的混合物；助洗剂，包括不含磷酸盐的助洗剂(例如，沸石助洗剂，其例子包括沸石A、沸石X、沸石P和沸石MAP，含量在0重量％至小于10重量％的范围内)、磷酸盐助洗剂(例如，三聚磷酸钠，含量在0重量％至小于10重量％的范围内)、柠檬酸、柠檬酸盐、次氮基三乙酸、硅酸盐(例如，硅酸钠、硅酸钾或偏硅酸钠，含量在0重量％至小于10重量％的范围内；或层状硅酸盐(SKS-6))；碳酸盐(例如，碳酸钠和/或碳酸氢钠，含量在0重量％至小于80重量％的范围内)；以及漂白剂，包括光致漂白剂(photobleach)(如磺化酞菁锌、磺化酞菁铝、氧杂蒽染料，和这些物质的混合物)、疏水性或亲水性漂白活化剂(如十二烷酰基羟苯磺酸盐、癸酰基羟苯磺酸盐、癸酰基羟苯甲酸或其盐、3，5，5-三甲基己酰基羟苯磺酸盐、四乙酰乙二胺(TAED)、壬酰基羟苯磺酸盐(NOBS)、腈季化物(nitrilequat)，和这些物质的混合物)、过氧化氢源(例如，无机过氧化氢合物盐，其例子包括过硼酸盐、过碳酸盐、过硫酸盐、过磷酸盐或过硅酸盐的一水合钠盐或四水合钠盐)、预先形成的亲水性和/或疏水性过酸(例如，过羧酸及其盐、过碳酸及其盐、过亚氨酸及其盐、过氧单硫酸及其盐，和这些物质的混合物)和/或漂白催化剂(例如亚胺漂白促进剂，示例包括亚铵阳离子和聚离子、亚铵两性离子、改性胺、改性氧化胺、N-磺酰基亚胺、N-膦酰基亚胺、N-酰基亚胺、噻二唑二氧化物、全氟亚胺；环糖酮以及它们的混合物；含金属的漂白催化剂，例如铜、铁、钛、钌、钨、钼或锰阳离子连同辅助金属阳离子诸如锌或铝和多价螯合剂诸如乙二胺四乙酸、乙二胺四(亚甲基膦酸)及其水溶性盐)。
这种组合物可包含酶，例如，蛋白酶、淀粉酶、脂肪酶、纤维素酶、胆碱氧化酶、过氧化物酶/氧化酶、果胶酸裂解酶、甘露聚糖酶、角质酶、漆酶、磷脂酶、溶血磷脂酶、酰基转移酶、过水解酶、芳基酯酶，和这些酶的任意混合物。
这种组合物可任选包含附加的洗涤剂成分，包括香料微囊、用淀粉包封的谐香剂、调色剂、另外的聚合物(包括维持织物完整性的聚合物和阳离子聚合物)、锁住染料的成分、织物软化剂、增白剂(例如C.I.荧光增白剂)、絮凝剂、螯合剂、烷氧基化聚胺、织物沉积助剂和/或环糊精。
自动盘碟洗涤(ADW)洗涤剂组合物
示例性的ADW洗涤剂组合物包括：含量从0重量％至10重量％的非离子表面活性剂，包括乙氧基化非离子型表面活性剂、醇烷氧基化表面活性剂、环氧封端的聚(氧烷基化)醇或氧化胺表面活性剂；含量在5重量％至60重量％范围内的助洗剂，包括磷酸盐助洗剂(例如单磷酸盐、二磷酸盐、三聚磷酸盐或其它寡聚磷酸盐，三聚磷酸钠(STPP))和无磷助洗剂(例如基于氨基酸的化合物，包括甲基甘氨酸二乙酸(MGDA)及其盐和衍生物、谷氨酸-N，N-二乙酸(GLDA)及其盐和衍生物、亚氨基二琥珀酸(IDS)及其盐和衍生物、羧甲基菊粉及其盐和衍生物，次氮基三乙酸(NTA)、二亚乙基三胺五乙酸(DTPA)、β-丙氨酸二乙酸(B-ADA)以及这些物质的盐)；含量在0.5重量％至50重量％范围内的、聚羧酸的均聚物和共聚物及其部分或完全中和盐，单体聚羧酸和羟基羧酸及其盐；含量在0.1重量％至50重量％范围内的磺化/羧化聚合物(为产物提供尺寸稳定性)；含量在约0.1重量％至约10重量％范围内的干燥助剂(选自聚酯，尤其是阴离子聚酯，任选连同另外的具有有利于缩聚的3至6个官能团(通常是酸、醇或酯官能团)的单体，聚碳酸酯-，聚氨酯-和/或聚脲-聚有机硅氧烷化合物或其前体化合物(尤其是活性环状碳酸酯和尿素型))；含量在约1重量％至约20重量％范围内的硅酸盐(包括硅酸钠或硅酸钾，例如二硅酸钠、偏硅酸钠和结晶页硅酸盐)；无机漂白剂(例如过氧化氢合盐，如过硼酸盐、过碳酸盐、过磷酸盐、过硫酸盐和过硅酸盐)和有机漂白剂(例如有机过氧酸，包括二酰基和四酰基过氧化物，尤其是二过氧基十二烷二酸、二过氧基十四烷二酸、二过氧基十六烷二酸)；漂白活化剂(即，含量在约0.1重量％至约10重量％范围内的有机过酸前体)；漂白催化剂(例如，三氮杂环壬烷合锰及相关络合物，二吡啶基胺合Co、Cu、Mn、Fe及相关络合物，五胺乙酸合钴(III)及相关络合物)；含量在约0.1重量％至5重量％范围内的金属护理剂(例如，苯并三唑、金属盐及络合物，和/或硅酸盐)；酶，其含量为每克自动盘碟洗涤剂组合物含约0.01至5.0mg活性酶(例如，蛋白酶、淀粉酶、脂肪酶、纤维素酶、胆碱氧化酶、过氧化物酶/氧化酶、果胶酸裂解酶、甘露聚糖酶、角质酶、漆酶、磷脂酶、溶血磷脂酶、酰基转移酶、过水解酶、芳基酯酶，以及它们的任何混合物)；以及酶稳定剂组分(选自寡糖、多糖和无机二价金属盐)。
另外的洗涤剂组合物
在下面编号的段落中描述了可加入本发明脂解酶的另外的示例性洗涤剂制剂。
1)配制为颗粒料的洗涤剂组合物，其具有至少600g/L的堆密度，含约7％至约12％的直链烷基苯磺酸盐(按酸计)；约1％至约4％的醇乙氧基硫酸盐(例如，C12-18醇，1至2个环氧乙烷(EO)单元)或烷基硫酸盐(例如，C16-18烷基硫酸盐)；约5％至约9％的醇乙氧基化物(例如，C14-15醇，7个EO单元)；约14％至约20％的碳酸钠(例如，Na2CO3)；约2％至约6％的可溶性硅酸盐(例如，Na2O·2SiO2)；约15％至约22％的沸石(例如，NaAlSiO4)；0％至约6％的硫酸钠(例如，Na2SO4)；约0％至约15％的柠檬酸钠/柠檬酸(例如，C6H5Na3O7/C6H8O7)；约11％至约18％的过硼酸钠(例如，NaBO3·H2O)；约2％至约6％的TAED；0％至约2％的羧甲基纤维素(CMC)；0％至3％的聚合物(例如，马来酸/丙烯酸共聚物、PVP、PEG)；0.0001％至0.1％的酶(按纯酶计)；和0％至5％的微量成分(例如，抑泡剂、香料、荧光增白剂、光致漂白剂)。
2)配制为颗粒料的洗涤剂组合物，其具有至少600g/L的堆密度，含约6％至约11％的直链烷基苯磺酸盐(按酸计)；约1％至约3％的醇乙氧基硫酸盐(例如，C12-18醇，1至2个EO单元)或烷基硫酸盐(例如，C16-18烷基硫酸盐)；约5％至约9％的醇乙氧基化物(例如，C14-15醇，7个EO单元)；约15％至约21％的碳酸钠(例如，Na2CO3)；约1％至约4％的可溶性硅酸盐(例如，Na2O·2SiO2)；约24％至约34％的沸石(例如，NaAlSiO4)；约4％至约10％的硫酸钠(例如，Na2SO4)；0％至约15％的柠檬酸钠/柠檬酸(如C6H5Na3O7/C6H8O7)；0％至约2％的羧甲基纤维素(CMC)；1％至6％的聚合物(如马来酸/丙烯酸共聚物、PVP、PEG)；0.0001％至0.1％的酶(按纯酶蛋白计)；和0％至5％的微量成分(如抑泡剂、香料)。
3)配制为颗粒料的洗涤剂组合物，其具有至少600g/L的堆密度，含约5％至约9％的直链烷基苯磺酸盐(按酸计)；约7％至约14％的醇乙氧基化物(例如，C12-15醇，7个EO单元)；约1至约3％的脂肪酸皂(例如，C16-22脂肪酸)；约10％至约17％的碳酸钠(Na2CO3)；约3％至约9％的可溶性硅酸盐(例如，Na2O·2SiO2)；约23％至约33％的沸石(NaAlSiO4)；0％至约4％的硫酸钠(例如，Na2SO4)；约8％至约16％的过硼酸钠(例如，NaBO3·H2O)；约2％至约8％的TAED；0％至约1％的膦酸盐(例如，EDTMPA)；0％至约2％的羧甲基纤维素(CMC)；0％至3％的聚合物(例如，马来酸/丙烯酸共聚物、PVP、PEG)；0.0001％至0.1％的酶(按纯酶蛋白计)；和0％至5％的微量成分(例如，抑泡剂、香料、荧光增白剂)。
4)配制为颗粒料的洗涤剂组合物，其具有至少600g/L的堆密度，含约8％至约12％的直链烷基苯磺酸盐(按酸计)；约10％至约25％的醇乙氧基化物(例如，C12-15醇，7个EO单元)；约14％至约22％的碳酸钠(Na2CO3)；约1％至约5％的可溶性硅酸盐(例如，Na2O·2SiO2)；约25％至约35％的沸石(例如，NaAlSiO4)；0％至约10％的硫酸钠(例如，Na2SO4)；0％至约2％的羧甲基纤维素(CMC)；1％至3％的聚合物(例如，马来酸/丙烯酸共聚物、PVP、PEG)；0.0001％至0.1％的酶(按纯酶蛋白计)；和0％至5％的微量成分(例如，抑泡剂、香料)。
5)含水液体洗涤剂组合物，含约15％至约21％的直链烷基苯磺酸盐(按酸计)；约12％至约18％的醇乙氧基化物(例如，C12-15醇，7个EO单元；或C12-15醇，5个EO单元)；约3％至约13％的脂肪酸皂(例如，油酸)；0％至约13％的烯基琥珀酸(C12-14烯基琥珀酸)；约8％至约18％的氨基乙醇；约2％至约8％的柠檬酸；0％至约3％的膦酸盐；0％至约3％的聚合物(例如，PVP、PEG)；0％至约2％的硼酸盐(例如，B4O7)；0％至约3％的乙醇；约8％至约14％的丙二醇；0.0001％至0.1％的酶(按纯酶蛋白计)；和0％至5％的微量成分(例如，分散剂、抑泡剂、香料、荧光增白剂)。
6)结构化含水液体洗涤剂组合物，含约15％至约21％的直链烷基苯磺酸盐(按酸计)；3％至9％的醇乙氧基化物(例如，C12-15醇，7个EO单元；或C12-15醇，5个EO单元)；约3％至约10％的脂肪酸皂(例如，油酸)；约14％至约22％的沸石(NaAlSiO4)；约9％至约18％的柠檬酸钾；0％至约2％的硼酸盐(例如，B4O7)；0％至约2％的羧甲基纤维素(CMC)；0％至约3％的聚合物(例如，PEG、PVP)；0％至约3％的锚定聚合物(例如，甲基丙烯酸月桂酯/丙烯酸共聚物，摩尔比25∶1，MW3800)；0％至约5％的甘油；0.0001％至0.1％的酶(按纯酶蛋白计)；和0％至5％的微量成分(例如，分散剂、抑泡剂、香料、荧光增白剂)。
7)配制为颗粒料的洗涤剂组合物，其具有至少600g/L的堆密度，含约5％至约10％的脂肪醇硫酸盐；约3％至约9％的乙氧基化脂肪酸单乙醇酰胺；0％至3％的脂肪酸皂；约5％至约10％的碳酸钠(例如，Na2CO3)；约1％至约4％的可溶性硅酸盐(例如，Na2O·2SiO2)；约20％至约40％的沸石(例如，NaAlSiO4)；约2％至约8％的硫酸钠(例如，Na2SO4)；约12％至约18％的过硼酸钠(例如，NaBO3·H2O)；约2％至约7％的TAED；约1％至约5％的聚合物(例如，马来酸/丙烯酸共聚物、PEG)；0.0001至0.1％的酶(按纯酶蛋白计)；和0％至5％的微量成分(例如，荧光增白剂、抑泡剂、香料)。
8)配制为颗粒料的洗涤剂组合物，含约8％至约14％的直链烷基苯磺酸盐(按酸计)；约5％至约11％的乙氧基化脂肪酸单乙醇酰胺；0％至约3％的脂肪酸皂；约4％至约10％的碳酸钠(例如，Na2CO3)；约1％至约4％的可溶性硅酸盐(Na2O·2SiO2)；约30％至约50％的沸石(例如，NaAlSiO4)；约3％至约11％的硫酸钠(例如，Na2SO4)；约5％至约12％的柠檬酸钠(例如，C6H5Na3O7)；约1％至约5％的聚合物(例如，PVP、马来酸/丙烯酸共聚物、PEG)；0.0001％至0.1％的酶(按纯酶蛋白计)；和0％至5％的微量成分(例如，抑泡剂、香料)。
9)配制为颗粒料的洗涤剂组合物，含约6％至约12％的直链烷基苯磺酸盐(按酸计)；约1％至约4％的非离子表面活性剂；约2％至约6％的脂肪酸皂；约14％至约22％的碳酸钠(例如，Na2CO3)；约18％至约32％的沸石(例如，NaAlSiO4)；约5％至约20％的硫酸钠(例如，Na2SO4)；约3％至约8％的柠檬酸钠(例如，C6H5Na3O7)；约4％至约9％的过硼酸钠(例如，NaBO3·H2O)；约1％至约5％的漂白活化剂(例如，NOBS或TAED)；0％至约2％的羧甲基纤维素(CMC)；约1％至约5％的聚合物(例如，聚羧酸酯或PEG)；0.0001％至0.1％的酶(按纯酶蛋白计)；和0％至5％的微量成分(例如，荧光增白剂、香料)。
10)含水液体洗涤剂组合物，含约15％至约23％的直链烷基苯磺酸盐(按酸计)；约8％至约15％的醇乙氧基硫酸盐(例如，C12-15醇，2至3个EO单元)；约3％至约9％的醇乙氧基化物(例如，C12-15醇，7个EO单元；或C12-15醇，5个EO单元)；0％至约3％的脂肪酸皂(例如，月桂酸)；约1％至约5％的氨基乙醇；约5％至约10％的柠檬酸钠；约2％至约6％的助水溶物(例如，甲苯磺酸钠)；0％至约2％的硼酸盐(例如，B4O7)；0％至约1％的羧甲基纤维素；约1％至约3％的乙醇；约2％至约5％的丙二醇；0.0001％至0.1％的酶(按纯酶蛋白计)；和0％至5％的微量成分(例如，聚合物、分散剂、香料、荧光增白剂)。
11)含水液体洗涤剂组合物，含约20％至约32％的直链烷基苯磺酸盐(按酸计)；6％至12％的醇乙氧基化物(例如，C12-15醇，7个EO单元；或C12-15醇，5个EO单元)；约2％至约6％的氨基乙醇；约8％至约14％的柠檬酸；约1％至约3％的硼酸盐(例如，B4O7)；0％至约3％的聚合物(例如，马来酸/丙烯酸共聚物，锚定聚合物例如，甲基丙烯酸月桂酯/丙烯酸共聚物)；约3％至约8％的甘油；0.0001％至0.1％的酶(按纯酶蛋白计)；和0％至5％的微量成分(例如，助水溶物、分散剂、香料、荧光增白剂)。
12)配制为颗粒料的洗涤剂组合物，其具有至少600g/L的堆密度，含约25％至约40％的阴离子表面活性剂(直链烷基苯磺酸盐、烷基硫酸盐、α-烯烃磺酸盐、α-磺基脂肪酸甲酯、链烷磺酸盐、皂)；约1％至约10％的非离子表面活性剂(例如，醇乙氧基化物)；约8％至约25％的碳酸钠(例如，Na2CO3)；约5％至约15％的可溶性硅酸盐(例如，Na2O·2SiO2)；0％至约5％的硫酸钠(例如，Na2SO4)；约15％至约28％的沸石(NaAlSiO4)；0％至约20％的过硼酸钠(例如，NaBO3·4H2O)；约0％至约5％的漂白活化剂(TAED或NOBS)；0.0001％至0.1％的酶(按纯酶蛋白计)；和0％至3％的微量成分(例如，香料、荧光增白剂)。
13)如上文的第1)至第12)所描述的洗涤剂组合物，其中所有或部分的直链烷基苯磺酸盐被(C12-C18)烷基硫酸盐替代。
14)配制为颗粒料的洗涤剂组合物，其具有至少600g/L的堆密度，含约9％至约15％的(C12-C18)烷基硫酸盐；约3％至约6％的醇乙氧基化物；约1％至约5％的多羟基烷基脂肪酸酰胺；约10％至约20％的沸石(例如，NaAlSiO4)；约10％至约20％的层状二硅酸盐(例如，赫希司特公司(Hoechst)出品的SK56)；约3％至约12％的碳酸钠(例如，Na2CO3)；0％至约6％的可溶性硅酸盐(例如，Na2O·2SiO2)；约4％至约8％的柠檬酸钠；约13％至约22％的过碳酸钠；约3％至约8％的TAED；0％至约5％的聚合物(例如，聚羧酸酯和PVP)；0.0001％至0.1％的酶(按纯酶蛋白计)；和0％至5％的微量成分(例如，荧光增白剂、光致漂白剂、香料、抑泡剂)。
15)配制为颗粒料的洗涤剂组合物，其具有至少600g/L的堆密度，含约4％至约8％的(C12-C18)烷基硫酸盐；约11％至约15％的醇乙氧基化物；约1％至约4％的皂；约35％至约45％的沸石MAP或沸石A；约2％至约8％的碳酸钠(Na2CO3)；0％至约4％的可溶性硅酸盐(例如，Na2O·2SiO2)；约13％至约22％的过碳酸钠；1％至8％的TAED；0％至约3％的羧甲基纤维素(CMC)；0％至约3％的聚合物(例如，聚羧酸酯和PVP)；0.0001％至0.1％的酶(按纯酶蛋白计)；和0％至3％的微量成分(例如，荧光增白剂、膦酸盐、香料)。
16)如上文的第1)至第15)项中所描述的洗涤剂制剂，其含稳定化的或包封的过酸，作为额外的组分，或作为之前规定的漂白体系的替代物。
17)如上文的第1)、3)、7)、9)和第12)项中所描述的洗涤剂组合物，其中过硼酸盐被过碳酸盐替代。
18)如上文的第1)、3)、7)、9)、12)、14)和第15)项中所描述的洗涤剂组合物，其额外含有锰催化剂。锰催化剂例如为“Efficientmanganesecatalystsforlow-temperaturebleaching”Nature369：637-639(1994)中描述的一种化合物。
19)配制为非水性洗涤剂液体的洗涤剂组合物，含液体非离子表面活性剂(例如，直链烷氧基化伯醇)、助洗剂体系(例如，磷酸盐)、一种或多种酶，和碱。该洗涤剂还可包含阴离子型表面活性剂和/或漂白体系。
如上，可以以洗涤剂中常规采用的浓度掺入本发明的脂解酶多肽。目前设想的是，洗涤剂组合物中添加的酶量可对应每升洗涤液添加0.00001至1.0mg脂解酶多肽(按纯酶蛋白计)。
洗涤剂组合物还可包含其他常规的洗涤剂成分，如抗絮凝剂、填充材料、泡沫抑制剂、耐腐蚀剂、污垢悬浮剂、螯合剂、抗污垢再沉积剂、脱水剂、染料、杀菌剂、荧光剂、增稠剂和香料。
洗涤剂组合物可被配制为手洗(手动)或机洗(自动)衣物洗涤剂组合物，包括适用于预处理脏污织物的衣物洗涤添加剂组合物和漂洗附加的织物软化剂组合物；或被配制为用于普通家庭硬质表面清洁操作的洗涤剂组合物；或被配制用于手动或自动盘碟操作。
本文描述的任一种清洁组合物都可包含任意多种附加的酶。一般来讲，所述酶应当与选择的洗涤剂相容(如就最佳pH，与其他酶成分和非酶成分等的相容性而言)，而且所述酶应当以有效量添加。提供下列酶作为示例。
蛋白酶：合适的蛋白酶包括源于动物、植物或微生物的蛋白酶。化学修饰的突变体或蛋白质工程突变体以及天然加工的蛋白质也包括在内。蛋白酶可为丝氨酸蛋白酶或金属蛋白酶、碱性微生物蛋白酶、胰蛋白酶样蛋白酶或糜蛋白酶样蛋白酶。碱性蛋白酶的示例是枯草杆菌蛋白酶，尤其是来源于芽胞杆菌属(Bacillus)的那些，例如，枯草杆菌蛋白酶Novo、枯草杆菌蛋白酶Carlsberg、枯草杆菌蛋白酶309、枯草杆菌蛋白酶147和枯草杆菌蛋白酶168(参见例如，WO89/06279)。胰蛋白酶样蛋白酶的示例是胰蛋白酶(例如源于猪或牛的胰蛋白酶)和镰孢属蛋白酶(参见例如，WO89/06270和WO94/25583)。可用的蛋白酶的示例还包括但不限于WO92/19729、WO98/20115、WO98/20116和WO98/34946中所述的变体。可商购获得的蛋白酶包括但不限于： PRIMASETM、DURALASETM、KANNASETM和BLAZETM(诺和诺德公司(NovoNordiskA/S)和诺维信公司)；MAXACALTM、MAXAPEMTM、PURAFECTOXPTM、FN2TM和FN3TM(丹尼斯克(美国)公司(DaniscoUSInc.))。其他示例性蛋白酶包括源自解淀粉芽孢杆菌的NprE和源自纤维单胞菌属物种菌株69B4的ASP。
脂肪酶：合适的脂肪酶包括源于细菌或真菌的脂肪酶。包括化学修饰的突变体、蛋白水解改性的突变体或蛋白质工程突变体。可用的脂肪酶的示例包括但不限于来源于腐质霉属(同义词嗜热丝孢菌属)的脂肪酶，例如来源于疏棉状腐质霉(H.lanuginosa)(疏棉状嗜热丝孢菌(T.lanuginosus))(参见例如，EP258068和EP305216)、来源于特异腐质霉(H.insolens)(参见例如，WO96/13580)的脂肪酶；假单胞菌属脂肪酶，例如，来源于产碱假单胞菌(P.alcaligenes)或类产碱假单胞菌(P.pseudoalcaligenes)(参见例如，EP218272)、洋葱假单胞菌(P.cepacia)(参见例如，EP331376)、施氏假单孢菌(P.stutzeri)(参见例如，GB1,372,034)、荧光假单胞菌(P.fluorescens)、假单胞菌属菌株SD705(参见例如，WO95/06720和WO96/27002)、威斯康星假单胞菌(P.wisconsinensis)(参见例如，WO96/12012)的脂肪酶；芽孢杆菌属脂肪酶，例如来源于枯草芽孢杆菌(参见例如，Dartois等人，BiochemicaetBiophysicaActa，1131：253-360(1993))、嗜热脂肪芽孢杆菌(参见例如，JP64/744992)或短小芽孢杆菌(参见例如，WO91/16422)的脂肪酶。设想用于制剂中的额外的脂肪酶变体包括例如WO92/05249、WO94/01541、WO95/35381、WO96/00292、WO95/30744、WO94/25578、WO95/14783、WO95/22615、WO97/04079、WO97/07202、EP407225和EP260105中所述的那些。一些可商购获得的脂肪酶包括和LIPOLASEULTRATM(诺和诺德公司和诺维信公司)。
聚酯酶：组合物中可包含合适的聚酯酶，例如WO01/34899、WO01/14629和US6933140中所述的那些。
淀粉酶：组合物可与其他淀粉酶混合，例如非生产增强性淀粉酶。所述淀粉酶可包括可商购获得的淀粉酶，例如但不限于 和BANTM(诺和诺德公司和诺维信公司)；和(得自丹尼斯克(美国)公司)。
纤维素酶：可将纤维素酶添加到组合物中。合适的纤维素酶包括源于细菌或真菌的纤维素酶。包括化学修饰的突变体或蛋白质工程突变体。合适的纤维素酶包括来源于芽孢杆菌属、假单胞菌属、腐质霉属、镰孢属、梭孢壳属(Thielavia)、支顶孢属(Acremonium)的纤维素酶，例如，由例如美国专利4,435,307、5,648,263、5,691,178、5,776,757和WO89/09259中所公开的特异腐质霉、嗜热毁丝霉(Myceliophthorathermophila)和尖孢镰孢菌(Fusariumoxysporum)产生的真菌纤维素酶。设想使用的示例性纤维素酶是对纺织物具有颜色护理有益效果的纤维素酶。此类纤维素酶的示例为例如EP0495257、EP0531372、WO96/11262、WO96/29397和WO98/08940中所述的纤维素酶。其他例子为纤维素酶变体，例如WO94/07998、WO98/12307、WO95/24471、PCT/DK98/00299、EP531315、美国专利5,457,046、5,686,593和5,763,254中所述的那些。市售纤维素酶包括和(诺和诺德公司和诺维信公司)；和(丹尼斯克(美国)公司)；以及KAC-500(B)TM(花王公司)。
过氧化物酶/氧化酶：设想用于组合物中的合适的过氧化物酶/氧化酶包括源于植物、细菌或真菌的那些。包括化学修饰的突变体或蛋白质工程突变体。可用的过氧化物酶的示例包括WO93/24618、WO95/10602和WO98/15257中所述的来源于鬼伞属(Coprinus)，例如来源于灰盖鬼伞(C.cinereus)的过氧化物酶，及其变体。可商购获得的过氧化物酶包括例如GUARDZYMETM(诺和诺德公司和诺维信公司)。
洗涤剂组合物还可包含2，6-β-D-果聚糖水解酶，该酶可有效去除/清洁家居和/或工业纺织物/衣物上存在的生物膜。
可通过添加分开的含有一种或多种酶的添加剂，或通过添加含有所有这些酶的混合添加剂，将洗涤剂酶包含于洗涤剂组合物中。洗涤剂添加剂，即分开的添加剂或组合添加剂，可被配制为例如颗粒料、液体、浆料等。示例性洗涤剂添加剂制剂包括但不限于颗粒料(尤其是非粉尘颗粒)、液体(尤其是稳定化的液体)或浆料。
例如，可如美国专利4,106,991和4,661,452中所公开的，制备无尘颗粒，并且可任选以本领域已知的方法进行涂覆。蜡质包衣材料的示例是聚环氧乙烷产品(例如，聚乙二醇，PEG)，平均摩尔重量为1,000至20,000；具有16至50个环氧乙烷单元的乙氧基化壬基酚；乙氧基化脂肪醇，其中醇含有12至20个碳原子，并且其中有15至80个环氧乙烷单元；脂肪醇；脂肪酸；以及脂肪酸的甘油单酯、甘油二酯和甘油三酯。适合于应用流化床技术的成膜包衣材料的示例在例如GB1483591中给出。例如，可根据已建立的方法，通过添加多元醇例如丙二醇、糖或糖醇、乳酸或硼酸使液体酶制品稳定化。可按照EP238,216中所公开的方法制备受保护的酶。
洗涤剂组合物可为任意便利的形式，例如，条棒状、片状、粉末状、颗粒状、糊状或液态。液体洗涤剂可为水性的，通常含有最多约70％的水和0％至约30％的有机溶剂。也可以设想含有约30％或更少的水的致密洗涤剂凝胶。洗涤剂组合物可以任选包含一种或多种表面活性剂，该表面活性剂可为非离子型，包括半极性和/或阴离子型和/或阳离子型和/或两性离子型。表面活性剂可以约0.1重量％至约60重量％的宽范围存在。
当包含在其中时，洗涤剂将一般含约1％至约40％的阴离子型表面活性剂，例如直链烷基苯磺酸盐、α-烯烃磺酸盐、烷基硫酸盐(脂肪醇硫酸盐)、醇乙氧基硫酸盐、仲链烷磺酸盐、α-磺基脂肪酸甲酯、烷基或烯基琥珀酸，或者皂。
当包含在其中时，洗涤剂将一般含约0.2％至约40％的非离子型表面活性剂，例如醇乙氧基化物、壬基酚乙氧基化物、烷基多苷、烷基二甲基胺氧化物、乙氧基化脂肪酸单乙醇酰胺、脂肪酸单乙醇酰胺、多羟基烷基脂肪酸酰胺或葡糖胺的N-酰基N-烷基衍生物(“葡糖酰胺”)。
洗涤剂组合物可含0％至约65％的洗涤剂助洗剂或络合剂，例如沸石、二磷酸盐、三磷酸盐、膦酸盐、碳酸盐、柠檬酸盐、次氮基三乙酸、乙二胺四乙酸(EDTA)、二亚乙基三胺五乙酸、烷基或烯基琥珀酸、可溶性硅酸盐或层状硅酸盐(如，赫希司特公司出品的SKS-6)。
洗涤剂可包含一种或多种聚合物。示例性聚合物包括羧甲基纤维素(CMC)、聚乙烯吡咯烷酮(PVP)、聚乙二醇(PEG)、聚乙烯醇(PVA)、聚(乙烯基吡啶-N-氧化物)、聚(乙烯基咪唑)、聚羧酸脂如聚丙烯酸脂、马来酸/丙烯酸共聚物，以及甲基丙烯酸月桂酯/丙烯酸共聚物。
可用常规的稳定剂稳定洗涤剂组合物中的酶，这些稳定剂为例如多元醇(如丙二醇或甘油)；糖或糖醇；乳酸；硼酸或硼酸衍生物(如芳族硼酸酯)；或苯基硼酸衍生物(如4-甲酰基苯基硼酸)。该组合物可根据WO92/19709和WO92/19708所述方法进行配制。
预期在所述洗涤剂组合物中，尤其是酶变体，可按照每升洗涤液体约0.01至约100mg的酶蛋白的量加入(例如，每升洗涤液体约0.05至约5.0mg的酶蛋白或每升洗涤液体0.1至约1.0mg的酶蛋白)。
有利地包括本发明的脂解酶多肽的代表性洗涤剂制剂包括，如见于WO2013063460，第78-152页中的洗涤剂制剂，尤其是第94至152页的表格，据此以引用的方式并入。脂解酶通常以0.00001％至10％酶蛋白的水平(以组合物的重量计)掺入到洗涤剂组合物中。在一些实施方案中，洗涤剂组合物包含多于0.0001％、0.001％、0.01％或0.1％的脂解酶(以组合物的重量计)。在一些实施方案中，洗涤剂组合物包含少于1％、0.1％、0.01％或0.001％的脂解酶(以组合物的重量计)。
评估洗涤剂组合物中脂解酶活性的方法
许多脂解清洁测定法是本领域已知的，包括布样和微布样测定法。所附的实施例仅描述了几个此类测定。
为了进一步说明所述组合物和方法及它们的优点，下面给出的具体实施例应理解为举例说明性的，而不是限制性的。
制备和使用清洁组合物的方法
本发明的清洁组合物被配制成任何合适的形式并通过配制者所选择的任何合适方法制备(参见例如，美国专利5,879,584、5,691,297、5,574,005、5,569,645、5,565,422、5,516,448、5,489,392、5,486,303、4,515,705、4,537,706、4,515,707、4,550,862、4,561,998、4,597,898、4,968,451、5,565,145、5,929,022、6,294,514和6,376,445)。
在一些实施方案中，本发明的清洁组合物以单位剂型提供，包括小片、胶囊、小囊、小袋和多隔室小袋。在一些实施方案中，对单位剂量形式进行设计以提供多隔室小袋(或其他单位剂量形式)内的成分的控制释放。合适的单位剂量和控制释放形式是本领域已知的(参见例如，EP2100949、WO02/102955、美国专利4,765,916、4,972,017和WO04/111178关于适用于单位剂量和控制释放形式的材料)。在一些实施方案中，单位剂型以使用水溶性薄膜或水溶性小袋包裹的片剂提供。EP2100947中提供了各种单位剂量形式，并且这些形式是本领域已知的。
使用方法
在一些实施方案中，本发明的清洁组合物用于清洁表面(例如，盘碟)、衣物、硬质表面、隐形眼镜等。在一些实施方案中，表面的至少一部分与本发明的清洁组合物的至少一个实施方案接触，所述组合物为纯形式或稀释于洗涤液中，随后任选洗涤和/或冲洗所述表面。出于本发明的目的，“洗涤”包括但不限于擦洗和机械洗涤。在一些实施方案中，本发明的清洁组合物在溶液中以约500ppm至约15,000ppm的浓度使用。在一些实施方案中，其中洗涤溶剂是水时，水温通常在约5℃至约90℃的范围内。
本发明提供了用于清洁或洗涤需要清洁的物品或表面(例如，硬质表面)的方法，包括但不限于，用于清洁或洗涤盘碟物品、餐具物品、织物物品、衣物物品、个人护理物品等的方法，以及用于清洁或洗涤硬质或柔软表面(例如，物品的硬质表面)的方法。
在一些实施方案中，本发明提供了一种用于清洁需要清洁的物品、物体或表面的方法，该方法包括将物品或表面(或期望清洁的物品或表面的部分)与至少一种本发明变体脂肪酶脂解酶或本发明的组合物接触足以清洁所述物品、物体或表面至所需程度的时间，和/或在适于和/或有效用于清洁所述物品、物体或表面至所需程度的条件下进行接触。一些此类方法还包括用水冲洗物品、物体或表面。在一些此类方法中，清洁组合物为盘碟洗涤剂组合物，待清洁的物品或物体为盘碟物品或餐具物品。如本文所用，“盘碟物品”为通常用于供给食物或食用食物的物品。盘碟物品可以是但不限于(例如)盘子、碟子、杯子、碗等。如本文所用，“餐具”是一更广义的术语，其包括但不限于(例如)盘碟、刀具、餐刀、餐叉、匙、筷子、玻璃器具、罐壶、调味汁碟、饮用容器、盛菜物品等。预期“餐具物品”包括这些或相似的用于供应食物或食用食物的物品中的任何一种。在一些此类方法中，清洁组合物为自动盘碟洗涤剂组合物或手洗盘碟洗涤剂组合物，待清洁的物品或物体为盘碟或餐具物品。在一些此类方法中，清洁组合物为衣物洗涤剂组合物(例如，粉末衣物洗涤剂组合物或液体衣物洗涤剂组合物)，并且待清洁的物品为织物物品。在一些其他实施方案中，清洁组合物为衣物预处理组合物。
在一些实施方案中，本发明提供用于清洁或洗涤任选分别需要清洁或洗涤的织物物品的方法。在一些实施方案中，该方法包括：提供包含变体脂解酶的组合物(包括但不限于织物或衣物清洁组合物)，以及需要清洁的织物物品或衣物物品，将该织物物品或衣物物品(或期望清洁的物品的部分)与组合物在足以或有效用于清洁或洗涤该织物或衣物物品至所需程度的条件下接触。
在一些实施方案中，本发明提供了一种用于清洁或洗涤任选需要清洁的物品或表面(例如，硬质表面)的方法，该方法包括提供待清洁或洗涤的物品或表面，并且将该物品或表面(或期望清洁或洗涤的物品或表面的部分)与至少一种本发明脂肪酶变体或本发明的组合物(包含至少一种此类脂肪酶变体)接触足以清洁或洗涤该物品或表面至所需程度的时间和/或在足以或有效用于清洁或洗涤该物品或表面至所需程度的条件下接触。此类组合物包括但不限于，例如，本发明的清洁组合物或洗涤剂组合物(例如，手洗盘碟洗涤剂组合物、手洗盘碟清洁组合物、衣物洗涤剂或织物洗涤剂或衣物或织物清洁组合物、液体衣物洗涤剂、液体衣物清洁组合物、粉末衣物洗涤剂组合物、粉末衣物清洁组合物、自动盘碟洗涤剂组合物、衣物助洗剂清洁组合物或洗涤剂组合物、衣物清洁添加剂和衣物预洗剂组合物等)。在一些实施方案中，将该方法重复一次或多次，尤其是如果需要额外的清洁或洗涤的话。例如，在一些情况下，该方法任选还包括使物品或表面与至少一种变体脂解酶或组合物保持接触足以或有效用于清洁或洗涤物品或表面至所需程度的一段时间。在一些实施方案中，该方法还包括用水和/或其他液体冲洗物品或表面。在一些实施方案中，这些方法还包括再次用至少一种本发明的变体脂解酶或本发明的组合物接触物品或表面，并且使该物品或表面与至少一种变体脂解酶或组合物保持接触足以清洁或洗涤物品或表面至所需程度的一段时间。在一些实施方案中，清洁组合物为盘碟洗涤剂组合物，待清洁的物品为盘碟物品或餐具物品。在本发明方法的一些实施方案中，清洁组合物为自动盘碟洗涤剂组合物或手洗盘碟洗涤剂组合物，待清洁的物品为盘碟或餐具物品。在该方法的一些实施方案中，清洁组合物为衣物洗涤剂组合物，待清洁的物品为织物物品。
本发明还提供了在自动盘碟洗涤机中清洁餐具或盘碟物品的方法，该方法包括提供自动盘碟洗涤机，在该机器中放入足以清洁餐具或盘碟物品的一定量的包含至少一种本发明脂肪酶变体或本发明组合物的自动盘碟洗涤组合物(例如，通过将该组合物置于机器中的合适的或提供的洗涤剂隔室或分配器中)，将盘碟或餐具物品放入该机器中，运行该机器以清洁该餐具或盘碟物品(例如按照制造商的说明书)。在一些实施方案中，该方法包括本文所述的任何自动盘碟洗涤组合物，该组合物包括但不限于本文提供的至少一种脂肪酶变体。待使用的自动盘碟洗涤组合物的量可根据制造商的说明书或建议容易地确定，可采用任何形式的包含至少一种本发明变体脂解酶的自动盘碟洗涤组合物(例如，液态、粉末状、固态、凝胶状、片状等形式)，包括本文所述的任何组合物在内。
本发明还提供了用于清洁任选地需要清洁的表面、物品或物体的方法，该方法包括将物品或表面(或期望清洁的物品或表面的部分)与至少一种本发明脂肪酶变体或本发明清洁组合物(所述组合物为纯形式或稀释于洗涤液中)接触足够的时间，和/或在足以或有效清洁或洗涤物品或表面至所需程度的条件下接触。然后如果需要，可以(任选地)洗涤和/或冲洗表面、物品或物体。出于本发明的目的，“洗涤”包括但不限于(例如)擦洗和机械搅动。在一些实施方案中，清洁组合物在溶液(例如，水溶液)中以约500ppm至约15,000ppm的浓度使用。当洗涤溶剂是水时，水温通常在约5℃至约90℃的范围内，并且当所述表面、物品或物体包括织物时，水与织物的质量比通常为约1∶1至约30∶1。
本发明还提供了在洗衣机中清洁衣物或织物物品的方法，该方法包括提供洗衣机，将足以清洁衣物或织物物品的一定量的包含至少一种本发明脂肪酶变体的衣物洗涤剂组合物置于洗衣机中(例如，通过将该组合物置于洗衣机中的合适的或提供的洗涤剂隔室或分配器中)，将衣物或织物物品放入洗衣机中，运转洗衣机以清洁衣物或织物物品(例如，按照制造商的说明书)。本发明的方法包括本文所述的任何衣物洗涤剂组合物，其包含但不限于本文所提供的任何脂肪酶变体中的至少一种。待使用的衣物洗涤剂组合物的量可根据制造商的说明书或建议容易地确定，可采用任何形式的包含至少一种本发明变体脂解酶的衣物洗涤剂组合物(例如，固态、粉末状、液态、片状、凝胶状等形式)，包括本文所述的任何组合物在内。
本发明还提供了改善羧酸酯键水解的方法，例如长链甘油三酯中的羧酸酯键水解。水解反应可应用于多种行业，包括洗涤剂清洁、食品、香料行业、药物的生物催化拆分、化学品生产、生物传感器开发、生物修复和化妆品及香水行业。
在一些实施方案中，可将本发明的变体用来在有机溶剂中进行酯交换，例如用于生产类可可脂(例如，通过棕榈油的酯交换)和母乳脂肪替代品，以及用于制备甘油酯产品(例如，生产黄油或人造黄油)。在一些实施方案中，本发明的变体可用于水解牛油。在一些实施方案中，本发明的变体可用于改良食品风味，例如通过短链脂肪酸和醇合成酯。在一些实施方案中，本发明的变体可用来帮助去除脂肪从而生产瘦肉、或帮助香肠发酵、或改良其他食品的风味(例如，奶酪、大米、豆浆或苹果酒)(参见US20110262591)。在一些实施方案中，本发明的变体可用于烘培(参见WO1994004035、EP585988、EP785994)。
在一些实施方案中，本发明的变体可通过水解脂质生产肥皂。在一些实施方案中，本发明的变体可用于个人护理用途(例如，化妆品)，其中该变体可以帮助生产润肤剂，例如肉豆蔻酸异丙酯、棕榈酸异丙酯或棕榈酸-2-乙基己基酯。在一些实施方案中，本发明的变体可用于帮助消化、治疗胃肠道紊乱、消化不良，以及消化系统变态反应的皮肤表现。
在一些实施方案中，本发明的变体可用于拆分药物化合物的外消旋混合物。在一些实施方案中，本发明的变体可用于生产生物柴油。
在一些实施方案中，本发明的变体可用于生产可生物降解的聚合物，例如油酸1-丁酯或三羟甲基丙烷酯。
在一些实施方案中，本发明的变体可用在纺织工业中去除上浆润滑剂，改善染色性能。在一些实施方案中，本发明的变体可用于脱油脂，例如用于动物兽皮、羊皮或羊毛的脱油脂。在一些实施方案中，本发明的变体可用于增加纸浆的制浆率。本发明的变体也可用来在纸浆和造纸工业中控制树脂(pitch)(参见EP374700)。
在一些实施方案中，本发明的变体可用于碳水化合物水溶液或浆料的脱胶，以改善其过滤性，特别是淀粉水解产物，尤其是小麦淀粉水解产物(因为其难以过滤并且会产生浑浊的滤液)。可使用本领域熟知的方法来进行所述处理。参见，例如EP219,269、EP808,903和美国专利6,103,505。
本发明还提供根据美国专利6,558,715的在烘培中的使用方法。
实验
本发明将在以下实施例中进一步详细描述，所述实施例并不旨在以任何方式限制权利要求中的本发明范围。
实施例1
方法
以下测定法为在后述实施例中使用的标准测定法。有时具体方案要求与这些标准测定法有一定偏差。在这些情况下，实施例中将明确与下面这些标准测定方案的偏差。
A.性能指数
酶的性能指数(PI)将相同蛋白质浓度下变体的性能(测量值)与标准酶的性能(理论值或测量值)相比较。此外，可使用标准酶的朗格缪尔方程的参数计算理论值。
性能指数(PI)大于1(PI＞1)表示变体性能相比亲本序列(如SEQIDNO：1)或参考序列(如SEQIDNO：2，，诺维信公司)提高；PI等于1(PI＝1)表示变体性能与标准酶表现相同；PI小于1(PI＜1)表示变体性能比标准酶差。
B.对硝基苯基酯水解测定法
测定TLL变体对于具有不同烷基链长的三种不同对硝基苯基(pNP)酯底物的脂肪酶活性，以确定TLL变体的链长偏好性。表1-1提供了pNP酯底物的详细信息。
使用预悬浮于乙醇(5％)中的0.8mMpNP酯，在0.05MHEPES(调节pH至8.2)中，或在0.05MMES(调节pH至6.0)中，制得含pNP酯底物的反应乳液。混合pNP酯/缓冲液悬浮液，转移到96孔微量滴定板(MTP)的各孔中，每孔盛有酶样品，总体积为200μL。调整酶样品的稀释度及其转移体积以保持反应在线性范围内。在3分钟内，在OD405nm处监测释放产生的pNP，并用空白值(不含酶)校正。使用pNP标准曲线计算每秒产生的pNP产物，然后相对于孔中添加的酶样品进行归一化(μmolpNP/s/mg添加的酶)。在pH6.0使用辛酸对硝基苯酯的情况下，混合pNP酯/缓冲液悬浮液，转移到96孔微量滴定板(MTP)的各孔中，每孔盛有酶样品，总体积为150μL。密封滴定板，置于iEMS振荡器(赛默飞世尔科技公司(Thermoscientific))中，在25℃温度下、以900rpm振荡10分钟。温育后，添加50μl含0.5％TritonX-100的0.2MHEPES(pH8.2)。在OD405nm处读取释放产生的pNP，并用空白值(不含酶)校正。通过比较变体酶与参考TLL酶(SEQIDNO：1或2)对特定pNP酯底物的水解，以确定水解性能指数。
通过计算在烷基链长不同的成对底物上的比活性的比值，来确定脂肪酶的优选链长。用脂肪酶水解棕榈酸对硝基苯酯的比活性除以该脂肪酶水解丁酸对硝基苯酯的比活性，若比值大于1，表明优选较长的链长；若比值小于1，表明优选较短的烷基链长。将变体酶水解特定pNP酯底物对的比活性比值与标准酶水解该特定pNP酯底物对的比活性比值进行比较，由此衡量TLL变体的优选烷基链长的变化水平。
C.洗涤剂稳定性测定法
将TLL变体在重垢型液体(HDL)洗涤剂(商品名为汰渍冷水洗衣精(Tidecoldwaterliquid)(美国宝洁公司(P&G)，经热处理))的10％(v/v)溶液中在升高的温度下进行胁迫处理，由此监测TLL变体的加速洗涤剂稳定性。
在96孔PCR板中，用汰渍冷水洗衣精的10％(v/v)溶液将脂肪酶的原酶(rawferment)稀释20倍。混合后，将2μL转移到盛有198μLpNP辛酸酯底物的96孔板板孔中，按B部分描述的方法测量活性，得到未经受胁迫处理的活性值。密封PCR板，置于PCR仪中42℃温育30分钟。结束温育后，将PCR板在4℃温度下冷却3分钟，然后再次测量活性。将4μL经温育的混合物转移到盛有196μlpNP辛酸酯/缓冲液悬浮液的96孔板上，按B部分描述的方法测量酶变体的活性，得到经胁迫处理后的活性值。
将变体酶经胁迫处理后的活性值与未经受胁迫处理的活性值的比值和参考TLL酶(SEQIDNO：1或2)的该比值进行比较，确定洗涤剂稳定性的性能指数。
D.热稳定性测定法
在50mMHEPES，pH8.2(具有1ppm枯草杆菌蛋白酶BPN’-Y217L蛋白酶)中，在升高的温度下胁迫处理TLL变体，由此监测TLL变体的加速热稳定性。
将95μL含蛋白酶的50mMHEPES(pH8.2)转移到盛有5μL酶样品的96孔PCR板板孔中。混合后，将2μL缓冲液/脂肪酶混合物转移到盛有198μlpNP辛酸酯/缓冲液悬浮液的96孔板上，按B部分描述的方法测量酶变体的活性。
密封PCR板，置于PCR仪中69℃温育30分钟。温育后，将PCR板在4℃温度下冷却3分钟，然后测量活性。将4μL经温育的混合物转移到盛有196μlpNP辛酸酯/缓冲液悬浮液的96孔板上，按B部分描述的方法测量酶变体的活性。
将加热后的酶活性除以加热前的酶活性，计算出热稳定性活性比值，以剩余活性百分比表示。将变体酶的这种活性比值与经受类似处理的参考TLL酶(SEQIDNO：1或2)的这种活性比值进行比较，确定与加速热稳定性相关的性能指数。
E.LAS稳定性测定法
将TLL变体放入用HEPES缓冲液(pH8)稀释的0.1％LAS中，使其经受胁迫处理，由此监测TLL变体的加速LAS稳定性；其中，LAS为直链烷基苯磺酸盐，具体地讲，为十二烷基苯磺酸钠，西格玛公司目录号为289957。
将80μL0.1％LAS(w/v)(pH8.2)转移到盛有20μL酶样品的96孔PCR板板孔中。混合后，将2μL缓冲液/脂肪酶混合物转移到盛有198μlpNP辛酸酯/缓冲液悬浮液的96孔板上，按B部分描述的方法测量酶变体的活性。
密封PCR板，置于PCR仪中25℃温育30分钟。温育后，将PCR板在4℃温度下冷却3分钟，然后测量活性。将4μL经温育的混合物转移到盛有196μlpNP辛酸酯/缓冲液悬浮液的96孔板上，按B部分描述的方法测量酶变体的活性。
将在LAS中温育后的酶活性除以没有LAS时的酶活性，计算出LAS稳定性活性比值，以剩余活性百分比表示。将酶变体的这种活性比值与经受类似处理的参考TLL酶(SEQIDNO：1或2)的这种活性比值进行比较，确定与LAS稳定性相关的性能指数。
F.CS-61微布样测定法
利用微布样测定法检测脂肪酶变体的清洁性能。CS-61布样是预先用牛油和红色染料(荷兰试验材料中心(CenterforTestmaterial，CFT))染污的棉布样，在96孔板的孔中使用。把布样切成多个5mm直径的小片，放入MTP的每个孔中。
在两种洗涤剂背景下检测脂肪酶变体的性能，分别为：全剂量汰渍冷水洗衣精(经95℃热处理3小时，最终剂量：0.92g/l)，半剂量汰渍冷水洗衣精(经95℃热处理3小时)加辅剂(正十二烷基-β-D-麦芽吡喃糖苷)(洗涤剂最终剂量：0.46g/L，辅剂：0.272μM)。
在MTP板上进行培养，然后用密理博公司(Millipore)的滤器过滤培养物的培养液，获得待检测的脂肪酶变体样品。所用的缓冲液为20mMHEPES(最终浓度)(pH8.2)，将水硬度调至6gpg(Ca∶Mg＝2∶1)。向96孔板上每个装有布样的孔中添加246μL体积的HDL洗涤剂溶液(如上所述)。然后向每个孔添加4μL体积的酶样品，引发反应。密封96孔板，置于iEMS振荡器(赛默飞世尔科技公司)中，在30℃温度下、以900rpm振荡30分钟。温育后，使用Hydrospeed洗板机(澳大利亚的帝肯公司(Tecan，Austria))，用去离子水冲洗织物三次，50℃干燥过夜。用扫描仪(MiCrotekScanMaker900)对冲洗并干燥过的织物进行RGB测量，量化去污效果。图像被导入PhotoshopCSII，在该软件中使用驯鹿图像公司(ReindeerGraphics)研发的IPTK5.0，从含布样的区域提取RGB值。使用以下公式，计算经洗涤织物相对于未经洗涤织物的污垢去除百分率(SRI)值：
污垢去除百分率(SRI)＝(ΔE/ΔE初始)*100
其中
将酶变体的SRI与参考TLL酶(SEQIDNO：1或2)在与变体相同酶剂量情况下的SRI进行比较，计算与清洁性能相关的性能指数。利用朗缪尔拟合，计算标准酶在酶剂量与变体相同情况下的SRI。
G.织物粘附性测定法
使用微布样测定法检测TLL变体在洗涤剂溶液中对棉织物的粘附性，洗涤剂溶液为0.46g/l热灭活的汰渍冷水洗衣精，以20mMHEPES(pH8.2)缓冲，水硬度为6gpg(Ca∶Mg＝2∶1)。把漂白过的棉布样EMPA221切成多个5mm直径的小片，放入96孔板(3641，康宁公司(Corning))的每个孔中。向板上每个装有布样的孔中添加95μL体积的前述洗涤剂溶液。为开始测定，将5μL原酶混入各孔，共用三块板进行平行测定。将板密封，置于iEMS振荡器(赛默飞世尔科技公司)中，在30℃温度下、以900rpm振荡10分钟。温育后，使用Hydrospeed洗板机(澳大利亚的帝肯公司)，用自来水将该微布样板冲洗三次。向冲洗过的板上的每个孔添加200μLpNP辛酸酯/缓冲液悬浮液，以测量织物结合酶的程度。然后将板置于iEMS振荡器(赛默飞世尔科技公司)中30℃温育5分钟；取150μL溶液转移到新96孔板中，测量405nm处的OD值并扣除空白。将每种TLL变体的该活性水平与经受类似处理的参考TLL酶(SEQIDNO：1或2)的该活性水平进行比较，确定与织物粘附性相关的性能指数。
H.洗涤剂
使用的市售洗涤剂为：
汰渍冷水洗衣精(宝洁公司)。该洗涤剂于2010年8月购买；经热处理(95℃处理3小时)，目的是使产品配方中的酶失活。
I.蛋白质测定法
将每孔盛有60μL过滤的培养液的96孔圆底MTP(5042-1385，安捷伦公司(Agilent)，0.5mL聚丙烯)用于高效液相色谱(HPLC)蛋白质测定法。取每种样品各10μL，注入配备AcquityUPLCBEH300C4色谱柱(1.7μm，2.1mm×50mm)(沃特世公司(Waters))的Agilent1290(HewlettPackard)(U)HPLC。使用含0.1％三氟乙酸的MiliQ水，蛋白质结合在色谱柱上，然后使用最高达70％的乙腈(含0.1％三氟乙酸)梯度，对蛋白质进行洗脱。测量220nm处的吸光度值，然后使用ChemStation软件(安捷伦科技公司(AgilentTechnologies))对洗脱峰进行积分。基于纯化的TLL-蛋白的标准曲线来确定TLL变体的蛋白浓度。
实施例2
产生TLL变体
A.生成TLL组合文库
为在芽孢杆菌细胞中生产，将pHYT-TLLwt质粒DNA(美国专利申请61/713,436描述了这种质粒DNA)用作模板，以在成熟区(美国专利申请61/713,436中的SEQIDNO：3)的多个预选位点产生组合文库。委托GeneArt股份公司(德国雷根斯堡市)按标准方案构建组合文库。组合文库由含有编码TLL变体序列的表达质粒的转化枯草芽孢杆菌细胞组成。每个受关注位点的对应密码子保持野生型，或被至少一种非野生型氨基酸的密码子替换。
SEQIDNO：3示出合成的TLL基因的核苷酸序列。
gctagcgcagctggcaaagaagttagccaagatctgttcaaccaattcaaccttttcgctcaatactctgcagctgcttactgcggaaagaacaacgatgcacctgctggtactaacatcacttgcacaggtaacgcatgtcctgaagtagaaaaagctgatgctacatttctttactcttttgaagatagcggcgtcggcgatgttaccggtttcttagctctggataacacaaacaaacttatcgtccttagcttcagaggctctcgctcaatcgaaaactggatcggtaaccttaattttgacttgaaagaaatcaacgatatctgctctggttgccgtggccatgacggattcacatcatcttggagaagcgtcgcagacacgcttcgccaaaaagtagaagatgccgtacgcgaacacccagattacagagtagttttcacaggtcactctcttggcggagctttagcaacagtagcaggcgctgatctccgcggtaacggatacgacattgatgtcttctcttacggcgctccgcgcgtcggtaacagagcgtttgctgaatttttaactgtacaaacaggcggaactctttatcgcatcactcacacaaacgatattgtcccgcgcttacctccgagagaatttggttactcacacagctctcctgaatactggatcaaaagcggtacattggtacctgttactcgaaacgatatcgtcaaaattgaaggaattgacgccaccggcggcaacaaccaaccgaacatccctgacatcccggcacacctttggtacttcggcttaatcggaacatgcctttaaaagctt
为在真菌细胞中生产，订购了合成的DNA序列(美国英杰公司)，这些序列含有来自丝状真菌疏绵状嗜热丝孢菌(Thermomyceslanuginosus)的脂肪酶的编码区的变体，并携带attB1和attB2位点以容许BP重组克隆进pDonor221载体(美国英杰公司)。pDonor-TLL质粒的序列经供应商核实。经由LR技术将所有变体与目的载体pTTTpyr2重组，从而得到最终的表达质粒pTTTpyrG2-TLL变体。在大肠杆菌TOP10菌株中生成质粒，将其纯化，然后用于真菌转化(下文将进一步描述)。
表达载体含有里氏木霉cbhI启动子和终止子区域，从而使得所关注的基因能够进行强的诱导型表达；此外，还包含构巢曲霉(Aspergillusnidulans)amdS和pyr2选择标记，从而赋予转化体在含乙酰胺但不含尿苷的基本培养基中生长的能力。由于源自里氏木霉的端粒区域，所以该质粒在所述真菌细胞中自维持。使用复制型质粒可以导致转化率升高，而且可以避免整合型真菌转化中观察到的基因座依赖性表达的问题。
采用PEG-原生质体方法，用质粒DNA转化缺失了主要纤维素酶的里氏木霉菌株(Δcbh1Δcbh2Δegl1Δegl2Δegl3Δbgl1pyr2-)。为制备原生质体，将孢子放入含5mM尿苷的木霉属基本培养基(MM)(20g/L葡萄糖，15g/LKH2PO4，pH4.5，5g/L(NH4)2SO4，0.6g/LMgSO4·7H2O，0.6g/LCaCl2·2H2O，1ml1000X里氏木霉痕量元素溶液(5g/LFeSO4·7H2O，1.4g/LZnSO4·7H2O，1.6g/LMnSO4·H2O，3.7g/LCoCl2·6H2O))，在28℃温度下以150rpm振荡速度培养16至24小时。离心收集萌发的孢子，用50mg/mlGlucanexG200(瑞士的诺维信公司)溶液处理以溶解掉真菌细胞壁。然后，采用标准方法(如的等人[Gene61(1987)155-164]所述的方法)进一步制备原生质体。
将总体积50μl、含有大约1μgDNA和5×106个原生质体的转化混合物与200μl25％PEG溶液混合，用等体积的1.2M山梨糖醇/10mMTris(pH7.5)/10mMCaCl2溶液稀释，接着与1ml含1M山梨糖醇和10mMNH4Cl的3％琼脂糖基本培养基(MM)(20g/L葡萄糖，15g/LKH2PO4，1g/LCaCl2·2H2O，1g/LMgSO4·7H2O，pH4.5，2.5ml/L400X里氏木霉痕量元素溶液(175g/L柠檬酸，200g/LFeSO4π7H2O，16g/LZnSO4·7H2O，3.2g/LCuSO4·5H2O，1.4g/LMnSO4·H2O，0.8g/L硼酸))混合。转化体生长后，合并各孔中的孢子，放入装有含10mM乙酰胺(用于额外的选择压力)的MM的新板上重新贴壁(repatch)。一旦形成孢子，收获孢子并用于在24孔微量滴定板中(用于筛选)或摇瓶中(用于验证研究)于如下生产培养基中接种液体培养物：37g/L葡萄糖，1g/L槐糖，9g/L酪蛋白氨基酸(casminoacids)，10g/L(NH4)2SO4，5g/LKH2PO4，1g/LCaCl2·2H2O，1g/LMgSO4·7H2O，33g/L1，4-哌嗪二丙磺酸，pH5.5，2.5ml/L400X里氏木霉痕量元素。在24孔MTP中加入1ml生产培养基以生产变体。对于摇瓶，按比例增加体积。
在28℃和80％湿度，200rpm振荡培养板6天。过滤收集培养物上清液。对于更大规模发酵和纯化，将选定转化体的孢子接种到2L摇瓶内的500ml同样的生产培养基中，28℃搅动培养6天，然后过滤、浓缩。利用SDS-PAGE分析，监测变体的表达。
下表2-1显示具有位置组合的组合文库，其中在所述位置进行了置换。
B.生成TLL合成基因变体
将pHYT-TLLwt质粒DNA(在美国专利申请61/713,436中描述)用作模板来产生变体，每种变体在成熟区(美国专利申请61/713,436中的SEQIDNO：3)的预选位点处分别具有一组固定的突变。委托GeneArt股份公司(德国雷根斯堡市)利用基因合成技术生成这些变体。这些变体由含有编码TLL变体序列的表达质粒的转化枯草芽孢杆菌细胞组成。GeneArt公司提供在96孔板中的变体，每孔一种变体，培养物冷冻在甘油中。下表2-2列出了以这种方式生成的变体。
表2-2：TLL合成基因变体的列表：A018K/D027S，A018K/E045F，A018K/L075D，A018K/L075Q，A018K/T189D，D027E/G163P，D027N/E056K，D027N/N233Q，D027S/N033D，D027S/P256T，D111A/L264R，D130A/L264R，D130A/T189Q，D130A/V187N，D130A/V187T，E045F/N073R，E045F/T189D，F051T/L075R，G023K/D130A，G023K/L264R，G023Q/L075Q，K024A/L075Q，K024A/V154I，L075Q/D130A，L075Q/G156W，L075R/L264R，L227M/L264R，N011K/D027S，N033D/E045F，N073R/L075D，N094R/G156W，P029E/N033D，S058M/L264R，V187T/L264R，A018K/A049V/L075Q，A018K/A049V/V187T，A018K/D027S/E045F，A018K/D027S/N073R，A018K/D027S/T189D，A018K/E045F/N073R，A018K/E045F/T189D，A018K/E045F/V187T，A018K/G023K/G156W，A018K/G023K/L075Q，A018K/G023Q/L075Q，A018K/K024A/V154I，A018K/L075D/T189D，A018K/L075Q/G156W，A018K/L075Q/V187T，A018K/N033D/L075D，A018K/N033D/T189D，A018K/N073R/L075D，A018K/P029E/T189D，A018K/V154I/G156W，A049V/L075Q/T189D，A049V/V187T/T189D，D027E/S058M/G163P，D027N/N233Q/P256T，D027N/S058M/P256T，D027Q/N233Q/P256T，D027Q/S058M/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/N073R/L075D，D027S/N233Q/P256T，D027S/S058M/P256T，D111A/D130A/L264R，D130A/T189Q/L264R，D130A/V154I/G156W，D130A/V187N/L264R，D130A/V187T/L264R，E045F/L075D/T189D，E045F/N073R/L075D，E056K/D130A/L264R，E056K/D130A/T189Q，F051T/I252Q/L264R，G023K/D130A/L264R，G023K/D130A/V187T，G023K/E056K/V187T，G023K/L075R/L264R，G023K/V187N/L264R，G023K/V187T/L264R，G023Q/A049V/T189D，G023Q/D111A/L264R，G023Q/D130A/G156W，G023Q/E045F/V077I，G023Q/E056K/L075R，G023Q/K024A/G156W，G023Q/L075Q/V077I，G023Q/L075Q/V187T，G163P/L227M/L264R，I090F/N233Q/P256T，K024A/D130A/V154I，K024A/L075Q/G156W，K024A/L075Q/V077I，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/D130A/V187T，L075Q/G156W/T189D，L075Q/G156W/V187N，L075Q/G156W/V187T，L075Q/N094R/V154I，L075Q/V077I/T189D，L075Q/V154I/V187T，L075Q/V187N/L264R，L075Q/V187T/L264R，L075Q/V187T/T189D，L075R/D130A/L264R，L075R/D130A/V187T，L075R/V187N/L264R，L075R/V187T/L264R，N011K/D027S/S058M，N011K/N233Q/P256T，N033D/E045F/L075D，N033D/E045F/N073R，N033D/N073R/T189D，N073R/L075D/T189D，N094R/D130A/V187T，P029E/E045F/L075D，P029E/L075D/T189D，P029E/N033D/E045F，P029E/N033D/L075D，P029E/N073R/L075D，Q004D/D027S/P256T，Q004D/N233Q/P256T，Q004D/S058M/N233Q，S058M/L227M/L264R，V077I/D130A/V154I，V077I/V187A/T189D，V187N/T189Q/L264R，A018K/A049V/L075Q/T189D，A018K/A049V/L075Q/V187T，A018K/D027S/E045F/N073R，A018K/D027S/E045F/T189D，A018K/D027S/L075D/T189D，A018K/D027S/N033D/L075D，A018K/D027S/P029E/T189D，A018K/D111A/G156W/T189Q，A018K/D130A/G156W/V187T，A018K/E045F/A049V/G156W，A018K/E045F/A049V/V187T，A018K/E045F/L075D/T189D，A018K/E045F/L075Q/T189D，A018K/E045F/L075Q/V187T，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/T189D，A018K/G023Q/A049V/V187T，A018K/G023Q/E045F/T189D，A018K/G023Q/E045F/V187T，A018K/G023Q/G156W/V187T，A018K/G023Q/L075Q/T189D，A018K/G023Q/L075Q/V077I，A018K/G023Q/L075Q/V187T，A018K/G023Q/L075R/D130A，A018K/G023Q/V077I/G156W，A018K/G023Q/V077I/V187T，A018K/G023Q/V187T/T189D，A018K/K024A/D130A/G156W，A018K/L075Q/G156W/T189D，A018K/L075Q/G156W/V187T，A018K/L075Q/N094R/D111A，A018K/L075Q/N094R/D130A，A018K/L075Q/N094R/V187Q，A018K/L075Q/V077I/N094R，A018K/L075Q/V187T/T189D，A018K/N033D/L075D/T189D，A018K/N073R/L075D/T189D，A018K/P029E/N033D/L075D，A018K/P029E/N033D/T189D，A018K/P029E/N073R/L075D，A018K/V154I/G156W/V187T，A049V/L075Q/V077I/T189D，A049V/L075Q/V187T/T189D，D027E/D048Q/G163P/L264R，D027E/D130A/N233Q/L264R，D027E/D137Q/G163P/L227M，D027E/D137Q/L227M/L264R，D027E/G163P/L227M/L264R，D027E/L227M/N233Q/L264R，D027E/S058M/G163P/L264R，D027N/E056K/N233Q/P256T，D027N/E056K/S058M/P256T，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027Q/S058M/I090F/P256T，D027S/D130A/I252Q/L264R，D027S/E045F/L075D/T189D，D027S/E045F/N073R/L075D，D027S/E045F/N073R/T189D，D027S/E056K/D111A/V154I，D027S/E056K/I090F/P256T，D027S/N033D/E045F/N073R，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/L075D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/L075D，D027S/P029E/E045F/N073R，D027S/P029E/E045F/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D027S/P029E/N033D/N073R，D027S/P029E/N033D/T189D，D048Q/D130A/G163P/L264R，D048Q/D137Q/G163P/L227M，D048Q/G163P/N233Q/L264R，D111A/D130A/V154I/G156W，D111A/D130A/V154I/L264R，D111A/D130A/V187T/L264R，D130A/G163P/L227M/L264R，D130A/G163P/N233Q/L264R，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，E056K/D130A/V187N/L264R，E056K/D130A/V187T/L264R，E056K/L075Q/V187N/L264R，F051T/D130A/I252Q/L264R，F051T/L075G/I252Q/L264R，F051T/L075Q/I252Q/L264R，G023K/D027Q/F051T/L075Q，G023K/D027S/L075Q/L264R，G023K/D111A/D130A/L264R，G023K/D130A/G156W/T189Q，G023K/D130A/V187N/L264R，G023K/D130A/V187T/L264R，G023K/D130A/V187T/T189Q，G023K/E056K/D130A/L264R，G023K/E056K/D130A/V187N，G023K/E056K/L075R/L264R，G023K/E056K/V187T/L264R，G023K/K024A/V154I/V187N，G023K/L075Q/D130A/V187N，G023K/L075Q/G156W/L264R，G023K/L075Q/G156W/V187N，G023K/L075R/D130A/L264R，G023K/L075R/V187N/L264R，G023K/L075R/V187T/L264R，G023Q/A049V/L075Q/T189D，G023Q/A049V/L075Q/V077I，G023Q/A049V/V077I/G156W，G023Q/D027S/D111A/G156W，G023Q/E045F/A049V/T189D，G023Q/E045F/A049V/V187T，G023Q/E045F/L075Q/G156W，G023Q/E045F/L075Q/V187T，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/K024A/D130A/G156W，G023Q/K024A/L075R/G156W，G023Q/K024A/L075R/V154I，G023Q/K024A/V077I/G156W，G023Q/L075Q/D130A/L264R，G023Q/L075Q/G156W/V187N，G023Q/L075Q/G156W/V187T，G023Q/L075Q/V187T/L264R，G023Q/V154I/G156W/V187N，G091Q/V187T/I252Q/L264R，K024A/D130A/V154I/V187T，K024A/L075Q/D130A/G156W，K024A/L075Q/D130A/V154I，K024A/L075Q/V187T/T189Q，K024A/L075R/G156W/V187N，L075G/D130A/V187T/I252Q，L075G/G091Q/V187N/L264R，L075G/V187H/I252Q/L264R，L075Q/D130A/G156W/V187N，L075Q/D130A/V154I/G156W，L075Q/G156W/V187T/L264R，L075Q/V077I/D130A/V187Q，L075Q/V077I/G156W/V187N，L075Q/V077I/N094R/G156W，L075Q/V077I/V154I/V187Q，L075Q/V077I/V187T/T189D，L075Q/V187N/T189Q/L264R，L075R/D111A/V154I/G156W，L075R/D130A/V154I/L264R，L075R/D130A/V187T/L264R，N011K/D027N/E056K/S058M，N011K/D027N/S058M/P256T，N011K/D027Q/S058M/P256T，N011K/D027S/E056K/P256T，N011K/D027S/N233Q/P256T，N011K/D027S/S058M/N233Q，N011K/E056K/N233Q/P256T，N011K/E056K/S058M/P256T，N011K/G023K/L075Q/D111A，N011K/S058M/N233Q/P256T，N033D/E045F/N073R/T189A，P029E/E045F/L075D/T189D，P029E/E045F/N073R/L075D，P029E/E045F/N073R/T189D，P029E/N033D/E045F/L075D，P029E/N033D/L075D/T189D，P029E/N073R/L075D/T189D，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/D027S/I090F/P256T，Q004D/N011K/D027N/P256T，Q004D/N011K/D027Q/N233Q，Q004D/N011K/D027S/P256T，Q004D/N011K/E056K/S058M，S058M/D137Q/GI63P/N233Q，S058M/G163P/L227M/L264R，S058M/G163P/N233Q/L264R，S058M/L075Q/G091Q/I252Q，S058M/L075Q/I252Q/L264R，A018K/A049V/L075Q/V187T/T189D，A018K/D027S/E045F/L075D/T189D，A018K/D027S/E045F/N073R/D137V，A018K/D027S/N033D/E045F/T189D，A018K/D027S/N033D/L075D/T189D，A018K/D027S/N033D/N073R/T189D，A018K/D027S/N073R/L075D/T189D，A018K/D027S/P029E/E045F/T189D，A018K/D027S/P029E/N033D/L075D，A018K/D027S/P029E/N073R/L075D，A018K/D130A/G156W/V187N/L264R，A018K/D130A/G156W/V187T/L264R，A018K/E045F/A049V/L075Q/G156W，A018K/E045F/A049V/L075Q/T189D，A018K/E045F/A049V/L075Q/V187T，A018K/E045F/N073R/L075D/T189D，A018K/G023K/D130A/V154I/G156W，A018K/G023K/K024A/D130A/G156W，A018K/G023K/K024A/D130A/V154I，A018K/G023K/K024A/L075R/N094R，A018K/G023K/L075Q/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/A049V/L075Q/G156W，A018K/G023Q/A049V/L075Q/V077I，A018K/G023Q/A049V/L075Q/V187T，A018K/G023Q/A049V/V077I/V187T，A018K/G023Q/E045F/A049V/L075Q，A018K/G023Q/E045F/A049V/V187T，A018K/G023Q/E045F/L075Q/V187T，A018K/G023Q/E045F/V077I/T189D，A018K/G023Q/K024A/D130A/V154I，A018K/G023Q/K024A/D130A/V187Q，A018K/G023Q/K024A/L075Q/G156W，A018K/G023Q/L075Q/G156W/V187T，A018K/G023Q/L075Q/V187T/T189D，A018K/G023Q/V077I/D130A/G156W，A018K/G023Q/V077I/V187T/T189D，A018K/K024A/L075Q/D130A/L264R，A018K/K024A/L075Q/V154I/G156W，A018K/K024A/L075Q/V154I/T189D，A018K/K024A/L075R/D130A/V154I，A018K/K024A/L075R/N094R/D130A，A018K/K024A/N094R/D130A/V187N，A018K/K024A/N094R/G156W/V187T，A018K/L075Q/D111A/D130A/V187T，A018K/L075Q/G156W/V187T/T189D，A018K/L075Q/N094R/D130A/V187N，A018K/L075Q/N094R/G156W/V187N，A018K/L075Q/V077I/N094R/G156W，A018K/L075Q/V187T/T189Q/L264R，A018K/N033D/E045F/L075D/T189D，A018K/P029E/E045F/N073R/L075D，A018K/P029E/N033D/N073R/L075D，A018K/V077I/G156W/V187T/T189D，D027E/D048Q/D137Q/L227M/L264R，D027E/D048Q/G163P/N233Q/L264R，D027E/D048Q/L227M/N233Q/L264R，D027E/D048Q/S058M/G163P/N233Q，D027E/D130A/D137Q/G163P/L264R，D027E/D130A/G163P/L227M/L264R，D027E/D130A/G163P/N233Q/L264R，D027E/D137Q/G163P/L227M/L264R，D027E/D137Q/G163P/N233Q/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L264R，D027E/S058M/D137Q/G163P/N233Q，D027E/S058M/G163P/L227M/L264R，D027N/E056K/S058M/N233Q/P256T，D027N/S058M/I090F/N233Q/P256T，D027Q/E056K/I090F/N233Q/P256T，D027Q/E056K/S058M/N233Q/P256T，D027Q/F051T/L075Q/D130A/L264R，D027S/E056K/D111A/V187N/L264R，D027S/E056K/S058M/N233Q/P256T，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D027S/L075Q/D111A/D130A/V187N，D027S/L075R/V154I/T189Q/L264R，D027S/N033D/E045F/N073R/L075D，D027S/N033D/N073R/L075D/T189D，D027S/P029E/E045F/N073R/L075D，D027S/P029E/E045F/N073R/T189D，D027S/P029E/N033D/E045F/N073R，D027S/P029E/N033D/N073R/L075D，D027S/P029E/N073R/L075D/T189D，D048Q/D130A/D137Q/G163P/L264R，D048Q/D130A/G163P/L227M/L264R，D048Q/D137Q/G163P/L227M/L264R，D048Q/D137Q/G163P/N233Q/L264R，D048Q/S058M/D130A/N233Q/L264R，D048Q/S058M/D137Q/G163P/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/D137Q/N233Q/L264R，D048Q/S058M/G163P/L227M/L264R，D048Q/S058M/G163P/N233Q/L264R，D130A/D137Q/G163P/L227M/L264R，D130A/D137Q/G163P/N233Q/L264R，D130A/G163P/L227M/N233Q/L264R，D130A/V154I/G156W/V187N/T189Q，E045F/A049V/L075Q/V187T/T189D，E056K/D130A/V187H/I252Q/L264R，E056K/L075G/V187N/I252Q/L264R，E056K/L075Q/G156W/T189Q/L264R，E056K/L075R/D130A/V187N/L264R，E056K/L075R/D130A/V187T/L264R，E056K/N094R/G156W/V187N/L264R，E056K/S058M/I090F/N233Q/P256T，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/G091Q/D130A/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/L264R，F051T/L075Q/V187N/I252Q/L264R，F051T/L075R/V187N/I252Q/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/D027S/E056K/V187T/L264R，G023K/D111A/V154I/V187T/L264R，G023K/E056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K/G023K/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/G023K/L075R/V077I/N094R/D130A/G156W/V187Q，A018K/G023Q/E045F/L075Q/V077I/G156W/V187T/T189D，A018K/G023Q/K024A/L075Q/V077I/D130A/G156W/V187Q，A018K/G023Q/K024A/L075R/D130A/V154I/G156W/V187N，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，A018K/G023Q/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/K024A/L075Q/V077I/N094R/D130A/V154I/G156W，A018K/K024A/L075R/D130A/V154I/G156W/V187T/L264R，A018K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/E056K/L075G/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/D027S/F051T/E056K/L075R/D130A/V187T/L264R，G023K/D027S/F051T/S058M/L075Q/V187H/I252Q/L264R，G023K/E056K/L075Q/D130A/V154I/V187N/T189Q/L264R，G023K/F051T/E056K/L075R/G091Q/D130A/V187T/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187T/L264R，G023K/K024A/L075R/N094R/V154I/G156W/V187T/T189Q，G023K/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，G023K/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/E056K/S058M/L075R/G091Q/V187N/I252Q，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/E056K/L075Q/D111A/V154I/V187N/T189Q/L264R，G023Q/E056K/L075Q/N094R/D111A/G156W/T189Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/K024A/L075Q/V077I/D130A/V154I/G156W/V187N，G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/V187N，G023Q/L075Q/D111A/D130A/V154I/G156W/V187N/L264R，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，G023Q/L075Q/N094R/D130A/V154I/G156W/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，K024A/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，N011K/G023K/D027S/L075Q/D111A/D130A/V154I/T189Q，N011K/G023K/D027S/L075R/V154I/G156W/V187N/T189Q，N011K/G023K/E056K/L075Q/D130A/V154I/T189Q/L264R，N011K/G023K/L075Q/D111A/G156W/V187N/T189Q/L264R，N011K/G023Q/D027S/L075Q/N094R/V154I/G156W/T189Q，N011K/G023Q/D027S/N094R/V154I/G156W/T189Q/L264R，N011K/G023Q/L075Q/D111A/D130A/V154I/V187N/L264R，N011K/G023Q/L075Q/N094R/V154I/G156W/T189Q/L264R，A018K/G023K/K024A/L075Q/N094R/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/V154I/G156W/V187N，A018K/G023K/K024A/L075Q/N094R/D130A/V187T/T189Q/L264K，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，A018K/G023Q/D027S/L075Q/V077I/R108K/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/G156W/V187H，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/P029E/S058M/V077I/R108K/H135F/G156W，A018K/G023Q/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W/V187N，A018K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V187N/T189Q/L264R，G023K/D027S/F051T/E056K/S058M/L075Q/G091Q/V187N/L264R，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/D027S/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023K/L075Q/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/F051T/E056K/L075G/G091Q/V187N/I252Q/L264R，G023Q/D027S/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，G023Q/D027S/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/G023K/D027S/E056K/L075R/D111A/G156W/V187N/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，N011K/G023Q/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/E045F/A049V/S058M/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/S058M/L075Q/V077I/R108K/H135F/G156W/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023K/D027S/F051T/E056K/S058M/L075G/G091Q/D130A/I252Q/L264R，G023Q/D027S/F051T/E056K/L075R/G091Q/D130A/V187H/I252Q/L264R，G023Q/D027S/F051T/E056K/S058M/L075R/G091Q/D130A/V187H/I252Q，G023Q/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023K/L075R/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，N011K/G023Q/D027S/E056K/L075R/D130A/V154I/G156W/T189Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，N011K/G023Q/L075Q/N094R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T，N011K/G023Q/D027S/L075Q/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/H135F/V187T/T189D，A018K/G023Q/D027S/P029E/N033D/E045F/A049V/S058M/N073S/K074S/L075Q/V077I/N101D/R108K/H135F/D137V/G156W/V187T/T189D
实施例3
TLL组合变体的酯水解活性
按实施例1所述的程序，分析测定了按实施例2所述方法生成的TLL变体作用于丁酸对硝基苯酯、辛酸对硝基苯酯、棕榈酸对硝基苯酯这三种底物的脂肪酶活性。计算了变体相比TLL(SEQIDNO：1)或参考序列SEQIDNO：2的性能指数。
表3-1：下面示出pH8时，相比TLL(SEQIDNO：1)，水解丁酸对硝基苯酯的性能指数＞1的TLL变体。
A018K/L075D，D027N/E056K，D027S/N033D，D130A/L264R，D130A/V187N，E045F/N073R，K024A/L075Q，K024A/V154I，L075Q/D130A，L075R/L264R，P029E/N033D，V187T/L264R，A018K/E045F/N073R，A018K/G023K/G156W，A018K/G023K/L075Q，A018K/L075D/T189D，A018K/V154I/G156W，A049V/L075Q/T189D，A049V/V187T/T189D，D027E/S058M/G163P，D027N/N233Q/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D130A/V187T/L264R，E045F/L075D/T189D，E056K/D130A/T189Q，G023K/E056K/V187T，G023Q/A049V/T189D，G023Q/D111A/L264R，G023Q/L075Q/V187T，G163P/L227M/L264R，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/G156W/V187N，L075Q/V187N/L264R，L075R/D130A/L264R，L075R/D130A/V187T，L075R/V187T/L264R，N073R/L075D/T189D，P029E/N033D/E045F，V077I/D130A/V154I，V187N/T189Q/L264R，A018K/D027S/E045F/N073R，A018K/D027S/N033D/L075D，A018K/E045F/L075D/T189D，A018K/E045F/L075Q/T189D，A018K/G023K/D111A/T189Q，A018K/G023Q/E045F/T189D，A018K/G023Q/E045F/V187T，A018K/G023Q/L075R/D130A，A018K/G023Q/V077I/V187T，A018K/L075Q/G156W/V187T，A018K/L075Q/N094R/D130A，A018K/L075Q/V077I/N094R，A018K/P029E/N073R/L075D，D027E/D048Q/G163P/L264R，D027E/D137Q/G163P/L227M，D027E/S058M/G163P/L264R，D027N/E056K/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/E045F/N073R/L075D，D027S/E045F/N073R/T189D，D027S/N033D/E045F/N073R，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/N073R，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D048Q/G163P/N233Q/L264R，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，G023K/D027Q/F051T/L075Q，G023K/L075Q/G156W/V187N，G023K/L075R/D130A/L264R，G023Q/A049V/L075Q/V077I，G023Q/A049V/V077I/G156W，G023Q/D027S/D111A/G156W，G023Q/K024A/L075R/G156W，G023Q/K024A/L075R/V154I，G023Q/L075Q/D130A/L264R，G023Q/V154I/G156W/V187N，K024A/L075Q/D130A/V154I，K024A/L075R/G156W/V187N，L075G/G091Q/V187N/L264R，L075Q/D130A/G156W/V187N，L075Q/V077I/G156W/V187N，N011K/D027S/E056K/P256T，N011K/E056K/N233Q/P256T，N011K/S058M/N233Q/P256T，N033D/E045F/N073R/T189A，P029E/N033D/L075D/T189D，P029E/N073R/L075D/T189D，Q004D/D027N/S058M/P256T，Q004D/D027S/I090F/P256T，Q004D/N011K/D027Q/N233Q，S058M/D137Q/G163P/N233Q，S058M/L075Q/G091Q/I252Q，A018K/A049V/L075Q/V187T/T189D，A018K/D027S/E045F/L075D/T189D，A018K/D027S/E045F/N073R/D137V，A018K/D027S/N033D/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/G023K/K024A/L075R/N094R，A018K/G023Q/E045F/A049V/V187T，A018K/G023Q/E045F/L075Q/V187T，A018K/G023Q/K024A/D130A/V154I，A018K/G023Q/K024A/L075Q/G156W，A018K/G023Q/L075Q/G156W/V187T，A018K/G023Q/L075Q/V187T/T189D，A018K/G023Q/V077I/D130A/G156W，A018K/G023Q/V077I/V187T/T189D，A018K/K024A/L075Q/D130A/L264R，A018K/K024A/L075Q/V154I/T189D，A018K/K024A/L075R/D130A/V154I，A018K/K024A/N094R/D130A/V187N，A018K/L075Q/D111A/D130A/V187T，A018K/L075Q/N094R/D130A/V187N，A018K/L075Q/N094R/G156W/V187N，A018K/P029E/N033D/N073R/L075D，A018K/V077I/G156W/V187T/T189D，D027E/S058M/D130A/G163P/L264R，D027E/S058M/D137Q/G163P/N233Q，D027E/S058M/G163P/L227M/L264R，D027N/S058M/I090F/N233Q/P256T，D027S/E056K/D111A/V187N/L264R，D027S/E056K/S058M/N233Q/P256T，D027S/L075Q/D111A/D130A/V187N，D027S/L075R/V154I/T189Q/L264R，D027S/P029E/N033D/E045F/N073R，D048Q/D130A/G163P/L227M/L264R，D048Q/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/G163P/L227M/L264R，D048Q/S058M/G163P/N233Q/L264R，D130A/D137Q/G163P/N233Q/L264R，E045F/A049V/L075Q/V187T/T189D，F051T/S058M/L075Q/G091Q/I252Q，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/K024A/L075R/D130A/G156W，G023K/L075Q/D111A/D130A/V187T，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187N/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/A049V/L075Q/G156W/V187T，G023Q/E045F/A049V/L075Q/V077I，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/K024A/L075R/D130A/V154I，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/D111A/V154I/V187T，K024A/L075R/G156W/V187N/T189Q，K024A/L075R/V154I/G156W/V187Q，L075Q/D130A/V187T/T189Q/L264R，L075Q/N094R/D130A/G156W/V187T，L075R/D111A/D130A/V187N/T189Q，L075R/D130A/V154I/T189Q/L264R，L075R/D130A/V187T/T189Q/L264R，N011K/D027N/E056K/N233Q/P256T，N011K/D027Q/E056K/S058M/N233Q，N011K/D027Q/I090F/N233Q/P256T，N011K/D027S/E056K/N233Q/P256T，N011K/G023K/D111A/G156W/L264R，P029E/N033D/E045F/L075D/T189D，Q004D/D027S/E056K/N233Q/P256T，Q004D/D027S/S058M/N233Q/P256T，Q004D/N011K/D027Q/E056K/N233Q，S058M/D130A/D137Q/G163P/L264R，S058M/D130A/G163P/L227M/L264R，S058M/G163P/L227M/N233Q/L264R，A018K/D027S/N033D/N073R/L075D/T189D，A018K/G023K/K024A/N094R/D130A/V154I，A018K/G023K/L075Q/D130A/G156W/V187N，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023K/L075R/N094R/D130A/V187N，A018K/G023Q/A049V/L075Q/V077I/G156W，A018K/G023Q/A049V/V077I/V187T/T189D，A018K/G023Q/E045F/A049V/G156W/V187T，A018K/G023Q/E045F/A049V/L075Q/T189D，A018K/G023Q/E045F/G156W/V187T/T189D，A018K/G023Q/E045F/L075Q/G156C/V187T，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/G023Q/L075Q/G156W/V187T/T189D，A018K/G023Q/L075Q/V077I/N094R/V154I，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/N094R/V154I/V187N，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/K024A/L075R/D130A/V187N/T189Q，A018K/K024A/V077I/N094R/D130A/G156W，A018K/L075Q/D111A/V154I/T189Q/L264R，A018K/L075Q/D130A/V187N/T189Q/L264R，A018K/N033D/E045F/N073R/L075D/T189D，A018K/N094R/D111A/D130A/V154I/V187N，A018K/P029E/N033D/E045F/L075D/T189D，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/E056K/S058M/I090F/N233Q/P256T，D027Q/F051T/L075Q/D130A/V187H/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D027S/P029E/N033D/E045F/N073R/T189D，D048Q/S058M/D137Q/G163P/N233Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/G091Q/D130A/V187H/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/N094R/D130A/G156W，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/K024A/L075Q/D130A/V154I/G156W，G023Q/L075Q/N094R/D111A/G156W/L264R，G023Q/L075R/V154I/G156W/V187N/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/N094R/D130A/V154I/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，K024A/L075Q/V154I/V187T/T189Q/L264R，L075Q/D111A/D130A/V154I/G156W/T189Q，L075Q/D130A/V154I/G156W/V187N/L264R，L075Q/N094R/D111A/G156W/T189Q/L264R，L075R/D130A/V154I/G156W/V187N/L264R，N011K/D027N/E056K/S058M/N233Q/P256T，N011K/D027S/E056K/S058M/I090F/P256T，Q004D/D027N/E056K/S058M/N233Q/P256T，Q004D/N011K/D027Q/E056K/S058M/N233Q，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，S058M/D130A/G163P/L227M/N233Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/E045F/L075Q/V077I/G156W/V187T/T189D，A018K/G023K/K024A/L075Q/D130A/G156W/V187N，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/K024A/L075R/V154I/G156W/V187Q，A018K/G023K/K024A/N094R/V154I/V187T/L264R，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/E045F/A049V/L075Q/G156W/V187T，A018K/G023Q/E045F/L075Q/G156W/V187T/T189D，A018K/G023Q/K024A/L075Q/V077I/N094R/V154I，A018K/G023Q/K024A/L075R/N094R/G156W/V187N，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/L075Q/D111A/D130A/V154I/G156W/L264R，A018K/L075Q/N094R/D111A/V154I/V187T/L264R，D027E/D048Q/S058M/D130A/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/N233Q/L264R，D027Q/F051T/L075G/D130A/V187H/I252Q/L264R，D027S/E056K/L075R/D111A/G156W/V187N/L264R，D027S/F051T/L075G/G091Q/V187T/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，E056K/L075Q/D111A/D130A/G156W/V187N/T189Q，G023K/D027Q/F051T/L075G/D130A/V187H/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023K/D027S/L075Q/N094R/V154I/G156W/T189Q，G023K/K024A/L075Q/D111A/D130A/T189Q/L264R，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023Q/E056K/L075Q/D111A/G156W/V187N/L264R，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/L075Q/D111A/D130A/V154I/G156W/L264R，G023Q/L075Q/D130A/V154I/V187N/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，K024A/L075Q/D111A/D130A/V154I/G156W/L264R，K024A/L075R/D130A/V154I/G156W/T189Q/L264R，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，N011K/D027S/D111A/D130A/V154I/V187N/T189Q，N011K/E056K/L075Q/D111A/D130A/V154I/L264R，Q004D/N011K/D027N/S058M/I090F/N233Q/P256T，Q004D/N011K/D027Q/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023K/L075R/V077I/N094R/D130A/G156W/V187Q，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/E056K/L075Q/D130A/V154I/V187N/T189Q/L264R，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/V187N，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，G023Q/L075Q/N094R/D130A/V154I/G156W/T189Q/L264R，K024A/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，N011K/G023K/D027S/L075Q/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075Q/N094R/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023K/L075Q/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，G023Q/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，N011K/G023Q/D027S/E056K/L075R/D130A/V154I/G156W/T189Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/N033D/E045F/A049V/S058M/N073S/K074S/L075Q/V0771/N101D/R108K/H135F/D137V/G156W/V187T/T189D
表3-2：下面示出相比TLL(SEQIDNO：1)表达性能指数≥0.5且pH8水解丁酸对硝基苯酯的性能指数＞1的TLL变体。
A018K/L075D，D027S/N033D，D130A/L264R，D130A/V187N，E045F/N073R，K024A/V154I，L075R/L264R，P029E/N033D，V187T/L264R，A018K/E045F/N073R，A018K/L075D/T189D，A018K/V154I/G156W，D027E/S058M/G163P，D027S/L075Q/G091Q，D027S/N033D/T189D，D130A/V187T/L264R，G023K/E056K/V187T，G023Q/A049V/T189D，G163P/L227M/L264R，L075G/D130A/V187H，L075Q/V187N/L264R，L075R/D130A/L264R，L075R/D130A/V187T，L075R/V187T/L264R，P029E/N033D/E045F，V187N/T189Q/L264R，A018K/E045F/L075D/T189D，A018K/G023K/D111A/T189Q，A018K/L075Q/V077I/N094R，A018K/P029E/N073R/L075D，D027E/D048Q/G163P/L264R，D027E/D137Q/G163P/L227M，D027E/S058M/G163P/L264R，D027Q/E056K/N233Q/P256T，D027S/E045F/N073R/T189D，D027S/N033D/E045F/N073R，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/N073R，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D048Q/G163P/N233Q/L264R，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，G023K/D027Q/F051T/L075Q，G023K/L075R/D130A/L264R，G023Q/A049V/V077I/G156W，G023Q/K024A/L075R/G156W，G023Q/L075Q/D130A/L264R，L075G/G091Q/V187N/L264R，N033D/E045F/N073R/T189A，P029E/N073R/L075D/T189D，Q004D/D027N/S058M/P256T，S058M/D137Q/G163P/N233Q，S058M/L075Q/G091Q/I252Q，A018K/D027S/E045F/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/G023Q/K024A/L075Q/G156W，A018K/G023Q/V077I/D130A/G156W，A018K/K024A/L075R/D130A/V154I，A018K/K024A/N094R/D130A/V187N，A018K/P029E/N033D/N073R/L075D，D027E/S058M/D130A/G163P/L264R，D027E/S058M/D137Q/G163P/N233Q，D027E/S058M/G163P/L227M/L264R，D027S/E056K/D111A/V187N/L264R，D027S/L075R/V154I/T189Q/L264R，D048Q/D130A/G163P/L227M/L264R，D048Q/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/G163P/L227M/L264R，D048Q/S058M/G163P/N233Q/L264R，D130A/D137Q/G163P/N233Q/L264R，E045F/A049V/L075Q/V187T/T189D，F051T/S058M/L075Q/G091Q/I252Q，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187N/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/V154I/G156W/V187Q，N011K/G023K/D111A/G156W/L264R，S058M/D130A/D137Q/G163P/L264R，S058M/D130A/G163P/L227M/L264R，S058M/G163P/L227M/N233Q/L264R，A018K/D027S/N033D/N073R/L075D/T189D，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D048Q/S058M/D137Q/G163P/N233Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/G091Q/D130A/V187H/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/F051T/D130A/V187T/I252Q/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/N094R/D130A/V154I/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，S058M/D130A/G163P/L227M/N233Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，D027E/D048Q/S058M/D130A/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T
表3-3：下面示出相比TLL(SEQIDNO：1)表达性能指数≥0.5且pH8水解丁酸对硝基苯酯的性能指数≥最大PI值的50％的TLL变体。
D130A/L264R，A018K/E045F/N073R，A018K/L075D/T189D，D027S/N033D/T189D，L075G/D130A/V187H，L075Q/V187N/L264R，L075R/D130A/L264R，L075R/D130A/V187T，A018K/E045F/L075D/T189D，A018K/L075Q/V077I/N094R，A018K/P029E/N073R/L075D，D027S/N033D/L075D/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，G023K/D027Q/F051T/L075Q，G023K/L075R/D130A/L264R，G023Q/K024A/L075R/G156W，L075G/G091Q/V187N/L264R，P029E/N073R/L075D/T189D，A018K/D027S/E045F/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/G023Q/K024A/L075Q/G156W，A018K/K024A/L075R/D130A/V154I，A018K/K024A/N094R/D130A/V187N，A018K/P029E/N033D/N073R/L075D，D027S/L075R/V154I/T189Q/L264R，D130A/D137Q/G163P/N233Q/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/N094R/G156W/V187N/T189Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/V154I/G156W/V187Q，N011K/G023K/D111A/G156W/L264R，A018K/D027S/N033D/N073R/L075D/T189D，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R
表3-4：下面示出，相比参考序列SEQIDNO：2，pH8水解丁酸对硝基苯酯的性能指数＞1的TLL变体。
A018K/L075D，D027N/E056K，D027S/N033D，D130A/L264R，D130A/V187N，E045F/N073R，K024A/L075Q，K024A/V154I，L075Q/D130A，L075R/L264R，P029E/N033D，V187T/L264R，A018K/E045F/N073R，A018K/G023K/G156W，A018K/G023K/L075Q，A018K/L075D/T189D，A018K/V154I/G156W，A049V/L075Q/T189D，A049V/V187T/T189D，D027E/S058M/G163P，D027N/N233Q/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D130A/V187T/L264R，E045F/L075D/T189D，E056K/D130A/T189Q，G023K/D130A/V187T，G023K/E056K/V187T，G023Q/A049V/T189D，G023Q/D111A/L264R，G023Q/L075Q/V187T，G163P/L227M/L264R，K024A/D130A/V154I，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/G156W/V187N，L075Q/V187N/L264R，L075R/D130A/L264R，L075R/D130A/V187T，L075R/V187T/L264R，N073R/L075D/T189D，N094R/D130A/V187T，P029E/N033D/E045F，Q004D/D027S/P256T，V077I/D130A/V154I，V187N/T189Q/L264R，A018K/D027S/E045F/N073R，A018K/D027S/N033D/L075D，A018K/E045F/L075D/T189D，A018K/E045F/L075Q/T189D，A018K/G023K/D111A/T189Q，A018K/G023Q/E045F/T189D，A018K/G023Q/E045F/V187T，A018K/G023Q/L075R/D130A，A018K/G023Q/V077I/V187T，A018K/L075Q/G156W/V187T，A018K/L075Q/N094R/D130A，A018K/L075Q/V077I/N094R，A018K/P029E/N073R/L075D，D027E/D048Q/G163P/L264R，D027E/D137Q/G163P/L227M，D027E/S058M/G163P/L264R，D027N/E056K/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/E045F/N073R/L075D，D027S/E045F/N073R/T189D，D027S/N033D/E045F/N073R，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/N073R，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D048Q/G163P/N233Q/L264R，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，G023K/D027Q/F051T/L075Q，G023K/K024A/V154I/V187N，G023K/L075Q/G156W/V187N，G023K/L075R/D130A/L264R，G023Q/A049V/L075Q/V077I，G023Q/A049V/V077I/G156W，G023Q/D027S/D111A/G156W，G023Q/E045F/L075Q/V187T，G023Q/K024A/L075R/G156W，G023Q/K024A/L075R/V154I，G023Q/L075Q/D130A/L264R，G023Q/V154I/G156W/V187N，K024A/L075Q/D130A/V154I，K024A/L075R/G156W/V187N，L075G/G091Q/V187N/L264R，L075Q/D130A/G156W/V187N，L075Q/V077I/G156W/V187N，N011K/D027S/E056K/P256T，N011K/E056K/N233Q/P256T，N011K/S058M/N233Q/P256T，N033D/E045F/N073R/T189A，P029E/N033D/L075D/T189D，P029E/N073R/L075D/T189D，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027S/I090F/P256T，Q004D/N011K/D027Q/N233Q，S058M/D137Q/G163P/N233Q，S058M/L075Q/G091Q/I252Q，A018K/A049V/L075Q/V187T/T189D，A018K/D027S/E045F/L075D/T189D，A018K/D027S/E045F/N073R/D137V，A018K/D027S/N033D/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/G023K/K024A/L075R/N094R，A018K/G023Q/E045F/A049V/V187T，A018K/G023Q/E045F/L075Q/V187T，A018K/G023Q/K024A/D130A/V154I，A018K/G023Q/K024A/L075Q/G156W，A018K/G023Q/L075Q/G156W/V187T，A018K/G023Q/L075Q/V187T/T189D，A018K/G023Q/V077I/D130A/G156W，A018K/G023Q/V077I/V187T/T189D，A018K/K024A/L075Q/D130A/L264R，A018K/K024A/L075Q/V154I/T189D，A018K/K024A/L075R/D130A/V154I，A018K/K024A/N094R/D130A/V187N，A018K/L075Q/D111A/D130A/V187T，A018K/L075Q/N094R/D130A/V187N，A018K/L075Q/N094R/G156W/V187N，A018K/P029E/N033D/N073R/L075D，A018K/V077I/G156W/V187T/T189D，D027E/S058M/D130A/G163P/L264R，D027E/S058M/D137Q/G163P/N233Q，D027E/S058M/G163P/L227M/L264R，D027N/S058M/I090F/N233Q/P256T，D027S/E056K/D111A/V187N/L264R，D027S/E056K/S058M/N233Q/P256T，D027S/L075Q/D111A/D130A/V187N，D027S/L075R/V154I/T189Q/L264R，D027S/P029E/N033D/E045F/N073R，D048Q/D130A/G163P/L227M/L264R，D048Q/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/G163P/L227M/L264R，D048Q/S058M/G163P/N233Q/L264R，D130A/D137Q/G163P/N233Q/L264R，E045F/A049V/L075Q/V187T/T189D，F051T/L075G/G091Q/D130A/L264R，F051T/L075Q/G091Q/D130A/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/K024A/L075R/D130A/G156W，G023K/L075Q/D111A/D130A/V187T，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187N/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/A049V/L075Q/G156W/V187T，G023Q/E045F/A049V/L075Q/V077I，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/K024A/L075R/D130A/V154I，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/D111A/V154I/V187T，K024A/L075R/G156W/V187N/T189Q，K024A/L075R/V154I/G156W/V187Q，L075Q/D130A/V187T/T189Q/L264R，L075Q/N094R/D130A/G156W/V187T，L075R/D111A/D130A/V187N/T189Q，L075R/D130A/V154I/T189Q/L264R，L075R/D130A/V187T/T189Q/L264R，N011K/D027N/E056K/N233Q/P256T，N011K/D027Q/E056K/S058M/N233Q，N011K/D027Q/I090F/N233Q/P256T，N011K/D027S/E056K/N233Q/P256T，N011K/G023K/D111A/G156W/L264R，P029E/N033D/E045F/L075D/T189D，Q004D/D027S/E056K/N233Q/P256T，Q004D/D027S/S058M/N233Q/P256T，Q004D/N011K/D027Q/E056K/N233Q，S058M/D130A/D137Q/G163P/L264R，S058M/D130A/G163P/L227M/L264R，S058M/G163P/L227M/N233Q/L264R，A018K/D027S/N033D/N073R/L075D/T189D，A018K/G023K/K024A/N094R/D130A/V154I，A018K/G023K/L075Q/D130A/G156W/V187N，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023K/L075R/N094R/D130A/V187N，A018K/G023Q/A049V/L075Q/V077I/G156W，A018K/G023Q/A049V/V077I/V187T/T189D，A018K/G023Q/E045F/A049V/G156W/V187T，A018K/G023Q/E045F/A049V/L075Q/T189D，A018K/G023Q/E045F/G156W/V187T/T189D，A018K/G023Q/E045F/L075Q/G156C/V187T，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/G023Q/L075Q/G156W/V187T/T189D，A018K/G023Q/L075Q/V077I/N094R/V154I，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/N094R/V154I/V187N，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/K024A/L075R/D130A/V187N/T189Q，A018K/K024A/V077I/N094R/D130A/G156W，A018K/L075Q/D111A/V154I/T189Q/L264R，A018K/L075Q/D130A/V187N/T189Q/L264R，A018K/N033D/E045F/N073R/L075D/T189D，A018K/N094R/D111A/D130A/V154I/V187N，A018K/P029E/N033D/E045F/L075D/T189D，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/E056K/S058M/I090F/N233Q/P256T，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027Q/F051T/L075Q/D130A/V187H/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D027S/P029E/N033D/E045F/N073R/T189D，D048Q/S058M/D137Q/G163P/N233Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/G091Q/D130A/V187H/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/N094R/D130A/G156W，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/L075Q/D130A/V154I/G156W，G023Q/L075Q/N094R/D111A/G156W/L264R，G023Q/L075R/V154I/G156W/V187N/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/N094R/D130A/V154I/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，K024A/L075Q/V154I/V187T/T189Q/L264R，L075Q/D111A/D130A/V154I/G156W/T189Q，L075Q/D130A/V154I/G156W/V187N/L264R，L075Q/N094R/D111A/G156W/T189Q/L264R，L075R/D130A/V154I/G156W/V187N/L264R，N011K/D027N/E056K/S058M/N233Q/P256T，N011K/D027S/E056K/S058M/I090F/P256T，N011K/G023Q/L075Q/D130A/V187N/T189Q，Q004D/D027N/E056K/S058M/N233Q/P256T，Q004D/N011K/D027Q/E056K/S058M/N233Q，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，S058M/D130A/G163P/L227M/N233Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/E045F/L075Q/V077I/G156W/V187T/T189D，A018K/G023K/K024A/L075Q/D130A/G156W/V187N，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/K024A/L075R/V154I/G156W/V187Q，A018K/G023K/K024A/N094R/V154I/V187T/L264R，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/E045F/A049V/L075Q/G156W/V187T，A018K/G023Q/E045F/L075Q/G156W/V187T/T189D，A018K/G023Q/K024A/L075Q/V077I/N094R/V154I，A018K/G023Q/K024A/L075R/N094R/G156W/V187N，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/L075Q/D111A/D130A/V154I/G156W/L264R，A018K/L075Q/N094R/D111A/V154I/V187T/L264R，D027E/D048Q/D130A/D137Q/G163P/N233Q/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/D048Q/S058M/D130A/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/D137Q/G163P/N233Q/L264R，D027Q/F051T/L075G/D130A/V187H/I252Q/L264R，D027S/E056K/L075R/D111A/G156W/V187N/L264R，D027S/F051T/L075G/G091Q/V187T/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，E056K/L075Q/D111A/D130A/G156W/V187N/T189Q，G023K/D027Q/F051T/L075G/D130A/V187H/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023K/D027S/L075Q/N094R/V154I/G156W/T189Q，G023K/K024A/L075Q/D111A/D130A/T189Q/L264R，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023Q/E056K/L075Q/D111A/G156W/V187N/L264R，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/L075Q/D111A/D130A/V154I/G156W/L264R，G023Q/L075Q/D130A/V154I/V187N/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，K024A/L075Q/D111A/D130A/V154I/G156W/L264R，K024A/L075R/D130A/V154I/G156W/T189Q/L264R，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，N011K/D027S/D111A/D130A/V154I/V187N/T189Q，N011K/E056K/L075Q/D111A/D130A/V154I/L264R，Q004D/N011K/D027N/S058M/I090F/N233Q/P256T，Q004D/N011K/D027Q/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023K/L075R/V077I/N094R/D130A/G156W/V187Q，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/E056K/L075Q/D130A/V154I/V187N/T189Q/L264R，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/V187N，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，G023Q/L075Q/N094R/D130A/V154I/G156W/T189Q/L264R，K024A/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，N011K/G023K/D027S/L075Q/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075Q/N094R/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023K/L075Q/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，G023Q/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T7T189Q，N011K/G023Q/D027S/E056K/L075R/D130A/V154I/G156W/T189Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/H135F/V187T/T189D，A018K/G023Q/D027S/P029E/N033D/E045F/A049V/S058M/N073S/K074S/L075Q/V077I/N101D/R108K/H135F/D137V/G156W/V187T/T189D
表3-5：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比SEQIDNO：2在pH8水解丁酸对硝基苯酯的性能指数≥1的TLL变体。
A018K/L075D，D027S/N033D，D130A/L264R，D130A/V187N，E045F/N073R，K024A/V154I，L075R/L264R，P029E/N033D，V187T/L264R，A018K/E045F/N073R，A018K/L075D/T189D，A018K/V154I/G156W，D027E/S058M/G163P，D027S/L075Q/G091Q，D027S/N033D/T189D，D130A/V187T/L264R，G023K/D130A/V187T，G023K/E056K/V187T，G023Q/A049V/T189D，G163P/L227M/L264R，K024A/D130A/V154I，L075G/D130A/V187H，L075Q/V187N/L264R，L075R/D130A/L264R，L075R/D130A/V187T，L075R/V187T/L264R，N094R/D130A/V187T，P029E/N033D/E045F，Q004D/D027S/P256T，V187N/T189Q/L264R，A018K/E045F/L075D/T189D，A018K/G023K/D111A/T189Q，A018K/L075Q/V077I/N094R，A018K/P029E/N073R/L075D，D027E/D048Q/G163P/L264R，D027E/D137Q/G163P/L227M，D027E/S058M/G163P/L264R，D027Q/E056K/N233Q/P256T，D027S/E045F/N073R/T189D，D027S/N033D/E045F/N073R，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/N073R，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D048Q/G163P/N233Q/L264R，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，G023K/D027Q/F051T/L075Q，G023K/K024A/V154I/V187N，G023K/L075R/D130A/L264R，G023Q/A049V/V077I/G156W，G023Q/K024A/L075R/G156W，G023Q/L075Q/D130A/L264R，L075G/G091Q/V187N/L264R，N033D/E045F/N073R/T189A，P029E/N073R/L075D/T189D，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，S058M/D137Q/G163P/N233Q，S058M/L075Q/G091Q/I252Q，A018K/D027S/E045F/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/G023Q/K024A/L075Q/G156W，A018K/G023Q/V077I/D130A/G156W，A018K/K024A/L075R/D130A/V154I，A018K/K024A/N094R/D130A/V187N，A018K/P029E/N033D/N073R/L075D，D027E/S058M/D130A/G163P/L264R，D027E/S058M/D137Q/G163P/N233Q，D027E/S058M/G163P/L227M/L264R，D027S/E056K/D111A/V187N/L264R，D027S/L075R/V154I/T189Q/L264R，D048Q/D130A/G163P/L227M/L264R，D048Q/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/G163P/L227M/L264R，D048Q/S058M/G163P/N233Q/L264R，D130A/D137Q/G163P/N233Q/L264R，E045F/A049V/L075Q/V187T/T189D，F051T/L075G/G091Q/D130A/L264R，F051T/L075Q/G091Q/D130A/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187N/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/V154I/G156W/V187Q，N011K/G023K/D111A/G156W/L264R，S058M/D130A/D137Q/G163P/L264R，S058M/D130A/G163P/L227M/L264R，S058M/G163P/L227M/N233Q/L264R，A018K/D027S/N033D/N073R/L075D/T189D，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D048Q/S058M/D137Q/G163P/N233Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/G091Q/D130A/V187H/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/N094R/D130A/V154I/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，S058M/D130A/G163P/L227M/N233Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，D027E/D048Q/D130A/D137Q/G163P/N233Q/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/D048Q/S058M/D130A/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/I264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T
表3-6：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比SEQIDNO：2在pH8水解丁酸对硝基苯酯的性能指数≥最大PI值的50％的TLL变体。
D130A/L264R，A018K/E045F/N073R，A018K/L075D/T189D，D027S/N033D/T189D，L075G/D130A/V187H，L075Q/V187N/L264R，L075R/D130A/L264R，L075R/D130A/V187T，A018K/E045F/L075D/T189D，A018K/L075Q/V077I/N094R，A018K/P029E/N073R/L075D，D027S/N033D/L075D/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，G023K/D027Q/F051T/L075Q，G023K/L075R/D130A/L264R，G023Q/K024A/L075R/G156W，L075G/G091Q/V187N/L264R，P029E/N073R/L075D/T189D，A018K/D027S/E045F/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/G023Q/K024A/L075Q/G156W，A018K/K024A/L075R/D130A/V154I，A018K/K024A/N094R/D130A/V187N，A018K/P029E/N033D/N073R/L075D，D027S/L075R/V154I/T189Q/L264R，D130A/D137Q/G163P/N233Q/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/N094R/G156W/V187N/T189Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/V154I/G156W/V187Q，N011K/G023K/D111A/G156W/L264R，A018K/D027S/N033D/N073R/L075D/T189D，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R
表3-7：下面示出，相比TLL(SEQIDNO：1)，pH8水解辛酸对硝基苯酯(辛酸酯)的性能指数＞1的TLL变体。
A018K/L075Q，D027S/N033D，D130A/L264R，K024A/L075Q，L227M/L264R，P029E/N033D，A018K/G023K/L075Q，A018K/G023Q/L075Q，A018K/L075D/T189D，A018K/L075Q/G156W，A018K/L075Q/V187T，A018K/N073R/L075D，A018K/P029E/T189D，A049V/V187T/T189D，D027S/N033D/T189D，E045F/N073R/L075D，G023K/V187N/L264R，G023Q/L075Q/V187T，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/D130A/V187T，L075Q/G156W/T189D，L075Q/G156W/V187N，L075Q/V187N/L264R，L075R/D130A/L264R，L075R/D130A/V187T，N073R/L075D/T189D，P029E/N033D/E045F，V077I/V187A/T189D，A018K/A049V/L075Q/T189D，A018K/D027S/N033D/L075D，A018K/D027S/P029E/T189D，A018K/E045F/L075D/T189D，A018K/E045F/L075Q/V187T，A018K/G023K/D111A/T189Q，A018K/G023Q/E045F/V187T，A018K/G023Q/G156W/V187T，A018K/G023Q/L075Q/V077I，A018K/G023Q/L075Q/V187T，A018K/G023Q/L075R/D130A，A018K/G023Q/V077I/V187T，A018K/G023Q/V187T/T189D，A018K/L075Q/G156W/V187T，A018K/L075Q/N094R/D111A，A018K/L075Q/V077I/N094R，A018K/P029E/N033D/T189D，D027E/D137Q/G163P/L227M，D027E/L227M/N233Q/L264R，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D130A/G163P/L227M/L264R，G023K/D027Q/F051T/L075Q，G023K/L075Q/G156W/V187N，G023K/L075R/D130A/L264R，G023Q/E045F/A049V/T189D，G023Q/E045F/L075Q/G156W，G023Q/K024A/L075R/V154I，K024A/L075Q/D130A/G156W，K024A/L075Q/V187T/T189Q，K024A/L075R/G156W/V187N，L075Q/D130A/G156W/V187N，L075Q/V077I/D130A/V187Q，L075Q/V077I/V187T/T189D，L075R/D130A/V154I/L264R，N011K/D027S/E056K/P256T，N011K/S058M/N233Q/P256T，P029E/E045F/L075D/T189D，P029E/N033D/L075D/T189D，P029E/N073R/L075D/T189D，Q004D/N011K/D027N/P256T，A018K/D027S/N033D/L075D/T189D，A018K/D027S/P029E/E045F/T189D，A018K/D027S/P029E/N033D/L075D，A018K/D027S/P029E/N073R/L075D，A018K/G023K/K024A/L075R/N094R，A018K/G023Q/A049V/L075Q/V187T，A018K/G023Q/E045F/L075Q/V187T，A018K/G023Q/K024A/D130A/V154I，A018K/G023Q/K024A/L075Q/G156W，A018K/G023Q/L075Q/G156W/V187T，A018K/K024A/L075Q/D130A/L264R，A018K/K024A/L075Q/V154I/G156W，A018K/K024A/N094R/D130A/V187N，A018K/L075Q/D111A/D130A/V187T，A018K/L075Q/N094R/D130A/V187N，A018K/L075Q/N094R/G156W/V187N，A018K/L075Q/V077I/N094R/G156W，A018K/P029E/N033D/N073R/L075D，D027E/D048Q/S058M/G163P/N233Q，D027E/D130A/G163P/N233Q/L264R，D027S/E056K/S058M/N233Q/P256T，D027S/L075R/V154I/T189Q/L264R，D027S/P029E/N033D/E045F/N073R，D130A/D137Q/G163P/N233Q/L264R，E056K/L075Q/G156W/T189Q/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/K024A/L075R/D130A/G156W，G023K/L075Q/D111A/D130A/V187T，G023K/L075Q/V077I/D130A/G156W，G023K/N094R/G156W/V187N/T189Q，G023Q/A049V/L075Q/G156W/V187T，G023Q/A049V/L075Q/V187T/T189D，G023Q/D027S/V154I/V187N/L264R，G023Q/E045F/L075Q/V077I/V187T，G023Q/K024A/L075Q/G156W/V187Q，G023Q/K024A/L075Q/V077I/V187Q，G023Q/K024A/V077I/D130A/V154I，G023Q/L075Q/V077I/D130A/G156W，G023Q/L075Q/V077I/G156W/V187N，K024A/L075R/V154I/G156W/V187Q，L075R/D130A/V154I/T189Q/L264R，L075R/D130A/V187T/T189Q/L264R，N011K/D027N/E056K/N233Q/P256T，N011K/G023K/D111A/G156W/L264R，N011K/G023K/E056K/L075Q/T189Q，A018K/D027S/N033D/N073R/L075D/T189D，A018K/D027S/P029E/N033D/E045F/L075D，A018K/D027S/P029E/N073R/L075D/T189D，A018K/G023K/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075R/D130A/V187N，A018K/G023K/K024A/N094R/D130A/V154I，A018K/G023K/L075Q/D130A/G156W/V187N，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023K/L075R/N094R/D130A/V187N，A018K/G023K/L075R/V077I/D130A/V154I，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/G023Q/L075Q/G156W/V187T/T189D，A018K/G023Q/L075Q/V077I/N094R/V154I，A018K/G023Q/L075Q/V077I/V187T/T189D，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/N094R/V154I/V187N，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/K024A/L075R/D130A/V187N/T189Q，A018K/L075Q/D130A/V154I/T189Q/L264R，A018K/L075Q/D130A/V187N/T189Q/L264R，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027Q/F051T/L075Q/D130A/V187H/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/P029E/N033D/E045F/N073R/T189D，D048Q/S058M/D130A/G163P/N233Q/L264R，E045F/A049V/L075Q/V077I/V187T/T189D，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/K024A/L075Q/N094R/D130A/G156W，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/L075Q/D111A/D130A/V154I/V187N，G023K/L075Q/N094R/V154I/V187T/L264R，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/E045F/A049V/L075Q/V077I/V187T，G023Q/E045F/L075Q/V077I/G156W/V187T，G023Q/K024A/L075Q/D130A/V154I/G156W，G023Q/L075R/V154I/G156W/V187N/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/D130A/G156W/T189Q/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，K024A/L075Q/V154I/V187N/T189Q/L264R，K024A/L075Q/V154I/V187T/T189Q/L264R，L075R/D130A/V154I/G156W/V187N/L264R，N011K/D027N/E056K/S058M/N233Q/P256T，N011K/D027S/E056K/S058M/N233Q/P256T，N011K/E056K/L075Q/D130A/V187N/T189Q，Q004D/D027N/E056K/S058M/N233Q/P256T，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/D130A/G156W/V187N，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075Q/V077I/N094R/G156W，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/K024A/L075R/V154I/G156W/V187Q，A018K/G023K/K024A/N094R/V154I/V187T/L264R，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/E045F/A049V/L075Q/G156W/V187T，A018K/G023Q/K024A/L075Q/V077I/N094R/V154I，A018K/G023Q/K024A/L075R/D130A/V154I/V187N，A018K/G023Q/K024A/L075R/V077I/D130A/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/D111A/G156W/V187N/L264R，A018K/K024A/L075Q/D130A/G156W/V187T/T189Q，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/L075Q/D111A/V154I/V187T/T189Q/L264R，A018K/L075Q/N094R/D111A/V154I/V187T/L264R，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/L075G/D130A/V187H/I252Q/L264R，D027S/F051T/L075G/G091Q/V187T/I252Q/L264R，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，E056K/L075Q/D111A/D130A/G156W/V187N/T189Q，G023K/D027Q/F051T/L075G/D130A/V187H/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023K/K024A/L075Q/D111A/D130A/T189Q/L264R，G023K/K024A/L075Q/D111A/V154I/G156W/V187N，G023K/L075R/N094R/V154I/G156W/V187N/L264R，G023Q/D027S/L075Q/G091Q/V187T/I252Q/L264R，G023Q/E056K/L075Q/D111A/G156W/V187N/L264R，G023Q/E056K/L075Q/D130A/V154I/G156W/T189Q，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/L075Q/D111A/D130A/V154I/G156W/L264R，G023Q/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/L075Q/D130A/V154I/V187N/T189Q/L264R，K024A/L075Q/D111A/D130A/V154I/G156W/L264R，K024A/L075Q/D111A/D130A/V187N/T189Q/L264R，A018K/G023K/K024A/L075Q/D130Y/V154I/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023K/K024A/L075R/D111A/V154I/V187T/L264R，A018K/G023K/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/G023Q/K024A/L075Q/V077I/D130A/G156W/V187Q，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，A018K/K024A/L075Q/V077I/N094R/D130A/V154I/G156W，A018K/K024A/L075R/D130A/V154I/G156W/V187T/L264R，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/E056K/L075G/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/K024A/L075R/N094R/V154I/G156W/V187T/T189Q，G023K/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，G023Q/D027S/E056K/S058M/L075R/G091Q/V187N/I252Q，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/E056K/L075Q/N094R/D111A/G156W/T189Q/L264R，G023Q/K024A/L075Q/V077I/D130A/V154I/G156W/V187N，G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/V187N，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，K024A/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，N011K/G023Q/D027S/L075Q/N094R/V154I/G156W/T189Q，A018K/G023K/K024A/L075Q/N094R/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023K/L075Q/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023K/D027S/F051T/E056K/S058M/L075G/G091Q/D130A/I252Q/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D
表3-8：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且pH8水解辛酸对硝基苯酯(辛酸酯)的性能指数＞1的TLL变体。
D027S/N033D，D130A/L264R，L227M/L264R，P029E/N033D，A018K/L075D/T189D，A018K/P029E/T189D，D027S/N033D/T189D，G023K/V187N/L264R，L075G/D130A/V187H，L075Q/V187N/L264R，L075R/D130A/L264R，L075R/D130A/V187T，P029E/N033D/E045F，A018K/D027S/P029E/T189D，A018K/E045F/L075D/T189D，A018K/G023K/D111A/T189Q，A018K/L075Q/V077I/N094R，D027E/D137Q/G163P/L227M，D027E/L227M/N233Q/L264R，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D130A/G163P/L227M/L264R，G023K/D027Q/F051T/L075Q，G023K/L075R/D130A/L264R，P029E/N073R/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/G023Q/K024A/L075Q/G156W，A018K/K024A/N094R/D130A/V187N，A018K/P029E/N033D/N073R/L075D，D027E/D130A/G163P/N233Q/L264R，D027S/L075R/V154I/T189Q/L264R，D130A/D137Q/G163P/N233Q/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/N094R/G156W/V187N/T189Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/V154I/G156W/V187Q，N011K/G023K/D111A/G156W/L264R，A018K/D027S/N033D/N073R/L075D/T189D，A018K/D027S/P029E/N033D/E045F/L075D，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/V077I/N094R/D130A/V187N，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R
表3-9：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且pH8水解辛酸对硝基苯酯(辛酸酯)的性能指数≥最大PI值的50％的TLL变体。
L227M/L264R，A018K/L075D/T189D，A018K/P029E/T189D，D027S/N033D/T189D，G023K/V187N/L264R，L075R/D130A/V187T，A018K/D027S/P029E/T189D，A018K/L075Q/V077I/N094R，D027E/L227M/N233Q/L264R，D027S/N033D/L075D/T189D，D027S/P029E/E045F/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D130A/G163P/L227M/L264R，G023K/L075R/D130A/L264R，P029E/N073R/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/K024A/N094R/D130A/V187N，A018K/P029E/N033D/N073R/L075D，D027E/D130A/G163P/N233Q/L264R，D027S/L075R/V154I/T189Q/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/N094R/G156W/V187N/T189Q，K024A/L075R/V154I/G156W/V187Q，A018K/D027S/P029E/N033D/E045F/L075D，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，K024A/L075Q/D111A/V154I/V187N/T189Q，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R
表3-10：下面示出，相比参考序列SEQIDNO：2，在pH8水解辛酸对硝基苯酯(辛酸酯)的性能指数＞1的TLL变体。
A018K/L075Q，D130A/L264R，K024A/L075Q，L227M/L264R，A018K/G023K/L075Q，A018K/G023Q/L075Q，A018K/L075D/T189D，A018K/L075Q/G156W，A018K/L075Q/V187T，A018K/N073R/L075D，A018K/P029E/T189D，A049V/V187T/T189D，D027S/N033D/T189D，E045F/N073R/L075D，G023K/V187N/L264R，G023Q/L075Q/V187T，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/D130A/V187T，L075Q/G156W/T189D，L075Q/G156W/V187N，L075R/D130A/L264R，L075R/D130A/V187T，N073R/L075D/T189D，V077I/V187A/T189D，A018K/A049V/L075Q/T189D，A018K/D027S/N033D/L075D，A018K/D027S/P029E/T189D，A018K/E045F/L075Q/V187T，A018K/G023K/D111A/T189Q，A018K/G023Q/L075Q/V077I，A018K/G023Q/L075Q/V187T，A018K/G023Q/L075R/D130A，A018K/G023Q/V187T/T189D，A018K/L075Q/N094R/D111A，A018K/L075Q/V077I/N094R，A018K/P029E/N033D/T189D，D027E/L227M/N233Q/L264R，D027S/N033D/L075D/T189D，D027S/P029E/E045F/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D130A/G163P/L227M/L264R，G023K/D027Q/F051T/L075Q，G023K/L075R/D130A/L264R，G023Q/E045F/L075Q/G156W，G023Q/K024A/L075R/V154I，K024A/L075Q/D130A/G156W，K024A/L075Q/V187T/T189Q，K024A/L075R/G156W/V187N，L075Q/D130A/G156W/V187N，L075Q/V077I/D130A/V187Q，L075Q/V077I/V187T/T189D，L075R/D130A/V154I/L264R，P029E/E045F/L075D/T189D，P029E/N033D/L075D/T189D，P029E/N073R/L075D/T189D，Q004D/N011K/D027N/P256T，A018K/D027S/N033D/L075D/T189D，A018K/D027S/P029E/E045F/T189D，A018K/D027S/P029E/N033D/L075D，A018K/D027S/P029E/N073R/L075D，A018K/G023K/K024A/L075R/N094R，A018K/G023Q/E045F/L075Q/V187T，A018K/G023Q/K024A/L075Q/G156W，A018K/G023Q/L075Q/G156W/V187T，A018K/K024A/L075Q/V154I/G156W，A018K/K024A/N094R/D130A/V187N，A018K/L075Q/D111A/D130A/V187T，A018K/L075Q/N094R/D130A/V187N，A018K/L075Q/N094R/G156W/V187N，A018K/L075Q/V077I/N094R/G156W，A018K/P029E/N033D/N073R/L075D，D027E/D048Q/S058M/G163P/N233Q，D027E/D130A/G163P/N233Q/L264R，D027S/L075R/V154I/T189Q/L264R，D027S/P029E/N033D/E045F/N073R，D130A/D137Q/G163P/N233Q/L264R，E056K/L075Q/G156W/T189Q/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/K024A/L075R/D130A/G156W，G023K/L075Q/D111A/D130A/V187T，G023K/L075Q/V077I/D130A/G156W，G023K/N094R/G156W/V187N/T189Q，G023Q/A049V/L075Q/G156W/V187T，G023Q/A049V/L075Q/V187T/T189D，G023Q/E045F/L075Q/V077I/V187T，G023Q/K024A/L075Q/G156W/V187Q，G023Q/K024A/L075Q/V077I/V187Q，G023Q/K024A/V077I/D130A/V154I，G023Q/L075Q/V077I/G156W/V187N，K024A/L075R/V154I/G156W/V187Q，L075R/D130A/V187T/T189Q/L264R，N011K/D027N/E056K/N233Q/P256T，N011K/G023K/E056K/L075Q/T189Q，A018K/D027S/N033D/N073R/L075D/T189D，A018K/D027S/P029E/N033D/E045F/L075D，A018K/D027S/P029E/N073R/L075D/T189D，A018K/G023K/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075R/D130A/V187N，A018K/G023K/K024A/N094R/D130A/V154I，A018K/G023K/L075Q/D130A/G156W/V187N，A018K/G023K/L075R/N094R/D130A/V187N，A018K/G023K/L075R/V077I/D130A/V154I，A018K/G023Q/L075Q/V077I/N094R/V154I，A018K/G023Q/L075Q/V077I/V187T/T189D，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/N094R/V154I/V187N，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/K024A/L075R/D130A/V187N/T189Q，A018K/L075Q/D130A/V154I/T189Q/L264R，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027Q/F051T/L075Q/D130A/V187H/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D048Q/S058M/D130A/G163P/N233Q/L264R，E045F/A049V/L075Q/V077I/V187T/T189D，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/K024A/L075Q/N094R/D130A/G156W，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/L075Q/D111A/D130A/V154I/V187N，G023K/L075Q/N094R/V154I/V187T/L264R，G023Q/K024A/L075Q/D130A/V154I/G156W，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/D130A/G156W/T189Q/L264R，K024A/L075Q/V154I/V187N/T189Q/L264R，K024A/L075Q/V154I/V187T/T189Q/L264R，L075R/D130A/V154I/G156W/V187N/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/D130A/G156W/V187N，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/N094R/G156W，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/K024A/L075R/V154I/G156W/V187Q，A018K/G023K/K024A/N094R/V154I/V187T/L264R，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/K024A/L075Q/V077I/N094R/V154I，A018K/G023Q/K024A/L075R/D130A/V154I/V187N，A018K/G023Q/K024A/L075R/V077I/D130A/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/D111A/G156W/V187N/L264R，A018K/K024A/L075Q/D130A/G156W/V187T/T189Q，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/L075Q/D111A/V154I/V187T/T189Q/L264R，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，E056K/L075Q/D111A/D130A/G156W/V187N/T189Q，G023K/D027Q/F051T/L075G/D130A/V187H/L264R，G023K/K024A/L075Q/D111A/V154I/G156W/V187N，G023K/L075R/N094R/V154I/G156W/V187N/L264R，G023Q/D027S/L075Q/G091Q/V187T/I252Q/L264R，G023Q/E056K/L075Q/D111A/G156W/V187N/L264R，G023Q/E056K/L075Q/D130A/V154I/G156W/T189Q，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/L075Q/D130A/V154I/V187N/T189Q/L264R，K024A/L075Q/D111A/D130A/V154I/G156W/L264R，K024A/L075Q/D111A/D130A/V187N/T189Q/L264R，A018K/G023K/K024A/L075Q/D130Y/V154I/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023K/K024A/L075R/D111A/V154I/V187T/L264R，A018K/G023K/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/G023Q/K024A/L075Q/V077I/D130A/G156W/V187Q，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，A018K/K024A/L075Q/V077I/N094R/D130A/V154I/G156W，A018K/K024A/L075R/D130A/V154I/G156W/V187T/L264R，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/E056K/L075G/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/K024A/L075R/N094R/V154I/G156W/V187T/T189Q，G023K/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，G023Q/D027S/E056K/S058M/L075R/G091Q/V187N/I252Q，G023Q/E056K/L075Q/N094R/D111A/G156W/T189Q/L264R，G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/V187N，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，K024A/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，N011K/G023Q/D027S/L075Q/N094R/V154I/G156W/T189Q，A018K/G023K/K024A/L075Q/N094R/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023K/D027S/F051T/E056K/S058M/L075G/G091Q/D130A/I252Q/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D
表3-11：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比SEQIDNO：2在pH8水解辛酸对硝基苯酯(辛酸酯)的性能指数＞1的TLL变体。
D130A/L264R，L227M/L264R，A018K/L075D/T189D，A018K/P029E/T189D，D027S/N033D/T189D，G023K/V187N/L264R，L075G/D130A/V187H，L075R/D130A/L264R，L075R/D130A/V187T，A018K/D027S/P029E/T189D，A018K/G023K/D111A/T189Q，A018K/L075Q/V077I/N094R，D027E/L227M/N233Q/L264R，D027S/N033D/L075D/T189D，D027S/P029E/E045F/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D130A/G163P/L227M/L264R，G023K/D027Q/F051T/L075Q，G023K/L075R/D130A/L264R，P029E/N073R/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/G023Q/K024A/L075Q/G156W，A018K/K024A/N094R/D130A/V187N，A018K/P029E/N033D/N073R/L075D，D027E/D130A/G163P/N233Q/L264R，D027S/L075R/V154I/T189Q/L264R，D130A/D137Q/G163P/N233Q/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/N094R/G156W/V187N/T189Q，G023Q/K024A/L075Q/G156W/V187Q，K024A/L075R/V154I/G156W/V187Q，A018K/D027S/N033D/N073R/L075D/T189D，A018K/D027S/P029E/N033D/E045F/L075D，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，K024A/L075Q/D111A/V154I/V187N/T189Q，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R
表3-12：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比SEQIDNO：2在pH8水解辛酸对硝基苯酯(辛酸酯)的性能指数≥最大PI值的50％的TLL变体
L227M/L264R，A018K/L075D/T189D，A018K/P029E/T189D，D027S/N033D/T189D，G023K/V187N/L264R，L075R/D130A/V187T，A018K/D027S/P029E/T189D，A018K/L075Q/V077I/N094R，D027E/L227M/N233Q/L264R，D027S/N033D/L075D/T189D，D027S/P029E/E045F/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D130A/G163P/L227M/L264R，G023K/L075R/D130A/L264R，P029E/N073R/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/K024A/N094R/D130A/V187N，A018K/P029E/N033D/N073R/L075D，D027E/D130A/G163P/N233Q/L264R，D027S/L075R/V154I/T189Q/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/N094R/G156W/V187N/T189Q，K024A/L075R/V154I/G156W/V187Q，A018K/D027S/P029E/N033D/E045F/L075D，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，K024A/L075Q/D111A/V154I/V187N/T189Q，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R
表3-13：下面示出，相比TLL(SEQIDNO：1)，在pH8水解棕榈酸对硝基苯酯的性能指数＞1的TLL变体。
A018K/E045F，D027S/N033D，D130A/L264R，D130A/V187T，G023K/L264R，K024A/L075Q，L075R/L264R，S058M/L264R，V187T/L264R，A018K/A049V/L075Q，A018K/D027S/E045F，A018K/E045F/T189D，A018K/L075D/T189D，A018K/L075Q/V187T，A018K/N033D/T189D，A018K/N073R/L075D，A018K/P029E/T189D，A049V/V187T/T189D，D027N/N233Q/P256T，D027Q/N233Q/P256T，D027S/N033D/T189D，D111A/D130A/L264R，D130A/V187N/L264R，F051T/I252Q/L264R，G023K/D130A/L264R，G023K/E056K/V187T，G023K/L075R/L264R，G023K/V187N/L264R，G023K/V187T/L264R，G023Q/L075Q/V187T，G163P/L227M/L264R，K024A/L075Q/V077I，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/G156W/V187N，L075Q/V187N/L264R，L075Q/V187T/L264R，L075R/D130A/L264R，L075R/D130A/V187T，P029E/N073R/L075D，Q004D/D027S/P256T，V187N/T189Q/L264R，A018K/D027S/N033D/L075D，A018K/E045F/L075D/T189D，A018K/G023K/D111A/T189Q，A018K/G023Q/L075R/D130A，A018K/G023Q/V077I/V187T，A018K/L075Q/N094R/V187Q，A018K/L075Q/V077I/N094R，A018K/N073R/L075D/T189D，A018K/P029E/N073R/L075D，D027E/D048Q/G163P/L264R，D027E/D130A/N233Q/L264R，D027E/D137Q/G163P/L227M，D027E/G163P/L227M/L264R，D027Q/S058M/I090F/P256T，D027S/D130A/I252Q/L264R，D027S/E045F/N073R/L075D，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D048Q/G163P/N233Q/L264R，E056K/D130A/V187N/L264R，F051T/D130A/I252Q/L264R，F051T/L075G/I252Q/L264R，F051T/L075Q/I252Q/L264R，G023K/D027Q/F051T/L075Q，G023K/D130A/V187N/L264R，G023K/D130A/V187T/L264R，G023K/E056K/L075R/L264R，G023K/L075Q/D130A/V187N，G023K/L075Q/G156W/V187N，G023K/L075R/D130A/L264R，G023K/L075R/V187T/L264R，G023Q/E045F/A049V/T189D，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/K024A/L075R/V154I，G023Q/L075Q/D130A/L264R，G023Q/L075Q/V187T/L264R，G091Q/V187T/I252Q/L264R，K024A/D130A/V154I/V187T，K024A/L075Q/D130A/G156W，K024A/L075R/G156W/V187N，L075G/D130A/V187T/I252Q，L075G/V187H/I252Q/L264R，L075Q/D130A/G156W/V187N，L075Q/V077I/D130A/V187Q，N011K/S058M/N233Q/P256T，P029E/E045F/N073R/T189D，P029E/N033D/L075D/T189D，P029E/N073R/L075D/T189D，Q004D/D027N/E056K/P256T，Q004D/N011K/D027S/P256T，S058M/G163P/L227M/L264R，S058M/G163P/N233Q/L264R，S058M/L075Q/I252Q/L264R，A018K/D027S/N033D/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/D130A/G156W/V187N/L264R，A018K/G023K/K024A/L075R/N094R，A018K/G023Q/E045F/A049V/V187T，A018K/G023Q/E045F/L075Q/V187T，A018K/G023Q/E045F/V077I/T189D，A018K/G023Q/K024A/L075Q/G156W，A018K/G023Q/L075Q/G156W/V187T，A018K/K024A/L075Q/D130A/L264R，A018K/K024A/L075Q/V154I/G156W，A018K/K024A/L075R/N094R/D130A，A018K/L075Q/D111A/D130A/V187T，A018K/L075Q/N094R/D130A/V187N，A018K/L075Q/N094R/G156W/V187N，A018K/L075Q/V077I/N094R/G156W，A018K/P029E/N033D/N073R/L075D，D027E/D048Q/D137Q/L227M/L264R，D027E/D048Q/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L264R，D027E/D130A/G163P/L227M/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L264R，D027E/S058M/G163P/L227M/L264R，D027S/E056K/S058M/N233Q/P256T，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D027S/L075R/V154I/T189Q/L264R，D027S/N033D/E045F/N073R/L075D，D027S/N033D/N073R/L075D/T189D，D027S/P029E/N073R/L075D/T189D，D048Q/D130A/G163P/L227M/L264R，D048Q/D137Q/G163P/L227M/L264R，D048Q/D137Q/G163P/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/D137Q/N233Q/L264R，D048Q/S058M/G163P/N233Q/L264R，D130A/D137Q/G163P/L227M/L264R，D130A/D137Q/G163P/N233Q/L264R，E056K/L075G/V187N/I252Q/L264R，E056K/L075Q/G156W/T189Q/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/L264R，F051T/L075Q/V187N/I252Q/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187N/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/D111A/D130A/V187T，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187N/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/A049V/L075Q/G156W/V187T，G023Q/E056K/V187N/I252Q/L264R，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/K024A/L075Q/G156W/V187N，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075G/G091Q/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，G023Q/L075Q/V077I/G156W/V187N，K024A/L075R/V154I/G156W/V187Q，L075G/D130A/V187T/I252Q/L264R，L075Q/V154I/V187N/T189Q/L264R，L075R/D130A/V187T/T189Q/L264R，N011K/G023K/D111A/G156W/L264R，N011K/G023K/E056K/L075Q/T189Q，S058M/D130A/D137Q/G163P/L264R，S058M/D130A/G163P/L227M/L264R，S058M/G163P/L227M/N233Q/L264R，A018K/D027S/N033D/N073R/L075D/T189D，A018K/G023K/L075Q/D130A/G156W/V187N，A018K/G023Q/L075Q/G156W/V187T/T189D，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/D111A/V187N/T189Q/L264R，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075R/D130A/V187N/T189Q，A018K/K024A/V077I/N094R/D130A/G156W，A018K/L075Q/D130A/V187N/T189Q/L264R，D027E/D048Q/D130A/D137H/G163P/L264R，D027E/D048Q/D130A/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L227M/L264R，D027E/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027Q/F051T/L075Q/D130A/V187H/L264R，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/S058M/D130A/G163P/L227M/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/L264R，D048Q/S058M/D137Q/G163P/N233Q/L264R，D048Q/S058M/G163P/L227M/N233Q/L264R，F051T/E056K/D130A/V187N/I252Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023K/K024A/L075Q/N094R/D130A/G156W，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/L075Q/D111A/D130A/V154I/V187N，G023K/L075Q/N094R/V154I/V187T/L264R，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/L075Q/D130A/V187T/I252Q，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/F051T/G091Q/V187N/I252Q/L264R，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/L075Q/D130A/V154I/G156W，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/L075G/G091Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/V187N/I252Q/L264R，G023Q/L075R/V154I/G156W/V187N/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/V154I/V187T/T189Q/L264R，L075Q/D130A/V154I/G156W/V187N/L264R，L075Q/G091Q/D130A/V187H/I252Q/L264R，L075R/D130A/V154I/G156W/V187N/L264R，N011K/D027Q/E056K/S058M/N233Q/P256T，N011K/D027S/E056K/S058M/N233Q/P256T，N011K/E056K/L075Q/D130A/V187N/T189Q，Q004D/D027N/E056K/S058M/N233Q/P256T，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，S058M/D130A/G163P/L227M/N233Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/D130A/G156W/V187N，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/K024A/L075R/V154I/G156W/V187Q，A018K/G023K/K024A/N094R/V154I/V187T/L264R，A018K/G023Q/K024A/L075Q/V077I/N094R/V154I，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/D130A/V154I/G156W/V187T/L264R，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，D027E/D048Q/D130A/D137Q/G163P/N233Q/L264R，D027E/D048Q/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/D048Q/S058M/D130A/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/L075G/D130A/V187H/I252Q/L264R，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/L075G/G091Q/V187T/I252Q/L264R，D027S/F051T/S058M/L075Q/V187N/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，E056K/L075Q/D111A/D130A/G156W/V187N/T189Q，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/D027Q/F051T/L075G/D130A/V187H/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023K/K024A/L075Q/D111A/D130A/T189Q/L264R，G023K/L075R/N094R/V154I/G156W/V187N/L264R，G023Q/D027S/F051T/E056K/D130A/I252Q/L264R，G023Q/D027S/L075Q/G091Q/V187T/I252Q/L264R，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/E056K/L075Q/D111A/G156W/V187N/L264R，G023Q/E056K/L075Q/D130A/V154I/G156W/T189Q，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/K024A/L075Q/V077I/D130A/G156W/V187N，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/L075Q/D111A/D130A/V154I/G156W/L264R，G023Q/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/L075Q/D130A/V154I/V187N/T189Q/L264R，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，K024A/L075Q/D111A/D130A/V154I/G156W/L264R，Q004D/N011K/D027N/S058M/I090F/N233Q/P256T，A018K/G023K/K024A/L075Q/D130Y/V154I/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023Q/K024A/L075Q/V077I/D130A/G156W/V187Q，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，D027E/D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/E056K/L075G/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/D027S/F051T/S058M/L075Q/V187H/I252Q/L264R，G023K/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/E056K/L075Q/N094R/D111A/G156W/T189Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/V187N，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，K024A/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，N011K/G023Q/D027S/L075Q/N094R/V154I/G156W/T189Q，N011K/G023Q/L075Q/D111A/D130A/V154I/V187N/L264R，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W/V187N，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023K/D027S/F051T/E056K/S058M/L075G/G091Q/D130A/I252Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T
表3-14：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且在pH8水解棕榈酸对硝基苯酯的性能指数＞1的TLL变体。
A018K/E045F，D027S/N033D，D130A/L264R，D130A/V187T，G023K/L264R，L075R/L264R，S058M/L264R，V187T/L264R，A018K/L075D/T189D，A018K/N033D/T189D，A018K/P029E/T189D，D027S/N033D/T189D，D111A/D130A/L264R，D130A/V187N/L264R，F051T/I252Q/L264R，G023K/D130A/L264R，G023K/E056K/V187T，G023K/L075R/L264R，G023K/V187N/L264R，G023K/V187T/L264R，G163P/L227M/L264R，L075G/D130A/V187H，L075Q/V187N/L264R，L075Q/V187T/L264R，L075R/D130A/L264R，L075R/D130A/V187T，Q004D/D027S/P256T，V187N/T189Q/L264R，A018K/E045F/L075D/T189D，A018K/G023K/D111A/T189Q，A018K/L075Q/V077I/N094R，A018K/P029E/N073R/L075D，D027E/D048Q/G163P/L264R，D027E/D130A/N233Q/L264R，D027E/D137Q/G163P/L227M，D027E/G163P/L227M/L264R，D027S/D130A/I252Q/L264R，D027S/N033D/E045E/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D048Q/G163P/N233Q/L264R，E056K/D130A/V187N/L264R，F051T/D130A/I252Q/L264R，F051T/L075G/I252Q/L264R，F051T/L075Q/I252Q/L264R，G023K/D027Q/F051T/L075Q，G023K/D130A/V187N/L264R，G023K/D130A/V187T/L264R，G023K/E056K/L075R/L264R，G023K/L075R/D130A/L264R，G023K/L075R/V187T/L264R，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/L075Q/D130A/L264R，G023Q/L075Q/V187T/L264R，G091Q/V187T/I252Q/L264R，K024A/D130A/V154I/V187T，L075G/D130A/V187T/I252Q，L075G/V187H/I252Q/L264R，P029E/N073R/L075D/T189D，Q004D/D027N/E056K/P256T，Q004D/N011K/D027S/P256T，S058M/G163P/L227M/L264R，S058M/G163P/N233Q/L264R，S058M/L075Q/I252Q/L264R，A018K/D027S/P029E/N033D/L075D，A018K/D130A/G156W/V187N/L264R，A018K/G023Q/K024A/L075Q/G156W，A018K/P029E/N033D/N073R/L075D，D027E/D048Q/D137Q/L227M/L264R，D027E/D048Q/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L264R，D027E/D130A/G163P/L227M/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L264R，D027E/S058M/G163P/L227M/L264R，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D027S/L075R/V154I/T189Q/L264R，D048Q/D130A/G163P/L227M/L264R，D048Q/D137Q/G163P/L227M/L264R，D048Q/D137Q/G163P/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/D137Q/N233Q/L264R，D048Q/S058M/G163P/N233Q/L264R，D130A/D137Q/G163P/L227M/L264R，D130A/D137Q/G163P/N233Q/L264R，E056K/L075G/V187N/I252Q/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/L264R，F051T/L075Q/V187N/I252Q/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187N/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187N/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/E056K/V187N/I252Q/L264R，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075G/G091Q/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，K024A/L075R/V154I/G156W/V187Q，L075G/D130A/V187T/I252Q/L264R，N011K/G023K/D111A/G156W/L264R，S058M/D130A/D137Q/G163P/L264R，S058M/D130A/G163P/L227M/L264R，S058M/G163P/L227M/N233Q/L264R，A018K/D027S/N033D/N073R/L075D/T189D，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/D111A/V187N/T189Q/L264R，A018K/K024A/L075Q/N094R/V154I/G156W，D027E/D048Q/D130A/D137H/G163P/L264R，D027E/D048Q/D130A/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L227M/L264R，D027E/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/S058M/D130A/G163P/L227M/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/L264R，D048Q/S058M/D137Q/G163P/N233Q/L264R，D048Q/S058M/G163P/L227M/N233Q/L264R，F051T/E056K/D130A/V187N/I252Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/L075Q/D130A/V187T/I252Q，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/F051T/G091Q/V187N/I252Q/L264R，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/L075G/G091Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/V187N/I252Q/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，L075Q/G091Q/D130A/V187H/I252Q/L264R，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，S058M/D130A/G163P/L227M/N233Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/D130A/V154I/G156W/V187T/L264R，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，D027E/D048Q/D130A/D137Q/G163P/N233Q/L264R，D027E/D048Q/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/D048Q/S058M/D130A/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/S058M/L075Q/V187N/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023Q/D027S/F051T/E056K/D130A/I252Q/L264R，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，D027E/D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/D027S/F051T/S058M/L075Q/V187H/I252Q/L264R，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T
表3-15：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且在pH8水解棕榈酸对硝基苯酯的性能指数≥最大PI值的50％的TLL变体
D130A/L264R，D130A/V187T，A018K/L075D/T189D，A018K/P029E/T189D，D027S/N033D/T189D，G023K/V187N/L264R，L075G/D130A/V187H，L075Q/V187N/L264R，L075R/D130A/L264R，L075R/D130A/V187T，Q004D/D027S/P256T，A018K/L075Q/V077I/N094R，D027S/D130A/I252Q/L264R，D027S/P029E/E045F/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，F051T/L075Q/I252Q/L264R，G023K/L075R/D130A/L264R，G023K/L075R/V187T/L264R，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G091Q/V187T/I252Q/L264R，L075G/V187H/I252Q/L264R，P029E/N073R/L075D/T189D，Q004D/N011K/D027S/P256T，S058M/G163P/L227M/L264R，S058M/L075Q/I252Q/L264R，A018K/D027S/P029E/N033D/L075D，A018K/G023Q/K024A/L075Q/G156W，A018K/P029E/N033D/N073R/L075D，D027E/D048Q/G163P/N233Q/L264R，D027E/D130A/G163P/L227M/L264R，D027E/G163P/L227M/N233Q/L264R，D027S/F051T/V187N/I252Q/L264R，D027S/L075R/V154I/T189Q/L264R，D048Q/D137Q/G163P/N233Q/L264R，D130A/D137Q/G163P/N233Q/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075Q/V187N/I252Q/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187N/L264R，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075Q/G091Q/I252Q/L264R，K024A/L075R/V1541/G156W/V187Q，S058M/G163P/L227M/N233Q/L264R，A018K/D027S/N033D/N073R/L075D/T189D，A018K/K024A/L075Q/N094R/V154I/G156W，D027E/D048Q/D130A/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，F051T/E056K/D130A/V187N/I252Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023Q/L075Q/G091Q/V187N/I252Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027S/F051T/S058M/L075Q/V187N/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/D027S/F051T/E056K/D130A/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R
表3-16：下面示出，相比参考序列SEQIDNO：2，在pH8水解棕榈酸对硝基苯酯的性能指数＞1的TLL变体。
A018K/E045F，D027S/N033D，D111A/L264R，D130A/L264R，D130A/V187T，G023K/L264R，K024A/L075Q，L075R/L264R，S058M/L264R，V187T/L264R，A018K/A049V/L075Q，A018K/D027S/E045F，A018K/E045F/T189D，A018K/L075D/T189D，A018K/L075Q/V187T，A018K/N033D/T189D，A018K/N073R/L075D，A018K/P029E/T189D，A049V/V187T/T189D，D027N/N233Q/P256T，D027Q/N233Q/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D111A/D130A/L264R，D130A/V187N/L264R，E045F/L075D/T189D，F051T/I252Q/L264R，G023K/D130A/L264R，G023K/E056K/V187T，G023K/L075R/L264R，G023K/V187N/L264R，G023K/V187T/L264R，G023Q/E056K/L075R，G023Q/L075Q/V187T，G163P/L227M/L264R，K024A/L075Q/V077I，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/G156W/V187N，L075Q/V187N/L264R，L075Q/V187T/L264R，L075R/D130A/L264R，L075R/D130A/V187T，L075R/V187T/L264R，N073R/L075D/T189D，P029E/N073R/L075D，Q004D/D027S/P256T，V187N/T189Q/L264R，A018K/D027S/N033D/L075D，A018K/E045F/L075D/T189D，A018K/G023K/D111A/T189Q，A018K/G023Q/L075R/D130A，A018K/G023Q/V077I/V187T，A018K/G023Q/V187T/T189D，A018K/L075Q/N094R/V187Q，A018K/L075Q/V077I/N094R，A018K/N073R/L075D/T189D，A018K/P029E/N073R/L075D，D027E/D048Q/G163P/L264R，D027E/D130A/N233Q/L264R，D027E/D137Q/G163P/L227M，D027E/G163P/L227M/L264R，D027E/S058M/G163P/L264R，D027Q/S058M/I090F/P256T，D027S/D130A/I252Q/L264R，D027S/E045F/N073R/L075D，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D048Q/G163P/N233Q/L264R，D111A/D130A/V154I/L264R，E056K/D130A/V187N/L264R，F051T/D130A/I252Q/L264R，F051T/L075G/I252Q/L264R，F051T/L075Q/I252Q/L264R，G023K/D027Q/F051T/L075Q，G023K/D130A/V187N/L264R，G023K/D130A/V187T/L264R，G023K/E056K/L075R/L264R，G023K/L075Q/D130A/V187N，G023K/L075Q/G156W/V187N，G023K/L075R/D130A/L264R，G023K/L075R/V187T/L264R，G023Q/E045F/A049V/T189D，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/K024A/L075R/V154I，G023Q/L075Q/D130A/L264R，G023Q/L075Q/V187T/L264R，G091Q/V187T/I252Q/L264R，K024A/D130A/V154I/V187T，K024A/L075Q/D130A/G156W，K024A/L075R/G156W/V187N，L075G/D130A/V187T/I252Q，L075G/V187H/I252Q/L264R，L075Q/D130A/G156W/V187N，L075Q/V077I/D130A/V187Q，N011K/S058M/N233Q/P256T，P029E/E045F/N073R/T189D，P029E/N033D/L075D/T189D，P029E/N073R/L075D/T189D，Q004D/D027N/E056K/P256T，Q004D/N011K/D027S/P256T，S058M/G163P/L227M/L264R，S058M/G163P/N233Q/L264R，S058M/L075Q/G091Q/I252Q，S058M/L075Q/I252Q/L264R，A018K/D027S/N033D/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/D130A/G156W/V187N/L264R，A018K/G023K/K024A/L075R/N094R，A018K/G023Q/A049V/L075Q/V077I，A018K/G023Q/E045F/A049V/V187T，A018K/G023Q/E045F/L075Q/V187T，A018K/G023Q/E045F/V077I/T189D，A018K/G023Q/K024A/L075Q/G156W，A018K/G023Q/L075Q/G156W/V187T，A018K/G023Q/L075Q/V187T/T189D，A018K/K024A/L075Q/D130A/L264R，A018K/K024A/L075Q/V154I/G156W，A018K/K024A/L075R/N094R/D130A，A018K/L075Q/D111A/D130A/V187T，A018K/L075Q/N094R/D130A/V187N，A018K/L075Q/N094R/G156W/V187N，A018K/L075Q/V077I/N094R/G156W，A018K/P029E/N033D/N073R/L075D，D027E/D048Q/D137Q/L227M/L264R，D027E/D048Q/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L264R，D027E/D130A/G163P/L227M/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L264R，D027E/S058M/D137Q/G163P/N233Q，D027E/S058M/G163P/L227M/L264R，D027S/E056K/S058M/N233Q/P256T，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D027S/L075R/V154I/T189Q/L264R，D027S/N033D/E045F/N073R/L075D，D027S/N033D/N073R/L075D/T189D，D027S/P029E/N073R/L075D/T189D，D048Q/D130A/G163P/L227M/L264R，D048Q/D137Q/G163P/L227M/L264R，D048Q/D137Q/G163P/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/D137Q/N233Q/L264R，D048Q/S058M/G163P/N233Q/L264R，D130A/D137Q/G163P/L227M/L264R，D130A/D137Q/G163P/N233Q/L264R，E056K/L075G/V187N/I252Q/L264R，E056K/L075Q/G156W/T189Q/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/G091Q/D130A/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/L264R，F051T/L075Q/V187N/I252Q/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187N/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/D111A/D130A/V187T，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187N/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/A049V/L075Q/G156W/V187T，G023Q/E056K/V187N/I252Q/L264R，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/K024A/L075Q/G156W/V187N，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075G/G091Q/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，G023Q/L075Q/V077I/G156W/V187N，K024A/L075R/V154I/G156W/V187Q，L075G/D130A/V187T/I252Q/L264R，L075Q/V154I/V187N/T189Q/L264R，L075R/D130A/V154I/T189Q/L264R，L075R/D130A/V187T/T189Q/L264R，N011K/G023K/D111A/G156W/L264R，N011K/G023K/E056K/L075Q/T189Q，S058M/D130A/D137Q/G163P/L264R，S058M/D130A/G163P/L227M/L264R，S058M/G163P/L227M/N233Q/L264R，A018K/D027S/N033D/N073R/L075D/T189D，A018K/G023K/K024A/N094R/D130A/V154I，A018K/G023K/L075Q/D130A/G156W/V187N，A018K/G023Q/L075Q/G156W/V187T/T189D，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/D111A/V187N/T189Q/L264R，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075R/D130A/V187N/T189Q，A018K/K024A/V077I/N094R/D130A/G156W，A018K/L075Q/D130A/V187N/T189Q/L264R，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D048Q/D130A/D137H/G163P/L264R，D027E/D048Q/D130A/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L227M/L264R，D027E/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027Q/F051T/L075Q/D130A/V187H/L264R，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/S058M/D130A/G163P/L227M/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/L264R，D048Q/S058M/D137Q/G163P/N233Q/L264R，D048Q/S058M/G163P/L227M/N233Q/L264R，F051T/E056K/D130A/V187N/I252Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/E056K/L075R/D130A/V187T/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023K/K024A/L075Q/N094R/D130A/G156W，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/L075Q/D111A/D130A/V154I/V187N，G023K/L075Q/N094R/V154I/V187T/L264R，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/L075Q/D130A/V187T/I252Q，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/F051T/G091Q/V187N/I252Q/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/L075Q/D130A/V154I/G156W，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/L075G/G091Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/V187N/I252Q/L264R，G023Q/L075R/V154I/G156W/V187N/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/V154I/V187T/T189Q/L264R，K024A/L075R/D111A/V154I/G156W/V187N，L075Q/D130A/V154I/G156W/V187N/L264R，L075Q/G091Q/D130A/V187H/I252Q/L264R，L075R/D130A/V154I/G156W/V187N/L264R，N011K/D027Q/E056K/S058M/N233Q/P256T，N011K/D027S/E056K/S058M/N233Q/P256T，N011K/E056K/L075Q/D130A/V187N/T189Q，Q004D/D027N/E056K/S058M/N233Q/P256T，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，S058M/D130A/G163P/L227M/N233Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/D130A/G156W/V187N，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/K024A/L075R/V154I/G156W/V187Q，A018K/G023K/K024A/N094R/V154I/V187T/L264R，A018K/G023Q/K024A/L075Q/V077I/N094R/V154I，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/D130A/V154I/G156W/V187T/L264R，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，A018K/L075Q/D130A/G156W/V187N/T189Q/L264R，D027E/D048Q/D130A/D137Q/G163P/N233Q/L264R，D027E/D048Q/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/D048Q/S058M/D130A/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/L075G/D130A/V187H/I252Q/L264R，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/L075G/G091Q/V187T/I252Q/L264R，D027S/F051T/S058M/L075Q/V187N/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，E056K/L075Q/D111A/D130A/G156W/V187N/T189Q，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/D027Q/F051T/L075G/D130A/V187H/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023K/K024A/L075Q/D111A/D130A/T189Q/L264R，G023K/L075R/N094R/V154I/G156W/V187N/L264R，G023Q/D027S/F051T/E056K/D130A/I252Q/L264R，G023Q/D027S/L075Q/G091Q/V187T/I252Q/L264R，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/E056K/L075Q/D111A/G156W/V187N/L264R，G023Q/E056K/L075Q/D130A/V154I/G156W/T189Q，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/K024A/L075Q/V077I/D130A/G156W/V187N，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/L075Q/D111A/D130A/V154I/G156W/L264R，G023Q/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/L075Q/D130A/V154I/V187N/T189Q/L264R，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，K024A/L075Q/D111A/D130A/V154I/G156W/L264R，Q004D/N011K/D027N/S058M/I090F/N233Q/P256T，A018K/G023K/K024A/L075Q/D130Y/V154I/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023Q/K024A/L075Q/V077I/D130A/G156W/V187Q，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，D027E/D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/E056K/L075G/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/D027S/F051T/S058M/L075Q/V187H/I252Q/L264R，G023K/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/E056K/L075Q/N094R/D111A/G156W/T189Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/V187N，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，K024A/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，N011K/G023Q/D027S/L075Q/N094R/V154I/G156W/T189Q，N011K/G023Q/L075Q/D111A/D130A/V154I/V187N/L264R，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W/V187N，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023K/D027S/F051T/E056K/S058M/L075G/G091Q/D130A/I252Q/L264R，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T
表3-17：下面示出，相比TLL(SEQIDN0：1)表达性能指数≥0.5，且相比SEQIDNO：2在pH8水解棕榈酸对硝基苯酯的性能指数＞1的TLL变体。
A018K/E045F，D027S/N033D，D111A/L264R，D130A/L264R，D130A/V187T，G023K/L264R，1075R/L264R，S058M/L264R，V187T/L264R，A018K/L075D/T189D，A018K/N033D/T189D，A018K/P029E/T189D，D027S/L075Q/G091Q，D027S/N033D/T189D，D111A/D130A/L264R，D130A/V187N/L264R，F051T/I252Q/L264R，G023K/D130A/L264R，G023K/E056K/V187T，G023K/L075R/L264R，G023K/V187N/L264R，G023K/V187T/L264R，G163P/L227M/L264R，L075G/D130A/V187H，L075Q/V187N/L264R，L075Q/V187T/L264R，L075R/D130A/L264R，L075R/D130A/V187T，L075R/V187T/L264R，Q004D/D027S/P256T，V187N/T189Q/L264R，A018K/E045F/L075D/T189D，A018K/G023K/D111A/T189Q，A018K/L075Q/V077I/N094R，A018K/P029E/N073R/L075D，D027E/D048Q/G163P/L264R，D027E/D130A/N233Q/L264R，D027E/D137Q/G163P/L227M，D027E/G163P/L227M/L264R，D027E/S058M/G163P/L264R，D027S/D130A/I252Q/L264R，D027S/N033D/E045E/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D048Q/G163P/N233Q/L264R，E056K/D130A/V187N/L264R，F051T/D130A/I252Q/L264R，F051T/L075G/I252Q/L264R，F051T/L075Q/I252Q/L264R，G023K/D027Q/F051T/L075Q，G023K/D130A/V187N/L264R，G023K/D130A/V187T/L264R，G023K/E056K/L075R/L264R，G023K/L075R/D130A/L264R，G023K/L075R/V187T/L264R，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/L075Q/D130A/L264R，G023Q/L075Q/V187T/L264R，G091Q/V187T/I252Q/L264R，K024A/D130A/V154I/V187T，L075G/D130A/V187T/I252Q，L075G/V187H/I252Q/L264R，P029E/N073R/L075D/T189D，Q004D/D027N/E056K/P256T，Q004D/N011K/D027S/P256T，S058M/G163P/L227M/L264R，S058M/G163P/N233Q/L264R，S058M/L075Q/G091Q/I252Q，S058M/L075Q/I252Q/L264R，A018K/D027S/P029E/N033D/L075D，A018K/D130A/G156W/V187N/L264R，A018K/G023Q/K024A/L075Q/G156W，A018K/P029E/N033D/N073R/L075D，D027E/D048Q/D137Q/L227M/L264R，D027E/D048Q/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L264R，D027E/D130A/G163P/L227M/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L264R，D027E/S058M/D137Q/G163P/N233Q，D027E/S058M/G163P/L227M/L264R，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D027S/L075R/V154I/T189Q/L264R，D048Q/D130A/G163P/L227M/L264R，D048Q/D137Q/G163P/L227M/L264R，D048Q/D137Q/G163P/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/D137Q/N233Q/L264R，D048Q/S058M/G163P/N233Q/L264R，D130A/D137Q/G163P/L227M/L264R，D130A/D137Q/G163P/N233Q/L264R，E056K/L075G/V187N/I252Q/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/G091Q/D130A/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/L264R，F051T/L075Q/V187N/I252Q/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187N/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187N/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/E056K/V187N/I252Q/L264R，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075G/G091Q/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，K024A/L075R/V154I/G156W/V187Q，L075G/D130A/V187T/I252Q/L264R，N011K/G023K/D111A/G156W/L264R，S058M/D130A/D137Q/G163P/L264R，S058M/D130A/G163P/L227M/L264R，S058M/G163P/L227M/N233Q/L264R，A018K/D027S/N033D/N073R/L075D/T189D，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/D111A/V187N/T189Q/L264R，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D048Q/D130A/D137H/G163P/L264R，D027E/D048Q/D130A/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L227M/L264R，D027E/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/S058M/D130A/G163P/L227M/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/L264R，D048Q/S058M/D137Q/G163P/N233Q/L264R，D048Q/S058M/G163P/L227M/N233Q/L264R，F051T/E056K/D130A/V187N/I252Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/E056K/L075R/D130A/V187T/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/L075Q/D130A/V187T/I252Q，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/F051T/G091Q/V187N/I252Q/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/L075G/G091Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/V187N/I252Q/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，L075Q/G091Q/D130A/V187H/I252Q/L264R，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，S058M/D130A/G163P/L227M/N233Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/D130A/V154I/G156W/V187T/L264R，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，D027E/D048Q/D130A/D137Q/G163P/N233Q/L264R，D027E/D048Q/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/D048Q/S058M/D130A/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/S058M/L075Q/V187N/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023Q/D027S/F051T/E056K/D130A/I252Q/L264R，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，D027E/D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/D027S/F051T/S058M/L075Q/V187H/I252Q/L264R，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T表3-18：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比SEQIDNO：2在pH8水解棕榈酸对硝基苯酯的性能指数≥最大PI值的50％的TLL变体。
D130A/L264R，D130A/V187T，A018K/L075D/T189D，A018K/P029E/T189D，D027S/N033D/T189D，G023K/V187N/L264R，L075G/D130A/V187H，L075Q/V187N/L264R，L075R/D130A/L264R，L075R/D130A/V187T，Q004D/D027S/P256T，A018K/L075Q/V077I/N094R，D027S/D130A/I252Q/L264R，D027S/P029E/E045F/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，F051T/L075Q/I252Q/L264R，G023K/L075R/D130A/L264R，G023K/L075R/V187T/L264R，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G091Q/V187T/I252Q/L264R，L075G/V187H/I252Q/L264R，P029E/N073R/L075D/T189D，Q004D/N011K/D027S/P256T，S058M/G163P/L227M/L264R，S058M/L075Q/I252Q/L264R，A018K/D027S/P029E/N033D/L075D，A018K/G023Q/K024A/L075Q/G156W，A018K/P029E/N033D/N073R/L075D，D027E/D048Q/G163P/N233Q/L264R，D027E/D130A/G163P/L227M/L264R，D027E/G163P/L227M/N233Q/L264R，D027S/F051T/V187N/I252Q/L264R，D027S/L075R/V154I/T189Q/L264R，D048Q/D137Q/G163P/N233Q/L264R，D130A/D137Q/G163P/N233Q/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075Q/V187N/I252Q/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187N/L264R，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075Q/G091Q/I252Q/L264R，K024A/L075R/V154I/G156W/V187Q，S058M/G163P/L227M/N233Q/L264R，A018K/D027S/N033D/N073R/L075D/T189D，A018K/K024A/L075Q/N094R/V154I/G156W，D027E/D048Q/D130A/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，F051T/E056K/D130A/V187N/I252Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023Q/L075Q/G091Q/V187N/I252Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027S/F051T/S058M/L075Q/V187N/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/D027S/F051T/E056K/D130A/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R
表3-19：下面示出，相比TLL(SEQIDNO：1)，在pH6水解辛酸对硝基苯酯(辛酸酯)的性能指数＞1的TLL变体。
D027E/G163P，K024A/L075Q，P029E/N033D，A018K/D027S/E045F，A018K/L075D/T189D，A018K/L075Q/V187T，A018K/N073R/L075D，A018K/P029E/T189D，A049V/V187T/T189D，D027S/N033D/T189D，G023K/V187N/L264R，G023Q/L075Q/V187T，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/G156W/T189D，L075Q/G156W/V187N，L075R/D130A/L264R，L075R/D130A/V187T，N073R/L075D/T189D，P029E/N033D/E045F，P029E/N033D/L075D，A018K/D027S/N033D/L075D，A018K/D027S/P029E/T189D，A018K/G023Q/A049V/T189D，A018K/G023Q/L075R/D130A，A018K/G023Q/V077I/V187T，A018K/L075Q/V077I/N094R，A018K/P029E/N033D/T189D，D027E/D137Q/L227M/L264R，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D130A/G163P/L227M/L264R，G023K/D027Q/F051T/L075Q，G023K/L075Q/G156W/V187N，G023K/L075R/D130A/L264R，G023Q/E045F/A049V/T189D，G023Q/K024A/L075R/V154I，K024A/L075Q/D130A/V154I，K024A/L075R/G156W/V187N，L075Q/D130A/G156W/V187N，L075Q/V077I/G156W/V187N，L075Q/V077I/V187T/T189D，P029E/E045F/L075D/T189D，P029E/N033D/L075D/T189D，P029E/N073R/L075D/T189D，A018K/D027S/N033D/L075D/T189D，A018K/D027S/P029E/E045F/T189D，A018K/D027S/P029E/N033D/L075D，A018K/D027S/P029E/N073R/L075D，A018K/G023K/K024A/L075R/N094R，A018K/G023Q/E045F/A049V/L075Q，A018K/G023Q/E045F/L075Q/V187T，A018K/G023Q/K024A/D130A/V154I，A018K/G023Q/K024A/L075Q/G156W，A018K/G023Q/L075Q/G156W/V187T，A018K/K024A/L075R/D130A/V154I，A018K/K024A/N094R/D130A/V187N，A018K/L075Q/D111A/D130A/V187T，A018K/L075Q/N094R/G156W/V187N，A018K/P029E/N033D/N073R/L075D，D027E/D048Q/S058M/G163P/N233Q，D027S/L075R/V154I/T189Q/L264R，D130A/D137Q/G163P/N233Q/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/K024A/L075R/D130A/G156W，G023K/L075Q/D111A/D130A/V187T，G023K/L075Q/V077I/D130A/G156W，G023K/N094R/G156W/V187N/T189Q，G023Q/A049V/L075Q/G156W/V187T，G023Q/D027S/V154I/V187N/L264R，G023Q/K024A/L075Q/G156W/V187Q，K024A/L075R/V154I/G156W/V187Q，L075R/D130A/V154I/T189Q/L264R，L075R/D130A/V187T/T189Q/L264R，N011K/D027N/E056K/N233Q/P256T，N011K/G023K/E056K/L075Q/T189Q，A018K/D027S/P029E/N033D/E045F/L075D，A018K/D027S/P029E/N073R/L075D/T189D，A018K/G023K/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/N094R/D130A/V154I，A018K/G023K/L075Q/D130A/G156W/V187N，A018K/G023K/L075R/N094R/D130A/V187N，A018K/G023Q/E045F/A049V/L075Q/V187T，A018K/G023Q/L075Q/G156W/V187T/T189D，A018K/G023Q/L075Q/V077I/N094R/V154I，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/N094R/V154I/V187N，A018K/K024A/L075R/D130A/V187N/T189Q，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027Q/E056K/S058M/I090F/N233Q/P256T，D027Q/F051T/L075Q/D130A/V187H/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/P029E/N033D/E045F/N073R/T189D，G023K/K024A/L075Q/N094R/D130A/G156W，G023K/K024A/L075Q/V154I/G156W/V187Q，G023Q/K024A/L075Q/D130A/V154I/G156W，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/V154I/V187T/T189Q/L264R，L075R/D130A/V154I/G156W/V187N/L264R，N011K/E056K/L075Q/D130A/V187N/T189Q，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/D130A/G156W/V187N，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/K024A/L075R/V154I/G156W/V187Q，A018K/G023Q/E045F/L075Q/V077I/G156W/V187T，A018K/G023Q/K024A/L075Q/V077I/N094R/V154I，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，E056K/L075Q/D111A/D130A/G156W/V187N/T189Q，G023K/D027Q/F051T/L075G/D130A/V187H/L264R，G023K/L075R/N094R/V154I/G156W/V187N/L264R，G023Q/D027S/L075Q/G091Q/V187T/I252Q/L264R，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/L075Q/D130A/V154I/G156W/T189Q/L264R，A018K/G023K/K024A/L075Q/D130Y/V154I/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，G023K/D027S/E056K/L075G/D130A/V187T/I252Q/L264R，G023K/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，A018K/G023K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，G023K/D027S/F051T/E056K/S058M/L075G/G091Q/D130A/I252Q/L264R
表3-20：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且在pH6水解辛酸对硝基苯酯(辛酸酯)的性能指数＞1的TLL变体。
D027E/G163P，P029E/N033D，A018K/L075D/T189D，A018K/P029E/T189D，D027S/N033D/T189D，G023K/V187N/L264R，L075G/D130A/V187H，L075R/D130A/L264R，L075R/D130A/V187T，P029E/N033D/E045F，A018K/D027S/P029E/T189D，A018K/L075Q/V077I/N094R，D027E/D137Q/L227M/L264R，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D130A/G163P/L227M/L264R，G023K/D027Q/F051T/L075Q，G023K/L075R/D130A/L264R，P029E/N073R/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/G023Q/K024A/L075Q/G156W，A018K/K024A/L075R/D130A/V154I，A018K/K024A/N094R/D130A/V187N，A018K/P029E/N033D/N073R/L075D，D027S/L075R/V154I/T189Q/L264R，D130A/D137Q/G163P/N233Q/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/N094R/G156W/V187N/T189Q，G023Q/K024A/L075Q/G156W/V187Q，K024A/L075R/V154I/G156W/V187Q，A018K/D027S/P029E/N033D/E045F/L075D，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，G023K/K024A/L075Q/V154I/G156W/V187Q，K024A/L075Q/D111A/V154I/V187N/T189Q，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H
表3-21：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且在pH6水解辛酸对硝基苯酯(辛酸酯)的性能指数≥最大PI值的50％的TLL变体。
A018K/P029E/T189D，D027S/N033D/T189D，G023K/V187N/L264R，L075R/D130A/V187T，A018K/D027S/P029E/T189D，A018K/L075Q/V077I/N094R，D027S/P029E/E045F/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D130A/G163P/L227M/L264R，P029E/N073R/L075D/T189D，A018K/G023Q/K024A/L075Q/G156W，A018K/K024A/L075R/D130A/V154I，G023K/E056K/D130A/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，K024A/L075R/V154I/G156W/V187Q，A018K/D027S/P029E/N033D/E045F/L075D，A018K/K024A/L075Q/N094R/V154I/G156W，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W
表3-22：下面示出，相比参考序列SEQIDNO：2，在pH6水解辛酸对硝基苯酯(辛酸酯)的性能指数＞1的TLL变体。
D027E/G163P，K024A/L075Q，A018K/D027S/E045F，A018K/L075D/T189D，A018K/L075Q/V187T，A018K/N073R/L075D，A018K/P029E/T189D，A049V/V187T/T189D，D027S/N033D/T189D，G023K/V187N/L264R，G023Q/L075Q/V187T，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/G156W/T189D，L075Q/G156W/V187N，L075R/D130A/L264R，L075R/D130A/V187T，N073R/L075D/T189D，P029E/N033D/L075D，A018K/D027S/N033D/L075D，A018K/D027S/P029E/T189D，A018K/G023Q/A049V/T189D，A018K/G023Q/L075R/D130A，A018K/G023Q/V077I/V187T，A018K/L075Q/V077I/N094R，A018K/P029E/N033D/T189D，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D130A/G163P/L227M/L264R，G023K/D027Q/F051T/L075Q，G023K/L075Q/G156W/V187N，G023K/L075R/D130A/L264R，G023Q/E045F/A049V/T189D，G023Q/K024A/L075R/V154I，K024A/L075Q/D130A/V154I，K024A/L075R/G156W/V187N，L075Q/D130A/G156W/V187N，L075Q/V077I/G156W/V187N，L075Q/V077I/V187T/T189D，P029E/E045F/L075D/T189D，P029E/N033D/L075D/T189D，P029E/N073R/L075D/T189D，A018K/D027S/N033D/L075D/T189D，A018K/D027S/P029E/E045F/T189D，A018K/D027S/P029E/N033D/L075D，A018K/D027S/P029E/N073R/L075D，A018K/G023K/K024A/L075R/N094R，A018K/G023Q/E045F/A049V/L075Q，A018K/G023Q/E045F/L075Q/V187T，A018K/G023Q/K024A/D130A/V154I，A018K/G023Q/K024A/L075Q/G156W，A018K/G023Q/L075Q/G156W/V187T，A018K/K024A/L075R/D130A/V154I，A018K/K024A/N094R/D130A/V187N，A018K/L075Q/D111A/D130A/V187T，A018K/L075Q/N094R/G156W/V187N，A018K/P029E/N033D/N073R/L075D，D027E/D048Q/S058M/G163P/N233Q，D027S/L075R/V154I/T189Q/L264R，D130A/D137Q/G163P/N233Q/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/K024A/L075R/D130A/G156W，G023K/L075Q/D111A/D130A/V187T，G023K/L075Q/V077I/D130A/G156W，G023K/N094R/G156W/V187N/T189Q，G023Q/A049V/L075Q/G156W/V187T，G023Q/K024A/L075Q/G156W/V187Q，K024A/L075R/V154I/G156W/V187Q，L075R/D130A/V154I/T189Q/L264R，L075R/D130A/V187T/T189Q/L264R，N011K/D027N/E056K/N233Q/P256T，N011K/G023K/E056K/L075Q/T189Q，A018K/D027S/P029E/N033D/E045F/L075D，A018K/D027S/P029E/N073R/L075D/T189D，A018K/G023K/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/N094R/D130A/V154I，A018K/G023K/L075Q/D130A/G156W/V187N，A018K/G023K/L075R/N094R/D130A/V187N，A018K/G023Q/E045F/A049V/L075Q/V187T，A018K/G023Q/L075Q/G156W/V187T/T189D，A018K/G023Q/L075Q/V077I/N094R/V154I，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/N094R/V154I/V187N，A018K/K024A/L075R/D130A/V187N/T189Q，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027Q/E056K/S058M/I090F/N233Q/P256T，D027Q/F051T/L075Q/D130A/V187H/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/P029E/N033D/E045F/N073R/T189D，G023K/K024A/L075Q/N094R/D130A/G156W，G023K/K024A/L075Q/V154I/G156W/V187Q，G023Q/K024A/L075Q/D130A/V154I/G156W，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/V154I/V187T/T189Q/L264R，L075R/D130A/V154I/G156W/V187N/L264R，N011K/E056K/L075Q/D130A/V187N/T189Q，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/D130A/G156W/V187N，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/K024A/L075R/V154I/G156W/V187Q，A018K/G023Q/E045F/L075Q/V077I/G156W/V187T，A018K/G023Q/K024A/L075Q/V077I/N094R/V154I，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，E056K/L075Q/D111A/D130A/G156W/V187N/T189Q，G023K/D027Q/F051T/L075G/D130A/V187H/L264R，G023K/L075R/N094R/V154I/G156W/V187N/L264R，G023Q/D027S/L075Q/G091Q/V187T/I252Q/L264R，G023Q/L075Q/D130A/V154I/G156W/T189Q/L264R，A018K/G023K/K024A/L075Q/D130Y/V154I/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，G023K/D027S/E056K/L075G/D130A/V187T/I252Q/L264R，G023K/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，A018K/G023K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，G023K/D027S/F051T/E056K/S058M/L075G/G091Q/D130A/I252Q/L264R
表3-23：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比SEQIDNO：2在pH6水解辛酸对硝基苯酯(辛酸酯)的性能指数＞1的TLL变体。
D027E/G163P，A018K/L075D/T189D，A018K/P029E/T189D，D027S/N033D/T189D，G023K/V187N/L264R，L075G/D130A/V187H，L075R/D130A/L264R，L075R/D130A/V187T，A018K/D027S/P029E/T189D，A018K/L075Q/V077I/N094R，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D130A/G163P/L227M/L264R，G023K/D027Q/F051T/L075Q，G023K/L075R/D130A/L264R，P029E/N073R/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/G023Q/K024A/L075Q/G156W，A018K/K024A/L075R/D130A/V154I，A018K/K024A/N094R/D130A/V187N，A018K/P029E/N033D/N073R/L075D，D027S/L075R/V154I/T189Q/L264R，D130A/D137Q/G163P/N233Q/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/N094R/G156W/V187N/T189Q，G023Q/K024A/L075Q/G156W/V187Q，K024A/L075R/V154I/G156W/V187Q，A018K/D027S/P029E/N033D/E045F/L075D，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，G023K/K024A/L075Q/V154I/G156W/V187Q，K024A/L075Q/D111A/V154I/V187N/T189Q，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H
表3-24：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比SEQIDNO：2在pH6水解辛酸对硝基苯酯(辛酸酯)的性能指数≥最大PI值的50％的TLL变体。
A018K/P029E/T189D，D027S/N033D/T189D，G023K/V187N/L264R，L075R/D130A/V187T，A018K/D027S/P029E/T189D，A018K/L075Q/V077I/N094R，D027S/P029E/E045F/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D130A/G163P/L227M/L264R，P029E/N073R/L075D/T189D，A018K/G023Q/K024A/L075Q/G156W，A018K/K024A/L075R/D130A/V154I，G023K/E056K/D130A/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，K024A/L075R/V154I/G156W/V187Q，A018K/D027S/P029E/N033D/E045F/L075D，A018K/K024A/L075Q/N094R/V154I/G156W，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W
表3-25：下面示出，相比TLL(SEQIDNO：1)，pNPP/pNPB或pNPO/pNPB比活性比值增大的TLL变体。
A018K/E045F，D111A/L264R，D130A/V187T，G023K/L264R，L075R/L264R，S058M/L264R，V187T/L264R，A018K/E045F/T189D，A018K/G023K/L075Q，A018K/N033D/T189D，D027N/N233Q/P256T，D027Q/N233Q/P256T，D027Q/S058M/P256T，D111A/D130A/L264R，D130A/V154I/G156W，D130A/V187N/L264R，F051T/I252Q/L264R，G023K/D130A/L264R，G023K/E056K/V187T，G023K/L075R/L264R，G023K/V187T/L264R，G163P/L227M/L264R，L075Q/V187T/L264R，Q004D/D027S/P256T，A018K/D130A/G156W/V187T，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/V187T，A018K/G023Q/V077I/G156W，D027E/D048Q/G163P/L264R，D027E/D130A/N233Q/L264R，D027E/G163P/L227M/L264R，D027S/D130A/I252Q/L264R，D027S/E045F/N073R/L075D，D027S/N033D/E045F/T189D，D048Q/D130A/G163P/L264R，D111A/D130A/V154I/G156W，D111A/D130A/V187T/L264R，E056K/D130A/V187N/L264R，F051T/D130A/I252Q/L264R，F051T/L075G/I252Q/L264R，F051T/L075Q/I252Q/L264R，G023K/D130A/V187N/L264R，G023K/D130A/V187T/T189Q，G023K/E056K/L075R/L264R，G023K/L075R/V187T/L264R，G023Q/E045F/A049V/V187T，G023Q/E045F/L075Q/G156W，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/L075Q/D130A/L264R，G023Q/L075Q/V187T/L264R，G091Q/V187T/I252Q/L264R，K024A/D130A/V154I/V187T，L075G/D130A/V187T/I252Q，L075G/V187H/I252Q/L264R，N011K/S058M/N233Q/P256T，P029E/E045F/N073R/T189D，Q004D/D027N/E056K/P256T，Q004D/N011K/D027N/P256T，Q004D/N011K/D027S/P256T，S058M/G163P/L227M/L264R，S058M/G163P/N233Q/L264R，S058M/L075Q/I252Q/L264R，A018K/D130A/G156W/V187N/L264R，A018K/E045F/A049V/L075Q/T189D，A018K/G023K/K024A/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/A049V/L075Q/V187T，A018K/G023Q/E045F/V077I/T189D，A018K/G023Q/K024A/L075Q/G156W，A018K/K024A/N094R/G156W/V187T，A018K/L075Q/N094R/D130A/V187N，D027E/D048Q/D137Q/L227M/L264R，D027E/D048Q/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L264R，D027E/D130A/G163P/L227M/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L264R，D027N/E056K/S058M/N233Q/P256T，D027S/E056K/S058M/N233Q/P256T，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D027S/L075R/V154I/T189Q/L264R，D027S/N033D/E045F/N073R/L075D，D027S/P029E/N033D/E045F/N073R，D048Q/D130A/G163P/L227M/L264R，D048Q/D137Q/G163P/L227M/L264R，D048Q/D137Q/G163P/N233Q/L264R，D048Q/S058M/D137Q/N233Q/L264R，D130A/D137Q/G163P/L227M/L264R，D130A/V154I/G156W/V187N/T189Q，E056K/L075G/V187N/I252Q/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/G091Q/D130A/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/L264R，F051T/L075Q/V187N/I252Q/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/E056K/D130A/V187N/L264R，G023K/K024A/D111A/D130A/V187T，G023K/K024A/D111A/G156W/T189Q，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187N/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/E045F/A049V/V077I/T189D，G023Q/E045F/L075Q/V077I/V187T，G023Q/E056K/V187N/I252Q/L264R，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/L075G/G091Q/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，K024A/L075R/V154I/G156W/V187Q，L075G/D130A/V187T/I252Q/L264R，L075Q/D111A/D130A/V187T/T189Q，L075Q/V077I/D130A/G156W/V187N，N011K/D027N/S058M/N233Q/P256T，N011K/D027Q/S058M/N233Q/P256T，S058M/D130A/G163P/L227M/L264R，S058M/G163P/L227M/N233Q/L264R，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/E045F/L075Q/G156W/V187T，A018K/G023Q/K024A/V077I/V154I/V187N，A018K/G023Q/K024A/V154I/G156W/V187Q，A018K/K024A/D111A/V187N/T189Q/L264R，A018K/K024A/V077I/N094R/D130A/G156W，D027E/D048Q/D130A/D137H/G163P/L264R，D027E/D048Q/D130A/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D130A/D137Q/G163P/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L227M/L264R，D027E/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/F051T/G091Q/D130A/I252Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/S058M/D130A/G163P/L227M/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/L264R，D048Q/S058M/G163P/L227M/N233Q/L264R，F051T/E056K/D130A/V187N/I252Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/V187T/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/L075Q/D130A/V187T/I252Q，G023Q/F051T/G091Q/V187N/I252Q/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/L075G/G091Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/V187N/I252Q/L264R，G023Q/L075R/V154I/G156W/V187N/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/D130A/G156W/T189Q/L264R，L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/D027Q/E056K/S058M/N233Q/P256T，N011K/D027S/E056K/S058M/N233Q/P256T，Q004D/D027N/E056K/S058M/N233Q/P256T，Q004D/N011K/D027N/I090F/N233Q/P256T，Q004D/N011K/D027S/I090F/N233Q/P256T，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/G163P/L227M/N233Q/L264R，A018K/G023Q/E045F/A049V/L075Q/G156W/V187T，A018K/K024A/D130A/V154I/G156W/V187T/L264R，A018K/K024A/N094R/D130A/G156W/V187T/L264R，A018K/L075Q/D130A/G156W/V187N/T189Q/L264R，D027E/D048Q/D130A/D137Q/G163P/N233Q/L264R，D027E/D048Q/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/S058M/L075Q/V187N/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/S058M/G091Q/V187N/I252Q，G023Q/D027S/F051T/E056K/D130A/I252Q/L264R，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，Q004D/N011K/D027N/S058M/I090F/N233Q/P256T，D027E/D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/S058M/L075Q/V187H/I252Q/L264R，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，A018K/G023K/K024A/L075Q/N094R/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/V154I/G156W/V187N，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R
表3-26：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且pNPP/pNPB或pNPO/pNPB比活性比值增大的TLL变体。
A018K/E045F，D111A/L264R，D130A/V187T，G023K/L264R，L075R/L264R，S058M/L264R，V187T/L264R，A018K/N033D/T189D，D027Q/S058M/P256T，D111A/D130A/L264R，D130A/V154I/G156W，D130A/V187N/L264R，F051T/I252Q/L264R，G023K/D130A/L264R，G023K/E056K/V187T，G023K/L075R/L264R，G023K/V187T/L264R，G163P/L227M/L264R，L075Q/V187T/L264R，Q004D/D027S/P256T，A018K/D130A/G156W/V187T，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/V187T，A018K/G023Q/V077I/G156W，D027E/D048Q/G163P/L264R，D027E/D130A/N233Q/L264R，D027E/G163P/L227M/L264R，D027S/D130A/I252Q/L264R，D027S/N033D/E045F/T189D，D048Q/D130A/G163P/L264R，D111A/D130A/V154I/G156W，D111A/D130A/V187T/L264R，E056K/D130A/V187N/L264R，F051T/D130A/I252Q/L264R，F051T/L075G/I252Q/L264R，F051T/L075Q/I252Q/L264R，G023K/D130A/V187N/L264R，G023K/D130A/V187T/T189Q，G023K/E056K/L075R/L264R，G023K/L075R/V187T/L264R，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/L075Q/D130A/L264R，G023Q/L075Q/V187T/L264R，G091Q/V187T/I252Q/L264R，K024A/D130A/V154I/V187T，L075G/D130A/V187T/I252Q，L075G/V187H/I252Q/L264R，Q004D/D027N/E056K/P256T，Q004D/N011K/D027S/P256T，S058M/G163P/L227M/L264R，S058M/G163P/N233Q/L264R，S058M/L075Q/I252Q/L264R，A018K/D130A/G156W/V187N/L264R，A018K/G023K/K024A/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/K024A/L075Q/G156W，A018K/K024A/N094R/G156W/V187T，D027E/D048Q/D137Q/L227M/L264R，D027E/D048Q/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L264R，D027E/D130A/G163P/L227M/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L264R，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D027S/L075R/V154I/T189Q/L264R，D048Q/D130A/G163P/L227M/L264R，D048Q/D137Q/G163P/L227M/L264R，D048Q/D137Q/G163P/N233Q/L264R，D048Q/S058M/D137Q/N233Q/L264R，D130A/D137Q/G163P/L227M/L264R，D130A/V154I/G156W/V187N/T189Q，E056K/L075G/V187N/I252Q/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/G091Q/D130A/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/L264R，F051T/L075Q/V187N/I252Q/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/E056K/D130A/V187N/L264R，G023K/K024A/D111A/D130A/V187T，G023K/K024A/D111A/G156W/T189Q，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187N/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/E056K/V187N/I252Q/L264R，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/L075G/G091Q/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，K024A/L075R/V154I/G156W/V187Q，L075G/D130A/V187T/I252Q/L264R，N011K/D027N/S058M/N233Q/P256T，S058M/D130A/G163P/L227M/L264R，S058M/G163P/L227M/N233Q/L264R，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/K024A/V077I/V154I/V187N，A018K/G023Q/K024A/V154I/G156W/V187Q，A018K/K024A/D111A/V187N/T189Q/L264R，D027E/D048Q/D130A/D137H/G163P/L264R，D027E/D048Q/D130A/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D130A/D137Q/G163P/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L227M/L264R，D027E/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/F051T/G091Q/D130A/I252Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/S058M/D130A/G163P/L227M/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/L264R，D048Q/S058M/G163P/L227M/N233Q/L264R，F051T/E056K/D130A/V187N/I252Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/V187T/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/L075Q/D130A/V187T/I252Q，G023Q/F051T/G091Q/V187N/I252Q/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/L075G/G091Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/V187N/I252Q/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，L075Q/G091Q/D130A/V187H/I252Q/L264R，Q004D/N011K/D027N/I090F/N233Q/P256T，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/G163P/L227M/N233Q/L264R，A018K/K024A/D130A/V154I/G156W/V187T/L264R，A018K/K024A/N094R/D130A/G156W/V187T/L264R，D027E/D048Q/D130A/D137Q/G163P/N233Q/L264R，D027E/D048Q/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/S058M/L075Q/V187N/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/S058M/G091Q/V187N/I252Q，G023Q/D027S/F051T/E056K/D130A/I252Q/L264R，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/S058M/L075Q/V187H/I252Q/L264R，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R
表3-27：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且pNPP/pNPB或pNPO/pNPB≥最大比活性比值的50％的TLL变体。
D130A/V187T，D027Q/S058M/P256T，F051T/I252Q/L264R，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/V187T，D027S/D130A/I252Q/L264R，D027S/N033D/E045F/T189D，F051T/D130A/I252Q/L264R，F051T/L075G/I252Q/L264R，F051T/L075Q/I252Q/L264R，G023K/D130A/V187T/T189Q，G023K/L075R/V187T/L264R，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/L075Q/V187T/L264R，G091Q/V187T/I252Q/L264R，L075G/V187H/I252Q/L264R，Q004D/N011K/D027S/P256T，S058M/G163P/L227M/L264R，S058M/L075Q/I252Q/L264R，D027E/D048Q/D137Q/L227M/L264R，D027E/D048Q/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L264R，D027E/D130A/G163P/L227M/L264R，D027E/G163P/L227M/N233Q/L264R，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D048Q/D137Q/G163P/N233Q/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/V187N/I252Q/L264R，G023K/E056K/D130A/V187N/L264R，G023K/K024A/D111A/G156W/T189Q，G023Q/E056K/V187N/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，N011K/D027N/S058M/N233Q/P256T，A018K/G023Q/K024A/V077I/V154I/V187N，A018K/K024A/D111A/V187N/T189Q/L264R，D027E/D048Q/D130A/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，F051T/E056K/D130A/V187N/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023Q/F051T/G091Q/V187N/I252Q/L264R，G023Q/L075G/G091Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/V187N/I252Q/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，L075Q/G091Q/D130A/V187H/I252Q/L264R，A018K/K024A/D130A/V154I/G156W/V187T/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027S/F051T/S058M/L075Q/V187N/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/S058M/G091Q/V187N/I252Q，G023Q/D027S/F051T/E056K/D130A/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/S058M/L075Q/V187H/I252Q/L264R，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R
表3-28：下面示出，相比参考序列SEQIDNO：2，pNPP/pNPB或pNPO/pNPB比活性比值增大的TLL变体。
A018K/E045F，D130A/V187T，G023K/L264R，S058M/L264R，A018K/E045F/T189D，A018K/G023K/L075Q，D027N/N233Q/P256T，D027Q/N233Q/P256T，D130A/V187N/L264R，F051T/I252Q/L264R，G023K/D130A/L264R，G023K/V187T/L264R，Q004D/D027S/P256T，D027E/D130A/N233Q/L264R，D027E/G163P/L227M/L264R，D027S/D130A/I252Q/L264R，D027S/E045F/N073R/L075D，D048Q/D130A/G163P/L264R，E056K/D130A/V187N/L264R，F051T/D130A/I252Q/L264R，F051T/L075G/I252Q/L264R，F051T/L075Q/I252Q/L264R，G023K/D130A/V187N/L264R，G023K/E056K/L075R/L264R，G023K/L075R/V187T/L264R，G023Q/E045F/A049V/V187T，G023Q/E045F/L075Q/G156W，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/L075Q/V187T/L264R，G091Q/V187T/I252Q/L264R，L075G/D130A/V187T/I252Q，L075G/V187H/I252Q/L264R，N011K/S058M/N233Q/P256T，P029E/E045F/N073R/T189D，Q004D/N011K/D027S/P256T，S058M/G163P/L227M/L264R，S058M/L075Q/I252Q/L264R，A018K/D130A/G156W/V187N/L264R，A018K/G023Q/E045F/V077I/T189D，D027E/D048Q/D137Q/L227M/L264R，D027E/D048Q/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L264R，D027E/D130A/G163P/L227M/L264R，D027E/G163P/L227M/N233Q/L264R，D027S/E056K/S058M/N233Q/P256T，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D027S/N033D/E045F/N073R/L075D，D027S/P029E/N033D/E045F/N073R，D048Q/D137Q/G163P/N233Q/L264R，D048Q/S058M/D137Q/N233Q/L264R，D130A/D137Q/G163P/L227M/L264R，E056K/L075G/V187N/I252Q/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/V187N/I252Q/L264R，G023K/E056K/D130A/V187N/L264R，G023K/L075R/D130A/V187N/L264R，G023Q/E045F/A049V/V077I/T189D，G023Q/E045F/L075Q/V077I/V187T，G023Q/E056K/V187N/I252Q/L264R，G023Q/L075G/G091Q/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，L075G/D130A/V187T/I252Q/L264R，S058M/G163P/L227M/N233Q/L264R，A018K/G023Q/K024A/V154I/G156W/V187Q，A018K/K024A/D111A/V187N/T189Q/L264R，A018K/K024A/V077I/N094R/D130A/G156W，D027E/D048Q/D130A/D137H/G163P/L264R，D027E/D048Q/D130A/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L227M/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027Q/F051T/G091Q/D130A/I252Q/L264R，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/S058M/D130A/G163P/L227M/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/L264R，D048Q/S058M/G163P/L227M/N233Q/L264R，F051T/E056K/D130A/V187N/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/F051T/G091Q/V187N/I252Q/L264R，G023Q/L075G/G091Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/V187N/I252Q/L264R，G023Q/L075R/V154I/G156W/V187N/L264R，K024A/L075Q/D130A/G156W/T189Q/L264R，L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/D027Q/E056K/S058M/N233Q/P256T，N011K/D027S/E056K/S058M/N233Q/P256T，Q004D/D027N/E056K/S058M/N233Q/P256T，Q004D/N011K/D027S/I090F/N233Q/P256T，A018K/K024A/D130A/V154I/G156W/V187T/L264R，A018K/K024A/N094R/D130A/G156W/V187T/L264R，D027E/D048Q/D130A/D137Q/G163P/N233Q/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/S058M/L075Q/V187N/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/S058M/G091Q/V187N/I252Q，G023Q/D027S/F051T/E056K/D130A/I252Q/L264R，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，Q004D/N011K/D027N/S058M/I090F/N233Q/P256T，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/S058M/L075Q/V187H/I252Q/L264R，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，A018K/G023K/K024A/L075Q/N094R/D130A/V154I/G156W/V187N，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R
表3-29：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比SEQIDNO：2而言pNPP/pNPB或pNPO/pNPB比活性比值增大的TLL变体。
A018K/E045F，D130A/V187T，G023K/L264R，S058M/L264R，D130A/V187N/L264R，F051T/I252Q/L264R，G023K/D130A/L264R，G023K/V187T/L264R，Q004D/D027S/P256T，D027E/D130A/N233Q/L264R，D027E/G163P/L227M/L264R，D027S/D130A/I252Q/L264R，D048Q/D130A/G163P/L264R，E056K/D130A/V187N/L264R，F051T/D130A/I252Q/L264R，F051T/L075G/I252Q/L264R，F051T/L075Q/I252Q/L264R，G023K/D130A/V187N/L264R，G023K/E056K/L075R/L264R，G023K/L075R/V187T/L264R，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/L075Q/V187T/L264R，G091Q/V187T/I252Q/L264R，L075G/D130A/V187T/I252Q，L075G/V187H/I252Q/L264R，Q004D/N011K/D027S/P256T，S058M/G163P/L227M/L264R，S058M/L075Q/I252Q/L264R，A018K/D130A/G156W/V187N/L264R，D027E/D048Q/D137Q/L227M/L264R，D027E/D048Q/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L264R，D027E/D130A/G163P/L227M/L264R，D027E/G163P/L227M/N233Q/L264R，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D048Q/D137Q/G163P/N233Q/L264R，D048Q/S058M/D137Q/N233Q/L264R，D130A/D137Q/G163P/L227M/L264R，E056K/L075G/V187N/I252Q/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/V187N/I252Q/L264R，G023K/E056K/D130A/V187N/L264R，G023K/L075R/D130A/V187N/L264R，G023Q/E056K/V187N/I252Q/L264R，G023Q/L075G/G091Q/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，L075G/D130A/V187T/I252Q/L264R，S058M/G163P/L227M/N233Q/L264R，A018K/G023Q/K024A/V154I/G156W/V187Q，A018K/K024A/D111A/V187N/T189Q/L264R，D027E/D048Q/D130A/D137H/G163P/L264R，D027E/D048Q/D130A/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L227M/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027Q/F051T/G091Q/D130A/I252Q/L264R，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/S058M/D130A/G163P/L227M/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/L264R，D048Q/S058M/G163P/L227M/N233Q/L264R，F051T/E056K/D130A/V187N/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/F051T/G091Q/V187N/I252Q/L264R，G023Q/L075G/G091Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/V187N/I252Q/L264R，L075Q/G091Q/D130A/V187H/I252Q/L264R，A018K/K024A/D130A/V154I/G156W/V187T/L264R，A018K/K024A/N094R/D130A/G156W/V187T/L264R，D027E/D048Q/D130A/D137Q/G163P/N233Q/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/S058M/L075Q/V187N/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/S058M/G091Q/V187N/I252Q，G023Q/D027S/F051T/E056K/D130A/I252Q/L264R，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/S058M/L075Q/V187H/I252Q/L264R，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R
表3-30：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比SEQIDNO：2而言pNPP/pNPB或pNPO/pNPB≥最大比活性比值的50％的TLL变体。
D130A/V187T，F051T/I252Q/L264R，D027S/D130A/I252Q/L264R，F051T/D130A/I252Q/L264R，F051T/L075G/I252Q/L264R，F051T/L075Q/I252Q/L264R，G023K/L075R/V187T/L264R，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/L075Q/V187T/L264R，G091Q/V187T/I252Q/L264R，L075G/V187H/I252Q/L264R，Q004D/N011K/D027S/P256T，S058M/G163P/L227M/L264R，S058M/L075Q/I252Q/L264R，D027E/D048Q/D137Q/L227M/L264R，D027E/D048Q/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L264R，D027E/D130A/G163P/L227M/L264R，D027E/G163P/L227M/N233Q/L264R，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D048Q/D137Q/G163P/N233Q/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/V187N/I252Q/L264R，G023K/E056K/D130A/V187N/L264R，G023Q/E056K/V187N/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，A018K/K024A/D111A/V187N/T189Q/L264R，D027E/D048Q/D130A/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，F051T/E056K/D130A/V187N/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023Q/F051T/G091Q/V187N/I252Q/L264R，G023Q/L075G/G091Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/V187N/I252Q/L264R，L075Q/G091Q/D130A/V187H/I252Q/L264R，A018K/K024A/D130A/V154I/G156W/V187T/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027S/F051T/S058M/L075Q/V187N/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/D027S/F051T/E056K/D130A/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/S058M/L075Q/V187H/I252Q/L264R，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R
表3-31：下面示出，相比TLL(SEQIDNO：1)，pNPB/pNPP比活性比值增大的TLL变体。
A018K/L075D，A018K/T189D，D027N/E056K，D027N/N233Q，D027S/N033D，D027S/P256T，D130A/L264R，D130A/V187N，E045F/N073R，K024A/L075Q，K024A/V154I，L075Q/D130A，N094R/G156W，P029E/N033D，A018K/A049V/V187T，A018K/E045F/N073R，A018K/E045F/V187T，A018K/G023K/G156W，A018K/K024A/V154I，A018K/L075D/T189D，A049V/V187T/T189D，D027E/S058M/G163P，D027N/S058M/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/S058M/P256T，D130A/V187T/L264R，E045F/L075D/T189D，E056K/D130A/L264R，G023K/D130A/V187T，G023Q/A049V/T189D，G023Q/D130A/G156W，G023Q/K024A/G156W，G023Q/L075Q/V187T，K024A/D130A/V154I，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/G156W/V187N，L075Q/V187N/L264R，L075R/D130A/L264R，L075R/D130A/V187T，L075R/V187T/L264R，N011K/D027S/S058M，N011K/N233Q/P256T，N073R/L075D/T189D，N094R/D130A/V187T，P029E/N033D/E045F，V187N/T189Q/L264R，A018K/D027S/N033D/L075D，A018K/D111A/G156W/T189Q，A018K/E045F/A049V/G156W，A018K/E045F/A049V/V187T，A018K/E045F/L075D/T189D，A018K/E045F/L075Q/T189D，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/V187T，A018K/G023Q/E045F/T189D，A018K/G023Q/E045F/V187T，A018K/G023Q/L075R/D130A，A018K/G023Q/V077I/V187T，A018K/L075Q/G156W/V187T，A018K/L075Q/N094R/D130A，A018K/L075Q/V077I/N094R，A018K/P029E/N073R/L075D，A018K/V154I/G156W/V187T，D027E/D137Q/G163P/L227M，D027E/S058M/G163P/L264R，D027N/E056K/N233Q/P256T，D027N/E056K/S058M/P256T，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/E045F/L075D/T189D，D027S/E045F/N073R/T189D，D027S/E056K/D111A/V154I，D027S/E056K/I090F/P256T，D027S/N033D/E045F/N073R，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/L075D，D027S/P029E/E045F/N073R，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/G163P/N233Q/L264R，D111A/D130A/V154I/G156W，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，E056K/D130A/V187T/L264R，G023K/D027Q/F051T/L075Q，G023K/D130A/G156W/T189Q，G023K/K024A/V154I/V187N，G023K/L075Q/G156W/V187N，G023K/L075R/D130A/L264R，G023Q/A049V/V077I/G156W，G023Q/D027S/D111A/G156W，G023Q/K024A/D130A/G156W，G023Q/K024A/L075R/V154I，G023Q/K024A/V077I/G156W，G023Q/V154I/G156W/V187N，K024A/L075R/G156W/V187N，L075G/G091Q/V187N/L264R，L075Q/D130A/G156W/V187N，L075Q/V077I/G156W/V187N，L075Q/V187N/T189Q/L264R，L075R/D130A/V187T/L264R，N011K/D027N/E056K/S058M，N011K/D027Q/S058M/P256T，N011K/D027S/E056K/P256T，N011K/D027S/N233Q/P256T，N011K/E056K/N233Q/P256T，N033D/E045F/N073R/T189A，P029E/E045F/N073R/L075D，P029E/N033D/L075D/T189D，P029E/N073R/L075D/T189D，Q004D/D027N/S058M/P256T，Q004D/D027S/I090F/P256T，S058M/D137Q/G163P/N233Q，S058M/L075Q/G091Q/I252Q，A018K/A049V/L075Q/V187T/T189D，A018K/D027S/E045F/L075D/T189D，A018K/D027S/N033D/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/E045F/A049V/L075Q/V187T，A018K/G023K/D130A/V154I/G156W，A018K/G023K/K024A/D130A/G156W，A018K/G023K/K024A/L075R/N094R，A018K/G023Q/A049V/L075Q/G156W，A018K/G023Q/A049V/L075Q/V187T，A018K/G023Q/E045F/A049V/V187T，A018K/G023Q/E045F/L075Q/V187T，A018K/G023Q/K024A/D130A/V154I，A018K/G023Q/K024A/D130A/V187Q，A018K/G023Q/L075Q/G156W/V187T，A018K/G023Q/L075Q/V187T/T189D，A018K/G023Q/V077I/D130A/G156W，A018K/G023Q/V077I/V187T/T189D，A018K/K024A/L075Q/D130A/L264R，A018K/K024A/L075Q/V154I/T189D，A018K/K024A/N094R/D130A/V187N，A018K/K024A/N094R/G156W/V187T，A018K/L075Q/D111A/D130A/V187T，A018K/L075Q/N094R/G156W/V187N，A018K/P029E/E045F/N073R/L075D，A018K/P029E/N033D/N073R/L075D，A018K/V077I/G156W/V187T/T189D，D027E/S058M/D137Q/G163P/N233Q，D027E/S058M/G163P/L227M/L264R，D027Q/E056K/I090F/N233Q/P256T，D027Q/E056K/S058M/N233Q/P256T，D027S/P029E/N033D/E045F/N073R，D048Q/S058M/D130A/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/G163P/L227M/L264R，D048Q/S058M/G163P/N233Q/L264R，D130A/D137Q/G163P/N233Q/L264R，D130A/G163P/L227M/N233Q/L264R，E045F/A049V/L075Q/V187T/T189D，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/K024A/D111A/D130A/V187T，G023K/K024A/L075R/D130A/G156W，G023K/L075Q/D111A/D130A/V187T，G023K/L075Q/D130A/V154I/T189Q，G023K/N094R/G156W/V187N/T189Q，G023Q/A049V/L075Q/G156W/V187T，G023Q/E045F/A049V/G156W/T189D，G023Q/E045F/A049V/L075Q/V077I，G023Q/K024A/L075Q/G156W/V187Q，G023Q/K024A/L075R/D130A/V154I，G023Q/K024A/L075R/V077I/D130A，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/D111A/V154I/V187T，K024A/L075R/G156W/V187N/T189Q，L075Q/D130A/V187T/T189Q/L264R，L075Q/N094R/D130A/G156W/V187T，L075R/D130A/V154I/T189Q/L264R，L075R/D130A/V187T/T189Q/L264R，N011K/D027N/S058M/N233Q/P256T，N011K/D027Q/E056K/S058M/N233Q，N011K/D027Q/S058M/N233Q/P256T，N011K/D027S/E056K/N233Q/P256T，N011K/D027S/I090F/N233Q/P256T，N011K/G023K/D111A/G156W/L264R，P029E/N033D/E045F/L075D/T189D，Q004D/D027N/E056K/S058M/P256T，Q004D/D027N/I090F/N233Q/P256T，Q004D/D027Q/E056K/S058M/P256T，Q004D/N011K/D027N/N233Q/P256T，S058M/D130A/D137Q/G163P/L264R，S058M/D137Q/G163P/L227M/L264R，V077I/D130A/V154I/G156W/V187N，A018K/D027S/N033D/E045F/N073R/L075D，A018K/D027S/N033D/N073R/L075D/T189D，A018K/G023K/K024A/N094R/D130A/V154I，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023K/L075Q/D130A/G156W/V187N，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023K/L075R/N094R/D130A/V187N，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/A049V/L075Q/V077I/G156W，A018K/G023Q/A049V/V077I/V187T/T189D，A018K/G023Q/E045F/A049V/G156W/V187T，A018K/G023Q/E045F/G156W/V187T/T189D，A018K/G023Q/E045F/L075Q/G156C/V187T，A018K/G023Q/E045F/L075Q/G156W/V187T，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/G023Q/L075Q/G156W/V187T/T189D，A018K/G023Q/L075Q/V077I/N094R/V154I，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/N094R/V154I/V187N，A018K/K024A/L075R/D130A/V187N/T189Q，A018K/L075Q/D111A/V154I/G156W/V187N，A018K/L075Q/D111A/V154I/T189Q/L264R，A018K/L075Q/D130A/V187N/T189Q/L264R，A018K/N094R/D111A/D130A/V154I/V187N，A018K/P029E/N033D/E045F/L075D/T189D，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/F051T/L075Q/D130A/V187H/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D027S/P029E/N033D/E045F/N073R/L075D，D027S/P029E/N033D/E045F/N073R/T189D，D027S/P029E/N033D/N073R/L075D/T189D，D048Q/S058M/D130A/D137Q/G163P/L264R，D048Q/S058M/D137Q/G163P/N233Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/F051T/G091Q/D130A/V187H/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/N094R/D130A/G156W，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/D027S/N094R/V154I/G156W/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/K024A/L075Q/D130A/V154I/G156W，G023Q/K024A/V077I/D130A/V154I/G156W，G023Q/L075Q/N094R/D111A/G156W/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/N094R/D130A/V154I/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，K024A/L075Q/V154I/V187T/T189Q/L264R，L075Q/D111A/D130A/V154I/G156W/T189Q，L075Q/D130A/V154I/G156W/V187N/L264R，L075Q/N094R/D111A/G156W/T189Q/L264R，L075R/D130A/V154I/G156W/V187N/L264R，N011K/D027N/E056K/I090F/N233Q/P256T，N011K/D027S/E056K/I090F/N233Q/P256T，Q004D/N011K/D027N/S058M/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，S058M/D130A/D137Q/G163P/N233Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/D111A/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/K024A/L075Q/D130A/G156W/V187N，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/K024A/L075R/V154I/G156W/V187Q，A018K/G023K/K024A/N094R/V154I/V187T/L264R，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/E045F/L075Q/G156W/V187T/T189D，A018K/G023Q/K024A/L075Q/V077I/N094R/V154I，A018K/G023Q/K024A/L075R/N094R/G156W/V187N，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，A018K/L075Q/N094R/D111A/V154I/V187T/L264R，D027E/D048Q/S058M/D130A/G163P/L227M/L264R，D027Q/F051T/L075G/D130A/V187H/I252Q/L264R，D027S/E056K/L075R/D111A/G156W/V187N/L264R，D027S/F051T/L075G/G091Q/V187T/I252Q/L264R，D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，E056K/L075Q/D111A/D130A/G156W/V187N/T189Q，G023K/D027Q/F051T/L075G/D130A/V187H/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023K/F051T/E056K/L075R/D130A/V187T/I252Q，G023K/K024A/L075Q/D111A/D130A/T189Q/L264R，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023K/L075Q/D111A/D130A/V154I/G156W/L264R，G023K/L075Q/N094R/D130A/V154I/V187N/L264R，G023Q/E056K/L075Q/D111A/G156W/V187N/L264R，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/L075Q/D111A/D130A/V154I/G156W/L264R，G023Q/L075Q/D130A/V154I/V187N/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，K024A/L075Q/D111A/D130A/V154I/G156W/L264R，K024A/L075R/D130A/V154I/G156W/T189Q/L264R，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，N011K/D027S/D111A/D130A/V154I/V187N/T189Q，N011K/E056K/L075Q/D111A/D130A/V154I/L264R，N011K/G023Q/L075Q/V154I/V187N/T189Q/L264R，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/L075R/V077I/N094R/D130A/G156W/V187Q，A018K/G023Q/E045F/L075Q/V077I/G156W/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/D027S/F051T/E056K/L075R/D130A/V187T/L264R，G023K/E056K/L075Q/D130A/V154I/V187N/T189Q/L264R，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/V187N，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，G023Q/L075Q/N094R/D130A/V154I/G156W/T189Q/L264R，K024A/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，A018K/G023K/K024A/L075Q/N094R/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023K/L075Q/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，G023Q/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，N011K/G023Q/D027S/E056K/L075R/D130A/V154I/G156W/T189Q/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T，N011K/G023Q/D027S/L075Q/N094R/D111A/V154I/G156W/V187N/T189Q/L264R
表3-32：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且pNPB/pNPP比活性比值增大的TLL变体。
A018K/L075D，D027S/N033D，D027S/P256T，D130A/L264R，D130A/V187N，E045F/N073R，K024A/V154I，N094R/G156W，P029E/N033D，A018K/A049V/V187T，A018K/E045F/N073R，A018K/E045F/V187T，A018K/K024A/V154I，A018K/L075D/T189D，D027E/S058M/G163P，D027N/S058M/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/S058M/P256T，D130A/V187T/L264R，E056K/D130A/L264R，G023K/D130A/V187T，G023Q/A049V/T189D，G023Q/D130A/G156W，G023Q/K024A/G156W，K024A/D130A/V154I，L075G/D130A/V187H，L075Q/V187N/L264R，L075R/D130A/L264R，L075R/D130A/V187T，L075R/V187T/L264R，N011K/N233Q/P256T，N094R/D130A/V187T，P029E/N033D/E045F，V187N/T189Q/L264R，A018K/D111A/G156W/T189Q，A018K/E045F/A049V/G156W，A018K/E045F/L075D/T189D，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/V187T，A018K/L075Q/V077I/N094R，A018K/P029E/N073R/L075D，D027E/D137Q/G163P/L227M，D027E/S058M/G163P/L264R，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/E045F/L075D/T189D，D027S/E045F/N073R/T189D，D027S/N033D/E045F/N073R，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/N073R，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/G163P/N233Q/L264R，D111A/D130A/V154I/G156W，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，E056K/D130A/V187T/L264R，G023K/D027Q/F051T/L075Q，G023K/D130A/G156W/T189Q，G023K/K024A/V154I/V187N，G023K/L075R/D130A/L264R，G023Q/A049V/V077I/G156W，G023Q/K024A/D130A/G156W，G023Q/K024A/V077I/G156W，L075G/G091Q/V187N/L264R，L075R/D130A/V187T/L264R，N011K/D027Q/S058M/P256T，N011K/D027S/N233Q/P256T，N033D/E045F/N073R/T189A，P029E/N073R/L075D/T189D，Q004D/D027N/S058M/P256T，S058M/D137Q/G163P/N233Q，S058M/L075Q/G091Q/I252Q，A018K/D027S/E045F/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/G023K/D130A/V154I/G156W，A018K/G023K/K024A/D130A/G156W，A018K/G023Q/K024A/D130A/V187Q，A018K/G023Q/V077I/D130A/G156W，A018K/K024A/N094R/D130A/V187N，A018K/K024A/N094R/G156W/V187T，A018K/P029E/N033D/N073R/L075D，D027E/S058M/D137Q/G163P/N233Q，D027E/S058M/G163P/L227M/L264R，D048Q/S058M/D130A/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/G163P/L227M/L264R，D048Q/S058M/G163P/N233Q/L264R，D130A/D137Q/G163P/N233Q/L264R，E045F/A049V/L075Q/V187T/T189D，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/K024A/D111A/D130A/V187T，G023K/N094R/G156W/V187N/T189Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075Q/V077I/D130A/G156W，N011K/D027N/S058M/N233Q/P256T，N011K/D027S/I090F/N233Q/P256T，N011K/G023K/D111A/G156W/L264R，Q004D/N011K/D027N/N233Q/P256T，S058M/D130A/D137Q/G163P/L264R，S058M/D137Q/G163P/L227M/L264R，V077I/D130A/V154I/G156W/V187N，A018K/D027S/N033D/E045F/N073R/L075D，A018K/D027S/N033D/N073R/L075D/T189D，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L264R，D048Q/S058M/D137Q/G163P/N233Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/F051T/G091Q/D130A/V187H/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/D027S/N094R/V154I/G156W/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/K024A/V077I/D130A/V154I/G156W，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/N094R/D130A/V154I/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，N011K/D027S/E056K/I090F/N233Q/P256T，Q004D/N011K/D027N/S058M/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，S058M/D130A/D137Q/G163P/N233Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/D111A/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，D027E/D048Q/S058M/D130A/G163P/L227M/L264R，D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，N011K/G023Q/L075Q/V154I/V187N/T189Q/L264R，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V1541/G156W/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135E/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T
表3-33：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且pNPB/pNPP≥最大比活性比值的50％的TLL变体。
A018K/L075D，D027S/P256T，E045F/N073R，K024A/V154I，N094R/G156W，A018K/E045F/N073R，A018K/L075D/T189D，D027S/N033D/T189D，E056K/D130A/L264R，G023Q/A049V/T189D，K024A/D130A/V154I，L075R/D130A/L264R，L075R/D130A/V187T，N094R/D130A/V187T，P029E/N033D/E045F，A018K/E045F/A049V/G156W，A018K/E045F/L075D/T189D，A018K/P029E/N073R/L075D，D027Q/E056K/N233Q/P256T，D027S/E045F/L075D/T189D，D027S/E045F/N073R/T189D，D027S/N033D/E045F/N073R，D027S/N033D/L075D/T189D，D027S/P029E/E045F/N073R，D027S/P029E/N033D/L075D，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，G023K/D027Q/F051T/L075Q，G023K/L075R/D130A/L264R，G023Q/A049V/V077I/G156W，L075G/G091Q/V187N/L264R，N011K/D027S/N233Q/P256T，N033D/E045F/N073R/T189A，P029E/N073R/L075D/T189D，S058M/D137Q/G163P/N233Q，A018K/D027S/E045F/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/G023Q/V077I/D130A/G156W，A018K/K024A/N094R/D130A/V187N，A018K/P029E/N033D/N073R/L075D，E045F/A049V/L075Q/V187T/T189D，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/K024A/D111A/D130A/V187T，G023K/N094R/G156W/V187N/T189Q，G023Q/L075Q/V077I/D130A/G156W，N011K/D027S/I090F/N233Q/P256T，Q004D/N011K/D027N/N233Q/P256T，V077I/D130A/V154I/G156W/V187N，A018K/D027S/N033D/E045F/N073R/L075D，A018K/D027S/N033D/N073R/L075D/T189D，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/K024A/L075R/N094R/G156W，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027S/E056K/S058M/V187T/I252Q/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023Q/D027S/N094R/V154I/G156W/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/K024A/V077I/D130A/V154I/G156W，K024A/L075Q/V077I/N094R/D130A/V187N，Q004D/N011K/D027N/S058M/N233Q/P256T，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，N011K/G023Q/L075Q/V154I/V187N/T189Q/L264R，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D表3-34：下面示出，相比参考序列SEQIDNO：2，pNPB/pNPP比活性比值增大的TLL变体。
A018K/L075D，D027N/E056K，D027S/N033D，D027S/P256T，D130A/V187N，E045F/N073R，K024A/L075Q，K024A/V154I，L075Q/D130A，N094R/G156W，P029E/N033D，A018K/E045F/N073R，A018K/E045F/V187T，A018K/G023K/G156W，A018K/K024A/V154I，A018K/L075D/T189D，A049V/V187T/T189D，D027E/S058M/G163P，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/S058M/P256T，D130A/V187T/L264R，E045F/L075D/T189D，E056K/D130A/L264R，G023K/D130A/V187T，G023Q/A049V/T189D，G023Q/K024A/G156W，G023Q/L075Q/V187T，K024A/D130A/V154I，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/G156W/V187N，L075R/D130A/L264R，L075R/D130A/V187T，L075R/V187T/L264R，N011K/D027S/S058M，N073R/L075D/T189D，N094R/D130A/V187T，P029E/N033D/E045F，A018K/D027S/N033D/L075D，A018K/D111A/G156W/T189Q，A018K/E045F/A049V/G156W，A018K/E045F/L075D/T189D，A018K/E045F/L075Q/T189D，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/V187T，A018K/G023Q/E045F/T189D，A018K/G023Q/E045F/V187T，A018K/G023Q/L075R/D130A，A018K/G023Q/V077I/V187T，A018K/L075Q/G156W/V187T，A018K/L075Q/N094R/D130A，A018K/L075Q/V077I/N094R，A018K/P029E/N073R/L075D，A018K/V154I/G156W/V187T，D027E/D137Q/G163P/L227M，D027N/E056K/N233Q/P256T，D027N/E056K/S058M/P256T，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/E045F/L075D/T189D，D027S/E045F/N073R/T189D，D027S/E056K/D111A/V154I，D027S/E056K/I090F/P256T，D027S/N033D/E045F/N073R，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/L075D，D027S/P029E/E045F/N073R，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，G023K/D027Q/F051T/L075Q，G023K/D130A/G156W/T189Q，G023K/K024A/V154I/V187N，G023K/L075Q/G156W/V187N，G023K/L075R/D130A/L264R，G023Q/A049V/V077I/G156W，G023Q/D027S/D111A/G156W，G023Q/K024A/L075R/V154I，G023Q/V154I/G156W/V187N，K024A/L075R/G156W/V187N，L075G/G091Q/V187N/L264R，L075Q/D130A/G156W/V187N，L075Q/V077I/G156W/V187N，L075Q/V187N/T189Q/L264R，N011K/D027N/E056K/S058M，N011K/D027Q/S058M/P256T，N011K/D027S/E056K/P256T，N011K/D027S/N233Q/P256T，N011K/E056K/N233Q/P256T，N033D/E045F/N073R/T189A，P029E/E045F/N073R/L075D，P029E/N033D/L075D/T189D，P029E/N073R/L075D/T189D，Q004D/D027N/S058M/P256T，Q004D/D027S/I090F/P256T，S058M/D137Q/G163P/N233Q，A018K/A049V/L075Q/V187T/T189D，A018K/D027S/E045F/L075D/T189D，A018K/D027S/N033D/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/E045F/A049V/L075Q/V187T，A018K/G023K/D130A/V154I/G156W，A018K/G023K/K024A/L075R/N094R，A018K/G023Q/A049V/L075Q/G156W，A018K/G023Q/A049V/L075Q/V187T，A018K/G023Q/E045F/A049V/V187T，A018K/G023Q/E045F/L075Q/V187T，A018K/G023Q/K024A/D130A/V154I，A018K/G023Q/K024A/D130A/V187Q，A018K/G023Q/L075Q/G156W/V187T，A018K/G023Q/L075Q/V187T/T189D，A018K/G023Q/V077I/D130A/G156W，A018K/G023Q/V077I/V187T/T189D，A018K/K024A/L075Q/D130A/L264R，A018K/K024A/L075Q/V154I/T189D，A018K/K024A/N094R/D130A/V187N，A018K/L075Q/D111A/D130A/V187T，A018K/L075Q/N094R/G156W/V187N，A018K/P029E/E045F/N073R/L075D，A018K/P029E/N033D/N073R/L075D，A018K/V077I/G156W/V187T/T189D，D027E/S058M/D137Q/G163P/N233Q，D027Q/E056K/I090F/N233Q/P256T，D027Q/E056K/S058M/N233Q/P256T，D027S/P029E/N033D/E045F/N073R，D048Q/S058M/D130A/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/G163P/L227M/L264R，D048Q/S058M/G163P/N233Q/L264R，D130A/G163P/L227M/N233Q/L264R，E045F/A049V/L075Q/V187T/T189D，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/K024A/D111A/D130A/V187T，G023K/K024A/L075R/D130A/G156W，G023K/L075Q/D111A/D130A/V187T，G023K/L075Q/D130A/V154I/T189Q，G023K/N094R/G156W/V187N/T189Q，G023Q/A049V/L075Q/G156W/V187T，G023Q/E045F/A049V/G156W/T189D，G023Q/E045F/A049V/L075Q/V077I，G023Q/K024A/L075Q/G156W/V187Q，G023Q/K024A/L075R/D130A/V154I，G023Q/K024A/L075R/V077I/D130A，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/D111A/V154I/V187T，K024A/L075R/G156W/V187N/T189Q，L075Q/D130A/V187T/T189Q/L264R，L075Q/N094R/D130A/G156W/V187T，L075R/D130A/V154I/T189Q/L264R，N011K/D027Q/E056K/S058M/N233Q，N011K/D027Q/S058M/N233Q/P256T，N011K/D027S/E056K/N233Q/P256T，N011K/D027S/I090F/N233Q/P256T，N011K/G023K/D111A/G156W/L264R，P029E/N033D/E045F/L075D/T189D，Q004D/D027N/E056K/S058M/P256T，Q004D/D027N/I090F/N233Q/P256T，Q004D/D027Q/E056K/S058M/P256T，Q004D/N011K/D027N/N233Q/P256T，S058M/D130A/D137Q/G163P/L264R，S058M/D137Q/G163P/L227M/L264R，V077I/D130A/V154I/G156W/V187N，A018K/D027S/N033D/E045F/N073R/L075D，A018K/D027S/N033D/N073R/L075D/T189D，A018K/G023K/K024A/N094R/D130A/V154I，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023K/L075Q/D130A/G156W/V187N，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023K/L075R/N094R/D130A/V187N，A018K/G023Q/A049V/L075Q/V077I/G156W，A018K/G023Q/A049V/V077I/V187T/T189D，A018K/G023Q/E045F/A049V/G156W/V187T，A018K/G023Q/E045F/G156W/V187T/T189D，A018K/G023Q/E045F/L075Q/G156C/V187T，A018K/G023Q/E045F/L075Q/G156W/V187T，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/G023Q/L075Q/G156W/V187T/T189D，A018K/G023Q/L075Q/V077I/N094R/V154I，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/N094R/V154I/V187N，A018K/K024A/L075R/D130A/V187N/T189Q，A018K/L075Q/D111A/V154I/G156W/V187N，A018K/L075Q/D111A/V154I/T189Q/L264R，A018K/L075Q/D130A/V187N/T189Q/L264R，A018K/N094R/D111A/D130A/V154I/V187N，A018K/P029E/N033D/E045F/L075D/T189D，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/F051T/L075Q/D130A/V187H/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D027S/P029E/N033D/E045F/N073R/L075D，D027S/P029E/N033D/E045F/N073R/T189D，D027S/P029E/N033D/N073R/L075D/T189D，D048Q/S058M/D130A/D137Q/G163P/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/N094R/D130A/G156W，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/D027S/N094R/V154I/G156W/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/K024A/L075Q/D130A/V154I/G156W，G023Q/K024A/V077I/D130A/V154I/G156W，G023Q/L075Q/N094R/D111A/G156W/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/N094R/D130A/V154I/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，L075Q/D111A/D130A/V154I/G156W/T189Q，L075Q/D130A/V154I/G156W/V187N/L264R，L075Q/N094R/D111A/G156W/T189Q/L264R，L075R/D130A/V154I/G156W/V187N/L264R，N011K/D027N/E056K/I090F/N233Q/P256T，Q004D/N011K/D027N/S058M/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，S058M/D130A/D137Q/G163P/N233Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/D130A/G156W/V187N，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/K024A/L075R/V154I/G156W/V187Q，A018K/G023K/K024A/N094R/V154I/V187T/L264R，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/E045F/L075Q/G156W/V187T/T189D，A018K/G023Q/K024A/L075Q/V077I/N094R/V154I，A018K/G023Q/K024A/L075R/N094R/G156W/V187N，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/L075Q/N094R/D111A/V154I/V187T/L264R，D027S/E056K/L075R/D111A/G156W/V187N/L264R，D027S/F051T/L075G/G091Q/V187T/I252Q/L264R，D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，E056K/L075Q/D111A/D130A/G156W/V187N/T189Q，G023K/D027Q/F051T/L075G/D130A/V187H/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023K/F051T/E056K/L075R/D130A/V187T/I252Q，G023K/K024A/L075Q/D111A/D130A/T189Q/L264R，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023K/L075Q/D111A/D130A/V154I/G156W/L264R，G023K/L075Q/N094R/D130A/V154I/V187N/L264R，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/L075Q/D111A/D130A/V154I/G156W/L264R，G023Q/L075Q/D130A/V154I/V187N/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，K024A/L075Q/D111A/D130A/V154I/G156W/L264R，K024A/L075R/D130A/V154I/G156W/T189Q/L264R，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，N011K/D027S/D111A/D130A/V154I/V187N/T189Q，N011K/E056K/L075Q/D111A/D130A/V154I/L264R，N011K/G023Q/L075Q/V154I/V187N/T189Q/L264R，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/L075R/V077I/N094R/D130A/G156W/V187Q，A018K/G023Q/E045F/L075Q/V077I/G156W/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/D027S/F051T/E056K/L075R/D130A/V187T/L264R，G023K/E056K/L075Q/D130A/V154I/V187N/T189Q/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/V187N，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，G023Q/L075Q/N094R/D130A/V154I/G156W/T189Q/L264R，K024A/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，A018K/G023K/K024A/L075Q/N094R/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023K/L075Q/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，G023Q/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，N011K/G023K/L075R/D111A/D130A/V154I/5/187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，N011K/G023Q/D027S/E056K/L075R/D130A/V154I/G156W/T189Q/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，N011K/G023Q/D027S/L075Q/N094R/D111A/V154I/G156W/V187N/T189Q/L264R
表3-35：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比参考序列SEQIDNO：2而言pNPB/pNPP比活性比值增大的TLL变体
A018K/L075D，D027S/N033D，D027S/P256T，D130A/V187N，E045F/N073R，K024A/V154I，N094R/G156W，P029E/N033D，A018K/E045F/N073R，A018K/E045F/V187T，A018K/K024A/V154I，A018K/L075D/T189D，D027E/S058M/G163P，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/S058M/P256T，D130A/V187T/L264R，E056K/D130A/L264R，G023K/D130A/V187T，G023Q/A049V/T189D，G023Q/K024A/G156W，K024A/D130A/V154I，L075G/D130A/V187H，L075R/D130A/L264R，L075R/D130A/V187T，L075R/V187T/L264R，N094R/D130A/V187T，P029E/N033D/E045F，A018K/D111A/G156W/T189Q，A018K/E045F/A049V/G156W，A018K/E045F/L075D/T189D，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/V187T，A018K/L075Q/V077I/N094R，A018K/P029E/N073R/L075D，D027E/D137Q/G163P/L227M，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/E045F/L075D/T189D，D027S/E045F/N073R/T189D，D027S/N033D/E045F/N073R，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/N073R，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，G023K/D027Q/F051T/L075Q，G023K/D130A/G156W/T189Q，G023K/K024A/V154I/V187N，G023K/L075R/D130A/L264R，G023Q/A049V/V077I/G156W，L075G/G091Q/V187N/L264R，N011K/D027Q/S058M/P256T，N011K/D027S/N233Q/P256T，N033D/E045F/N073R/T189A，P029E/N073R/L075D/T189D，Q004D/D027N/S058M/P256T，S058M/D137Q/G163P/N233Q，A018K/D027S/E045F/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/G023K/D130A/V154I/G156W，A018K/G023Q/K024A/D130A/V187Q，A018K/G023Q/V077I/D130A/G156W，A018K/K024A/N094R/D130A/V187N，A018K/P029E/N033D/N073R/L075D，D027E/S058M/D137Q/G163P/N233Q，D048Q/S058M/D130A/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/G163P/L227M/L264R，D048Q/S058M/G163P/N233Q/L264R，E045F/A049V/L075Q/V187T/T189D，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/K024A/D111A/D130A/V187T，G023K/N094R/G156W/V187N/T189Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075Q/V077I/D130A/G156W，N011K/D027S/I090F/N233Q/P256T，N011K/G023K/D111A/G156W/L264R，Q004D/N011K/D027N/N233Q/P256T，S058M/D130A/D137Q/G163P/L264R，S058M/D137Q/G163P/L227M/L264R，V077I/D130A/V154I/G156W/V187N，A018K/D027S/N033D/E045F/N073R/L075D，A018K/D027S/N033D/N073R/L075D/T189D，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/D027S/N094R/V154I/G156W/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/K024A/V077I/D130A/V154I/G156W，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/N094R/D130A/V154I/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，Q004D/N011K/D027N/S058M/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，S058M/D130A/D137Q/G163P/N233Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，N011K/G023Q/L075Q/V154I/V187N/T189Q/L264R，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D
表3-36：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比参考序列SEQIDNO：2而言pNPB/pNPP≥最大比活性比值的50％的TLL变体。
A018K/L075D，D027S/P256T，E045F/N073R，K024A/V154I，N094R/G156W，A018K/E045F/N073R，A018K/L075D/T189D，D027S/N033D/T189D，E056K/D130A/L264R，G023Q/A049V/T189D，K024A/D130A/V154I，L075R/D130A/L264R，L075R/D130A/V187T，N094R/D130A/V187T，P029E/N033D/E045F，A018K/E045F/A049V/G156W，A018K/E045F/L075D/T189D，A018K/P029E/N073R/L075D，D027Q/E056K/N233Q/P256T，D027S/E045F/L075D/T189D，D027S/E045F/N073R/T189D，D027S/N033D/E045F/N073R，D027S/N033D/L075D/T189D，D027S/P029E/E045F/N073R，D027S/P029E/N033D/L075D，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，G023K/D027Q/F051T/L075Q，G023K/L075R/D130A/L264R，G023Q/A049V/V077I/G156W，L075G/G091Q/V187N/L264R，N011K/D027S/N233Q/P256T，N033D/E045F/N073R/T189A，P029E/N073R/L075D/T189D，S058M/D137Q/G163P/N233Q，A018K/D027S/E045F/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/G023Q/V077I/D130A/G156W，A018K/K024A/N094R/D130A/V187N，A018K/P029E/N033D/N073R/L075D，E045F/A049V/L075Q/V187T/T189D，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/K024A/D111A/D130A/V187T，G023K/N094R/G156W/V187N/T189Q，G023Q/L075Q/V077I/D130A/G156W，N011K/D027S/I090F/N233Q/P256T，Q004D/N011K/D027N/N233Q/P256T，V077I/D130A/V154I/G156W/V187N，A018K/D027S/N033D/E045F/N073R/L075D，A018K/D027S/N033D/N073R/L075D/T189D，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/K024A/L075R/N094R/G156W，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027S/E056K/S058M/V187T/I252Q/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023Q/D027S/N094R/V154I/G156W/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/K024A/V077I/D130A/V154I/G156W，K024A/L075Q/V077I/N094R/D130A/V187N，Q004D/N011K/D027N/S058M/N233Q/P256T，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，N011K/G023Q/L075Q/V154I/V187N/T189Q/L264R，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D
实施例4
TLL组合变体的热稳定性
按实施例1所述的程序(热稳定性测定法)，分析测定了按实施例2所述方法生成的TLL变体的热稳定性。计算了变体相比TLL(SEQIDNO：1)或参考序列SEQIDNO：2的性能指数。
表4-1：下面示出，在热稳定性测定过程中，相比TLL(SEQIDNO：1)性能指数＞1的TLL变体。
A018K/D027S，A018K/L075D，A018K/L075Q，A018K/T189D，D027E/G163P，D027N/E056K，D027N/N233Q，D027S/N033D，D027S/P256T，D111A/L264R，D130A/L264R，D130A/T189Q，D130A/V187N，D130A/V187T，E045F/N073R，G023K/D130A，G023K/L264R，G023Q/L075Q，K024A/L075Q，K024A/V154I，L075Q/D130A，L075Q/G156W，L075R/L264R，L227M/L264R，N011K/D027S，N073R/L075D，N094R/G156W，P029E/N033D，S058M/L264R，V187T/L264R，A018K/A049V/L075Q，A018K/A049V/V187T，A018K/D027S/E045F，A018K/D027S/N073R，A018K/D027S/T189D，A018K/E045F/T189D，A018K/E045F/V187T，A018K/G023K/G156W，A018K/G023K/L075Q，A018K/G023Q/L075Q，A018K/K024A/V154I，A018K/L075D/T189D，A018K/L075Q/G156W，A018K/L075Q/V187T，A018K/N033D/L075D，A018K/N033D/T189D，A018K/N073R/L075D，A018K/P029E/T189D，A018K/V154I/G156W，A049V/V187T/T189D，D027E/S058M/G163P，D027N/N233Q/P256T，D027N/S058M/P256T，D027Q/N233Q/P256T，D027Q/S058M/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/N073R/L075D，D027S/N233Q/P256T，D027S/S058M/P256T，D111A/D130A/L264R，D130A/T189Q/L264R，D130A/V154I/G156W，D130A/V187N/L264R，D130A/V187T/L264R，E045F/L075D/T189D，E045F/N073R/L075D，E056K/D130A/L264R，E056K/D130A/T189Q，G023K/D130A/L264R，G023K/D130A/V187T，G023K/E056K/V187T，G023K/L075R/L264R，G023K/V187N/L264R，G023K/V187T/L264R，G023Q/A049V/T189D，G023Q/D111A/L264R，G023Q/D130A/G156W，G023Q/E056K/L075R，G023Q/K024A/G156W，G023Q/L075Q/V187T，G163P/L227M/L264R，K024A/D130A/V154I，K024A/L075Q/G156W，K024A/L075Q/V077I，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/D130A/V187T，L075Q/G156W/T189D，L075Q/G156W/V187N，L075Q/G156W/V187T，L075Q/N094R/V154I，L075Q/V077I/T189D，L075Q/V154I/V187T，L075Q/V187T/L264R，L075Q/V187T/T189D，L075R/D130A/L264R，L075R/D130A/V187T，L075R/V187N/L264R，L075R/V187T/L264R，N011K/D027S/S058M，N011K/N233Q/P256T，N033D/N073R/T189D，N073R/L075D/T189D，N094R/D130A/V187T，P029E/E045F/L075D，P029E/L075D/T189D，P029E/N033D/L075D，P029E/N073R/L075D，Q004D/D027S/P256T，Q004D/N233Q/P256T，S058M/L227M/L264R，V077I/D130A/V154I，V077I/V187A/T189D，V187N/T189Q/L264R，A018K/A049V/L075Q/T189D，A018K/A049V/L075Q/V187T，A018K/D027S/E045F/N073R，A018K/D027S/E045F/T189D，A018K/D027S/N033D/L075D，A018K/D027S/P029E/T189D，A018K/D111A/G156W/T189Q，A018K/D130A/G156W/V187T，A018K/E045F/A049V/G156W，A018K/E045F/A049V/V187T，A018K/E045F/L075D/T189D，A018K/E045F/L075Q/V187T，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/T189D，A018K/G023Q/A049V/V187T，A018K/G023Q/E045F/T189D，A018K/G023Q/E045F/V187T，A018K/G023Q/G156W/V187T，A018K/G023Q/L075Q/T189D，A018K/G023Q/L075Q/V077I，A018K/G023Q/L075Q/V187T，A018K/G023Q/L075R/D130A，A018K/G023Q/V077I/G156W，A018K/G023Q/V077I/V187T，A018K/G023Q/V187T/T189D，A018K/K024A/D130A/G156W，A018K/L075Q/G156W/T189D，A018K/L075Q/G156W/V187T，A018K/L075Q/N094R/D111A，A018K/L075Q/N094R/D130A，A018K/L075Q/N094R/V187Q，A018K/L075Q/V077I/N094R，A018K/L075Q/V187T/T189D，A018K/N033D/L075D/T189D，A018K/N073R/L075D/T189D，A018K/P029E/N033D/L075D，A018K/P029E/N033D/T189D，A018K/P029E/N073R/L075D，A049V/L075Q/V187T/T189D，D027E/D048Q/G163P/L264R，D027E/D130A/N233Q/L264R，D027E/D137Q/G163P/L227M，D027E/D137Q/L227M/L264R，D027E/G163P/L227M/L264R，D027E/L227M/N233Q/L264R，D027E/S058M/G163P/L264R，D027N/E056K/N233Q/P256T，D027N/E056K/S058M/P256T，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/E056K/D111A/V154I，D027S/N033D/E045F/N073R，D027S/N033D/L075D/T189D，D027S/N033D/N073R/L075D，D027S/N033D/N073R/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D027S/P029E/N033D/T189D，D048Q/D130A/G163P/L264R，D048Q/G163P/N233Q/L264R，D111A/D130A/V154I/G156W，D111A/D130A/V154I/L264R，D130A/G163P/L227M/L264R，D130A/G163P/N233Q/L264R，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，E056K/D130A/V187N/L264R，E056K/D130A/V187T/L264R，E056K/L075Q/V187N/L264R，F051T/D130A/I252Q/L264R，F051T/L075G/I252Q/L264R，G023K/D027Q/F051T/L075Q，G023K/D027S/L075Q/L264R，G023K/D111A/D130A/L264R，G023K/D130A/G156W/T189Q，G023K/D130A/V187N/L264R，G023K/D130A/V187T/L264R，G023K/D130A/V187T/T189Q，G023K/E056K/D130A/L264R，G023K/E056K/L075R/L264R，G023K/K024A/V154I/V187N，G023K/L075Q/D130A/V187N，G023K/L075Q/G156W/L264R，G023K/L075Q/G156W/V187N，G023K/L075R/D130A/L264R，G023K/L075R/V187T/L264R，G023Q/A049V/L075Q/T189D，G023Q/A049V/L075Q/V077I，G023Q/A049V/V077I/G156W，G023Q/D027S/D111A/G156W，G023Q/E045F/A049V/T189D，G023Q/E045F/A049V/V187T，G023Q/E045F/L075Q/G156W，G023Q/K024A/D130A/G156W，G023Q/K024A/L075R/G156W，G023Q/K024A/L075R/V154I，G023Q/K024A/V077I/G156W，G023Q/L075Q/D130A/L264R，G023Q/L075Q/G156W/V187N，G023Q/L075Q/G156W/V187T，G023Q/V154I/G156W/V187N，G091Q/V187T/I252Q/L264R，K024A/D130A/V154I/V187T，K024A/L075Q/D130A/G156W，K024A/L075Q/D130A/V154I，K024A/L075Q/V187T/T189Q，K024A/L075R/G156W/V187N，L075G/D130A/V187T/I252Q，L075G/G091Q/V187N/L264R，L075Q/D130A/G156W/V187N，L075Q/D130A/V154I/G156W，L075Q/G156W/V187T/L264R，L075Q/V077I/D130A/V187Q，L075Q/V077I/G156W/V187N，L075Q/V077I/N094R/G156W，L075Q/V077I/V154I/V187Q，L075Q/V077I/V187T/T189D，L075R/D111A/V154I/G156W，L075R/D130A/V154I/L264R，L075R/D130A/V187T/L264R，N011K/D027N/E056K/S058M，N011K/D027Q/S058M/P256T，N011K/D027S/E056K/P256T，N011K/D027S/N233Q/P256T，N011K/E056K/N233Q/P256T，N011K/G023K/L075Q/D111A，N011K/S058M/N233Q/P256T，P029E/E045F/L075D/T189D，P029E/E045F/N0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94R/D130A/V154I/V187N/L264R，G023K/L075Q/N094R/V154I/G156W/V187T/L264R，G023K/L075R/N094R/V154I/G156W/V187N/L264R，G023Q/D027S/L075Q/V154I/G156W/V187N/L264R，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/E045F/L075Q/V077I/G156W/V187T/T189D，G023Q/E056K/L075Q/D111A/G156W/V187N/L264R，G023Q/E056K/L075Q/D130A/V154I/G156W/T189Q，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/L075R/G091Q/V187N/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/K024A/L075Q/V077I/D130A/G156W/V187N，G023Q/K024A/L075Q/V077I/D130A/V154I/G156W，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/L075Q/D111A/D130A/V154I/G156W/L264R，G023Q/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/L075Q/D130A/V154I/V187N/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，G023Q/L075Q/V077I/N094R/D130A/G156W/V187Q，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，K024A/L075Q/D111A/D130A/V154I/G156W/L264R，K024A/L075Q/D111A/D130A/V187N/T189Q/L264R，K024A/L075R/D130A/V154I/G156W/T189Q/L264R，L075Q/D111A/D130A/G156W/V187N/T189Q/L264R，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，N011K/D027S/D111A/D130A/V154I/V187N/T189Q，N011K/E056K/L075Q/D111A/D130A/V154I/L264R，N011K/E056K/L075Q/D130A/V154I/V187N/T189Q，N011K/G023K/L075Q/D111A/D130A/V154I/V187N，N011K/G023Q/L075Q/V154I/V187N/T189Q/L264R，Q004D/N011K/D027Q/E056K/S058M/N233Q/P256T，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/D130Y/V154I/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023K/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/G023K/L075R/V077I/N094R/D130A/G156W/V187Q，A018K/G023Q/E045F/L075Q/V077I/G156W/V187T/T189D，A018K/G023Q/K024A/L075Q/V077I/D130A/G156W/V187Q，A018K/G023Q/K024A/L075R/D130A/V154I/G156W/V187N，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，A018K/G023Q/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/K024A/L075Q/V077I/N094R/D130A/V154I/G156W，A018K/K024A/L075R/D130A/V154I/G156W/V187T/L264R，A018K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/E056K/L075G/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/D027S/F051T/E056K/L075R/D130A/V187T/L264R，G023K/E056K/L075Q/D130A/V154I/V187N/T189Q/L264R，G023K/F051T/E056K/L075R/G091Q/D130A/V187T/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187T/L264R，G023K/K024A/L075R/N094R/V154I/G156W/V187T/T189Q，G023K/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，G023K/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/E056K/S058M/L075R/G091Q/V187N/I252Q，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/E056K/L075Q/D111A/V154I/V187N/T189Q/L264R，G023Q/E056K/L075Q/N094R/D111A/G156W/T189Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/K024A/L075Q/V077I/D130A/V154I/G156W/V187N，G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/V187N，G023Q/L075Q/D111A/D130A/V154I/G156W/V187N/L264R，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，K024A/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，N011K/G023K/D027S/L075Q/D111A/D130A/V154I/T189Q，N011K/G023K/D027S/L075R/V154I/G156W/V187N/T189Q，N011K/G023K/E056K/L075Q/D130A/V154I/T189Q/L264R，N011K/G023K/L075Q/D111A/G156W/V187N/T189Q/L264R，N011K/G023Q/D027S/L075Q/N094R/V154I/G156W/T189Q，N011K/G023Q/D027S/N094R/V154I/G156W/T189Q/L264R，N011K/G023Q/L075Q/D111A/D130A/V154I/V187N/L264R，N011K/G023Q/L075Q/N094R/V154I/G156W/T189Q/L264R，A018K/G023K/K024A/L075Q/N094R/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/V154I/G156W/V187N，A018K/G023K/K024A/L075Q/N094R/D130A/V187T/T189Q/L264K，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，A018K/G023Q/D027S/L075Q/V077I/R108K/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/G156W/V187H，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/P029E/S058M/V077I/R108K/H135F/G156W，A018K/G023Q/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W/V187N，A018K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V187N/T189Q/L264R，G023K/D027S/F051T/E056K/S058M/L075Q/G091Q/V187N/L264R，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/D027S/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023K/L075Q/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/F051T/E056K/L075G/G091Q/V187N/I252Q/L264R，G023Q/D027S/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，G023Q/D027S/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，N011K/G023K/D027S/E056K/L075R/D111A/G156W/V187N/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，N011K/G023Q/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/E045F/A049V/S058M/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/S058M/L075Q/V077I/R108K/H135F/G156W/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023Q/D027S/F051T/E056K/L075R/G091Q/D130A/V187H/I252Q/L264R，G023Q/D027S/F051T/E056K/S058M/L075R/G091Q/D130A/V187H/I252Q，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023K/L075R/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，N011K/G023Q/D027S/E056K/L075R/D130A/V154I/G156W/T189Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T，N011K/G023Q/D027S/L075Q/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D
表4-2：下面示出，相比TLL(SEQIDNO：1)，表达性能指数≥0.5，且在热稳定性测定中的性能指数＞1的TLL变体。
A018K/L075D，D027E/G163P，D027S/N033D，D027S/P256T，D111A/L264R，D130A/L264R，D130A/V187N，D130A/V187T，E045F/N073R，G023K/L264R，K024A/V154I，L075R/L264R，L227M/L264R，N094R/G156W，P029E/N033D，S058M/L264R，V187T/L264R，A018K/A049V/V187T，A018K/E045F/V187T，A018K/K024A/V154I，A018K/L075D/T189D，A018K/N033D/T189D，A018K/P029E/T189D，A018K/V154I/G156W，D027E/S058M/G163P，D027N/S058M/P256T，D027Q/S058M/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/S058M/P256T，D111A/D130A/L264R，D130A/V154I/G156W，D130A/V187N/L264R，D130A/V187T/L264R，E056K/D130A/L264R，G023K/D130A/L264R，G023K/D130A/V187T，G023K/E056K/V187T，G023K/L075R/L264R，G023K/V187N/L264R，G023K/V187T/L264R，G023Q/A049V/T189D，G023Q/D130A/G156W，G023Q/K024A/G156W，G163P/L227M/L264R，K024A/D130A/V154I，L075G/D130A/V187H，L075Q/V187T/L264R，L075R/D130A/L264R，L075R/D130A/V187T，L075R/V187N/L264R，L075R/V187T/L264R，N011K/N233Q/P256T，N094R/D130A/V187T，Q004D/D027S/P256T，V187N/T189Q/L264R，A018K/D027S/P029E/T189D，A018K/D111A/G156W/T189Q，A018K/D130A/G156W/V187T，A018K/E045F/A049V/G156W，A018K/E045F/L075D/T189D，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/V187T，A018K/G023Q/V077I/G156W，A018K/K024A/D130A/G156W，A018K/L075Q/V077I/N094R，A018K/N033D/L075D/T189D，A018K/P029E/N073R/L075D，D027E/D048Q/G163P/L264R，D027E/D130A/N233Q/L264R，D027E/D137Q/G163P/L227M，D027E/D137Q/L227M/L264R，D027E/G163P/L227M/L264R，D027E/L227M/N233Q/L264R，D027E/S058M/G163P/L264R，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/N033D/E045F/N073R，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D048Q/G163P/N233Q/L264R，D111A/D130A/V154I/G156W，D130A/G163P/L227M/L264R，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，E056K/D130A/V187N/L264R，E056K/D130A/V187T/L264R，F051T/D130A/I252Q/L264R，F051T/L075G/I252Q/L264R，G023K/D027Q/F051T/L075Q，G023K/D130A/G156W/T189Q，G023K/D130A/V187N/L264R，G023K/D130A/V187T/L264R，G023K/D130A/V187T/T189Q，G023K/E056K/L075R/L264R，G023K/K024A/V154I/V187N，G023K/L075R/D130A/L264R，G023K/L075R/V187T/L264R，G023Q/A049V/V077I/G156W，G023Q/K024A/D130A/G156W，G023Q/K024A/L075R/G156W，G023Q/K024A/V077I/G156W，G023Q/L075Q/D130A/L264R，G091Q/V187T/I252Q/L264R，K024A/D130A/V154I/V187T，L075G/D130A/V187T/I252Q，L075G/G091Q/V187N/L264R，L075R/D130A/V187T/L264R，N011K/D027Q/S058M/P256T，N011K/D027S/N233Q/P256T，P029E/N073R/L075D/T189D，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/N011K/D027S/P256T，S058M/D137Q/G163P/N233Q，S058M/G163P/L227M/L264R，S058M/G163P/N233Q/L264R，S058M/L075Q/G091Q/I252Q，S058M/L075Q/I252Q/L264R，A018K/D027S/E045F/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/D130A/G156W/V187N/L264R，A018K/G023K/D130A/V154I/G156W，A018K/G023K/K024A/D130A/G156W，A018K/G023K/K024A/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/K024A/D130A/V187Q，A018K/G023Q/K024A/L075Q/G156W，A018K/K024A/L075R/D130A/V154I，A018K/K024A/N094R/D130A/V187N，A018K/K024A/N094R/G156W/V187T，A018K/P029E/N033D/N073R/L075D，D027E/D048Q/D137Q/L227M/L264R，D027E/D048Q/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L264R，D027E/D130A/G163P/L227M/L264R，D027E/D130A/G163P/N233Q/L264R，D027E/D137Q/G163P/N233Q/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L264R，D027E/S058M/D137Q/G163P/N233Q，D027E/S058M/G163P/L227M/L264R，D027Q/F051T/L075Q/D130A/L264R，D027S/E056K/D111A/V187N/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D048Q/D130A/G163P/L227M/L264R，D048Q/D137Q/G163P/L227M/L264R，D048Q/D137Q/G163P/N233Q/L264R，D048Q/S058M/D130A/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/D137Q/N233Q/L264R，D048Q/S058M/G163P/L227M/L264R，D048Q/S058M/G163P/N233Q/L264R，D130A/D137Q/G163P/L227M/L264R，D130A/D137Q/G163P/N233Q/L264R，D130A/V154I/G156W/V187N/T189Q，E045F/A049V/L075Q/V187T/T189D，E056K/L075G/V187N/I252Q/L264R，E056K/L075R/D130A/V187T/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/G091Q/D130A/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187N/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187N/L264R，G023K/E056K/L075R/V187T/L264R，G023K/K024A/D111A/D130A/V187T，G023K/K024A/D111A/G156W/T189Q，G023K/L075Q/V077I/D130A/G156W，G023K/L075Q/V187H/I252Q/L264R，G023K/L075R/D130A/V187N/L264R，G023K/L075R/D130A/V187T/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075G/G091Q/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/V154I/G156W/V187Q，L075G/D130A/V187T/I252Q/L264R，N011K/D027N/S058M/N233Q/P256T，N011K/D027S/I090F/N233Q/P256T，N011K/G023K/D111A/G156W/L264R，Q004D/N011K/D027N/N233Q/P256T，S058M/D130A/D137Q/G163P/L264R，S058M/D130A/G163P/L227M/L264R，S058M/D137Q/G163P/L227M/L264R，S058M/D137Q/G163P/L227M/N233Q，S058M/G163P/L227M/N233Q/L264R，V077I/D130A/V154I/G156W/V187N，A018K/D027S/N033D/E045F/N073R/L075D，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/G023Q/K024A/V077I/V154I/V187N，A018K/G023Q/K024A/V154I/G156W/V187Q，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D048Q/D130A/D137H/G163P/L264R，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D130A/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/N233Q/L264R，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/D137Q/G163P/N233Q/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L227M/L264R，D027E/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/F051T/G091Q/D130A/I252Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/D130A/G163P/L227M/N233Q/L264R，D048Q/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L264R，D048Q/S058M/D130A/G163P/L227M/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/L264R，D048Q/S058M/D137Q/G163P/N233Q/L264R，D048Q/S058M/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/E056K/L075R/D130A/V187T/L264R，G023K/F051T/G091Q/D130A/V187H/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/L075G/D130A/V187T/I252Q/L264R，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/L075Q/D130A/V187T/I252Q，G023Q/D027S/N094R/V154I/G156W/T189Q，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/K024A/V077I/D130A/V154I/G156W，G023Q/L075G/G091Q/D130A/I252Q/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/N094R/D130A/V154I/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/D027S/E056K/I090F/N233Q/P256T，Q004D/N011K/D027N/I090F/N233Q/P256T，Q004D/N011K/D027N/S058M/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，S058M/D130A/G163P/L227M/N233Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/D111A/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/D130A/V154I/G156W/V187T/L264R，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/K024A/N094R/D130A/G156W/V187T/L264R，D027E/D048Q/D130A/D137Q/G163P/N233Q/L264R，D027E/D048Q/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/D048Q/S058M/D130A/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/L264R，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/D137Q/L227M/N233Q/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/S058M/G091Q/V187N/I252Q，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，N011K/G023Q/L075Q/V154I/V187N/T189Q/L264R，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T
表4-3：下面示出，相比TLL(SEQIDNO：1)，表达性能指数≥0.5，且在热稳定性测定中性能指数≥最大PI值的50％的TLL变体。
E045F/N073R，A018K/E045F/V187T，A018K/V154I/G156W，D111A/D130A/L264R，V187N/T189Q/L264R，A018K/D111A/G156W/T189Q，A018K/D130A/G156W/V187T，A018K/E045F/A049V/G156W，A018K/G023Q/V077I/G156W，A018K/D027S/E045F/L075D/T189D，A018K/D130A/G156W/V187N/L264R，A018K/G023K/D130A/V154I/G156W，A018K/G023Q/K024A/L075Q/G156W，D027S/E056K/D111A/V187N/L264R，G023K/K024A/D111A/D130A/V187T，G023K/L075Q/V187H/I252Q/L264R，N011K/G023K/D111A/G156W/L264R，S058M/D137Q/G163P/L227M/N233Q，S058M/G163P/L227M/N233Q/L264R，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/K024A/V154I/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，D048Q/S058M/D130A/D137Q/G163P/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/L075G/D130A/V187T/I252Q/L264R，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，A018K/D111A/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/K024A/N094R/D130A/G156W/V187T/L264R，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/I075Q/V077I/N094R/D130A/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T
表4-4：下面示出，在热稳定性测定中，相比参考序列SEQIDNO：2，性能指数＞1的TLL变体。
A018K/D027S，A018K/L075D，A018K/L075Q，A018K/T189D，D027E/G163P，D027S/N033D，D027S/P256T，D111A/L264R，D130A/L264R，D130A/T189Q，D130A/V187N，D130A/V187T，E045F/N073R，F051T/L075R，G023K/D130A，G023K/L264R，G023Q/L075Q，K024A/L075Q，K024A/V154I，L075Q/D130A，L075Q/G156W，L075R/L264R，L227M/L264R，N011K/D027S，N073R/L075D，P029E/N033D，S058M/L264R，V187T/L264R，A018K/A049V/L075Q，A018K/A049V/V187T，A018K/D027S/N073R，A018K/D027S/T189D，A018K/E045F/V187T，A018K/G023K/G156W，A018K/G023K/L075Q，A018K/G023Q/L075Q，A018K/K024A/V154I，A018K/L075D/T189D，A018K/L075Q/G156W，A018K/L075Q/V187T，A018K/N033D/L075D，A018K/N033D/T189D，A018K/N073R/L075D，A018K/P029E/T189D，A018K/V154I/G156W，D027E/S058M/G163P，D027N/N233Q/P256T，D027N/S058M/P256T，D027Q/N233Q/P256T，D027Q/S058M/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/N073R/L075D，D027S/N233Q/P256T，D027S/S058M/P256T，D111A/D130A/L264R，D130A/T189Q/L264R，D130A/V154I/G156W，D130A/V187N/L264R，D130A/V187T/L264R，E045F/L075D/T189D，E045F/N073R/L075D，E056K/D130A/L264R，E056K/D130A/T189Q，G023K/D130A/L264R，G023K/D130A/V187T，G023K/E056K/V187T，G023K/L075R/L264R，G023K/V187N/L264R，G023K/V187T/L264R，G023Q/A049V/T189D，G023Q/D111A/L264R，G023Q/D130A/G156W，G023Q/E056K/L075R，G023Q/K024A/G156W，G023Q/L075Q/V187T，G163P/L227M/L264R，K024A/D130A/V154I，K024A/L075Q/G156W，K024A/L075Q/V077I，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/D130A/V187T，L075Q/G156W/T189D，L075Q/G156W/V187N，L075Q/G156W/V187T，L075Q/N094R/V154I，L075Q/V077I/T189D，L075Q/V154I/V187T，L075Q/V187T/L264R，L075Q/V187T/T189D，L075R/D130A/L264R，L075R/D130A/V187T，L075R/V187N/L264R，L075R/V187T/L264R，N011K/D027S/S058M，N011K/N233Q/P256T，N073R/L075D/T189D，N094R/D130A/V187T，P029E/E045F/L075D，P029E/L075D/T189D，P029E/N033D/L075D，P029E/N073R/L075D，Q004D/D027S/P256T，Q004D/N233Q/P256T，S058M/L227M/L264R，V077I/D130A/V154I，V187N/T189Q/L264R，A018K/A049V/L075Q/T189D，A018K/A049V/L075Q/V187T，A018K/D027S/E045F/N073R，A018K/D027S/E045F/T189D，A018K/D027S/N033D/L075D，A018K/D027S/P029E/T189D，A018K/D111A/G156W/T189Q，A018K/D130A/G156W/V187T，A018K/E045F/A049V/G156W，A018K/E045F/A049V/V187T，A018K/E045F/L075D/T189D，A018K/E045F/L075Q/V187T，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/T189D，A018K/G023Q/A049V/V187T，A018K/G023Q/E045F/T189D，A018K/G023Q/E045F/V187T，A018K/G023Q/G156W/V187T，A018K/G023Q/L075Q/T189D，A018K/G023Q/L075Q/V077I，A018K/G023Q/L075Q/V187T，A018K/G023Q/L075R/D130A，A018K/G023Q/V077I/G156W，A018K/G023Q/V077I/V187T，A018K/G023Q/V187T/T189D，A018K/K024A/D130A/G156W，A018K/L075Q/G156W/T189D，A018K/L075Q/G156W/V187T，A018K/L075Q/N094R/D111A，A018K/L075Q/N094R/D130A，A018K/L075Q/N094R/V187Q，A018K/L075Q/V077I/N094R，A018K/N033D/L075D/T189D，A018K/N073R/L075D/T189D，A018K/P029E/N033D/T189D，A018K/P029E/N073R/L075D，A049V/L075Q/V187T/T189D，D027E/D048Q/G163P/L264R，D027E/D130A/N233Q/L264R，D027E/D137Q/G163P/L227M，D027E/D137Q/L227M/L264R，D027E/G163P/L227M/L264R，D027E/L227M/N233Q/L264R，D027E/S058M/G163P/L264R，D027N/E056K/N233Q/P256T，D027N/E056K/S058M/P256T，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/D130A/I252Q/L264R，D027S/E056K/D111A/V154I，D027S/N033D/E045F/N073R，D027S/N033D/L075D/T189D，D027S/N033D/N073R/L075D，D027S/N033D/N073R/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D027S/P029E/N033D/T189D，D048Q/D130A/G163P/L264R，D048Q/G163P/N233Q/L264R，D111A/D130A/V154I/G156W，D111A/D130A/V154I/L264R，D130A/G163P/L227M/L264R，D130A/G163P/N233Q/L264R，E045F/A049V/G156W/V187T，E056K/D130A/V187N/L264R，E056K/D130A/V187T/L264R，E056K/L075Q/V187N/L264R，F051T/D130A/I252Q/L264R，F051T/L075G/I252Q/L264R，G023K/D027Q/F051T/L075Q，G023K/D027S/L075Q/L264R，G023K/D111A/D130A/L264R，G023K/D130A/G156W/T189Q，G023K/D130A/V187N/L264R，G023K/D130A/V187T/L264R，G023K/D130A/V187T/T189Q，G023K/E056K/D130A/L264R，G023K/E056K/L075R/L264R，G023K/K024A/V154I/V187N，G023K/L075Q/D130A/V187N，G023K/L075Q/G156W/L264R，G023K/L075Q/G156W/V187N，G023K/L075R/D130A/L264R，G023K/L075R/V187T/L264R，G023Q/A049V/L075Q/T189D，G023Q/A049V/L075Q/V077I，G023Q/A049V/V077I/G156W，G023Q/D027S/D111A/G156W，G023Q/E045F/A049V/T189D，G023Q/E045F/A049V/V187T，G023Q/E045F/L075Q/G156W，G023Q/K024A/L075R/G156W，G023Q/K024A/L075R/V154I，G023Q/K024A/V077I/G156W，G023Q/L075Q/D130A/L264R，G023Q/L075Q/G156W/V187N，G023Q/L075Q/G156W/V187T，G023Q/V154I/G156W/V187N，G091Q/V187T/I252Q/L264R，K024A/D130A/V154I/V187T，K024A/L075Q/D130A/G156W，K024A/L075Q/D130A/V154I，K024A/L075Q/V187T/T189Q，K024A/L075R/G156W/V187N，L075G/D130A/V187T/I252Q，L075G/G091Q/V187N/L264R，L075G/V187H/I252Q/L264R，L075Q/D130A/G156W/V187N，L075Q/D130A/V154I/G156W，L075Q/G156W/V187T/L264R，L075Q/V077I/D130A/V187Q，L075Q/V077I/G156W/V187N，L075Q/V077I/N094R/G156W，L075Q/V077I/V154I/V187Q，L075Q/V077I/V187T/T189D，L075R/D111A/V154I/G156W，L075R/D130A/V154I/L264R，L075R/D130A/V187T/L264R，N011K/D027N/E056K/S058M，N011K/D027Q/S058M/P256T，N011K/D027S/E056K/P256T，N011K/D027S/N233Q/P256T，N011K/E056K/N233Q/P256T，N011K/G023K/L075Q/D111A，P029E/E045F/N073R/L075D，P029E/N073R/L075D/T189D，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/N011K/D027N/P256T，Q004D/N011K/D027Q/N233Q，Q004D/N011K/D027S/P256T，S058M/D1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64R，K024A/L075Q/D111A/D130A/V154I/G156W/L264R，K024A/L075Q/D111A/D130A/V187N/T189Q/L264R，K024A/L075R/D130A/V154I/G156W/T189Q/L264R，L075Q/D111A/D130A/G156W/V187N/T189Q/L264R，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，N011K/E056K/L075Q/D111A/D130A/V154I/L264R，N011K/E056K/L075Q/D130A/V154I/V187N/T189Q，N011K/G023K/L075Q/D111A/D130A/V154I/V187N，Q004D/N011K/D027Q/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/D130Y/V154I/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023K/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/G023K/L075R/V077I/N094R/D130A/G156W/V187Q，A018K/G023Q/E045F/L075Q/V077I/G156W/V187T/T189D，A018K/G023Q/K024A/L075Q/V077I/D130A/G156W/V187Q，A018K/G023Q/K024A/L075R/D130A/V154I/G156W/V187N，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，A018K/G023Q/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/K024A/L075Q/V077I/N094R/D130A/V154I/G156W，A018K/K024A/L075R/D130A/V154I/G156W/V187T/L264R，A018K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/E056K/L075G/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/D027S/F051T/E056K/L075R/D130A/V187T/L264R，G023K/E056K/L075Q/D130A/V154I/V187N/T189Q/L264R，G023K/F051T/E056K/L075R/G091Q/D130A/V187T/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187T/L264R，G023K/K024A/L075R/N094R/V154I/G156W/V187T/T189Q，G023K/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，G023K/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/E056K/S058M/L075R/G091Q/V187N/I252Q，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/E056K/L075Q/N094R/D111A/G156W/T189Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/K024A/L075Q/V077I/D130A/V154I/G156W/V187N，G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/V187N，G023Q/L075Q/D111A/D130A/V154I/G156W/V187N/L264R，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，K024A/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，N011K/G023K/D027S/L075Q/D111A/D130A/V154I/T189Q，N011K/G023K/D027S/L075R/V154I/G156W/V187N/T189Q，N011K/G023K/E056K/L075Q/D130A/V154I/T189Q/L264R，N011K/G023K/L075Q/D111A/G156W/V187N/T189Q/L264R，N011K/G023Q/D027S/L075Q/N094R/V154I/G156W/T189Q，N011K/G023Q/D027S/N094R/V154I/G156W/T189Q/L264R，N011K/G023Q/L075Q/D111A/D130A/V154I/V187N/L264R，N011K/G023Q/L075Q/N094R/V154I/G156W/T189Q/L264R，A018K/G023K/K024A/L075Q/N094R/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/V154I/G156W/V187N，A018K/G023K/K024A/L075Q/N094R/D130A/V187T/T189Q/L264K，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，A018K/G023Q/D027S/L075Q/V077I/R108K/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/G156W/V187H，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/P029E/S058M/V077I/R108K/H135F/G156W，A018K/G023Q/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W/V187N，A018K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V187N/T189Q/L264R，G023K/D027S/F051T/E056K/S058M/L075Q/G091Q/V187N/L264R，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/D027S/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023K/L075Q/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/F051T/E056K/L075G/G091Q/V187N/I252Q/L264R，G023Q/D027S/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，G023Q/D027S/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，N011K/G023K/D027S/E056K/L075R/D111A/G156W/V187N/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，N011K/G023Q/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/E045F/A049V/S058M/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/S058M/L075Q/V077I/R108K/H135F/G156W/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023Q/D027S/F051T/E056K/L075R/G091Q/D130A/V187H/I252Q/L264R，G023Q/D027S/F051T/E056K/S058M/L075R/G091Q/D130A/V187H/I252Q，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023K/L075R/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，N011K/G023Q/D027S/E056K/L075R/D130A/V154I/G156W/T189Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T，N011K/G023Q/D027S/L075Q/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D
表4-5：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比参考序列SEQIDNO：2在热稳定性测定过程中性能指数＞1的TLL变体。
A018K/L075D，D027E/G163P，D027S/N033D，D027S/P256T，D111A/L264R，D130A/L264R，D130A/V187N，D130A/V187T，E045F/N073R，F051T/L075R，G023K/L264R，K024A/V154I，L075R/L264R，L227M/L264R，P029E/N033D，S058M/L264R，V187T/L264R，A018K/A049V/V187T，A018K/E045F/V187T，A018K/K024A/V154I，A018K/L075D/T189D，A018K/N033D/T189D，A018K/P029E/T189D，A018K/V154I/G156W，D027E/S058M/G163P，D027N/S058M/P256T，D027Q/S058M/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/S058M/P256T，D111A/D130A/L264R，D130A/V154I/G156W，D130A/V187N/L264R，D130A/V187T/L264R，E056K/D130A/L264R，G023K/D130A/L264R，G023K/D130A/V187T，G023K/E056K/V187T，G023K/L075R/L264R，G023K/V187N/L264R，G023K/V187T/L264R，G023Q/A049V/T189D，G023Q/D130A/G156W，G023Q/K024A/G156W，G163P/L227M/L264R，K024A/D130A/V154I，L075G/D130A/V187H，L075Q/V187T/L264R，L075R/D130A/L264R，L075R/D130A/V187T，L075R/V187N/L264R，L075R/V187T/L264R，N011K/N233Q/P256T，N094R/D130A/V187T，Q004D/D027S/P256T，V187N/T189Q/L264R，A018K/D027S/P029E/T189D，A018K/D111A/G156W/T189Q，A018K/D130A/G156W/V187T，A018K/E045F/A049V/G156W，A018K/E045F/L075D/T189D，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/V187T，A018K/G023Q/V077I/G156W，A018K/K024A/D130A/G156W，A018K/L075Q/V077I/N094R，A018K/N033D/L075D/T189D，A018K/P029E/N073R/L075D，D027E/D048Q/G163P/L264R，D027E/D130A/N233Q/L264R，D027E/D137Q/G163P/L227M，D027E/D137Q/L227M/L264R，D027E/G163P/L227M/L264R，D027E/L227M/N233Q/L264R，D027E/S058M/G163P/L264R，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/D130A/I252Q/L264R，D027S/N033D/E045F/N073R，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D048Q/G163P/N233Q/L264R，D111A/D130A/V154I/G156W，D130A/G163P/L227M/L264R，E045F/A049V/G156W/V187T，E056K/D130A/V187N/L264R，E056K/D130A/V187T/L264R，F051T/D130A/I252Q/L264R，F051T/L075G/I252Q/L264R，G023K/D027Q/F051T/L075Q，G023K/D130A/G156W/T189Q，G023K/D130A/V187N/L264R，G023K/D130A/V187T/L264R，G023K/D130A/V187T/T189Q，G023K/E056K/L075R/L264R，G023K/K024A/V154I/V187N，G023K/L075R/D130A/L264R，G023K/L075R/V187T/L264R，G023Q/A049V/V077I/G156W，G023Q/K024A/L075R/G156W，G023Q/K024A/V077I/G156W，G023Q/L075Q/D130A/L264R，G091Q/V187T/I252Q/L264R，K024A/D130A/V154I/V187T，L075G/D130A/V187T/I252Q，L075G/G091Q/V187N/L264R，L075G/V187H/I252Q/L264R，L075R/D130A/V187T/L264R，N011K/D027Q/S058M/P256T，N011K/D027S/N233Q/P256T，P029E/N073R/L075D/T189D，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/N011K/D027S/P256T，S058M/D137Q/G163P/N233Q，S058M/G163P/L227M/L264R，S058M/G163P/N233Q/L264R，S058M/L075Q/G091Q/I252Q，S058M/L075Q/I252Q/L264R，A018K/D027S/E045F/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/D130A/G156W/V187N/L264R，A018K/G023K/D130A/V154I/G156W，A018K/G023K/K024A/D130A/G156W，A018K/G023K/K024A/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/K024A/D130A/V187Q，A018K/G023Q/K024A/L075Q/G156W，A018K/K024A/L075R/D130A/V154I，A018K/K024A/N094R/D130A/V187N，A018K/K024A/N094R/G156W/V187T，A018K/P029E/N033D/N073R/L075D，D027E/D048Q/D137Q/L227M/L264R，D027E/D048Q/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L264R，D027E/D130A/G163P/L227M/L264R，D027E/D130A/G163P/N233Q/L264R，D027E/D137Q/G163P/N233Q/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L264R，D027E/S058M/D137Q/G163P/N233Q，D027E/S058M/G163P/L227M/L264R，D027Q/F051T/L075Q/D130A/L264R，D027S/E056K/D111A/V187N/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D048Q/D130A/G163P/L227M/L264R，D048Q/D137Q/G163P/L227M/L264R，D048Q/D137Q/G163P/N233Q/L264R，D048Q/S058M/D130A/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/D137Q/N233Q/L264R，D048Q/S058M/G163P/L227M/L264R，D048Q/S058M/G163P/N233Q/L264R，D130A/D137Q/G163P/L227M/L264R，D130A/D137Q/G163P/N233Q/L264R，D130A/V154I/G156W/V187N/T189Q，E045F/A049V/L075Q/V187T/T189D，E056K/L075G/V187N/I252Q/L264R，E056K/L075R/D130A/V187T/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/G091Q/D130A/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/L264R，F051T/L075Q/V187N/I252Q/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187N/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/V187N/L264R，G023K/E056K/L075R/V187T/L264R，G023K/K024A/D111A/D130A/V187T，G023K/K024A/D111A/G156W/T189Q，G023K/L075Q/V077I/D130A/G156W，G023K/L075Q/V187H/I252Q/L264R，G023K/L075R/D130A/V187N/L264R，G023K/L075R/D130A/V187T/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075G/G091Q/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/V154I/G156W/V187Q，L075G/D130A/V187T/I252Q/L264R，N011K/D027N/S058M/N233Q/P256T，N011K/G023K/D111A/G156W/L264R，Q004D/N011K/D027N/N233Q/P256T，S058M/D130A/D137Q/G163P/L264R，S058M/D130A/G163P/L227M/L264R，S058M/D137Q/G163P/L227M/L264R，S058M/D137Q/G163P/L227M/N233Q，S058M/G163P/L227M/N233Q/L264R，V077I/D130A/V154I/G156W/V187N，A018K/D027S/N033D/E045F/N073R/L075D，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/G023Q/K024A/V077I/V154I/V187N，A018K/G023Q/K024A/V154I/G156W/V187Q，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D048Q/D130A/D137H/G163P/L264R，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D130A/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/N233Q/L264R，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/D137Q/G163P/N233Q/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L227M/L264R，D027E/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/F051T/G091Q/D130A/I252Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/D130A/G163P/L227M/N233Q/L264R，D048Q/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L264R，D048Q/S058M/D130A/G163P/L227M/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/L264R，D048Q/S058M/D137Q/G163P/N233Q/L264R，D048Q/S058M/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/E056K/L075R/D130A/V187T/L264R，G023K/F051T/G091Q/D130A/V187H/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/L075G/D130A/V187T/I252Q/L264R，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/L075Q/D130A/V187T/I252Q，G023Q/D027S/N094R/V154I/G156W/T189Q，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/K024A/V077I/D130A/V154I/G156W，G023Q/L075G/G091Q/D130A/I252Q/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/N094R/D130A/V154I/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，L075Q/G091Q/D130A/V187H/I252Q/L264R，Q004D/N011K/D027N/I090F/N233Q/P256T，Q004D/N011K/D027N/S058M/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，S058M/D130A/G163P/L227M/N233Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/D111A/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/D130A/V154I/G156W/V187T/L264R，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/K024A/N094R/D130A/G156W/V187T/L264R，D027E/D048Q/D130A/D137Q/G163P/N233Q/L264R，D027E/D048Q/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/D048Q/S058M/D130A/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/L264R，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/D137Q/L227M/N233Q/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/S058M/G091Q/V187N/I252Q，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T
表4-6：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比参考序列SEQIDNO：2在热稳定性测定中性能指数≥最大PI值的50％的TLL变体。
A018K/L075D，D027E/G163P，E045F/N073R，L227M/L264R，A018K/E045F/V187T，A018K/L075D/T189D，G163P/L227M/L264R，L075R/V187N/L264R，A018K/E045F/A049V/G156W，D027E/D048Q/G163P/L264R，D027E/D130A/N233Q/L264R，D027E/G163P/L227M/L264R，D027E/L227M/N233Q/L264R，D027E/S058M/G163P/L264R，D027S/P029E/N033D/E045F，D048Q/G163P/N233Q/L264R，D130A/G163P/L227M/L264R，F051T/D130A/I252Q/L264R，F051T/L075G/I252Q/L264R，G023K/D130A/V187T/L264R，G023Q/L075Q/D130A/L264R，L075G/G091Q/V187N/L264R，S058M/G163P/L227M/L264R，S058M/G163P/N233Q/L264R，A018K/D027S/E045F/L075D/T189D，D027E/D130A/D137Q/G163P/L264R，D027E/D130A/G163P/L227M/L264R，D027E/D130A/G163P/N233Q/L264R，D027E/D137Q/G163P/N233Q/L264R，D027E/S058M/D130A/G163P/L264R，D027E/S058M/G163P/L227M/L264R，D027S/E056K/D111A/V187N/L264R，D048Q/D130A/G163P/L227M/L264R，D048Q/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/N233Q/L264R，D048Q/S058M/D137Q/N233Q/L264R，D048Q/S058M/G163P/L227M/L264R，D048Q/S058M/G163P/N233Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/G091Q/D130A/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/E056K/L075R/V187N/L264R，G023K/L075Q/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/L075Q/D130A/I252Q/L264R，L075G/D130A/V187T/I252Q/L264R，S058M/D130A/G163P/L227M/L264R，S058M/D137Q/G163P/L227M/L264R，S058M/D137Q/G163P/L227M/N233Q，D027E/D048Q/D130A/D137H/G163P/L264R，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/N233Q/L264R，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/D137Q/G163P/N233Q/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L227M/L264R，D027E/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/F051T/G091Q/D130A/I252Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D027S/F051T/L075Q/D130A/I252Q/L264R，D048Q/D130A/G163P/L227M/N233Q/L264R，D048Q/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233Q/L264R，D048Q/S058M/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/L075G/D130A/V187T/I252Q/L264R，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/F051T/L075Q/D130A/V187H/I252Q，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，S058M/D130A/G163P/L227M/N233Q/L264R，D027E/D048Q/D130A/D137Q/G163P/N233Q/L264R，D027E/D048Q/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D130A/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/L264R，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/D137Q/L227M/N233Q/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，D027E/D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D
实施例5
TLL组合变体的洗涤剂稳定性
按实施例1所述的程序(洗涤剂稳定性测定法)，分析测定了按实施例2所述生成的TLL变体的洗涤剂稳定性。计算了变体相比TLL(SEQIDNO：1)或参考序列SEQIDNO：2的性能指数。
表5-1：下面示出，相比TLL(SEQIDNO：1)，在洗涤剂稳定性测定中性能指数＞1的TLL变体。
A018K/D027S，A018K/L075Q，A018K/T189D，D027E/G163P，D027N/E056K，D027N/N233Q，D027S/N033D，D027S/P256T，D111A/L264R，D130A/T189Q，D130A/V187N，G023K/D130A，G023Q/L075Q，K024A/L075Q，K024A/V154I，L075Q/D130A，L075Q/G156W，N011K/D027S，N073R/L075D，N094R/G156W，P029E/N033D，A018K/A049V/L075Q，A018K/A049V/V187T，A018K/D027S/E045F，A018K/D027S/N073R，A018K/D027S/T189D，A018K/E045F/T189D，A018K/E045F/V187T，A018K/G023K/G156W，A018K/G023K/L075Q，A018K/G023Q/L075Q，A018K/K024A/V154I，A018K/L075Q/G156W，A018K/L075Q/V187T，A018K/N033D/T189D，A018K/N073R/L075D，A018K/V154I/G156W，A049V/L075Q/T189D，A049V/V187T/T189D，D027E/S058M/G163P，D027N/N233Q/P256T，D027N/S058M/P256T，D027Q/N233Q/P256T，D027Q/S058M/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/N073R/L075D，D027S/N233Q/P256T，D027S/S058M/P256T，D111A/D130A/L264R，D130A/T189Q/L264R，D130A/V154I/G156W，D130A/V187T/L264R，E045F/L075D/T189D，E045F/N073R/L075D，E056K/D130A/L264R，G023K/D130A/L264R，G023K/D130A/V187T，G023K/V187N/L264R，G023K/V187T/L264R，G023Q/A049V/T189D，G023Q/D111A/L264R，G023Q/D130A/G156W，G023Q/E056K/L075R，G023Q/K024A/G156W，G023Q/L075Q/V187T，I090F/N233Q/P256T，K024A/D130A/V154I，K024A/L075Q/G156W，K024A/L075Q/V077I，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/D130A/V187T，L075Q/G156W/T189D，L075Q/G156W/V187N，L075Q/G156W/V187T，L075Q/V077I/T189D，L075Q/V154I/V187T，L075Q/V187N/L264R，L075Q/V187T/T189D，L075R/D130A/V187T，N011K/D027S/S058M，N011K/N233Q/P256T，N033D/E045F/L075D，N033D/N073R/T189D，N073R/L075D/T189D，N094R/D130A/V187T，P029E/E045F/L075D，P029E/L075D/T189D，P029E/N033D/E045F，P029E/N033D/L075D，P029E/N073R/L075D，Q004D/D027S/P256T，Q004D/S058M/N233Q，V077I/D130A/V154I，V077I/V187A/T189D，V187N/T189Q/L264R，A018K/A049V/L075Q/T189D，A018K/A049V/L075Q/V187T，A018K/D027S/L075D/T189D，A018K/D027S/N033D/L075D，A018K/D027S/P029E/T189D，A018K/D111A/G156W/T189Q，A018K/D130A/G156W/V187T，A018K/E045F/A049V/G156W，A018K/E045F/A049V/V187T，A018K/E045F/L075Q/T189D，A018K/E045F/L075Q/V187T，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/T189D，A018K/G023Q/A049V/V187T，A018K/G023Q/E045F/T189D，A018K/G023Q/E045F/V187T，A018K/G023Q/G156W/V187T，A018K/G023Q/L075Q/T189D，A018K/G023Q/L075Q/V077I，A018K/G023Q/L075Q/V187T，A018K/G023Q/L075R/D130A，A018K/G023Q/V077I/G156W，A018K/G023Q/V077I/V187T，A018K/G023Q/V187T/T189D，A018K/K024A/D130A/G156W，A018K/L075Q/G156W/T189D，A018K/L075Q/G156W/V187T，A018K/L075Q/N094R/D111A，A018K/L075Q/N094R/D130A，A018K/L075Q/N094R/V187Q，A018K/L075Q/V077I/N094R，A018K/N033D/L075D/T189D，A018K/N073R/L075D/T189D，A018K/P029E/N033D/L075D，A018K/P029E/N033D/T189D，A018K/V154I/G156W/V187T，A049V/L075Q/V077I/T189D，A049V/L075Q/V187T/T189D，D027E/D137Q/G163P/L227M，D027N/E056K/N233Q/P256T，D027N/E056K/S058M/P256T，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027Q/S058M/I090F/P256T，D027S/D130A/I252Q/L264R，D027S/E056K/D111A/V154I，D027S/E056K/I090F/P256T，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/L075D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/L075D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D027S/P029E/N033D/N073R，D027S/P029E/N033D/T189D，D111A/D130A/V154I/G156W，D111A/D130A/V154I/L264R，D111A/D130A/V187T/L264R，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，E056K/D130A/V187T/L264R，E056K/L075Q/V187N/L264R，G023K/D027Q/F051T/L075Q，G023K/D027S/L075Q/L264R，G023K/D111A/D130A/L264R，G023K/D130A/G156W/T189Q，G023K/D130A/V187N/L264R，G023K/D130A/V187T/L264R，G023K/D130A/V187T/T189Q，G023K/K024A/V154I/V187N，G023K/L075Q/D130A/V187N，G023K/L075Q/G156W/L264R，G023K/L075Q/G156W/V187N，G023K/L075R/D130A/L264R，G023K/L075R/V187T/L264R，G023Q/A049V/L075Q/T189D，G023Q/A049V/V077I/G156W，G023Q/E045F/A049V/T189D，G023Q/E045F/A049V/V187T，G023Q/E045F/L075Q/G156W，G023Q/E045F/L075Q/V187T，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/K024A/D130A/G156W，G023Q/K024A/L075R/G156W，G023Q/K024A/L075R/V154I，G023Q/K024A/V077I/G156W，G023Q/L075Q/D130A/L264R，G023Q/L075Q/G156W/V187N，G023Q/L075Q/G156W/V187T，G023Q/L075Q/V187T/L264R，G023Q/V154I/G156W/V187N，K024A/D130A/V154I/V187T，K024A/L075Q/D130A/G156W，K024A/L075Q/D130A/V154I，K024A/L075Q/V187T/T189Q，K024A/L075R/G156W/V187N，L075G/D130A/V187T/I252Q，L075Q/D130A/G156W/V187N，L075Q/D130A/V154I/G156W，L075Q/G156W/V187T/L264R，L075Q/V077I/D130A/V187Q，L075Q/V077I/G156W/V187N，L075Q/V077I/N094R/G156W，L075Q/V077I/V154I/V187Q，L075Q/V077I/V187T/T189D，L075R/D111A/V154I/G156W，L075R/D130A/V154I/L264R，N011K/D027N/E056K/S058M，N011K/D027N/S058M/P256T，N011K/D027Q/S058M/P256T，N011K/D027S/E056K/P256T，N011K/D027S/N233Q/P256T，N011K/D027S/S058M/N233Q，N011K/E056K/N233Q/P256T，N011K/G023K/L075Q/D111A，N011K/S058M/N233Q/P256T，P029E/E045F/L075D/T189D，P029E/E045F/N073R/L075D，P029E/E045F/N073R/T189D，P029E/N033D/E045F/L075D，P029E/N033D/L075D/T189D，P029E/N073R/L075D/T189D，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/D027S/I090F/P256T，Q004D/N011K/D027N/P256T，Q004D/N011K/D027Q/N233Q，Q004D/N011K/D027S/P256T，Q004D/N011K/E056K/S058M，S058M/D137Q/G163P/N233Q，S058M/L075Q/G091Q/I252Q，A018K/A049V/L075Q/V187T/T189D，A018K/D027S/N033D/L075D/T189D，A018K/D027S/N033D/N073R/T189D，A018K/D027S/P029E/E045F/T189D，A018K/D027S/P029E/N0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18K/G023K/K024A/L075Q/N094R/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/V154I/G156W/V187N，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023Q/D027S/L075Q/V077I/R108K/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/G156W/V187H，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/P029E/S058M/V077I/R108K/H135F/G156W，A018K/G023Q/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W/V187N，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，G023K/D027S/F051T/E056K/S058M/L075Q/G091Q/V187N/L264R，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023K/L075Q/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/F051T/E056K/L075G/G091Q/V187N/I252Q/L264R，G023Q/D027S/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，N011K/G023K/D027S/E056K/L075R/D111A/G156W/V187N/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，N011K/G023Q/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/E045F/A049V/S058M/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/S058M/L075Q/V077I/R108K/H135F/G156W/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023Q/D027S/F051T/E056K/L075R/G091Q/D130A/V187H/I252Q/L264R，G023Q/D027S/F051T/E056K/S058M/L075R/G091Q/D130A/V187H/I252Q，G023Q/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023Q/D027S/E056K/L075R/D130A/V154I/G156W/T189Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T，N011K/G023Q/D027S/L075Q/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/N033D/E045F/A049V/S058M/N073S/K074S/L075Q/V077I/N101D/R108K/H135F/D137V/G156W/V187T/T189D
表5-2：下面示出，相比TLL(SEQIDNO：1)，表达性能指数≥0.5，且在洗涤剂稳定性测定中性能指数＞1的TLL变体。
D027E/G163P，D027S/N033D，D027S/P256T，D111A/L264R，D130A/V187N，K024A/V154I，N094R/G156W，P029E/N033D，A018K/A049V/V187T，A018K/E045F/V187T，A018K/K024A/V1541，A018K/N033D/T189D，A018K/V154I/G156W，D027E/S058M/G163P，D027N/S058M/P256T，D027Q/S058M/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/S058M/P256T，D111A/D130A/L264R，D130A/V154I/G156W，D130A/V187T/L264R，E056K/D130A/L264R，G023K/D130A/L264R，G023K/D130A/V187T，G023K/V187N/L264R，G023K/V187T/L264R，G023Q/A049V/T189D，G023Q/D130A/G156W，G023Q/K024A/G156W，I090F/N233Q/P256T，K024A/D130A/V154I，L075G/D130A/V187H，L075Q/V187N/L264R，L075R/D130A/V187T，N011K/N233Q/P256T，N094R/D130A/V187T，P029E/N033D/E045F，Q004D/D027S/P256T，Q004D/S058M/N233Q，V187N/T189Q/L264R，A018K/D027S/P029E/T189D，A018K/D111A/G156W/T189Q，A018K/D130A/G156W/V187T，A018K/E045F/A049V/G156W，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/V187T，A018K/G023Q/V077I/G156W，A018K/K024A/D130A/G156W，A018K/L075Q/V077I/N094R，A018K/N033D/L075D/T189D，D027E/D137Q/G163P/L227M，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/D130A/I252Q/L264R，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D111A/D130A/V154I/G156W，D111A/D130A/V187T/L264R，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，E056K/D130A/V187T/L264R，G023K/D027Q/F051T/L075Q，G023K/D130A/G156W/T189Q，G023K/D130A/V187N/L264R，G023K/D130A/V187T/L264R，G023K/D130A/V187T/T189Q，G023K/K024A/V154I/V187N，G023K/L075R/D130A/L264R，G023K/L075R/V187T/L264R，G023Q/A049V/V077I/G156W，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/K024A/D130A/G156W，G023Q/K024A/L075R/G156W，G023Q/K024A/V077I/G156W，G023Q/L075Q/D130A/L264R，G023Q/L075Q/V187T/L264R，K024A/D130A/V154I/V187T，L075G/D130A/V187T/I252Q，N011K/D027Q/S058M/P256T，N011K/D027S/N233Q/P256T，P029E/N073R/L075D/T189DQ004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/N011K/D027S/P256T，S058M/D137Q/G163P/N233Q，S058M/L075Q/G091Q/I252Q，A018K/D027S/P029E/N033D/L075D，A018K/G023K/D130A/V154I/G156W，A018K/G023K/K024A/D130A/G156W，A018K/G023K/K024A/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/K024A/D130A/V187Q，A018K/G023Q/K024A/L075Q/G156W，A018K/K024A/L075R/D130A/V154I，A018K/K024A/N094R/G156W/V187T，D027E/S058M/D137Q/G163P/N233Q，D027S/E056K/D111A/V187N/L264R，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D048Q/S058M/D130A/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/G163P/N233Q/L264R，D130A/V154I/G156W/V187N/T189Q，E045F/A049V/L075Q/V187T/T189D，E056K/L075R/D130A/V187T/L264R，F051T/L075G/G091Q/D130A/L264R，F051T/L075Q/G091Q/D130A/L264R，F051T/L075Q/V187N/I252Q/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187N/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187N/L264R，G023K/E056K/L075R/V187T/L264R，G023K/K024A/D111A/D130A/V187T，G023K/K024A/D111A/G156W/T189Q，G023K/L075Q/V077I/D130A/G156W，G023K/L075Q/V187H/I252Q/L264R，G023K/L075R/D130A/V187N/L264R，G023K/L075R/D130A/V187T/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075G/G091Q/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/V154I/G156W/V187Q，N011K/D027N/S058M/N233Q/P256T，N011K/D027S/I090F/N233Q/P256T，N011K/G023K/D111A/G156W/L264R，Q004D/N011K/D027N/N233Q/P256T，S058M/G163P/L227M/N233Q/L264R，V077I/D130A/V154I/G156W/V187N，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/G023Q/K024A/V077I/V154I/V187N，A018K/G023Q/K024A/V154I/G156W/V187Q，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/N094R/D111A/D130A/V154I/V187N，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/F051T/G091Q/D130A/I252Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L264R，D048Q/S058M/D137Q/G163P/L227M/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/E056K/L075R/D130A/V187T/L264R，G023K/F051T/G091Q/D130A/V187H/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/L075Q/D130A/V187T/I252Q，G023Q/D027S/N094R/V154I/G156W/T189Q，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/K024A/V077I/D130A/V154I/G156W，G023Q/L075G/G091Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/V187N/I252Q/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/N094R/D130A/V154I/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/D027S/E056K/I090F/N233Q/P256T，Q004D/N011K/D027N/I090F/N233Q/P256T，Q004D/N011K/D027N/S058M/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/D111A/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/D137Q/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/S058M/G091Q/V187N/I252Q，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，N011K/G023Q/L075Q/V154I/V187N/T189Q/L264R，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T
表5-3：下面示出，相比TLL(SEQIDNO：1)，表达性能指数≥0.5，且在洗涤剂稳定性测定中性能指数≥最大PI值的50％的TLL变体。
D027S/P256T，K024A/V154I，A018K/A049V/V187T，A018K/K024A/V154I，A018K/V154I/G156W，D027N/S058M/P256T，D027S/S058M/P256T，G023Q/A049V/T189D，L075Q/V187N/L264R，N011K/N233Q/P256T，P029E/N033D/E045F，Q004D/D027S/P256T，A018K/D111A/G156W/T189Q，A018K/D130A/G156W/V187T，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/V187T，A018K/G023Q/V077I/G156W，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/N033D/E045F/T189D，D111A/D130A/V154I/G156W，E045F/L075Q/G156W/V187T，G023K/D130A/G156W/T189Q，G023K/D130A/V187T/T189Q，G023K/K024A/V154I/V187N，G023Q/A049V/V077I/G156W，G023Q/K024A/L075R/G156W，K024A/D130A/V154I/V187T，N011K/D027Q/S058M/P256T，N011K/D027S/N233Q/P256T，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/N011K/D027S/P256T，A018K/G023K/D130A/V154I/G156W，A018K/G023K/K024A/D130A/G156W，A018K/G023K/K024A/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/K024A/L075Q/G156W，D027S/E056K/D111A/V187N/L264R，D130A/V154I/G156W/V187N/T189Q，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187T/L264R，G023K/K024A/D111A/D130A/V187T，G023K/K024A/D111A/G156W/T189Q，G023K/L075Q/V077I/D130A/G156W，G023K/L075Q/V187H/I252Q/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/V154I/G156W/V187Q，N011K/D027N/S058M/N233Q/P256T，N011K/D027S/I090F/N233Q/P256T，N011K/G023K/D111A/G156W/L264R，Q004D/N011K/D027N/N233Q/P256T，S058M/G163P/L227M/N233Q/L264R，V077I/D130A/V154I/G156W/V187N，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/K024A/V077I/V154I/V187N，A018K/G023Q/K024A/V154I/G156W/V187Q，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，D048Q/S058M/D130A/D137Q/G163P/L264R，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/V187T/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/D027S/N094R/V154I/G156W/T189Q，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/K024A/V077I/D130A/V154I/G156W，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/N094R/D130A/V154I/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，N011K/D027S/E056K/I090F/N233Q/P256T，Q004D/N011K/D027N/I090F/N233Q/P256T，Q004D/N011K/D027N/S058M/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，A018K/D111A/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，G023K/D027S/F051T/S058M/G091Q/V187N/I252Q，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/I075Q/V077I/N094R/D130A/G156W/V187N，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T
表5-4：下面示出，相比参考序列SEQIDNO：2，在洗涤剂稳定性测定中性能指数＞1的TLL变体。
A018K/D027S，A018K/L075D，A018K/L075Q，D027E/G163P，D027N/E056K，D027N/N233Q，D027S/N033D，D027S/P256T，D111A/L264R，D130A/L264R，G023K/D130A，G023Q/L075Q，K024A/L075Q，K024A/V154I，L075Q/D130A，L075Q/G156W，N073R/L075D，P029E/N033D，A018K/A049V/L075Q，A018K/A049V/V187T，A018K/D027S/N073R，A018K/D027S/T189D，A018K/E045F/T189D，A018K/G023K/G156W，A018K/G023K/L075Q，A018K/G023Q/L075Q，A018K/K024A/V154I，A018K/L075D/T189D，A018K/L075Q/G156W，A018K/L075Q/V187T，A018K/N033D/L075D，A018K/N033D/T189D，A018K/N073R/L075D，A018K/P029E/T189D，A018K/V154I/G156W，A049V/L075Q/T189D，A049V/V187T/T189D，D027E/S058M/G163P，D027N/N233Q/P256T，D027N/S058M/P256T，D027Q/N233Q/P256T，D027Q/S058M/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/N073R/L075D，D027S/N233Q/P256T，D027S/S058M/P256T，D130A/V187T/L264R，E045F/L075D/T189D，E045F/N073R/L075D，E056K/D130A/L264R，G023K/D130A/L264R，G023K/D130A/V187T，G023K/V187N/L264R，G023K/V187T/L264R，G023Q/A049V/T189D，G023Q/E056K/L075R，G023Q/L075Q/V187T，I090F/N233Q/P256T，K024A/L075Q/G156W，K024A/L075Q/V077I，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/D130A/V187T，L075Q/G156W/T189D，L075Q/G156W/V187N，L075Q/G156W/V187T，L075Q/V077I/T189D，L075Q/V154I/V187T，L075Q/V187N/L264R，L075Q/V187T/T189D，L075R/D130A/V187T，N011K/D027S/S058M，N011K/N233Q/P256T，N033D/E045F/L075D，N033D/N073R/T189D，N073R/L075D/T189D，N094R/D130A/V187T，P029E/E045F/L075D，P029E/L075D/T189D，P029E/N033D/E045F，P029E/N033D/L075D，P029E/N073R/L075D，Q004D/D027S/P256T，S058M/L227M/L264R，V077I/D130A/V154I，A018K/A049V/L075Q/T189D，A018K/D027S/L075D/T189D，A018K/D027S/N033D/L075D，A018K/D027S/P029E/T189D，A018K/D111A/G156W/T189Q，A018K/D130A/G156W/V187T，A018K/E045F/L075D/T189D，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/T189D，A018K/G023Q/A049V/V187T，A018K/G023Q/E045F/T189D，A018K/G023Q/E045F/V187T，A018K/G023Q/G156W/V187T，A018K/G023Q/L075Q/T189D，A018K/G023Q/L075Q/V077I，A018K/G023Q/L075Q/V187T，A018K/G023Q/L075R/D130A，A018K/G023Q/V077I/G156W，A018K/G023Q/V077I/V187T，A018K/G023Q/V187T/T189D，A018K/L075Q/G156W/T189D，A018K/L075Q/G156W/V187T，A018K/L075Q/N094R/D130A，A018K/N033D/L075D/T189D，A018K/N073R/L075D/T189D，A018K/P029E/N033D/L075D，A018K/P029E/N033D/T189D，A018K/V154I/G156W/V187T，A049V/L075Q/V077I/T189D，A049V/L075Q/V187T/T189D，D027E/D137Q/G163P/L227M，D027N/E056K/N233Q/P256T，D027N/E056K/S058M/P256T，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/D130A/I252Q/L264R，D027S/E056K/D111A/V154I，D027S/E056K/I090F/P256T，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/L075D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/L075D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D027S/P029E/N033D/N073R，D027S/P029E/N033D/T189D，D111A/D130A/V154I/G156W，D111A/D130A/V154I/L264R，E045F/L075Q/G156W/V187T，E056K/D130A/V187N/L264R，E056K/D130A/V187T/L264R，E056K/L075Q/VI87N/L264R，F051T/L075Q/I252Q/L264R，G023K/D027Q/F051T/L075Q，G023K/D027S/L075Q/L264R，G023K/D111A/D130A/L264R，G023K/D130A/G156W/T189Q，G023K/D130A/V187N/L264R，G023K/D130A/V187T/L264R，G023K/D130A/V187T/T189Q，G023K/K024A/V154I/V187N，G023K/L075Q/D130A/V187N，G023K/L075Q/G156W/L264R，G023K/L075Q/G156W/V187N，G023K/L075R/D130A/L264R，G023K/L075R/V187T/L264R，G023Q/A049V/L075Q/T189D，G023Q/A049V/V077I/G156W，G023Q/E045F/A049V/T189D，G023Q/E045F/L075Q/G156W，G023Q/E045F/L075Q/V187T，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/K024A/L075R/G156W，G023Q/K024A/L075R/V154I，G023Q/K024A/V077I/G156W，G023Q/L075Q/D130A/L264R，G023Q/L075Q/G156W/V187N，G023Q/L075Q/G156W/V187T，G023Q/L075Q/V187T/L264R，G023Q/V154I/G156W/V187N，K024A/D130A/V154I/V187T，K024A/L075Q/D130A/G156W，K024A/L075Q/D130A/V154I，K024A/L075Q/V187T/T189Q，K024A/L075R/G156W/V187N，L075G/D130A/V187T/I252Q，L075G/G091Q/V187N/L264R，L075Q/D130A/G156W/V187N，L075Q/D130A/V154I/G156W，L075Q/G156W/V187T/L264R，L075Q/V077I/D130A/V187Q，L075Q/V077I/G156W/V187N，L075Q/V077I/N094R/G156W，L075Q/V077I/V154I/V187Q，L075Q/V077I/V187T/T189D，L075R/D111A/V154I/G156W，L075R/D130A/V154I/L264R，N011K/D027N/E056K/S058M，N011K/D027Q/S058M/P256T，N011K/D027S/E056K/P256T，N011K/D027S/N233Q/P256T，N011K/D027S/S058M/N233Q，N011K/E056K/N233Q/P256T，N011K/G023K/L075Q/D111A，N011K/S058M/N233Q/P256T，P029E/E045F/L075D/T189D，P029E/E045F/N073R/L075D，P029E/N033D/E045F/L075D，P029E/N033D/L075D/T189D，P029E/N073R/L075D/T189D，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/D027S/I090F/P256T，Q004D/N011K/D027N/P256T，Q004D/N011K/D027S/P256T，S058M/D137Q/G163P/N233Q，S058M/G163P/L227M/L264R，S058M/G163P/N233Q/L264R，S058M/L075Q/G091Q/I252Q，S058M/L075Q/I252Q/L264R，A018K/A049V/L075Q/V187T/T189D，A018K/D027S/N033D/L075D/T189D，A018K/D027S/N033D/N073R/T189D，A018K/D027S/P029E/E045F/T189D，A018K/D027S/P029E/N033D/L075D，A018K/D027S/P029E/N073R/L075D，A018K/E045F/A049V/L075Q/T189D，A018K/G023K/D130A/V154I/G156W，A018K/G023K/K024A/D130A/G156W，A018K/G023K/K024A/D130A/V154I，A018K/G023K/K024A/L075R/N094R，A018K/G023K/L075Q/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/A049V/L075Q/G156W，A018K/G023Q/A049V/L075Q/V187T，A018K/G023Q/E045F/L075Q/V187T，A018K/G023Q/K024A/D130A/V154I，A018K/G023Q/K024A/L075Q/G156W，A018K/G023Q/L075Q/G156W/V187T，A018K/G023Q/L0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I252Q，G023K/K024A/L075Q/D111A/D130A/T189Q/L264R，G023K/K024A/L075Q/D111A/V154I/G156W/V187N，G023K/K024A/L075Q/D130A/V187T/T189Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N，G023K/K024A/L075Q/V077I/D130A/G156W/V187Q，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023K/L075Q/D111A/D130A/V154I/G156W/L264R，G023K/L075Q/D111A/V154I/G156W/T189Q/L264R，G023K/L075Q/D130A/G156W/V187N/T189Q/L264R，G023K/L075Q/N094R/V154I/G156W/V187T/L264R，G023K/L075R/N094R/V154I/G156W/V187N/L264R，G023Q/D027S/L075Q/V154I/G156W/V187N/L264R，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/E045F/L075Q/V077I/G156W/V187T/T189D，G023Q/E056K/L075Q/D111A/G156W/V187N/L264R，G023Q/E056K/L075Q/D130A/V154I/G156W/T189Q，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/L075R/G091Q/V187N/I252Q/L264R，G023Q/K024A/L075Q/V077I/D130A/G156W/V187N，G023Q/K024A/L075Q/V077I/D130A/V154I/G156W，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/L075Q/D111A/D130A/V154I/G156W/L264R，G023Q/L075Q/D130A/V154I/V187N/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，G023Q/L075Q/V077I/N094R/D130A/G156W/V187Q，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，K024A/L075Q/D111A/D130A/V154I/G156W/L264R，K024A/L075R/D130A/V154I/G156W/T189Q/L264R，L075Q/D111A/D130A/G156W/V187N/T189Q/L264R，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，N011K/E056K/L075Q/D111A/D130A/V154I/L264R，N011K/E056K/L075Q/D130A/V154I/V187N/T189Q，N011K/G023K/L075Q/D111A/D130A/V154I/V187N，Q004D/N011K/D027Q/E056K/S058M/N233Q/P256T，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/D130Y/V154I/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023K/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/G023K/L075R/V077I/N094R/D130A/G156W/V187Q，A018K/G023Q/K024A/L075Q/V077I/D130A/G156W/V187Q，A018K/G023Q/K024A/L075R/D130A/V154I/G156W/V187N，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，A018K/G023Q/L075Q/V077I/D130A/V154I/G156W/V187N，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/E056K/L075G/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/D027S/F051T/E056K/L075R/D130A/V187T/L264R，G023K/E056K/L075Q/D130A/V154I/V187N/T189Q/L264R，G023K/F051T/E056K/L075R/G091Q/D130A/V187T/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187T/L264R，G023K/K024A/L075R/N094R/V154I/G156W/V187T/T189Q，G023K/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，G023K/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/E056K/S058M/L075R/G091Q/V187N/I252Q，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/E056K/L075Q/N094R/D111A/G156W/T189Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/K024A/L075Q/V077I/D130A/V154I/G156W/V187N，G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/V187N，G023Q/L075Q/D111A/D130A/V154I/G156W/V187N/L264R，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，N011K/G023K/D027S/L075R/V154I/G156W/V187N/T189Q，N011K/G023K/E056K/L075Q/D130A/V154I/T189Q/L264R，N011K/G023Q/D027S/L075Q/N094R/V154I/G156W/T189Q，N011K/G023Q/D027S/N094R/V154I/G156W/T189Q/L264R，N011K/G023Q/L075Q/N094R/V154I/G156W/T189Q/L264R，A018K/G023K/K024A/L075Q/N094R/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/V154I/G156W/V187N，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/L075Q/V077I/R108K/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/G156W/V187H，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/P029E/S058M/V077I/R108K/H135F/G156W，A018K/G023Q/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W/V187N，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，G023K/D027S/F051T/E056K/S058M/L075Q/G091Q/V187N/L264R，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023K/L075Q/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/F051T/E056K/L075G/G091Q/V187N/I252Q/L264R，G023Q/D027S/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/G023K/D027S/E056K/L075R/D111A/G156W/V187N/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，N011K/G023Q/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/S058M/L075Q/V077I/R108K/H135F/G156W/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023Q/D027S/F051T/E056K/L075R/G091Q/D130A/V187H/I252Q/L264R，G023Q/D027S/F051T/E056K/S058M/L075R/G091Q/D130A/V187H/I252Q，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023Q/D027S/E056K/L075R/D130A/V154I/G156W/T189Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T，N011K/G023Q/D027S/L075Q/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D
表5-5：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比参考序列SEQIDNO：2在洗涤剂稳定性测定中性能指数＞1的TLL变体。
A018K/L075D，D027F/G163P，D027S/N033D，D027S/P256T，D111A/L264R，D130A/L264R，K024A/V154I，P029E/N033D，A018K/A049V/V187T，A018K/K024A/V154I，A018K/L075D/T189D，A018K/N033D/T189D，A018K/P029E/T189D，A018K/V154I/G156W，D027E/S058M/G163P，D027N/S058M/P256T，D027Q/S058M/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/S058M/P256T，D130A/V187T/L264R，E056K/D130A/L264R，G023K/D130A/L264R，G023K/D130A/V187T，G023K/V187N/L264R，G023K/V187T/L264R，G023Q/A049V/T189D，I090F/N233Q/P256T，L075G/D130A/V187H，L075Q/V187N/L264R，L075R/D130A/V187T，N011K/N233Q/P256T，N094R/D130A/V187T，P029E/N033D/E045F，Q004D/D027S/P256T，A018K/D027S/P029E/T189D，A018K/D111A/G156W/T189Q，A018K/D130A/G156W/V187T，A018K/E045F/L075D/T189D，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/V187T，A018K/G023Q/V077I/G156W，A018K/N033D/L075D/T189D，D027E/D137Q/G163P/L227M，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/D130A/I252Q/L264R，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029F/L075D/T189D，D027S/P029E/N033D/L075D，D111A/D130A/V154I/G156W，E045F/L075Q/G156W/V187T，E056K/D130A/V187N/L264R，E056K/D130A/V187T/L264R，F051T/L075Q/I252Q/L264R，G023K/D027Q/F051T/L075Q，G023K/D130A/G156W/T189Q，G023K/D130A/V187N/L264R，G023K/D130A/V187T/L264R，G023K/D130A/V187T/T189Q，G023K/K024A/V154I/V187N，G023K/L075R/D130A/L264R，G023K/L075R/V187T/L264R，G023Q/A049V/V077I/G156W，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/K024A/L075R/G156W，G023Q/K024A/V077I/G156W，G023Q/L075Q/D130A/L264R，G023Q/L075Q/V187T/L264R，K024A/D130A/V154I/V187T，L075G/D130A/V187T/I252Q，L075G/G091Q/V187N/L264R，N011K/D027Q/S058M/P256T，N011K/D027S/N233Q/P256T，P029E/N073R/L075D/T189D，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/N011K/D027S/P256T，S058M/D137Q/G163P/N233Q，S058M/G163P/L227M/L264R，S058M/G163P/N233Q/L264R，S058M/L075Q/G091Q/I252Q，S058M/L075Q/I252Q/L264R，A018K/D027S/P029E/N033D/L075D，A018K/G023K/D130A/V154I/G156W，A018K/G023K/K024A/D130A/G156W，A018K/G023K/K024A/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/K024A/L075Q/G156W，A018K/K024A/L075R/D130A/V154I，D027E/D048Q/G163P/N233Q/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/N233Q，D027S/E056K/D111A/V187N/L264R，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D048Q/D137Q/G163P/N233Q/L264R，D048Q/S058M/D130A/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/D137Q/N233Q/L264R，D048Q/S058M/G163P/L227M/L264R，D048Q/S058M/G163P/N233Q/L264R，D130A/D137Q/G163P/N233Q/L264R，D130A/V154I/G156W/V187N/T189Q，E056K/L075R/D130A/V187T/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/G091Q/D130A/L264R，F051T/L075Q/G091Q/D130A/L264R，F051T/L075Q/V187N/I252Q/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187N/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187N/L264R，G023K/E056K/L075R/V187T/L264R，G023K/K024A/D111A/D130A/V187T，G023K/K024A/D111A/G156W/T189Q，G023K/L075Q/V077I/D130A/G156W，G023K/L075Q/V187H/I252Q/L264R，G023K/L075R/D130A/V187N/L264R，G023K/L075R/D130A/V187T/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/D027S/L075G/I252Q/L264R，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075G/G091Q/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/V154I/G156W/V187Q，N011K/D027N/S058M/N233Q/P256T，N011K/D027S/I090F/N233Q/P256T，N011K/G023K/D111A/G156W/L264R，Q004D/N011K/D027N/N233Q/P256T，S058M/D130A/D137Q/G163P/L264R，S058M/D130A/G163P/L227M/L264R，S058M/G163P/L227M/N233Q/L264R，V077I/D130A/V154I/G156W/V187N，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/K024A/V077I/V154I/V187N，A018K/G023Q/K024A/V154I/G156W/V187Q，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/F051T/G091Q/D130A/I252Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/L264R，D048Q/S058M/D137Q/G163P/N233Q/L264R，D048Q/S058M/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/E056K/L075R/D130A/V187T/L264R，G023K/F051T/G091Q/D130A/V187H/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/L075Q/D130A/V187T/I252Q，G023Q/D027S/N094R/V154I/G156W/T189Q，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/K024A/V077I/D130A/V154I/G156W，G023Q/L075G/G091Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/V187N/I252Q/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/N094R/D130A/V154I/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/D027S/E056K/I090F/N233Q/P256T，Q004D/N011K/D027N/I090F/N233Q/P256T，Q004D/N011K/D027N/S058M/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，S058M/D130A/G163P/L227M/N233Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/D111A/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，D027E/D048Q/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/D048Q/S058M/D130A/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/D137Q/L227M/N233Q/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/S058M/G091Q/V187N/I252Q，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T
表5-6：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比参考序列SEQIDNO：2在洗涤剂稳定性测定中性能指数≥最大PI值的50％的TLL变体。
D027S/N033D，D027S/L075Q/G091Q，D027S/N033D/T189D，E056K/D130A/L264R，G023K/D130A/L264R，G023K/D130A/V187T，G023K/V187N/L264R，G023K/V187T/L264R，L075G/D130A/V187H，A018K/D027S/P029E/T189D，D027Q/E056K/N233Q/P256T，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，E056K/D130A/V187T/L264R，G023K/D027Q/F051T/L075Q，G023K/D130A/V187N/L264R，G023K/K024A/V154I/V187N，G023K/L075R/D130A/L264R，G023K/L075R/V187T/L264R，L075G/D130A/V187T/I252Q，P029E/N073R/L075D/T189D，Q004D/D027Q/N233Q/P256T，Q004D/N011K/D027S/P256T，A018K/D027S/P029E/N033D/L075D，A018K/G023K/D130A/V154I/G156W，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187N/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/V187N/L264R，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/L075Q/V187H/I252Q/L264R，G023K/L075R/D130A/V187N/L264R，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/L075G/G091Q/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，A018K/G023K/L075Q/V077I/G156W/V187Q，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/F051T/G091Q/D130A/I252Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/E056K/L075R/D130A/V187T/L264RG023K/F051T/G091Q/D130A/V187H/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/L075Q/D130A/V187T/I252Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/L075G/G091Q/D130A/I252Q/L264R，N011K/D027S/E056K/I090F/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q
实施例6
TLL组合变体的LAS稳定性
按实施例1所述的程序(LAS稳定性测定法)，分析测定了按实施例2所述生成的TLL变体的LAS稳定性。计算了变体相比TLL(SEQIDNO：1)或参考序列SEQIDNO：2的性能指数。
表6-1：下面示出，相比TLL(SEQIDNO：1)，在LAS稳定性测定中性能指数＞1的TLL变体。
A018K/L075Q，A018K/T189D，D027E/G163P，D027N/E056K，D027N/N233Q，D027S/N033D，D027S/P256T，D111A/L264R，D130A/T189Q，D130A/V187N，D130A/V187T，G023K/D130A，G023Q/L075Q，K024A/L075Q，K024A/V154I，L075Q/D130A，L075Q/G156W，N011K/D027S，N073R/L075D，N094R/G156W，P029E/N033D，A018K/A049V/L075Q，A018K/A049V/V187T，A018K/D027S/E045F，A018K/D027S/N073R，A018K/D027S/T189D，A018K/E045F/T189D，A018K/E045F/V187T，A018K/G023K/G156W，A018K/G023K/L075Q，A018K/G023Q/L075Q，A018K/K024A/V154I，A018K/L075Q/G156W，A018K/L075Q/V187T，A018K/N033D/T189D，A018K/N073R/L075D，A018K/P029E/T189D，A018K/V154I/G156W，A049V/V187T/T189D，D027E/S058M/G163P，D027N/N233Q/P256T，D027N/S058M/P256T，D027Q/N233Q/P256T，D027Q/S058M/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/N073R/L075D，D027S/N233Q/P256T，D027S/S058M/P256T，D111A/D130A/L264R，D130A/V154I/G156W，E045F/L075D/T189D，E056K/D130A/L264R，G023K/D130A/V187T，G023K/V187N/L264R，G023K/V187T/L264R，G023Q/A049V/T189D，G023Q/D130A/G156W，G023Q/E056K/L075R，G023Q/K024A/G156W，G023Q/L075Q/V187T，I090F/N233Q/P256T，K024A/D130A/V154I，K024A/L075Q/G156W，K024A/L075Q/V077I，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/D130A/V187T，L075Q/G156W/T189D，L075Q/G156W/V187N，L075Q/G156W/V187T，L075Q/N094R/V154I，L075Q/V077I/T189D，L075Q/V154I/V187T，L075Q/V187N/L264R，L075Q/V187T/T189D，L075R/D130A/L264R，L075R/D130A/V187T，N011K/D027S/S058M，N011K/N233Q/P256T，N033D/E045F/L075D，N033D/N073R/T189D，N073R/L075D/T189D，N094R/D130A/V187T，P029E/L075D/T189D，P029E/N033D/E045F，P029E/N033D/L075D，P029E/N073R/L075D，Q004D/D027S/P256T，Q004D/S058M/N233Q，V077I/D130A/V154I，V077I/V187A/T189D，A018K/A049V/L075Q/T189D，A018K/D027S/N033D/L075D，A018K/D027S/P029E/T189D，A018K/D111A/G156W/T189Q，A018K/D130A/G156W/V187T，A018K/E045F/A049V/G156W，A018K/E045F/A049V/V187T，A018K/E045F/L075Q/T189D，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/T189D，A018K/G023Q/A049V/V187T，A018K/G023Q/E045F/T189D，A018K/G023Q/E045F/V187T，A018K/G023Q/G156W/V187T，A018K/G023Q/L075Q/T189D，A018K/G023Q/L075Q/V077I，A018K/G023Q/L075Q/V187T，A018K/G023Q/L075R/D130A，A018K/G023Q/V077I/G156W，A018K/G023Q/V077I/V187T，A018K/G023Q/V187T/T189D，A018K/K024A/D130A/G156W，A018K/L075Q/G156W/T189D，A018K/L075Q/G156W/V187T，A018K/L075Q/N094R/D111A，A018K/L075Q/N094R/D130A，A018K/L075Q/N094R/V187Q，A018K/L075Q/V077I/N094R，A018K/L075Q/V187T/T189D，A018K/N033D/L075D/T189D，A018K/N073R/L075D/T189D，A018K/P029E/N033D/L075D，A018K/P029E/N033D/T189D，A049V/L075Q/V077I/T189D，A049V/L075Q/V187T/T189D，D027N/E056K/N233Q/P256T，D027N/E056K/S058M/P256T，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/E056K/D111A/V154I，D027S/E056K/I090F/P256T，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/L075D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/L075D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D027S/P029E/N033D/N073R，D027S/P029E/N033D/T189D，D048Q/D137Q/G163P/L227M，D111A/D130A/V154I/G156W，D111A/D130A/V154I/L264R，D111A/D130A/V187T/L264R，E045F/L075Q/G156W/V187T，E056K/D130A/V187T/L264R，E056K/L075Q/V187N/L264R，F051T/D130A/I252Q/L264R，G023K/D027Q/F051T/L075Q，G023K/D027S/L075Q/L264R，G023K/D111A/D130A/L264R，G023K/D130A/G156W/T189Q，G023K/D130A/V187T/L264R，G023K/D130A/V187T/T189Q，G023K/E056K/L075R/L264R，G023K/K024A/V154I/V187N，G023K/L075Q/D130A/V187N，G023K/L075Q/G156W/L264R，G023K/L075Q/G156W/V187N，G023Q/A049V/L075Q/T189D，G023Q/A049V/L075Q/V077I，G023Q/A049V/V077I/G156W，G023Q/D027S/D111A/G156W，G023Q/E045F/A049V/T189D，G023Q/E045F/A049V/V187T，G023Q/E045F/L075Q/G156W，G023Q/E045F/L075Q/V187T，G023Q/K024A/D130A/G156W，G023Q/K024A/L075R/G156W，G023Q/K024A/L075R/V154I，G023Q/K024A/V077I/G156W，G023Q/L075Q/G156W/V187N，G023Q/L075Q/G156W/V187T，G023Q/V154I/G156W/V187N，K024A/D130A/V154I/V187T，K024A/L075Q/D130A/G156W，K024A/L075Q/D130A/V154I，K024A/L075Q/V187T/T189Q，K024A/L075R/G156W/V187N，L075G/D130A/V187T/I252Q，L075Q/D130A/G156W/V187N，L075Q/D130A/V154I/G156W，L075Q/G156W/V187T/L264R，L075Q/V077I/D130A/V187Q，L075Q/V077I/G156W/V187N，L075Q/V077I/N094R/G156W，L075Q/V077I/V154I/V187Q，L075Q/V077I/V187T/T189D，L075R/D111A/V154I/G156W，L075R/D130A/V154I/L264R，N011K/D027N/E056K/S058M，N011K/D027N/S058M/P256T，N011K/D027Q/S058M/P256T，N011K/D027S/E056K/P256T，N011K/D027S/N233Q/P256T，N011K/E056K/N233Q/P256T，N011K/E056K/S058M/P256T，N011K/G023K/L075Q/D111A，N011K/S058M/N233Q/P256T，P029E/E045F/L075D/T189D，P029E/E045F/N073R/L075D，P029E/E045F/N073R/T189D，P029E/N033D/L075D/T189D，P029E/N073R/L075D/T189D，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/D027S/I090F/P256T，Q004D/N011K/D027N/P256T，Q004D/N011K/D027Q/N233Q，Q004D/N011K/D027S/P256T，S058M/L075Q/G091Q/I252Q，A018K/A049V/L075Q/V187T/T189D，A018K/D027S/N033D/L075D/T189D，A018K/D027S/N033D/N073R/T189D，A018K/D027S/P029E/E045F/T189D，A018K/D027S/P029E/N033D/L075D，A018K/D027S/P029E/N073R/L075D，A018K/E045F/A049V/L075Q/G156W，A018K/G023K/D130A/V154I/G156W，A018K/G023K/K024A/D130A/G156W，A018K/G023K/K024A/D130A/V154I，A018K/G023K/K024A/L075R/N094R，A018K/G023K/L075Q/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/A049V/L075Q/G156W，A018K/G023Q/A049V/L075Q/V077I，A018K/G023Q/A049V/L075Q/V187T，A018K/G023Q/A049V/V077I/V187T，A018K/G0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D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/K024A/L075Q/V077I/D130A/G156W/V187N，G023Q/K024A/L075Q/V077I/D130A/V154I/G156W，G023Q/L075Q/D111A/D130A/V154I/G156W/L264R，G023Q/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/L075Q/D130A/V154I/V187N/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，G023Q/L075Q/V077I/N094R/D130A/G156W/V187Q，K024A/L075Q/D111A/D130A/V154I/G156W/L264R，K024A/L075Q/D111A/D130A/V187N/T189Q/L264R，K024A/L075R/D130A/V154I/G156W/T189Q/L264R，L075Q/D111A/D130A/G156W/V187N/T189Q/L264R，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，N011K/E056K/L075Q/D111A/D130A/V154I/L264R，N011K/E056K/L075Q/D130A/V154I/V187N/T189Q，N011K/G023K/L075Q/D111A/D130A/V154I/V187N，N011K/G023Q/L075Q/V154I/V187N/T189Q/L264R，Q004D/D027N/E056K/S058M/I090F/N233Q/P256T，Q004D/N011K/D027N/S058M/I090F/N233Q/P256T，Q004D/N011K/D027Q/E056K/S058M/N233Q/P256T，Q004D/N011K/D027Q/S058M/I090F/N233Q/P256T，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/D130Y/V154I/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023K/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/G023K/L075R/V077I/N094R/D130A/G156W/V187Q，A018K/G023Q/E045F/L075Q/V077I/G156W/V187T/T189D，A018K/G023Q/K024A/L075Q/V077I/D130A/G156W/V187Q，A018K/G023Q/K024A/L075R/D130A/V154I/G156W/V187N，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，A018K/G023Q/L075Q/V077I/D130A/V154I/G156W/V187N，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/D027S/F051T/E056K/L075R/D130A/V187T/L264R，G023K/E056K/L075Q/D130A/V154I/V187N/T189Q/L264R，G023K/F051T/E056K/L075R/G091Q/D130A/V187T/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187T/L264R，G023K/K024A/L075R/N094R/V154I/G156W/V187T/T189Q，G023K/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，G023K/L0750/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/E056K/S058M/L075R/G091Q/V187N/I252Q，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/E056K/L075Q/D111A/V154I/V187N/T189Q/L264R，G023Q/E056K/L075Q/N094R/D111A/G156W/T189Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/K024A/L075Q/V077I/D130A/V154I/G156W/V187N，G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/V187N，G023Q/L075Q/D111A/D130A/V154I/G156W/V187N/L264R，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，N011K/G023K/D027S/L075Q/D111A/D130A/V154I/T189Q，N011K/G023K/D027S/L075R/V154I/G156W/V187N/T189Q，N011K/G023K/E056K/L075Q/D130A/V154I/T189Q/L264R，N011K/G023Q/D027S/L075Q/N094R/V154I/G156W/T189Q，N011K/G023Q/D027S/N094R/V154I/G156W/T189Q/L264R，N011K/G023Q/L075Q/N094R/V154I/G156W/T189Q/L264R，A018K/G023K/K024A/L075Q/N094R/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023Q/D027S/L075Q/V077I/R108K/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/G156W/V187H，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/P029E/S058M/V077I/R108K/H135F/G156W，A018K/G023Q/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W/V187N，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V187N/T189Q/L264R，G023K/D027S/F051T/E056K/S058M/L075Q/G091Q/V187N/L264R，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/D027S/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023K/L075Q/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/F051T/E056K/L075G/G091Q/V187N/I252Q/L264R，G023Q/D027S/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，G023Q/D027S/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，N011K/G023K/D027S/E056K/L075R/D111A/G156W/V187N/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，N011K/G023Q/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/E045F/A049V/S058M/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077T/R108K/H135F/V187H，A018K/G023Q/D027S/S058M/L075Q/V077I/R108K/H135F/G156W/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023K/D027S/F051T/E056K/S058M/L075G/G091Q/D130A/I252Q/L264R，G023Q/D027S/F051T/E056K/L075R/G091Q/D130A/V187H/I252Q/L264R，G023Q/D027S/F051T/E056K/S058M/L075R/G091Q/D130A/V187H/I252Q，G023Q/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023K/L075R/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，N011K/G023Q/D027S/E056K/L075R/D130A/V154I/G156W/T189Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，N011K/G023Q/L075Q/N094R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T，N011K/G023Q/D027S/L075Q/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/N033D/E045F/A049V/S058M/N073S/K074S/L075Q/V077I/N101D/R108K/H135F/D137V/G156W/V187T/T189D
表6-2：下面示出，相比TLL(SEQIDNO：1)，表达性能指数≥0.5，且在LAS稳定性测定中性能指数＞1的TLL变体。
D027E/G163P，D027S/N033D，D027S/P256T，D111A/L264R，D130A/V187N，D130A/V187T，K024A/V154I，N094R/G156W，P029E/N033D，A018K/A049V/V187T，A018K/E045F/V187T，A018K/K024A/V154I，A018K/N033D/T189D，A018K/P029E/T189D，A018K/V154I/G156W，D027E/S058M/G163P，D027N/S058M/P256T，D027Q/S058M/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/S058M/P256T，D111A/D130A/L264R，D130A/V154I/G156W，E056K/D130A/L264R，G023K/D130A/V187T，G023K/V187N/L264R，G023K/V187T/L264R，G023Q/A049V/T189D，G023Q/D130A/G156W，G023Q/K024A/G156W，I090F/N233Q/P256T，K024A/D130A/V154I，L075G/D130A/V187H，L075Q/V187N/L264R，L075R/D130A/L264R，L075R/D130A/V187T，N011K/N233Q/P256T，N094R/D130A/V187T，P029E/N033D/E045F，Q004D/D027S/P256T，Q004D/S058M/N233Q，A018K/D027S/P029E/T189D，A018K/D111A/G156W/T189Q，A018K/D130A/G156W/V187T，A018K/E045F/A049V/G156W，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/V187T，A018K/G023Q/V077I/G156W，A018K/K024A/D130A/G156W，A018K/L075Q/V077I/N094R，A018K/N033D/L075D/T189D，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/D137Q/G163P/L227M，D111A/D130A/V154I/G156W，D111A/D130A/V187T/L264R，E045F/L075Q/G156W/V187T，E056K/D130A/V187T/L264R，F051T/D130A/I252Q/L264R，G023K/D027Q/F051T/L075Q，G023K/D130A/G156W/T189Q，G023K/D130A/V187T/L264R，G023K/D130A/V187T/T189Q，G023K/E056K/L075R/L264R，G023K/K024A/V154I/V187N，G023Q/A049V/V077I/G156W，G023Q/K024A/D130A/G156W，G023Q/K024A/L075R/G156W，G023Q/K024A/V077I/G156W，K024A/D130A/V154I/V187T，L075G/D130A/V187T/I252Q，N011K/D027Q/S058M/P256T，N011K/D027S/N233Q/P256T，P029E/N073R/L075D/T189D，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/N011K/D027S/P256T，S058M/L075Q/G091Q/I252Q，A018K/D027S/P029E/N033D/L075D，A018K/G023K/D130A/V154I/G156W，A018K/G023K/K024A/D130A/G156W，A018K/G023K/K024A/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/K024A/D130A/V187Q，A018K/G023Q/K024A/L075Q/G156W，A018K/K024A/L075R/D130A/V154I，A018K/K024A/N094R/D130A/V187N，A018K/K024A/N094R/G156W/V187T，D027E/D048Q/D137Q/L227M/L264R，D027E/D048Q/G163P/N233Q/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/N233Q，D027S/E056K/D111A/V187N/L264R，D027S/L075R/V154I/T189Q/L264R，D048Q/S058M/G163P/N233Q/L264R，D130A/V154I/G156W/V187N/T189Q，E056K/L075R/D130A/V187T/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187N/L264R，G023K/E056K/L075R/V187T/L264R，G023K/K024A/D111A/D130A/V187T，G023K/K024A/D111A/G156W/T189Q，G023K/L075Q/V077I/D130A/G156W，G023K/L075Q/V187H/I252Q/L264R，G023K/L075R/D130A/V187N/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/D027S/L075G/I252Q/L264R，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/V154I/G156W/V187Q，N011K/D027N/S058M/N233Q/P256T，N011K/D027S/I090F/N233Q/P256T，N011K/G023K/D111A/G156W/L264R，Q004D/N011K/D027N/N233Q/P256T，S058M/G163P/L227M/N233Q/L264R，V077I/D130A/V154I/G156W/V187N，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/G023Q/K024A/V077I/V154I/V187N，A018K/G023Q/K024A/V154I/G156W/V187Q，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D137Q/G163P/L227M/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/F051T/G091Q/D130A/I252Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D048Q/D130A/G163P/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/E056K/L075R/D130A/V187T/L264R，G023K/F051T/G091Q/D130A/V187H/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/N094R/V154I/G156W/T189Q，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/K024A/V077I/D130A/V154I/G156W，G023Q/L075Q/G091Q/V187N/I252Q/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/N094R/D130A/V154I/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，N011K/D027S/E056K/I090F/N233Q/P256T，Q004D/N011K/D027N/I090F/N233Q/P256T，Q004D/N011K/D027N/S058M/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/D111A/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/S058M/G091Q/V187N/I252Q，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，N011K/G023Q/L075Q/V154I/V187N/T189Q/L264R，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T
表6-3：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且在LAS稳定性测定中性能指数≥最大PI值的50％的TLL变体。
D027S/P256T，D130A/V187T，K024A/V154I，A018K/A049V/V187T，A018K/E045F/V187T，A018K/K024A/V154I，A018K/V154I/G156W，D027N/S058M/P256T，D027Q/S058M/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/S058M/P256T，D130A/V154I/G156W，G023K/D130A/V187T，G023K/V187T/L264R，G023Q/A049V/T189D，I090F/N233Q/P256T，K024A/D130A/V154I，L075Q/V187N/L264R，L075R/D130A/L264R，N011K/N233Q/P256T，N094R/D130A/V187T，Q004D/D027S/P256T，Q004D/S058M/N233Q，A018K/D027S/P029E/T189D，A018K/D111A/G156W/T189Q，A018K/D130A/G156W/V187T，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/V187T，A018K/G023Q/V077I/G156W，A018K/K024A/D130A/G156W，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/D137Q/G163P/L227M，D111A/D130A/V154I/G156W，E045F/L075Q/G156W/V187T，F051T/D130A/I252Q/L264R，G023K/D027Q/F051T/L075Q，G023K/D130A/G156W/T189Q，G023K/D130A/V187T/T189Q，G023K/E056K/L075R/L264R，G023K/K024A/V154I/V187N，G023Q/A049V/V077I/G156W，G023Q/K024A/D130A/G156W，G023Q/K024A/L075R/G156W，G023Q/K024A/V077I/G156W，K024A/D130A/V154I/V187T，N011K/D027Q/S058M/P256T，N011K/D027S/N233Q/P256T，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/N011K/D027S/P256T，S058M/L075Q/G091Q/I252Q，A018K/D027S/P029E/N033D/L075D，A018K/G023K/D130A/V154I/G156W，A018K/G023K/K024A/D130A/G156W，A018K/G023K/K024A/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/K024A/D130A/V187Q，A018K/G023Q/K024A/L075Q/G156W，A018K/K024A/L075R/D130A/V154I，A018K/K024A/N094R/G156W/V187T，D027E/D048Q/D137Q/L227M/L264R，D027E/G163P/L227M/N233Q/L264R，D027S/E056K/D111A/V187N/L264R，D027S/L075R/V154I/T189Q/L264R，D130A/V154I/G156W/V187N/T189Q，E056K/L075R/D130A/V187T/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/D027S/E056K/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187N/L264R，G023K/E056K/L075R/V187T/L264R，G023K/K024A/D111A/D130A/V187T，G023K/K024A/D111A/G156W/T189Q，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187N/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/D027S/L075G/I252Q/L264R，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/V154I/G156W/V187Q，N011K/D027N/S058M/N233Q/P256T，N011K/D027S/I090F/N233Q/P256T，N011K/G023K/D111A/G156W/L264R，Q004D/N011K/D027N/N233Q/P256T，V077I/D130A/V154I/G156W/V187N，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/G023Q/K024A/V077I/V154I/V187N，A018K/G023Q/K024A/V154I/G156W/V187Q，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/N094R/D111A/D130A/V154I/V187N，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D048Q/D130A/G163P/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/D027S/N094R/V154I/G156W/T189Q，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/K024A/V077I/D130A/V154I/G156W，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/N094R/D130A/V154I/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，N011K/D027S/E056K/I090F/N233Q/P256T，Q004D/N011K/D027N/I090F/N233Q/P256T，Q004D/N011K/D027N/S058M/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，A018K/D111A/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，G023K/D027S/F051T/S058M/G091Q/V187N/I252Q，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，N011K/G023Q/L075Q/V154I/V187N/T189Q/L264R，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T
表6-4：下面示出，相比参考序列SEQIDNO：2，在LAS稳定性测定中性能指数＞1的TLL变体。
D130A/V187T，N011K/D027S，A018K/D027S/N073R，D027N/N233Q/P256T，D027S/L075Q/G091Q，E045F/L075D/T189D，G023K/V187T/L264R，I090F/N233Q/P256T，N033D/E045F/L075D，N033D/N073R/T189D，Q004D/S058M/N233Q，A018K/L075Q/G156W/T189D，A018K/L075Q/V187T/T189D，A049V/L075Q/V077I/T189D，D027S/E056K/D111A/V154I，D027S/N033D/L075D/T189D，D027S/N033D/N073R/L075D，D027S/P029E/E045F/L075D，D027S/P029E/N033D/N073R，D048Q/D137Q/G163P/L227M，G023K/D027S/L075Q/L264R，G023K/E056K/L075R/L264R，G023Q/A049V/L075Q/V077I，G023Q/D027S/D111A/G156W，K024A/L075Q/D130A/V154I，N011K/D027Q/S058M/P256T，N011K/E056K/N233Q/P256T，N011K/G023K/L075Q/D111A，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/N011K/D027N/P256T，Q004D/N011K/D027S/P256T，D027E/D048Q/D137Q/L227M/L264R，D027E/G163P/L227M/N233Q/L264R，D027N/S058M/I090F/N233Q/P256T，D027S/P029E/N033D/N073R/L075D，E056K/D130A/V187H/I252Q/L264R，E056K/L075R/D130A/V187T/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/L075R/V187N/L264R，G023K/E056K/L075R/V187T/L264R，G023K/K024A/D111A/D130A/V187T，G023Q/D027S/L075G/I252Q/L264R，G023Q/K024A/L075Q/D130A/G156W，L075R/D111A/D130A/V187N/T189Q，N011K/D027N/E056K/N233Q/P256T，N011K/D027N/E056K/S058M/P256T，N011K/D027N/S058M/N233Q/P256T，N011K/D027Q/E056K/S058M/N233Q，N011K/D027Q/I090F/N233Q/P256T，N011K/D027Q/S058M/N233Q/P256T，N011K/D027S/E056K/N233Q/P256T，N011K/G023K/E056K/L075Q/T189Q，Q004D/D027S/E056K/N233Q/P256T，Q004D/N011K/D027Q/S058M/N233Q，A018K/D027S/P029E/N033D/L075D/T189D，A018K/G023K/D111A/D130A/V154I/T189Q，D027Q/E056K/S058M/D130A/I252Q/L264R，D027S/E056K/S058M/V187T/I252Q/L264R，D027S/G091Q/D130A/V187N/I252Q/L264R，D027S/L075Q/D111A/G156W/V187N/T189Q，D048Q/D130A/G163P/L227M/N233Q/L264R，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/L075Q/V077I/D130A/G156W/V187Q，G023Q/D027S/F051T/L075Q/V187T/I252Q，G023Q/D027S/L075R/D111A/V187N/L264R，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，N011K/D027N/E056K/S058M/N233Q/P256T，N011K/D027Q/E056K/S058M/N233Q/P256T，N011K/D027Q/S058M/I090F/N233Q/P256T，N011K/D027S/E056K/S058M/N233Q/P256T，N011K/G023K/D027S/V154I/G156W/T189Q，N011K/G023Q/L075Q/D130A/V187N/T189Q，P029E/N033D/K074S/L075D/N101D/T189D，Q004D/D027S/E056K/S058M/N233Q/P256T，Q004D/N011K/D027N/I090F/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，D027Q/F051T/L075Q/G091Q/D130A/V187T/I252Q，G023K/D027S/F051T/S058M/G091Q/V187N/I252Q，G023K/D027S/L075Q/N094R/V154I/G156W/T189Q，G023K/F051T/L075Q/G091Q/D130A/V187H/I252Q，G023K/L075Q/D111A/D130A/V154I/G156W/L264R，G023Q/L075Q/D130A/V154I/V187N/T189Q/L264R，N011K/E056K/L075Q/D130A/V154I/V187N/T189Q，N011K/G023K/L075Q/D111A/D130A/V154I/V187N，Q004D/N011K/D027N/S058M/I090F/N233Q/P256T，Q004D/N011K/D027Q/E056K/S058M/N233Q/P256T，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/D027S/F051T/E056K/L075R/D130A/V187T/L264R，G023K/F051T/E056K/L075R/G091Q/D130A/V187T/L264R，G023Q/D027S/E056K/S058M/L075R/G091Q/V187N/I252Q，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，N011K/G023K/D027S/L075Q/D111A/D130A/V154I/T189Q，A018K/G023Q/D027S/L075Q/V077I/R108K/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/G156W/V187H，A018K/G023Q/D027S/P029E/S058M/V077I/R108K/H135F/G156W，A018K/G023Q/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，G023K/D027S/F051T/E056K/S058M/L075Q/G091Q/V187N/L264R，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/D027S/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023Q/D027S/F051T/E056K/L075G/G091Q/V187N/I252Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/V187T/T189D，A018K/G023Q/D027S/S058M/L075Q/V077I/R108K/H135F/G156W/V187H，G023K/D027S/F051T/E056K/S058M/L075G/G091Q/D130A/I252Q/L264R，G023Q/D027S/F051T/E056K/L075R/G091Q/D130A/V187H/I252Q/L264R，G023Q/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023Q/D027S/E056K/L075R/D130A/V154I/G156W/T189Q/L264R
表6-5：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比参考序列SEQIDNO：2在LAS稳定性测定中性能指数＞1的TLL变体。
D130A/V187T，D027S/L075Q/G091Q，G023K/V187T/L264R，I090F/N233Q/P256T，Q004D/S058M/N233Q，D027S/N033D/L075D/T189D，D048Q/D137Q/G163P/L227M，G023K/E056K/L075R/L264R，N011K/D027Q/S058M/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/N011K/D027S/P256T，D027E/D048Q/D137Q/L227M/L264R，D027E/G163P/L227M/N233Q/L264R，E056K/L075R/D130A/V187T/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/L075R/V187N/L264R，G023K/E056K/L075R/V187T/L264R，G023K/K024A/D111A/D130A/V187T，G023Q/D027S/L075G/I252Q/L264R，N011K/D027N/S058M/N233Q/P256T，D027Q/E056K/S058M/D130A/I252Q/L264R，D027S/E056K/S058M/V187T/I252Q/L264R，D048Q/D130A/G163P/L227M/N233Q/L264R，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，Q004D/N011K/D027N/I090F/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，G023K/D027S/F051T/S058M/G091Q/V187N/I252Q，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/I264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/1252Q，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q
表6-6：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比参考序列SEQIDNO：2在LAS稳定性测定中性能指数≥最大PI值的50％的TLL变体。
D130A/V187T，G023K/V187T/L264R，I090F/N233Q/P256T，Q004D/S058M/N233Q，D048Q/D137Q/G163P/L227M，G023K/E056K/L075R/L264R，Q004D/D027Q/N233Q/P256T，Q004D/N011K/D027S/P256T，D027E/D048Q/D137Q/L227M/L264R，D027E/G163P/L227M/N233Q/L264R，E056K/L075R/D130A/V187T/L264R，G023K/E056K/L075R/V187N/L264R，G023K/K024A/D111A/D130A/V187T，G023Q/D027S/L075G/I252Q/L264R，N011K/D027N/S058M/N233Q/P256T，D027Q/E056K/S058M/D130A/I252Q/L264R，D027S/E056K/S058M/V187T/I252Q/L264R，D048Q/D130A/G163P/L227M/N233Q/L264R，G023Q/E056K/L075Q/D130A/V154I/L264R，Q004D/N011K/D027S/S058M/N233Q/P256T，G023K/D027S/F051T/S058M/G091Q/V187N/I252Q，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q
实施例7
TLL组合变体的织物粘附性
按实施例1所述的程序(粘附性测定法)，分析测定了按实施例2所述生成的TLL变体在洗涤剂溶液中对棉织物的粘附性。计算了变体相比TLL(SEQIDNO：1)或参考序列SEQIDNO：2粘附性增大或减小的性能指数。
表7-1：下面示出，相比TLL(SEQIDNO：1)，在织物粘附性测定中性能指数＞1的TLL变体。
A018K/L075Q，D111A/L264R，D130A/L264R，D130A/V187N，D130A/V187T，G023K/L264R，L075R/L264R，S058M/L264R，V187T/L264R，A018K/A049V/V187T，A018K/G023Q/L075Q，A018K/K024A/V154I，A018K/L075Q/G156W，A018K/N033D/T189D，A018K/P029E/T189D，D111A/D130A/L264R，D130A/V187N/L264R，D130A/V187T/L264R，F051T/I252Q/L264R，G023K/D130A/L264R，G023K/D130A/V187T，G023K/E056K/V187T，G023K/L075R/L264R，G023K/V187N/L264R，G023K/V187T/L264R，G023Q/A049V/T189D，G023Q/L075Q/V187T，G163P/L227M/L264R，K024A/L075Q/V077I，L075Q/G156W/T189D，L075Q/G156W/V187N，L075Q/G156W/V187T，L075Q/V154I/V187T，L075Q/V187N/L264R，L075Q/V187T/T189D，L075R/D130A/L264R，L075R/D130A/V187T，N073R/L075D/T189D，P029E/L075D/T189D，A018K/D027S/P029E/T189D，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/V187T，A018K/G023Q/L075Q/T189D，A018K/G023Q/L075Q/V187T，A018K/G023Q/L075R/D130A，A018K/K024A/D130A/G156W，D027E/D130A/N233Q/L264R，D027E/G163P/L227M/L264R，D027E/S058M/G163P/L264R，D027S/D130A/I252Q/L264R，D027S/N033D/E045F/T189D，D027S/P029E/L075D/T189D，D048Q/D130A/G163P/L264R，D048Q/G163P/N233Q/L264R，D111A/D130A/V154I/G156W，D111A/D130A/V187T/L264R，E056K/D130A/V187N/L264R，E056K/D130A/V187T/L264R，F051T/L075G/I252Q/L264R，F051T/L075Q/I252Q/L264R，G023K/D130A/V187N/L264R，G023K/D130A/V187T/L264R，G023K/D130A/V187T/T189Q，G023K/E056K/L075R/L264R，G023K/K024A/V154I/V187N，G023K/L075Q/D130A/V187N，G023K/L075R/V187T/L264R，G023Q/A049V/L075Q/T189D，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/L075Q/D130A/L264R，G023Q/L075Q/G156W/V187N，G023Q/L075Q/V187T/L264R，G091Q/V187T/I252Q/L264R，K024A/D130A/V154I/V187T，K024A/L075R/G156W/V187N，L075G/D130A/V187T/I252Q，L075G/V187H/I252Q/L264R，L075Q/D130A/G156W/V187N，L075Q/V077I/D130A/V187Q，L075Q/V077I/V154I/V187Q，L075R/D111A/V154I/G156W，L075R/D130A/V154I/L264R，P029E/N033D/L075D/T189D，S058M/G163P/L227M/L264R，S058M/G163P/N233Q/L264R，S058M/L075Q/G091Q/I252Q，S058M/L075Q/I252Q/L264R，A018K/D130A/G156W/V187N/L264R，A018K/G023K/K024A/D130A/G156W，A018K/G023K/K024A/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/K024A/D130A/V187Q，A018K/G023Q/L075Q/G156W/V187T，A018K/K024A/L075R/N094R/D130A，A018K/K024A/N094R/D130A/V187N，A018K/K024A/N094R/G156W/V187T，A018K/L075Q/D111A/D130A/V187T，D027E/D048Q/D137Q/L227M/L264R，D027E/D130A/G163P/L227M/L264R，D027E/D130A/G163P/N233Q/L264R，D027E/D137Q/G163P/N233Q/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L264R，D027E/S058M/G163P/L227M/L264R，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D027S/L075R/V154I/T189Q/L264R，D027S/P029E/N073R/L075D/T189D，D048Q/D130A/G163P/L227M/L264R，D048Q/D137Q/G163P/L227M/L264R，D048Q/S058M/D137Q/N233Q/L264R，D130A/D137Q/G163P/L227M/L264R，D130A/D137Q/G163P/N233Q/L264R，D130A/V154I/G156W/V187N/T189Q，E056K/L075G/V187N/I252Q/L264R，E056K/L075Q/G156W/T189Q/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/G091Q/D130A/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/L264R，F051T/L075Q/V187N/I252Q/L264R，F051T/L075R/V187N/I252Q/L264R，G023K/E056K/D130A/V187N/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/D111A/D130A/V187T，G023K/L075Q/D130A/G156W/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187N/L264R，G023K/L075R/D130A/V187T/L264R，G023Q/E056K/V187N/I252Q/L264R，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/K024A/L075Q/G156W/V187N，G023Q/K024A/L075R/G156W/V187Q，G023Q/L075G/G091Q/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，G023Q/L075Q/V077I/G156W/V187N，G091Q/D130A/V187H/I252Q/L264R，K024A/L075Q/D111A/G156W/V187T，K024A/L075Q/V077I/G156W/V187N，K024A/L075Q/V077I/V154I/V187N，K024A/L075R/V154I/G156W/V187Q，L075G/D130A/V187T/I252Q/L264R，L075Q/D111A/V187N/T189Q/L264R，L075Q/V154I/V187N/T189Q/L264R，L075R/D130A/V154I/T189Q/L264R，L075R/D130A/V187T/T189Q/L264R，N011K/G023K/D111A/G156W/L264R，S058M/D130A/G163P/L227M/L264R，A018K/A049V/L075Q/G156W/V187T/T189D，A018K/G023K/K024A/L075R/D130A/V187N，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023K/L075Q/D130A/G156W/V187N，A018K/G023K/L075Q/G156W/V187N/L264R，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/K024A/V077I/V154I/V187N，A018K/G023Q/K024A/V154I/G156W/V187Q，A018K/G023Q/L075Q/G156W/V187T/T189D，A018K/G023Q/L075Q/V077I/D130A/V187N，A018K/G023Q/L075Q/V077I/G156W/V187N，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/D111A/V187N/T189Q/L264R，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/K024A/L075R/D130A/V187N/T189Q，A018K/L075Q/D130A/V154I/T189Q/L264R，A018K/L075Q/D130A/V187N/T189Q/L264R，A018K/L075R/D130A/V154I/G156W/V187N，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D048Q/D130A/D137H/G163P/L264R，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D130A/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D130A/D137Q/G163P/N233Q/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L227M/L264R，D027E/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/F051T/G091Q/D130A/I252Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027Q/F051T/L075Q/D130A/V187H/L264R，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/G163P/L227M/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/L264R，D048Q/S058M/D137Q/G163P/N233Q/L264R，D111A/D130A/V154I/G156W/V187N/L264R，F051T/E056K/D130A/V187N/I252Q/L264R，F051T/E056K/L075Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，F051T/L075R/D130A/V187N/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187T/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/L075Q/D111A/D130A/V154I/V187N，G023K/L075Q/N094R/V154I/V187T/L264R，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/L075Q/D130A/V187T/I252Q，G023Q/F051T/G091Q/V187N/I252Q/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/L075G/G091Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/V187N/I252Q/L264R，G023Q/L075R/V154I/G156W/V187N/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，K024A/L075Q/V154I/V187N/T189Q/L264R，K024A/L075Q/V154I/V187T/T189Q/L264R，K024A/N094R/V154I/G156W/T189Q/L264R，L075Q/D111A/D130A/V154I/V187N/T189Q，L075Q/D130A/V154I/G156W/V187N/L264R，L075Q/G091Q/D130A/V187H/I252Q/L264R，L075R/D130A/V154I/G156W/V187N/L264R，N011K/E056K/L075Q/D130A/V187N/T189Q，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，S058M/D130A/G163P/L227M/N233Q/L264R，A018K/D111A/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/K024A/L075Q/D130A/G156W/V187N，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/N094R/G156W，A018K/G023K/K024A/N094R/V154I/V187T/L264R，A018K/G023Q/K024A/L075Q/V077I/N094R/V154I，A018K/G023Q/K024A/L075R/D130A/V154I/V187N，A018K/K024A/D130A/V154I/G156W/V187T/L264R，A018K/K024A/L075Q/D111A/G156W/V187N/L264R，A018K/K024A/L075Q/D130A/G156W/V187T/T189Q，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，A018K/K024A/N094R/D130A/G156W/V187T/L264R，A018K/L075Q/D111A/V154I/V187T/T189Q/L264R，D027E/D048Q/D130A/D137Q/G163P/N233Q/L264R，D027E/D048Q/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/D048Q/S058M/D130A/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/L075G/D130A/V187H/I252Q/L264R，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/L075G/G091Q/V187T/I252Q/L264R，D027S/F051T/S058M/L075Q/V187N/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/D027Q/F051T/L075G/D130A/V187H/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023K/K024A/L075Q/D111A/D130A/T189Q/L264R，G023K/K024A/L075Q/D111A/V154I/G156W/V187N，G023K/K024A/L075Q/D130A/V187T/T189Q/L264R，G023K/K024A/L075Q/V077I/D130A/G156W/V187Q，G023K/L075R/N094R/V154I/G156W/V187N/L264R，G023Q/D027S/F051T/E056K/D130A/I252Q/L264R，G023Q/D027S/L075Q/G091Q/V187T/I252Q/L264R，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/E056K/L075Q/D111A/G156W/V187N/L264R，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/L075R/G091Q/V187N/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/K024A/L075Q/V077I/D130A/G156W/V187N，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/L075Q/D130A/V154I/V187N/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，K024A/L075Q/D111A/D130A/V154I/G156W/L264R，L075Q/D111A/D130A/G156W/V187N/T189Q/L264R，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，A018K/G023K/K024A/L075Q/D130Y/V154I/V187N/T189Q，A018K/G023K/K024A/L075R/D111A/V154I/V187T/L264R，A018K/G023Q/K024A/L075Q/V077I/D130A/G156W/V187Q，A018K/K024A/L075R/D130A/V154I/G156W/V187T/L264R，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/E056K/L075G/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/S058M/L075Q/V187H/I252Q/L264R，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，K024A/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，N011K/G023K/D027S/L075R/V154I/G156W/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/V187T/T189Q/L264K，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023K/L075Q/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023K/D027S/F051T/E056K/S058M/L075G/G091Q/D130A/I252Q/L264R，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R
表7-2：下面示出，相比TLL(SEQIDNO：1)，表达性能指数≥0.5，且在织物粘附性测定中性能指数＞1的TLL变体。D111A/L264R，D130A/L264R，D130A/V187N，D130A/V187T，G023K/L264R，L075R/L264R，S058M/L264R，V187T/L264R，A018K/A049V/V187T，A018K/K024A/V154I，A018K/N033D/T189D，A018K/P029E/T189D，D111A/D130A/L264R，D130A/V187N/L264R，D130A/V187T/L264R，F051T/1252Q/L264R，G023K/D130A/L264R，G023K/D130A/V187T，G023K/E056K/V187T，G023K/L075R/L264R，G023K/V187N/L264R，G023K/V187T/L264R，G023Q/A049V/T189D，G163P/L227M/L264R，L075Q/V187N/L264R，L075R/D130A/L264R，L075R/D130A/V187T，A018K/D027S/P029E/T189D，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/V187T，A018K/K024A/D130A/G156W，D027E/D130A/N233Q/L264R，D027E/G163P/L227M/L264R，D027E/S058M/G163P/L264R，D027S/D130A/1252Q/L264R，D027S/N033D/E045F/T189D，D027S/P029E/L075D/T189D，D048Q/D130A/G163P/L264R，D048Q/G163P/N233Q/L264R，D111A/D130A/V154I/G156W，D111A/D130A/V187T/L264R，E056K/D130A/V187N/L264R，E056K/D130A/V187T/L264R，F051T/L075G/1252Q/L264R，F051T/L075Q/1252Q/L264R，G023K/D130A/V187N/L264R，G023K/D130A/V187T/L264R，G023K/D130A/V187T/T189Q，G023K/E056K/L075R/L264R，G023K/K024A/V154I/V187N，G023K/L075R/V187T/L264R，G023Q/F051T/1252Q/L264R，G023Q/G091Q/1252Q/L264R，G023Q/L075Q/D130A/L264R，G023Q/L075Q/V187T/L264R，G091Q/V187T/1252Q/L264R，K024A/D130A/V154I/V187T，L075G/D130A/V187T/1252Q，L075G/V187H/1252Q/L264R，S058M/G163P/L227M/L264R，S058M/G163P/N233Q/L264R，S058M/L075Q/G091Q/1252Q，S058M/L075Q/1252Q/L264R，A018K/D130A/G156W/V187N/L264R，A018K/G023K/K024A/D130A/G156W，A018K/G023K/K024A/D130A/V154I，A018K/G023K/V0771/D130A/V187N，A018K/G023Q/K024A/D130A/V187Q，A018K/K024A/N094R/D130A/V187N，A018K/K024A/N094R/G156W/V187T，D027E/D048Q/D137Q/L227M/L264R，D027E/D130A/G163P/L227M/L264R，D027E/D130A/G163P/N233Q/L264R，D027E/D137Q/G163P/N233Q/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L264R，D027E/S058M/G163P/L227M/L264R，D027S/F051T/V187N/1252Q/L264R，D027S/L075G/D130A/1252Q/L264R，D027S/L075G/G091Q/1252Q/L264R，D027S/L075R/V1541/T189Q/L264R，D048Q/D130A/G163P/L227M/L264R，D048Q/D137Q/G163P/L227M/L264R，D048Q/S058M/D137Q/N233Q/L264R，D130A/D137Q/G163P/L227M/L264R，D130A/D137Q/G163P/N233Q/L264R，D130A/V154I/G156W/V187N/T189Q，E056K/L075G/V187N/I252Q/L264R，F05/T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/I264R，F051T/L075G/G091Q/D130A/L264R，F051T/L075G/V187N/1252Q/I264R，F051T/L075Q/G091Q/D130A/L264R，F051T/L075Q/V187N/I252Q/L264R，G023K/E056K/D130A/V187N/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187N/L264R，G023K/L075R/D130A/V187T/L264R，G023Q/E056K/V187N/I252Q/L264R，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/L075G/G091Q/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，K024A/L075R/V154I/G156W/V187Q，L075G/D130A/V187T/I252Q/L264R，N011K/G023K/D111A/G156W/L264R，S058M/D130A/G163P/L227M/L264R，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/K024A/V077I/V154I/V187N，A018K/G023Q/K024A/V154I/G156W/V187Q，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/D111A/V187N/T189Q/L264R，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D048Q/D130A/D137H/G163P/L264R，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D130A/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D130A/D137Q/G163P/N233Q/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L227M/L264R，D027E/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/F051T/G091Q/D130A/I252Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/G163P/L227M/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/L264R，D048Q/S058M/D137Q/G163P/N233Q/L264R，F051T/E056K/D130A/V187N/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187T/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/L075Q/D130A/V187T/I252Q，G023Q/F051T/G091Q/V187N/I252Q/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/L075G/G091Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/V187N/I252Q/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，L075Q/G091Q/D130A/V187H/I252Q/L264R，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，S058M/D130A/G163P/L227M/N233Q/L264R，A018K/D111A/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/K024A/D130A/V154I/G156W/V187T/L264R，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，A018K/K024A/N094R/D130A/G156W/V187T/L264R，D027E/D048Q/D130A/D137Q/G163P/N233Q/L264R，D027E/D048Q/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/D048Q/S058M/D130A/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/L075Q/D130A/V187T/I252Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/S058M/L075Q/V187N/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023Q/D027S/F051T/E056K/D130A/I252Q/L264R，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/S058M/L075Q/V187H/I252Q/L264R，G023Q/D027Q/F051T/L075Q/G091Q/D130A/I252Q/L264R，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T//T189Q，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R
表7-3：下面示出，相比TLL(SEQIDNO：1)，表达性能指数≥0.5，且在织物粘附性测定中性能指数≥最大PI值的50％的TLL变体。
D111A/L264R，D130A/L264R，D130A/V187N，D130A/V187T，G023K/L264R，L075R/L264R，S058M/L264R，V187T/L264R，A018K/A049V/V187T，A018K/K024A/V154I，D111A/D130A/L264R，D130A/V187N/L264R，D130A/V187T/L264R，F051T/I252Q/L264R，G023K/D130A/L264R，G023K/E056K/V187T，G023K/L075R/L264R，G023K/V187T/L264R，L075Q/V187N/L264R，L075R/D130A/L264R，A018K/G023K/D111A/T189Q，D027E/D130A/N233Q/L264R，D027S/D130A/I252Q/L264R，D048Q/D130A/G163P/L264R，D111A/D130A/V187T/L264R，E056K/D130A/V187N/L264R，F051T/L075G/I252Q/L264R，F051T/L075Q/I252Q/L264R，G023K/D130A/V187N/L264R，G023K/D130A/V187T/T189Q，G023K/E056K/L075R/L264R，G023K/K024A/V154I/V187N，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/L075Q/D130A/L264R，G023Q/L075Q/V187T/L264R，G091Q/V187T/I252Q/L264R，K024A/D130A/V154I/V187T，L075G/D130A/V187T/I252Q，S058M/G163P/L227M/L264R，S058M/L075Q/I252Q/L264R，A018K/D130A/G156W/V187N/L264R，A018K/G023K/K024A/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/K024A/D130A/V187Q，A018K/K024A/N094R/D130A/V187N，D027E/D048Q/D137Q/L227M/L264R，D027E/D130A/G163P/L227M/L264R，D027E/D130A/G163P/N233Q/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L264R，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D027S/L075R/V154I/T189Q/L264R，D048Q/S058M/D137Q/N233Q/L264R，D130A/D137Q/G163P/L227M/L264R，D130A/D137Q/G163P/N233Q/L264R，D130A/V154I/G156W/V187N/T189Q，E056K/L075G/V187N/I252Q/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/V187N/I252Q/L264R，G023K/E056K/D130A/V187N/L264R，G023K/E056K/D130A/V187T/L264R，G023K/L075R/D130A/V187N/L264R，G023Q/E056K/V187N/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，L075G/D130A/V187T/I252Q/L264R，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/K024A/V077I/V154I/V187N，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/D111A/V187N/T189Q/L264R，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D048Q/D130A/D137H/G163P/L264R，D027E/S058M/D130A/G163P/L227M/L264R，D027E/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/S058M/D130A/G163P/L227M/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，F051T/E056K/D130A/V187N/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/L075Q/D130A/V187T/I252Q，G023Q/F051T/G091Q/V187N/I252Q/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/L075G/G091Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/V187N/I252Q/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，L075Q/G091Q/D130A/V187H/I252Q/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，A018K/D111A/D130A/V154I/G156W/V187T/T189Q，A018K/K024A/D130A/V154I/G156W/V187T/L264R，A018K/K024A/N094R/D130A/G156W/V187T/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264RD027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/S058M/L075Q/V187N/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/D027S/F051T/E056K/D130A/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/S058M/L075Q/V187H/I252Q/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/1252Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R
表7-4：下面示出，相比参考序列SEQIDNO：2，在织物粘附性测定中性能指数＞1的TLL变体。
D111A/L264R，D130A/L264R，D130A/V187N，D130A/V187T，G023K/L264R，L075R/L264R，S058M/L264R，V187T/L264R，A018K/A049V/V187T，A018K/K024A/V154I，A018K/N033D/T189D，D111A/D130A/L264R，D130A/V187N/L264R，D130A/V187T/L264R，F051T/I252Q/L264R，G023K/D130A/L264R，G023K/E056K/V187T，G023K/L075R/L264R，G023K/V187T/L264R，G023Q/L075Q/V187T，L075Q/G156W/V187N，L075Q/G156W/V187T，L075Q/V187N/L264R，L075R/D130A/L264R，A018K/D027S/P029E/T189D，A018K/G023K/D111A/T189Q，A018K/G023Q/L075Q/T189D，A018K/G023Q/L075Q/V187T，A018K/G023Q/L075R/D130A，D027E/D130A/N233Q/L264R，D027E/G163P/L227M/L264R，D027S/D130A/I252Q/L264R，D048Q/D130A/G163P/L264R，D111A/D130A/V154I/G156W，D111A/D130A/V187T/L264R，E056K/D130A/V187N/L264R，E056K/D130A/V187T/L264R，F051T/L075G/I252Q/L264R，F051T/L075Q/I252Q/L264R，G023K/D130A/V187N/L264R，G023K/D130A/V187T/T189Q，G023K/E056K/L075R/L264R，G023K/K024A/V154I/V187N，G023K/L075Q/D130A/V187N，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/L075Q/D130A/L264R，G023Q/L075Q/V187T/L264R，G091Q/V187T/I252Q/L264R，K024A/D130A/V154I/V187T，K024A/L075R/G156W/V187N，L075G/D130A/V187T/I252Q，L075G/V187H/I252Q/L264R，L075Q/V077I/D130A/V187Q，P029E/N033D/L075D/T189D，S058M/G163P/L227M/L264R，S058M/L075Q/I252Q/L264R，A018K/D130A/G156W/V187N/L264R，A018K/G023K/K024A/D130A/G156W，A018K/G023K/K024A/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/K024A/D130A/V187Q，A018K/G023Q/L075Q/G156W/V187T，A018K/K024A/N094R/D130A/V187N，A018K/L075Q/D111A/D130A/V187T，D027E/D048Q/D137Q/L227M/L264R，D027E/D130A/G163P/L227M/L264R，D027E/D130A/G163P/N233Q/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L264R，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D027S/L075R/V154I/T189Q/L264R，D027S/P029E/N073R/L075D/T189D，D048Q/S058M/D137Q/N233Q/L264R，D130A/D137Q/G163P/L227M/L264R，D130A/D137Q/G163P/N233Q/L264R，D130A/V154I/G156W/V187N/T189Q，E056K/L075G/V187N/I252Q/L264R，E056K/L075Q/G156W/T189Q/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/G091Q/D130A/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/V187N/I252Q/L264R，G023K/E056K/D130A/V187N/L264R，G023K/E056K/D130A/V187T/L264R，G023K/L075Q/D130A/G156W/L264R，G023K/L075R/D130A/V187N/L264R，G023Q/E056K/V187N/I252Q/L264R，G023Q/K024A/L075Q/G156W/V187N，G023Q/K024A/L075R/G156W/V187Q，G023Q/L075G/G091Q/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，G023Q/L075Q/V077I/G156W/V187N，G091Q/D130A/V187H/I252Q/L264R，K024A/L075Q/D111A/G156W/V187T，L075G/D130A/V187T/I252Q/L264R，L075Q/D111A/V187N/T189Q/L264R，L075R/D130A/V154I/T189Q/L264R，L075R/D130A/V187T/T189Q/L264R，S058M/D130A/G163P/L227M/L264R，A018K/G023K/K024A/L075R/D130A/V187N，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023K/L075Q/D130A/G156W/V187N，A018K/G023K/L075Q/G156W/V187N/L264R，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/K024A/V077I/V154I/V187N，A018K/G023Q/L075Q/G156W/V187T/T189D，A018K/G023Q/L075Q/V077I/G156W/V187N，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/D111A/V187N/T189Q/L264R，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/K024A/L075R/D130A/V187N/T189Q，A018K/L075Q/D130A/V154I/T189Q/L264R，A018K/L075Q/D130A/V187N/T189Q/L264R，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D048Q/D130A/D137H/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L227M/L264R，D027E/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/F051T/L075Q/D130A/V187H/L264R，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/S058M/D130A/G163P/L227M/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，F051T/E056K/D130A/V187N/I252Q/L264R，F051T/E056K/L075Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，F051T/L075R/D130A/V187N/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/L075Q/D111A/D130A/V154I/V187N，G023K/L075Q/N094R/V154I/V187T/L264R，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/L075Q/D130A/V187T/I252Q，G023Q/F051T/G091Q/V187N/I252Q/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/L075G/G091Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/V187N/I252Q/L264R，G023Q/L075R/V154I/G156W/V187N/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，L075Q/D130A/V154I/G156W/V187N/L264R，L075Q/G091Q/D130A/V187H/I252Q/L264R，L075R/D130A/V154I/G156W/V187N/L264R，N011K/E056K/L075Q/D130A/V187N/T189Q，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，A018K/D111A/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/K024A/L075Q/D130A/G156W/V187N，A018K/G023K/K024A/N094R/V154I/V187T/L264R，A018K/G023Q/K024A/L075R/D130A/V154I/V187N，A018K/K024A/D130A/V154I/G156W/V187T/L264R，A018K/K024A/L075Q/D111A/G156W/V187N/L264R，A018K/K024A/L075Q/D130A/G156W/V187T/T189Q，A018K/K024A/N094R/D130A/G156W/V187T/L264R，A018K/L075Q/D111A/V154I/V187T/T189Q/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/L075G/D130A/V187H/I252Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/L075G/G091Q/V187T/I252Q/L264R，D027S/F051T/S058M/L075Q/V187N/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/D027Q/F051T/L075G/D130A/V187H/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q/L264R，G023K/K024A/L075Q/D111A/V154I/G156W/V187N，G023K/K024A/L075Q/D130A/V187T/T189Q/L264R，G023K/K024A/L075Q/V077I/D130A/G156W/V187Q，G023Q/D027S/F051T/E056K/D130A/I252Q/L264R，G023Q/D027S/L075Q/G091Q/V187T/I252Q/L264R，G023Q/E056K/L075Q/D111A/G156W/V187N/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/L075R/G091Q/V187N/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，K024A/L075Q/D111A/D130A/V154I/G156W/L264R，L075Q/D111A/D130A/G156W/V187N/T189Q/L264R，A018K/G023K/K024A/L075Q/D130Y/V154I/V187N/T189Q，A018K/G023K/K024A/L075R/D111A/V154I/V187T/L264R，A018K/G023Q/K024A/L075Q/V077I/D130A/G156W/V187Q，A018K/K024A/L075R/D130A/V154I/G156W/V187T/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/E056K/L075G/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/S058M/L075Q/V187H/I252Q/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，K024A/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，N011K/G023K/D027S/L075R/V154I/G156W/V187N/T189Q，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023Q/D027S/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R
表7-5：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比参考序列SEQIDNO：2在织物粘附性测定中性能指数＞1的TLL变体。
D111A/L264R，D130A/L264R，D130A/V187N，D130A/V187T，G023K/L264R，L075R/L264R，S058M/L264R，V187T/L264R，A018K/A049V/V187T，A018K/K024A/V154I，A018K/N033D/T189D，D111A/D130A/L264R，D130A/V187N/L264R，D130A/V187T/L264R，F051T/I252Q/L264R，G023K/D130A/L264R，G023K/E056K/V187T，G023K/L075R/L264R，G023K/V187T/L264R，L075Q/V187N/L264R，L075R/D130A/L264R，A018K/D027S/P029E/T189D，A018K/G023K/D111A/T189Q，D027E/D130A/N233Q/L264R，D027E/G163P/L227M/L264R，D027S/D130A/I252Q/L264R，D048Q/D130A/G163P/L264R，D111A/D130A/V154I/G156W，D111A/D130A/V187T/L264R，E056K/D130A/V187N/L264R，E056K/D130A/V187T/L264R，F051T/L075G/I252Q/L264R，F051T/L075Q/I252Q/L264R，G023K/D130A/V187N/L264R，G023K/D130A/V187T/T189Q，G023K/E056K/L075R/L264R，G023K/K024A/V154I/V187N，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/L075Q/D130A/L264R，G023Q/L075Q/V187T/L264R，G091Q/V187T/I252Q/L264R，K024A/D130A/V154I/V187T，L075G/D130A/V187T/I252Q，L075G/V187H/I252Q/L264R，S058M/G163P/L227M/L264R，S058M/L075Q/I252Q/L264R，A018K/D130A/G156W/V187N/L264R，A018K/G023K/K024A/D130A/G156W，A018K/G023K/K024A/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/K024A/D130A/V187Q，A018K/K024A/N094R/D130A/V187N，D027F/D048Q/D137Q/L227M/L264R，D027E/D130A/G163P/L227M/L264R，D027E/D130A/G163P/N233Q/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L264R，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D027S/L075R/V154I/T189Q/L264R，D048Q/S058M/D137Q/N233Q/L264R，D130A/D137Q/G163P/L227M/L264R，D130A/D137Q/G163P/N233Q/L264R，D130A/V154I/G156W/V187N/T189Q，E056K/L075G/V187N/I252Q/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/G091Q/D130A/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/V187N/I252Q/L264R，G023K/E056K/D130A/V187N/L264R，G023K/E056K/D130A/V187T/L264R，G023K/L075R/D130A/V187N/L264R，G023Q/E056K/V187N/I252Q/L264R，G023Q/L075G/G091Q/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，L075G/D130A/V187T/I252Q/L264R，S058M/D130A/G163P/L227M/L264R，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/K024A/V077I/V154I/V187N，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/D111A/V187N/T189Q/L264R，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D048Q/D130A/D137H/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L227M/L264R，D027E/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/S058M/D130A/G163P/L227M/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，F051T/E056K/D130A/V187N/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/L075Q/D130A/V187T/I252Q，G023Q/F051T/G091Q/V187N/I252Q/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/L075G/G091Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/V187N/I252Q/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，L075Q/G091Q/D130A/V187H/I252Q/L264R，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，A018K/D111A/D130A/V154I/G156W/V187T/T189Q，A018K/K024A/D130A/V154I/G156W/V187T/L264R，A018K/K024A/N094R/D130A/G156W/V187T/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/S058M/L075Q/V187N/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/D027S/F051T/E056K/D130A/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/S058M/L075Q/V187H/I252Q/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R
表7-6：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比参考序列SEQIDNO：2在织物粘附性测定中性能指数≥最大PI值的50％的TLL变体。
D111A/L264R，D130A/L264R，D130A/V187N，D130A/V187T，G023K/L264R，L075R/L264R，S058M/L264R，V187T/L264R，A018K/A049V/V187T，A018K/K024A/V154I，D111A/D130A/L264R，D130A/V187N/L264R，D130A/V187T/L264R，F051T/I252Q/L264R，G023K/D130A/L264R，G023K/E056K/V187T，G023K/L075R/L264R，G023K/V187T/L264R，L075Q/V187N/L264R，L075R/D130A/L264R，A018K/G023K/D111A/T189Q，D027E/D130A/N233Q/L264R，D027S/D130A/I252Q/L264R，D048Q/D130A/G163P/L264R，D111A/D130A/V187T/L264R，E056K/D130A/V187N/L264R，F051T/L075G/I252Q/L264R，F051T/L075Q/I252Q/L264R，G023K/D130A/V187N/L264R，G023K/D130A/V187T/T189Q，G023K/E056K/L075R/L264R，G023K/K024A/V154I/V187N，G023Q/F051T/I252Q/L264R，G023Q/G091Q/I252Q/L264R，G023Q/L075Q/D130A/L264R，G023Q/L075Q/V187T/L264R，G091Q/V187T/I252Q/L264R，K024A/D130A/V154I/V187T，L075G/D130A/V187T/I252Q，S058M/G163P/L227M/L264R，S058M/L075Q/I252Q/L264R，A018K/D130A/G156W/V187N/L264R，A018K/G023K/K024A/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/K024A/D130A/V187Q，A018K/K024A/N094R/D130A/V187N，D027E/D048Q/D137Q/L227M/L264R，D027E/D130A/G163P/L227M/L264R，D027E/D130A/G163P/N233Q/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L264R，D027S/F051T/V187N/I252Q/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D027S/L075R/V154I/T189Q/L264R，D048Q/S058M/D137Q/N233Q/L264R，D130A/D137Q/G163P/L227M/L264R，D130A/D137Q/G163P/N233Q/L264R，D130A/V154I/G156W/V187N/T189Q，E056K/L075G/V187N/I252Q/L264R，F051T/D130A/V187T/I252Q/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/V187N/I252Q/L264R，G023K/E056K/D130A/V187N/L264R，G023K/E056K/D130A/V187T/L264R，G023K/L075R/D130A/V187N/L264R，G023Q/E056K/V187N/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/I252Q/L264R，L075G/D130A/V187T/I252Q/L264R，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/K024A/V077I/V154I/V187N，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/D111A/V187N/T189Q/L264R，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D048Q/D130A/D137H/G163P/L264R，D027E/S058M/D130A/G163P/L227M/L264R，D027E/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/S058M/D130A/G163P/L227M/L264R，D048Q/S058M/D130A/L227M/N233Q/L264R，F051T/E056K/D130A/V187N/I252Q/L264R，F051T/L075G/G091Q/V187H/I252Q/L264R，F051T/L075G/G091Q/V187T/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/L075Q/D130A/I252Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，G023Q/D027S/F051T/L075Q/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/L075Q/D130A/V187T/I252Q，G023Q/F051T/G091Q/V187N/I252Q/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/V077I/D130A/G156W/V187N，G023Q/L075G/G091Q/D130A/I252Q/L264R，G023Q/L075Q/G091Q/V187N/I252Q/L264R，K024A/L075Q/D111A/V154I/V187N/T189Q，L075Q/G091Q/D130A/V187H/I252Q/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，A018K/D111A/D130A/V154I/G156W/V187T/T189Q，A018K/K024A/D130A/V154I/G156W/V187T/L264R，A018K/K024A/N094R/D130A/G156W/V187T/L264R，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/G163P/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/S058M/L075Q/V187N/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/D027S/F051T/E056K/D130A/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/F051T/S058M/G091Q/D130A/I252Q/L264R，G023Q/L075G/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/S058M/L075Q/V187H/I252Q/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075G/G091Q/V187T/L264R，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，G023Q/F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023Q/S058M/L075R/G091Q/D130A/V187H/I252Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023Q/F051T/S058M/L075Q/G091Q/D130A/V187H/I252Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R
表7-7：下面示出，相比TLL(SEQIDNO：1)，在织物粘附性测定中性能指数＜1的TLL变体。
A018K/D027S，A018K/E045F，A018K/L075D，A018K/T189D，D027E/G163P，D027N/E056K，D027N/N233Q，D027S/N033D，D027S/P256T，D130A/T189Q，E045F/N073R，E045F/T189D，F051T/L075R，G023K/D130A，G023Q/L075Q，K024A/L075Q，K024A/V154I，L075Q/D130A，L075Q/G156W，L227M/L264R，N011K/D027S，N033D/E045F，N073R/L075D，N094R/G156W，P029E/N033D，A018K/A049V/L075Q，A018K/D027S/E045F，A018K/D027S/N073R，A018K/D027S/T189D，A018K/E045F/N073R，A018K/E045F/T189D，A018K/E045F/V187T，A018K/G023K/G156W，A018K/G023K/L075Q，A018K/L075D/T189D，A018K/L075Q/V187T，A018K/N033D/L075D，A018K/N073R/L075D，A018K/V154I/G156W，A049V/L075Q/T189D，A049V/V187T/T189D，D027E/S058M/G163P，D027N/N233Q/P256T，D027N/S058M/P256T，D027Q/N233Q/P256T，D027Q/S058M/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/N073R/L075D，D027S/N233Q/P256T，D027S/S058M/P256T，D130A/T189Q/L264R，D130A/V154I/G156W，E045F/L075D/T189D，E045F/N073R/L075D，E056K/D130A/L264R，E056K/D130A/T189Q，G023Q/D111A/L264R，G023Q/D130A/G156W，G023Q/E045F/V077I，G023Q/E056K/L075R，G023Q/K024A/G156W，G023Q/L075Q/V077I，I090F/N233Q/P256T，K024A/D130A/V154I，K024A/L075Q/G156W，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/D130A/V187T，L075Q/N094R/V154I，L075Q/V077I/T189D，L075Q/V187T/L264R，L075R/V187N/L264R，L075R/V187T/L264R，N011K/D027S/S058M，N011K/N233Q/P256T，N033D/E045F/L075D，N033D/E045F/N073R，N033D/N073R/T189D，N094R/D130A/V187T，P029E/E045F/L075D，P029E/N033D/E045F，P029E/N033D/L075D，P029E/N073R/L075D，Q004D/D027S/P256T，Q004D/N233Q/P256T，Q004D/S058M/N233Q，S058M/L227M/L264R，V077I/D130A/V154I，V077I/V187A/T189D，V187N/T189Q/L264R，A018K/A049V/L075Q/T189D，A018K/A049V/L075Q/V187T，A018K/D027S/E045F/N073R，A018K/D027S/E045F/T189D，A018K/D027S/L075D/T189D，A018K/D027S/N033D/L075D，A018K/D111A/G156W/T189Q，A018K/D130A/G156W/V187T，A018K/E045F/A049V/G156W，A018K/E045F/A049V/V187T，A018K/E045F/L075D/T189D，A018K/E045F/L075Q/T189D，A018K/E045F/L075Q/V187T，A018K/G023Q/A049V/T189D，A018K/G023Q/E045F/T189D，A018K/G023Q/E045F/V187T，A018K/G023Q/G156W/V187T，A018K/G023Q/L075Q/V077I，A018K/G023Q/V077I/G156W，A018K/G023Q/V077I/V187T，A018K/G023Q/V187T7T189D，A018K/L075Q/G156W/T189D，A018K/L075Q/G156W/V187T，A018K/L075Q/N094R/D111A，A018K/L075Q/N094R/D130A，A018K/L075Q/N094R/V187Q，A018K/L075Q/V077I/N094R，A018K/L075Q/V187T/T189D，A018K/N033D/L075D/T189D，A018K/N073R/L075D/T189D，A018K/P029E/N033D/L075D，A018K/P029E/N033D/T189D，A018K/P029E/N073R/L075D，A018K/V154I/G156W/V187T，A049V/L075Q/V077I/T189D，A049V/L075Q/V187T/T189D，D027E/D048Q/G163P/L264R，D027E/D137Q/G163P/L227M，D027E/D137Q/L227M/L264R，D027E/L227M/N233Q/L264R，D027N/E056K/N233Q/P256T，D027N/E056K/S058M/P256T，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027Q/S058M/I090F/P256T，D027S/E045F/L075D/T189D，D027S/E045F/N073R/L075D，D027S/E045F/N073R/T189D，D027S/E056K/D111A/V154I，D027S/E056K/I090F/P256T，D027S/N033D/E045F/N073R，D027S/N033D/L075D/T189D，D027S/N033D/N073R/L075D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/L075D，D027S/P029E/E045F/N073R，D027S/P029E/E045F/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D027S/P029E/N033D/N073R，D027S/P029E/N033D/T189D，D048Q/D137Q/G163P/L227M，D111A/D130A/V154I/L264R，D130A/G163P/L227M/L264R，D130A/G163P/N233Q/L264R，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，E056K/L075Q/V187N/L264R，F051T/D130A/I252Q/L264R，G023K/D027Q/F051T/L075Q，G023K/D027S/L075Q/L264R，G023K/D111A/D130A/L264R，G023K/D130A/G156W/T189Q，G023K/E056K/D130A/L264R，G023K/E056K/D130A/V187N，G023K/E056K/V187T/L264R，G023K/L075Q/G156W/L264R，G023K/L075Q/G156W/V187N，G023K/L075R/D130A/L264R，G023K/L075R/V187N/L264R，G023Q/A049V/L075Q/V077I，G023Q/A049V/V077I/G156W，G023Q/D027S/D111A/G156W，G023Q/E045F/A049V/T189D，G023Q/E045F/A049V/V187T，G023Q/E045F/L075Q/G156W，G023Q/E045F/L075Q/V187T，G023Q/K024A/D130A/G156W，G023Q/K024A/L075R/G156W，G023Q/K024A/L075R/V154I，G023Q/K024A/V077I/G156W，G023Q/L075Q/G156W/V187T，G023Q/V154I/G156W/V187N，K024A/L075Q/D130A/G156W，K024A/L075Q/D130A/V154I，K024A/L075Q/V187T/T189Q，L075G/G091Q/V187N/L264R，L075Q/D130A/V154I/G156W，L075Q/G156W/V187T/L264R，L075Q/V077I/G156W/V187N，L075Q/V077I/N094R/G156W，L075Q/V077I/V187T/T189D，L075Q/V187N/T189Q/L264R，L075R/D130A/V187T/L264R，N011K/D027N/E056K/S058M，N011K/D027N/S058M/P256T，N011K/D027Q/S058M/P256T，N011K/D027S/E056K/P256T，N011K/D027S/N233Q/P256T，N011K/D027S/S058M/N233Q，N011K/E056K/N233Q/P256T，N011K/E056K/S058M/P256T，N011K/G023K/L075Q/D111A，N011K/S058M/N233Q/P256T，N033D/E045F/N073R/T189A，P029E/E045F/L075D/T189D，P029E/E045F/N073R/L075D，P029E/E045F/N073R/T189D，P029E/N033D/E045F/L075D，P029E/N073R/L075D/T189D，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/D027S/I090F/P256T，Q004D/N011K/D027N/P256T，Q004D/N011K/D027Q/N233Q，Q004D/N011K/D027S/P256T，Q004D/N011K/E056K/S058M，S058M/D137Q/G163P/N233Q，A018K/A049V/L075Q/V187T/T189D，A018K/D027S/E045F/L075D/T189D，A018K/D027S/E045F/N073R/D137V，A018K/D027S/N033D/E045F/T189D，A018K/D027S/N033D/L075D/T189D，A018K/D027S/N033D/N073R/T189D，A018K/D027S/N073R/L075D/T189D，A018K/D027S/P029E/E045F/T189D，A018K/D027S/P029E/N033D/L075D，A018K/D027S/P029E/N073R/L075D，A018K/D130A/G156W/V187T/L264R，A018K/E045F/A049V/L075Q/G156W，A018K/E045F/A049V/L075Q/T189D，A0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N233Q/P256T，N011K/D027S/D111A/D130A/V154I/V187N/T189Q，N011K/E056K/L075Q/D111A/D130A/V154I/L264R，N011K/E056K/L075Q/D130A/V154I/V187N/T189Q，N011K/G023K/L075Q/D111A/D130A/V154I/V187N，N011K/G023Q/L075Q/V154I/V187N/T189Q/L264R，Q004D/D027N/E056K/S058M/I090F/N233Q/P256T，Q004D/N011K/D027N/S058M/I090F/N233Q/P256T，Q004D/N011K/D027Q/E056K/S058M/N233Q/P256T，Q004D/N011K/D027Q/S058M/I090F/N233Q/P256T，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023K/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/G023K/L075R/V077I/N094R/D130A/G156W/V187Q，A018K/G023Q/E045F/L075Q/V077I/G156W/V187T/T189D，A018K/G023Q/K024A/L075R/D130A/V154I/G156W/V187N，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，A018K/G023Q/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/K024A/L075Q/V077I/N094R/D130A/V154I/G156W，A018K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/D027S/F051T/E056K/L075R/D130A/V187T/L264R，G023K/E056K/L075Q/D130A/V154I/V187N/T189Q/L264R，G023K/F051T/E056K/L075R/G091Q/D130A/V187T/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187T/L264R，G023K/K024A/L075R/N094R/V154I/G156W/V187T/T189Q，G023K/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，G023K/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/D027S/E056K/S058M/L075R/G091Q/V187N/I252Q，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/E056K/L075Q/D111A/V154I/V187N/T189Q/L264R，G023Q/E056K/L075Q/N094R/D111A/G156W/T189Q/L264R，G023Q/K024A/L075Q/V077I/D130A/V154I/G156W/V187N，G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/V187N，G023Q/L075Q/D111A/D130A/V154I/G156W/V187N/L264R，G023Q/L075Q/N094R/D130A/V154I/G156W/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，N011K/G023K/D027S/L075Q/D111A/D130A/V154I/T189Q，N011K/G023K/E056K/L075Q/D130A/V154I/T189Q/L264R，N011K/G023K/L075Q/D111A/G156W/V187N/T189Q/L264R，N011K/G023Q/D027S/L075Q/N094R/V154I/G156W/T189Q，N011K/G023Q/D027S/N094R/V154I/G156W/T189Q/L264R，N011K/G023Q/L075Q/D111A/D130A/V154I/V187N/L264R，N011K/G023Q/L075Q/N094R/V154I/G156W/T189Q/L264R，A018K/G023K/K024A/L075Q/N094R/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/V154I/G156W/V187N，A018K/G023Q/D027S/L075Q/V077I/R108K/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/G156W/V187H，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/P029E/S058M/V077I/R108K/H135F/G156W，A018K/G023Q/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W/V187N，A018K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V187N/T189Q/L264R，G023K/D027S/F051T/E056K/S058M/L075Q/G091Q/V187N/L264R，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/D027S/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023Q/D027S/F051T/E056K/L075G/G091Q/V187N/I252Q/L264R，G023Q/D027S/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，G023Q/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，N011K/G023K/D027S/E056K/L075R/D111A/G156W/V187N/L264R，N011K/G023Q/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/E045F/A049V/S058M/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/S058M/L075Q/V077I/R108K/H135F/G156W/V187H，G023Q/D027S/F051T/E056K/L075R/G091Q/D130A/V187H/I252Q/L264R，G023Q/D027S/F051T/E056K/S058M/L075R/G091Q/D130A/V187H/I252Q，G023Q/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，N011K/G023K/L075R/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，N011K/G023Q/D027S/E056K/L075R/D130A/V154I/G156W/T189Q/L264R，N011K/G023Q/L075Q/N094R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T，N011K/G023Q/D027S/L075Q/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/H135F/V187T/T189D，A018K/G023Q/D027S/P029E/N033D/E045F/A049V/S058M/N073S/K074S/L075Q/V077I/N101D/R108K/H135F/D137V/G156W/V187T/T189D
表7-8：下面示出，相比TLL(SEQIDNO：1)，表达性能指数≥0.5，且在织物粘附性测定中性能指数＜1的TLL变体。
A018K/E045F，A018K/L075D，D027E/G163P，D027S/N033D，D027S/P256T，E045F/N073R，F051T/L075R，K024A/V154I，L227M/L264R，N033D/E045F，N094R/G156W，P029E/N033D，A018K/E045F/N073R，A018K/E045F/V187T，A018K/L075D/T189D，A018K/V154I/G156W，D027E/S058M/G163P，D027N/S058M/P256T，D027Q/S058M/P256T，D027S/L075Q/G091Q，D027S/N033D/T189D，D027S/S058M/P256T，D130A/V154I/G156W，E056K/D130A/L264R，G023Q/D130A/G156W，G023Q/K024A/G156W，I090F/N233Q/P256T，K024A/D130A/V154I，L075G/D130A/V187H，L075Q/V187T/L264R，L075R/V187N/L264R，L075R/V187T/L264R，N011K/N233Q/P256T，N094R/D130A/V187T，P029E/N033D/E045F，Q004D/D027S/P256T，Q004D/S058M/N233Q，V187N/T189Q/L264R，A018K/D111A/G156W/T189Q，A018K/D130A/G156W/V187T，A018K/E045F/A049V/G156W，A018K/E045F/L075D/T189D，A018K/G023Q/V077I/G156W，A018K/L075Q/V077I/N094R，A018K/N033D/L075D/T189D，A018K/P029E/N073R/L075D，D027E/D048Q/G163P/L264R，D027E/D137Q/G163P/L227M，D027E/D137Q/L227M/L264R，D027E/L227M/N233Q/L264R，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/E045F/L075D/T189D，D027S/E045F/N073R/T189D，D027S/N033D/E045F/N073R，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/N073R，D027S/P029E/E045F/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D137Q/G163P/L227M，D130A/G163P/L227M/L264R，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，F051T/D130A/I252Q/L264R，G023K/D027Q/F051T/L075Q，G023K/D130A/G156W/T189Q，G023K/E056K/D130A/V187N，G023K/E056K/V187T/L264R，G023K/L075R/D130A/L264R，G023Q/A049V/V077I/G156W，G023Q/K024A/D130A/G156W，G023Q/K024A/L075R/G156W，G023Q/K024A/V077I/G156W，L075G/G091Q/V187N/L264R，L075R/D130A/V187T/L264R，N011K/D027Q/S058M/P256T，N011K/D027S/N233Q/P256T，N033D/E045F/N073R/T189A，P029E/N073R/L075D/T189D，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/N011K/D027S/P256T，S058M/D137Q/G163P/N233Q，A018K/D027S/E045F/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/E045F/N073R/L075D/T189D，A018K/G023K/D130A/V154I/G156W，A018K/G023Q/K024A/L075Q/G156W，A018K/G023Q/V077I/D130A/G156W，A018K/K024A/L075R/D130A/V154I，A018K/P029E/N033D/N073R/L075D，D027E/D048Q/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L264R，D027E/S058M/D137Q/G163P/N233Q，D027Q/F051T/L075Q/D130A/L264R，D027S/E056K/D111A/V187N/L264R，D048Q/D137Q/G163P/N233Q/L264R，D048Q/S058M/D130A/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D048Q/S058M/G163P/L227M/L264R，D048Q/S058M/G163P/N233Q/L264R，E045F/A049V/L075Q/V187T/T189D，E056K/L075R/D130A/V187N/L264R，E056K/L075R/D130A/V187T/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/D027S/E056K/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187N/L264R，G023K/K024A/D111A/D130A/V187T，G023K/K024A/D111A/G156W/T189Q，G023K/L075Q/V187H/I252Q/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/D027S/L075G/I252Q/L264R，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075Q/V077I/D130A/G156W，N011K/D027N/S058M/N233Q/P256T，N011K/D027S/I090F/N233Q/P256T，Q004D/N011K/D027N/N233Q/P256T，S058M/D130A/D137Q/G163P/L264R，S058M/D137Q/G163P/L227M/L264R，S058M/D137Q/G163P/L227M/N233Q，S058M/G163P/L227M/N233Q/L264R，V077I/D130A/V154I/G156W/V187N，A018K/D027S/N033D/E045F/N073R/L075D，A018K/D027S/N033D/N073R/L075D/T189D，A018K/D027S/P029E/N033D/E045F/L075D，A018K/G023Q/K024A/L075R/N094R/G156W，D027E/D048Q/D137Q/G163P/N233Q/L264R，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/D130A/L227M/N233Q，D027E/D048Q/S058M/G163P/L227M/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027S/E056K/S058M/V187T/I252Q/L264R，D048Q/D130A/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L264R，D048Q/S058M/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/F051T/G091Q/D130A/V187H/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/L075G/D130A/V187T/I252Q/L264R，G023Q/D027S/N094R/V154I/G156W/T189Q，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/K024A/V077I/D130A/V154I/G156W，K024A/L075Q/N094R/D130A/V154I/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，N011K/D027S/E056K/I090F/N233Q/P256T，Q004D/N011K/D027N/I090F/N233Q/P256T，Q004D/N011K/D027N/S058M/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/S058M/G091Q/V187N/I252Q，G023K/K024A/N094R/V154I/G156W/V187N/T189Q，G023Q/F051T/E056K/L075R/G091Q/V187H/I252Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，N011K/G023Q/L075Q/V154I/V187N/T189Q/L264R，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，D027E/D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/1252Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T
表7-9：下面示出，相比TLL(SEQIDNO：1)，表达性能指数≥0.5，且在织物粘附性测定中性能指数≤最大PI值的50％的TLL变体。
A018K/E045F，A018K/L075D，D027E/G163P，D027S/P256T，E045F/N073R，F051T/L075R，K024A/V154I，L227M/L264R，N033D/E045F，A018K/E045F/N073R，A018K/E045F/V187T，A018K/V154I/G156W，D027N/S058M/P256T，D027Q/S058M/P256T，D027S/L075Q/G091Q，D027S/S058M/P256T，E056K/D130A/L264R，G023Q/K024A/G156W，I090F/N233Q/P256T，N011K/N233Q/P256T，N094R/D130A/V187T，P029E/N033D/E045F，Q004D/D027S/P256T，Q004D/S058M/N233Q，A018K/D111A/G156W/T189Q，A018K/E045F/A049V/G156W，A018K/E045F/L075D/T189D，A018K/P029E/N073R/L075D，D027E/D137Q/L227M/L264R，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/E045F/L075D/T189D，D027S/E045F/N073R/T189D，D027S/N033D/E045F/N073R，D027S/N033D/L075D/T189D，D027S/P029E/E045F/N073R，D027S/P029E/E045F/T189D，D027S/P029E/N033D/E045F，D048Q/D137Q/G163P/L227M，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，G023K/E056K/D130A/V187N，G023K/E056K/V187T/L264R，G023Q/K024A/L075R/G156W，L075G/G091Q/V187N/L264R，N011K/D027Q/S058M/P256T，N011K/D027S/N233Q/P256T，N033D/E045F/N073R/T189A，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/N011K/D027S/P256T，A018K/D027S/E045F/L075D/T189D，A018K/E045F/N073R/L075D/T189D，A018K/K024A/L075R/D130A/V154I，D027E/D130A/D137Q/G163P/L264R，D027Q/F051T/L075Q/D130A/L264R，D027S/E056K/D111A/V187N/L264R，D048Q/D137Q/G163P/N233Q/L264R，D048Q/S058M/G163P/L227M/L264R，E045F/A049V/L075Q/V187T/T189D，E056K/L075R/D130A/V187N/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/L075R/V187N/L264R，G023K/K024A/D111A/D130A/V187T，G023K/L075Q/V187H/I252Q/L264R，G023Q/D027S/L075G/I252Q/L264R，G023Q/L075Q/V077I/D130A/G156W，N011K/D027N/S058M/N233Q/P256T，N011K/D027S/I090F/N233Q/P256T，Q004D/N011K/D027N/N233Q/P256T，S058M/D137Q/G163P/L227M/N233Q，V077I/D130A/V154I/G156W/V187N，A018K/D027S/N033D/E045F/N073R/L075D，A018K/D027S/N033D/N073R/L075D/T189D，A018K/D027S/P029E/N033D/E045F/L075D，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/D130A/L227M/N233Q，D027Q/E056K/S058M/D130A/I252Q/L264R，D027S/E056K/S058M/V187T/I252Q/L264R，D048Q/D130A/G163P/L227M/N233Q/L264R，D048Q/S058M/G163P/L227M/N233Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/L075G/D130A/V187T/I252Q/L264R，G023Q/D027S/N094R/V154I/G156W/T189Q，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/K024A/V077I/D130A/V154I/G156W，N011K/D027S/E056K/I090F/N233Q/P256T，Q004D/N011K/D027N/S058M/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，D048Q/S058M/D130A/D137Q/L227M/N233Q/L264R，G023Q/F051T/E056K/L075R/G091Q/V187H/I252Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，N011K/G023Q/L075Q/V154I/V187N/T189Q/L264R，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G0230/D027S/E045F/A049V/S058M/N073S/L0750/R108K/V187T/T189D
表7-10：下面示出，相比参考序列SEQIDNO：2，在织物粘附性测定中性能指数＜1的TLL变体。
A018K/D027S，A018K/E045F，A018K/L075D，A018K/T189D，D027E/G163P，D027N/E056K，D027N/N233Q，D027S/P256T，D130A/T189Q，E045F/N073R，E045F/T189D，F051T/L075R，G023K/D130A，G023Q/L075Q，K024A/V154I，L227M/L264R，N011K/D027S，N033D/E045F，N073R/L075D，A018K/A049V/L075Q，A018K/D027S/E045F，A018K/D027S/N073R，A018K/D027S/T189D，A018K/E045F/N073R，A018K/E045F/T189D，A018K/E045F/V187T，A018K/G023K/G156W，A018K/G023K/L075Q，A018K/N033D/L075D，A018K/N073R/L075D，A018K/V154I/G156W，A049V/L075Q/T189D，A049V/V187T/T189D，D027N/N233Q/P256T，D027N/S058M/P256T，D027Q/N233Q/P256T，D027Q/S058M/P256T，D027S/L075Q/G091Q，D027S/N073R/L075D，D027S/N233Q/P256T，D027S/S058M/P256T，D130A/V154I/G156W，E045F/L075D/T189D，E045F/N073R/L075D，E056K/D130A/L264R，E056K/D130A/T189Q，G023Q/D111A/L264R，G023Q/E045F/V077I，G023Q/E056K/L075R，G023Q/K024A/G156W，G023Q/L075Q/V077I，I090F/N233Q/P256T，K024A/L075Q/G156W，L075Q/D130A/V187T，L075Q/N094R/V154I，L075Q/V077I/T189D，N011K/D027S/S058M，N011K/N233Q/P256T，N033D/E045F/L075D，N033D/E045F/N073R，N033D/N073R/T189D，N094R/D130A/V187T，P029E/E045F/L075D，P029E/N033D/E045F，P029E/N033D/L075D，P029E/N073R/L075D，Q004D/D027S/P256T，Q004D/N233Q/P256T，Q004D/S058M/N233Q，S058M/L227M/L264R，V077I/D130A/V154I，V077I/V187A/T189D，A018K/A049V/L075Q/T189D，A018K/D027S/E045F/N073R，A018K/D027S/E045F/T189D，A018K/D027S/L075D/T189D，A018K/D027S/N033D/L075D，A018K/D111A/G156W/T189Q，A018K/E045F/A049V/G156W，A018K/E045F/A049V/V187T，A018K/E045F/L075D/T189D，A018K/E045F/L075Q/T189D，A018K/E045F/L075Q/V187T，A018K/G023Q/A049V/T189D，A018K/G023Q/E045F/T189D，A018K/G023Q/G156W/V187T，A018K/G023Q/V077I/V187T，A018K/L075Q/G156W/T189D，A018K/L075Q/G156W/V187T，A018K/L075Q/N094R/D111A，A018K/L075Q/N094R/D130A，A018K/L075Q/V187T/T189D，A018K/N073R/L075D/T189D，A018K/P029E/N033D/T189D，A018K/P029E/N073R/L075D，A018K/V154I/G156W/V187T，A049V/L075Q/V077I/T189D，A049V/L075Q/V187T/T189D，D027E/D137Q/L227M/L264R，D027N/E056K/N233Q/P256T，D027N/E056K/S058M/P256T，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027Q/S058M/I090F/P256T，D027S/E045F/L075D/T189D，D027S/E045F/N073R/L075D，D027S/E045F/N073R/T189D，D027S/E056K/D111A/V154I，D027S/E056K/I090F/P256T，D027S/N033D/E045F/N073R，D027S/N033D/L075D/T189D，D027S/N033D/N073R/L075D，D027S/P029E/E045F/L075D，D027S/P029E/E045F/N073R，D027S/P029E/E045F/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/N073R，D048Q/D137Q/G163P/L227M，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，G023K/D027S/L075Q/L264R，G023K/D111A/D130A/L264R，G023K/E056K/D130A/L264R，G023K/E056K/D130A/V187N，G023K/E056K/V187T/L264R，G023K/L075Q/G156W/L264R，G023K/L075R/V187N/L264R，G023Q/A049V/L075Q/V077I，G023Q/D027S/D111A/G156W，G023Q/E045F/A049V/T189D，G023Q/E045F/A049V/V187T，G023Q/E045F/L075Q/G156W，G023Q/E045F/L075Q/V187T，G023Q/K024A/L075R/G156W，G023Q/V154I/G156W/V187N，K024A/L075Q/D130A/G156W，K024A/L075Q/D130A/V154I，L075G/G091Q/V187N/L264R，L075Q/D130A/V154I/G156W，L075Q/G156W/V187T/L264R，L075Q/V077I/G156W/V187N，L075Q/V077I/V187T/T189D，L075Q/V187N/T189Q/L264R，N011K/D027N/E056K/S058M，N011K/D027N/S058M/P256T，N011K/D027Q/S058M/P256T，N011K/D027S/E056K/P256T，N011K/D027S/N233Q/P256T，N011K/D027S/S058M/N233Q，N011K/E056K/N233Q/P256T，N011K/E056K/S058M/P256T，N011K/G023K/L075Q/D111A，N011K/S058M/N233Q/P256T，N033D/E045F/N073R/T189A，P029E/E045F/L075D/T189D，P029E/E045F/N073R/L075D，P029E/E045F/N073R/T189D，P029E/N033D/E045F/L075D，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/D027S/I090F/P256T，Q004D/N011K/D027N/P256T，Q004D/N011K/D027Q/N233Q，Q004D/N011K/D027S/P256T，Q004D/N011K/E056K/S058M，A018K/A049V/L075Q/V187T/T189D，A018K/D027S/E045F/L075D/T189D，A018K/D027S/E045F/N073R/D137V，A018K/D027S/N033D/E045F/T189D，A018K/D027S/N033D/N073R/T189D，A018K/D027S/N073R/L075D/T189D，A018K/D027S/P029E/E045F/T189D，A018K/D027S/P029E/N073R/L075D，A018K/D130A/G156W/V187T/L264R，A018K/E045F/A049V/L075Q/G156W，A018K/E045F/A049V/L075Q/T189D，A018K/E045F/A049V/L075Q/V187T，A018K/E045F/N073R/L075D/T189D，A018K/G023K/L075Q/D130A/V154I，A018K/G023Q/A049V/L075Q/G156W，A018K/G023Q/A049V/L075Q/V077I，A018K/G023Q/A049V/L075Q/V187T，A018K/G023Q/A049V/V077I/V187T，A018K/G023Q/E045F/A049V/L075Q，A018K/G023Q/E045F/A049V/V187T，A018K/G023Q/E045F/L075Q/V187T，A018K/G023Q/E045F/V077I/T189D，A018K/G023Q/K024A/D130A/V1541，A018K/G023Q/V077I/V187T/T189D，A018K/K024A/L075Q/V154I/T189D，A018K/K024A/L075R/D130A/V154I，A018K/L075Q/G156W/V187T/T189D，A018K/L075Q/V187T/T189Q/L264R，A018K/N033D/E045F/L075D/T189D，A018K/P029E/E045F/N073R/L075D，A018K/V077I/G156W/V187T/T189D，D027E/D048Q/L227M/N233Q/L264R，D027E/D048Q/S058M/G163P/N233Q，D027E/D130A/D137Q/G163P/L264R，D027E/D137Q/G163P/L227M/L264R，D027N/E056K/S058M/N233Q/P256T，D027N/S058M/I090F/N233Q/P256T，D027Q/E056K/I090F/N233Q/P256T，D027Q/E056K/S058M/N233Q/P256T，D027Q/F051T/L075Q/D130A/L264R，D027S/E056K/D111A/V187N/L264R，D027S/E056K/S058M/N233Q/P256T，D027S/L075Q/D111A/D130A/V187N，D027S/N033D/E045F/N073R/L075D，D027S/P029E/E045F/N073R/L075D，D027S/P029E/E045F/N073R/T189D，D027S/P029E/N033D/E045F/N073R，D027S/P029E/N033D/N073R/L075D，D048Q/D130A/D137Q/G163P/L264R，D048Q/D137Q/G163P/N233Q/L264R，D048Q/S058M/D137Q/G163P/L264R，D048Q/S058M/G163P/L227M/L264R，D130A/G163P/L227M/N233Q/L264R，E045F/A049V/L075Q/V187T/T189D，E056K/D130A/V187H/I252Q/L264R，E056K/L075R/D130A/V187N/L264R，E056K/N094R/G156W/V187N/L264R，E056K/S058M/I090F/N233Q/P256T，F051T/S058M/L075Q/G091Q/I252Q，G023K/D027S/E056K/V187T/L264R，G023K/D111A/V154I/V187T/L264R，G023K/E056K/L075R/V187N/L264R，G023K/K024A/D111A/D130A/V187T，G023K/K024A/L075R/D130A/G156W，G023K/L075Q/D130A/V154I/T189Q，G023K/L075Q/V187H/I252Q/L264R，G023Q/A049V/L075Q/G156W/T189D，G023Q/A049V/L075Q/V187T/T189D，G023Q/D027S/D111A/T189Q/L264R，G023Q/D027S/L075G/I252Q/L264R，G023Q/D027S/V154I/V187N/L264R，G023Q/E045F/A049V/G156W/T189D，G023Q/E045F/A049V/L075Q/V077I，G023Q/E045F/A049V/V077I/T189D，G023Q/E045F/L075Q/G156W/T189D，G023Q/E045F/L075Q/V077I/V187T，G023Q/K024A/L075Q/D130A/G156W，G023Q/K024A/L075R/D130A/V154I，G023Q/K024A/L075R/V077I/D130A，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/D111A/V154I/V187T，K024A/L075R/G156W/V187N/T189Q，L075Q/D111A/D130A/V187T/T189Q，L075Q/D130A/V187T/T189Q/L264R，L075Q/N094R/D130A/G156W/V187T，L075Q/V077I/D130A/G156W/V187N，L075R/D111A/D130A/V187N/T189Q，L075R/V077I/N094R/V154I/G156W，N011K/D027N/E056K/N233Q/P256T，N011K/D027N/E056K/S058M/P256T，N011K/D027N/S058M/N233Q/P256T，N011K/D027Q/E056K/S058M/N233Q，N011K/D027Q/I090F/N233Q/P256T，N011K/D027Q/S058M/I090F/P256T，N011K/D027Q/S058M/N233Q/P256T，N011K/D027S/E056K/N233Q/P256T，N011K/D027S/I090F/N233Q/P256T，N011K/G023Q/L075Q/V187N/T189Q，P029E/N033D/E045F/L075D/T189D，P029E/N033D/K074S/L075D/T189D，Q004D/D027N/E056K/S058M/P256T，Q004D/D027N/I090F/N233Q/P256T，Q004D/D027Q/E056K/N233Q/P256T，Q004D/D027Q/E056K/S058M/P256T，Q004D/D027S/E056K/N233Q/P256T，Q004D/D027S/E056K/S058M/P256T，Q004D/D027S/S058M/N233Q/P256T，Q004D/N011K/D027N/N233Q/P256T，Q004D/N011K/D027Q/E056K/N233Q，Q004D/N011K/D027Q/S058M/N233Q，Q004D/N011K/I090F/N233Q/P256T，S058M/D137Q/G163P/L227M/N233Q，V077I/D130A/V154I/G156W/V187N，A018K/D027S/N033D/E045F/L075D/T189D，A018K/D027S/N033D/E045F/N073R/L075D，A018K/D027S/N033D/N073R/L075D/T189D，A018K/D027S/P029E/N033D/E045F/L075D，A018K/D027S/P029E/N033D/L075D/T189D，A018K/D027S/P029E/N073R/L075D/T189D，A018K/E045F/A049V/L075Q/V077I/V187T，A018K/G023K/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075R/D111A/L264R，A018K/G023K/L075R/N094R/D130A/V187N，A018K/G023K/L075R/V077I/D130A/V154I，A018K/G023Q/A049V/L075Q/V077I/G156W，A018K/G023Q/A049V/V077I/V187T/T189D，A018K/G023Q/E045F/A049V/G156W/V187T，A018K/G023Q/E045F/A049V/L075Q/T189D，A018K/G023Q/E045F/A049V/L075Q/V077I，A018K/G023Q/E045F/A049V/L075Q/V187T，A018K/G023Q/E045F/G156W/V187T/T189D，A018K/G023Q/E045F/L075Q/G156C/V187T，A018K/G023Q/E045F/L075Q/G156W/V187T，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/G023Q/L075Q/V077I/D130A/G156W，A018K/G023Q/L075Q/V077I/N094R/V154I，A018K/K024A/V077I/N094R/D130A/G156W，A018K/L075Q/D111A/V154I/G156W/V187N，A018K/L075Q/D111A/V154I/T189Q/L264R，A018K/N033D/E045F/N073R/L075D/T189D，A018K/P029E/N033D/E045F/L075D/T189D，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/D130A/L227M/N233Q，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/E056K/S058M/I090F/N233Q/P256T，D027S/E056K/S058M/V187T/I252Q/L264R，D027S/G091Q/D130A/V187N/I252Q/L264R，D027S/L075Q/D111A/G156W/V187N/T189Q，D027S/P029E/N033D/E045F/N073R/L075D，D027S/P029E/N033D/E045F/N073R/T189D，D027S/P029E/N033D/N073R/L075D/T189D，D048Q/D130A/G163P/L227M/N233Q/L264R，D048Q/S058M/G163P/L227M/N233Q/L264R，E045F/A049V/L075Q/V077I/V187T/T189D，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/G156W/V187T/T189Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/L075G/D130A/V187T/I252Q/L264R，G023K/L075Q/V077I/D130A/G156W/V187Q，G023Q/A049V/L075Q/G156W/V187T/T189D，G023Q/D027S/F051T/L075Q/V187T/I252Q，G023Q/D027S/L075Q/D111A/G156W/L264R，G023Q/D027S/L075R/D111A/V187N/L264R，G023Q/D027S/N094R/V154I/G156W/T189Q，G023Q/E045F/A049V/L075Q/G156W/V187T，G023Q/E045F/A049V/L075Q/V077I/V187T，G023Q/E045F/L075Q/V077I/G156W/V187T，G023Q/E045F/L075Q/V077I/V187T/T189D，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/K024A/L075Q/D130A/V154I/G156W，G023Q/K024A/V077I/D130A/V154I/G156W，G023Q/L075Q/N094R/D111A/G156W/L264R，G023Q/L075Q/V077I/N094R/D130A/V187Q，K024A/D111A/D130A/V187T/T189Q/L264R，K024A/L075Q/D130A/G156W/T189Q/L264R，K024A/L075R/D111A/V154I/G156W/V187N，L075Q/D111A/D130A/V154I/G156W/T189Q，L075Q/N094R/D111A/G156W/T189Q/L264R，N011K/D027N/E056K/I090F/N233Q/P256T，N011K/D027N/E056K/S058M/N233Q/P256T，N011K/D027N/S058M/I090F/N233Q/P256T，N011K/D027Q/E056K/S058M/N233Q/P256T，N011K/D027Q/S058M/I090F/N233Q/P256T，N011K/D027S/E056K/I090F/N233Q/P256T，N011K/D027S/E056K/S058M/I090F/P256T，N011K/D027S/E056K/S058M/N233Q/P256T，N011K/D027S/S058M/I090F/N233Q/P256T，N011K/G023K/D027S/V154I/G156W/T189Q，N011K/G023Q/L075Q/D130A/V187N/T189Q，N011K/G023Q/L075Q/N094R/D130A/L264R，P029E/N033D/K074S/L075D/N101D/T189D，Q004D/D027N/E056K/S058M/N233Q/P256T，Q004D/D027S/E056K/S058M/N233Q/P256T，Q004D/N011K/D027N/S058M/N233Q/P256T，Q004D/N011K/D027Q/E056K/S058M/N233Q，Q004D/N011K/D027S/I090F/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，A018K/E045F/L075Q/V077I/G156W/V187T/T189D，A018K/G023Q/E045F/A049V/L075Q/G156W/V187T，A018K/G023Q/E045F/L075Q/G156W/V187T/T189D，A018K/G023Q/E045F/L075Q/V077I/G156W/V187T，A018K/G023Q/K024A/L075R/N094R/G156W/V187N，A018K/L075Q/D111A/D130A/V154I/G156W/L264R，A018K/L075Q/D130A/G156W/V187N/T189Q/L264R，A018K/L075Q/N094R/D111A/V154I/V187T/L264R，D027E/S058M/D130A/D137Q/G163P/N233Q/L264R，D027Q/F051T/L075Q/G091Q/D130A/V187T/I252Q，D027S/E056K/L075R/D111A/G156W/V187N/L264R，D027S/L075R/V154I/G156W/V187N/T189Q/L264R，D027S/P029E/N033D/E045F/N073R/L075D/T189D，D048Q/S058M/D130A/D137Q/L227M/N233Q/L264R，E056K/L075Q/D111A/D130A/G156W/V187N/T189Q，G023K/D027S/L075Q/N094R/V154I/G156W/T189Q，G023K/F051T/E056K/L075R/D130A/V187T/I252Q，G023K/F051T/L075Q/G091Q/D130A/V187H/I252Q，G023K/K024A/L075Q/N094R/V154I/G156W/V187N，G023K/L075Q/D111A/D130A/V154I/G156W/L264R，G023K/L075Q/D111A/V154I/G156W/T189Q/L264R，G023K/L075Q/D130A/G156W/V187N/T189Q/L264R，G023K/L075Q/N094R/D130A/V154I/V187N/L264R，G023Q/D027S/L075Q/V154I/G156W/V187N/L264R，G023Q/E045F/A049V/V077I/G156W/V187T/T189D，G023Q/E045F/L075Q/V077I/G156W/V187T/T189D，G023Q/F051T/E056K/L075R/G091Q/V187H/I252Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，K024A/L075Q/D111A/D130A/V187N/T189Q/L264R，K024A/L075R/D130A/V154I/G156W/T189Q/L264R，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，N011K/D027S/D111A/D130A/V154I/V187N/T189Q，N011K/E056K/L075Q/D111A/D130A/V154I/L264R，N011K/E056K/L075Q/D130A/V154I/V187N/T189Q，N011K/G023K/L075Q/D111A/D130A/V154I/V187N，N011K/G023Q/L075Q/V154I/V187N/T189Q/L264R，Q004D/D027N/E056K/S058M/I090F/N233Q/P256T，Q004D/N011K/D027N/S058M/I090F/N233Q/P256T，Q004D/N011K/D027Q/E056K/S058M/N233Q/P256T，Q004D/N011K/D027Q/S058M/I090F/N233Q/P256T，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/L075R/V077I/N094R/D130A/G156W/V187Q，A018K/G023Q/E045F/L075Q/V077I/G156W/V187T/T189D，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，A018K/G023Q/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/E056K/L075R/D130A/V187T/L264R，G023K/E056K/L075Q/D130A/V154I/V187N/T189Q/L264R，G023K/F051T/E056K/L075R/G091Q/D130A/V187T/L264R，G023K/K024A/L075R/N094R/V154I/G156W/V187T/T189Q，G023K/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，G023Q/D027S/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/E056K/L075Q/D111A/V154I/V187N/T189Q/L264R，G023Q/L075Q/D111A/D130A/V154I/G156W/V187N/L264R，G023Q/L075Q/N094R/D130A/V154I/G156W/T189Q/L264R，N011K/G023K/D027S/L075Q/D111A/D130A/V154I/T189Q，N011K/G023K/E056K/L075Q/D130A/V154I/T189Q/L264R，N011K/G023K/L075Q/D111A/G156W/V187N/T189Q/L264R，N011K/G023Q/D027S/L075Q/N094R/V154I/G156W/T189Q，N011K/G023Q/L075Q/D111A/D130A/V154I/V187N/L264R，A018K/G023K/K024A/L075Q/N094R/D130A/V154I/G156W/V187N，A018K/G023Q/D027S/L075Q/V077I/R108K/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/G156W/V187H，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/P029E/S058M/V077I/R108K/H135F/G156W，A018K/G023Q/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V187N/T189Q/L264R，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/D027S/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023Q/D027S/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，N011K/G023K/D027S/E056K/L075R/D111A/G156W/V187N/L264R，A018K/G023Q/D027S/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/E045F/A049V/S058M/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/S058M/L075Q/V077I/R108K/H135F/G156W/V187H，G023Q/D027S/F051T/E056K/S058M/L075R/G091Q/D130A/V187H/I252Q，G023Q/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，N011K/G023K/L075R/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，N011K/G023Q/D027S/E056K/L075R/D130A/V154I/G156W/T189Q/L264R，N011K/G023Q/L075Q/N094R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，N011K/G023Q/D027S/L075Q/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/H135F/V187T/T189D，A018K/G023Q/D027S/P029E/N033D/E045F/A049V/S058M/N073S/K074S/L075Q/V077I/N101D/R108K/H135F/D137V/G156W/V187T/T189D
表7-11：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比参考序列SEQIDNO：2在织物粘附性测定中性能指数＜1的TLL变体。
A018K/E045F，A018K/L075D，D027E/G163P，D027S/P256T，E045F/N073R，F051T/L075R，K024A/V154I，L227M/L264R，N033D/E045F，A018K/E045F/N073R，A018K/E045F/V187T，A018K/V154I/G156W，D027N/S058M/P256T，D027Q/S058M/P256T，D027S/L075Q/G091Q，D027S/S058M/P256T，D130A/V154I/G156W，E056K/D130A/L264R，G023Q/K024A/G156W，I090F/N233Q/P256T，N011K/N233Q/P256T，N094R/D130A/V187T，P029E/N033D/E045F，Q004D/D027S/P256T，Q004D/S058M/N233Q，A018K/D111A/G156W/T189Q，A018K/E045F/A049V/G156W，A018K/E045F/L075D/T189D，A018K/P029E/N073R/L075D，D027E/D137Q/L227M/L264R，D027N/S058M/N233Q/P256T，D027Q/E056K/N233Q/P256T，D027S/E045F/L075D/T189D，D027S/E045F/N073R/T189D，D027S/N033D/E045F/N073R，D027S/N033D/L075D/T189D，D027S/P029E/E045F/N073R，D027S/P029E/E045F/T189D，D027S/P029E/N033D/E045F，D048Q/D137Q/G163P/L227M，E045F/A049V/G156W/V187T，E045F/L075Q/G156W/V187T，G023K/E056K/D130A/V187N，G023K/E056K/V187T/L264R，G023Q/K024A/L075R/G156W，L075G/G091Q/V187N/L264R，N011K/D027Q/S058M/P256T，N011K/D027S/N233Q/P256T，N033D/E045F/N073R/T189A，Q004D/D027N/E056K/P256T，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/N011K/D027S/P256T，A018K/D027S/E045F/L075D/T189D，A018K/E045F/N073R/L075D/T189D，A018K/K024A/L075R/D130A/V154I，D027E/D130A/D137Q/G163P/L264R，D027Q/F051T/L075Q/D130A/L264R，D027S/E056K/D111A/V187N/L264R，D048Q/D137Q/G163P/N233Q/L264R，D048Q/S058M/G163P/L227M/L264R，E045F/A049V/L075Q/V187T/T189D，E056K/L075R/D130A/V187N/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/D027S/E056K/V187T/L264R，G023K/E056K/L075R/V187N/L264R，G023K/K024A/D111A/D130A/V187T，G023K/L075Q/V187H/I252Q/L264R，G023Q/D027S/L075G/I252Q/L264R，G023Q/L075Q/V077I/D130A/G156W，N011K/D027N/S058M/N233Q/P256T，N011K/D027S/I090F/N233Q/P256T，Q004D/N011K/D027N/N233Q/P256T，S058M/D137Q/G163P/L227M/N233Q，V077I/D130A/V154I/G156W/V187N，A018K/D027S/N033D/E045F/N073R/L075D，A018K/D027S/N033D/N073R/L075D/T189D，A018K/D027S/P029E/N033D/E045F/L075D，A018K/G023Q/K024A/L075R/N094R/G156W，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/D130A/L227M/N233Q，D027Q/E056K/S058M/D130A/I252Q/L264R，D027S/E056K/S058M/V187T/I252Q/L264R，D048Q/D130A/G163P/L227M/N233Q/L264R，D048Q/S058M/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/L075G/D130A/V187T/I252Q/L264R，G023Q/D027S/N094R/V154I/G156W/T189Q，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/E056K/L075Q/D130A/V154I/L264R，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/K024A/V077I/D130A/V154I/G156W，N011K/D027S/E056K/I090F/N233Q/P256T，Q004D/N011K/D027N/S058M/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，D048Q/S058M/D130A/D137Q/L227M/N233Q/L264R，G023Q/F051T/E056K/L075R/G091Q/V187H/I252Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，N011K/G023Q/L075Q/V154I/V187N/T189Q/L264R，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D
表7-12：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比参考序列SEQIDNO：2在织物粘附性测定中性能指数≤最大PI值的50％的TLL变体。
A018K/E045F，A018K/L075D，D027E/G163P，E045F/N073R，F051T/L075R，K024A/V154I，L227M/L264R，N033D/E045F，A018K/E045F/N073R，A018K/V154I/G156W，D027S/L075Q/G091Q，D027S/S058M/P256T，E056K/D130A/L264R，I090F/N233Q/P256T，N011K/N233Q/P256T，Q004D/S058M/N233Q，A018K/D111A/G156W/T189Q，A018K/E045F/L075D/T189D，D027E/D137Q/L227M/L264R，D027Q/E056K/N233Q/P256T，D027S/E045F/L075D/T189D，D027S/E045F/N073R/T189D，D027S/N033D/E045F/N073R，D027S/P029E/E045F/N073R，D027S/P029E/N033D/E045F，D048Q/D137Q/G163P/L227M，G023K/E056K/D130A/V187N，G023K/E056K/V187T/L264R，G023Q/K024A/L075R/G156W，L075G/G091Q/V187N/L264R，N033D/E045F/N073R/T189A，Q004D/D027N/S058M/P256T，Q004D/D027Q/N233Q/P256T，Q004D/D027Q/S058M/P256T，Q004D/N011K/D027S/P256T，A018K/D027S/E045F/L075D/T189D，A018K/E045F/N073R/L075D/T189D，A018K/K024A/L075R/D130A/V154I，D027Q/F051T/L075Q/D130A/L264R，D048Q/D137Q/G163P/N233Q/L264R，E045F/A049V/L075Q/V187T/T189D，E056K/L075R/D130A/V187N/L264R，G023K/E056K/L075R/V187N/L264R，G023K/K024A/D111A/D130A/V187T，G023K/L075Q/V187H/I252Q/L264R，G023Q/D027S/L075G/I252Q/L264R，N011K/D027N/S058M/N233Q/P256T，N011K/D027S/I090F/N233Q/P256T，S058M/D137Q/G163P/L227M/N233Q，A018K/D027S/N033D/E045F/N073R/L075D，A018K/D027S/N033D/N073R/L075D/T189D，A018K/D027S/P029E/N033D/E045F/L075D，D027E/D048Q/S058M/D130A/G163P/L264R，D027E/D048Q/S058M/D130A/L227M/N233Q，D027S/E056K/S058M/V187T/I252Q/L264R，D048Q/D130A/G163P/L227M/N233Q/L264R，D048Q/S058M/G163P/L227M/N233Q/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/L075G/D130A/V187T/I252Q/L264R，G023Q/E056K/D111A/D130A/V187N/T189Q，G023Q/K024A/V077I/D130A/V154I/G156W，N011K/D027S/E056K/I090F/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，D048Q/S058M/D130A/D137Q/L227M/N233Q/L264R，G023Q/F051T/E056K/L075R/G091Q/V187H/I252Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，N011K/D027N/E056K/S058M/I090F/N233Q/P256T，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，G023Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D
实施例8
TLL组合变体的清洁性能
按实施例1所述的程序(CS-61微布样测定法)，使用半剂量辅剂或全剂量，分析测定了按实施例2所述生成的TLL变体的清洁性能。计算了变体相比TLL(SEQIDNO：1)或参考序列SEQIDNO：2的性能指数。
表8-1：下面示出，相比TLL(SEQIDNO：1)，使用半剂量辅剂在CS-61微布样测定中性能指数＞1的TLL变体。
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108K/H135F/V187T/T189D，A018K/G023Q/D027S/P029E/N033D/E045F/A049V/S058M/N073S/K074S/L075Q/V077I/N101D/R108K/H135F/D137V/G156W/V187T/T189D
表8-2：下面示出，相比TLL(SEQIDNO：1)，表达性能指数≥0.5，且在使用半剂量+辅剂的CS-61微布样测定中性能指数＞1的TLL变体。
D027S/N033D，D130A/L264R，D130A/V187T，L075R/L264R，N033D/E045F，P029E/N033D，V187T/L264R，A018K/E045F/V187T，A018K/L075D/T189D，A018K/P029E/T189D，D027E/S058M/G163P，D027S/L075Q/G091Q，D027S/N033D/T189D，D111A/D130A/L264R，F051T/I252Q/L264R，G023K/E056K/V187T，G023K/V187N/L264R，G023Q/A049V/T189D，G163P/L227M/L264R，L075G/D130A/V187H，L075Q/V187N/L264R，L075Q/V187T/L264R，L075R/D130A/L264R，L075R/D130A/V187T，L075R/V187N/L264R，P029E/N033D/E045F，V187N/T189Q/L264R，A018K/D027S/P029E/T189D，A018K/D130A/G156W/V187T，A018K/L075Q/V077I/N094R，A018K/N033D/L075D/T189D，D027E/D130A/N233Q/L264R，D027E/D137Q/G163P/L227M，D027E/D137Q/L227M/L264R，D027E/L227M/N233Q/L264R，D027E/S058M/G163P/L264R，D027S/D130A/I252Q/L264R，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D048Q/G163P/N233Q/L264R，D130A/G163P/L227M/L264R，E056K/D130A/V187N/L264R，F051T/D130A/I252Q/L264R，F051T/L075G/I252Q/L264R，F051T/L075Q/I252Q/L264R，G023K/D027Q/F051T/L075Q，G023K/D130A/G156W/T189Q，G023K/D130A/V187T/L264R，G023K/D130A/V187T/T189Q，G023K/E056K/V187T/L264R，G023K/K024A/V154I/V187N，G023K/L075R/D130A/L264R，G023Q/A049V/V077I/G156W，G023Q/L075Q/D130A/L264R，G023Q/L075Q/V187T/L264R，K024A/D130A/V154I/V187T，L075G/D130A/V187T/I252Q，L075G/V187H/I252Q/L264R，L075R/D130A/V187T/L264R，N011K/D027S/N233Q/P256T，P029E/N073R/L075D/T189D，S058M/D137Q/G163P/N233Q，S058M/G163P/L227M/L264R，S058M/G163P/N233Q/L264R，S058M/L075Q/G091Q/I252Q，A018K/D027S/P029E/N033D/L075D，A018K/D130A/G156W/V187N/L264R，A018K/E045F/N073R/L075D/T189D，A018K/G023K/D130A/V154I/G156W，A018K/G023K/K024A/D130A/G156W，A018K/G023Q/K024A/D130A/V187Q，A018K/G023Q/K024A/L075Q/G156W，A018K/G023Q/V077I/D130A/G156W，A018K/K024A/N094R/D130A/V187N，A018K/P029E/N033D/N073R/L075D，D027E/D130A/G163P/N233Q/L264R，D027E/D137Q/G163P/N233Q/L264R，D027E/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L264R，D027E/S058M/G163P/L227M/L264R，D027S/E056K/D111A/V187N/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075G/G091Q/I252Q/L264R，D027S/L075R/V154I/T189Q/L264R，D048Q/D130A/G163P/L227M/L264R，D048Q/D137Q/G163P/L227M/L264R，D048Q/D137Q/G163P/N233Q/L264R，D048Q/S058M/D130A/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D130A/D137Q/G163P/L227M/L264R，D130A/D137Q/G163P/N233Q/L264R，E056K/L075G/V187N/I252Q/L264R，E056K/L075R/D130A/V187T/L264R，F051T/L075G/D130A/I252Q/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/L264R，F051T/L075Q/V187N/I252Q/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187N/L264R，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/L075Q/V187H/I252Q/L264R，G023K/L075R/D130A/V187T/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/E056K/V187N/I252Q/L264R，G023Q/F051T/L075Q/G091Q/I252Q，G023Q/L075G/G091Q/I252Q/L264R，G023Q/L075Q/D130A/I252Q/L264R，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/V154I/G156W/V187Q，L075G/D130A/V187T/I252Q/L264R，N011K/G023K/D111A/G156W/L264R，S058M/D130A/D137Q/G163P/L264R，S058M/D130A/G163P/L227M/L264R，S058M/D137Q/G163P/L227M/L264R，S058M/G163P/L227M/N233Q/L264R，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/K024A/V077I/V154I/V187N，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/D111A/V187N/T189Q/L264R，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D130A/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/N233Q/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/D137Q/G163P/N233Q/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/G163P/L227M/L264R，D027E/S058M/D130A/G163P/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027E/S058M/D137Q/G163P/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/F051T/G091Q/D130A/I252Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027Q/L075Q/D130A/V187T/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D027S/F051T/L075Q/D130A/I252Q/L264R，D027S/L075Q/G091Q/V187H/I252Q/L264R，D048Q/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L264R，D048Q/S058M/D130A/G163P/L227M/L264R，D048Q/S058M/D137Q/G163P/L227M/L264R，D048Q/S058M/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/D130A/V154I/G156W/V187T/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/F051T/G091Q/D130A/V187H/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/D027S/L075G/D130A/V187H/L264R，G023Q/D027S/L075Q/D130A/V187T/I252Q，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/F051T/G091Q/V187N/I252Q/L264R，G023Q/F051T/L075G/G091Q/V187N/I252Q，G023Q/K024A/D130A/V154I/G156W/V187Q，G023Q/K024A/V077I/D130A/G156W/V187N，K024A/L075Q/N094R/D130A/V154I/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，L075Q/G091Q/D130A/V187H/I252Q/L264R，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，S058M/D130A/G163P/L227M/N233Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，D027E/D048Q/S058M/D130A/D137Q/G163P/L264R，D027E/D048Q/S058M/D130A/G163P/L227M/L264R，D027E/D048Q/S058M/D130A/G163P/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/L264R，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/L264R，D027E/S058M/D137Q/G163P/L227M/N233Q/L264R，D027Q/F051T/S058M/L075Q/G091Q/I252Q/L264R，D027Q/L075G/G091Q/D130A/V187H/I252Q/L264R，D027S/F051T/S058M/L075R/D130A/I252Q/L264R，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/S058M/D130A/G163P/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/V187N/I252Q/L264R，F051T/L075R/G091Q/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/S058M/L075Q/D130A/V187N，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/E056K/L075R/D111A/D130A/V187N/T189Q，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/V187H/I252Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，G023Q/S058M/L075G/G091Q/D130A/I252Q/L264R，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D027E/D048Q/S058M/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027Q/F051T/S058M/L075R/D130A/V187T/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/S058M/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187H/I252Q/L264R，G023Q/F051T/L075Q/G091Q/D130A/V187T/I252Q/L264R，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，D027Q/F051T/S058M/L075G/G091Q/D130A/V187T/I252Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T
表8-3：下面示出，相比TLL(SEQIDNO：1)，表达性能指数≥0.5，且在使用半剂量辅剂的CS-61微布样测定中性能指数≥最大PI值的50％的TLL变体。
P029E/N033D，A018K/L075D/T189D，A018K/P029E/T189D，D027S/N033D/T189D，G023K/V187N/L264R，G023Q/A049V/T189D，L075G/D130A/V187H，L075Q/V187N/L264R，L075R/D130A/V187T，L075R/V187N/L264R，P029E/N033D/E045F，A018K/D027S/P029E/T189D，A018K/L075Q/V077I/N094R，A018K/N033D/L075D/T189D，D027E/D137Q/G163P/L227M，D027E/L227M/N233Q/L264R，D027S/N033D/L075D/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D130A/G163P/L227M/L264R，G023K/D027Q/F051T/L075Q，G023K/D130A/V187T/L264R，G023Q/A049V/V077I/G156W，L075G/D130A/V187T/I252Q，L075R/D130A/V187T/L264R，P029E/N073R/L075D/T189D，S058M/D137Q/G163P/N233Q，A018K/G023K/K024A/D130A/G156W，A018K/G023Q/K024A/D130A/V187Q，A018K/K024A/N094R/D130A/V187N，A018K/P029E/N033D/N073R/L075D，D027E/D130A/G163P/N233Q/L264R，D027E/D137Q/G163P/N233Q/L264R，D130A/D137Q/G163P/N233Q/L264R，E056K/L075R/D130A/V187T/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187N/L264R，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187T/L264R，K024A/L075R/V154I/G156W/V187Q，N011K/G023K/D111A/G156W/L264R，S058M/D130A/G163P/L227M/L264R，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/N233Q/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D048Q/D137Q/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/F051T/D130A/V187T/I252Q/L264R，G023Q/K024A/V077I/D130A/G156W/V187N，K024A/L075Q/V077I/N094R/D130A/V187N，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，D027E/D048Q/S058M/D130A/G163P/L227M/L264R，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T
表8-4：下面示出，相比参考序列SEQIDNO：2，在使用半剂量辅剂的CS-61微布样测定中性能指数＞1的TLL变体。
A018K/L075Q，A018K/T189D，G023K/D130A，K024A/L075Q，L075Q/D130A，L075Q/G156W，P029E/N033D，A018K/A049V/L075Q，A018K/D027S/N073R，A018K/D027S/T189D，A018K/E045F/T189D，A018K/G023K/L075Q，A018K/G023Q/L075Q，A018K/L075Q/G156W，A018K/L075Q/V187T，A018K/N033D/L075D，A018K/N073R/L075D，A018K/P029E/T189D，A049V/V187T/T189D，D027S/N033D/T189D，D027S/N233Q/P256T，E056K/D130A/T189Q，G023K/V187N/L264R，G023Q/A049V/T189D，G023Q/D111A/L264R，G023Q/E056K/L075R，G023Q/L075Q/V077I，G023Q/L075Q/V187T，K024A/L075Q/V077I，L075G/D130A/V187H，L075Q/D130A/V187T，L075Q/G156W/T189D，L075Q/G156W/V187N，L075Q/V077I/T189D，L075Q/V154I/V187T，L075Q/V187T/T189D，L075R/D130A/V187T，L075R/V187N/L264R，N073R/L075D/T189D，P029E/E045F/L075D，P029E/L075D/T189D，P029E/N033D/E045F，P029E/N073R/L075D，Q004D/N233Q/P256T，S058M/L227M/L264R，V077I/V187A/T189D，A018K/A049V/L075Q/T189D，A018K/A049V/L075Q/V187T，A018K/D027S/N033D/L075D，A018K/D027S/P029E/T189D，A018K/E045F/L075Q/T189D，A018K/G023Q/E045F/T189D，A018K/G023Q/E045F/V187T，A018K/G023Q/L075Q/T189D，A018K/G023Q/L075Q/V077I，A018K/G023Q/L075Q/V187T，A018K/G023Q/V187T/T189D，A018K/L075Q/N094R/D111A，A018K/L075Q/N094R/V187QA018K/L075Q/V077I/N094R，A018K/N033D/L075D/T189D，A018K/N073R/L075D/T189D，A049V/L075Q/V077I/T189D，A049V/L075Q/V187T/T189D，D027E/D137Q/G163P/L227M，D027S/N033D/L075D/T189D，D027S/P029E/E045F/L075D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D027S/P029E/N033D/T189D，D048Q/D130A/G163P/L264R，D130A/G163P/L227M/L264R，D130A/G163P/N233Q/L264R，E056K/L075Q/V187N/L264R，G023K/D130A/V187T/L264R，G023K/L075Q/D130A/V187N，G023K/L075Q/G156W/L264R，G023K/L075Q/G156W/V187N，G023K/L075R/V187N/L264R，G023Q/A049V/L075Q/T189D，G023Q/A049V/L075Q/V077I，G023Q/D027S/D111A/G156W，G023Q/E045F/A049V/T189D，G023Q/E045F/L075Q/G156W，G023Q/E045F/L075Q/V187T，G023Q/K024A/L075R/V154I，G023Q/L075Q/G156W/V187N，G023Q/L075Q/G156W/V187T，K024A/L075Q/V187T/T189Q，K024A/L075R/G156W/V187N，L075G/D130A/V187T/I252Q，L075Q/D130A/G156W/V187N，L075Q/V077I/D130A/V187Q，L075Q/V077I/G156W/V187N，L075Q/V077I/N094R/G156W，L075Q/V077I/VI54I/V187Q，L075Q/V077I/V187T/T189D，L075R/D130A/V154I/L264R，N011K/E056K/S058M/P256T，P029E/E045F/L075D/T189D，P029E/E045F/N073R/L075D，P029E/E045F/N073R/T189D，P029E/N033D/E045F/L075D，P029E/N033D/L075D/T189D，P029E/N073R/L075D/T189D，Q004D/D027S/I090F/P256T，Q004D/N011K/D027N/P256T，A018K/D027S/E045F/N073R/D137V，A018K/D027S/N033D/E045F/T189D，A018K/D027S/N033D/L075D/T189D，A018K/D027S/N073R/L075D/T189D，A018K/D027S/P029E/E045F/T189D，A018K/D027S/P029E/N073R/L075D，A018K/G023K/K024A/D130A/G156W，A018K/G023K/K024A/L075R/N094R，A018K/G023K/L075Q/D130A/V154I，A018K/G023Q/A049V/L075Q/G156W，A018K/G023Q/L075Q/G156W/V187T，A018K/G023Q/L075Q/V187T/T189D，A018K/K024A/L075Q/D130A/L264R，A018K/K024A/L075Q/V154I/G156W，A018K/K024A/L075R/N094R/D130A，A018K/K024A/N094R/D130A/V187N，A018K/L075Q/D111A/D130A/V187T，A018K/L075Q/N094R/G156W/V187N，A018K/L075Q/V077I/N094R/G156W，A018K/N033D/E045F/L075D/T189D，A018K/P029E/E045F/N073R/L075D，D027E/D130A/G163P/N233Q/L264R，D027E/D137Q/G163P/L227M/L264R，D027E/D137Q/G163P/N233Q/L264R，D027S/N033D/N073R/L075D/T189D，D027S/P029E/N033D/N073R/L075D，D027S/P029E/N073R/L075D/T189D，D048Q/D130A/D137Q/G163P/L264R，D130A/D137Q/G163P/N233Q/L264R，E056K/D130A/V187H/I252Q/L264R，E056K/L075Q/G156W/T189Q/L264R，E056K/L075R/D130A/V187T/L264R，G023K/D027S/E056K/V187T/L264R，G023K/D111A/V154I/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187N/L264R，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/D111A/D130A/V187T，G023K/L075Q/D130A/G156W/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187T/L264R，G023Q/A049V/L075Q/G156W/T189D，G023Q/A049V/L075Q/G156W/V187T，G023Q/A049V/L075Q/V187T/T189D，G023Q/D027S/V154I/V187N/L264R，G023Q/E045F/L075Q/V077I/V187T，G023Q/K024A/L075Q/G156W/V187N，G023Q/K024A/L075Q/V077I/V187Q，G023Q/K024A/L075R/G156W/V187Q，G023Q/K024A/V077I/D130A/V154I，G023Q/L075Q/V077I/G156W/V187N，G091Q/D130A/V187H/I252Q/L264R，K024A/L075Q/D111A/G156W/V187T，K024A/L075Q/V077I/G156W/V187N，K024A/L075Q/V077I/V154I/V187N，K024A/L075R/V154I/G156W/V187Q，L075Q/D111A/V187N/T189Q/L264R，L075Q/V154I/V187N/T189Q/L264R，L075R/D111A/D130A/V187N/T189Q，L075R/D130A/V154I/T189Q/L264R，L075R/D130A/V187T/T189Q/L264R，L075R/G156W/V187T/T189Q/L264R，N011K/G023K/D111A/G156W/L264R，N011K/G023K/E056K/L075Q/T189Q，P029E/N033D/E045F/L075D/T189D，P029E/N033D/K074S/L075D/T189D，P029E/N033D/N073R/L075D/T189D，Q004D/D027Q/E056K/N233Q/P256T，Q004D/N011K/D027Q/E056K/N233Q，Q004D/N011K/I090F/N233Q/P256T，A018K/A049V/L075Q/G156W/V187T/T189D，A018K/D027S/N033D/E045F/L075D/T189D，A018K/D027S/P029E/N073R/L075D/T189D，A018K/G023K/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075R/D130A/V187N，A018K/G023K/L075Q/D130A/G156W/V187N，A018K/G023K/L075Q/G156W/V187N/L264R，A018K/G023K/L075R/V077I/D130A/V154I，A018K/G023Q/A049V/L075Q/V077I/V187T，A018K/G023Q/E045F/A049V/L075Q/T189D，A018K/G023Q/L075Q/G156W/V187T/T189D，A018K/G023Q/L075Q/V077I/D130A/V187N，A018K/G023Q/L075Q/V077I/G156W/V187N，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/N094R/V154I/V187N，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/K024A/L075R/D130A/V187N/T189Q，A018K/L075Q/D130A/V154I/T189Q/L264R，A018K/L075Q/D130A/V187N/T189Q/L264R，A018K/L075R/D130A/V154I/G156W/V187N，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/N233Q/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/F051T/L075G/D130A/I252Q/L264R，D027Q/F051T/L075Q/D130A/V187H/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D027S/P029E/N033D/E045F/N073R/T189D，D027S/P029E/N033D/N073R/L075D/T189D，D048Q/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/G163P/N233Q/L264R，D111A/D130A/V154I/G156W/V187N/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/L075Q/D111A/D130A/V154I/V187N，G023K/L075Q/N094R/V154I/V187T/L264R，G023K/L075Q/V077I/D130A/G156W/V187Q，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/A049V/L075Q/G156W/V187T/T189D，G023Q/D027Q/E056K/L075R/I252Q/L264R，G023Q/D027S/F051T/L075Q/V187T/I252Q，G023Q/E045F/L075Q/V077I/G156W/V187T，G023Q/E045F/L075Q/V077I/V187T/T189D，G023Q/K024A/L075Q/D130A/V154I/V187N，G023Q/L075Q/V077I/N094R/D130A/V187Q，K024A/D111A/D130A/V187T/T189Q/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，K024A/L075Q/V154I/V187N/T189Q/L264R，K024A/N094R/V154I/G156W/T189Q/L264R，L075Q/D111A/D130A/V154I/V187N/T189Q，L075Q/N094R/D111A/G156W/T189Q/L264R，N011K/D027S/S058M/I090F/N233Q/P256T，N011K/E056K/L075Q/D130A/V187N/T189Q，N011K/G023K/D027S/V154I/G156W/T189Q，N011K/G023Q/L075Q/D130A/V187N/T189Q，P029E/N033D/K074S/L075D/N101D/T189D，Q004D/N011K/D027Q/E056K/S058M/N233Q，A018K/G023K/K024A/L075Q/V077I/N094R/G156W，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/K024A/L075R/V154I/G156W/V187Q，A018K/G023K/K024A/N094R/V154I/V187T/L264R，A018K/G023Q/K024A/L075Q/V077I/D130A/G156W，A018K/G023Q/K024A/L075Q/V077I/N094R/V154I，A018K/G023Q/K024A/L075R/D130A/V154I/V187N，A018K/G023Q/K024A/L075R/V077I/D130A/G156W，A018K/K024A/L075Q/D111A/G156W/V187N/L264R，A018K/K024A/L075Q/D130A/G156W/V187T/T189Q，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，A018K/L075Q/D111A/D130A/V154I/G156W/L264R，A018K/L075Q/D111A/V154I/V187T/T189Q/L264R，A018K/L075Q/N094R/D111A/V154I/V187T/L264R，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/N233Q/L264R，D027Q/F051T/L075G/D130A/V187H/I252Q/L264R，D027Q/F051T/L075Q/G091Q/D130A/V187T/I252Q，D027S/F051T/L075G/G091Q/V187T/I252Q/L264R，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，E056K/L075Q/D111A/D130A/G156W/V187N/T189Q，G023K/D027Q/F051T/L075G/D130A/V187H/L264R，G023K/D027S/L075Q/N094R/V154I/G156W/T189Q，G023K/F051T/E056K/L075R/D130A/V187T/I252Q，G023K/K024A/L075Q/D111A/V154I/G156W/V187N，G023K/K024A/L075Q/D130A/V187T/T189Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N，G023K/K024A/L075Q/V077I/D130A/G156W/V187Q，G023K/L075Q/N094R/V154I/G156W/V187T/L264R，G023K/L075R/N094R/V154I/G156W/V187N/L264R，G023Q/D027S/L075Q/G091Q/V187T/I252Q/L264R，G023Q/E045F/A049V/V077I/G156W/V187T/T189D，G023Q/E045F/L075Q/V077I/G156W/V187T/T189D，G023Q/E056K/L075Q/D130A/V154I/G156W/T189Q，G023Q/F051T/L075R/G091Q/V187N/I252Q/L264R，G023Q/K024A/L075Q/V077I/D130A/G156W/V187N，G023Q/K024A/L075Q/V077I/D130A/V154I/G156W，G023Q/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/L075Q/D130A/V154I/V187N/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，G023Q/L075Q/V077I/N094R/D130A/G156W/V187Q，L075Q/D111A/D130A/G156W/V187N/T189Q/L264R，N011K/E056K/L075Q/D130A/V154I/V187N/T189Q，N011K/G023K/L075Q/D111A/D130A/V154I/V187N，Q004D/N011K/D027Q/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/D130Y/V154I/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023K/K024A/L075R/D111A/V154I/V187T/L264R，A018K/G023K/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/G023Q/E045F/L075Q/V077I/G156W/V187T/T189D，A018K/G023Q/K024A/L075Q/V077I/D130A/G156W/V187Q，A018K/G023Q/K024A/L075R/D130A/V154I/G156W/V187N，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，A018K/G023Q/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/K024A/L075Q/V077I/N094R/D130A/V154I/G156W，A018K/K024A/L075R/D130A/V154I/G156W/V187T/L264R，A018K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/E056K/L075G/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/D027S/F051T/E056K/L075R/D130A/V187T/L264R，G023K/F051T/E056K/L075R/G091Q/D130A/V187T/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187T/L264R，G023K/K024A/L075R/N094R/V154I/G156W/V187T/T189Q，G023K/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075R/G091Q/V187N/I252Q，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/E056K/L075Q/D111A/V154I/V187N/T189Q/L264R，G023Q/K024A/L075Q/V077I/D130A/V154I/G156W/V187N，G023Q/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，N011K/G023K/E056K/L075Q/D130A/V154I/T189Q/L264R，N011K/G023Q/D027S/L075Q/N094R/V154I/G156W/T189Q，N011K/G023Q/D027S/N094R/V154I/G156W/T189Q/L264R，N011K/G023Q/L075Q/N094R/V154I/G156W/T189Q/L264R，A018K/G023K/K024A/L075Q/N094R/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，A018K/G023Q/D027S/P029E/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/G156W/V187H，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/P029E/S058M/V077I/R108K/H135F/G156W，A018K/G023Q/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W/V187N，A018K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，G023K/D027S/F051T/E056K/S058M/L075Q/G091Q/V187N/L264R，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/D027S/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，N011K/G023Q/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/S058M/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023K/D027S/F051T/E056K/S058M/L075G/G091Q/D130A/I252Q/L264R，G023Q/D027S/F051T/E056K/L075R/G091Q/D130A/V187H/I252Q/L264R，G023Q/D027S/F051T/E056K/S058M/L075R/G091Q/D130A/V187H/I252Q，G023Q/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，N011K/G023K/L075R/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，N011K/G023Q/L075Q/N094R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/H135F/V187T/T189D，A018K/G023Q/D027S/P029E/N033D/E045F/A049V/S058M/N073S/K074S/L075Q/V077I/N101D/R108K/H135F/D137V/G156W/V187T/T189D
表8-5：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比参考序列SEQIDNO：2在使用半剂量辅剂的CS-61微布样测定中性能指数＞1的TLL变体A018K/P029E/T189D，D027S/N033D/T189D，G023K/V187N/L264R，G023Q/A049V/T189D，L075G/D130A/V187H，L075R/D130A/V187T，L075R/V187N/L264R，P029E/N033D/E045F，A018K/D027S/P029E/T189D，A018K/L075Q/V077I/N094R，A018K/N033D/L075D/T189D，D027E/D137Q/G163P/L227M，D027S/N033D/L075D/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D130A/G163P/L227M/L264R，G023K/D130A/V187T/L264R，L075G/D130A/V187T/I252Q，P029E/N073R/L075D/T189D，A018K/G023K/K024A/D130A/G156W，A018K/K024A/N094R/D130A/V187N，D027E/D130A/G163P/N233Q/L264R，D027E/D137Q/G163P/N233Q/L264R，D130A/D137Q/G163P/N233Q/L264R，E056K/L075R/D130A/V187T/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187N/L264R，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187T/L264R，K024A/L075R/V154I/G156W/V187Q，N011K/G023K/D111A/G156W/L264R，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/N233Q/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D048Q/D137Q/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/N094R/D111A/G156W/V187T/L264R，K024A/L075Q/V077I/N094R/D130A/V187N，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，F051T/E056K/L075G/G091Q/D130A/V187H/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/G023Q/E056K/L075R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R
表8-6：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比参考序列SEQIDNO：2在使用半剂量+辅剂的CS-61微布样测定中性能指数≥最大PI值的D027S/N033D/T189D，G023K/V187N/L264R，L075R/D130A/V187T，D048Q/D130A/G163P/L264R，D130A/G163P/L227M/L264R，P029E/N073R/L075D/T189D，A018K/K024A/N094R/D130A/V187N，D027E/D130A/G163P/N233Q/L264R，D027E/D137Q/G163P/N233Q/L264R，D130A/D137Q/G163P/N233Q/L264R，E056K/L075R/D130A/V187T/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/L075R/V187N/L264R，G023K/E056K/L075R/V187T/L264R，G023K/L075R/D130A/V187T/L264R，K024A/L075R/V154I/G156W/V187Q，A018K/K024A/L075Q/N094R/V154I/G156W，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D048Q/D137Q/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/N094R/D111A/G156W/V187T/L264R，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R
表8-7：下面示出，相比TLL(SEQIDNO：1)，在使用全剂量的CS-61微布样测定中性能指数＞1的TLL变体。
A018K/D027S，A018K/L075Q，A018K/T189D，D027N/E056K，D027S/N033D，D130A/L264R，D130A/V187N，E045F/T189D，G023K/D130A，G023K/L264R，K024A/L075Q，K024A/V154I，L075Q/D130A，L075Q/G156W，L227M/L264R，N033D/E045F，P029E/N033D，V187T/L264R，A018K/A049V/L075Q，A018K/D027S/E045F，A018K/D027S/N073R，A018K/D027S/T189D，A018K/E045F/T189D，A018K/G023K/G156W，A018K/G023K/L075Q，A018K/G023Q/L075Q，A018K/K024A/V154I，A018K/L075D/T189D，A018K/L075Q/G156W，A018K/L075Q/V187T，A018K/N033D/L075D，A018K/N073R/L075D，A018K/P029E/T189D，A018K/V154I/G156W，A049V/V187T/T189D，D027E/S058M/G163P，D027Q/N233Q/P256T，D027S/N033D/T189D，D027S/N233Q/P256T，D111A/D130A/L264R，E045F/N073R/L075D，E056K/D130A/T189Q，G023K/D130A/L264R，G023K/D130A/V187T，G023K/E056K/V187T，G023K/V187N/L264R，G023Q/A049V/T189D，G023Q/D111A/L264R，G023Q/E056K/L075R，G023Q/K024A/G156W，G023Q/L075Q/V077I，G023Q/L075Q/V187T，K024A/L075Q/G156W，K024A/L075Q/V077I，L075G/D130A/V187H，L075Q/D111A/D130A，L075Q/D130A/V187T，L075Q/G156W/T189D，L075Q/G156W/V187N，L075Q/G156W/V187T，L075Q/V077I/T189D，L075Q/V154I/V187T，L075Q/V187N/L264R，L075Q/V187T/L264R，L075Q/V187T/T189D，L075R/D130A/L264R，L075R/D130A/V187T，L075R/V187N/L264R，L075R/V187T/L264R，N011K/D027S/S058M，N033D/E045F/N073R，N033D/N073R/T189D，N073R/L075D/T189D，P029E/E045F/L075D，P029E/L075D/T189D，P029E/N033D/E045F，P029E/N033D/L075D，P029E/N073R/L075D，S058M/L227M/L264R，V077I/D130A/V154I，V077I/V187A/T189D，V187N/T189Q/L264R，A018K/A049V/L075Q/T189D，A018K/A049V/L075Q/V187T，A018K/D027S/E045F/N073R，A018K/D027S/E045F/T189D，A018K/D027S/N033D/L075D，A018K/D027S/P029E/T189D，A018K/E045F/L075D/T189D，A018K/E045F/L075Q/T189D，A018K/E045F/L075Q/V187T，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/T189D，A018K/G023Q/A049V/V187T，A018K/G023Q/E045F/T189D，A018K/G023Q/E045F/V187T，A018K/G023Q/G156W/V187T，A018K/G023Q/L075Q/T189D，A018K/G023Q/L075Q/V077I，A018K/G023Q/L075Q/V187T，A018K/G023Q/L075R/D130A，A018K/G023Q/V077I/V187T，A018K/G023Q/V187T/T189D，A018K/L075Q/G156W/T189D，A018K/L075Q/G156W/V187T，A018K/L075Q/N094R/D111A，A018K/L075Q/N094R/V187Q，A018K/L075Q/V077I/N094R，A018K/L075Q/V187T/T189D，A018K/N033D/L075D/T189D，A018K/N073R/L075D/T189D，A018K/P029E/N033D/T189D，A018K/P029E/N073R/L075D，A049V/L075Q/V077I/T189D，A049V/L075Q/V187T/T189D，D027E/D137Q/G163P/L227M，D027E/D137Q/L227M/L264R，D027E/L227M/N233Q/L264R，D027S/E056K/D111A/V154I，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/L075D，D027S/P029E/E045F/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D027S/P029E/N033D/T189D，D048Q/D130A/G163P/L264R，D111A/D130A/V154I/L264R，D130A/G163P/L227M/L264R，D130A/G163P/N233Q/L264R，E056K/D130A/V187N/L264R，E056K/D130A/V187T/L264R，E056K/L075Q/V187N/L264R，G023K/D027Q/F051T/L075Q，G023K/D027S/L075Q/L264R，G023K/D130A/V187T/L264R，G023K/E056K/D130A/L264R，G023K/E056K/V187T/L264R，G023K/K024A/V154I/V187N，G023K/L075Q/D130A/V187N，G023K/L075Q/G156W/L264R，G023K/L075Q/G156W/V187N，G023K/L075R/D130A/L264R，G023K/L075R/V187N/L264R，G023K/L075R/V187T/L264R，G023Q/A049V/L075Q/T189D，G023Q/A049V/L075Q/V077I，G023Q/D027S/D111A/G156W，G023Q/E045F/A049V/T189D，G023Q/E045F/A049V/V187T，G023Q/E045F/L075Q/G156W，G023Q/E045F/L075Q/V187T，G023Q/K024A/L075R/G156W，G023Q/K024A/L075R/V154I，G023Q/L075Q/G156W/V187N，G023Q/L075Q/G156W/V187T，G023Q/L075Q/V187T/L264R，K024A/D130A/V154I/V187T，K024A/L075Q/D130A/G156W，K024A/L075Q/V187T/T189Q，K024A/L075R/G156W/V187N，L075G/D130A/V187T/I252Q，L075Q/D130A/G156W/V187N，L075Q/D130A/V154I/G156W，L075Q/V077I/D130A/V187Q，L075Q/V077I/G156W/V187N，L075Q/V077I/N094R/G156W，L075Q/V077I/V154I/V187Q，L075Q/V077I/V187T/T189D，L075R/D111A/V154I/G156W，L075R/D130A/V154I/L264R，N011K/D027N/S058M/P256T，N011K/D027S/E056K/P256T，N011K/D027S/N233Q/P256T，N011K/G023K/L075Q/D111A，P029E/E045F/L075D/T189D，P029E/E045F/N073R/L075D，P029E/E045F/N073R/T189D，P029E/N033D/E045F/L075D，P029E/N033D/L075D/T189D，P029E/N073R/L075D/T189D，Q004D/D027S/I090F/P256T，Q004D/N011K/D027N/P256T，Q004D/N011K/D027Q/N233Q，S058M/D137Q/G163P/N233Q，S058M/G163P/N233Q/L264R，S058M/L075Q/G091Q/I252Q，A018K/D027S/E045F/N073R/D137V，A018K/D027S/N033D/E045F/T189D，A018K/D027S/N033D/L075D/T189D，A018K/D027S/N073R/L075D/T189D，A018K/D027S/P029E/E045F/T189D，A018K/D027S/P029E/N033D/L075D，A018K/D027S/P029E/N073R/L075D，A018K/D130A/G156W/V187N/L264R，A018K/E045F/A049V/L075Q/V187T，A018K/E045F/N073R/L075D/T189D，A018K/G023K/D130A/V154I/G156W，A018K/G023K/K024A/D130A/G156W，A018K/G023K/K024A/L075R/N094R，A018K/G023K/L075Q/D130A/V154I，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/A049V/L075Q/G156W，A018K/G023Q/A049V/L075Q/V077I，A018K/G023Q/A049V/L075Q/V187T，A018K/G023Q/A049V/V077I/V187T，A018K/G023Q/E045F/A049V/L075Q，A018K/G023Q/E045F/A049V/V187T，A018K/G023Q/E045F/L075Q/V187T，A018K/G023Q/E045F/V077I/T189D，A018K/G023Q/K024A/D130A/V154I，A018K/G023Q/K024A/D130A/V187Q，A018K/G023Q/K024A/L075Q/G156W，A018K/G023Q/L075Q/G156W/V187T，A018K/G023Q/L075Q/V187T/T189D，A018K/G023Q/V077I/D130A/G156W，A018K/G023Q/V077I/V187T/T189D，A018K/K024A/L075Q/D130A/L264R，A018K/K024A/L075Q/V154I/G156W，A018K/K024A/L075Q/V154I/T189D，A018K/K024A/L075R/D130A/V154I，A018K/K024A/L075R/N094R/D130A，A018K/K024A/N0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T189Q/L264R，L075Q/D111A/D130A/G156W/V187N/T189Q/L264R，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，N011K/D027S/D111A/D130A/V154I/V187N/T189Q，N011K/E056K/L075Q/D130A/V154I/V187N/T189Q，N011K/G023K/L075Q/D111A/D130A/V154I/V187N，Q004D/N011K/D027Q/E056K/S058M/N233Q/P256T，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/D130Y/V154I/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023K/K024A/L075R/D111A/V154I/V187T/L264R，A018K/G023K/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/G023K/L075R/V077I/N094R/D130A/G156W/V187Q，A018K/G023Q/E045F/L075Q/V077I/G156W/V187T/T189D，A018K/G023Q/K024A/L075Q/V077I/D130A/G156W/V187Q，A018K/G023Q/K024A/L075R/D130A/V154I/G156W/V187N，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，A018K/G023Q/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/K024A/L075Q/V077I/N094R/D130A/V154I/G156W，A018K/K024A/L075R/D130A/V154I/G156W/V187T/L264R，A018K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/E056K/L075G/D130A/V187T/I252Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/D027S/F051T/E056K/L075R/D130A/V187T/L264R，G023K/F051T/E056K/L075R/G091Q/D130A/V187T/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187T/L264R，G023K/K024A/L075R/N094R/V154I/G156W/V187T/T189Q，G023K/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，G023K/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075R/G091Q/V187N/I252Q，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，G023Q/E056K/L075Q/D111A/V154I/V187N/T189Q/L264R，G023Q/E056K/L075Q/N094R/D111A/G156W/T189Q/L264R，G023Q/K024A/L075Q/V077I/D130A/V154I/G156W/V187N，G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/V187N，G023Q/L075Q/N094R/D111A/D130A/G156W/T189Q/L264R，G023Q/L075Q/N094R/D130A/V154I/G156W/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，K024A/D111A/D130A/V154I/G156W/V187T/T189Q/L264R，N011K/G023K/D027S/L075R/V154I/G156W/V187N/T189Q，N011K/G023K/E056K/L075Q/D130A/V154I/T189Q/L264R，N011K/G023Q/D027S/L075Q/N094R/V154I/G156W/T189Q，N011K/G023Q/D027S/N094R/V154I/G156W/T189Q/L264R，N011K/G023Q/L075Q/N094R/V154I/G156W/T189Q/L264R，A018K/G023K/K024A/L075Q/N094R/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，A018K/G023Q/D027S/L075Q/V077I/R108K/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/G156W/V187H，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/P029E/S058M/V077I/R108K/H135F/G156W，A018K/G023Q/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W/V187N，A018K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V187N/T189Q/L264R，G023K/D027S/F051T/E056K/S058M/L075Q/G091Q/V187N/L264R，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/D027S/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/F051T/E056K/L075G/G091Q/V187N/I252Q/L264R，G023Q/D027S/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，N011K/G023Q/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/E045F/A049V/S058M/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/S058M/L075Q/V077I/R108K/H135F/G156W/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023K/D027S/F051T/E056K/S058M/L075G/G091Q/D130A/I252Q/L264R，G023Q/D027S/F051T/E056K/L075R/G091Q/D130A/V187H/I252Q/L264R，G023Q/D027S/F051T/E056K/S058M/L075R/G091Q/D130A/V187H/I252Q，G023Q/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，N011K/G023K/L075R/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，N011K/G023Q/L075Q/N094R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T，N011K/G023Q/D027S/L075Q/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/H135F/V187T/T189D，A018K/G023Q/D027S/P029E/N033D/E045F/A049V/S058M/N073S/K074S/L075Q/V077I/N101D/R108K/H135F/D137V/G156W/V187T/T189D
表8-8：下面示出，相比TLL(SEQIDNO：1)，表达性能指数≥0.5，且在使用全剂量的CS-61微布样测定中性能指数＞1的TLL变体。
D027S/N033D，D130A/L264R，D130A/V187N，G023K/L264R，K024A/V154I，L227M/L264R，N033D/E045F，P029E/N033D，V187T/L264R，A018K/K024A/V154I，A018K/L075D/T189D，A018K/P029E/T189D，A018K/V154I/G156WD027E/S058M/G163P，D027S/N033D/T189D，D111A/D130A/L264R，G023K/D130A/L264RG023K/D130A/V187T，G023K/E056K/V187T，G023K/V187N/L264R，G023Q/A049V/T189D，G023Q/K024A/G156W，L075G/D130A/V187H，L075Q/V187N/L264R，L075Q/V187T/L264RL075R/D130A/L264R，L075R/D130A/V187T，L075R/V187N/L264R，L075R/V187T/L264RP029E/N033D/E045F，V187N/T189Q/L264R，A018K/D027S/P029E/T189D，A018K/E045F/L075D/T189D，A018K/G023K/D111A/T189Q，A018K/G023Q/A049V/V187T，A018K/L075Q/V077I/N094R，A018K/N033D/L075D/T189D，A018K/P029E/N073R/L075D，D027E/D137Q/G163P/L227M，D027E/D137Q/L227M/L264R，D027E/L227M/N233Q/L264R，D027S/N033D/E045F/T189D，D027S/N033D/L075D/T189D，D027S/N033D/N073R/T189D，D027S/P029E/E045F/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D130A/G163P/L227M/L264R，E056K/D130A/V187N/L264R，E056K/D130A/V187T/L264R，G023K/D027Q/F051T/L075Q，G023K/D130A/V187T/L264R，G023K/E056K/V187T/L264R，G023K/K024A/V154I/V187N，G023K/L075R/D130A/L264R，G023K/L075R/V187T/L264R，G023Q/K024A/L075R/G156W，G023Q/L075Q/V187T/L264R，K024A/D130A/V154I/V187T，L075G/D130A/V187T/I252Q，N011K/D027S/N233Q/P256T，P029E/N073R/L075D/T189D，S058M/D137Q/G163P/N233Q，S058M/G163P/N233Q/L264R，S058M/L075Q/G091Q/I252Q，A018K/D027S/P029E/N033D/L075D，A018K/D130A/G156W/V187N/L264R，A018K/E045F/N073R/L075D/T189D，A018K/G023K/D130A/V154I/G156W，A018K/G023K/K024A/D130A/G156W，A018K/G023K/V077I/D130A/V187N，A018K/G023Q/K024A/D130A/V187Q，A018K/G023Q/K024A/L075Q/G156W，A018K/G023Q/V077I/D130A/G156W，A018K/K024A/L075R/D130A/V1541，A018K/K024A/N094R/D130A/V187N，A018K/P029E/N033D/N073R/L075D，D027E/D130A/G163P/L227M/L264R，D027E/D130A/G163P/N233Q/L264R，D027E/D137Q/G163P/N233Q/L264R，D027E/S058M/D137Q/G163P/N233Q，D027S/E056K/D111A/V187N/L264R，D027S/L075G/D130A/I252Q/L264R，D027S/L075R/V154I/T189Q/L264R，D048Q/S058M/D130A/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233E，D130A/D137Q/G163P/N233Q/L264R，E045F/A049V/L075Q/V187T/T189D，E056K/L075R/D130A/V187T/L264R，F051T/L075G/V187N/I252Q/L264R，F051T/L075Q/G091Q/D130A/L264R，F051T/S058M/L075Q/G091Q/I252Q，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187N/L264R，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/L075Q/V187H/I252Q/L264R，G023K/L075R/D130A/V187T/L264R，G023K/N094R/G156W/V187N/T189Q，G023Q/K024A/L075Q/G156W/V187Q，G023Q/L075Q/V077I/D130A/G156W，K024A/L075R/V154I/G156W/V187Q，N011K/G023K/D111A/G156W/L264R，S058M/D130A/D137Q/G163P/L264R，S058M/G163P/L227M/N233Q/L264R，A018K/D027S/N033D/N073R/L075D/T189D，A018K/D027S/P029E/N033D/E045F/L075D，A018K/G023K/K024A/V077I/G156W/V187Q，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/A049V/G156W/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/G023Q/N094R/D130A/G156W/V187Q，A018K/K024A/D111A/V187N/T189Q/L264R，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/N094R/D111A/D130A/V154I/V187N，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/N233Q/L264R，D027E/D048Q/S058M/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/L227M/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/F051T/L075G/G091Q/V187N/I252Q，D027Q/S058M/L075R/D130A/V187T/I252Q，D027S/E056K/S058M/V187T/I252Q/L264R，D048Q/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D137Q/G163P/N233Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/D027Q/F051T/E056K/S058M/L075Q，G023K/D027S/L075R/D130A/V187T/I252Q，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/F051T/D130A/V187N/I252Q/L264R，G023K/K024A/L075Q/D111A/D130A/T189Q，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/K024A/L075R/D130A/V154I/V187N，G023K/L075G/D130A/V187T/I252Q/L264R，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/F051T/L075Q/D130A/V187H/I252Q，G023Q/K024A/V077I/D130A/V154I/G156W，K024A/L075Q/V077I/N094R/D130A/V187N，Q004D/N011K/D027N/I090F/N233Q/P256T，Q004D/N011K/D027S/S058M/N233Q/P256T，S058M/D130A/D137Q/G163P/L227M/L264R，S058M/D130A/D137Q/G163P/N233Q/L264R，S058M/D130A/G163P/L227M/N233Q/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/N094R/V154I/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/D130A/V154I/G156W/V187T，A018K/K024A/L075Q/V077I/N094R/D130A/G156W，D027E/D130A/D137Q/G163P/L227M/N233Q/L264R，D027E/S058M/D130A/D137Q/G163P/L227M/L264R，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/L227M/N233Q/L264R，G023K/D027S/F051T/S058M/G091Q/V187N/I252Q，G023Q/D027S/S058M/L075Q/D130A/I252Q/L264R，G023Q/F051T/G091Q/D130A/V187H/I252Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187N，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075R/N094R/V154I/G156W/V187N/T189Q，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/G156W/V187T
表8-9：下面示出，相比TLL(SEQIDNO：1)，表达性能指数≥0.5，且在使用全剂量的CS-61微布样测定中性能指数≥最大PI值的50％的TLL变体。
A018K/P029E/T189D，A018K/V154I/G156W，D027S/N033D/T189D，G023K/V187N/L264R，L075R/D130A/V187T，L075R/V187N/L264R，A018K/D027S/P029E/T189D，A018K/L075Q/V077I/N094R，A018K/N033D/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，G023K/D130A/V187T/L264R，P029E/N073R/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/G023Q/K024A/L075Q/G156W，A018K/K024A/N094R/D130A/V187N，D027E/D130A/G163P/N233Q/L264R，D027E/D137Q/G163P/N233Q/L264R，D130A/D137Q/G163P/N233Q/L264R，E045F/A049V/L075Q/V187T/T189D，E056K/L075R/D130A/V187T/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187N/L264R，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187T/L264R，K024A/L075R/V154I/G156W/V187Q，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/K024A/L075Q/V077I/D130A/V187N，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D048Q/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/N094R/D111A/G156W/V187T/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/L227M/N233Q/L264R，N011K/A018K/K024A/V077I/V154I/G156W/T189Q，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，A018K/G023K/K024A/V077I/D130A/V154I/G156W/V187T/T189Q，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，N011K/G023K/L075R/V077I/D130A/V154I/G156W/V187T/T189Q，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，N011K/A018K/G023K/K024A/L075R/V077I/D130A/V154I/V187T/T189Q，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R
表8-10：下面示出，相比参考序列SEQIDNO：2，在使用全剂量的CS-61微布样测定中性能指数＞1的TLL变体。
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30A/V154I，G023Q/L075Q/V077I/G156W/V187N，G091Q/D130A/V187H/I252Q/L264R，K024A/L075Q/D111A/G156W/V187T，K024A/L075Q/V077I/G156W/V187N，K024A/L075Q/V077I/V154I/V187N，K024A/L075R/V154I/G156W/V187Q，L075Q/D111A/V187N/T189Q/L264R，L075Q/N094R/D130A/G156W/V187T，L075Q/V077I/D130A/G156W/V187N，L075Q/V154I/V187N/T189Q/L264R，L075R/D130A/V154I/T189Q/L264R，L075R/D130A/V187T/T189Q/L264R，L075R/G156W/V187T/T189Q/L264R，L075R/V077I/N094R/V154I/G156W，N011K/D027N/E056K/N233Q/P256T，N011K/D027N/E056K/S058M/P256T，N011K/G023K/D111A/G156W/L264R，N011K/G023K/E056K/L075Q/T189Q，P029E/N033D/E045F/L075D/T189D，P029E/N033D/K074S/L075D/T189D，P029E/N033D/N073R/L075D/T189D，Q004D/D027S/S058M/N233Q/P256T，Q004D/N011K/D027Q/E056K/N233Q，S058M/G163P/L227M/N233Q/L264R，A018K/A049V/L075Q/G156W/V187T/T189D，A018K/D027S/N033D/E045F/L075D/T189D，A018K/D027S/P029E/N073R/L075D/T189D，A018K/E045F/A049V/L075Q/V077I/V187T，A018K/G023K/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075R/D130A/V187N，A018K/G023K/K024A/N094R/D130A/V154I，A018K/G023K/L075Q/D130A/G156W/V187N，A018K/G023K/L075Q/G156W/V187N/L264R，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023K/L075R/N094R/D130A/V187N，A018K/G023K/L075R/V077I/D130A/V154I，A018K/G023Q/A049V/L075Q/V077I/V187T，A018K/G023Q/E045F/A049V/L075Q/T189D，A018K/G023Q/E045F/A049V/L075Q/V077I，A018K/G023Q/E045F/A049V/L075Q/V187T，A018K/G023Q/E045F/L075Q/G156W/V187T，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/G023Q/L075Q/G156W/V187T/T189D，A018K/G023Q/L075Q/V077I/D130A/G156W，A018K/G023Q/L075Q/V077I/D130A/V187N，A018K/G023Q/L075Q/V077I/G156W/V187N，A018K/G023Q/L075Q/V077I/V187T/T189D，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，A018K/K024A/L075R/D130A/V187N/T189Q，A018K/K024A/V077I/N094R/D130A/G156W，A018K/L075Q/D130A/V154I/T189Q/L264R，A018K/L075Q/D130A/V187N/T189Q/L264R，A018K/L075R/D130A/V154I/G156W/V187N，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D027Q/F051T/L075G/D130A/I252Q/L264R，D027Q/F051T/L075Q/D130A/V187H/L264R，D027S/P029E/N033D/N073R/L075D/T189D，D048Q/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/G163P/N233Q/L264R，D111A/D130A/V154I/G156W/V187N/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/K024A/L075Q/N094R/D130A/G156W，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/L075G/D130A/V187T/I252Q/L264R，G023K/L075Q/D111A/D130A/V154I/V187N，G023K/L075Q/N094R/V154I/V187T/L264R，G023K/L075Q/V077I/D130A/G156W/V187Q，G023K/N094R/D111A/G156W/V187T/L264R，G023Q/A049V/L075Q/G156W/V187T/T189D，G023Q/D027Q/E056K/L075R/I252Q/L264R，G023Q/D027S/F051T/L075Q/V187T/I252Q，G023Q/D027S/L075Q/D111A/G156W/L264R，G023Q/E045F/A049V/L075Q/G156W/V187T，G023Q/E045F/A049V/L075Q/V077I/V187T，G023Q/E045F/L075Q/V077I/V187T/T189D，G023Q/K024A/L075Q/D130A/V154I/G156W，G023Q/K024A/L075Q/D130A/V154I/V187N，G023Q/K024A/L075Q/V154I/G156W/V187N，G023Q/L075Q/V077I/N094R/D130A/V187Q，G023Q/L075R/V154I/G156W/V187N/L264R，K024A/D111A/D130A/V187T/T189Q/L264R，K024A/L075Q/V154I/G156W/T189Q/L264R，K024A/L075Q/V154I/V187N/T189Q/L264R，K024A/L075Q/V154I/V187T/T189Q/L264R，K024A/N094R/V154I/G156W/T189Q/L264R，L075Q/D111A/D130A/V154I/G156W/T189Q，L075Q/D111A/D130A/V154I/V187N/T189Q，L075Q/N094R/D111A/G156W/T189Q/L264R，N011K/D027N/S058M/I090F/N233Q/P256T，N011K/D027Q/E056K/S058M/N233Q/P256T，N011K/D027S/S058M/I090F/N233Q/P256T，N011K/E056K/L075Q/D130A/V187N/T189Q，N011K/G023K/D027S/V154I/G156W/T189Q，N011K/G023Q/L075Q/D130A/V187N/T189Q，N011K/G023Q/L075Q/N094R/D130A/L264R，P029E/N033D/K074S/L075D/N101D/T189D，Q004D/D027S/E056K/S058M/N233Q/P256T，Q004D/N011K/D027Q/E056K/S058M/N233Q，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/N094R/G156W，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/K024A/L075R/V154I/G156W/V187Q，A018K/G023K/K024A/N094R/V154I/V187T/L264R，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/E045F/A049V/L075Q/G156W/V187T，A018K/G023Q/E045F/L075Q/V077I/G156W/V187T，A018K/G023Q/K024A/L075Q/V077I/D130A/G156W，A018K/G023Q/K024A/L075Q/V077I/N094R/V154I，A018K/G023Q/K024A/L075R/D130A/V154I/V187N，A018K/G023Q/K024A/L075R/N094R/G156W/V187N，A018K/G023Q/K024A/L075R/V077I/D130A/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，A018K/K024A/L075Q/D111A/G156W/V187N/L264R，A018K/K024A/L075Q/D130A/G156W/V187T/T189Q，A018K/L075Q/D111A/V154I/V187T/T189Q/L264R，A018K/L075Q/N094R/D111A/V154I/V187T/L264R，D027E/S058M/D130A/D137Q/G163P/N233Q/L264R，D027Q/F051T/L075G/D130A/V187H/I252Q/L264R，D027Q/F051T/L075Q/G091Q/D130A/V187T/I252Q，D027S/F051T/L075G/G091Q/V187T/I252Q/L264R，D027S/P029E/N033D/E045F/N073R/L075D/T189D，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/L227M/N233Q/L264R，E056K/L075Q/D111A/D130A/G156W/V187N/T189Q，G023K/D027Q/F051T/L075G/D130A/V187H/L264R，G023K/D027S/L075Q/N094R/V154I/G156W/T189Q，G023K/F051T/E056K/L075R/D130A/V187T/I252Q，G023K/K024A/L075Q/D111A/V154I/G156W/V187N，G023K/K024A/L075Q/D130A/V187T/T189Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N，G023K/K024A/L075Q/V077I/D130A/G156W/V187Q，G023K/L075Q/D111A/V154I/G156W/T189Q/L264R，G023K/L075Q/D130A/G156W/V187N/T189Q/L264R，G023K/L075Q/N094R/V154I/G156W/V187T/L264R，G023K/L075R/N094R/V154I/G156W/V187N/L264R，G023Q/D027S/L075Q/G091Q/V187T/I252Q/L264R，G023Q/E045F/A049V/V077I/G156W/V187T/T189D，G023Q/E045F/L075Q/V077I/G156W/V187T/T189D，G023Q/E056K/L075Q/D130A/V154I/G156W/T189Q，G023Q/F051T/L075R/G091Q/V187N/I252Q/L264R，G023Q/K024A/L075Q/V077I/D130A/G156W/V187N，G023Q/K024A/L075Q/V077I/D130A/V154I/G156W，G023Q/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/L075Q/D130A/V154I/V187N/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/G156W/V187Q，K024A/L075Q/D111A/D130A/V187N/T189Q/L264R，L075Q/D111A/D130A/G156W/V187N/T189Q/L264R，N011K/D027S/D111A/D130A/V154I/V187N/T189Q，N011K/E056K/L075Q/D130A/V154I/V187N/T189Q，N011K/G023K/L075Q/D111A/D130A/V154I/V187N，Q004D/N011K/D027Q/E056K/S058M/N233Q/P256T，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/D130Y/V154I/V187N/T189Q，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023K/K024A/L075R/D111A/V154I/V187T/L264R，A018K/G023K/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/G023K/L075R/V077I/N094R/D130A/G156W/V187Q，A018K/G023Q/E045F/L075Q/V077I/G156W/V187T/T189D，A018K/G023Q/K024A/L075Q/V077I/D130A/G156W/V187Q，A018K/G023Q/K024A/L075R/D130A/V154I/G156W/V187N，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，A018K/G023Q/L075Q/V077I/D130A/V154I/G156W/V187N，A018K/K024A/L075Q/V077I/N094R/D130A/V154I/G156W，A018K/K024A/L075R/D130A/V154I/G156W/V187T/L264R，A018K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023K/D027S/F051T/E056K/L075R/D130A/V187T/L264R，G023K/F051T/E056K/L075R/G091Q/D130A/V187T/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187T/L264R，G023K/K024A/L075R/N094R/V154I/G156W/V187T/T189Q，G023K/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，G023K/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/E056K/S058M/L075R/G091Q/V187N/I252Q，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，G023Q/D027S/L075Q/D130A/V154I/G156W/T189Q/L264R，G023Q/E056K/L075Q/D111A/V154I/V187N/T189Q/L264R，G023Q/E056K/L075Q/N094R/D111A/G156W/T189Q/L264R，G023Q/L075Q/N094R/D130A/V154I/G156W/T189Q/L264R，G023Q/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，N011K/G023K/D027S/L075R/V154I/G156W/V187N/T189Q，N011K/G023K/E056K/L075Q/D130A/V154I/T189Q/L264R，N011K/G023Q/D027S/L075Q/N094R/V154I/G156W/T189Q，N011K/G023Q/D027S/N094R/V154I/G156W/T189Q/L264R，N011K/G023Q/L075Q/N094R/V154I/G156W/T189Q/L264R，A018K/G023K/K024A/L075Q/N094R/D111A/D130A/V154I/T189Q，A018K/G023K/K024A/L075R/V077I/D130A/V154I/G156W/V187N，A018K/G023K/L075Q/N094R/D111A/V154I/G156W/T189Q/L264R，A018K/G023Q/D027S/L075Q/V077I/R108K/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/G156W/V187H，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/P029E/S058M/V077I/R108K/H135F/G156W，A018K/G023Q/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/K024A/L075R/N094R/D130A/V154I/G156W/V187N，A018K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V154I/G156W/L264R，D027S/E056K/L075Q/N094R/D111A/D130A/V187N/T189Q/L264R，G023K/D027S/F051T/E056K/S058M/L075Q/G091Q/V187N/L264R，G023K/D027S/F051T/E056K/S058M/L075R/G091Q/V187H/I252Q，G023K/D027S/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/E056K/L075Q/D111A/D130A/V154I/G156W/T189Q/L264R，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，G023K/K024A/N094R/D130A/V154I/G156W/V187T/T189Q/L264R，G023Q/D027S/F051T/E056K/L075G/G091Q/V187N/I252Q/L264R，G023Q/D027S/L075Q/N094R/D130A/V154I/G156W/V187N/L264R，N011K/G023K/L075R/D111A/D130A/V154I/V187N/T189Q/L264R，N011K/G023Q/L075Q/D111A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/E045F/A049V/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/E045F/A049V/S058M/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023Q/D027S/S058M/L075Q/V077I/R108K/H135F/G156W/V187H，A018K/G023Q/K024A/L075Q/V077I/N094R/D130A/V154I/G156W/V187Q，G023K/D027S/F051T/E056K/S058M/L075G/G091Q/D130A/I252Q/L264R，G023Q/D027S/F051T/E056K/L075R/G091Q/D130A/V187H/I252Q/L264R，G023Q/D027S/F051T/E056K/S058M/L075R/G091Q/D130A/V187H/I252Q，G023Q/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，N011K/G023K/L075R/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，N011K/G023Q/L075Q/N094R/D111A/D130A/V154I/G156W/V187N/L264R，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R，N011K/G023Q/D027S/L075Q/N094R/D111A/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/V187T/T189D，A018K/G023Q/D027S/P029E/E045F/A049V/S058M/N073S/L075Q/R108K/H135F/V187T/T189D，A018K/G023Q/D027S/P029E/N033D/E045F/A049V/S058M/N073S/K074S/L075Q/V077I/N101D/R108K/H135F/D137V/G156W/V187T/T189D
表8-11：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比参考序列SEQIDNO：2在使用全剂量的CS-61微布样测定中性能指数＞1的TLL变体。
L227M/L264R，A018K/L075D/T189D，A018K/P029E/T189D，A018K/V154I/G156W，D027S/N033D/T189D，G023K/V187N/L264R，G023Q/A049V/T189D，L075G/D130A/V187H，L075Q/V187N/L264R，L075R/D130A/V187T，L075R/V187N/L264R，P029E/N033D/E045F，V187N/T189Q/L264R，A018K/D027S/P029E/T189D，A018K/L075Q/V077I/N094R，A018K/N033D/L075D/T189D，D027S/N033D/L075D/T189D，D027S/P029E/L075D/T189D，D027S/P029E/N033D/E045F，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，D130A/G163P/L227M/L264R，G023K/D130A/V187T/L264R，P029E/N073R/L075D/T189D，A018K/D027S/P029E/N033D/L075D，A018K/G023Q/K024A/L075Q/G156W，A018K/K024A/L075R/D130A/V154I，A018K/K024A/N094R/D130A/V187N，A018K/P029E/N033D/N073R/L075D，D027E/D130A/G163P/L227M/L264R，D027E/D130A/G163P/N233Q/L264R，D027E/D137Q/G163P/N233Q/L264R，D027S/L075R/V154I/T189Q/L264R，D130A/D137Q/G163P/N233Q/L264R，E045F/A049V/L075Q/V187T/T189D，E056K/L075R/D130A/V187T/L264R，G023K/D027S/E056K/V187T/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/D130A/V187T，G023K/E056K/L075R/V187N/L264R，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/L075Q/V187H/I252Q/L264R，G023K/L075R/D130A/V187T/L264R，G023K/N094R/G156W/V187N/T189Q，K024A/L075R/V154I/G156W/V187Q，N011K/G023K/D111A/G156W/L264R，S058M/G163P/L227M/N233Q/L264R，A018K/G023K/L075Q/V077I/G156W/V187Q，A018K/G023Q/K024A/L075R/N094R/G156W，A018K/K024A/L075Q/N094R/V154I/G156W，A018K/K024A/L075Q/V077I/D130A/V187N，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D027E/D137Q/G163P/L227M/N233Q/L264R，D027Q/E056K/S058M/D130A/I252Q/L264R，D048Q/D137Q/G163P/L227M/N233Q/L264R，F051T/L075G/G091Q/D130A/I252Q/L264R，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/E056K/L075R/D130A/T189Q/L264R，G023K/E056K/L075R/D130A/V187N/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，G023K/L075G/D130A/V187T/I252Q/L264R，G023K/N094R/D111A/G156W/V187T/L264R，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075Q/N094R/G156W/V187Q，A018K/G023K/K024A/L075R/N094R/D130A/V154I，A018K/G023K/L075Q/V077I/N094R/V154I/G156W，A018K/G023Q/L075R/V077I/N094R/G156W/V187N，D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，D048Q/S058M/D130A/D137Q/L227M/N233Q/L264R，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/N094R/D130A/G156W/V187Q，A018K/G023K/K024A/L075Q/V077I/D130A/V154I/G156W，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，A018K/G023Q/K024A/L075R/V077I/N094R/D130A/G156W，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q，G023Q/D027S/E056K/L075R/D111A/V154I/G156W/L264R，G023Q/D027S/F051T/S058M/G091Q/D130A/V187N/I252Q，A018K/G023Q/D027S/P029E/L075Q/V077I/R108K/H135F/V187H，G023K/K024A/L075Q/N094R/V154I/G156W/V187N/T189Q/L264R，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/H135F/G156W/V187T，A018K/G023Q/D027S/P029E/S058M/L075Q/V077I/R108K/H135F/V187H，A018K/G023K/L075R/N094R/D111A/D130A/V154I/G156W/V187N/T189Q/L264R
表8-12：下面示出，相比TLL(SEQIDNO：1)表达性能指数≥0.5，且相比参考序列SEQIDNO：2在使用全剂量的CS-61微布样测定中性能指数≥最大PI值的50％的TLL变体。
A018K/P029E/T189D，D027S/N033D/T189D，L075R/D130A/V187T，L075R/V187N/L264R，A018K/D027S/P029E/T189D，A018K/L075Q/V077I/N094R，A018K/N033D/L075D/T189D，D027S/P029E/N033D/L075D，D048Q/D130A/G163P/L264R，P029E/N073R/L075D/T189D，A018K/D027S/P029E/N033D/L075D，D027E/D130A/G163P/N233Q/L264R，D027E/D137Q/G163P/N233Q/L264R，D130A/D137Q/G163P/N233Q/L264R，E056K/L075R/D130A/V187T/L264R，G023K/E056K/D130A/V187T/L264R，G023K/E056K/L075R/V187N/L264R，G023K/E056K/L075R/V187T/L264R，G023K/L075Q/V077I/D130A/G156W，G023K/L075R/D130A/V187T/L264R，K024A/L075R/V154I/G156W/V187Q，A018K/K024A/L075Q/V077I/D130A/V187N，D027E/D048Q/D130A/D137Q/G163P/L264R，D027E/D048Q/D137Q/G163P/L227M/L264R，D027E/D048Q/D137Q/G163P/N233Q/L264R，D027E/D130A/D137Q/G163P/L227M/L264R，D027E/D130A/G163P/L227M/N233Q/L264R，D048Q/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/L075R/V187N/I252Q/L264R，G023K/K024A/L075Q/V154I/G156W/V187Q，A018K/D027S/P029E/N033D/N073R/L075D/T189D，A018K/G023K/K024A/L075R/N094R/D130A/V154I，D048Q/S058M/D130A/D137Q/L227M/N233Q/L264R，Q004D/N011K/D027S/E056K/S058M/N233Q/P256T，A018K/G023K/K024A/L075Q/V077I/N094R/D130A/G156W，D027E/D048Q/D130A/D137Q/G163P/L227M/N233Q/L264R，G023K/D027S/F051T/E056K/L075Q/G091Q/V187H/I252Q

Claims (71)

1.一种脂解酶变体或其活性片段，所述脂解酶变体或其活性片段与亲本脂解酶相比包含至少两、三、四、五、六、七、八、九或十或更多个氨基酸修饰，其中第一氨基酸修饰在所述脂解酶变体中选自第4、11、18、23、24、27、29、33、45、48、49、51、56、58、73、75、77、90、91、94、111、130、137、154、156、163、187、189、227、233、252、256和264位的位置，其中所述变体的氨基酸位置是根据与SEQIDNO:1中示出的疏棉状嗜热丝孢菌(Thermomyceslanuginosa)脂肪酶的氨基酸序列的对应性进行编号的。
2.根据权利要求1所述的脂解酶变体或其活性片段，其中所述相比于亲本脂解酶的至少两、三、四、五、六、七、八、九或十或更多个氨基酸修饰在所述脂解酶变体中选自第4、11、18、23、24、27、29、33、45、48、49、51、56、58、73、75、77、90、91、94、111、130、137、154、156、163、187、189、227、233、252、256和264位的位置，其中所述变体的氨基酸位置是根据与SEQIDNO:1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。
3.一种脂解酶变体或其活性片段，所述脂解酶变体或其活性片段与亲本脂解酶相比存在两、三、四、五、六、七、八、九或十或更多个氨基酸修饰，其中所述两、三、四、五、六、七、八、九或十或更多个氨基酸修饰在所述脂解酶变体中选自第4、11、18、23、24、27、29、33、45、48、49、51、56、58、73、75、77、90、91、94、111、130、137、154、156、163、187、189、227、233、252、256和264位的位置，其中所述变体的氨基酸位置是根据与SEQIDNO:1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。
4.根据权利要求1至3中任一项所述的脂解酶变体或其活性片段，其中所述变体或其活性片段包含表2-1的氨基酸修饰，其中所述变体的氨基酸位置是根据与SEQIDNO:1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。
5.根据权利要求1至4中任一项所述的脂解酶变体或其活性片段，其中所述变体或活性片段具有脂解活性。
6.根据权利要求1至5中任一项所述的脂解酶变体或其活性片段，其中相对于所述亲本脂解酶，所述变体或活性片段水解丁酸对硝基苯酯、辛酸对硝基苯酯或棕榈酸对硝基苯酯的性能指数(pI)大于1.0。
7.根据权利要求6所述的脂解酶变体或其活性片段，其中所述性能指数是使用实施例1的丁酸对硝基苯酯、辛酸对硝基苯酯或棕榈酸对硝基苯酯测定法测得的。
8.根据权利要求7所述的脂解酶变体或其活性片段，其中所述性能指数在pH8时测得。
9.根据权利要求1至8中任一项所述的脂解酶变体或其活性片段，其中所述变体包含表3-1中所示的氨基酸修饰，其中所述变体的氨基酸位置是根据与SEQIDNO:1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。
10.根据权利要求9所述的脂解酶变体或其活性片段，其中相对于具有SEQIDNO:1序列的参考脂解酶，所述变体或活性片段水解丁酸对硝基苯酯的性能指数(pI)大于1.0。
11.根据权利要求10所述的脂解酶变体或其活性片段，其中所述性能指数是使用实施例1的丁酸对硝基苯酯测定法测得的。
12.根据权利要求11所述的脂解酶变体或其活性片段，其中所述性能指数在pH8时测得。
13.根据权利要求1至8中任一项所述的脂解酶变体或其活性片段，其中所述变体包含表3-7中所示的氨基酸修饰，其中所述变体的氨基酸位置是根据与SEQIDNO:1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。
14.根据权利要求13所述的脂解酶变体或其活性片段，其中相对于具有SEQIDNO:1序列的参考脂解酶，所述变体或活性片段水解辛酸对硝基苯酯的性能指数(pI)大于1.0。
15.根据权利要求14所述的脂解酶变体或其活性片段，其中所述性能指数是使用实施例1的辛酸对硝基苯酯测定法测得的。
16.根据权利要求15所述的脂解酶变体或其活性片段，其中所述性能指数在pH8时测得。
17.根据权利要求1至8中任一项所述的脂解酶变体或其活性片段，其中所述变体包含表3-13中所示的氨基酸修饰，其中所述变体的氨基酸位置是根据与SEQIDNO:1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。
18.根据权利要求17所述的脂解酶变体或其活性片段，其中相对于具有SEQIDNO:1序列的参考脂解酶，所述变体或活性片段水解棕榈酸对硝基苯酯的性能指数(pI)大于1.0。
19.根据权利要求18所述的脂解酶变体或其活性片段，其中所述性能指数是使用实施例1的棕榈酸对硝基苯酯测定法测得的。
20.根据权利要求19所述的脂解酶变体或其活性片段，其中所述性能指数在pH8时测得。
21.根据权利要求1至8中任一项所述的脂解酶变体或其活性片段，其中所述变体包含表3-25中所示的氨基酸修饰，其中所述变体的氨基酸位置是根据与SEQIDNO:1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。
22.根据权利要求9所述的脂解酶变体或其活性片段，其中与SEQIDNO:1相比，所述变体或活性片段具有增加的pNPP/pNPB或pNPO/pNPB比活性比值。
23.根据权利要求1至8中任一项所述的脂解酶变体或其活性片段，其中所述变体包含表3-31中所示的氨基酸修饰，其中所述变体的氨基酸位置是根据与SEQIDNO:1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。
24.根据权利要求9所述的脂解酶变体或其活性片段，其中与SEQIDNO:1相比，所述变体或活性片段具有增加的pNPB/pNPP比活性比值。
25.根据权利要求1至8中任一项所述的脂解酶变体或其活性片段，其中所述变体包含表4-1中所示的氨基酸修饰，其中所述变体的氨基酸位置是根据与SEQIDNO:1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。
26.根据权利要求25所述的脂解酶变体或其活性片段，其中相对于具有SEQIDNO:1序列的参考脂解酶，所述变体或活性片段的热稳定性的性能指数(pI)大于1.0。
27.根据权利要求1至8中任一项所述的脂解酶变体或其活性片段，其中所述变体包含表5-1中所示的氨基酸修饰，其中所述变体的氨基酸位置是根据与SEQIDNO:1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。
28.根据权利要求27所述的脂解酶变体或其活性片段，其中相对于具有SEQIDNO:1序列的参考脂解酶，所述变体或活性片段的洗涤剂稳定性的性能指数(pI)大于1.0。
29.根据权利要求1至8中任一项所述的脂解酶变体或其活性片段，其中所述变体包含表6-1中所示的氨基酸修饰，其中所述变体的氨基酸位置是根据与SEQIDNO:1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。
30.根据权利要求29所述的脂解酶变体或其活性片段，其中相对于具有SEQIDNO:1序列的参考脂解酶，所述变体或活性片段的LAS稳定性的性能指数(pI)大于1.0。
31.根据权利要求1至8中任一项所述的脂解酶变体或其活性片段，其中所述变体包含表7-1中所示的氨基酸修饰，其中所述变体的氨基酸位置是根据与SEQIDNO:1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。
32.根据权利要求31所述的脂解酶变体或其活性片段，其中相对于具有SEQIDNO:1序列的参考脂解酶，所述变体或活性片段的织物粘附性的性能指数(pI)大于1.0。
33.根据权利要求32所述的脂解酶变体或其活性片段，其中所述性能指数是使用实施例1的织物粘附性测定法测得的。
34.根据权利要求1至8中任一项所述的脂解酶变体或其活性片段，其中所述变体包含表7-7中所示的氨基酸修饰，其中所述变体的氨基酸位置是根据与SEQIDNO:1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。
35.根据权利要求34所述的脂解酶变体或其活性片段，其中相对于具有SEQIDNO:1序列的参考脂解酶，所述变体或活性片段的织物粘附性的性能指数(pI)小于1.0。
36.根据权利要求35所述的脂解酶变体或其活性片段，其中所述性能指数是使用实施例1的织物粘附性测定法测得的。
37.根据权利要求1至8中任一项所述的脂解酶变体或其活性片段，其中所述变体包含表8-1中所示的氨基酸修饰，其中所述变体的氨基酸位置是根据与SEQIDNO:1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。
38.根据权利要求37所述的脂解酶变体或其活性片段，其中相对于具有SEQIDNO:1序列的参考脂解酶，所述变体或活性片段的清洁性能的性能指数(pI)大于1.0。
39.根据权利要求38所述的脂解酶变体或其活性片段，其中所述性能指数是使用实施例1的CS-61微布样测定法，利用半剂量+辅剂测得的。
40.根据权利要求1至8中任一项所述的脂解酶变体或其活性片段，其中所述变体包含表8-7中所示的氨基酸修饰，其中所述变体的氨基酸位置是根据与SEQIDNO:1中示出的疏棉状嗜热丝孢菌脂肪酶的氨基酸序列的对应性进行编号的。
41.根据权利要求40所述的脂解酶变体或其活性片段，其中相对于具有SEQIDNO:1序列的参考脂解酶，所述变体或活性片段的清洁性能的性能指数(pI)大于1.0。
42.根据权利要求41所述的脂解酶变体或其活性片段，其中所述性能指数是使用实施例1的CS-61微布样测定法，利用全剂量测得的。
43.根据权利要求1至42中任一项所述的脂解酶变体或其活性片段，其中所述变体与脂解酶TLL同源物具有至少50％的同一性。
44.根据权利要求43所述的脂解酶变体，其中所述脂解酶TLL同源物来自选自以下的属：嗜热丝孢菌属(Thermomyces)、疣孢菌属(Verrucosispora)、糖单孢菌属(Saccharomonospora)、链霉菌属(Streptomyces)、小单孢菌属(Micromonospora)、链孢囊菌属(Streptosporangium)、拟无枝酸菌属(Amycolatopsis)、纤维单胞菌属(Cellulomonas)、放线束丝菌属(Actinosynnema)、克里布所菌属(Kribbella)、高温单孢菌属(Thermomonospora)、异常球菌属(Deinococcus)、动球菌属(Kineococcus)、拟诺卡氏菌属(Nocardiopsis)、弗兰克氏菌属(Frankia)、琼斯氏菌属(Jonesia)、假单胞菌属(Pseudomonas)、食酸菌属(Acidovorax)和类诺卡氏菌科(Nocardioidaceae)。
45.根据权利要求44所述的脂解酶变体，其中所述脂解酶TLL同源物来自嗜热丝孢菌属。
46.根据权利要求43或44所述的脂解酶变体，其中所述脂解酶TLL同源物为SEQIDNO:1中示出的疏棉状嗜热丝孢菌脂肪酶。
47.一种组合物，所述组合物包含至少一种根据权利要求1至46中任一项所述的脂解酶变体。
48.根据权利要求47所述的组合物，其中所述组合物是清洁组合物。
49.根据权利要求47或48所述的组合物，其中所述组合物是颗粒状、粉末状、固态、条棒状、液态、片状、凝胶状或糊状组合物。
50.根据权利要求47或48所述的组合物，其中所述组合物是单位剂量组合物。
51.根据权利要求47至50中任一项所述的组合物，其中所述清洁组合物是洗涤剂组合物。
52.根据权利要求47至51中任一项所述的组合物，其中所述组合物为衣物洗涤剂组合物、盘碟洗涤剂组合物或硬质表面清洁组合物。
53.根据权利要求52所述的组合物，其中所述盘碟洗涤剂是手洗盘碟洗涤剂组合物或机洗盘碟洗涤剂组合物。
54.根据权利要求52所述的组合物，其中所述清洁组合物是衣物洗涤剂组合物。
55.根据权利要求47至51中任一项所述的组合物，其中所述清洁组合物是衣物洗涤剂添加剂。
56.根据权利要求47至55中任一项所述的组合物，所述组合物还包含至少一种漂白剂。
57.根据权利要求47至56中任一项所述的组合物，其中所述清洁组合物不含磷酸盐。
58.根据权利要求47至56中任一项所述的组合物，其中所述清洁组合物含磷酸盐。
59.根据权利要求47至58中任一项所述的组合物，所述组合物还包含至少一种附加的酶。
60.根据权利要求59所述的组合物，其中所述附加的酶选自蛋白酶、半纤维素酶、纤维素酶、过氧化物酶、脂解酶、金属脂解酶、木聚糖酶、脂肪酶、磷脂酶、酯酶、过水解酶、角质酶、果胶酶、果胶酸裂解酶、甘露聚糖酶、角蛋白酶、还原酶、氧化酶、酚氧化酶、脂氧合酶、木质素酶、支链淀粉酶、鞣酸酶、戊聚糖酶、麦拉宁酶、β-葡聚糖酶、阿拉伯糖苷酶、透明质酸酶、软骨素酶、漆酶和淀粉酶。
61.一种水解脂肪酸酯或甘油三酯的方法，所述方法包括使所述脂肪酸酯或甘油三酯与根据权利要求1至46中任一项所述的脂解酶变体接触。
62.一种清洁方法，所述方法包括使表面或物品与包含至少一种根据权利要求1至46中任一项所述的脂解酶变体的组合物接触。
63.一种清洁方法，所述方法包括使表面或物品与根据权利要求47至60中任一项所述的组合物接触。
64.根据权利要求62或63所述的方法，所述方法还包括在分别使所述表面或物品与所述清洁组合物接触之后，冲洗所述表面或物品。
65.根据权利要求62至64中任一项所述的方法，其中所述物品为盘碟。
66.根据权利要求62至64中任一项所述的方法，其中所述物品为织物。
67.根据权利要求62至66中任一项所述的方法，所述方法还包括在使所述表面或物品与所述清洁组合物接触之后冲洗所述表面或物品的步骤。
68.根据权利要求67所述的方法，所述方法还包括在冲洗所述表面或物品之后干燥所述表面或物品的步骤。
69.一种清洁表面或物品的方法，所述方法包括：提供根据权利要求47至60中任一项所述的清洁组合物以及需要清洁的表面或物品；在适于清洁所述表面或物品的条件下，使所述清洁组合物与所述需要清洁的表面或物品接触，以产生清洁过的表面或物品。
70.根据权利要求69所述的方法，所述方法还包括冲洗所述清洁过的表面或物品以产生冲洗过的表面或物品的步骤。
71.根据权利要求69或70所述的方法，所述方法还包括干燥所述冲洗过的表面或物品的步骤。
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