CN111019916B - 一种d-氨基酸氧化酶突变体及其应用 - Google Patents

一种d-氨基酸氧化酶突变体及其应用 Download PDF

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CN111019916B
CN111019916B CN201811408085.1A CN201811408085A CN111019916B CN 111019916 B CN111019916 B CN 111019916B CN 201811408085 A CN201811408085 A CN 201811408085A CN 111019916 B CN111019916 B CN 111019916B
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田振华
程占冰
徐艳冰
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Shanghai Qizhou Ziyue Biotechnology Co ltd
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Abstract

本发明公开了一种D‑氨基酸氧化酶突变体，其序列包括将SEQ ID NO.1所示序列或与SEQ ID NO.1至少有76％同一性的序列的第54位氨基酸残基N、第58位氨基酸残基F、第211位氨基酸残基C和第213位氨基酸残基M进行突变后的序列；所述D‑氨基酸氧化酶突变体具有比野生型D‑氨基酸氧化酶高的酶活性、酶活稳定性和/或耐铵性。本发明还公开了所述D‑氨基酸氧化酶突变体在制备2‑氧代‑4‑(羟基甲基氧膦基)丁酸中的应用。本发明的D‑氨基酸氧化酶突变体酶活性很高、酶活稳定性提高和/或耐铵性增强，进而降低成本，利于工业化生产。

Description

一种D-氨基酸氧化酶突变体及其应用
技术领域
本发明属于生物技术领域，具体涉及一种D-氨基酸氧化酶突变体及其在制备2-氧代-4-(羟基甲基氧膦基)丁酸(PPO)中的应用。
背景技术
草铵膦是由赫斯特公司80年代开发的广谱触杀型除草剂。目前世界上的三大除草剂是草甘膦、草铵膦、百草枯，相对于草甘膦和百草枯，草铵膦具有优异的除草性能及较小的副作用。草铵膦有两种光学异构体，分别为D-草铵膦和L-草铵膦，但是只有L-草铵膦具有除草活性，因此发展L-草铵膦的方法对于提高原子经济性、降低使用成本、减轻环境压力具有重要意义。
目前，制备L-草铵膦的方法有很多是采用2-氧代-4-(羟基甲基氧膦基)丁酸(PPO)为底物，经转氨酶催化制备L-草铵膦的方法，其中US5221737A和EP0344683A记载了用谷氨酸作为氨基供体，自相应的酮酸4-(羟甲基氧膦基)-2-氧代丁酸通过来源于大肠杆菌的氨基转氨酶作用得到L-草铵膦的方法，反应体系中需要等量或过量的氨基供体谷氨酸，使产物难以纯化。CN1284858C对上述方法进行了改进，采用天冬氨酸作为氨基供体，自相应的酮酸4-(羟甲基氧膦基)-2-氧代丁酸通过天冬氨酸转氨酶作用得到L-草铵膦的方法，此方法中天冬氨酸转变为草酰乙酸，而草酰乙酸在含水介质中不稳定，并自发的脱羧为丙酮酸，丙酮酸可通过酶促反应除去，使得逆反应不能进行，反应只需要等摩尔的氨基供体和氨基受体。但是使用转氨酶的方法中所用的氨基供体多为氨基酸，成本较高。
Figure GDA0001914066270000011
另外还有采用2-氧代-4-(羟基甲基氧膦基)丁酸(PPO)为底物，经氨基酸脱氢酶催化，制备L-草铵膦的方法，如CN106978453A，其采用无机氨基供体，使得产物分离简单，成本降低。
Figure GDA0001914066270000012
以上都采用PPO为反应原料，但是PPO成本很高，从而导致生产L-草铵膦的成本很高，因此，US9834802B公开了以DL-草铵膦为原料制备L-草铵膦的方法，先由D-氨基酸氧化酶(DAAO)氧化D-草铵膦得到PPO，再由PPO经转氨酶催化得到L-草铵膦。
Figure GDA0001914066270000021
但是以DL-草铵膦为原料，仍然存在一个问题，因为市场上一般采用Strecker方法生产D,L-草铵膦，需要使用过量的氯化铵，所以生产的D,L-草铵膦都为铵盐，同时还有过量的氯化铵存在。铵离子的存在对DAAO酶的酶活有抑制作用，从而降低了酶活。例如本发明人发现D,L-草铵膦铵盐投料浓度高时，高浓度的铵离子对上述US9834802B中效果最佳的突变体的酶活有很大的抑制作用。
因此迫切需要寻找一种DAAO酶，不受铵离子的抑制，从而提高DAAO酶的酶活稳定性，进而提高D,L-草铵膦铵盐的投料浓度。
发明内容
本发明所要解决的技术问题是为了克服现有技术中D-氨基酸氧化酶的酶活性不高、酶活稳定性差或耐铵性差等缺陷，本发明提供了一种D-氨基酸氧化酶突变体以及其在制备2-氧代-4-(羟基甲基氧膦基)丁酸中的应用。本发明的D-氨基酸氧化酶突变体酶活性很高、酶活稳定性提高和/或耐铵性增强，进而降低成本，利于工业化生产。
本发明人在实验初期发现高浓度的铵离子对US9834802B中效果最佳的突变体的酶活有很大的抑制作用，因此筛选了其他与US9834802B中所述野生型序列的同源性比较高的一些序列的54、58、213位的突变体，发现铵离子对同源性为76％的N2DAAO序列的抑制作用最弱，因此选用N2DAAO序列进行进一步突变。本发明人经过大量筛选后发现，引入C211位点的突变时，所得的突变体在铵离子的作用下酶活性有所提高，进而对包括C211在内的位点进行组合突变并构建突变体文库，从中筛选出本发明所述的D-氨基酸氧化酶突变体。后续对US9834802B中所述野生型的序列同样进行包括C211位点在内的组合突变，惊奇地发现所得突变体确实对铵离子的耐受性有所提高，从而验证了C211位点对不同D-氨基酸氧化酶的耐铵性均有很大的影响，故C211位连同54、58、213位的突变可显著提高D-氨基酸氧化酶在铵离子存在环境下的酶活和酶活稳定性。
为解决上述技术问题，本发明旨在提供一种D-氨基酸氧化酶突变体，所述D-氨基酸氧化酶突变体的序列包括将SEQ ID NO.1所示序列或与SEQ ID NO.1至少有76％同一性的序列的第54位氨基酸残基N、第58位氨基酸残基F、第211位氨基酸残基C和第213位氨基酸残基M进行突变后的序列；所述D-氨基酸氧化酶突变体具有比野生型D-氨基酸氧化酶高的酶活性、酶活稳定性和/或耐铵性。
本发明所述耐铵性是指反应过程中本发明所述突变体对铵离子的耐受性增强，所述耐受性增强主要体现在本发明突变体在铵离子浓度较高时酶活稳定性高(即酶活损失少)，酶活仍然较高。
较佳地，所述与SEQ ID NO.1至少有76％同一性的序列如SEQ ID NO.3所示。
较佳地，所述D-氨基酸氧化酶突变体的序列包括将SEQ ID NO.1所示的第54位氨基酸残基N突变为A、C、G、S、T或V；和/或，第58位氨基酸残基F突变为亲水性或小位阻的氨基酸残基；和/或，第211位氨基酸残基C突变为A、D、G、H、L、M、S或Y；和/或，第213位氨基酸残基M突变为A、C、F、L、R、S、T、V或W的序列。
根据本发明，所述的亲水性或小位阻是指突变后的氨基酸残基和野生序列中氨基酸残基相比更亲水或空间位阻更小。所述的氨基酸可以为修饰或未修饰的天然氨基酸；本发明以天然氨基酸为例。
较佳地，所述第54位氨基酸残基N突变为V、第58位氨基酸残基F突变为Q、第211位氨基酸残基C突变为A、D、G、H、L、M、S或Y，和第213位氨基酸残基M突变为A、C、F、L、R、S、T、V或W；更佳地，所述第211位氨基酸残基C突变为A、G、M、S、Y或L，和/或，所述第213位氨基酸残基M突变为F、L、R、T或W；进一步更佳地，所述第211位氨基酸残基C突变为L或M，和/或，所述第213位氨基酸残基M突变为T。
较佳地，所述第211位氨基酸残基C突变为L，第213位氨基酸残基M突变为T，第54位氨基酸残基N突变为A、C、G、S、T或V，和第58位氨基酸残基F突变为A、G、H、K或Q；更佳地，所述第54位氨基酸残基N突变为A、G、S或T，和/或，所述第58位氨基酸残基F突变为H、K或Q；进一步更佳地，所述第54位氨基酸残基N突变为A，和/或，所述第58位氨基酸残基F突变为H或K；进一步更佳地，所述D-氨基酸氧化酶突变体的序列还包括将SEQ ID NO.1所示的第56位氨基酸残基T突变为N、S或L，所述第56位氨基酸残基T优选突变为N。
上述大写英文单字母代表如本领域技术人员熟知的氨基酸，根据本发明，在此代表的是相应的氨基酸残基。
较佳地，所述D-氨基酸氧化酶突变体的氨基酸序列如SEQ ID NO.5、SEQ ID NO.7、SEQ ID NO.9、SEQ ID NO.11、SEQ ID NO.13、SEQ ID NO.15、SEQ ID NO.17、SEQ ID NO.19、SEQ ID NO.21、SEQ ID NO.23、SEQ ID NO.25、SEQ ID NO.27、SEQ ID NO.29、SEQ IDNO.31、SEQ ID NO.33、SEQ ID NO.35、SEQ ID NO.37SEQ ID NO.39、SEQ ID NO.41、SEQ IDNO.43、SEQ ID NO.45、SEQ ID NO.47、SEQ ID NO.49、SEQ ID NO.51、SEQ ID NO.53、SEQ IDNO.55、SEQ ID NO.57、SEQ ID NO.59、SEQ ID NO.61、SEQ ID NO.63、SEQ ID NO.65、SEQ IDNO.68、SEQ ID NO.70、SEQ ID NO.72、SEQ ID NO.75、SEQ ID NO.79、SEQ ID NO.82、SEQ IDNO.90、SEQ ID NO.100、或SEQ ID NO.102所示。
较佳地，编码所述D-氨基酸氧化酶突变体的核苷酸序列如SEQ ID NO.6、SEQ IDNO.8、SEQ ID NO.10、SEQ ID NO.12、SEQ ID NO.14、SEQ ID NO.16、SEQ ID NO.18、SEQ IDNO.20、SEQ ID NO.22、SEQ ID NO.24、SEQ ID NO.26、SEQ ID NO.28、SEQ ID NO.30、SEQ IDNO.32、SEQ ID NO.34、SEQ ID NO.36、SEQ ID NO.38、SEQ ID NO.40、SEQ ID NO.42、SEQ IDNO.44、SEQ ID NO.46、SEQ ID NO.48、SEQ ID NO.50、SEQ ID NO.52、SEQ ID NO.54、SEQ IDNO.56、SEQ ID NO.58、SEQ ID NO.60、SEQ ID NO.62、SEQ ID NO.64、SEQ ID NO.66、SEQ IDNO.67、SEQ ID NO.69、SEQ ID NO.71、SEQ ID NO.73、SEQ ID NO.74、SEQ ID NO.76、SEQ IDNO.77、SEQ ID NO.78、SEQ ID NO.80、SEQ ID NO.81、SEQ ID NO.83、SEQ ID NO.84、SEQ IDNO.85、SEQ ID NO.86、SEQ ID NO.87、SEQ ID NO.88、SEQ ID NO.89、SEQ ID NO.91、SEQ IDNO.101、或SEQ ID NO.103所示。
为解决上述技术问题，本发明旨在提供一种分离的核酸，所述核酸编码上述的D-氨基酸氧化酶突变体。
为解决上述技术问题，本发明旨在提供一种包含上述核酸的重组表达载体。
为解决上述技术问题，本发明旨在提供一种包含上述核酸或上述重组表达载体的转化体。
为解决上述技术问题，本发明旨在提供一种上述的D-氨基酸氧化酶突变体在制备2-氧代-4-(羟基甲基氧膦基)丁酸中的应用。
本发明所述酶活性包括酶的比酶活和酶活的特性。
在符合本领域常识的基础上，上述各优选条件，可任意组合，即得本发明各较佳实例。
本发明所用试剂和原料均市售可得。
本发明的积极进步效果在于：本发明的D-氨基酸氧化酶突变体酶活性很高、酶活稳定性提高和/或耐铵性增强，进而降低成本，利于工业化生产。
具体实施方式
下面通过实施例的方式进一步说明本发明，但并不因此将本发明限制在所述的实施例范围之中。下列实施例中未注明具体条件的实验方法，按照常规方法和条件，或按照商品说明书选择。
本发明中的实验方法如无特别说明均为常规方法，基因克隆操作具体可参加J.萨姆布鲁克等编的《分子克隆实验指南》。
本发明中的氨基酸简写符号如无特殊说明均为本领域常规，具体简写符号对应的氨基酸如表1所示。
表1
氨基酸名称 三字母符号 单字母符号 氨基酸名称 三字母符号 单字母符号
丙氨酸(alanine) Ala A 亮氨酸(leucine) Leu L
精氨酸(arginine) Arg R 赖氨酸(lysine) Lys K
天冬酰胺(asparagine) Asn N 甲硫氨酸(methionine) Met M
天冬氨酸(aspartic acid) Asp D 笨丙氨酸(phenylalanine) Phe F
半胱氨酸(cysteine) Cys C 脯氨酸(proline) Pro P
谷氨酰胺(glutanine) Gln Q 丝胺酸(serine) Ser S
谷氨酸(glutamic acid) Glu E 苏氨酸(threonine) Thr T
甘氨酸(Glicine) Gly G 色氨酸(tryptophan) Trp W
组氨酸(histidine) His H 酪氨酸(tyrosine) Tyr Y
异亮氨酸(isoleucine) Ile I 颉氨酸(valine) Val V
所述氨基酸对应的密码子也为本领域常规，具体氨基酸与密码子的对应关系如表2所示。
表2
Figure GDA0001914066270000051
Figure GDA0001914066270000061
Pet28a和bugbuster protein extraction reagent购买自Novagen公司；NdeI酶、HindIII酶购买自Thermo Fisher公司，BL21感受态细胞购买自北京鼎国昌盛生物技术有限责任公司；过氧化氢酶购买自山东丰泰生物科技有限公司。
实施例1野生型D-氨基酸氧化酶(DAAO)的制备
从NCBI检索到GenBank登录号为KWU45700的来源于Rhodotorula sp.JG-1b的N2DAAO酶，全合成野生型(wt)N2DAAO酶基因，基因合成公司为苏州金唯智生物科技有限公司(苏州工业园区星湖街218号生物纳米科技园C3楼)。
将合成得到的wtN2DAAO酶基因连接质粒pET28a(具体方法参见J.Am.Chem.Soc.,2017,139(32),11241-11247)，酶切位点NdeI和HindIII，将酶连好的载体，转化至宿主大肠杆菌BL21感受态细胞；将其接种LB液体培养基于37℃下，200rpm摇床培养，待OD600至0.8左右时，取菌液加入终浓度为25％的无菌甘油编号后，置于-80℃低温冰箱保藏备用。
LB液体培养基组成：蛋白胨10g/L，酵母粉5g/L，NaCl 10g/L，用去离子水溶解后定容，121℃灭菌20min，待用；
将上述置于-80℃低温冰箱保藏备用的含有酶基因的工程菌在经平皿划线活化后，挑单菌落接种至含50μg/ml卡那霉素的5ml LB液体培养基中，37℃震荡培养12h，按2％接种量转接至150ml同样含50μg/ml卡那霉素的新鲜LB液体培养基中，37℃震荡至OD600达到0.8左右时，降温至30℃，加入IPTG至其终浓度为0.5mM，诱导培养16h，培养结束后，将培养液10000rpm离心10min，弃上清液，收集菌体，置于-20℃冰箱中保存，待用。
将培养结束后收集到的菌体，用50mM pH8.0磷酸缓冲液洗涤两次，之后重悬于50mLpH8.0的磷酸缓冲液中，均质破碎，破碎液离心去除沉淀，得到含重组wtN2DAAO酶的粗酶液。
实施例2 D-氨基酸氧化酶(DAAO)突变体文库(211位、213位)的构建
在实施例1中所述的wtN2DAAO序列的基础上突变了第54、58位(具体为N54V、F58Q)之后得到了突变的D-氨基酸氧化酶序列，根据该序列合成基因N2DAAO(N54V、F58Q)，基因合成公司为苏州金唯智生物科技有限公司(苏州工业园区星湖街218号生物纳米科技园C3楼)。然后以NdeI和HindIII酶切位点引入质粒pET28a，构建质粒pET28a-N2DAAO(质粒构建方法参见J.Am.Chem.Soc.,2017,139(32),11241-11247)。以质粒pET28a-N2DAAO为模板，PCR扩增目的条带。
其中，针对突变的D-氨基酸氧化酶序列(N54V、F58Q)的第211位、213位进行突变的突变体文库构建设计PCR的引物序列，具体如表3所示：
表3
Figure GDA0001914066270000071
其中，N代表A、G、C、T中任何一种核苷酸，M代表A或C，K代表G或T；其根据所述位点需突变成的氨基酸的编码核苷酸来选择，如A166-正向引物中的NNK可以代表AAG(赖氨酸)、AAT(天冬氨酸)、AGG(精氨酸)或AGT(丝氨酸)等，具体氨基酸所对应的核苷酸可参见表2。
PCR扩增体系为：
试剂 用量(μL)
2×PCR buffer(含高保真酶) 25
正向引物 1
反向引物 1
模板 1
去离子水 22
PCR扩增程序如下：
Figure GDA0001914066270000081
对PCR产物进行DpnI消化，37℃，2h。反应完成转化至BL21感受态细胞，涂布在含有50μg/mL卡纳霉素的LB培养基，37℃培养过夜，收菌，得到包含突变体文库的转化子。
实施例3高通量筛选突变体文库
按照如下实验步骤进行筛选：
将实施例2中所得转化子接种96孔板培养，加IPTG 30℃过夜诱导。之后收菌，加bugbuster protein extraction reagent裂解，离心得DAAO突变体酶液。
酶标仪检测方法：取100μL pH为8.0的100mM底物(消旋草铵膦，购自上海阿拉丁生化科技股份有限公司)，再加50μL的显色液(含60mg/mL的TBHBA(3-羟基-2,4,6-三溴苯甲酸)和100mg/mL的4-AAP(4-氨基安替比林))和25μL HRP(辣根过氧化物酶，0.1mg/mL)，最后加25μL的上述DAAO突变体酶液，得酶标板200μL反应体系，在30℃，pH 8.0的条件下对其进行分析。分别在0min和20min记录510nm处的吸光度，取差值，以野生型为参照系，筛选阳性克隆子。
将所筛选出的阳性克隆子进行培养，方法如下：
LB液体培养基组成：蛋白胨10g/L，酵母粉5g/L，NaCl 10g/L，用去离子水溶解后定容，121℃灭菌20min，待用。
挑单克隆接种至含50μg/ml卡纳霉素的5ml LB液体培养基中，37℃震荡培养12h。按2％接种量转接至150ml同样含50μg/ml卡纳霉素的新鲜LB液体培养基中，37℃震荡至OD600值达到0.8左右时，加入IPTG至其终浓度为0.5mM，30℃诱导培养16h。培养结束后，将培养液10000rpm离心10min，弃上清液，收集菌体，置于-20℃超低温冰箱中保存，待用。
将培养结束后收集到的菌体，用50mM pH 8.0的磷酸缓冲液洗涤菌体两次，之后将菌体重悬于pH 8.0的磷酸缓冲液中，低温高压均质破碎，破碎液离心去除沉淀，得到的上清液为初步的含重组DAAO突变体粗酶液。
突变体复筛的酶活检测方法为：
5mL反应体系中，加入1mL 500mM的D，L-草铵膦铵盐，0.25mL的上述初步的含重组DAAO突变体粗酶液，1.25mL辣根过氧化物酶(HRP)，2.5mL显色染料溶液(含60mg/mL的TBHBA和100mg/mL的4-AAP)，反应介质为pH为8.0的磷酸氢二钠-磷酸二氢钠缓冲液，于30℃摇床震荡反应，每隔2min，取反应液扫描510nm下的吸光值，做吸光度和时间(min)的酶反应动力学曲线，根据曲线斜率计算酶活。结果如表4所示。
单位酶活的定义：在特定反应条件(30℃)下，每分钟生成1μmol H2O2所需要的酶量，酶活单位是U。
比酶活为每毫克酶蛋白所含的活力单位，计算公式：酶活/蛋白含量，单位是U/mg或U/g。
表4
Figure GDA0001914066270000091
Figure GDA0001914066270000101
备注：表格中*代表比酶活在0-0.10U/mg之间，**代表比酶活在0.1-1.0U/mg之间，***代表比酶活在1.0-20U/mg之间，****代表比酶活在20-50U/mg之间。
由表4可以看出，所得突变体的比酶活均高于野生型N2DAAO的比酶活。其中比酶活最高的为DAAO氧化酶突变体11，其突变位点第54位的N突变为V，第58位的F突变为Q，第211位的C突变为L，第213位的M突变为T。
实施例4针对实施例3中的DAAO氧化酶突变体11的第54位、56位、58位进行突变的突变体文库的构建
针对DAAO氧化酶突变体11的第54位、56位、58位突变体文库构建所设计的引物序列如表5所示。
表5
Figure GDA0001914066270000102
其中，N代表A、G、C、T中任何一种核苷酸，M代表A或C，K代表G或T；其根据所述位点需突变成的氨基酸的编码核苷酸来选择，如A166-正向引物中的NNK可以代表AAG(赖氨酸)、AAT(天冬氨酸)、AGG(精氨酸)或AGT(丝氨酸)等，具体氨基酸所对应的核苷酸可参见表2。
根据文献J.Am.Chem.Soc.,2017,139(32),11241-11247中公开的方法构建了质粒模板pET28a-DAAO氧化酶突变体12，以pET28a-DAAO氧化酶突变体11为模板，进行PCR扩增目的条带。
扩增反应体系为：
试剂 用量(μL)
2XPCR buffer(含高保真酶) 25
引物F 1
引物R 1
模板 1
去离子水 22
扩增程序如下：
Figure GDA0001914066270000111
对PCR产物进行DpnI消化，37℃，2h。反应完成转化至BL21感受态细胞，涂布在含有50μg/mL卡纳霉素的LB培养基，37℃培养过夜，收菌，得到包含突变体文库的转化子。
实施例5酶活稳定性检测与NH4 +离子浓度影响检测
将实施例4中所得转化子接种96孔板培养，加IPTG 30℃过夜诱导。之后收菌，加bugbuster protein extraction reagent裂解，离心得突变子酶液。
将突变子酶液在50℃处理2h。按照实施例3中所述酶标仪检测方法分析阳性克隆的热稳定突变效果，筛选阳性克隆子。
将所选阳性克隆子进行培养，方法如下：
LB液体培养基组成：蛋白胨10g/L，酵母粉5g/L，NaCl 10g/L，用去离子水溶解后定容，121℃灭菌20min，待用。
挑单克隆接种至含50μg/ml卡纳霉素的5ml LB液体培养基中，37℃震荡培养12h。按2％接种量转接至150ml同样含50μg/ml卡纳霉素的新鲜LB液体培养基中，37℃震荡至OD600值达到0.8左右时，加入IPTG至其终浓度为0.5mM，30℃诱导培养16h。培养结束后，将培养液10000rpm离心10min，弃上清液，收集菌体，置于-20℃超低温冰箱中保存，待用。
将培养结束后收集到的菌体，用50mM pH 8.0磷酸缓冲液洗涤菌体两次，之后将菌体重悬于pH 8.0的磷酸缓冲液中，低温高压均质破碎，破碎液离心去除沉淀，得到的上清液为含重组DAAO突变体粗酶液。
对所得的DAAO突变体粗酶液进行突变体复筛的酶活检测，方法同实施例3。所得结果如表6所示，其中2h后剩余酶活％数值反应了突变体酶活的稳定性。
表6
Figure GDA0001914066270000121
Figure GDA0001914066270000131
备注：表格中酶活这一列中，*代表酶活在0-0.10U/mg之间，**代表比酶活在0.1-2.0U/mg之间，***代表比酶活在2.0-3.5U/mg之间，****代表比酶活在3.5-10U/mg之间。
2h后剩余酶活这一列中，*代表2h后剩余酶活在0-50％之间，**代表2h后剩余酶活在50-80％之间，***代表2h后剩余酶活在80-100％之间。
表中有些突变体的氨基酸序列一样，但是DNA序列不一样，其密码子不相同，相关的氨基酸序列和DNA序列详见序列表。
结果显示，除突变体34以外，上述其他突变体的酶活和酶活稳定性均比野生型高。其中，突变体36、突变体41、突变体42、突变体43、突变体45、突变体46、突变体49、突变体50的酶活较突变体11有较大的提高，且酶活稳定性也有较大的提高。此外由表中可以看出，上述突变体氨基酸序列相同，核苷酸序列不同时，其对铵离子的耐受性不同。
接着将上述所得突变体进行NH4 +离子浓度影响检测，方法如下：
在如实施例3中所述的酶标仪检测方法中的酶标板200μL反应体系中，加入终浓度为2M的NH4Cl，分析加入NH4Cl前后的酶活，确认NH4Cl浓度对酶活的影响。所得酶活数据如表7所示：
表7
Figure GDA0001914066270000141
Figure GDA0001914066270000151
备注：表格中*代表酶活残留比例在0-0.30之间，**代表酶活残留比例在0.3-0.5之间，***代表酶活残留比例在0.5-0.6之间，****代表酶活残留比例在0.6-1之间。
以上结果显示，上述突变体对铵离子的耐受性均比野生型高。其中，突变体42、突变体49的酶活、酶活稳定性、且对铵离子的耐受性较突变体11有较大的提高。此外由表中可以看出，上述突变体氨基酸序列相同，核苷酸序列不同时，其对铵离子的耐受性不同。
实施例6其它DAAO酶211位和213位组合突变的酶活
与GenBank登录号为KWU45700的序列(SEQ ID NO.1)的同一性为76％的rtDAAO酶，其来源于Rhodosporidium toruloidesUniProtKB/Swiss-Prot P80324，序列如SEQ IDNO.3所示。在该序列的基础上突变了第54、58和213位(具体为N54V、F58Q、M213S)之后得到了突变的D-氨基酸氧化酶序列，根据该序列合成基因rtDAAO(N54V-F58Q-M213S)，基因合成公司同上。针对此rtDAAO(N54V-F58Q-M213S)酶定点突变54位为A，56位为N，58位为H，211位为L、213位为T，得到如表8所示的突变体。对所得突变体进行突变体复筛的酶活检测，方法同实施例3，计算酶活和比酶活数据如表8所示。
按照如实施例3中所述的酶标仪检测方法中的酶标板200μL反应体系检测所得突变体的耐铵性，加入终浓度为2M的NH4Cl，分析加入NH4Cl前后的酶活，确认NH4Cl浓度对酶活的影响。所得酶活数据如表9所示。
表8
Figure GDA0001914066270000152
Figure GDA0001914066270000161
表9
Figure GDA0001914066270000162
由上表可以看出，突变体56(N54A-F58H-C211L-M213T)的耐铵性有所提高；突变体57(N54A-T56N-F58H-C211L-M213T)在酶活变化不大的情况下，提高了铵耐受性。
SEQUENCE LISTING
<110> 上海弈柯莱生物医药科技有限公司
<120> 一种D-氨基酸氧化酶突变体及其应用
<130> P180115286C
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ctgaccgacg gtccgcgtca ggctaaatgg gaagaatcta ccttcaaaaa atgggttgaa 240
ctggttccga ccggtcacgc tatgtggctg aaaggtaccc gtcgtttcgc tcagaacgaa 300
gacggtctgc tgggtcactg gtacaaagac atcaccccga actaccgtcc gctgccgtct 360
tctgaatgcc cgccgggtgc tatcggtgtt acctacgaca ccctgtctgt tcacgctccg 420
aaatactgcc agtacctggc tcgtgaactg cagaaactgg gtgctacctt cgaacgtcgt 480
accgttacct ctctggaaca ggctttcgac ggtgctgacc tggttgttaa cgctaccggt 540
ctgggtgcta aatctatcgc tggtatcgac gaccaggctg ctgaaccgat ccgtggtcag 600
accgttctgg ttaaatctcc gtgcaaacgt tgcaccatgg actcttctga cccggcttct 660
ccggcttaca tcatcccgcg tccgggtggt gaagttatct gcggtggtac ctacggtgtt 720
ggtgactggg acctgtctgt taacccggaa accgttcagc gtatcctgaa acactgcctg 780
cgtctggacc cgaccatctc ttctgacggt accatcgaag gtatcgaagt tctgcgtcac 840
aacgttggtc tgcgtccggc tcgtcgtggt ggtccgcgtg ttgaagctga acgtatcgtt 900
ctgccgctgg accgtaccaa atctccgctg tctctgggtc gtggttctgc tcgtgctgct 960
aaagaaaaag aagttaccct ggttcacgct tacggtttct cttctgctgg ttaccagcag 1020
tcttggggtg ctgctgaaga cgttgctcag ctggttgacg aagctttcca gcgttaccac 1080
ggtgctgctc gtgaatctaa actg 1104
<210> 5
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N2DAAO(N54V-F58Q-M213S)
<400> 5
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Cys Thr Ser Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 6
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N2DAAO(N54V-F58Q-M213S)
<400> 6
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc tgtaccagcg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 7
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211S-M213T
<400> 7
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Ser Thr Thr Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 8
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211S-M213T
<400> 8
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc agtaccacgg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 9
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211A-M213T
<400> 9
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Ala Thr Thr Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 10
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211A-M213T
<400> 10
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc gcgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 11
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211G-M213F
<400> 11
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Gly Thr Phe Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 12
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211G-M213F
<400> 12
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ggtacctttg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 13
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211M-M213R
<400> 13
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Met Thr Arg Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 14
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211M-M213R
<400> 14
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc acgacccatg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 15
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211G-M213T
<400> 15
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Gly Thr Thr Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 16
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211G-M213T
<400> 16
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc gggaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 17
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211A-M213R
<400> 17
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Ala Thr Arg Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 18
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211A-M213R
<400> 18
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc gctacccgag acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 19
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211G-M213S
<400> 19
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Gly Thr Ser Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 20
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211G-M213S
<400> 20
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc gggacctcgg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 21
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211D-M213V
<400> 21
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Asp Thr Val Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 22
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211D-M213V
<400> 22
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc gataccgtgg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 23
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211L-M213T
<400> 23
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Leu Thr Thr Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 24
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211L-M213T
<400> 24
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 25
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q- C211A-M213C
<400> 25
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Ala Thr Cys Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 26
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q- C211A-M213C
<400> 26
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc gctacctgtg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 27
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q- C211A-M213L
<400> 27
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Ala Thr Leu Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 28
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q- C211A-M213L
<400> 28
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc gcgacccttg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 29
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q- C211G-M213L
<400> 29
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Gly Thr Leu Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 30
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q- C211G-M213L
<400> 30
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc gggaccattg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 31
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q- C211M-M213S
<400> 31
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Met Thr Ser Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 32
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q- C211M-M213S
<400> 32
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc atgacctctg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 33
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q- C211A-M213S
<400> 33
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Ala Thr Ser Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 34
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q- C211A-M213S
<400> 34
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc gcgaccagtg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 35
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q- C211H-M213L
<400> 35
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg His Thr Leu Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 36
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q- C211H-M213L
<400> 36
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc cataccctgg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 37
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211S-M213C
<400> 37
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Ser Thr Cys Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 38
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211S-M213C
<400> 38
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc tcgacctgtg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 39
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211M-M213L
<400> 39
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Met Thr Leu Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 40
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211M-M213L
<400> 40
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc atgaccctgg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 41
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211S-M213L
<400> 41
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Ser Thr Leu Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 42
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211S-M213L
<400> 42
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc tctaccttgg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 43
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211Y-M213F
<400> 43
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Tyr Thr Phe Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 44
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211Y-M213F
<400> 44
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc tatacctttg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 45
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211S-M213V
<400> 45
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Ser Thr Val Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 46
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211S-M213V
<400> 46
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc tctaccgtgg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 47
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211M-M213T
<400> 47
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Met Thr Thr Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 48
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211M-M213T
<400> 48
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc atgaccacgg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 49
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211G-M213W
<400> 49
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Gly Thr Trp Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 50
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211G-M213W
<400> 50
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc gggacctggg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 51
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211A-M213A
<400> 51
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Ala Thr Ala Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 52
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211A-M213A
<400> 52
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc gcgaccgctg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 53
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211A-M213V
<400> 53
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Ala Thr Val Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 54
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V-F58Q-C211A-M213V
<400> 54
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc gcgaccgtgg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 55
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54C- T56T- F58G-C211L-M213T
<400> 55
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Cys Trp Thr Pro Gly Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Leu Thr Thr Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 56
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54C- T56T- F58G-C211L-M213T
<400> 56
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcat gttggacccc ggggatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 57
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54S- T56T- F58H-C211L-M213T
<400> 57
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Ser Trp Thr Pro His Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Leu Thr Thr Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 58
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54S- T56T- F58H-C211L-M213T
<400> 58
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcaa gttggacccc gcatatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 59
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54G- T56T- F58K-C211L-M213T
<400> 59
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Gly Trp Thr Pro Lys Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Leu Thr Thr Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 60
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54G- T56T- F58K-C211L-M213T
<400> 60
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag ggtggacccc gaagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 61
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54A- T56T- F58H-C211L-M213T
<400> 61
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Ala Trp Thr Pro His Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Leu Thr Thr Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 62
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54A- T56T- F58H-C211L-M213T
<400> 62
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag cttggacccc gcatatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 63
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54A- T56T- F58K-C211L-M213T
<400> 63
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Ala Trp Thr Pro Lys Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Leu Thr Thr Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 64
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54A- T56T- F58K-C211L-M213T
<400> 64
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag cgtggacccc gaagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 65
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54C- T56T- F58A-C211L-M213T
<400> 65
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Cys Trp Thr Pro Ala Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Leu Thr Thr Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 66
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54C- T56T- F58A-C211L-M213T
<400> 66
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcat gttggacccc ggcgatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 67
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54A- T56T- F58H-C211L-M213T
<400> 67
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag cttggacccc gcatatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 68
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54G- T56T- F58A-C211L-M213T
<400> 68
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Gly Trp Thr Pro Ala Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Leu Thr Thr Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 69
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54G- T56T- F58A-C211L-M213T
<400> 69
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag ggtggacccc ggctatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 70
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54V- T56T- F58H-C211L-M213T
<400> 70
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro His Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Leu Thr Thr Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 71
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54V- T56T- F58H-C211L-M213T
<400> 71
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag tttggacccc gcatatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 72
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54T- T56T- F58Q-C211L-M213T
<400> 72
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Thr Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Leu Thr Thr Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 73
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54T- T56T- F58Q-C211L-M213T
<400> 73
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcaa cttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 74
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54A- T56T- F58H-C211L-M213T
<400> 74
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag cgtggacccc gcatatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 75
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> 54A- T56T- F58Q-C211L-M213T
<400> 75
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Ala Trp Thr Pro Gln Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Leu Thr Thr Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 76
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> 54A- T56T- F58Q-C211L-M213T
<400> 76
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag cttggacccc gcagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 77
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54A- T56T- F58K-C211L-M213
<400> 77
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag cgtggactcc gaagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 78
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54A- T56T- F58K-C211L-M213T
<400> 78
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag cgtggacgcc gaagatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 79
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54A- T56N- F58H-C211L-M213T
<400> 79
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Ala Trp Asn Pro His Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Leu Thr Thr Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 80
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54A- T56N- F58H-C211L-M213T
<400> 80
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag cttggaatcc gcatatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 81
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54A- T56N- F58H-C211L-M213T
<400> 81
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag cttggaatcc gcatatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 82
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54A- T56S- F58H-C211L-M213T
<400> 82
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Ala Trp Ser Pro His Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Leu Thr Thr Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 83
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54A- T56S- F58H-C211L-M213T
<400> 83
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag cttggtcgcc gcatatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 84
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54A- T56S- F58H-C211L-M213T
<400> 84
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag cttggtcgcc gcatatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 85
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54A- T56S- F58H-C211L-M213T
<400> 85
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag cgtggtcgcc gcatatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 86
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54A- T56T- F58H-C211L-M213T
<400> 86
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag cgtggacgcc gcatatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 87
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54A- T56T- F58H-C211L-M213T
<400> 87
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag cgtggactcc gcatatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 88
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54A- T56S- F58H-C211L-M213T
<400> 88
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag cgtggagtcc gcatatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 89
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54A- T56N- F58H-C211L-M213T
<400> 89
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag cgtggaatcc gcatatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 90
<211> 372
<212> PRT
<213> Artificial Sequence
<220>
<223> N54A- T56L- F58H-C211L-M213T
<400> 90
Met Thr Gln Asn Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Cys Ala Leu Ala Leu Ala Gln Lys Gly Tyr Lys Val His Val
20 25 30
Val Ala Arg Asp Leu Pro Glu Asp Thr Val Ala Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Ala Trp Leu Pro His Met Ser Lys Glu Ala Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ala Thr Phe Lys Gln Trp Val Asp
65 70 75 80
Phe Val Pro Gln Gly Leu Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Glu Thr Glu Ala Asp Leu Leu Gly His Trp Tyr Lys Asp Ile Val
100 105 110
Pro Asn Tyr Arg His Leu Asn Pro Ser Asp Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val Asn Ala Pro Lys Phe Cys Gln
130 135 140
Tyr Leu Gln Arg Glu Ala Gln Lys Leu Gly Val Thr Phe Glu Arg Arg
145 150 155 160
Leu Val Thr Ser Leu Glu Gln Ile Ala Asp Gly Ala Asp Leu Ile Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Val Glu Asp Gln
180 185 190
Glu Val Glu Pro Ile Arg Gly Gln Thr Val Leu Ile Lys Ser Asn Cys
195 200 205
Lys Arg Leu Thr Thr Asp Ser Ser Asp Pro Lys Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Leu Val
225 230 235 240
Gly Asn Tyr Asp Leu Ser Val Asp Pro Ala Thr Ile Pro Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Ser Ile Ser Thr Asp Gly Thr Leu
260 265 270
Glu Gly Ile Glu Ile Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Leu Glu Arg Val Ser Phe Pro Leu Lys
290 295 300
Arg Gly Gln Ser Leu Leu Ala Leu Gly Thr Ala Lys Ala Ala Glu Gly
305 310 315 320
Lys Ala Ser Arg Thr Val Pro Val Val His Ala Tyr Gly Phe Ser Ser
325 330 335
Ala Gly Tyr Gln Gln Gly Trp Gly Ala Ala Leu Glu Val Arg Asp Leu
340 345 350
Val Asp Gln Ala Ile Gly Ser Ser Ser Ser Ala Ser Ser Gly Arg Tyr
355 360 365
Leu Ala Lys Leu
370
<210> 91
<211> 1116
<212> DNA
<213> Artificial Sequence
<220>
<223> N54A- T56L- F58H-C211L-M213T
<400> 91
atgacccaga ataagcgcgt ggttgttctg ggcagtggcg ttattggtct gagctgtgct 60
ttagctttag cccagaaggg ctataaagtg catgtggttg cccgcgatct gccggaagac 120
acagtggccc agacctttgc aagcccttgg gccggtgcag cgtggcttcc gcatatgagc 180
aaagaagctg gtccgcgtca agctaaatgg gaggaagcaa ccttcaaaca gtgggtggac 240
ttcgttccgc aaggtctggc aatgtggctg aaaggtaccc gccgctttgc agaaaccgaa 300
gccgatctgc tgggccattg gtataaggac atcgtgccga attaccgcca tctgaacccg 360
agcgattgtc cgccgggcgc aattggcgtg acctacgata ctttaagcgt taacgccccg 420
aagttctgtc agtatctgca gcgcgaagca cagaaactgg gcgtgacctt cgaacgccgt 480
ttagttacct ctttagagca gattgcagat ggcgccgatc tgatcgttaa tgcaaccggt 540
ctgggcgcca aaagcatcgc cggtgtggaa gatcaagaag tggaacctat ccgcggccaa 600
acagtgctga ttaagagcaa ctgtaaacgc ctgaccactg acagtagtga cccgaaaagc 660
ccggcctaca ttattccgcg cccgggtggt gaagtgatct gtggtggtac ctatttagtg 720
ggtaactatg atttaagcgt ggatccggcc accattccgc gcattttaaa acattgtctg 780
cgtttagacc cgagtattag caccgacggc acactggaag gcatcgaaat tttacgccat 840
aacgttggtc tgcgtcccgc tcgtcgtggt ggtccgcgtg tggaattaga acgcgtgagc 900
ttcccgctga agcgcggtca gagtctgtta gctttaggca ccgccaaagc agcagaaggt 960
aaagcaagcc gtaccgtgcc ggttgttcat gcctatggct tcagcagcgc cggttaccaa 1020
caaggttggg gcgcagcttt agaagttcgt gatctggtgg accaagctat tggtagcagc 1080
agtagtgcca gtagtggccg ttatttagcc aaactg 1116
<210> 92
<211> 368
<212> PRT
<213> Artificial Sequence
<220>
<223> rtDAAO(N54V-F58Q-M213S)
<400> 92
Met His Ser Gln Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Ser Ala Leu Ile Leu Ala Arg Lys Gly Tyr Ser Val His Ile
20 25 30
Leu Ala Arg Asp Leu Pro Glu Asp Val Ser Ser Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Val Trp Thr Pro Gln Met Thr Leu Thr Asp Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ser Thr Phe Lys Lys Trp Val Glu
65 70 75 80
Leu Val Pro Thr Gly His Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Gln Asn Glu Asp Gly Leu Leu Gly His Trp Tyr Lys Asp Ile Thr
100 105 110
Pro Asn Tyr Arg Pro Leu Pro Ser Ser Glu Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val His Ala Pro Lys Tyr Cys Gln
130 135 140
Tyr Leu Ala Arg Glu Leu Gln Lys Leu Gly Ala Thr Phe Glu Arg Arg
145 150 155 160
Thr Val Thr Ser Leu Glu Gln Ala Phe Asp Gly Ala Asp Leu Val Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Ile Asp Asp Gln
180 185 190
Ala Ala Glu Pro Ile Arg Gly Gln Thr Val Leu Val Lys Ser Pro Cys
195 200 205
Lys Arg Cys Thr Ser Asp Ser Ser Asp Pro Ala Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Gly Val
225 230 235 240
Gly Asp Trp Asp Leu Ser Val Asn Pro Glu Thr Val Gln Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Thr Ile Ser Ser Asp Gly Thr Ile
260 265 270
Glu Gly Ile Glu Val Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Ala Glu Arg Ile Val Leu Pro Leu Asp
290 295 300
Arg Thr Lys Ser Pro Leu Ser Leu Gly Arg Gly Ser Ala Arg Ala Ala
305 310 315 320
Lys Glu Lys Glu Val Thr Leu Val His Ala Tyr Gly Phe Ser Ser Ala
325 330 335
Gly Tyr Gln Gln Ser Trp Gly Ala Ala Glu Asp Val Ala Gln Leu Val
340 345 350
Asp Glu Ala Phe Gln Arg Tyr His Gly Ala Ala Arg Glu Ser Lys Leu
355 360 365
<210> 93
<211> 1104
<212> DNA
<213> Artificial Sequence
<220>
<223> rtDAAO(N54V-F58Q-M213S)
<400> 93
atgcactctc agaaacgtgt tgttgttctg ggttctggtg ttatcggtct gtcttctgct 60
ctgatcctgg ctcgtaaagg ttactctgtt cacatcctgg ctcgtgacct gccggaagac 120
gtttcttctc agaccttcgc ttctccgtgg gctggtgctg tttggacccc gcagatgacc 180
ctgaccgacg gtccgcgtca ggctaaatgg gaagaatcta ccttcaaaaa atgggttgaa 240
ctggttccga ccggtcacgc tatgtggctg aaaggtaccc gtcgtttcgc tcagaacgaa 300
gacggtctgc tgggtcactg gtacaaagac atcaccccga actaccgtcc gctgccgtct 360
tctgaatgcc cgccgggtgc tatcggtgtt acctacgaca ccctgtctgt tcacgctccg 420
aaatactgcc agtacctggc tcgtgaactg cagaaactgg gtgctacctt cgaacgtcgt 480
accgttacct ctctggaaca ggctttcgac ggtgctgacc tggttgttaa cgctaccggt 540
ctgggtgcta aatctatcgc tggtatcgac gaccaggctg ctgaaccgat ccgtggtcag 600
accgttctgg ttaaatctcc gtgcaaacgt tgcacctctg actcttctga cccggcttct 660
ccggcttaca tcatcccgcg tccgggtggt gaagttatct gcggtggtac ctacggtgtt 720
ggtgactggg acctgtctgt taacccggaa accgttcagc gtatcctgaa acactgcctg 780
cgtctggacc cgaccatctc ttctgacggt accatcgaag gtatcgaagt tctgcgtcac 840
aacgttggtc tgcgtccggc tcgtcgtggt ggtccgcgtg ttgaagctga acgtatcgtt 900
ctgccgctgg accgtaccaa atctccgctg tctctgggtc gtggttctgc tcgtgctgct 960
aaagaaaaag aagttaccct ggttcacgct tacggtttct cttctgctgg ttaccagcag 1020
tcttggggtg ctgctgaaga cgttgctcag ctggttgacg aagctttcca gcgttaccac 1080
ggtgctgctc gtgaatctaa actg 1104
<210> 94
<211> 368
<212> PRT
<213> Artificial Sequence
<220>
<223> rtDAAO(N54A-F58H)
<400> 94
Met His Ser Gln Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Ser Ala Leu Ile Leu Ala Arg Lys Gly Tyr Ser Val His Ile
20 25 30
Leu Ala Arg Asp Leu Pro Glu Asp Val Ser Ser Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Ala Trp Thr Pro His Met Thr Leu Thr Asp Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ser Thr Phe Lys Lys Trp Val Glu
65 70 75 80
Leu Val Pro Thr Gly His Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Gln Asn Glu Asp Gly Leu Leu Gly His Trp Tyr Lys Asp Ile Thr
100 105 110
Pro Asn Tyr Arg Pro Leu Pro Ser Ser Glu Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val His Ala Pro Lys Tyr Cys Gln
130 135 140
Tyr Leu Ala Arg Glu Leu Gln Lys Leu Gly Ala Thr Phe Glu Arg Arg
145 150 155 160
Thr Val Thr Ser Leu Glu Gln Ala Phe Asp Gly Ala Asp Leu Val Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Ile Asp Asp Gln
180 185 190
Ala Ala Glu Pro Ile Arg Gly Gln Thr Val Leu Val Lys Ser Pro Cys
195 200 205
Lys Arg Cys Thr Met Asp Ser Ser Asp Pro Ala Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Gly Val
225 230 235 240
Gly Asp Trp Asp Leu Ser Val Asn Pro Glu Thr Val Gln Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Thr Ile Ser Ser Asp Gly Thr Ile
260 265 270
Glu Gly Ile Glu Val Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Ala Glu Arg Ile Val Leu Pro Leu Asp
290 295 300
Arg Thr Lys Ser Pro Leu Ser Leu Gly Arg Gly Ser Ala Arg Ala Ala
305 310 315 320
Lys Glu Lys Glu Val Thr Leu Val His Ala Tyr Gly Phe Ser Ser Ala
325 330 335
Gly Tyr Gln Gln Ser Trp Gly Ala Ala Glu Asp Val Ala Gln Leu Val
340 345 350
Asp Glu Ala Phe Gln Arg Tyr His Gly Ala Ala Arg Glu Ser Lys Leu
355 360 365
<210> 95
<211> 1104
<212> DNA
<213> Artificial Sequence
<220>
<223> rtDAAO(N54A-F58H)
<400> 95
atgcactctc agaaacgtgt tgttgttctg ggttctggtg ttatcggtct gtcttctgct 60
ctgatcctgg ctcgtaaagg ttactctgtt cacatcctgg ctcgtgacct gccggaagac 120
gtttcttctc agaccttcgc ttctccgtgg gctggtgctg cttggacccc gcacatgacc 180
ctgaccgacg gtccgcgtca ggctaaatgg gaagaatcta ccttcaaaaa atgggttgaa 240
ctggttccga ccggtcacgc tatgtggctg aaaggtaccc gtcgtttcgc tcagaacgaa 300
gacggtctgc tgggtcactg gtacaaagac atcaccccga actaccgtcc gctgccgtct 360
tctgaatgcc cgccgggtgc tatcggtgtt acctacgaca ccctgtctgt tcacgctccg 420
aaatactgcc agtacctggc tcgtgaactg cagaaactgg gtgctacctt cgaacgtcgt 480
accgttacct ctctggaaca ggctttcgac ggtgctgacc tggttgttaa cgctaccggt 540
ctgggtgcta aatctatcgc tggtatcgac gaccaggctg ctgaaccgat ccgtggtcag 600
accgttctgg ttaaatctcc gtgcaaacgt tgcaccatgg actcttctga cccggcttct 660
ccggcttaca tcatcccgcg tccgggtggt gaagttatct gcggtggtac ctacggtgtt 720
ggtgactggg acctgtctgt taacccggaa accgttcagc gtatcctgaa acactgcctg 780
cgtctggacc cgaccatctc ttctgacggt accatcgaag gtatcgaagt tctgcgtcac 840
aacgttggtc tgcgtccggc tcgtcgtggt ggtccgcgtg ttgaagctga acgtatcgtt 900
ctgccgctgg accgtaccaa atctccgctg tctctgggtc gtggttctgc tcgtgctgct 960
aaagaaaaag aagttaccct ggttcacgct tacggtttct cttctgctgg ttaccagcag 1020
tcttggggtg ctgctgaaga cgttgctcag ctggttgacg aagctttcca gcgttaccac 1080
ggtgctgctc gtgaatctaa actg 1104
<210> 96
<211> 368
<212> PRT
<213> Artificial Sequence
<220>
<223> rtDAAO(N54A-56N-F58H)
<400> 96
Met His Ser Gln Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Ser Ala Leu Ile Leu Ala Arg Lys Gly Tyr Ser Val His Ile
20 25 30
Leu Ala Arg Asp Leu Pro Glu Asp Val Ser Ser Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Ala Trp Asn Pro His Met Thr Leu Thr Asp Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ser Thr Phe Lys Lys Trp Val Glu
65 70 75 80
Leu Val Pro Thr Gly His Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Gln Asn Glu Asp Gly Leu Leu Gly His Trp Tyr Lys Asp Ile Thr
100 105 110
Pro Asn Tyr Arg Pro Leu Pro Ser Ser Glu Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val His Ala Pro Lys Tyr Cys Gln
130 135 140
Tyr Leu Ala Arg Glu Leu Gln Lys Leu Gly Ala Thr Phe Glu Arg Arg
145 150 155 160
Thr Val Thr Ser Leu Glu Gln Ala Phe Asp Gly Ala Asp Leu Val Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Ile Asp Asp Gln
180 185 190
Ala Ala Glu Pro Ile Arg Gly Gln Thr Val Leu Val Lys Ser Pro Cys
195 200 205
Lys Arg Cys Thr Met Asp Ser Ser Asp Pro Ala Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Gly Val
225 230 235 240
Gly Asp Trp Asp Leu Ser Val Asn Pro Glu Thr Val Gln Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Thr Ile Ser Ser Asp Gly Thr Ile
260 265 270
Glu Gly Ile Glu Val Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Ala Glu Arg Ile Val Leu Pro Leu Asp
290 295 300
Arg Thr Lys Ser Pro Leu Ser Leu Gly Arg Gly Ser Ala Arg Ala Ala
305 310 315 320
Lys Glu Lys Glu Val Thr Leu Val His Ala Tyr Gly Phe Ser Ser Ala
325 330 335
Gly Tyr Gln Gln Ser Trp Gly Ala Ala Glu Asp Val Ala Gln Leu Val
340 345 350
Asp Glu Ala Phe Gln Arg Tyr His Gly Ala Ala Arg Glu Ser Lys Leu
355 360 365
<210> 97
<211> 1104
<212> DNA
<213> Artificial Sequence
<220>
<223> rtDAAO(N54A-56N-F58H)
<400> 97
atgcactctc agaaacgtgt tgttgttctg ggttctggtg ttatcggtct gtcttctgct 60
ctgatcctgg ctcgtaaagg ttactctgtt cacatcctgg ctcgtgacct gccggaagac 120
gtttcttctc agaccttcgc ttctccgtgg gctggtgctg cttggaaccc gcacatgacc 180
ctgaccgacg gtccgcgtca ggctaaatgg gaagaatcta ccttcaaaaa atgggttgaa 240
ctggttccga ccggtcacgc tatgtggctg aaaggtaccc gtcgtttcgc tcagaacgaa 300
gacggtctgc tgggtcactg gtacaaagac atcaccccga actaccgtcc gctgccgtct 360
tctgaatgcc cgccgggtgc tatcggtgtt acctacgaca ccctgtctgt tcacgctccg 420
aaatactgcc agtacctggc tcgtgaactg cagaaactgg gtgctacctt cgaacgtcgt 480
accgttacct ctctggaaca ggctttcgac ggtgctgacc tggttgttaa cgctaccggt 540
ctgggtgcta aatctatcgc tggtatcgac gaccaggctg ctgaaccgat ccgtggtcag 600
accgttctgg ttaaatctcc gtgcaaacgt tgcaccatgg actcttctga cccggcttct 660
ccggcttaca tcatcccgcg tccgggtggt gaagttatct gcggtggtac ctacggtgtt 720
ggtgactggg acctgtctgt taacccggaa accgttcagc gtatcctgaa acactgcctg 780
cgtctggacc cgaccatctc ttctgacggt accatcgaag gtatcgaagt tctgcgtcac 840
aacgttggtc tgcgtccggc tcgtcgtggt ggtccgcgtg ttgaagctga acgtatcgtt 900
ctgccgctgg accgtaccaa atctccgctg tctctgggtc gtggttctgc tcgtgctgct 960
aaagaaaaag aagttaccct ggttcacgct tacggtttct cttctgctgg ttaccagcag 1020
tcttggggtg ctgctgaaga cgttgctcag ctggttgacg aagctttcca gcgttaccac 1080
ggtgctgctc gtgaatctaa actg 1104
<210> 98
<211> 368
<212> PRT
<213> Artificial Sequence
<220>
<223> rtDAAO(C211L-M213T)
<400> 98
Met His Ser Gln Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Ser Ala Leu Ile Leu Ala Arg Lys Gly Tyr Ser Val His Ile
20 25 30
Leu Ala Arg Asp Leu Pro Glu Asp Val Ser Ser Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Asn Trp Thr Pro Phe Met Thr Leu Thr Asp Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ser Thr Phe Lys Lys Trp Val Glu
65 70 75 80
Leu Val Pro Thr Gly His Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Gln Asn Glu Asp Gly Leu Leu Gly His Trp Tyr Lys Asp Ile Thr
100 105 110
Pro Asn Tyr Arg Pro Leu Pro Ser Ser Glu Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val His Ala Pro Lys Tyr Cys Gln
130 135 140
Tyr Leu Ala Arg Glu Leu Gln Lys Leu Gly Ala Thr Phe Glu Arg Arg
145 150 155 160
Thr Val Thr Ser Leu Glu Gln Ala Phe Asp Gly Ala Asp Leu Val Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Ile Asp Asp Gln
180 185 190
Ala Ala Glu Pro Ile Arg Gly Gln Thr Val Leu Val Lys Ser Pro Cys
195 200 205
Lys Arg Leu Thr Thr Asp Ser Ser Asp Pro Ala Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Gly Val
225 230 235 240
Gly Asp Trp Asp Leu Ser Val Asn Pro Glu Thr Val Gln Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Thr Ile Ser Ser Asp Gly Thr Ile
260 265 270
Glu Gly Ile Glu Val Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Ala Glu Arg Ile Val Leu Pro Leu Asp
290 295 300
Arg Thr Lys Ser Pro Leu Ser Leu Gly Arg Gly Ser Ala Arg Ala Ala
305 310 315 320
Lys Glu Lys Glu Val Thr Leu Val His Ala Tyr Gly Phe Ser Ser Ala
325 330 335
Gly Tyr Gln Gln Ser Trp Gly Ala Ala Glu Asp Val Ala Gln Leu Val
340 345 350
Asp Glu Ala Phe Gln Arg Tyr His Gly Ala Ala Arg Glu Ser Lys Leu
355 360 365
<210> 99
<211> 1104
<212> DNA
<213> Artificial Sequence
<220>
<223> rtDAAO(C211L-M213T)
<400> 99
atgcactctc agaaacgtgt tgttgttctg ggttctggtg ttatcggtct gtcttctgct 60
ctgatcctgg ctcgtaaagg ttactctgtt cacatcctgg ctcgtgacct gccggaagac 120
gtttcttctc agaccttcgc ttctccgtgg gctggtgcta actggacccc gttcatgacc 180
ctgaccgacg gtccgcgtca ggctaaatgg gaagaatcta ccttcaaaaa atgggttgaa 240
ctggttccga ccggtcacgc tatgtggctg aaaggtaccc gtcgtttcgc tcagaacgaa 300
gacggtctgc tgggtcactg gtacaaagac atcaccccga actaccgtcc gctgccgtct 360
tctgaatgcc cgccgggtgc tatcggtgtt acctacgaca ccctgtctgt tcacgctccg 420
aaatactgcc agtacctggc tcgtgaactg cagaaactgg gtgctacctt cgaacgtcgt 480
accgttacct ctctggaaca ggctttcgac ggtgctgacc tggttgttaa cgctaccggt 540
ctgggtgcta aatctatcgc tggtatcgac gaccaggctg ctgaaccgat ccgtggtcag 600
accgttctgg ttaaatctcc gtgcaaacgt ctgaccaccg actcttctga cccggcttct 660
ccggcttaca tcatcccgcg tccgggtggt gaagttatct gcggtggtac ctacggtgtt 720
ggtgactggg acctgtctgt taacccggaa accgttcagc gtatcctgaa acactgcctg 780
cgtctggacc cgaccatctc ttctgacggt accatcgaag gtatcgaagt tctgcgtcac 840
aacgttggtc tgcgtccggc tcgtcgtggt ggtccgcgtg ttgaagctga acgtatcgtt 900
ctgccgctgg accgtaccaa atctccgctg tctctgggtc gtggttctgc tcgtgctgct 960
aaagaaaaag aagttaccct ggttcacgct tacggtttct cttctgctgg ttaccagcag 1020
tcttggggtg ctgctgaaga cgttgctcag ctggttgacg aagctttcca gcgttaccac 1080
ggtgctgctc gtgaatctaa actg 1104
<210> 100
<211> 368
<212> PRT
<213> Artificial Sequence
<220>
<223> rtDAAO(N54A-F58H-C211L-M213T)
<400> 100
Met His Ser Gln Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Ser Ala Leu Ile Leu Ala Arg Lys Gly Tyr Ser Val His Ile
20 25 30
Leu Ala Arg Asp Leu Pro Glu Asp Val Ser Ser Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Ala Trp Thr Pro His Met Thr Leu Thr Asp Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ser Thr Phe Lys Lys Trp Val Glu
65 70 75 80
Leu Val Pro Thr Gly His Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Gln Asn Glu Asp Gly Leu Leu Gly His Trp Tyr Lys Asp Ile Thr
100 105 110
Pro Asn Tyr Arg Pro Leu Pro Ser Ser Glu Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val His Ala Pro Lys Tyr Cys Gln
130 135 140
Tyr Leu Ala Arg Glu Leu Gln Lys Leu Gly Ala Thr Phe Glu Arg Arg
145 150 155 160
Thr Val Thr Ser Leu Glu Gln Ala Phe Asp Gly Ala Asp Leu Val Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Ile Asp Asp Gln
180 185 190
Ala Ala Glu Pro Ile Arg Gly Gln Thr Val Leu Val Lys Ser Pro Cys
195 200 205
Lys Arg Leu Thr Thr Asp Ser Ser Asp Pro Ala Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Gly Val
225 230 235 240
Gly Asp Trp Asp Leu Ser Val Asn Pro Glu Thr Val Gln Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Thr Ile Ser Ser Asp Gly Thr Ile
260 265 270
Glu Gly Ile Glu Val Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Ala Glu Arg Ile Val Leu Pro Leu Asp
290 295 300
Arg Thr Lys Ser Pro Leu Ser Leu Gly Arg Gly Ser Ala Arg Ala Ala
305 310 315 320
Lys Glu Lys Glu Val Thr Leu Val His Ala Tyr Gly Phe Ser Ser Ala
325 330 335
Gly Tyr Gln Gln Ser Trp Gly Ala Ala Glu Asp Val Ala Gln Leu Val
340 345 350
Asp Glu Ala Phe Gln Arg Tyr His Gly Ala Ala Arg Glu Ser Lys Leu
355 360 365
<210> 101
<211> 1104
<212> DNA
<213> Artificial Sequence
<220>
<223> rtDAAO(N54A-F58H-C211L-M213T)
<400> 101
atgcactctc agaaacgtgt tgttgttctg ggttctggtg ttatcggtct gtcttctgct 60
ctgatcctgg ctcgtaaagg ttactctgtt cacatcctgg ctcgtgacct gccggaagac 120
gtttcttctc agaccttcgc ttctccgtgg gctggtgctg cttggacccc gcacatgacc 180
ctgaccgacg gtccgcgtca ggctaaatgg gaagaatcta ccttcaaaaa atgggttgaa 240
ctggttccga ccggtcacgc tatgtggctg aaaggtaccc gtcgtttcgc tcagaacgaa 300
gacggtctgc tgggtcactg gtacaaagac atcaccccga actaccgtcc gctgccgtct 360
tctgaatgcc cgccgggtgc tatcggtgtt acctacgaca ccctgtctgt tcacgctccg 420
aaatactgcc agtacctggc tcgtgaactg cagaaactgg gtgctacctt cgaacgtcgt 480
accgttacct ctctggaaca ggctttcgac ggtgctgacc tggttgttaa cgctaccggt 540
ctgggtgcta aatctatcgc tggtatcgac gaccaggctg ctgaaccgat ccgtggtcag 600
accgttctgg ttaaatctcc gtgcaaacgt ctgaccaccg actcttctga cccggcttct 660
ccggcttaca tcatcccgcg tccgggtggt gaagttatct gcggtggtac ctacggtgtt 720
ggtgactggg acctgtctgt taacccggaa accgttcagc gtatcctgaa acactgcctg 780
cgtctggacc cgaccatctc ttctgacggt accatcgaag gtatcgaagt tctgcgtcac 840
aacgttggtc tgcgtccggc tcgtcgtggt ggtccgcgtg ttgaagctga acgtatcgtt 900
ctgccgctgg accgtaccaa atctccgctg tctctgggtc gtggttctgc tcgtgctgct 960
aaagaaaaag aagttaccct ggttcacgct tacggtttct cttctgctgg ttaccagcag 1020
tcttggggtg ctgctgaaga cgttgctcag ctggttgacg aagctttcca gcgttaccac 1080
ggtgctgctc gtgaatctaa actg 1104
<210> 102
<211> 368
<212> PRT
<213> Artificial Sequence
<220>
<223> rtDAAO(N54A-T56N-F58H-C211L-M213T)
<400> 102
Met His Ser Gln Lys Arg Val Val Val Leu Gly Ser Gly Val Ile Gly
1 5 10 15
Leu Ser Ser Ala Leu Ile Leu Ala Arg Lys Gly Tyr Ser Val His Ile
20 25 30
Leu Ala Arg Asp Leu Pro Glu Asp Val Ser Ser Gln Thr Phe Ala Ser
35 40 45
Pro Trp Ala Gly Ala Ala Trp Asn Pro His Met Thr Leu Thr Asp Gly
50 55 60
Pro Arg Gln Ala Lys Trp Glu Glu Ser Thr Phe Lys Lys Trp Val Glu
65 70 75 80
Leu Val Pro Thr Gly His Ala Met Trp Leu Lys Gly Thr Arg Arg Phe
85 90 95
Ala Gln Asn Glu Asp Gly Leu Leu Gly His Trp Tyr Lys Asp Ile Thr
100 105 110
Pro Asn Tyr Arg Pro Leu Pro Ser Ser Glu Cys Pro Pro Gly Ala Ile
115 120 125
Gly Val Thr Tyr Asp Thr Leu Ser Val His Ala Pro Lys Tyr Cys Gln
130 135 140
Tyr Leu Ala Arg Glu Leu Gln Lys Leu Gly Ala Thr Phe Glu Arg Arg
145 150 155 160
Thr Val Thr Ser Leu Glu Gln Ala Phe Asp Gly Ala Asp Leu Val Val
165 170 175
Asn Ala Thr Gly Leu Gly Ala Lys Ser Ile Ala Gly Ile Asp Asp Gln
180 185 190
Ala Ala Glu Pro Ile Arg Gly Gln Thr Val Leu Val Lys Ser Pro Cys
195 200 205
Lys Arg Leu Thr Thr Asp Ser Ser Asp Pro Ala Ser Pro Ala Tyr Ile
210 215 220
Ile Pro Arg Pro Gly Gly Glu Val Ile Cys Gly Gly Thr Tyr Gly Val
225 230 235 240
Gly Asp Trp Asp Leu Ser Val Asn Pro Glu Thr Val Gln Arg Ile Leu
245 250 255
Lys His Cys Leu Arg Leu Asp Pro Thr Ile Ser Ser Asp Gly Thr Ile
260 265 270
Glu Gly Ile Glu Val Leu Arg His Asn Val Gly Leu Arg Pro Ala Arg
275 280 285
Arg Gly Gly Pro Arg Val Glu Ala Glu Arg Ile Val Leu Pro Leu Asp
290 295 300
Arg Thr Lys Ser Pro Leu Ser Leu Gly Arg Gly Ser Ala Arg Ala Ala
305 310 315 320
Lys Glu Lys Glu Val Thr Leu Val His Ala Tyr Gly Phe Ser Ser Ala
325 330 335
Gly Tyr Gln Gln Ser Trp Gly Ala Ala Glu Asp Val Ala Gln Leu Val
340 345 350
Asp Glu Ala Phe Gln Arg Tyr His Gly Ala Ala Arg Glu Ser Lys Leu
355 360 365
<210> 103
<211> 1104
<212> DNA
<213> Artificial Sequence
<220>
<223> rtDAAO(N54A-T56N-F58H-C211L-M213T)
<400> 103
atgcactctc agaaacgtgt tgttgttctg ggttctggtg ttatcggtct gtcttctgct 60
ctgatcctgg ctcgtaaagg ttactctgtt cacatcctgg ctcgtgacct gccggaagac 120
gtttcttctc agaccttcgc ttctccgtgg gctggtgctg cttggaaccc gcacatgacc 180
ctgaccgacg gtccgcgtca ggctaaatgg gaagaatcta ccttcaaaaa atgggttgaa 240
ctggttccga ccggtcacgc tatgtggctg aaaggtaccc gtcgtttcgc tcagaacgaa 300
gacggtctgc tgggtcactg gtacaaagac atcaccccga actaccgtcc gctgccgtct 360
tctgaatgcc cgccgggtgc tatcggtgtt acctacgaca ccctgtctgt tcacgctccg 420
aaatactgcc agtacctggc tcgtgaactg cagaaactgg gtgctacctt cgaacgtcgt 480
accgttacct ctctggaaca ggctttcgac ggtgctgacc tggttgttaa cgctaccggt 540
ctgggtgcta aatctatcgc tggtatcgac gaccaggctg ctgaaccgat ccgtggtcag 600
accgttctgg ttaaatctcc gtgcaaacgt ctgaccaccg actcttctga cccggcttct 660
ccggcttaca tcatcccgcg tccgggtggt gaagttatct gcggtggtac ctacggtgtt 720
ggtgactggg acctgtctgt taacccggaa accgttcagc gtatcctgaa acactgcctg 780
cgtctggacc cgaccatctc ttctgacggt accatcgaag gtatcgaagt tctgcgtcac 840
aacgttggtc tgcgtccggc tcgtcgtggt ggtccgcgtg ttgaagctga acgtatcgtt 900
ctgccgctgg accgtaccaa atctccgctg tctctgggtc gtggttctgc tcgtgctgct 960
aaagaaaaag aagttaccct ggttcacgct tacggtttct cttctgctgg ttaccagcag 1020
tcttggggtg ctgctgaaga cgttgctcag ctggttgacg aagctttcca gcgttaccac 1080
ggtgctgctc gtgaatctaa actg 1104
<210> 104
<211> 39
<212> DNA
<213> Artificial Sequence
<220>
<223> 211-213正向引物
<220>
<221> misc_feature
<222> (16)..(17)
<223> n is a, c, g, or t
<220>
<221> misc_feature
<222> (18)
<223> k is g or t
<220>
<221> misc_feature
<222> (22)..(23)
<223> n is a, c, g, or t
<220>
<221> misc_feature
<222> (24)
<223> k is g or t
<400> 104
agcaactgta aacgcnnkac cnnkgacagt agtgacccg 39
<210> 105
<211> 39
<212> DNA
<213> Artificial Sequence
<220>
<223> 211-213反向引物
<220>
<221> misc_feature
<222> (16)
<223> m is a or c
<220>
<221> misc_feature
<222> (17)..(18)
<223> n is a, c, g, or t
<220>
<221> misc_feature
<222> (22)
<223> m is a or c
<220>
<221> misc_feature
<222> (23)..(24)
<223> n is a, c, g, or t
<400> 105
cgggtcacta ctgtcmnngg tmnngcgttt acagttgct 39
<210> 106
<211> 45
<212> DNA
<213> Artificial Sequence
<220>
<223> 54-56-58正向引物
<220>
<221> misc_feature
<222> (16)..(17)
<223> n is a, c, g, or t
<220>
<221> misc_feature
<222> (18)
<223> k is g or t
<220>
<221> misc_feature
<222> (22)..(23)
<223> n is a, c, g, or t
<220>
<221> misc_feature
<222> (24)
<223> k is g or t
<220>
<221> misc_feature
<222> (28)..(29)
<223> n is a, c, g, or t
<220>
<221> misc_feature
<222> (30)
<223> k is g or t
<400> 106
cccttgggcc ggtgcnnktt gnnkcccnnk gatgagcaaa gaagc 45
<210> 107
<211> 45
<212> DNA
<213> Artificial Sequence
<220>
<223> 54-56-58反向引物
<220>
<221> misc_feature
<222> (16)
<223> m is a or c
<220>
<221> misc_feature
<222> (17)..(18)
<223> n is a, c, g, or t
<220>
<221> misc_feature
<222> (22)
<223> m is a or c
<220>
<221> misc_feature
<222> (23)..(24)
<223> n is a, c, g, or t
<220>
<221> misc_feature
<222> (28)
<223> m is a or c
<220>
<221> misc_feature
<222> (29)..(30)
<223> n is a, c, g, or t
<400> 107
gcttctttgc tcatcmnngg gmnncaamnn gcaccggccc aaggg 45

Claims (6)

1.一种D-氨基酸氧化酶突变体，其特征在于，所述D-氨基酸氧化酶突变体的氨基酸序列如SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ ID NO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ ID NO:23、SEQ ID NO:25、SEQ IDNO:27、SEQ ID NO:29、SEQ ID NO:31、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、 SEQID NO:39、SEQ ID NO:41、SEQ ID NO:43、SEQ ID NO:45、SEQ ID NO:47、SEQ ID NO:49、SEQID NO:51、SEQ ID NO:53、SEQ ID NO:55、SEQ ID NO:57、SEQ ID NO:59、SEQ ID NO:61、SEQID NO:63、SEQ ID NO:65、SEQ ID NO:70、SEQ ID NO:72、SEQ ID NO:75、SEQ ID NO:79、SEQID NO:82、SEQ ID NO:90、SEQ ID NO:100，或SEQ ID NO:102所示。
2.如权利要求1所述的D-氨基酸氧化酶，其特征在于，编码所述D-氨基酸氧化酶突变体的核苷酸序列如SEQ ID NO:6、SEQ ID NO:8、SEQ ID NO:10、SEQ ID NO:12、SEQ ID NO:14、SEQ ID NO:16、SEQ ID NO:18、SEQ ID NO:20、SEQ ID NO:22、SEQ ID NO:24、SEQ ID NO:26、SEQ ID NO:28、SEQ ID NO:30、SEQ ID NO:32、SEQ ID NO:34、SEQ ID NO:36、SEQ IDNO:38、SEQ ID NO:40、SEQ ID NO:42、SEQ ID NO:44、SEQ ID NO:46、SEQ ID NO:48、SEQ IDNO:50、SEQ ID NO:52、SEQ ID NO:54、SEQ ID NO:56、SEQ ID NO:58、SEQ ID NO:60、SEQ IDNO:62、SEQ ID NO:64、SEQ ID NO:66、SEQ ID NO:67、SEQ ID NO:71、SEQ ID NO:73、SEQ IDNO:74、SEQ ID NO:76、SEQ ID NO:77、SEQ ID NO:78、SEQ ID NO:80、SEQ ID NO:81、SEQ IDNO:83、SEQ ID NO:84、SEQ ID NO:85、SEQ ID NO:86、SEQ ID NO:87、SEQ ID NO:88、SEQ IDNO:89、SEQ ID NO:91、SEQ ID NO:101，或SEQ ID NO:103所示。
3.一种分离的核酸，其特征在于，所述核酸编码如权利要求1或2所述的D-氨基酸氧化酶突变体。
4.一种包含如权利要求3所述的核酸的重组表达载体。
5.一种包含如权利要求3所述的核酸或如权利要求4所述的重组表达载体的转化体。
6.一种如权利要求1或2所述的D-氨基酸氧化酶突变体在制备2-氧代-4-(羟基甲基氧膦基)丁酸中的应用。
CN201811408085.1A 2018-11-23 2018-11-23 一种d-氨基酸氧化酶突变体及其应用 Active CN111019916B (zh)

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PCT/CN2019/120249 WO2020103928A1 (zh) 2018-11-23 2019-11-22 一种d-氨基酸氧化酶突变体及其应用
EP19887398.6A EP3885435A4 (en) 2018-11-23 2019-11-22 OXIDATIVE ENZYM MUTANT OF D-AMINO ACID AND USE THEREOF
US17/295,676 US11667897B2 (en) 2018-11-23 2019-11-22 D-amino acid oxidative enzyme mutant and application thereof

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