CN111556873A - N-乙酰神经氨酸的发酵生产 - Google Patents

N-乙酰神经氨酸的发酵生产 Download PDF

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CN111556873A
CN111556873A CN201880081537.1A CN201880081537A CN111556873A CN 111556873 A CN111556873 A CN 111556873A CN 201880081537 A CN201880081537 A CN 201880081537A CN 111556873 A CN111556873 A CN 111556873A
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S·詹尼温
D·瓦滕伯格
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Abstract

本申请公开了用于生产N‑乙酰神经氨酸的非天然存在的微生物、用于通过非天然存在的微生物的发酵生产N‑乙酰神经氨酸的方法以及包含已通过非天然存在的微生物的发酵生产的N‑乙酰神经氨酸的营养组合物。

Description

N-乙酰神经氨酸的发酵生产
本发明涉及能够生产N-乙酰神经氨酸的非天然存在的微生物、用于使用所述非天然存在的微生物的发酵生产N-乙酰神经氨酸的方法、通过发酵生产的N-乙酰神经氨酸的用途以及含有以这种方式生产的N-乙酰神经氨酸的产物。
背景技术
唾液酸(Sia)是具有九碳骨架的荷负电的单糖家族。在自然界中已经发现了50多种形式的α-酮酸。最丰富的唾液酸似乎是N-乙酰神经氨酸(NANA、NeuNAc、Neu5Ac)。
唾液酸是作为存在于脊椎动物和高等无脊椎动物的细胞表面上糖缀合物(糖蛋白和糖脂)中的聚糖的末端糖存在。唾液酸是致病细菌的脂多糖和荚膜多糖的成分,其包括大肠杆菌(Escherichia coli)K1、流感嗜血杆菌(Haemophilus influenzae)、杜克雷嗜血杆菌(Haemophilus ducreyi)、多杀性巴氏杆菌(Pateurella multocida)、淋病奈瑟氏菌(Neisseria gonorrhoeae)、脑膜炎奈瑟氏菌(Neisseria meningitidis)、空肠弯曲杆菌(Campylobacter jejuni)和无乳链球菌(Streptococcus agalactiae)。
唾液酸在许多生理和病理生理过程中起重要作用,包括胚胎神经系统的发育、转移、免疫应答的调节和细菌或病毒的感染。唾液酸是脑神经节苷脂和多唾液酸链(其修饰神经细胞粘附分子(NCAM))的基本成分,所述NCAM促进细胞间相互作用、神经元向外生长、突触连接的修饰和记忆形成。在仔猪中,富含唾液酸的饮食增加了脑唾液酸的水平和两种学习相关基因的表达。因此,饮食也能增强学习和记忆。
婴儿,特别是早产儿,由于在这个发育阶段的脑快速生长及其免疫系统的发育而对包括唾液酸在内的营养物质有很高的需求。唾液酸(特别是N-乙酰神经氨酸)的水平在人类母乳中很高(大约0.5g·L-1)。相比之下,婴儿配方物含有较低或甚至微不足道的N-乙酰神经氨酸。
因此,有必要提供足够质量和数量的唾液酸(特别是Neu5Ac),其质量和数量足以补充婴儿配方物和其他营养组合物。在这方面,过去已经公开了多种方法。
文献EP1484406A1记载了用于使用微生物生产N-乙酰神经氨酸的方法,与野生型菌株相比,该微生物具有产生Neu5Ac的能力,但具有有限的或没有分解Neu5Ac的能力,使得Neu5Ac积累在培养基中并可从培养基中回收。为了能够生产Neu5Ac,该微生物具有较强的N-乙酰神经氨酸合酶活性和/或N-乙酰葡糖胺2-差向异构酶活性。更具体地,对大肠杆菌细胞进行随机诱变,将细胞系(该细胞系在含葡萄糖的培养基上有利地生长,但在含N-乙酰神经氨酸的培养基上生长受限或不生长)用编码N-乙酰神经氨酸合酶和N-乙酰葡糖胺2-差向异构酶的表达质粒转化。经过一段时间的培养,通过离心沉淀细胞,将其以所谓“湿细胞”形式储存在-20℃下,解冻后根据需要使用。为生产N-乙酰神经氨酸,提供了一种反应混合物(30mL),包括90g·L-1N-乙酰葡糖胺、50g·L-1葡萄糖、10mL·L-1二甲苯和200g·L-1的所述湿细胞(在存在4g·L-1洗涤剂的情况下被透化)。体外反应完成后,通过HPLC评估Neu5Ac的形成。
文献WO94/29476A1公开了一种从N-乙酰-D-葡糖胺(NAG,GlcNAc)制备N-乙酰-D-神经氨酸的体外方法。在制备中,NAG通过碱催化的差向异构化作用转化为N-乙酰-D-甘露糖胺(NAM,ManNAc)。随后,NAM与丙酮酸在Neu5Ac-醛缩酶催化下反应生成Neu5Ac。从表达所述Neu5Ac-醛缩酶的重组大肠杆菌细胞中制备Neu5Ac-醛缩酶。通过将Eupergit-
Figure BDA0002543398380000021
小珠与所述重组大肠杆菌细胞粗提物混合,来固定醛缩酶。通过将所述固定化酶小珠加入到NAM和丙酮酸的混合物中,开始NAM向Neu5Ac的转化。在反应结束时,从反应混合物中分离出Neu5Ac。
在一种前述方法的替代方法中,EP0578825A1公开了一种用于通过在碱性条件下用N-乙酰神经氨酸裂解酶处理N-乙酰葡糖胺和丙酮酸的混合物来生产N-乙酰神经氨酸的体外方法。
US专利No.US 7,579,175公开了一种用于利用经透化的微生物生产N-乙酰神经氨酸的方法。该方法包括制备混合物,其含有:(i)具有N-乙酰神经氨酸醛缩酶活性或N-乙酰神经氨酸合成酶活性的微生物的培养物,或所述培养物的经处理的物质,(ii)能够产生丙酮酸的微生物的培养物或所述培养物的经处理的物质,或能够产生磷酸烯醇丙酮酸的微生物的培养物或所述培养物的经处理的物质,(iii)N-乙酰甘露糖胺,和(iv)形成丙酮酸或磷酸烯醇丙酮酸所必需的能源。该混合物是在水性介质中制备的,该水性介质包括螯合剂或表面活性剂,允许所述水性介质中形成和积累N-乙酰神经氨酸,然后从所述水性介质中回收N-乙酰神经氨酸。
上述方法的缺点是:(i)只能进行小规模生产;和(ii)需要过量的丙酮酸才能使反应平衡朝向Neu5Ac。此外,N-乙酰葡糖胺、N-乙酰甘露糖胺和磷酸烯醇丙酮酸是这些反应的昂贵底物。
国际公布WO2008/040717A2公开了一种用于生产唾液酸的方法,包括在培养基中培养微生物,其中所述微生物携带编码唾液酸合酶(NeuB)和UDP-GlcNAc差向异构酶(NeuC)的异源基因,其中所述微生物缺乏编码CMP-Neu5Ac合酶(NeuA)的基因,或其中任何编码CMP-Neu5Ac合酶(NeuA)的基因已被失活或缺失,以及其中编码唾液酸醛缩酶(NanA)、唾液酸转运蛋白(NanT)和任选地ManNAc激酶(NanK)的内源性基因已被缺失或失活。通过用冰醋酸进行沉淀,从培养物的上清液(2升)中纯化Neu5Ac。
国际公布WO2008/097366A2涉及产生唾液酸的代谢工程化大肠杆菌细胞。在所述细胞中,nanT(唾液酸转运蛋白)和nanA(唾液酸醛缩酶)基因被失活,并在所述nanT-nanA-大肠杆菌细胞中使用表达质粒引入和过表达促进B群脑膜炎奈瑟氏菌中唾液酸生物合成的neuC和neuB基因。此外,大肠杆菌葡糖胺合酶基因(glmS)与neuB和neuC共同过表达。
国际公布WO2012/083329A1公开了在木霉属(Trichoderma)真菌细胞中产生Neu5Ac的方法和试剂,所述细胞组成型表达N-乙酰葡糖胺2-差向异构酶和N-乙酰神经氨酸合酶。在GlcNAc存在下培养这种木霉属细胞,并用HPLC-MS分析菌丝中是否存在Neu5Ac。
中国专利申请CN106929461A公开了一种利用枯草芽孢杆菌(Bacillus subtilis)细胞生产N-乙酰神经氨酸的方法,该细胞表达编码葡糖胺-果糖-6-磷酸转氨酶、葡糖胺-6-磷酸N-乙酰转移酶、N-乙酰葡糖胺异构酶和N-乙酰神经氨酸合酶的基因。这些细胞还已缺失ptsG基因,该基因编码磷酸转移酶系统EIICBA的葡萄糖特异性成分。通过在含葡萄糖培养基中培养这些细胞,得到0.66g·L-1Neu5Ac的产量。
Zhu,D.及其同事(Zhu,D.et al.(2017)Biotechnol.Lett.39:227-234)报道了使用高拷贝数共表达载体在大肠杆菌中过表达PEP合成相关基因pck和ppsA,增加Neu5Ac的产生。
因此,一个目标是提供能够在工业规模上更有效地生产唾液酸,并使用廉价的碳源作为唯一碳源的微生物。
该目标是通过以下方法实现:提供一种非天然存在的微生物,其携带含有至少一种异源酶的唾液酸合成途径,其天然存在的唾液酸分解代谢途径失效,用于Neu5Ac生物合成的磷酸烯醇丙酮酸的可用性方面得到改善,并且能够利用发酵液中存在的单一廉价外源碳源,而不使用磷酸烯醇丙酮酸:磷酸转移酶系统来获得所述外源碳源。
发明内容
在第一方面,提供了一种用于生产Neu5Ac的非天然存在的微生物,其中所述非天然存在的微生物具有包括至少一种异源酶的唾液酸合成途径,其中天然存在的唾液酸分解代谢途径已被失效,其中至少一种用于输入在Neu5Ac发酵生产过程中未用作碳源的糖的磷酸转移酶系统已被失效,其中所述非天然存在的微生物可以利用发酵液中存在的外源碳源,而不使用磷酸转移酶系统来获取所述外源碳源。
在第二方面,提供了根据第一方面的非天然存在的微生物用于生产Neu5Ac的用途。
在第三方面,提供了一种用于使用根据第一方面的非天然存在的微生物的发酵生产Neu5Ac的方法。
在第四方面,提供了通过根据第二方面的方法生产的Neu5Ac。
在第五方面,提供了根据第四方面的Neu5Ac用于制备营养组合物用途。
在第六方面,提供了一种包含通过第三方面的方法生产的Neu5Ac的营养组合物。
附图说明
图1示出了说明生产Neu5Ac的代谢途径的示意图。
图2示出了说明生产Neu5Ac的额外和/或替代的代谢途径的示意图。
图3显示了一个柱形图,说明不同的非天然存在的大肠杆菌菌株的Neu5Ac产生水平。
图4显示了一个柱形图,说明了不同的非天然存在的大肠杆菌菌株的Neu5Ac产生水平。
具体实施方式
根据第一方面,提供了一种非天然存在的微生物,其能够产生Neu5Ac。所述非天然存在的微生物具有包括至少一种异源酶的唾液酸生物合成途径,所述异源酶以足以产生唾液酸的方式从异源核苷酸序列表达。所述微生物天然存在的唾液酸分解代谢途径已被失效。至少一种磷酸烯醇丙酮酸:糖磷酸转移酶系统也已被失效。非天然存在的微生物可以利用外源提供的碳源作为唯一的碳源,而不需要磷酸烯醇丙酮酸:糖磷酸转移酶系统来获得所述碳源。
本文所用的术语“非天然存在的微生物”是指一种微生物,它已被遗传工程化以引入至少一种异源核苷酸序列和/或其中在该微生物中天然存在的核苷酸序列已经被修饰,即改变、替换、插入或缺失。
本文所用的术语“异源”是指天然不具有一种化合物、多肽、蛋白质、酶或核苷酸序列的宿主生物中的所述化合物、多肽、蛋白质、酶、核酸分子或核苷酸序列(作为核酸分子的一部分)。“异源核苷酸序列”可能是一种基因或基因片段。术语“异源表达”是指异源基因或基因片段在天然不具有该基因或基因片段的宿主生物中的表达。异源基因表达导致宿主生物中存在异源多肽、蛋白质或酶。
非天然存在的微生物能够生产Neu5Ac。本文使用的术语“生产”是指通过微生物发酵生产Neu5Ac。“微生物发酵”可以理解为工业过程(通常大规模),其中所需的产物(例如Neu5Ac)是通过在含有营养物质的发酵液中培养微生物而产生的,使得所述微生物就可以将化合物转化为其他化合物。术语“大规模”和“工业”表明可通过发酵液体积超过100L、500L、1000L、5000L、10,000L、50,000L、100,000L或甚至200,000L的微生物发酵进行生产。
本文使用的术语“能够产生(capable of producing)”或“能够产生(able toproduce)”是指微生物产生Neu5Ac的能力,条件是在培养基或培养液中,并在允许微生物合成Neu5Ac的条件下培养。
唾液酸生物合成途径
非天然存在的微生物是用于生产Neu5Ac的微生物。因此,所述非天然存在的微生物能够产生Neu5Ac。非天然存在的微生物是一种已被遗传工程化而具有唾液酸生物合成途径的微生物。
在一个实施方案中,非天然存在的微生物的唾液酸生物合成途径包括至少一种选自以下的异源酶:谷氨酰胺-果糖-6-磷酸转氨酶、葡糖胺-6-磷酸N-乙酰转移酶、N-乙酰葡糖胺2-差向异构酶、N-乙酰神经氨酸合酶和类卤酸脱氢酶(HAD)超家族的糖磷酸酶。优选地,非天然存在的微生物是一种已被遗传工程化以包含一个或多个编码所述酶的基因的微生物。应当理解的是,宿主微生物(其已经携带一个或多个编码所述酶的基因,并以足以产生Neu5Ac的方式表达所述基因)不需要进行遗传工程化来完成唾液酸生物合成途径,但仍可进行遗传工程化以改变一个或多个所述基因的表达水平,以增加谷氨酰胺-果糖-6-磷酸转氨酶、葡糖胺-6-磷酸N-乙酰转移酶、N-乙酰葡糖胺-2-差向异构酶、N-乙酰神经氨酸合酶和/或类HAD超家族的糖磷酸酶的量,从而增加了非天然存在的微生物中Neu5Ac生物合成的速率。
谷氨酰胺-果糖-6-磷酸转氨酶(EC 2.6.1.16)利用谷氨酰胺催化果糖6-磷酸转化为葡糖胺-6-磷酸。这种酶反应通常被认为是己糖胺生物合成途径的第一步。谷氨酰胺-果糖-6-磷酸转氨酶的替代名称为D-果糖-6-磷酸转氨酶、GFAT、葡糖胺-6-磷酸合酶、磷酸己糖转氨酶和L-谷氨酰胺-D-果糖-6-磷酸酰胺转移酶。
在一个额外的和/或替代的实施方案中,非天然存在的微生物具有谷氨酰胺-果糖-6-磷酸转氨酶(GlmS),优选异源谷氨酰胺-果糖-6-磷酸转氨酶,更优选来自大肠杆菌的谷氨酰胺-果糖-6-磷酸转氨酶,或大肠杆菌GlmS的功能性变体。最优选地,功能性变体是一种形式的大肠杆菌GlmS,它同野生型酶一样显示出对葡糖胺-6-磷酸抑制的敏感性显著降低,例如由突变体glmS基因(glmS*54或glmS*)(参见SEQ ID NO:6)编码的。
本文所用的关于酶的术语“功能性变体”是指指定酶的不失去活性的多肽变体,其与指定酶的氨基酸序列有至少70%,优选至少80%,更优选至少90%,甚至更优选至少95%的同一性。这考虑到了衍生出这些多肽的基因组序列数据中存在某些变异的可能性,以及在不显著影响酶的催化活性的情况下,这些多肽中存在的一些氨基酸可以被替换的可能性。
术语“功能性变体”还包括指定酶的多肽变体,其代表不显著丧失催化活性的酶的截短变体。因此,截短变体的氨基酸序列可能不同于指定酶的氨基酸序列,其中缺少一个氨基酸、两个氨基酸或一段两个以上连续氨基酸。截短可以在指定酶的氨基末端(N端)、羧基末端(C端)和/或氨基酸序列内。
在一个额外的和/或替代的实施方案中,非天然存在的微生物包含一种核酸分子,该核酸分子包含编码谷氨酰胺-果糖-6-磷酸转氨酶的核苷酸序列。在一个额外的和/或替代的实施方案中,编码谷氨酰胺-果糖-6-磷酸转氨酶的核苷酸序列是一种异源核苷酸序列。在一个额外的和/或替代的实施方案中,编码谷氨酰胺-果糖-6-磷酸转氨酶的核苷酸序列编码大肠杆菌谷氨酰胺-果糖-6-磷酸转氨酶或其功能性变体。在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化以包含一种核酸分子(其包含编码谷氨酰胺-果糖-6-磷酸转氨酶或其功能性变体的核苷酸序列),和/或以包含谷氨酰胺-果糖-6-磷酸转氨酶或其功能性变体。
大肠杆菌谷氨酰胺-果糖-6-磷酸转氨酶(UniProtKB-P17169;SEQ ID NO:11)是由大肠杆菌glmS基因(SEQ ID NO:10)编码。在一个额外的和/或替代的实施方案中,非天然存在的微生物包含一种核酸分子,该核酸分子包含并表达编码大肠杆菌GlmS或其功能性变体的核苷酸序列,优选编码GlmS*的核苷酸序列(SEQ ID NO:12和SEQ ID NO:13)。
在一个额外的和/或替代的实施方案中,编码大肠杆菌GlmS或大肠杆菌GlmS的功能性变体之一的核苷酸序列与大肠杆菌glmS的序列同一性为至少80%、至少85%、至少90%、至少95%、至少98%或至少99%。
葡糖胺-6-磷酸N-乙酰转移酶(Gna1,EC 2.3.1.4)使用乙酰-CoA将葡糖胺-6-磷酸转化为N-乙酰葡糖胺-6-磷酸。这种酶促反应被认为是酿酒酵母(Saccharomycescerevisiae)中从α-D-葡糖胺6-磷酸合成N-乙酰-α-D-葡糖胺1-磷酸的子通路的第一步。Gna1也被称为磷酸葡糖胺乙酰化酶或磷酸葡糖胺转乙酰酶。
在一个额外的和/或替代的实施方案中,非天然存在的微生物包含葡糖胺-6-磷酸N-乙酰转移酶(Gna1),优选异源葡糖胺-6-磷酸N-乙酰转移酶,更优选来自酿酒酵母的葡糖胺-6-磷酸N-乙酰转移酶(UniProtKB-P43577,SEQ ID NO:15)或酿酒酵母Gna1的功能性变体。
在一个额外的和/或替代的实施方案中,非天然存在的微生物包含一种核酸分子,该核酸分子包含编码葡糖胺-6-磷酸N-乙酰转移酶的核苷酸序列。在一个额外的和/或替代的实施方案中,编码葡糖胺-6-磷酸N-乙酰转移酶的核苷酸序列是一种异源核苷酸序列。在一个额外的和/或替代的实施方案中,编码葡糖胺-6-磷酸N-乙酰转移酶的核苷酸序列编码酿酒酵母葡糖胺-6-磷酸N-乙酰转移酶或其功能性片段。然而,已知葡糖胺-6-磷酸N-乙酰转移酶、其推导出的氨基酸序列和编码这些葡糖胺-6-磷酸N-乙酰转移酶的核苷酸序列来自各种不同的物种,也可用作合适的葡糖胺-6-磷酸N-乙酰转移酶。在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化,以包含一种核酸分子(其包含编码葡糖胺-6-磷酸N-乙酰转移酶或其功能性变体的核苷酸序列),和/或以包含葡糖胺-6-磷酸N-乙酰转移酶或其功能性变体。
酿酒酵母葡糖胺-6-磷酸N-乙酰转移酶(UniProtKB-P43577;SEQ ID NO:15)是由酿酒酵母gna1基因(SEQ ID NO:14)编码。在一个额外的和/或替代的实施方案中,非天然存在的微生物包含核酸分子,该核酸分子包含并表达编码酿酒酵母Gna1或其功能性变体的核苷酸序列,优选编码酿酒酵母Gna1的核苷酸序列(SEQ ID NO:14)。
在一个额外的和/或替代的实施方案中,编码酿酒酵母Gna1或酿酒酵母Gna1的功能性变体之一的核苷酸序列与酿酒酵母gan1的序列同一性为至少80%、至少85%、至少90%、至少95%、至少98%或至少99%。
在一个额外的和/或替代的实施方案中,非天然存在的微生物表达类HAD超家族的糖磷酸酶,它催化N-乙酰葡糖胺-6-磷酸(GlcNAc6P)转化为N-乙酰葡糖胺(GlcNAc)。类HAD超家族是以细菌酶卤酸脱氢酶命名的,包括磷酸酶。类HAD超家族中催化GlcNAc6P转化为GlcNAc的合适的磷酸酶可选自果糖-1-磷酸磷酸酶(YqaB,UniProtKB-P77475)和α-D-葡萄糖1-磷酸磷酸酶(YihX,UniProtKB-P0A8Y3)。大肠杆菌YqaB和大肠杆菌YihX酶也被认为对GlcNAc6P起作用(Lee,S.-W.and Oh,M.-K.(2015)Metabolic Engineering 28:143-150)。在一个实施方案中,类HAD超家族中催化GlcNAc-6-磷酸转化为GlcNAc的糖磷酸酶是非天然存在的微生物中的异源酶。在一个额外的和/或替代的实施方案中,类HAD超家族中催化GlcNAc6P转化为GlcNAc的糖磷酸酶选自大肠杆菌YqaB、大肠杆菌YihX及其功能性变体。
在一个额外的和/或替代的实施方案中,非天然存在的微生物包含一种核酸分子,该核酸分子包含编码类HAD超家族中催化GlcNAc6P转化为GlcNAc的糖磷酸酶的核苷酸序列。在一个额外的和/或替代的实施方案中,编码类HAD超家族中催化GlcNAc6P转化为GlcNAc的糖磷酸酶的核苷酸序列是异源核苷酸序列。在一个额外的和/或替代的实施方案中,编码类HAD超家族中催化GlcNAc6P转化为GlcNAc的糖磷酸酶的核苷酸序列,编码大肠杆菌果糖-1-磷酸磷酸酶或大肠杆菌α-D-葡萄糖-1-磷酸磷酸酶或这两种酶之一的功能性片段。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化以包含一种核酸分子(其包含编码类HAD超家族中催化GlcNAc6P转化为GlcNAc的糖磷酸酶或所述HAD磷酸酶的功能性片段的核苷酸序列),和/或以包含类HAD超家族中催化GlcNAc6P转化为GlcNAc的糖磷酸酶或其功能性变体。
编码类HAD超家族中催化GlcNAc6P转化为GlcNAc的合适的糖磷酸酶的核苷酸序列可选自编码大肠杆菌YqaB、大肠杆菌YihX及其功能性变体的核苷酸序列。
大肠杆菌YqaB(SEQ ID NO:17)和大肠杆菌YihX(SEQ ID NO:19)分别由大肠杆菌基因yqaB(SEQ ID NO:16)和yihX(SEQ ID NO:18)编码。因此,在一个额外的和/或替代的实施方案中,非天然存在的微生物包含一种核酸分子,该核酸分子包含编码大肠杆菌YqaB、大肠杆菌YihX或这两种酶之一的功能性片段的核苷酸序列。
在一个额外的和/或替代的实施方案中,编码大肠杆菌YqaB或其功能性变体的核苷酸序列与大肠杆菌yqaB的序列同一性为至少80%、至少85%、至少90%、至少95%、至少98%或至少99%。
在一个额外的和/或替代的实施方案中,编码大肠杆菌YihX或其功能性变体的核苷酸序列与大肠杆菌yihX的序列同一性为至少80%、至少85%、至少90%、至少95%、至少98%或至少99%。
N-乙酰葡糖胺2-差向异构酶(EC 5.1.3.8)是一种催化N-乙酰葡糖胺(GlcNAc)转化为N-乙酰甘露糖胺(ManNAc)的酶。该酶是作用于碳水化合物及其衍生物的消旋酶。该酶类的系统名称为N-酰基-D-葡糖胺2-差向异构酶。该酶参与氨基-糖代谢和核苷酸-糖代谢。
在一个额外的和/或替代的实施方案中,非天然存在的微生物具有N-乙酰葡糖胺2-差向异构酶,优选异源N-乙酰葡糖胺2-差向异构酶。
在一个额外的和/或替代的实施方案中,N-乙酰葡糖胺2-差向异构酶来源于多变鱼腥蓝细菌(Anabena variabilis)、Acaryochloris sp.、念珠藻属种(Nostoc sp.)、点型念珠蓝细菌(Nostoc punctiforme)、卵形拟杆菌(Bacteroides ovatus)或集胞藻属种(Synechocystis sp.)或是其功能性变体。卵形拟杆菌ATCC 8483的N-乙酰葡糖胺2-差向异构酶(UniProtKB-A7LVG6,SEQ ID NO:21)是由基因BACOVA_01816(SEQ ID NO:20)编码。集胞藻属种(菌株PCC 6803)的N-乙酰葡糖胺2-差向异构酶(UniProtKB-P74124;SEQ ID NO:23)也称为肾素结合蛋白,是由slr1975基因(SEQ ID NO:22)编码。
在一个额外的和/或替代的实施方案中,非天然存在的微生物包含一种核酸分子,该核酸分子包含编码N-乙酰葡糖胺2-差向异构酶或其功能性变体的核苷酸序列。在一个额外的和/或替代的实施方案中,编码N-乙酰葡糖胺2-差向异构酶的核苷酸序列选自编码多变鱼腥蓝细菌、Acaryochloris sp.、念珠藻属种、点型念珠蓝细菌、卵形拟杆菌或集胞藻属种的N-乙酰葡糖胺2-差向异构酶及其功能性变体的核苷酸序列。在一个额外的和/或替代的实施方案中,编码N-乙酰葡糖胺2-差向异构酶的核苷酸序列为异源核苷酸序列。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化以包含一种核酸分子(其包含编码N-乙酰葡糖胺2-差向异构酶或其功能性变体的核苷酸序列),和/或以包含N-乙酰葡糖胺2-差向异构酶或其功能性变体。
在一个额外的和/或替代的实施方案中,编码N-乙酰葡糖胺2-差向异构酶的功能性变体之一的核苷酸序列与集胞藻属种slr1975基因的序列同一性为至少80%、至少85%、至少90%、至少95%、至少98%或至少99%。
在一个额外的和/或替代的实施方案中,非天然存在的微生物具有GlcNAc-6-磷酸差向异构酶活性和ManNAc-6-磷酸磷酸酶活性。
GlcNAc-6磷酸酶差向异构酶将GlcNAc-6-磷酸转化为ManNAc-6-磷酸,而ManNAc-6-磷酸磷酸酶则将ManNAc-6-磷酸去磷酸化以产生ManNAc。具有GlcNAc-6-磷酸差向异构酶活性和ManNAc-6-磷酸磷酸酶活性,为Neu5Ac生产提供了一种额外的或替代的途径,将GlcNAc-6-磷酸转化为ManNAc,如图2所示。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化,以具有编码GlcNAc-6-磷酸差向异构酶或其功能性变体的基因。优选地,非天然存在的微生物已被遗传工程化,以包含一种核酸分子,该核酸分子包含和表达编码GlcNAc-6-磷酸差向异构酶的核苷酸序列。
优选地,GlcNAc-6-磷酸差向异构酶来源于阴沟肠杆菌阴沟亚种(Enterobactercloacae subsp.cloacae)(SEQ ID NO:25)或其功能性变体。编码阴沟肠杆菌阴沟亚种的GlcNAc-6-磷酸差向异构酶的核苷酸序列为阴沟肠杆菌阴沟亚种ATCC 13047的nanE基因的蛋白质编码区(SEQ ID NO:24)。
在一个额外的和/或替代的实施方案中,编码GlcNAc-6-磷酸差向异构酶的功能性变体之一的核苷酸序列与阴沟肠杆菌阴沟亚种nanE基因的序列同一性为至少80%、至少85%、至少90%、至少95%、至少98%或至少99%的。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化,以具有编码ManNAc-6-磷酸磷酸酶或其功能性变体的基因。
N-乙酰神经氨酸合酶(EC 2.5.1.56)是一种利用磷酸烯醇丙酮酸(PEP)催化N-乙酰甘露糖胺(ManNAc)转化为Neu5Ac的酶。该N-乙酰神经氨酸合酶(NeuB)是由neuB基因编码。
在一个额外的和/或替代的实施方案中,非天然存在的微生物包含N-乙酰神经氨酸合酶或其功能性变体,优选异源N-乙酰神经氨酸合酶。在另一个实施方案中,N-乙酰神经氨酸合酶来源于空肠弯曲杆菌(SEQ ID NO:29)、无乳链球菌、分裂蛋白丁酸弧菌(Butyrivibrio proteoclasticus)、反刍兽甲烷短杆菌(Methanobrevibacterruminatium)、伍氏醋酸杆菌(Acetobacterium woodii)、Desulfobacula toluolica、大肠杆菌、变黑普雷沃氏菌(Prevotella nigescens)、Halorhabdus tiamatea、亚磷酸氧化产脱硫菌(Desulfotignum phosphitoxidans)或Candidatus Scalindua sp.、热液海源菌(Idomarina loihiensis)、具核梭杆菌(Fusobacterium nucleatum)或脑膜炎奈瑟氏菌。
在一个额外的和/或替代的实施方案中,非天然存在的微生物包含一种核酸分子,该核酸分子包含编码N-乙酰神经氨酸合酶或其功能性变体的核苷酸序列。在一个额外的和/或替代的实施方案中,编码N-乙酰神经氨酸合酶的核苷酸序列是一种异源核苷酸序列。在一个额外的和/或替代的实施方案中,编码N-乙酰神经氨酸合酶的核苷酸序列选自编码空肠弯曲杆菌NeuB(SEQ ID NO:28)、无乳链球菌NeuB、分裂蛋白丁酸弧菌NeuB、反刍兽甲烷短杆菌NeuB、伍氏醋酸杆菌NeuB、D.toluolica NeuB、大肠杆菌NeuB、变黑普雷沃氏菌NeuB、H.tiamatea NeuB、亚磷酸氧化产脱硫菌NeuB、Ca.scalindua sp.NeuB、热液海源菌NeuB、具核梭杆菌NeuB、脑膜炎奈瑟菌NeuB和其功能性变体的核苷酸序列。
在一个额外的和/或替代的实施方案中,编码N-乙酰神经氨酸合酶或N-乙酰神经氨酸合酶的功能性变体之一的核苷酸序列与编码空肠弯曲杆菌NeuB、无乳链球菌NeuB、分裂蛋白丁酸弧菌NeuB、反刍兽甲烷短杆菌NeuB、伍氏醋酸杆菌NeuB、D.toluolica NeuB、大肠杆菌NeuB、变黑普雷沃氏菌NeuB、H.tiamatea NeuB、亚磷酸氧化产脱硫菌NeuB、Ca.scalindua sp.NeuB、热液海源菌NeuB、具核梭杆菌NeuB、脑膜炎奈瑟菌NeuB的核苷酸序列之一的序列同一性为至少80%、至少85%、至少90%、至少95%、至少98%或至少99%。
唾液酸分解代谢途径
用于生产Neu5Ac的非天然存在的微生物不能利用Neu5Ac。在一个额外的实施方案中,非天然存在的微生物已被遗传工程化,使得其不利用唾液酸。因此,由非天然存在的微生物合成的Neu5Ac既未在天然存在的分解代谢途径中被降解,也未被掺入到脂多糖和/或多唾液酸中。相反,非天然存在的微生物能够将它合成的Neu5Ac分泌到培养基或发酵液中。
非天然存在的微生物能够产生Neu5Ac。为了能够产生Neu5Ac,天然存在的唾液酸分解代谢途径已被失效。破坏微生物中唾液酸分解代谢途径防止由该微生物合成的任何唾液酸被进一步代谢,从而提高了非天然存在的微生物可产生的唾液酸的产量。
在一个额外的和/或替代的实施方案中,通过遗传工程化微生物使天然存在的唾液酸分解代谢途径失效。
在一个额外的和/或替代的实施方案中,通过缺失或以其他方式突变一个或多个编码唾液酸分解代谢所需酶的基因来破坏天然存在的唾液酸分解代谢途径。因此,不再产生唾液酸分解代谢所需的酶,或产生的水平远低于正常水平,例如在野生型微生物中。例如,可以从基因组中缺失一个或多个编码唾液酸分解代谢所需酶的基因,使得根本不产生相应的酶。或者,控制基因表达的调控序列可以被替换或突变,使得所述基因不能被转录或翻译。这种对转录或翻译的削弱(impairment)包括转录或翻译的永久性削弱以及转录或翻译的短暂性削弱。即可以通过诱导或抑制转录或翻译来调节各个基因的转录或翻译。因此,可以在微生物培养过程中任何所需时间点诱导各个基因的表达,优选地通过在培养基中加入诱导各个基因表达的化合物(诱导剂)。在另一个实施方案中,可以在微生物培养过程中任何所需时间点抑制各个基因的表达,优选地通过在培养基中添加一种抑制各个基因表达的化合物(阻遏物),或者通过耗尽培养基中作为诱导剂的任何化合物。在一种不同的方法中,编码唾液酸分解代谢所需的酶的核苷酸序列可以这种方式被改变,即消除酶的活性。这可通过以下方法来实现:改变核苷酸序列以用终止子来替代原始核苷酸序列中的有义密码子(指定一个氨基酸),从而产生一个截短的多肽,该多肽缺乏唾液酸分解代谢所需的酶的活性,或者用另一个指定不同氨基酸的密码子替代有义密码子,从而产生唾液酸分解代谢所需的酶的非功能性变体。
在一个额外的和/或替代的实施方案中,进行破坏或改变以消除非天然存在的微生物中的唾液酸分解代谢的目标基因编码一种或多种选自N-乙酰甘露糖胺激酶、N-乙酰甘露糖胺-6-磷酸差向异构酶、N-乙酰神经氨酸醛缩酶和唾液酸通透酶的酶。
N-乙酰甘露糖胺激酶(EC 2.7.1.60)是一种磷酸化N-乙酰甘露糖胺以产生N-乙酰甘露糖胺-6-磷酸的酶。该N-乙酰甘露糖胺激酶是由nanK基因编码。大肠杆菌nanK的蛋白编码区的核苷酸序列用SEQ ID NO:30表示。
N-乙酰甘露糖胺-6-磷酸差向异构酶是一种将N-乙酰甘露糖胺-6-磷酸(ManNAc-6-P)转化为N-乙酰葡糖胺-6-磷酸(GlcNAc-6-P)的酶。这种酶促反应是从N-乙酰神经氨酸合成D-果糖6-磷酸的子通路的一步。该N-乙酰甘露糖胺-6-磷酸差向异构酶是由nanE基因编码。大肠杆菌nanE蛋白编码区的核苷酸序列用SEQ ID NO:32表示。
N-乙酰神经氨酸醛缩酶,又称N-乙酰神经氨酸裂解酶,催化N-乙酰神经氨酸的可逆羟醛裂解以形成丙酮酸和N-乙酰甘露糖胺(ManNAc)。N-乙酰神经氨酸醛缩酶是由nanA基因编码。大肠杆菌nanA蛋白编码区的核苷酸序列用SEQ ID NO:34表示。
唾液酸通透酶催化唾液酸跨细胞膜的质子依赖性转运。唾液酸通透酶能转运N-乙酰神经氨酸。唾液酸通透酶的变体还可以转运相关的唾液酸N-羟乙酰神经氨酸(Neu5Gc)和3-酮-3-脱氧-D-甘油-D-半乳糖壬酸(KDN)。虽然已知唾液酸通透酶在体外作为双向转运蛋白发挥功能,但它在体内负责细胞外Neu5Ac的细胞输入。唾液酸通透酶是由nanT基因编码。大肠杆菌nanT蛋白编码区的核苷酸序列用SEQ ID NO:36表示。破坏nanT可防止由非天然存在的微生物产生并分泌到培养基中的Neu5Ac的再输入。
在一个额外的和/或替代的实施方案中,与野生型微生物相比,非天然存在的微生物中至少一种选自以下的酶的活性被降低或消除:N-乙酰甘露糖胺激酶、N-乙酰甘露糖胺-6-磷酸差向异构酶、N-乙酰神经氨酸醛缩酶和唾液酸通透酶。在一个额外的和/或替代的实施方案中,微生物已被遗传工程化,以使这些酶的至少一种的活性被降低或消除。在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化,以完全缺失一个或多个编码N-乙酰甘露糖胺激酶、N-乙酰甘露糖胺-6-磷酸差向异构酶、N-乙酰神经氨酸醛缩酶和唾液酸通透酶的基因,以削弱这些基因中的一个或多个的表达,或者通过将突变引入该基因的蛋白编码区来消除一种或多种相应的酶的活性,使得由改变的核苷酸序列编码的多肽不具有由未改变的核苷酸序列编码的酶的酶活性。
在一个额外的和/或替代的实施方案中,非天然存在的微生物不具有N-乙酰葡糖胺-6-磷酸脱乙酰酶和N-乙酰葡糖胺-6-磷酸脱氨酶中的至少一种酶提供的酶活性。
在一个额外的和/或替代的实施方案中,在非天然存在的微生物中N-乙酰葡糖胺-6-磷酸脱乙酰酶和N-乙酰葡糖胺-6-磷酸脱氨酶中的至少一种已被失效。
N-乙酰葡糖胺-6-磷酸脱乙酰酶(EC 3.5.1.25)是参与氨基-糖-核苷酸的生物合成的第一步的酶。它催化N-乙酰葡糖胺-6-磷酸(GlcNAc-6-P)的N-乙酰基水解,以生成葡糖胺-6-磷酸和乙酸。N-乙酰葡糖胺-6-磷酸脱乙酰酶是由nagA基因编码。大肠杆菌nagA的蛋白编码区的核苷酸序列用SEQ ID NO:38表示。
葡糖胺-6-磷酸脱氨酶(EC 3.5.99.6)催化葡糖胺6-磷酸(GlcN6P)的可逆异构化-脱氨以生成果糖6-磷酸(Fru6P)。葡糖胺-6-磷酸脱氨酶是由nagB基因编码。大肠杆菌nagB蛋白编码区的核苷酸序列用SEQ ID NO:40表示。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化,以使N-乙酰葡糖胺-6-磷酸脱乙酰酶和/或葡糖胺-6-磷酸脱氨酶失效。在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化,以完全缺失一个或多个编码N-乙酰葡糖胺-6-磷酸脱乙酰酶和葡糖胺-6-磷酸脱氨酶的基因,以削弱这些基因中的一个或多个的表达,或者通过将突变引入这些基因中至少一个的蛋白编码区来消除一种或多种相应的酶的活性,使得由改变的核苷酸序列编码的多肽不具有由未改变的核苷酸序列编码的酶的酶活性。
磷酸转移酶碳水化合物转运系统
唾液酸的细胞内生产需要磷酸烯醇丙酮酸(PEP)。由于PEP参与糖酵解和糖异生,因此它是一种非常重要的代谢中间体。为了提高Neu5Ac的产量,对非天然存在的微生物进行遗传工程化,以为唾液酸生物合成提供了更好的PEP供应。为此目的,非天然存在的微生物已被遗传工程化,其中至少有一个PEP依赖的糖转运磷酸转移酶系统(PTS)已被失效,即相应的基因被缺失或破坏,或基因的表达被削弱。
适合破坏的PEP依赖的糖转运磷酸转移酶系统为GlcNAc通透酶,也称为蛋白质-Npi-磷酸-L-组氨酸:N-乙酰-D-葡糖胺Npi-磷酸转移酶(EC2.7.1.193),它是由nagE基因编码的。NagE(称为酶II)是PEP依赖的糖转运磷酸转移酶系统的成分。该系统同时将其底物从周质或细胞外间隙转运到细胞质中并磷酸化。缺失或破坏nagE或削弱其表达是有利的,因为这防止了通过消耗PEP来输入GlcNAc,否则将减少可用于唾液酸生产的PEP的量。因此,缺失或破坏nagE或削弱其表达增加了可以被非天然存在的微生物利用来产生Neu5Ac的胞内PEP池,从而增加非天然存在的微生物中Neu5Ac的产量,相对于可以生产唾液酸,但携带完整的功能性nagE基因的非天然存在的微生物而言。大肠杆菌nagE的蛋白编码区的核苷酸序列用SEQ ID NO:42表示。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化,以消除蛋白质-Npi-磷酸-L-组氨酸:N-乙酰-D-葡糖胺Npi-磷酸转移酶活性。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已经被遗传工程化,以完全缺失nagE基因,削弱其表达,或通过将突变引入nagE基因的蛋白编码区来消除NagE酶的活性,使得由改变的核苷酸序列编码的多肽不具有由未改变的核苷酸序列编码的酶的酶活性。
另一个或额外的适合破坏的用于输入碳水化合物的PEP依赖的糖转运磷酸转移酶系统为甘露糖通透酶。
ManXYZ——酶IIMan复合物(甘露糖PTS通透酶、蛋白质-Npi-磷酸组氨酸-D-甘露糖磷酸转移酶)输入外源己糖(甘露糖、葡萄糖、葡糖胺、果糖、2-脱氧葡萄糖、甘露糖胺、N-乙酰葡糖胺等)并将磷酸酯释放到细胞质中。这种酶也是PEP依赖的糖转运磷酸转移酶系统的成分。ManXYZ在三条多肽链中具有四个结构域,即ManX=IIABMan、ManY=IICMan和ManZ=IIDMan。它们是甘露糖PTS通透酶家族分离组(splinter group)的成员,该组与大多数其他PTS通透酶不同源。大肠杆菌manX、manY和manZ的蛋白编码区的核苷酸序列分别用SEQ IDNO:44、SEQ ID NO:46和SEQ ID NO:48表示。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已经被遗传工程化以消除蛋白质-Npi-磷酸-L-组氨酸:甘露糖Npi-磷酸转移酶的活性。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化,以完全缺失一个或多个编码ManX、ManY和ManZ的基因,削弱这些基因中的一个或多个的表达,或通过将突变引入所述基因的蛋白编码区来消除一种或多种相应的酶的活性,使得由改变的核苷酸序列编码的多肽不具有由未改变的核苷酸序列编码的酶的酶活性。
另一个或额外的适合破坏的PEP依赖的糖转运磷酸转移酶系统为葡萄糖转运蛋白。
葡萄糖特异性PTS转运蛋白(PtsG/Crr)吸收外源葡萄糖,将磷酸酯释放到细胞质中。酶IIGlc复合物在单个多肽链中具有两个结构域,结构域序为IIC-IIB(PtsG),它与另一个多肽链Crr或IIAGlc蛋白一起发挥作用。
缺失或破坏ptsG和/或crr或削弱其表达是有利的,因为这防止了通过消耗PEP来输入葡萄糖,否则将减少可用于唾液酸生产的PEP的数量。因此,缺失或破坏ptsG基因和/或crr基因或削弱其表达增加了可以被非天然存在的微生物利用来产生Neu5Ac的胞内PEP池,从而增加能够生产Neu5Ac的非天然存在的微生物的Neu5Ac产量,相对于可以生产唾液酸,但携带完整的功能性ptsG和/或crr基因的非天然存在的微生物而言。
大肠杆菌ptsG和crr的蛋白编码区的核苷酸序列分别用SEQ ID NO:50和SEQ IDNO:52表示。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化以消除PtsG/Crr活性。
在一个额外的和/或替代的实施方案中,非天然存在的微生物经过遗传工程化,完全缺失ptsG基因和/或crr基因,削弱ptsG基因和/或crr的表达,或通过将突变引入ptsG基因和/或crr基因的蛋白编码区来消除PtsG/Crr的活性,使得由改变的核苷酸序列编码的多肽不具有由未改变的核苷酸序列编码的酶的酶活性。
获取碳源
非天然存在的微生物需要碳源来生长、增殖和生产Neu5Ac。在一个额外的和/或替代的实施方案中,非天然存在的微生物可在廉价的唯一碳源上生长,例如葡萄糖或蔗糖。所述唯一碳源为非天然存在的微生物中唾液酸生物合成提供了浸提物(educt)。因此,对于Neu5Ac生产,没有必要在ManNAc、GlcNAc或葡糖胺(GlcN)存在下培养非天然存在的微生物。此外,非天然存在的微生物不需要PEP依赖的糖转运磷酸转移酶系统来输入该唯一的碳源,因此不需要利用PEP来获取该唯一的碳源。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化以利用蔗糖作为唯一的碳源。
在一个额外的和/或替代的实施方案中,非天然存在的微生物具有功能性蔗糖利用系统。所述功能性蔗糖利用系统使得能够将外源供应的蔗糖输入细胞并使蔗糖水解,使得产生的单糖葡萄糖和果糖可以被非天然存在的微生物的代谢以代谢方式利用并用于所需的Neu5Ac生产。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程以具有功能性蔗糖利用系统。在一个额外的和/或替代的实施方案中,非天然存在的微生物的蔗糖利用系统包括蔗糖质子同向转运系统、果糖激酶、转化酶和蔗糖操纵子阻遏物。
一种合适的蔗糖质子同向转运系统为由cscB基因编码的CscB,例如由大肠杆菌cscB基因(SEQ ID NO:54)编码的大肠杆菌CscB(SEQ ID NO:55)。
一种合适的果糖激酶(EC 2.7.1.4)为由cscK基因编码的CscK,例如由大肠杆菌的cscK基因(SEQ ID NO:56)编码的大肠杆菌的CscK(SEQ ID NO:57)。
水解β-D-呋喃果糖苷的末端非还原β-D-呋喃果糖苷残基的一种合适的转化酶(EC3.2.1.26)为CscA,例如由大肠杆菌的cscA基因(SEQ ID NO:58)编码的大肠杆菌的cscA(SEQ ID NO:59)。
一种合适的蔗糖操纵子阻遏物为由cscR基因编码的CscR,例如由大肠杆菌的cscR基因(SEQ ID NO:60)编码的大肠杆菌的CscR(SEQ ID NO:61)。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已经被遗传工程化以具有蔗糖质子同向转运系统、果糖激酶、转化酶和蔗糖操纵子阻遏物或这些蛋白质中任何一种的功能性变体。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化以具有一种核酸分子,该核酸分子包含编码蔗糖质子同向转运系统、果糖激酶、转化酶和蔗糖操纵子阻遏物的核苷酸序列,用于表达所述蔗糖质子同向转运系统、果糖激酶、转化酶和蔗糖操纵子阻遏物。在一个额外的和/或替代的实施方案中,非天然存在的微生物已经被遗传工程化来表达基因cscB、cscK、cscA,优选大肠杆菌基因cscB、cscK、cscA和cscR。
在一个额外的和/或替代的实施方案中,编码CscB、CscK、CscA或CscR的功能性变体的核苷酸序列分别与大肠杆菌cscB、cscK、cscA或cscR具有至少80%、至少85%、至少90%、至少95%、至少98%或至少99%的序列同一性。
能够产生Neu5Ac并携带功能性蔗糖利用系统的非天然存在的微生物可以在作为用于微生物代谢和Neu5Ac生物合成的唯一碳源的蔗糖存在下培养。蔗糖是一种廉价的糖,它作为通过发酵生产Neu5Ac的唯一碳源比其他唾液酸前体(如GlcNAc)更具成本效益。
另一种合适的允许非天然存在的微生物在唯一的碳源上生长,而不需要PEP依赖的糖转运磷酸转移酶系统的糖利用系统为LacY,其由lac操纵子的lacY基因编码。LacY是一种β-半乳糖苷通透酶,利用质子梯度在同一方向上跨细胞膜输入乳糖。细胞内乳糖可被β-半乳糖苷酶(LacZ)水解以在细胞内提供葡萄糖和半乳糖。lacZ基因也是lac操纵子的一部分。
在一个额外的和/或替代的实施方案中,非天然存在的微生物表达β-半乳糖苷通透酶和β-半乳糖苷酶。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化以表达β-半乳糖苷通透酶,优选大肠杆菌乳糖通透酶LacY(SEQ ID NO:63)或其功能性变体;和β-半乳糖苷酶,优选大肠杆菌LacZ(SEQ ID NO:65)或其功能性变体。在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化以携带一种核酸分子,该核酸分子包含编码β-半乳糖苷通透酶的核苷酸序列,优选编码大肠杆菌LacY(SEQ ID NO:62)或其功能性变体的核苷酸序列,和/或编码β-半乳糖苷酶的核苷酸序列,优选编码大肠杆菌LacZ(SEQ ID NO:64)或其功能性变体的核苷酸序列。
在一个额外的和/或替代的实施方案中,编码大肠杆菌LacY或其功能性变体的核苷酸序列与大肠杆菌lacY的序列同一性为至少80%、至少85%、至少90%、至少95%、至少98%或至少99%。
在一个额外的和/或替代的实施方案中,编码大肠杆菌LacZ或其功能性变体的核苷酸序列与大肠杆菌lacZ的序列同一性为至少80%、至少85%、至少90%、至少95%、至少98%或至少99%。
可以产生Neu5Ac并表达功能性β-半乳糖苷通透酶和功能性β-半乳糖苷酶的非天然存在的微生物允许在作为唯一碳源的乳糖上培养所述非天然存在的微生物。
在一个额外的和/或替代的实施方案中,非天然存在的微生物表达葡萄糖/H+-同向转运体。优选地,非天然存在的微生物已被遗传工程化,以携带一种核酸分子,该核酸分子包含编码葡萄糖/H+-同向转运体并允许在所述非天然存在的微生物中表达葡萄糖/H+-同向转运体的核苷酸序列。
合适的葡萄糖/H+-同向转运体选自表皮葡萄球菌(Staphylococcus epidermis)葡萄糖/H+-同向转运体(UniProtKB-A0A0U5QDM9;SEQ ID NO:67)、短乳杆菌(Lactobacillus brevis)葡萄糖/H+-同向转运体(UniProtKB-A0A0C1PU75,SEQ ID NO:69)及其功能性变体。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已经被遗传工程化以携带编码表皮葡萄球菌葡萄糖/H+-同向转运体或短乳杆菌葡萄糖/H+-同向转运体的核酸分子。优选地,非天然存在的微生物携带一种核酸分子,该核酸分子包含选自以下的核苷酸序列:SEQ ID NO:66、SEQ ID NO:68,以及与SEQ ID NO:66或SEQ ID NO:68具有至少80%、至少85%、至少90%、至少95%、至少98%或至少99%的序列同一性的核苷酸序列。
可以产生Neu5Ac并表达表皮葡萄球菌葡萄糖/H+-同向转运体或短乳杆菌葡萄糖/H+-同向转运体的非天然存在的微生物可以在作为唯一碳源的葡萄糖存在下培养,而不需要PEP来获得外源提供的葡萄糖。
额外的基因修饰
可以产生Neu5Ac的非天然存在的微生物可以任选地包括额外的特征,并可被遗传工程化以具有这些额外的特征。这些额外的特征被认为提高了非天然存在的微生物的生产力,获得更高的Neu5Ac产量。
在一个额外的和/或替代的实施方案中,非天然存在的微生物合成的PEP比野生型微生物多。在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化,以具有增强的PEP生物合成途径。优选地,非天然存在的微生物已被遗传工程化以具有增加的磷酸烯醇丙酮酸合酶活性,例如,其中编码磷酸烯醇丙酮酸合酶的ppsA基因被过表达和/或其中非天然存在的微生物包含允许表达磷酸烯醇丙酮酸合酶或其功能性变体的核苷酸序列的至少一个额外拷贝。过表达ppsA增加了细胞内PEP的合成,使得更多的PEP可用于唾液酸的生产。例如,一种合适的磷酸烯醇丙酮酸合酶为大肠杆菌的PpsA(SEQ ID NO:71)。
在一个额外的和/或替代的实施方案中,非天然存在的微生物包含一种核酸分子,该核酸分子包含编码大肠杆菌PpsA或其功能性变体的核苷酸序列。所述的编码大肠杆菌PpsA或其功能性变体的核苷酸序列与大肠杆菌ppsA基因(SEQ ID NO:70)的序列同一性为至少80%、至少85%、至少90%、至少95%、至少98%或至少99%。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化以过表达磷酸烯醇丙酮酸羧激酶。一种合适的磷酸烯醇丙酮酸羧激酶为大肠杆菌Pck(SEQ IDNO:73)。
磷酸烯醇丙酮酸羧激酶(EC 4.1.1.49)由pck基因编码,催化下列反应:草酰乙酸+ATP→磷酸烯醇丙酮酸+ADP+CO2。磷酸烯醇丙酮酸羧激酶参与糖异生。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化以过表达磷酸烯醇丙酮酸羧激酶和/或包含至少一种允许表达磷酸烯醇丙酮酸羧激酶或其功能性变体的额外的核苷酸序列。磷酸烯醇丙酮酸羧激酶的过表达增加了PEP的细胞内水平,使得更多的PEP可用于唾液酸的生产。
编码磷酸烯醇丙酮酸激酶或其功能性变体的额外的核苷酸序列可以是SEQ IDNO:72或与大肠杆菌pck基因(SEQ ID NO:72)的序列同一性为至少80%、至少85%、至少90%、至少95%、至少98%或至少99%的核苷酸序列。
在一个额外的和/或替代的实施方案中,非天然存在的微生物不具有功能性磷酸烯醇丙酮酸羧化酶(EC 4.1.1.31)。磷酸烯醇丙酮酸羧化酶形成草酰乙酸、三羧酸循环的四碳二羧酸源。磷酸烯醇丙酮酸羧化酶是由ppc基因编码。在大肠杆菌中,磷酸烯醇丙酮酸羧化酶(SEQ ID NO:27)是由pepC基因编码(SEQ ID NO:26)。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化以消除PEP羧化酶活性。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化缺失ppc基因或pepC基因,削弱其表达,或通过在ppc/pepC基因的蛋白编码区中引入突变而消除PEP羧化酶的活性,使得由改变的核苷酸序列编码的多肽不具有PEP羧化酶活性。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化,以降低或消除丙酮酸激酶活性。
丙酮酸激酶从二磷酸腺苷(ADP)和PEP生成三磷酸腺苷(ATP)。从ADP和PEP生成ATP是糖酵解的最后一步,这一步骤在生理条件下是不可逆的。许多肠杆菌科(Enterobacteriaceae)(包括大肠杆菌)有两种丙酮酸激酶亚型,即PykA(SEQ ID NO:75)和PykF(SEQ IC NO:77),它们在大肠杆菌中有37%的同一性。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化,以缺失一个或多个编码丙酮酸激酶的基因,优选pykA基因(SEQ ID NO:74)和/或pykF基因(SEQ ID NO:76),以削弱这些编码丙酮酸激酶的基因中的一个或多个的表达,或者通过在这些编码丙酮酸激酶的基因中一个或多个基因的蛋白编码区的核苷酸序列中引入一个或多个突变而消除至少一种丙酮酸激酶的活性,使得由改变的核苷酸序列编码的多肽不具有丙酮酸激酶活性。
在一个额外的和/或替代的实施方案中,非天然存在的微生物合成的谷氨酰胺比野生型更多。在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化以具有增强的谷氨酰胺生物合成途径。
谷氨酰胺合酶(GlnA)通过以下反应将谷氨酸转化为谷氨酰胺:ATP+L-谷氨酸+NH3=ADP+磷酸+L-谷氨酰胺。在大肠杆菌中,谷氨酰胺合酶(SEQ ID NO:79)是由glnA基因(SEQID NO:78)编码。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化以过表达谷氨酰胺合酶和/或包含至少一种允许表达谷氨酰胺合酶或其功能性变体的额外的核苷酸序列。谷氨酰胺合酶的过表达增加了谷氨酰胺的细胞内水平,其进而提高了果糖-6-磷酸(Frc-6P)向葡糖胺-6-磷酸(GlcN-6P)的细胞内转化。优选地,编码谷氨酰胺合酶或其功能性变体的额外的核苷酸序列可以是SEQ ID NO:78,或与大肠杆菌glnA基因具有至少80%、至少85%、至少90%、至少95%、至少98%或至少99%的序列同一性的核苷酸序列。
大肠杆菌的代谢模型证实,增强谷氨酰胺合成增强了Neu5Ac的产生。此外,一种未经遗传工程化以增强谷氨酰胺合成的产生Neu5Ac的大肠杆菌菌株(#NANA1)的转录组分析表明,与不能产生Neu5Ac的相关大肠杆菌菌株相比,谷氨酰胺合酶以更高水平表达。
在一个额外的和/或替代的实施方案中,与野生型微生物相比,非天然存在的微生物不携带功能性谷氨酸合酶或谷氨酸合酶活性较低。大肠杆菌谷氨酸合酶由两个亚基组成,即GltB(SEQ ID NO:81)和GltD(SEQ ID NO:83),通过消耗谷氨酰胺来合成谷氨酸。GltB是由gltB基因(SEQ ID NO:80)编码,GltD是由gltD基因(SEQ ID NO:82)编码。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化以缺失gltB基因和/或gltD基因,削弱这些基因中至少一种的表达,或通过在gltB基因和/或gltD基因的蛋白编码区中引入一种或多种突变来降低或消除谷氨酸合酶活性,使得由改变的核苷酸序列编码的多肽提供谷氨酸合酶的非功能性变体。
在一个额外的和/或替代的实施方案中,非天然存在的微生物不具有谷氨酰胺酶活性。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化,以缺失谷氨酰胺酶基因asnB、ybaS和yneH中的至少一种,以削弱这些基因中至少一种的表达,或通过在asnB、ybaS和/或yneH的蛋白质编码区中引入一种或多种突变来消除谷氨酰胺酶失活,使得由改变的核苷酸序列之一编码的多肽不具有谷氨酰胺酶活性。
AsnB是一种天冬酰胺合酶,利用谷氨酰胺催化天冬氨酸向天冬酰胺的ATP依赖性转化。大肠杆菌天冬酰胺合成酶AsnB(SEQ ID NO:85)是由大肠杆菌asnB基因(SEQ ID NO:84)编码。
YbaS(也称为GlsA1或Gls1)为谷氨酰胺酶1,是一种对L-谷氨酰胺具有高度选择性的谷氨酰胺酶。YbaS将L-谷氨酰胺转化为L-谷氨酸。大肠杆菌谷氨酰胺酶YbaS(SEQ ID NO:87)是由大肠杆菌ybaS基因(SEQ ID NO:86)编码。
YneH也称为GlsA2、GlsB或谷氨酰胺酶2,催化下列反应:L-谷氨酰胺+H2O=L-谷氨酸+NH3。大肠杆菌谷氨酰胺酶YneH(SEQ ID NO:89)是由大肠杆菌yneH基因(SEQ ID NO:88)编码。
在一个额外的和/或替代的实施方案中,与野生型微生物相比,非天然存在的微生物具有增加的谷氨酸脱氢酶活性。在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化以过表达谷氨酸脱氢酶和/或包含至少一种允许表达谷氨酸脱氢酶或其功能性变体的额外的核苷酸序列。
谷氨酸脱氢酶将谷氨酸转化为α-酮戊二酸。谷氨酸脱氢酶的过表达增加了α-酮戊二酸的形成,其进而可以通过谷氨酸合酶(例如由大肠杆菌gltD编码的谷氨酸成酶或其功能性变体)被转化为谷氨酸。然后谷氨酸可以通过谷氨酰胺合成酶(GlnA)或其功能性变体被转化为谷氨酰胺。
在一个额外的和/或替代的实施方案中,允许表达谷氨酸脱氢酶或其功能性变体的额外的核苷酸序列包括大肠杆菌谷氨酸脱氢酶GdhA(SEQ ID NO:91)的蛋白质编码区。编码谷氨酸脱氢酶或其功能性变体的核苷酸序列可以是SEQ ID NO:90,或与大肠杆菌gdhA基因(SEQ ID NO:90)具有至少80%、至少85%、至少90%、至少95%、至少98%或至少99%的序列同一性的核苷酸序列。
在一个额外的和/或替代的实施方案中,非天然存在的微生物无法合成脂多糖(LPS)和/或荚膜异多糖酸(colanic acid)。在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化,以消除LPS和/或荚膜异多糖酸的合成。
在一个实施方案中,非天然存在的微生物已经被遗传工程化来缺失wzxC基因,削弱wzxC基因的表达,或者通过在该基因的蛋白质编码区中引入一个或更多的突变而消除WzxC酶活性,使得由所述改变的核苷酸序列编码的多肽不具有WzxC的酶活性。WzxC是LPS生物合成所必需的,并编码一种假定的输出蛋白。大肠杆菌wzxC的核苷酸序列用SEQ ID NO:92表示,推导出的氨基酸序列用SEQ ID NO:93表示。
在一个额外的和/或替代的实施方案中,非天然存在的微生物不具有UDP-葡萄糖:十一异戊烯基磷酸葡萄糖-1-磷酸转移酶活性。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化而消除UDP-葡萄糖:十一异戊烯基磷酸葡萄糖-1-磷酸转移酶活性,优选地通过缺失wcaJ基因或其功能性变体,通过削弱wcaJ基因或其功能性变体的表达,或通过将突变引入该基因的蛋白质编码区而消除WcaJ酶的活性,使得由改变的核苷酸序列编码的多肽不具有WcaJ的酶活性。WcaJ编码UDP-葡萄糖:十一异戊烯基磷酸葡萄糖-1-磷酸转移酶。所述UDP-葡萄糖:十一异戊烯基磷酸葡萄糖-1-磷酸转移酶是荚膜异多糖酸生物合成的第一种酶。大肠杆菌wcaJ的核苷酸序列用SEQ ID NO:94表示,推导出的氨基酸序列用SEQ ID NO:95表示。
在一个额外的和/或替代的实施方案中,非天然存在的微生物可能不包含功能性β-半乳糖苷通透酶(LacY)和/或功能性β-半乳糖苷酶(LacZ),条件是非天然存在的微生物可以另一种非乳糖的唯一碳源上培养,例如在作为唯一碳源的蔗糖或葡萄糖上。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化,其中β-半乳糖苷通透酶基因(lacY)和/或β-半乳糖苷酶基因(lacZ)已被缺失,其中β-半乳糖苷通透酶基因和/或β-半乳糖苷酶基因的表达被削弱,或其中β-半乳糖苷通透酶基因和/或β-半乳糖苷酶基因的蛋白质编码区的核苷酸序列被改变,使得由所述改变的核苷酸序列编码的多肽不具有β-半乳糖苷通透酶和/或β-半乳糖苷酶的酶活性。
在一个额外的和/或替代的实施方案中,非天然存在的微生物不具有功能性YjhC。YjhC是由yjhC基因编码的氧化还原酶或其功能性变体。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化以消除YjhC氧化还原酶活性,优选地通过缺失yjhC基因,通过削弱yjhC基因的表达,或通过在yjhC基因的蛋白质编码区中引入一个或多个突变,使得由被改变的核苷酸序列编码的多肽不具有YjhC氧化还原酶活性。
大肠杆菌yjhC的核苷酸序列用SEQ ID NO:96表示,推导出的氨基酸序列用SEQ IDNO:97表示。
在一个额外的和/或替代的实施方案中,非天然存在的微生物不具有一种或多种以下酶活性:岩藻糖异构酶、墨角藻糖激酶和N-乙酰谷氨酰胺氨基酰化酶。在一个实施方案中,非天然存在的微生物已被遗传工程化,以消除这些酶活性中的一种或多种的活性。
岩藻糖异构酶将醛糖L-岩藻糖转化为相应的酮糖L-墨角藻糖(L-fuculose)。岩藻糖异构酶是从L-岩藻糖合成L-乳醛和磷酸甘油酮的子途径中的第一种酶。大肠杆菌岩藻糖异构酶Fucl(SEQ ID NO:99)是由大肠杆菌fucl基因(SEQ ID NO:98)编码。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化而消除岩藻糖异构酶活性,优选地通过缺失fucl基因,通过削弱fucl基因的表达或通过修饰fucl基因的蛋白质编码区,使得所述改变的核苷酸序列编码的多肽不具有岩藻糖异构酶活性。
墨角藻糖激酶(Fuculokinase)催化岩藻糖磷酸化。墨角藻糖激酶是从L-岩藻糖合成L-乳醛和磷酸甘油酮的子通路中的第二种酶。大肠杆菌墨角藻激酶FucK(SEQ ID NO:101)是由大肠杆菌fucK基因(SEQ ID NO:100)编码。大肠杆菌墨角藻糖激酶也可以较低效率磷酸化D-核酮糖、D-木糖和D-果糖。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化而消除岩藻糖异构酶活性,优选地通过缺失fucK基因,或通过削弱fucK基因的表达,或通过将突变引入fucK基因的蛋白质编码区,使得由所述改变的核苷酸序列编码的多肽不具有岩藻糖异构酶活性。
N-乙酰半乳糖胺-6-磷酸脱乙酰酶催化下列反应:N-乙酰-D-半乳糖胺6-磷酸+H2O→D-半乳糖胺6-磷酸+乙酸。N-乙酰半乳糖胺-6-磷酸脱乙酰酶是由agaA基因编码。与大肠杆菌菌株C和EC3132不同,K-12菌株不能在N-乙酰半乳糖胺和D-半乳糖胺上生长,因为它们携带缺失并因此缺乏特异性针对这些化合物的活性PTS系统。因此,在K-12菌株中,AgaA不参与这些化合物的降解。大肠杆菌AgaA(SEQ ID NO:103)是由大肠杆菌agaA基因(SEQ IDNO:102)编码。
在一个额外的和/或替代的实施方案中,非天然存在的微生物已被遗传工程化而消除N-乙酰半乳糖胺-6-磷酸脱乙酰酶活性,优选地通过缺失agaA基因,通过削弱agaA基因的表达,或通过将突变引入agaA基因的蛋白质编码区,使得由所述改变的核苷酸序列编码的多肽不具有N-乙酰半乳糖胺-6-磷酸脱乙酰酶活性。
非天然存在的微生物选自酵母菌、真菌和细菌。优选地,非天然存在的微生物是一种通常被认为是安全的(GRAS)的生物,例如由联邦药物管理局(FDA)确认的或由有资格的专家独立确定的,更优选原核微生物,最优选细菌微生物。适合生产Neu5Ac的细菌可选自以下属:芽孢杆菌属(Bacillus)、乳杆菌属(Lactobacillus)、乳球菌属(Lactococcus)、肠球菌属(Enterococcus)、双歧杆菌属(Bifidobacterium)、芽胞乳杆菌属(Sporolactobacillus)、小单孢菌属(Micromonospora)、微球菌属(Micrococcus)、红球菌属(Rhodococcus)和假单胞菌属(Pseudomonas)。适合的细菌种包括枯草芽孢杆菌(Bacillus subtilis)、地衣芽孢杆菌(Bacillus licheniformis)、凝结芽孢杆菌(Bacillus coagulans)、嗜热芽孢杆菌(Bacillus thermophilus)、侧孢芽孢杆菌(Bacillus laterosporus)、巨大芽孢杆菌(Bacillus megaterium)、蕈状芽孢杆菌(Bacillus mycoides)、短小芽孢杆菌(Bacillus pumilus)、迟缓芽孢杆菌(Bacilluslentus)、蜡样芽孢杆菌(Bacillus cereus)、环状芽孢杆菌(Bacillus circulans)、长双歧杆菌(Bifidobacterium longum)、婴儿双歧杆菌(Bifidobacterium infantis)、两歧双歧杆菌(Bifidobacterium bifidum)、弗氏柠檬酸杆菌(Citrobacter freundii)、解纤维素梭菌(Clostridium cellulolyticum)、杨氏梭菌(Clostridium ljungdahlii)、自产乙醇梭菌(Clostridium autoethanogenum)、丙酮丁醇梭菌(Clostridium acetobutylicum)、谷氨酸棒状杆菌(Corynebacterium glutamicum)、屎肠球菌(Enterococcus faecium)、嗜热肠球菌(Enterococcus thermophiles)、大肠杆菌、草生欧文氏菌(Erwinia herbicola)(成团泛菌(Pantoea agglomerans))、嗜酸乳杆菌(Lactobacillus acidophilus)、唾液乳杆菌(Lactobacillus salivarius)、胚牙乳杆菌(Lactobacillus plantarum)、瑞士乳杆菌(Lactobacillus helveticus)、德氏乳杆菌(Lactobacillus delbrueckii)、鼠李糖乳杆菌(Lactobacillus rhamnosus)、保加利亚乳杆菌(Lactobacillus bulgaricus)、卷曲乳杆菌(Lactobacillus crispatus)、加氏乳杆菌(Lactobacillus gasseri)、干酪乳杆菌(Lactobacillus casei)、罗伊氏乳杆菌(Lactobacillus reuteri)、詹氏乳杆菌(Lactobacillus jensenii)、乳酸乳球菌(Lactococcus lactis)、柠檬泛菌(Pantoeacitrea)、胡萝卜软腐果胶杆菌(Pectobacterium carotovorum)、费氏丙酸杆菌(Proprionibacterium freudenreichii)、荧光假单胞菌(Pseudomonas fluorescens)、铜绿假单胞菌(Pseudomonas aeruginosa)、嗜热链球菌(Streptococcus thermophiles)和野油菜黄单胞菌(Xanthomonas campestris)。
根据第二方面,提供了如前所述的非天然存在的微生物用于生产Neu5Ac的用途。非天然存在的微生物能够以工业规模产生Neu5Ac。本文所用的关于Neu5Ac生产的术语“能够(capable)”和“能够(able)”是指非天然存在的微生物合成Neu5Ac并将所述Neu5Ac分泌到发酵液中的能力,条件是所述非天然存在的微生物是在允许生产Neu5Ac的条件下培养的。这包括所述非天然存在的微生物增殖到高细胞密度和以大体积培养的能力,例如体积超过1,000L,优选10,000L,更优选80,000L,最优选200,000L。
根据第三方面,提供了一种用于通过微生物发酵生产Neu5Ac的方法。该方法包括以下步骤:
-提供一种能够产生Neu5Ac的非天然存在的微生物,优选如前所述的非天然存在的微生物;
-在允许通过所述微生物生产Neu5Ac的条件下,在发酵液中培养所述非天然存在的微生物;以及任选地
-从发酵液中回收Neu5Ac。
发酵液中含有至少一种用于所述非天然存在微生物的碳源。该碳源优选地选自葡萄糖、木糖、果糖、蔗糖、乳糖、甘油、合成气及其组合。
在一个额外的和/或替代的实施方案中,非天然存在的微生物是在不存在和/或不添加选自丙酮酸、葡糖胺、N-乙酰葡糖胺的一种或多种的情况下在发酵液中培养的。
该方法包括在发酵液中培养非天然存在的微生物期间回收由其产生的Neu5Ac的任选步骤。在已将非天然存在的微生物从发酵液中移除后,可以从发酵液中回收Neu5Ac,例如通过离心。随后,通过例如微滤、超滤、渗滤、模拟移动床型色谱、电渗析、反渗透、凝胶过滤、阴离子交换色谱法、阳离子交换色谱等合适的技术,可以从由此获得的澄清发酵液中进一步纯化Neu5Ac。
该方法适用于通过微生物发酵进行的Neu5Ac的大规模、经济上可持续的生产。
根据第四方面,提供了由如本文所述的微生物发酵产生的Neu5Ac。
根据第五方面,提供了如本文所述生产的Neu5Ac用于制备营养组合物的用途。
根据第六方面,提供了含有由第三方面的方法生产的Neu5Ac的营养组合物。
在一个额外的和/或替代的实施方案中,所述营养组合物还含有至少一种人乳低聚糖(HMO),优选至少一种中性HMO和/或至少一种酸性HMO。
中性HMO可选自2’-岩藻糖基乳糖(2’-FL)、3-岩藻糖基乳糖(3-FL)、乳-N-四糖(LNT)、乳-N-新四糖(LNnT)和乳-N-岩藻戊糖I(LNPFI)。
酸性HMO可选自唾液酸化HMO,优选地选自3’-唾液酸乳糖(3-SL)、6’-唾液酸乳糖(6-SL)、唾液酸乳-N-四糖a(LST-a)、唾液酸乳-N-四糖b(LST-b)、唾液酸乳-N-四糖c(LST-c)和双唾液酸乳-N-四糖(DSLNT)。
在另一个实施方案中,所述营养组合物选自药物制剂、婴儿配方物和膳食补充剂。
所述营养组合物可以液体形式或固体形式(包括但不限于粉末剂、颗粒剂(granule)、薄片(flake)和丸剂(pellet))存在。
在一个额外的和/或替代的实施方案中,营养组合物还包括微生物,优选益生菌微生物。对于婴儿食品应用,优选的微生物来源于健康人类的微生物群系或可以在其中找到。优选地,微生物选自以下属:双歧杆菌属、乳杆菌属、肠球菌属、链球菌属、葡萄球菌属(Staphylococcus)、消化链球菌属(Peptostreptococcus)、明串珠菌属(Leuconostoc)、梭菌属(Clostridium)、真细菌属(Eubacterium)、韦永氏球菌属(Veilonella)、梭杆菌属(Fusobacterium)、拟杆菌属(Bacterioides)、普氏菌属(Prevotella)、埃希氏菌属(Escherichia)、丙酸杆菌属(Propionibacterium)或酵母属(Saccharomyces),但其他的也可能是适当的。在一个额外的和/或替代的实施方案中,微生物选自短双歧杆菌(Bifidobacterium breve)、长双歧杆菌、乳酸双歧杆菌(Bifidobacterium lactis)、动物双歧杆菌(Bifidobacterium animalis)、两岐双岐杆菌、婴儿双岐杆菌、Bifidobacteriumaldolescentis、嗜酸乳杆菌、胚牙乳杆菌、唾液乳杆菌、干酪乳杆菌、加氏乳杆菌、罗伊氏乳杆菌、鼠李糖乳杆菌、胚牙乳杆菌、唾液乳杆菌、乳酸乳球菌、副干酪乳杆菌(Lactobacillusparacasei)、保加利亚乳杆菌、瑞士乳杆菌、发酵乳杆菌(Lactobacillus fermentum)、肠系膜明串珠菌(Leuconostoc mesenteroides)、大肠杆菌、屎肠球菌和嗜热链球菌(VSL#3)。
除了Neu5Ac与活生物体的组合外,Neu5Ac还可与有时用于益生菌领域的灭活培养物(例如间歇灭菌的(tyndalized)细菌)组合使用。这种灭活培养物可提供蛋白质、肽、低聚糖、细胞壁碎片和天然产物,导致对免疫系统的短期刺激。
在营养组合物中Neu5Ac和益生菌微生物的组合特别有利于在肠道中建立或重建适当的微生物群系,并促进与此相关的健康益处。
甚至更有利的是唾液酸与已确认的益生元(如低聚半乳糖(GOS)和/或低聚果糖(FOS),包括菊粉)的组合。
将就特定实施方案并参考附图来描述本发明,但本发明不仅限于此,而仅由权利要求来限定。此外,说明书和权利要求中的术语第一、第二和等用于区分相似的要素,而不必用于描述时间、空间、排行或任何其他方式的顺序。应该理解,如此使用的术语在适当的情况下是可互换的,并且本文描述的本发明的实施方案能够以不同于本文描述或举例说明的其他顺序操作。
应当注意的是,权利要求中使用的术语“包括”不应被解释为限于其后列出的方法;它不排除其他要素或步骤。因此,应被理解为指明所述特征、整数、步骤或成分的存在,但不排除存在或添加一个或多个其他特征、整数、步骤或成分或其组。因此,表述“包括装置A和B的设备”的范围不应限于仅由组件A和B组成的设备。这意味着,就本发明而言,该设备的唯一相关组件是A和B。
在本说明书中提及一个“实施方案”是指在本发明的至少一个实施方案中包括结合该实施方案描述的特定特性、结构或特征。因此,在本说明书中各处出现的短语“在一个实施方案中(in one embodiment)”或“在一个实施方案中(in an embodiment)”并不必然总是指相同的实施方案,而是可能指许多和/或不同的实施方案。此外,在一个或多个实施方案中,可以以任何适当的方式组合特定的特性、结构或特征,这对于本公开内容所属领域中的技术人员来说是显而易见的。
类似地,应当理解,在本发明的代表性实施方案的描述中,有时将本发明的各个特征组合在单个实施方案、附图或其描述中,以简化公开内容并促进对各个发明方面中的一个或多个的理解。然而,本公开内容的方法不应被解释为反映以下意图:所要求保护的发明需要比每个权利要求中明确列出的特征更多的特征。而是,如以下权利要求所反映的,本发明的方面可能少于任何前述公开的实施方案的所有特征的特征。因此,在详细说明后的权利要求在此明确纳入该详细说明中,其中每个权利要求独立地作为本发明的单独的实施方案。
此外,尽管本文描述的一些实施方案包括其他实施方案中包括的一些特征但不包括其他特征,但是不同实施方案的特征的组合意在落入本发明的范围内,并且形成不同的实施方案,如本领域技术人员将理解的那样。例如,在下文的权利要求中,任何要求保护的实施方案都可以任何组合使用。
此外,一些实施方案在本文中被描述为可以通过计算机系统的处理器或通过执行该功能的其他手段来应用的方法或方法要素(element)的组合。因此,具有用于执行这种方法或方法要素的必要指令的处理器构成用于执行该方法或方法要素的手段。此外,本文描述的装置实施方案的元件(element)是用于实施由该元件执行的功能,以实施本发明的手段的实例。
在本文提供的说明书和附图中,阐述了许多具体细节。然而,应当理解,可以在没有这些具体细节的情况下实施本发明的实施方案。在其他情况下,未详细示出公知的方法、结构和技术,以促进对说明书和附图的理解。
现在将通过对本发明的几个实施方案的详细描述来描述本发明。显然,在不脱离本发明的真实精神或技术优点的情况下,可以根据本领域技术人员的知识来配置本发明的其他实施方案,本发明仅由所附权利要求的条款来限制。
实施例
实施例1:生产N-乙酰神经氨酸的大肠杆菌BL21(DE3)菌株的代谢工程化
代谢工程化是通过特定内源性基因的诱变和缺失以及异源基因的基因组整合来实现的。通过使用错配寡核苷酸的诱变,使基因lacZ和araA失活,如Ellis等人所述(Proc.Natl.Acad.Sci.USA 98:6742-6746(2001))。
基因组缺失是按照Datsenko and Wanner(Proc.Natl.Acad.Sci.USA 97:6640-6645(2000))的方法产生的。为了防止N-乙酰葡糖胺的降解,从大肠杆菌菌株BL21(DE3)的基因组中缺失了以下基因:N-乙酰葡糖胺特异性PTS酶II(nagE)、N-乙酰葡糖胺-6-磷酸脱乙酰酶(nagA)和葡糖胺-6-磷酸脱氨酶(nagB)。还缺失了编码N-乙酰甘露糖胺激酶(nanK)、N-乙酰甘露糖胺-6-磷酸差向异构酶(nanE)、N-乙酰神经氨酸醛缩酶(nanA)和唾液酸通透酶(nanT)的整个N-乙酰神经氨酸分解代谢基因簇。还缺失了编码促进葡糖胺输入的磷酸烯醇丙酮酸依赖的磷酸转移酶系统的基因manX、manY和manZ。还缺失了wzxC-wcaJ基因。wcaJ基因编码UDP-葡萄糖:十一异戊烯基磷酸葡萄糖-1-磷酸转移酶,其催化荚膜异多糖酸合成的第一步(Stevenson et al.,J.Bacteriol.1996,178:4885-4893)。此外,缺失了基因fucl和fucK和agaA,其分别编码L-岩藻糖异构酶、L-墨角藻糖激酶和N-乙酰半乳糖胺-6-磷酸脱乙酰酶。
使用EZ-Tn5TM转座酶(Epicentre,USA)或水手转座酶Himar1的超活性C9-突变体,通过转座实现异源基因的基因组整合(Proc.Natl.Acad.Sci.1999,USA 96:11428-11433)。为了生产EZ-Tn5转座体,扩增目的基因和侧翼为FRT位点的抗生素抗性标记(或者,抗性标记基因的侧翼为lox66-lox71位点)。所得到的PCR产物在两个末端携带EZ-Tn5转座酶的19-bp嵌合端识别位点。对于使用Himar1转座酶的整合,将目的表达构建体(操纵子)与侧翼为FRT位点/lox66-lox71位点的抗生素抗性标记一起以类似方式克隆并转移到pEcomar载体中,其编码在阿拉伯糖诱导型启动子ParaB的控制下的水手转座酶Himar1的超活性C9-突变体。所有基因均经密码子优化以在大肠杆菌中表达,并由GenScript公司合成制备。
使用EZ-Tn5转座酶整合表达片段<Ptet-lacY-FRT-aadA-FRT>(SEQ ID NO:1)。在成功整合来自大肠杆菌K12 TG1(GenBank:ABN72583)的乳糖输入蛋白LacY的基因后,通过在质粒pCP20上编码的FLP重组酶从链霉素抗性克隆中去除抗性基因(Proc.Natl.Acad.Sci.2000,USA97:6640-6645)。来自大肠杆菌W(GenBank:CP002185.1)的csc基因簇(SEQ ID NO:2),包括使该菌株能够在作为唯一碳源的蔗糖上生长的蔗糖通透酶、果糖激酶、蔗糖水解酶和转录阻遏因子的基因(分别为基因cscB、cscK、cscA和cscR),也被插入到基因组中。利用质粒pEcomar-cscABKR通过转座将该基因簇整合到大肠杆菌BL21(DE3)菌株的基因组中。
将得到的菌株通过以下表达盒的基因组整合进一步修饰以产生Neu5Ac:<Ptet-slr1975-gna1-lox66-aacC1-lox71>(SEQ ID NO:3)、<Ptet-neuB-lox66-kanR-lox71>(SEQID NO:4)、<Ptet-slr1975-Pt5-neuB-FRT-dhfr-FRT>(SEQ ID NO:5)、<Ptet-glmS*-gna1-lox66-aacC1-lox71>(SEQ ID NO:6)和<Ptet-ppsA-lox66-aacC1-lox71>(SEQ ID NO:7)。除了SEQ ID NO:5的dhfr表达盒外,通过引入质粒pKD-Cre(SEQ ID NO:8),然后在含有100μg·mL-1氨苄西林和100mM L-阿拉伯糖的2YT琼脂平板上在30℃下筛选,将所有抗性标记基因都以逐步的方式从基因组中移除(在下一轮基因整合之前)。抗性克隆随后被转移到缺乏氨苄西林以及用于基因组整合的选择性抗生素的2YT琼脂平板上。将平板在42℃下孵育以愈合质粒的细胞。对氨苄西林和选择性抗生素敏感的克隆被用于进一步的实验和修饰。
基因slr1975(GenBank:BAL35720)编码集胞藻属种PCC6803 N-乙酰葡糖胺2-差向异构酶。基因gna1(GenBank:NP_116637)编码来自酿酒酵母的葡糖胺-6-磷酸乙酰转移酶。基因neuB(GenBank:AF305571)编码空肠弯曲杆菌的唾液酸合酶。基因glmS*是大肠杆菌L-谷氨酰胺:D-果糖-6-磷酸转氨酶基因的突变形式(Metab Eng.2005May;7(3):201-14)。基因ppsA(GenBank:ACT43527)编码大肠杆菌BL21(DE3)的磷酸烯醇丙酮酸合酶。
为了产生<Ptet-slr1975-gna1-lox66-aacC1-lox71>,将基因slr1975和gna1作为操纵子亚克隆到组成型启动子Ptet后面,并融合到庆大霉素抗性基因(侧翼为lox66/lox71位点),并通过平端连接插入pEcomar载体。使用载体pEcomar-slr1975-gna1-aacC1和水手转座酶Himar1的超活性C9-突变体将所得表达盒整合到基因组中,在阿拉伯糖诱导型启动子ParaB的控制下。
为了产生<Ptet-neuB-lox66-kanR-lox71>,将neuB克隆到组成型启动子Ptet后面,并融合到卡那霉素抗性基因(侧翼为lox66/lox71位点)。将得到的表达盒用EZ-Tn5转座酶整合到基因组中。为了产生<Ptet-slr1975-Pt5-neuB-FRT-dhfr-FRT>,将基因slr1975和neuB分别亚克隆到组成型启动子Ptet和Pt5后面,并融合到甲氧苄氨嘧啶(trimethoprim)抗性基因(侧翼为FRT位点)。将得到的表达盒用EZ-Tn5转座酶整合到基因组中。
通过将glmS*和gna1作为操纵子克隆到组成型启动子Ptet后面,产生表达盒<Ptet-glmS*-gna1-lox66-aacC1-lox71>。该构建体进一步融合到庆大霉素抗性基因(侧翼为lox66/lox71位点)。将得到的表达盒用EZ-Tn5转座酶整合到基因组中。
为了产生<Ptet-ppsA-lox66-aacC1-lox71>,将ppsA基因克隆到组成型启动子Ptet后面,并融合到庆大霉素抗性基因(侧翼为lox66/lox71位点)。将得到的表达盒用EZ-Tn5转座酶整合到基因组中。
累积的基因组修饰一起产生了Neu5Ac生产菌株大肠杆菌#NANA1。
实施例2:在分批发酵过程中生产N-乙酰神经氨酸
大肠杆菌BL21(DE3)菌株#NANA1在30℃下,在3L发酵罐(New Brunswick,Edison,USA)中培养,开始时使用1000mL矿物盐培养基,该培养基含有7g·L-1NH4H2PO4、7g·L- 1K2HPO4、2g·L-1KOH、0.3g·L-1柠檬酸、2g·L-1MgSO4x7·H2O、5g·L-1NH4Cl2和0.015g·L- 1CaCl2x6·H2O,补充1mL·L-1微量元素溶液(54.4g·L-1柠檬酸铁铵、9.8g·L-1MnCl2×4·H2O、1.6g·L-1CoCl2x6·H2O、1g·L-1CuCl2x2·H2O、1.9g·L-1H3BO3、9g·L-1ZnSO4x7·H2O、1.1g·L-1Na2MoO4x2·H2O、1.5g·L-1Na2SeO3、1.5·L-1NiSO4x6·H2O),并含有2%(m/v)蔗糖作为碳源以及抗生素博莱霉素(10μg·mL-1)。使用来自在相同的含蔗糖培养基中生长的预培养物的2.5%(v/v)接种物,开始培养。分批阶段结束的特征是溶氧水平上升。离开分批阶段后立即应用蔗糖进料。向50%(m/v)蔗糖进料中补充2g·L-1MgSO4x7·H2O、0.015g·L- 1CaCl2x6·H2O和1mL·L-1微量元素溶液。采用的补料速率为9.0至11.0mL·L-1,参照起始体积。充气维持在3L·min-1。通过控制搅拌速率,将溶氧保持在20-30%的饱和度。加入25%氨溶液使pH保持在7.0。
采用高效液相色谱(HPLC,Shimadzu)检测培养上清液中的Neu5Ac。该设备包含UV-VIS探测器,λ=210nm(SPD-10AVP,Shimadzu)和带有适当保护筒(guard cartidge)的RezexROA-有机酸H+分析柱(300x7.8mm)。在5mM H2SO4中,在50℃下进行等度洗脱,流速为0.5ml·min-1。将培养上清液离心,过滤灭菌,在95℃下加热5min。最后的离心之后,将5μL样品应用于柱上。使用商用标准(Carbosynth,Compton,UK),从标准曲线计算N-乙酰神经氨酸的浓度。孵育88h后,培养上清液中最终的Neu5Ac滴度为68.6g·L-1
实施例3:显示出提高的Neu5Ac生产能力的大肠杆菌菌株#NANA1的单基因敲除突变体的产生与培养
对大肠杆菌BL21(DE3)菌株#NANA1进行了进一步的修饰,产生了去除或破坏基因gltB、yjhC和ppC的缺失突变体。在96孔板中培养了菌株#NANA1、#NANA1△gltB、#NANA1△yjhC和#NANA1△ppc。因此,将菌株的单个菌落从琼脂平板转移到含有200μL实施例2中所述基本培养基的微量滴定板中,并在30℃下剧烈震荡孵育约20h。随后,将50μL培养液转移到深孔96孔板(2.0mL)中,每孔含有400μL基本培养基。
再孵育48小时后,停止培养,通过使用LC三重四极MS检测系统、多重反应监测(MRM)模式的质谱测定上清液中N-乙酰神经氨酸的含量。在四极1中选择和分析前体离子,使用氩气作为碰撞气体在碰撞室中发生碎片化,在四极3中选择碎片离子。用LC/MS级水以1:100稀释培养物上清液后,将1μl N-乙酰神经氨酸样品注入HPLC仪器。样品在具有Xbridge Amide保护筒(3.5μm,2.1×10mm)(Waters,USA)的XBridge Amide HPLC柱(3.5μm,2.1×50mm)(Waters,USA)上,在50℃下,在乙腈:H2O中,用10mM乙酸铵分离,流速为400μl.min-1。每次分离持续240秒。在电喷雾电离(ESI)正电离模式下,通过MRM分析了N-乙酰神经氨酸。质谱仪以单位分辨率操作。N-乙酰神经氨酸形成m/z309.2[M+H]的离子。将N-乙酰神经氨酸的前体离子在碰撞室中进一步碎片化为碎片离子m/z 292.20、m/z 274.15和m/z121.15。分别单独对每种分析物的碰撞能量、Q1和Q3预偏置(Pre Bias)进行优化。采用商用标准(Carbosynth,Compton,UK)建立了定量方法。
图3示出了培养菌株的相对Neu5Ac产量。与亲本菌株相比,菌株#NANA1的值设置为100%,单基因敲除突变体产生的N-乙酰神经氨酸多20%至25%。
实施例4:从发酵液中纯化Neu5Ac
发酵结束后,通过超滤,然后使用连续缠绕模块过滤器(截留值0.05μm)(CUT膜技术,Erkrath,Germany)和错流过滤器(截留值150kDa)(Microdyn-Nadir,Wiesbaden,Germany),将生物质与发酵培养基分离。获得了约1m3无细胞发酵培养基,其中含有超过19g·L-1唾液酸。
然后用离子交换色谱法对无细胞液体进行去离子化。首先,在一个体积为200L的H+形式的强阳离子交换剂(
Figure BDA0002543398380000351
S 2568(Lanxess AG,Cologne,Germany)上去除阳离子污染物。用NaOH将得到的pH约为1.5的溶液中和到7.0。在第二步中,用氯化物形式的强阴离子交换剂
Figure BDA0002543398380000352
S 6368 A(Lanxess AG,Cologne,Germany)从溶液中去除阴离子和不需要的色素。离子交换剂的床体积为200L。在错流过滤器(截留值150kDa)(Microdyn-Nadir GmbH,Wiesbaden,Germany)上使用第二个过滤步骤,去除源自酸化溶液的沉淀物。为了浓缩糖,将溶液在Dow
Figure BDA0002543398380000361
NF270-4040(INAQUA Vertriebsgesellschaft mbH,
Figure BDA0002543398380000362
Germany)上纳滤至体积的约1/4。然后将浓缩的Neu5Ac溶液在旋转蒸发器上进一步浓缩至浓度约为400g L-1或更高。在5℃下,用10倍过量的冰醋酸进行产物的特异性结晶,持续12-60小时。过滤固体级分,用乙醇洗涤,在40℃下干燥。将干燥的结晶产物进一步纯化。因此,每kg干燥产物溶于2L H2O中,用活性炭(CAS-No:7440-44-0,CarlRoth GmbH&Co.KG,Karlsruhe,Germany)处理。与活性炭分离后,将澄清溶液在50℃下蒸发浓缩,直到其凝固。将固体材料与99%乙醇混合,并在4℃下孵育至少16h。然后将固体级分过滤并在40℃下干燥。获得结晶白色产物的纯度大于95%,利用通过使用Rezex ROA-有机酸H+柱(Phenomenex,Aschaf fen burg,Germany)的HPLC测定的色谱图的曲线下面积测得。
实施例5:N-乙酰神经氨酸生产的替代途径
通过整合表达盒<Ptet-glmS*-gna1-lox66-aacC1-lox71>进一步修饰实施例1中描述的大肠杆菌BL21(DE3)菌株(ΔlacZ、ΔaraA、ΔnagABE、ΔnanATEK、ΔmanXYZ、ΔwcaJ、ΔfuclK、ΔagaA、lacY+、cscABKR+),得到能够合成N-乙酰葡糖胺的菌株(菌株A)。对菌株A进行修饰以生成用于生产N-乙酰神经氨酸的菌株。为此,将表达构建体<Ptet-slr1975-Pt5-neuB-FRT-dhfr-FRT>(SEQ ID NO:5)或<Ptet-EcnanE-Pt5-neuB-FRT-dhfr-FRT>(SEQ ID NO:9)分别整合到菌株A的基因组中,分别产生菌株B和C。EcnanE基因(GenBank:.YP_003614592)编码阴沟肠杆菌阴沟亚种ATCC 13047的N-酰基葡糖胺-6-磷酸2-差向异构酶。使用EZ-Tn5转座酶将所有表达盒整合到基因组中。
将这些菌株的单个菌落从琼脂平板转移到含有200μL实施例2中所述基本培养基的微量滴定板中,并在30℃下剧烈震荡培养约20h。随后,将50μL培养液转移到深孔96孔板(2.0mL)中,每孔含有400μL基本培养基。再孵育48小时后,停止培养,用质谱法测定上清液中N-乙酰神经氨酸水平。
仅在菌株B和C的培养上清液中可检测到Neu5Ac产生。在图4中,示出了培养菌株的相对Neu5Ac产量。在比较时,将菌株B的Neu5Ac产值设为100%。与菌株B相比,菌株C产生约7.5%的Neu5Ac。
实施例6:含有Neu5Ac的婴儿配方物的组成
婴儿配方物:脱脂乳
植物油(棕榈油、菜籽油、葵花籽油)
人乳低聚糖
L-岩藻糖
N-乙酰神经氨酸
脱脂奶粉
高山被孢霉(Mortierella alpine)的油
鱼油
碳酸钙
氯化钾
维生素C
氯化钠
维生素E
醋酸亚铁
硫酸锌
烟酸
D-泛酸钙
硫酸铜
维生素A
维生素B1
维生素B6
硫酸镁
碘酸钾
叶酸
维生素K
亚硒酸钠
维生素D
序列表
<110> Jennewein Biotechnologie GmbH
<120> N-乙酰神经氨酸的发酵生产
<130> P 1705 WO
<150> EP 17196925.6
<151> 2017-10-17
<160> 103
<170> PatentIn version 3.5
<210> 1
<211> 2851
<212> DNA
<213> 人工序列
<220>
<223> 表达片段
<400> 1
tggccagatg attaattcct aatttttgtt gacactctat cattgataga gttattttac 60
cactccctat cagtgataga gaaaagtgaa atgaatagtt cgacaaaaat ctagaaataa 120
ttttgtttaa ctttaagaag gagatataca aatgtactat ttaaaaaaca caaacttttg 180
gatgttcggt ttattctttt tcttttactt ttttatcatg ggagcctact tcccgttttt 240
cccgatttgg ctacatgaca tcaaccatat cagcaaaagt gatacgggta ttatttttgc 300
cgctatttct ctgttctcgc tattattcca accgctgttt ggtctgcttt ctgacaaact 360
cgggctgcgc aaatacctgc tgtggattat taccggcatg ttagtgatgt ttgcgccgtt 420
ctttattttt atcttcgggc cactgttaca atacaacatt ttagtaggat cgattgttgg 480
tggtatttat ctaggctttt gttttaacgc cggtgcgcca gcagtagagg catttattga 540
gaaagtcagc cgtcgcagta atttcgaatt tggtcgcgcg cggatgtttg gctgtgttgg 600
ctgggcgctg tgtgcctcga ttgtcggcat catgttcacc atcaataatc agtttgtttt 660
ctggctgggc tctggctgtg cactcatcct cgccgtttta ctctttttcg ccaaaacgga 720
tgcgccctct tctgccacgg ttgccaatgc ggtaggtgcc aaccattcgg catttagcct 780
taagctggca ctggaactgt tcagacagcc aaaactgtgg tttttgtcac tgtatgttat 840
tggcgtttcc tgcacctacg atgtttttga ccaacagttt gctaatttct ttacttcgtt 900
ctttgctacc ggtgaacagg gtacgcgggt atttggctac gtaacgacaa tgggcgaatt 960
acttaacgcc tcgattatgt tctttgcgcc actgatcatt aatcgcatcg gtgggaaaaa 1020
cgccctgctg ctggctggca ctattatgtc tgtacgtatt attggctcat cgttcgccac 1080
ctcagcgctg gaagtggtta ttctgaaaac gctgcatatg tttgaagtac cgttcctgct 1140
ggtgggctgc tttaaatata ttaccagcca gtttgaagtg cgtttttcag cgacgattta 1200
tctggtctgt ttctgcttct ttaagcaact ggcgatgatt tttatgtctg tactggcggg 1260
caatatgtat gaaagcatcg gtttccaggg cgcttatctg gtgctgggtc tggtggcgct 1320
gggcttcacc ttaatttccg tgttcacgct tagcggcccc ggcccgcttt ccctgctgcg 1380
tcgtcaggtg aatgaagtcg ctgggagcta agcggccgcg tcgacacgca aaaaggccat 1440
ccgtcaggat ggccttctgc ttaatttgat gcctggcagt ttatggcggg cgtcctgccc 1500
gccaccctcc gggccgttgc ttcgcaacgt tcaaatccgc tcccggcgga tttgtcctac 1560
tcaggagagc gttcaccgac aaacaacaga taaaacgaaa ggcccagtct ttcgactgag 1620
cctttcgttt tatttgatgc ctggcagttc cctactctcg catggggaga ccccacacta 1680
ccatcatgta tgaatatcct ccttagttcc tattccgaag ttcctattct ctagaaagta 1740
taggaacttc ggcgcgtcct acctgtgaca cgcgtgccgc agtctcacgc ccggagcgta 1800
gcgaccgagt gagctagcta tttgtttatt tttctaaata cattcaaata tgtatccgct 1860
catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgaggga 1920
agcggtgatc gccgaagtat cgactcaact atcagaggta gttggcgtca tcgagcgcca 1980
tctcgaaccg acgttgctgg ccgtacattt gtacggctcc gcagtggatg gcggcctgaa 2040
gccacacagt gatattgatt tgctggttac ggtgaccgta aggcttgatg aaacaacgcg 2100
gcgagctttg atcaacgacc ttttggaaac ttcggcttcc cctggagaga gcgagattct 2160
ccgcgctgta gaagtcacca ttgttgtgca cgacgacatc attccgtggc gttatccagc 2220
taagcgcgaa ctgcaatttg gagaatggca gcgcaatgac attcttgcag gtatcttcga 2280
gccagccacg atcgacattg atctggctat cttgctgaca aaagcaagag aacatagcgt 2340
tgccttggta ggtccagcgg cggaggaact ctttgatccg gttcctgaac aggatctatt 2400
tgaggcgcta aatgaaacct taacgctatg gaactcgccg cccgactggg ctggcgatga 2460
gcgaaatgta gtgcttacgt tgtcccgcat ttggtacagc gcagtaaccg gcaaaatcgc 2520
gccgaaggat gtcgctgccg actgggcaat ggagcgcctg ccggcccagt atcagcccgt 2580
catacttgaa gctagacagg cttatcttgg acaagaagaa gatcgcttgg cctcgcgcgc 2640
agatcagttg gaagaatttg tccactacgt gaaaggcgag atcaccaagg tagtcggcaa 2700
ataatgtcta acaattcgtt caagccgagg ggccgcaaga tccggccacg atgacccggt 2760
cgtcgggtac cggcagggcg gggcgtaagg cgcgccattt aaatgaagtt cctattccga 2820
agttcctatt ctctagaaag tataggaact t 2851
<210> 2
<211> 5226
<212> DNA
<213> 人工序列
<220>
<223> 大肠杆菌W csc基因簇,包括蔗糖通透酶(cscB)、果糖激酶(cscK)、蔗糖水解酶(cscA)和转录阻遏因子(cscR)的基因
<400> 2
acaggttggc tgataagtcc ccggtctggc agccgcgact gtaccagaac atgaatgagg 60
cgtttggatt aggcgattat tagcagggct aagcatttta ctattattat tttccggttg 120
agggatatag agctatcgac aacaaccgga aaaagtttac gtctatattg ctgaaggtac 180
aggcgtttcc ataactattt gctcgcgttt tttactcaag aagaaaatgc caaatagcaa 240
catcaggcag acaatacccg aaattgcgaa gaaaactgtc tggtagcctg cgtggtcaaa 300
gagtatccca gtcggcgttg aaagcagcac aatcccaagc gaactggcaa tttgaaaacc 360
aatcagaaag atcgtcgacg acaggcgctt atcaaagttt gccacgctgt atttgaagac 420
ggatatgaca caaagtggaa cctcaatggc atgtaacaac ttcactaatg aaataatcca 480
ggggttaacg aacagcgcgc aggaaaggat acgcaacgcc ataatcacaa ctccgataag 540
taatgcattt tttggcccta cccgattcac aaagaaagga ataatcgcca tgcacagcgc 600
ttcgagtacc acctggaatg agttgagata accatacagg cgcgttccta catcgtgtga 660
ttcgaataaa cctgaataaa agacaggaaa aagttgttga tcaaaaatgt tatagaaaga 720
ccacgtcccc acaataaata tgacgaaaac ccagaagttt cgatccttga aaactgcgat 780
aaaatcctct ttttttaccc ctcccgcatc tgccgctacg cactggtgat ccttatcttt 840
aaaacgcatg ttgatcatca taaatacagc gccaaatagc gagaccaacc agaagttgat 900
atggggactg atactaaaaa atatgccggc aaagaacgcg ccaatagcat agccaaaaga 960
tccccaggcg cgcgctgttc catattcgaa atgaaaattt cgcgccattt tttcggtgaa 1020
gctatcaagc aaaccgcatc ccgccagata ccccaagcca aaaaatagcg cccccagaat 1080
tagacctaca gaaaaattgc tttgcagtaa cggttcataa acgtaaatca taaacggtcc 1140
ggtcaagacc aggatgaaac tcatacacca gatgagcggt ttcttcagac cgagtttatc 1200
ctgaacgatg ccgtagaaca tcataaatag aatgctggta aactggttga ccgaataaag 1260
tgtacctaat tccgtccctg tcaaccctag atgtcctttc agccaaatag cgtataacga 1320
ccaccacagc gaccaggaaa taaaaaagag aaatgagtaa ctggatgcaa aacgatagta 1380
cgcatttctg aatggaatat tcagtgccat aattacctgc ctgtcgttaa aaaattcacg 1440
tcctatttag agataagagc gacttcgccg tttacttctc actattccag ttcttgtcga 1500
catggcagcg ctgtcattgc ccctttcgcc gttactgcaa gcgctccgca acgttgagcg 1560
agatcgataa ttcgtcgcat ttctctctca tctgtagata atcccgtaga ggacagacct 1620
gtgagtaacc cggcaacgaa cgcatctccc gcccccgtgc tatcgacaca attcacagac 1680
attccagcaa aatggtgaac ttgtcctcga taacagacca ccaccccttc tgcaccttta 1740
gtcaccaaca gcatggcgat ctcatactct tttgccaggg cgcatatatc ctgatcgttc 1800
tgtgtttttc cactgataag tcgccattct tcttccgaga gcttgacgac atccgccagt 1860
tgtagcgcct gccgcaaaca caagcggagc aaatgctcgt cttgccatag atcttcacga 1920
atattaggat cgaagctgac aaaacctccg gcatgccgga tcgccgtcat cgcagtaaat 1980
gcgctggtac gcgaaggctc ggcagacaac gcaattgaac agagatgtaa ccattcgcca 2040
tgtcgccagc agggcaagtc tgtcgtctct aaaaaaagat cggcactggg gcggaccata 2100
aacgtaaatg aacgttcccc ttgatcgttc agatcgacaa gcaccgtgga tgtccggtgc 2160
cattcatctt gcttcagata cgtgatatcg actccctcag ttagcagcgt tctttgcatt 2220
aacgcaccaa aaggatcatc ccccacccga cctataaacc cacttgttcc gcctaatctg 2280
gcgattccca ccgcaacgtt agctggcgcg ccgccaggac aaggcagtag gcgcccgtct 2340
gattctggca agagatctac gaccgcatcc cctaaaaccc atactttggc tgacattttt 2400
ttcccttaaa ttcatctgag ttacgcatag tgataaacct ctttttcgca aaatcgtcat 2460
ggatttacta aaacatgcat attcgatcac aaaacgtcat agttaacgtt aacatttgtg 2520
atattcatcg catttatgaa agtaagggac tttattttta taaaagttaa cgttaacaat 2580
tcaccaaatt tgcttaacca ggatgattaa aatgacgcaa tctcgattgc atgcggcgca 2640
aaacgcccta gcaaaacttc atgagcaccg gggtaacact ttctatcccc attttcacct 2700
cgcgcctcct gccgggtgga tgaacgatcc aaacggcctg atctggttta acgatcgtta 2760
tcacgcgttt tatcaacatc atccgatgag cgaacactgg gggccaatgc actggggaca 2820
tgccaccagc gacgatatga tccactggca gcatgagcct attgcgctag cgccaggaga 2880
cgataatgac aaagacgggt gtttttcagg tagtgctgtc gatgacaatg gtgtcctctc 2940
acttatctac accggacacg tctggctcga tggtgcaggt aatgacgatg caattcgcga 3000
agtacaatgt ctggctacca gtcgggatgg tattcatttc gagaaacagg gtgtgatcct 3060
cactccacca gaaggaatca tgcacttccg cgatcctaaa gtgtggcgtg aagccgacac 3120
atggtggatg gtagtcgggg cgaaagatcc aggcaacacg gggcagatcc tgctttatcg 3180
cggcagttcg ttgcgtgaat ggaccttcga tcgcgtactg gcccacgctg atgcgggtga 3240
aagctatatg tgggaatgtc cggacttttt cagccttggc gatcagcatt atctgatgtt 3300
ttccccgcag ggaatgaatg ccgagggata cagttaccga aatcgctttc aaagtggcgt 3360
aatacccgga atgtggtcgc caggacgact ttttgcacaa tccgggcatt ttactgaact 3420
tgataacggg catgactttt atgcaccaca aagcttttta gcgaaggatg gtcggcgtat 3480
tgttatcggc tggatggata tgtgggaatc gccaatgccc tcaaaacgtg aaggatgggc 3540
aggctgcatg acgctggcgc gcgagctatc agagagcaat ggcaaacttc tacaacgccc 3600
ggtacacgaa gctgagtcgt tacgccagca gcatcaatct gtctctcccc gcacaatcag 3660
caataaatat gttttgcagg aaaacgcgca agcagttgag attcagttgc agtgggcgct 3720
gaagaacagt gatgccgaac attacggatt acagctcggc actggaatgc ggctgtatat 3780
tgataaccaa tctgagcgac ttgttttgtg gcggtattac ccacacgaga atttagacgg 3840
ctaccgtagt attcccctcc cgcagcgtga cacgctcgcc ctaaggatat ttatcgatac 3900
atcatccgtg gaagtattta ttaacgacgg ggaagcggtg atgagtagtc gaatctatcc 3960
gcagccagaa gaacgggaac tgtcgcttta tgcctcccac ggagtggctg tgctgcaaca 4020
tggagcactc tggctactgg gttaacataa tatcaggtgg aacaacggat caacagcggg 4080
caagggatcc gcgtcactct tcccccttca cgaccttcaa taatatgcaa tgcagcttcc 4140
cgcccgataa tgtcatgtgg aagctgaatt gtggtcagcg gcggtaaaaa cagatgcccg 4200
acgccaacca gattatcaaa gcccattacg gcgacatcct gcgggattcg tacccccttc 4260
gccagaagaa cctgataagc cacaaaggct gcgcgatcgt taccacatat cagaacatca 4320
aaatctggtt tgcccggttt gaagtgggca ttgagtaaac ttgcgagatc ggtgtagtga 4380
tcatcacctg ttgccatgtg aaattgtttc acctcagcca gatctcgtcc agcatcacgc 4440
caggcctgct caaatccctg ccgacgatac cctgttgcca acgcactttc cggtagccag 4500
aagcataacg gttgacgata gcccgccgcg agcaaatgct gtgttgattc atattgtgca 4560
gtgtaatcat cagggatata actgggtaac gctgggtcat ccgccacaca gttcgccaat 4620
acaatatttt caccatacag agactcaggc agcgtgatat gtcgcagccc cattgtagta 4680
tagataatgc catccggacg gtgggcaagc agctgacgtg ccgcgcgggc agcgtcatct 4740
tcagaaaaaa tattgattaa aaaactattc cagccgaact cgctggcggt ttgctcaatg 4800
gcaagcagaa tatcaacaga gaaaggagtg gtagccgtgt cctgcgccag cacggcgaga 4860
gtcgacggct tacgtccttg agcgcgcatc ttacgggcgg aaagatcagg aacataattc 4920
agggtctgga ttgcctgcaa tacgcggtca cgcgttgcag gacgcacaga ttctgcatta 4980
tgcatcaccc gggagactgt catcatcgac actcccgcca ggcgtgcgac atcctttaat 5040
gaagccatac ccaagccgtt tgccgtaaaa cgggcactgt agcagaaaca gacgtcactg 5100
gcgagatcca acgccctatc acctgacaca gcaatacaat aaaaaataac aataattccc 5160
ggacaattgt ccccaattcc gcctctgttc tcgcattgta gaccggggac ttatcagcca 5220
acctgt 5226
<210> 3
<211> 3123
<212> DNA
<213> 人工序列
<220>
<223> 表达盒
<400> 3
ctgtctctta tacacatctc cggccagatg attaattcct aatttttgtt gacactctat 60
cattgataga gttattttac cactccctat cagtgataga gaaaagtgaa atgaatagtt 120
cgacaaaaat ctagaaataa ttttgtttaa ctttaagaag gagatataca aatgatcgct 180
caccgtcgtc aggaactggc tcaacagtat tatcaggctc tgcaccaaga tgtgctgccg 240
ttctgggaaa agtattcgct ggatcgtcaa ggcggtggct attttacctg cctggaccgc 300
aagggtcagg tttttgatac ggacaagttc atttggctgc aaaaccgtca agtgtggcaa 360
tttgcggttt tctacaatcg cctggaaccg aaaccgcagt ggctggaaat cgctcgtcat 420
ggtgcggatt ttctggcacg tcacggtcgt gatcaggacg gtaactggta tttcgccctg 480
gatcaggaag gcaaaccgct gcgccaaccg tacaatgtgt tttccgactg tttcgcggcg 540
atggcgttta gccagtatgc actggcttct ggtgctcaag aagcgaaggc cattgcactg 600
caagcgtata acaatgttct gcgtcgccag cataacccga aaggtcaata tgaaaagagt 660
tacccgggta cccgtccgct gaaatccctg gcagtgccga tgatcctggc taatctgacg 720
ctggaaatgg aatggctgct gccgccgacc acggtcgaag aagtgctggc ccagaccgtt 780
cgtgaagtca tgacggattt tctggacccg gaaattggcc tgatgcgcga agcagttacc 840
ccgacgggtg aatttgtcga ttcattcgaa ggccgcctgc tgaacccggg tcatggcatt 900
gaagcgatgt ggtttatgat ggatattgcc cagcgttcgg gtgaccgcca gctgcaagaa 960
caggctattg cggtggttct gaataccctg gaatatgcat gggatgaaga atttggtggc 1020
atcttttact tcctggaccg tcaaggtcac ccgccgcagc aactggaatg ggatcagaaa 1080
ctgtggtggg tccatctgga aaccctggtg gccctggcaa aaggtcacca ggcgacgggc 1140
caagaaaagt gctggcagtg gtttgaacgc gtgcatgatt atgcatggag ccactttgct 1200
gacccggaat atggtgaatg gttcggctac ctgaaccgtc gcggtgaagt gctgctgaat 1260
ctgaaaggtg gcaaatggaa gggctgcttc cacgttccgc gtgcgctgtg gctgtgtgcc 1320
gaaaccctgc aactgccggt ctcttaataa tcgaaggaga tacaacatga gcttacccga 1380
tggattttat ataaggcgaa tggaagaggg ggatttggaa caggtcactg agacgctaaa 1440
ggttttgacc accgtgggca ctattacccc cgaatccttc agcaaactca taaaatactg 1500
gaatgaagcc acagtatgga atgataacga agataaaaaa ataatgcaat ataaccccat 1560
ggtgattgtg gacaagcgca ccgagacggt tgccgctacg gggaatatca tcatcgaaag 1620
aaagatcatt catgaactgg ggctatgtgg ccacatcgag gacattgcag taaactccaa 1680
gtatcagggc caaggtttgg gcaagctctt gattgatcaa ttggtaacta tcggctttga 1740
ctacggttgt tataagatta ttttagattg cgatgagaaa aatgtcaaat tctatgaaaa 1800
atgtgggttt agcaacgcag gcgtggaaat gcaaattaga aaatagaata actagcataa 1860
acccccttgg ggcctctaaa cgggtcttga ggggtttttt gctgaaacca atttgcctgg 1920
cggcagtagc gcggtggtcc cacctgaccc catgccgaac tcagaagtga aacgccgtag 1980
cgccgatggt agtgtggggt ctccccatgc gagagtaggg aactgccagg catcaaataa 2040
aacgaaaggc tcagtcgaaa gactgggcct ttcgggatcc aggccggcct gttaagacgg 2100
ccagtgaatt cgagctcggt acctaccgtt cgtataatgt atgctatacg aagttatcga 2160
gctctagaga atgatcccct cattaggcca cacgttcaag tgcagcgcac accgtggaaa 2220
cggatgaagg cacgaaccca gttgacataa gcctgttcgg ttcgtaaact gtaatgcaag 2280
tagcgtatgc gctcacgcaa ctggtccaga accttgaccg aacgcagcgg tggtaacggc 2340
gcagtggcgg ttttcatggc ttgttatgac tgtttttttg tacagtctat gcctcgggca 2400
tccaagcagc aagcgcgtta cgccgtgggt cgatgtttga tgttatggag cagcaacgat 2460
gttacgcagc agcaacgatg ttacgcagca gggcagtcgc cctaaaacaa agttaggtgg 2520
ctcaagtatg ggcatcattc gcacatgtag gctcggccct gaccaagtca aatccatgcg 2580
ggctgctctt gatcttttcg gtcgtgagtt cggagacgta gccacctact cccaacatca 2640
gccggactcc gattacctcg ggaacttgct ccgtagtaag acattcatcg cgcttgctgc 2700
cttcgaccaa gaagcggttg ttggcgctct cgcggcttac gttctgccca ggtttgagca 2760
gccgcgtagt gagatctata tctatgatct cgcagtctcc ggcgagcacc ggaggcaggg 2820
cattgccacc gcgctcatca atctcctcaa gcatgaggcc aacgcgcttg gtgcttatgt 2880
gatctacgtg caagcagatt acggtgacga tcccgcagtg gctctctata caaagttggg 2940
catacgggaa gaagtgatgc actttgatat cgacccaagt accgccacct aacaattcgt 3000
tcaagccgag atcgtagaat ttcgacgacc tgcagccaag cataacttcg tataatgtat 3060
gctatacgaa cggtaggatc ctctagagtc gacctgcagg catgagatgt gtataagaga 3120
cag 3123
<210> 4
<211> 2965
<212> DNA
<213> 人工序列
<220>
<223> 表达盒
<400> 4
ctgtctctta tacacatctc cggccagatg attaattcct aatttttgtt gacactctat 60
cattgataga gttattttac cactccctat cagtgataga gaaaagtgaa atgaatagtt 120
cgacaaaaat ctagaaataa ttttgtttaa ctttaagaag gagatataca aatgaaagaa 180
atcaaaatcc agaacatcat catcagcgaa gaaaaagcgc cgctggttgt gccggaaatc 240
ggcattaacc ataatggtag tctggaactg gcaaaaatca tggtggatgc ggcctttagc 300
gccggtgcaa aaatcattaa acatcagacc cacattgtgg aagatgaaat gtctaaagca 360
gcgaaaaaag ttatcccggg caacgcgaaa atcagtatct acgaaatcat gcagaaatgc 420
gcgctggatt acaaagatga actggccctg aaagaatata ccgaaaaact gggtctggtg 480
tacctgtcta ccccgtttag tcgtgcgggt gcaaaccgtc tggaagatat gggtgttagt 540
gcgttcaaaa tcggcagcgg tgaatgtaac aattatccgc tgatcaaaca tattgccgca 600
tttaaaaaac cgatgattgt tagcaccggc atgaatagca tcgaatctat taaaccgacg 660
gtgaaaatcc tgctggataa cgaaattccg tttgttctga tgcataccac gaatctgtac 720
ccgaccccgc acaacctggt gcgtctgaat gccatgctgg aactgaaaaa agaattctct 780
tgcatggttg gtctgagtga tcacaccacg gataatctgg catgcctggg tgcagtggtt 840
ctgggtgcgt gtgtgctgga acgtcatttc accgatagca tgcaccgctc tggtccggat 900
attgtttgta gtatggatac gaaagcactg aaagaactga tcattcagag cgaacagatg 960
gcgatcattc gcggcaacaa tgaatctaaa aaagcggcca aacaggaaca ggtgaccatc 1020
gattttgcat tcgcgagtgt ggttagcatc aaagatatca aaaaaggcga agtgctgagc 1080
atggataata tttgggttaa acgtccgggt ctgggcggta tctctgcagc ggaatttgaa 1140
aacattctgg gcaaaaaagc actgcgcgat attgaaaatg atgcgcagct gtcttatgaa 1200
gatttcgcct aataaatcga tactagcata accccttggg gcctctaaac gcgtcgacac 1260
gcaaaaaggc catccgtcag gatggccttc tgcttaattt gatgcctggc agtttatggc 1320
gggcgtcctg cccgccaccc tccgggccgt tgcttcgcaa cgttcaaatc cgctcccggc 1380
ggatttgtcc tactcaggag agcgttcacc gacaaacaac agataaaacg aaaggcccag 1440
tctttcgact gagcctttcg ttttatttga tgcctggcag ttccctactc tcgcatgggg 1500
agaccccaca ctaccatccg gtatcgataa gcttgatggc gaaaggggga tgtgctgcaa 1560
ggcgattaag ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca 1620
gtgaattcga gctcggtacc taccgttcgt ataatgtatg ctatacgaag ttatcgagct 1680
ctagagaatg atcccctccc tcacgctgcc gcaagcactc agggcgcaag ggctgctaaa 1740
ggaagcggaa cacgtagaaa gccagtccgc agaaacggtg ctgaccccgg atgaatgtca 1800
gctactgggc tatctggaca agggaaaacg caagcgcaaa gagaaagcag gtagcttgca 1860
gtgggcttac atggcgatag ctagactggg cggttttatg gacagcaagc gaaccggaat 1920
tgccagctgg ggcgccctct ggtaaggttg ggaagccctg caaagtaaac tggatggctt 1980
tcttgccgcc aaggatctga tggcgcaggg gatcaagatc tgatcaagag acaggatgag 2040
gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg 2100
agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt 2160
tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc 2220
tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt 2280
gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag 2340
tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg 2400
ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag 2460
cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg 2520
atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc 2580
gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca 2640
tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc 2700
gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg 2760
ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct 2820
atcgccttct tgacgagttc ttctgagcgg gactctggga atttcgacga cctgcagcca 2880
agcataactt cgtataatgt atgctatacg aacggtagga tcctctagag tcgacctgca 2940
ggcatgagat gtgtataaga gacag 2965
<210> 5
<211> 3904
<212> DNA
<213> 人工序列
<220>
<223> 表达盒
<400> 5
ctgtctctta tacacatctc cggccagatg attaattcct aatttttgtt gacactctat 60
cattgataga gttattttac cactccctat cagtgataga gaaaagtgaa atgaatagtt 120
cgacaaaaat ctagaaataa ttttgtttaa ctttaagaag gagatataca aatgatcgct 180
caccgtcgtc aggaactggc tcaacagtat tatcaggctc tgcaccaaga tgtgctgccg 240
ttctgggaaa agtattcgct ggatcgtcaa ggcggtggct attttacctg cctggaccgc 300
aagggtcagg tttttgatac ggacaagttc atttggctgc aaaaccgtca agtgtggcaa 360
tttgcggttt tctacaatcg cctggaaccg aaaccgcagt ggctggaaat cgctcgtcat 420
ggtgcggatt ttctggcacg tcacggtcgt gatcaggacg gtaactggta tttcgccctg 480
gatcaggaag gcaaaccgct gcgccaaccg tacaatgtgt tttccgactg tttcgcggcg 540
atggcgttta gccagtatgc actggcttct ggtgctcaag aagcgaaggc cattgcactg 600
caagcgtata acaatgttct gcgtcgccag cataacccga aaggtcaata tgaaaagagt 660
tacccgggta cccgtccgct gaaatccctg gcagtgccga tgatcctggc taatctgacg 720
ctggaaatgg aatggctgct gccgccgacc acggtcgaag aagtgctggc ccagaccgtt 780
cgtgaagtca tgacggattt tctggacccg gaaattggcc tgatgcgcga agcagttacc 840
ccgacgggtg aatttgtcga ttcattcgaa ggccgcctgc tgaacccggg tcatggcatt 900
gaagcgatgt ggtttatgat ggatattgcc cagcgttcgg gtgaccgcca gctgcaagaa 960
caggctattg cggtggttct gaataccctg gaatatgcat gggatgaaga atttggtggc 1020
atcttttact tcctggaccg tcaaggtcac ccgccgcagc aactggaatg ggatcagaaa 1080
ctgtggtggg tccatctgga aaccctggtg gccctggcaa aaggtcacca ggcgacgggc 1140
caagaaaagt gctggcagtg gtttgaacgc gtgcatgatt atgcatggag ccactttgct 1200
gacccggaat atggtgaatg gttcggctac ctgaaccgtc gcggtgaagt gctgctgaat 1260
ctgaaaggtg gcaaatggaa gggctgcttc cacgttccgc gtgcgctgtg gctgtgtgcc 1320
gaaaccctgc aactgccggt ctcttaattt cgtcgacaca caggaaacat attaaaaatt 1380
aaaacctgca ggagtttaaa cgcggccgcg atatcgttgt aaaacgacgg ccagtgcaag 1440
aatcataaaa aatttatttg ctttcaggaa aatttttctg tataatagat tcataaattt 1500
gagagaggag tttttgtgag cggataacaa ttccccatct tagtatatta gttaagtata 1560
aatacacaag gagatataca tatgaaagaa atcaaaatcc agaacatcat catcagcgaa 1620
gaaaaagcgc cgctggttgt gccggaaatc ggcattaacc ataatggtag tctggaactg 1680
gcaaaaatca tggtggatgc ggcctttagc gccggtgcaa aaatcattaa acatcagacc 1740
cacattgtgg aagatgaaat gtctaaagca gcgaaaaaag ttatcccggg caacgcgaaa 1800
atcagtatct acgaaatcat gcagaaatgc gcgctggatt acaaagatga actggccctg 1860
aaagaatata ccgaaaaact gggtctggtg tacctgtcta ccccgtttag tcgtgcgggt 1920
gcaaaccgtc tggaagatat gggtgttagt gcgttcaaaa tcggcagcgg tgaatgtaac 1980
aattatccgc tgatcaaaca tattgccgca tttaaaaaac cgatgattgt tagcaccggc 2040
atgaatagca tcgaatctat taaaccgacg gtgaaaatcc tgctggataa cgaaattccg 2100
tttgttctga tgcataccac gaatctgtac ccgaccccgc acaacctggt gcgtctgaat 2160
gccatgctgg aactgaaaaa agaattctct tgcatggttg gtctgagtga tcacaccacg 2220
gataatctgg catgcctggg tgcagtggtt ctgggtgcgt gtgtgctgga acgtcatttc 2280
accgatagca tgcaccgctc tggtccggat attgtttgta gtatggatac gaaagcactg 2340
aaagaactga tcattcagag cgaacagatg gcgatcattc gcggcaacaa tgaatctaaa 2400
aaagcggcca aacaggaaca ggtgaccatc gattttgcat tcgcgagtgt ggttagcatc 2460
aaagatatca aaaaaggcga agtgctgagc atggataata tttgggttaa acgtccgggt 2520
ctgggcggta tctctgcagc ggaatttgaa aacattctgg gcaaaaaagc actgcgcgat 2580
attgaaaatg atgcgcagct gtcttatgaa gatttcgcct aaaataacta gcataacccc 2640
ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa accaatttgc ctggcggcag 2700
tagcgcggtg gtcccacctg accccatgcc gaactcagaa gtgaaacgcc gtagcgccga 2760
tggtagtgtg gggtctcccc atgcgagagt agggaactgc caggcatcaa ataaaacgaa 2820
aggctcagtc gaaagactgg gcctttcggg atccaggccg gcctgttaac gaattaatct 2880
tccgcggcgg tatcgataag cttgatatcg aattccgaag ttcctattct ctagaaagta 2940
taggaacttc aggtctgaag aggagtttac gtccagccaa gctagcttgg ctgcaggtcg 3000
tcgaaattct accgggtagg ggaggcgctt ttcccaaggc agtctggagc atgcgcttta 3060
gcagccccgc tgggcacttg gcgctacaca agtggcctct ggcctcgcac acattccaca 3120
tccaccggta ggcgccaacc ggctccgttc tttggtggcc ccttcgcgcc accttctact 3180
cctcccctag tcaggaagtt cccccccgcc ccgcagctcg cgtcgtgcag gacgtgacaa 3240
atggaagtag cacgtctcac tagtctcgtg cagatggaca gcaccgctga gcaatggaag 3300
cgggtaggcc tttggggcag cggccaatag cagctttgct ccttcgcttt ctgggctcag 3360
gggcgggctc agggggcggg gcgggcgccc gaaggtcctc cggaggcccg gcattctgca 3420
cgcttcaaaa gcgcacgtct gccgcgctgt tctcctcttc ctcatctccg ggcctttcga 3480
cctgcagcct gttgacaatt aatcatcggc atagtatatc ggcatagtat aatacgacaa 3540
ggtgaggaac taaaccatgg gtcaaagtag cgatgaagcc aacgctcccg ttgcagggca 3600
gtttgcgctt cccctgagtg ccacctttgg cttaggggat cgcgtacgca agaaatctgg 3660
tgccgcttgg cagggtcaag tcgtcggttg gtattgcaca aaactcactc ctgaaggcta 3720
tgcggtcgag tccgaatccc acccaggctc agtgcaaatt tatcctgtgg ctgcacttga 3780
acgtgtggcc taatgagggg atcaattctc tagagctcgc tgatcagaag ttcctattct 3840
ctagaaagta taggaacttc gatggcgcct catccctgaa gccaaagatg tgtataagag 3900
acag 3904
<210> 6
<211> 3793
<212> DNA
<213> 人工序列
<220>
<223> 表达盒
<400> 6
ctgtctctta tacacatctc cggccagatg attaattcct aatttttgtt gacactctat 60
cattgataga gttattttac cactccctat cagtgataga gaaaagtgaa atgaatagtt 120
cgacaaaaat ctagaaataa ttttgtttgg cgtcgagaag gagatagaaa atgtgcggta 180
tcgttggtgc tatcgcacag cgtgatgtag cgaaaatcct cctggaaggt ctgcgtcgtc 240
tcgaataccg tggttacgac tctgccggtc tggcagtagt ggatgcagaa ggtcacatga 300
ctcgtctgcg tcgtctgggt aaagtgcaga tgctcgcgca ggcggcggaa gaacacccac 360
tccacggtgg tacgggtatc gcacacactc gttgggcaac ccacggtgaa ccgtctgagg 420
tcaacgcaca cccgcatgtt agcgagcaca tcgtagtcgt tcacaacggt atcatcgaga 480
accacgaacc actccgtgag gaactcaaag cccgtggtta caccttcgta agcgaaaccg 540
acacggaagt tatcgcccac ctcgttaact gggaactcaa acagggtggt actctgcgtg 600
aagcagttct gcgtgccatt ccacagctgc gtggtgcata cggtaccgtg atcatggact 660
ctcgtcatcc ggataccctg ctcgccgcac gttctggttc tccactcgtt atcggtctgg 720
gtatgggtga gaacttcatc gcctctgatc agctggccct gctcccagtt acccgtcgct 780
tcatcttcct ggaagagggt gacatcgccg aaatcacccg tcgttccgtt aacatcttcg 840
acaaaacggg tgcggaagtt aaacgtcagg acatcgagtc taacctgcag tatgacgctg 900
gtgacaaagg catctaccgt cactacatgc agaaagagat ctacgaacag ccgaacgcga 960
tcaaaaacac cctgaccggt cgtatctctc acggtcaggt tgacctgtct gagctgggtc 1020
caaacgcgga cgaactcctg tccaaagtcg agcacatcca gatcctggct tgtggtacct 1080
cttacaactc cggtatggtt tctcgttact ggttcgaatc tctggcaggt atcccatgcg 1140
acgttgaaat cgcctccgaa ttccgttatc gtaaatctgc ggtacgtcgt aactccctca 1200
tgatcaccct gtctcagtct ggtgaaaccg ctgatactct ggcaggtctg cgtctcagca 1260
aagaactggg ttacctgggt tctctggcca tctgcaacgt tccgggttct agcctggttc 1320
gtgagtctgt gctggctctg atgaccaacg cgggtacgga gatcggtgtt gcctctacca 1380
aagcgttcac tacccagctc actgtcctgc tgatgctggt tgccaaactg tctcgtctca 1440
aaggcctcga cgctagcatc gaacacgaca tcgtacacgg tctgcaggcc ctcccatctc 1500
gtatcgagca gatgctgccg caggacaaac gtatcgaagc actggcagaa gacttcagcg 1560
acaaacacca cgcgctgttt ctgggtcgtg gtgaccagta cccaattgcg ctggaaggtg 1620
ccctgaaact gaaagagatc agctacatcc atgcagaggc atacgcagcg ggtgagctga 1680
aacatggtcc actggccctg atcgacgcag atatgccggt tattgtggtt gctccgaaca 1740
acggcctgct ggagaaactg aaatccaaca tcgaggaagt acgtgcgcgt ggtggtcagc 1800
tgtacgtgtt tgctgaccag gacgcgggtt tcgtttccag cgacaacatg cacatcatcg 1860
aaatgccgca tgttgaagag gtaatcgcgc caatcttcta caccgtaccg ctgcagctgc 1920
tggcgtacca tgtagccctg atcaaaggta cggacgttga ccagccgcgt aacctggcga 1980
aatccgtgac cgtggaataa cgaaggagat agaaccatga gcttacccga tggattttat 2040
ataaggcgaa tggaagaggg ggatttggaa caggtcactg agacgctaaa ggttttgacc 2100
accgtgggca ctattacccc cgaatccttc agcaaactca taaaatactg gaatgaagcc 2160
acagtatgga atgataacga agataaaaaa ataatgcaat ataaccccat ggtgattgtg 2220
gacaagcgca ccgagacggt tgccgctacg gggaatatca tcatcgaaag aaagatcatt 2280
catgaactgg ggctatgtgg ccacatcgag gacattgcag taaactccaa gtatcagggc 2340
caaggtttgg gcaagctctt gattgatcaa ttggtaacta tcggctttga ctacggttgt 2400
tataagatta ttttagattg cgatgagaaa aatgtcaaat tctatgaaaa atgtgggttt 2460
agcaacgcag gcgtggaaat gcaaattaga aaatagcatc cgtatcggaa acactagcat 2520
aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gctgaaacca atttgcctgg 2580
cggcagtagc gcggtggtcc cacctgaccc catgccgaac tcagaagtga aacgccgtag 2640
cgccgatggt agtgtggggt ctccccatgc gagagtaggg aactgccagg catcaaataa 2700
aacgaaaggc tcagtcgaaa gactgggcct ttcgcttcca caactttgta taataaagtt 2760
gtccccacgg ccagtgaatt cgagctcggt acctaccgtt cgtataatgt atgctatacg 2820
aagttatcga gctctagaga atgatcccct cattaggcca cacgttcaag tgcagcgcac 2880
accgtggaaa cggatgaagg cacgaaccca gttgacataa gcctgttcgg ttcgtaaact 2940
gtaatgcaag tagcgtatgc gctcacgcaa ctggtccaga accttgaccg aacgcagcgg 3000
tggtaacggc gcagtggcgg ttttcatggc ttgttatgac tgtttttttg tacagtctat 3060
gcctcgggca tccaagcagc aagcgcgtta cgccgtgggt cgatgtttga tgttatggag 3120
cagcaacgat gttacgcagc agcaacgatg ttacgcagca gggcagtcgc cctaaaacaa 3180
agttaggtgg ctcaagtatg ggcatcattc gcacatgtag gctcggccct gaccaagtca 3240
aatccatgcg ggctgctctt gatcttttcg gtcgtgagtt cggagacgta gccacctact 3300
cccaacatca gccggactcc gattacctcg ggaacttgct ccgtagtaag acattcatcg 3360
cgcttgctgc cttcgaccaa gaagcggttg ttggcgctct cgcggcttac gttctgccca 3420
ggtttgagca gccgcgtagt gagatctata tctatgatct cgcagtctcc ggcgagcacc 3480
ggaggcaggg cattgccacc gcgctcatca atctcctcaa gcatgaggcc aacgcgcttg 3540
gtgcttatgt gatctacgtg caagcagatt acggtgacga tcccgcagtg gctctctata 3600
caaagttggg catacgggaa gaagtgatgc actttgatat cgacccaagt accgccacct 3660
aacaattcgt tcaagccgag atcgtagaat ttcgacgacc tgcagccaag cataacttcg 3720
tataatgtat gctatacgaa cggtaggatc ctctagagtc gacctgcagg catgagatgt 3780
gtataagaga cag 3793
<210> 7
<211> 3847
<212> DNA
<213> 人工序列
<220>
<223> 表达盒
<400> 7
ctgtctctta tacacatctc cggccagatg attaattcct aatttttgtt gacactctat 60
cattgataga gttattttac cactccctat cagtgataga gaaaagtgaa atgaatagtt 120
cgacaaaaat ctagaaataa ttttgtttgg cgtcgagaag gagatagaac catgtccaac 180
aatggctcgt caccgctggt gctttggtat aaccaactcg gcatgaatga tgtagacagg 240
gttgggggca aaaatgcctc cctgggtgaa atgattacta acctttccgg aatgggtgtt 300
tccgttccga atggtttcgc cacaaccgcc gacgcgttta accagtttct ggaccaaagc 360
ggcgtaaacc agcgcattta tgaactgctg gataaaacgg atattgacga tgttactcag 420
cttgcgaaag cgggcgcgca aatccgccag tggattatcg acactccctt ccagcctgag 480
ctggaaaacg ccatcagcga agcctatgca cagctttctg ccgatgacga aaacgcctct 540
tttgcggtgc gctcctccgc caccgcagaa gatatgccgg acgcttcttt tgccggtcag 600
caggaaacct tcctcaacgt tcagggtttt gacgccgttc tcgtggcagt gaaacatgta 660
tttgcttctc tgtttaacga tcgcgccatc tcttatcgtg tgcaccaggg ttacgatcac 720
cgtggtgtgg cgctctccgc cggtgttcaa cggatggtgc gctctgacct cgcatcatct 780
ggcgtgatgt tctccattga taccgaatcc ggctttgacc aggtggtgtt tatcacttcc 840
gcatggggcc ttggtgagat ggtcgtgcag ggtgcggtta acccggatga gttttacgtg 900
cataaaccga cactggcggc gaatcgcccg gctatcgtgc gccgcaccat ggggtcgaaa 960
aaaatccgca tggtttacgc gccgacccag gagcacggca agcaggttaa aatcgaagac 1020
gtaccgcagg aacagcgtga catcttctcg ctgaccaacg aagaagtgca ggaactggca 1080
aaacaggccg tacaaattga gaaacactac ggtcgcccga tggatattga gtgggcgaaa 1140
gatggccaca ccggtaaact gttcattgtg caggcgcgtc cggaaaccgt gcgctcacgc 1200
ggtcaggtca tggagcgtta tacgctgcat tcacagggta agattatcgc cgaaggccgt 1260
gctatcggtc atcgcatcgg tgcgggtccg gtgaaagtca tccatgatat cagcgaaatg 1320
aaccgcatcg aacctggtga cgtgctggtc actgacatga ccgacccgga ctgggaaccg 1380
atcatgaaga aagcatctgc catcgtcacc aaccgtggcg gtcgtacctg tcacgcggcg 1440
atcatcgctc gtgaactggg cattccggcg gtagtgggct gtggtgatgc aacagaacgg 1500
atgaaagacg gtgagaacgt cactgtttct tgtgccgaag gtgataccgg ttacgtctat 1560
gcggagttgc tggaatttag cgtgaaaagc tccagcgtag aaacgatgcc ggatctgccg 1620
ttgaaagtga tgatgaacgt cggtaacccg gaccgagctt tcgacttcgc ctgtctgccg 1680
aacgaaggcg tgggacttgc gcgtctggaa tttatcatca accgtatgat tggcgtccac 1740
ccacgcgcac tgcttgagtt tgacgatcag gaaccgcagt tgcaaaacga aatccgcgag 1800
atgatgaaag gttttgattc tccgcgtgaa ttttacgttg gtcgtctgac tgaagggatc 1860
gcgacgctgg gtgccgcgtt ttatccgaag cgcgtcattg tccgtctctc tgattttaaa 1920
tcgaacgaat atgccaacct ggtcggtggt gagcgttacg agccagatga agagaacccg 1980
atgctcggct tccgtggcgc gggacgctat atttccgaca gcttccgcga ctgtttcgcg 2040
ctggagtgcg aagcagtgaa acgtgtgcgc aacgacatgg ggctgaccaa cgttgagatc 2100
atgatcccgt tcgtgcgaac cgtagatcag gcgaaagcgg tggttgagga actggcgcgt 2160
caggggctga aacgtggtga gaacgggctg aaaatcatca tgatgtgtga aatcccgtcc 2220
aacgccttgc tggccgagca gttcctcgaa tatttcgacg gcttctcaat tggctcaaac 2280
gacatgacgc agctggcgct cggtctggat cgtgactccg gcgtggtgtc tgaactgttc 2340
gatgagcgca acgatgcggt gaaagcactg ctgtcgatgg cgattcgtgc cgcgaagaaa 2400
cagggcaaat atgtcgggat ttgcggtcag ggtccgtccg accacgaaga ctttgccgca 2460
tggttgatgg aagaggggat cgatagcctg tctctgaacc cggacaccgt ggtgcaaacc 2520
tggttaagcc tggctgaact gaagaaataa catccgtatc ggaaacacta gcataacccc 2580
ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa accaatttgc ctggcggcag 2640
tagcgcggtg gtcccacctg accccatgcc gaactcagaa gtgaaacgcc gtagcgccga 2700
tggtagtgtg gggtctcccc atgcgagagt agggaactgc caggcatcaa ataaaacgaa 2760
aggctcagtc gaaagactgg gcctttcgct tccacaactt tgtataataa agttgtcccc 2820
acggccagtg aattcgagct cggtacctac cgttcgtata atgtatgcta tacgaagtta 2880
tcgagctcta gagaatgatc ccctcattag gccacacgtt caagtgcagc gcacaccgtg 2940
gaaacggatg aaggcacgaa cccagttgac ataagcctgt tcggttcgta aactgtaatg 3000
caagtagcgt atgcgctcac gcaactggtc cagaaccttg accgaacgca gcggtggtaa 3060
cggcgcagtg gcggttttca tggcttgtta tgactgtttt tttgtacagt ctatgcctcg 3120
ggcatccaag cagcaagcgc gttacgccgt gggtcgatgt ttgatgttat ggagcagcaa 3180
cgatgttacg cagcagcaac gatgttacgc agcagggcag tcgccctaaa acaaagttag 3240
gtggctcaag tatgggcatc attcgcacat gtaggctcgg ccctgaccaa gtcaaatcca 3300
tgcgggctgc tcttgatctt ttcggtcgtg agttcggaga cgtagccacc tactcccaac 3360
atcagccgga ctccgattac ctcgggaact tgctccgtag taagacattc atcgcgcttg 3420
ctgccttcga ccaagaagcg gttgttggcg ctctcgcggc ttacgttctg cccaggtttg 3480
agcagccgcg tagtgagatc tatatctatg atctcgcagt ctccggcgag caccggaggc 3540
agggcattgc caccgcgctc atcaatctcc tcaagcatga ggccaacgcg cttggtgctt 3600
atgtgatcta cgtgcaagca gattacggtg acgatcccgc agtggctctc tatacaaagt 3660
tgggcatacg ggaagaagtg atgcactttg atatcgaccc aagtaccgcc acctaacaat 3720
tcgttcaagc cgagatcgta gaatttcgac gacctgcagc caagcataac ttcgtataat 3780
gtatgctata cgaacggtag gatcctctag agtcgacctg caggcatgag atgtgtataa 3840
gagacag 3847
<210> 8
<211> 5554
<212> DNA
<213> 人工序列
<220>
<223> 质粒
<400> 8
catcgattta ttatgacaac ttgacggcta catcattcac tttttcttca caaccggcac 60
ggaactcgct cgggctggcc ccggtgcatt ttttaaatac ccgcgagaaa tagagttgat 120
cgtcaaaacc aacattgcga ccgacggtgg cgataggcat ccgggtggtg ctcaaaagca 180
gcttcgcctg gctgatacgt tggtcctcgc gccagcttaa gacgctaatc cctaactgct 240
ggcggaaaag atgtgacaga cgcgacggcg acaagcaaac atgctgtgcg acgctggcga 300
tatcaaaatt gctgtctgcc aggtgatcgc tgatgtactg acaagcctcg cgtacccgat 360
tatccatcgg tggatggagc gactcgttaa tcgcttccat gcgccgcagt aacaattgct 420
caagcagatt tatcgccagc agctccgaat agcgcccttc cccttgcccg gcgttaatga 480
tttgcccaaa caggtcgctg aaatgcggct ggtgcgcttc atccgggcga aagaaccccg 540
tattggcaaa tattgacggc cagttaagcc attcatgcca gtaggcgcgc ggacgaaagt 600
aaacccactg gtgataccat tcgcgagcct ccggatgacg accgtagtga tgaatctctc 660
ctggcgggaa cagcaaaata tcacccggtc ggcaaacaaa ttctcgtccc tgatttttca 720
ccaccccctg accgcgaatg gtgagattga gaatataacc tttcattccc agcggtcggt 780
cgataaaaaa atcgagataa ccgttggcct caatcggcgt taaacccgcc accagatggg 840
cattaaacga gtatcccggc agcaggggat cattttgcgc ttcagccata cttttcatac 900
tcccgccatt cagagaagaa accaattgtc catattgcat cagacattgc cgtcactgcg 960
tcttttactg gctcttctcg ctaaccaaac cggtaacccc gcttattaaa agcattctgt 1020
aacaaagcgg gaccaaagcc atgacaaaaa cgcgtaacaa aagtgtctat aatcacggca 1080
gaaaagtcca cattgattat ttgcacggcg tcacactttg ctatgccata gcatttttat 1140
ccataagatt agcggatcct acctgacgct ttttatcgca actctctact gtttctccat 1200
acccgttttt ttgggaattc gagctctaag gaggttataa aaaatgtcta atctgctgac 1260
ggtccaccaa aacctgccgg ctctgccggt cgatgctacc tctgatgaag ttcgcaaaaa 1320
cctgatggat atgtttcgtg atcgccaggc attcagcgaa catacctgga aaatgctgct 1380
gtccgtgtgc cgttcatggg cggcctggtg taaactgaac aatcgcaaat ggtttccggc 1440
ggaaccggaa gatgtccgtg actatctgct gtacctgcag gcccgcggtc tggcagttaa 1500
aacgatccag caacatctgg gccaactgaa tatgctgcac cgtcgctccg gtctgccgcg 1560
tccgagcgat tctaatgcgg tgtcactggt tatgcgtcgc attcgtaaag aaaacgtgga 1620
tgcaggcgaa cgcgctaaac aggcactggc ttttgaacgt accgatttcg accaagttcg 1680
ctcgctgatg gaaaacagcg atcgttgcca ggacatccgc aatctggcgt tcctgggtat 1740
tgcctataac accctgctgc gcattgcaga aatcgctcgt attcgcgtga aagatatcag 1800
ccgtacggac ggcggtcgca tgctgattca catcggccgt accaaaacgc tggtctctac 1860
cgcaggcgtg gaaaaagctc tgagtctggg tgtgacgaaa ctggttgaac gctggattag 1920
tgtctccggc gtggcggatg acccgaacaa ttacctgttt tgtcgtgttc gcaaaaatgg 1980
tgtcgcagct ccgtcagcca cctcgcagct gagcacgcgt gcactggaag gcatcttcga 2040
agctacccat cgcctgattt atggcgccaa agatgactcg ggtcaacgtt acctggcgtg 2100
gtctggtcac agtgcacgtg ttggtgccgc acgtgatatg gcccgtgccg gtgtttccat 2160
cccggaaatt atgcaggcag gcggttggac caacgttaat atcgtcatga actatattcg 2220
caatctggac tcggaaacgg gtgctatggt tcgcctgctg gaagacggtg actaatgagt 2280
gccggagttc atcgaaaaaa tggacgaggc actggctgaa attggttttg tatttgggga 2340
gcaatggcga tgacgcatcc tcacgataat atccgggtag gcgcaatcac tttcgtctac 2400
tccgttacaa agcgaggctg ggtatttccc ggcctttctg ttatccgaaa tccactgaaa 2460
gcacagcggc tggctgagga gataaataat aaacgagggg ctgtatgcac aaagcatctt 2520
ctgttgagtt aagaacgagt atcgagatgg cacatagcct tgctcaaatt ggaatcaggt 2580
ttgtgccaat accagtagaa acagacgaag aatccatggg tatggacagt tttccctttg 2640
atatgtaacg gtgaacagtt gttctacttt tgtttgttag tcttgatgct tcactgatag 2700
atacaagagc cataagaacc tcagatcctt ccgtatttag ccagtatgtt ctctagtgtg 2760
gttcgttgtt tttgcgtgag ccatgagaac gaaccattga gatcatactt actttgcatg 2820
tcactcaaaa attttgcctc aaaactggtg agctgaattt ttgcagttaa agcatcgtgt 2880
agtgtttttc ttagtccgtt acgtaggtag gaatctgatg taatggttgt tggtattttg 2940
tcaccattca tttttatctg gttgttctca agttcggtta cgagatccat ttgtctatct 3000
agttcaactt ggaaaatcaa cgtatcagtc gggcggcctc gcttatcaac caccaatttc 3060
atattgctgt aagtgtttaa atctttactt attggtttca aaacccattg gttaagcctt 3120
ttaaactcat ggtagttatt ttcaagcatt aacatgaact taaattcatc aaggctaatc 3180
tctatatttg ccttgtgagt tttcttttgt gttagttctt ttaataacca ctcataaatc 3240
ctcatagagt atttgttttc aaaagactta acatgttcca gattatattt tatgaatttt 3300
tttaactgga aaagataagg caatatctct tcactaaaaa ctaattctaa tttttcgctt 3360
gagaacttgg catagtttgt ccactggaaa atctcaaagc ctttaaccaa aggattcctg 3420
atttccacag ttctcgtcat cagctctctg gttgctttag ctaatacacc ataagcattt 3480
tccctactga tgttcatcat ctgagcgtat tggttataag tgaacgatac cgtccgttct 3540
ttccttgtag ggttttcaat cgtggggttg agtagtgcca cacagcataa aattagcttg 3600
gtttcatgct ccgttaagtc atagcgacta atcgctagtt catttgcttt gaaaacaact 3660
aattcagaca tacatctcaa ttggtctagg tgattttaat cactatacca attgagatgg 3720
gctagtcaat gataattact agtccttttc ctttgagttg tgggtatctg taaattctgc 3780
tagacctttg ctggaaaact tgtaaattct gctagaccct ctgtaaattc cgctagacct 3840
ttgtgtgttt tttttgttta tattcaagtg gttataattt atagaataaa gaaagaataa 3900
aaaaagataa aaagaataga tcccagccct gtgtataact cactacttta gtcagttccg 3960
cagtattaca aaaggatgtc gcaaacgctg tttgctcctc tacaaaacag accttaaaac 4020
cctaaaggct taagtagcac cctcgcaagc tcggttgcgg ccgcaatcgg gcaaatcgct 4080
gaatattcct tttgtctccg accatcaggc acctgagtcg ctgtcttttt cgtgacattc 4140
agttcgctgc gctcacggct ctggcagtga atgggggtaa atggcactac aggcgccttt 4200
tatggattca tgcaaggaaa ctacccataa tacaagaaaa gcccgtcacg ggcttctcag 4260
ggcgttttat ggcgggtctg ctatgtggtg ctatctgact ttttgctgtt cagcagttcc 4320
tgccctctga ttttccagtc tgaccacttc ggattatccc gtgacaggtc attcagactg 4380
gctaatgcac ccagtaaggc agcggtatca tcaacggggt ctgacgctca gtggaacgaa 4440
aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt 4500
ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac 4560
agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc 4620
atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc 4680
cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata 4740
aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc 4800
cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc 4860
aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca 4920
ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa 4980
gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca 5040
ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt 5100
tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt 5160
tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg 5220
ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga 5280
tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc 5340
agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg 5400
acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag 5460
ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg 5520
gttccgcgca catttccccg aaaagtgcca cctg 5554
<210> 9
<211> 3436
<212> DNA
<213> 人工序列
<220>
<223> 表达盒
<400> 9
ctgtctctta tacacatctc cggccagatg attaattcct aatttttgtt gacactctat 60
cattgataga gttattttac cactccctat cagtgataga gaaaagtgaa atgaatagtt 120
cgacaaaaat ctagaaataa ttttgtttaa ctttaagaag gagatataca aatgaagacc 180
gtgctggaca ccctgaaagg tcgcctggtt gtgagctgcc aagcgctgga aaatgagccg 240
ctgcatagcc cgtttattat gagccgtatg gcgctggcgg cgcgtcaggg tggtgcggcg 300
gcgatccgtg cgaacagcgt ggttgatatc gaggcgatta aggaacaagt taccctgccg 360
gtgatcggca tcattaagcg tgagtacccg gatagcgaag ttttcattac cgcgaccatg 420
aaagaggtgg acgaactgat gaccgtgagc ccggcgatca ttgcgctgga tgcgaccgac 480
cgtgcgcgtc cgggtggcga gagcctggcg atgctggtta cccgtatccg tacccgttat 540
ccgagcgtgc tgctgatggc ggatattgcg accgttgacg aagcggtgac cgcgcaggcg 600
ctgggtttcg attgcgttgg caccaccctg tacggttata ccgcgcagac cgtgggtcat 660
gcgctgccgg acgatgactg ccaatttctg aaagcggttc tggcggcggt taccgtgccg 720
gtggttgcgg aaggcaacgt ggacaccccg gaacgtgcgg cgcgttgcct ggcgctgggt 780
gcgcacatgg tggttgtggg tggcgcgatt acccgtccgc aacagattac cgaacgcttc 840
atggcggcga ttgatgcgca gagcaccgac cgtgcgtaat ttcgtcgaca cacaggaaac 900
atattaaaaa ttaaaacctg caggagttta aacgcggccg cgatatcgtt gtaaaacgac 960
ggccagtgca agaatcataa aaaatttatt tgctttcagg aaaatttttc tgtataatag 1020
attcataaat ttgagagagg agtttttgtg agcggataac aattccccat cttagtatat 1080
tagttaagta taaatacaca aggagatata catatgaaag aaatcaaaat ccagaacatc 1140
atcatcagcg aagaaaaagc gccgctggtt gtgccggaaa tcggcattaa ccataatggt 1200
agtctggaac tggcaaaaat catggtggat gcggccttta gcgccggtgc aaaaatcatt 1260
aaacatcaga cccacattgt ggaagatgaa atgtctaaag cagcgaaaaa agttatcccg 1320
ggcaacgcga aaatcagtat ctacgaaatc atgcagaaat gcgcgctgga ttacaaagat 1380
gaactggccc tgaaagaata taccgaaaaa ctgggtctgg tgtacctgtc taccccgttt 1440
agtcgtgcgg gtgcaaaccg tctggaagat atgggtgtta gtgcgttcaa aatcggcagc 1500
ggtgaatgta acaattatcc gctgatcaaa catattgccg catttaaaaa accgatgatt 1560
gttagcaccg gcatgaatag catcgaatct attaaaccga cggtgaaaat cctgctggat 1620
aacgaaattc cgtttgttct gatgcatacc acgaatctgt acccgacccc gcacaacctg 1680
gtgcgtctga atgccatgct ggaactgaaa aaagaattct cttgcatggt tggtctgagt 1740
gatcacacca cggataatct ggcatgcctg ggtgcagtgg ttctgggtgc gtgtgtgctg 1800
gaacgtcatt tcaccgatag catgcaccgc tctggtccgg atattgtttg tagtatggat 1860
acgaaagcac tgaaagaact gatcattcag agcgaacaga tggcgatcat tcgcggcaac 1920
aatgaatcta aaaaagcggc caaacaggaa caggtgacca tcgattttgc attcgcgagt 1980
gtggttagca tcaaagatat caaaaaaggc gaagtgctga gcatggataa tatttgggtt 2040
aaacgtccgg gtctgggcgg tatctctgca gcggaatttg aaaacattct gggcaaaaaa 2100
gcactgcgcg atattgaaaa tgatgcgcag ctgtcttatg aagatttcgc ctaaaataac 2160
tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgctg aaaccaattt 2220
gcctggcggc agtagcgcgg tggtcccacc tgaccccatg ccgaactcag aagtgaaacg 2280
ccgtagcgcc gatggtagtg tggggtctcc ccatgcgaga gtagggaact gccaggcatc 2340
aaataaaacg aaaggctcag tcgaaagact gggcctttcg ggatccaggc cggcctgtta 2400
acgaattaat cttccgcggc ggtatcgata agcttgatat cgaattccga agttcctatt 2460
ctctagaaag tataggaact tcaggtctga agaggagttt acgtccagcc aagctagctt 2520
ggctgcaggt cgtcgaaatt ctaccgggta ggggaggcgc ttttcccaag gcagtctgga 2580
gcatgcgctt tagcagcccc gctgggcact tggcgctaca caagtggcct ctggcctcgc 2640
acacattcca catccaccgg taggcgccaa ccggctccgt tctttggtgg ccccttcgcg 2700
ccaccttcta ctcctcccct agtcaggaag ttcccccccg ccccgcagct cgcgtcgtgc 2760
aggacgtgac aaatggaagt agcacgtctc actagtctcg tgcagatgga cagcaccgct 2820
gagcaatgga agcgggtagg cctttggggc agcggccaat agcagctttg ctccttcgct 2880
ttctgggctc aggggcgggc tcagggggcg gggcgggcgc ccgaaggtcc tccggaggcc 2940
cggcattctg cacgcttcaa aagcgcacgt ctgccgcgct gttctcctct tcctcatctc 3000
cgggcctttc gacctgcagc ctgttgacaa ttaatcatcg gcatagtata tcggcatagt 3060
ataatacgac aaggtgagga actaaaccat gggtcaaagt agcgatgaag ccaacgctcc 3120
cgttgcaggg cagtttgcgc ttcccctgag tgccaccttt ggcttagggg atcgcgtacg 3180
caagaaatct ggtgccgctt ggcagggtca agtcgtcggt tggtattgca caaaactcac 3240
tcctgaaggc tatgcggtcg agtccgaatc ccacccaggc tcagtgcaaa tttatcctgt 3300
ggctgcactt gaacgtgtgg cctaatgagg ggatcaattc tctagagctc gctgatcaga 3360
agttcctatt ctctagaaag tataggaact tcgatggcgc ctcatccctg aagccaaaga 3420
tgtgtataag agacag 3436
<210> 10
<211> 1830
<212> DNA
<213> 大肠杆菌(Escherichia coli)
<400> 10
atgtgtggaa ttgttggcgc gatcgcgcaa cgtgatgtag cagaaatcct tcttgaaggt 60
ttacgtcgtc tggaataccg cggatatgac tctgccggtc tggccgttgt tgatgcagaa 120
ggtcatatga cccgcctgcg tcgcctcggt aaagtccaga tgctggcaca ggcagcggaa 180
gaacatcctc tgcatggcgg cactggtatt gctcacactc gctgggcgac ccacggtgaa 240
ccttcagaag tgaatgcgca tccgcatgtt tctgaacaca ttgtggtggt gcataacggc 300
atcatcgaaa accatgaacc gctgcgtgaa gagctaaaag cgcgtggcta taccttcgtt 360
tctgaaaccg acaccgaagt gattgcccat ctggtgaact gggagctgaa acaaggcggg 420
actctgcgtg aggccgttct gcgtgctatc ccgcagctgc gtggtgcgta cggtacagtg 480
atcatggact cccgtcaccc ggataccctg ctggcggcac gttctggtag tccgctggtg 540
attggcctgg ggatgggcga aaactttatc gcttctgacc agctggcgct gttgccggtg 600
acccgtcgct ttatcttcct tgaagagggc gatattgcgg aaatcactcg ccgttcggta 660
aacatcttcg ataaaactgg cgcggaagta aaacgtcagg atatcgaatc caatctgcaa 720
tatgacgcgg gcgataaagg catttaccgt cactacatgc agaaagagat ctacgaacag 780
ccgaacgcga tcaaaaacac ccttaccgga cgcatcagcc acggtcaggt tgatttaagc 840
gagctgggac cgaacgccga cgaactgctg tcgaaggttg agcatattca gatcctcgcc 900
tgtggtactt cttataactc cggtatggtt tcccgctact ggtttgaatc gctagcaggt 960
attccgtgcg acgtcgaaat cgcctctgaa ttccgctatc gcaaatctgc cgtgcgtcgt 1020
aacagcctga tgatcacctt gtcacagtct ggcgaaaccg cggataccct ggctggcctg 1080
cgtctgtcga aagagctggg ttaccttggt tcactggcaa tctgtaacgt tccgggttct 1140
tctctggtgc gcgaatccga tctggcgcta atgaccaacg cgggtacaga aatcggcgtg 1200
gcatccacta aagcattcac cactcagtta actgtgctgt tgatgctggt ggcgaagctg 1260
tctcgcctga aaggtctgga tgcctccatt gaacatgaca tcgtgcatgg tctgcaggcg 1320
ctgccgagcc gtattgagca gatgctgtct caggacaaac gcattgaagc gctggcagaa 1380
gatttctctg acaaacatca cgcgctgttc ctgggccgtg gcgatcagta cccaatcgcg 1440
ctggaaggcg cattgaagtt gaaagagatc tcttacattc acgctgaagc ctacgctgct 1500
ggcgaactga aacacggtcc gctggcgcta attgatgccg atatgccggt tattgttgtt 1560
gcaccgaaca acgaattgct ggaaaaactg aaatccaaca ttgaagaagt tcgcgcgcgt 1620
ggcggtcagt tgtatgtctt cgccgatcag gatgcgggtt ttgtaagtag cgataacatg 1680
cacatcatcg agatgccgca tgtggaagag gtgattgcac cgatcttcta caccgttccg 1740
ctgcagctgc tggcttacca tgtcgcgctg atcaaaggca ccgacgttga ccagccgcgt 1800
aacctggcaa aatcggttac ggttgagtaa 1830
<210> 11
<211> 609
<212> PRT
<213> 大肠杆菌(Escherichia coli)
<400> 11
Met Cys Gly Ile Val Gly Ala Ile Ala Gln Arg Asp Val Ala Glu Ile
1 5 10 15
Leu Leu Glu Gly Leu Arg Arg Leu Glu Tyr Arg Gly Tyr Asp Ser Ala
20 25 30
Gly Leu Ala Val Val Asp Ala Glu Gly His Met Thr Arg Leu Arg Arg
35 40 45
Leu Gly Lys Val Gln Met Leu Ala Gln Ala Ala Glu Glu His Pro Leu
50 55 60
His Gly Gly Thr Gly Ile Ala His Thr Arg Trp Ala Thr His Gly Glu
65 70 75 80
Pro Ser Glu Val Asn Ala His Pro His Val Ser Glu His Ile Val Val
85 90 95
Val His Asn Gly Ile Ile Glu Asn His Glu Pro Leu Arg Glu Glu Leu
100 105 110
Lys Ala Arg Gly Tyr Thr Phe Val Ser Glu Thr Asp Thr Glu Val Ile
115 120 125
Ala His Leu Val Asn Trp Glu Leu Lys Gln Gly Gly Thr Leu Arg Glu
130 135 140
Ala Val Leu Arg Ala Ile Pro Gln Leu Arg Gly Ala Tyr Gly Thr Val
145 150 155 160
Ile Met Asp Ser Arg His Pro Asp Thr Leu Leu Ala Ala Arg Ser Gly
165 170 175
Ser Pro Leu Val Ile Gly Leu Gly Met Gly Glu Asn Phe Ile Ala Ser
180 185 190
Asp Gln Leu Ala Leu Leu Pro Val Thr Arg Arg Phe Ile Phe Leu Glu
195 200 205
Glu Gly Asp Ile Ala Glu Ile Thr Arg Arg Ser Val Asn Ile Phe Asp
210 215 220
Lys Thr Gly Ala Glu Val Lys Arg Gln Asp Ile Glu Ser Asn Leu Gln
225 230 235 240
Tyr Asp Ala Gly Asp Lys Gly Ile Tyr Arg His Tyr Met Gln Lys Glu
245 250 255
Ile Tyr Glu Gln Pro Asn Ala Ile Lys Asn Thr Leu Thr Gly Arg Ile
260 265 270
Ser His Gly Gln Val Asp Leu Ser Glu Leu Gly Pro Asn Ala Asp Glu
275 280 285
Leu Leu Ser Lys Val Glu His Ile Gln Ile Leu Ala Cys Gly Thr Ser
290 295 300
Tyr Asn Ser Gly Met Val Ser Arg Tyr Trp Phe Glu Ser Leu Ala Gly
305 310 315 320
Ile Pro Cys Asp Val Glu Ile Ala Ser Glu Phe Arg Tyr Arg Lys Ser
325 330 335
Ala Val Arg Arg Asn Ser Leu Met Ile Thr Leu Ser Gln Ser Gly Glu
340 345 350
Thr Ala Asp Thr Leu Ala Gly Leu Arg Leu Ser Lys Glu Leu Gly Tyr
355 360 365
Leu Gly Ser Leu Ala Ile Cys Asn Val Pro Gly Ser Ser Leu Val Arg
370 375 380
Glu Ser Asp Leu Ala Leu Met Thr Asn Ala Gly Thr Glu Ile Gly Val
385 390 395 400
Ala Ser Thr Lys Ala Phe Thr Thr Gln Leu Thr Val Leu Leu Met Leu
405 410 415
Val Ala Lys Leu Ser Arg Leu Lys Gly Leu Asp Ala Ser Ile Glu His
420 425 430
Asp Ile Val His Gly Leu Gln Ala Leu Pro Ser Arg Ile Glu Gln Met
435 440 445
Leu Ser Gln Asp Lys Arg Ile Glu Ala Leu Ala Glu Asp Phe Ser Asp
450 455 460
Lys His His Ala Leu Phe Leu Gly Arg Gly Asp Gln Tyr Pro Ile Ala
465 470 475 480
Leu Glu Gly Ala Leu Lys Leu Lys Glu Ile Ser Tyr Ile His Ala Glu
485 490 495
Ala Tyr Ala Ala Gly Glu Leu Lys His Gly Pro Leu Ala Leu Ile Asp
500 505 510
Ala Asp Met Pro Val Ile Val Val Ala Pro Asn Asn Glu Leu Leu Glu
515 520 525
Lys Leu Lys Ser Asn Ile Glu Glu Val Arg Ala Arg Gly Gly Gln Leu
530 535 540
Tyr Val Phe Ala Asp Gln Asp Ala Gly Phe Val Ser Ser Asp Asn Met
545 550 555 560
His Ile Ile Glu Met Pro His Val Glu Glu Val Ile Ala Pro Ile Phe
565 570 575
Tyr Thr Val Pro Leu Gln Leu Leu Ala Tyr His Val Ala Leu Ile Lys
580 585 590
Gly Thr Asp Val Asp Gln Pro Arg Asn Leu Ala Lys Ser Val Thr Val
595 600 605
Glu
<210> 12
<211> 1830
<212> DNA
<213> 大肠杆菌
<400> 12
atgtgcggta tcgttggtgc tatcgcacag cgtgatgtag cgaaaatcct cctggaaggt 60
ctgcgtcgtc tcgaataccg tggttacgac tctgccggtc tggcagtagt ggatgcagaa 120
ggtcacatga ctcgtctgcg tcgtctgggt aaagtgcaga tgctcgcgca ggcggcggaa 180
gaacacccac tccacggtgg tacgggtatc gcacacactc gttgggcaac ccacggtgaa 240
ccgtctgagg tcaacgcaca cccgcatgtt agcgagcaca tcgtagtcgt tcacaacggt 300
atcatcgaga accacgaacc actccgtgag gaactcaaag cccgtggtta caccttcgta 360
agcgaaaccg acacggaagt tatcgcccac ctcgttaact gggaactcaa acagggtggt 420
actctgcgtg aagcagttct gcgtgccatt ccacagctgc gtggtgcata cggtaccgtg 480
atcatggact ctcgtcatcc ggataccctg ctcgccgcac gttctggttc tccactcgtt 540
atcggtctgg gtatgggtga gaacttcatc gcctctgatc agctggccct gctcccagtt 600
acccgtcgct tcatcttcct ggaagagggt gacatcgccg aaatcacccg tcgttccgtt 660
aacatcttcg acaaaacggg tgcggaagtt aaacgtcagg acatcgagtc taacctgcag 720
tatgacgctg gtgacaaagg catctaccgt cactacatgc agaaagagat ctacgaacag 780
ccgaacgcga tcaaaaacac cctgaccggt cgtatctctc acggtcaggt tgacctgtct 840
gagctgggtc caaacgcgga cgaactcctg tccaaagtcg agcacatcca gatcctggct 900
tgtggtacct cttacaactc cggtatggtt tctcgttact ggttcgaatc tctggcaggt 960
atcccatgcg acgttgaaat cgcctccgaa ttccgttatc gtaaatctgc ggtacgtcgt 1020
aactccctca tgatcaccct gtctcagtct ggtgaaaccg ctgatactct ggcaggtctg 1080
cgtctcagca aagaactggg ttacctgggt tctctggcca tctgcaacgt tccgggttct 1140
agcctggttc gtgagtctgt gctggctctg atgaccaacg cgggtacgga gatcggtgtt 1200
gcctctacca aagcgttcac tacccagctc actgtcctgc tgatgctggt tgccaaactg 1260
tctcgtctca aaggcctcga cgctagcatc gaacacgaca tcgtacacgg tctgcaggcc 1320
ctcccatctc gtatcgagca gatgctgccg caggacaaac gtatcgaagc actggcagaa 1380
gacttcagcg acaaacacca cgcgctgttt ctgggtcgtg gtgaccagta cccaattgcg 1440
ctggaaggtg ccctgaaact gaaagagatc agctacatcc atgcagaggc atacgcagcg 1500
ggtgagctga aacatggtcc actggccctg atcgacgcag atatgccggt tattgtggtt 1560
gctccgaaca acggcctgct ggagaaactg aaatccaaca tcgaggaagt acgtgcgcgt 1620
ggtggtcagc tgtacgtgtt tgctgaccag gacgcgggtt tcgtttccag cgacaacatg 1680
cacatcatcg aaatgccgca tgttgaagag gtaatcgcgc caatcttcta caccgtaccg 1740
ctgcagctgc tggcgtacca tgtagccctg atcaaaggta cggacgttga ccagccgcgt 1800
aacctggcga aatccgtgac cgtggaataa 1830
<210> 13
<211> 609
<212> PRT
<213> 大肠杆菌
<400> 13
Met Cys Gly Ile Val Gly Ala Ile Ala Gln Arg Asp Val Ala Lys Ile
1 5 10 15
Leu Leu Glu Gly Leu Arg Arg Leu Glu Tyr Arg Gly Tyr Asp Ser Ala
20 25 30
Gly Leu Ala Val Val Asp Ala Glu Gly His Met Thr Arg Leu Arg Arg
35 40 45
Leu Gly Lys Val Gln Met Leu Ala Gln Ala Ala Glu Glu His Pro Leu
50 55 60
His Gly Gly Thr Gly Ile Ala His Thr Arg Trp Ala Thr His Gly Glu
65 70 75 80
Pro Ser Glu Val Asn Ala His Pro His Val Ser Glu His Ile Val Val
85 90 95
Val His Asn Gly Ile Ile Glu Asn His Glu Pro Leu Arg Glu Glu Leu
100 105 110
Lys Ala Arg Gly Tyr Thr Phe Val Ser Glu Thr Asp Thr Glu Val Ile
115 120 125
Ala His Leu Val Asn Trp Glu Leu Lys Gln Gly Gly Thr Leu Arg Glu
130 135 140
Ala Val Leu Arg Ala Ile Pro Gln Leu Arg Gly Ala Tyr Gly Thr Val
145 150 155 160
Ile Met Asp Ser Arg His Pro Asp Thr Leu Leu Ala Ala Arg Ser Gly
165 170 175
Ser Pro Leu Val Ile Gly Leu Gly Met Gly Glu Asn Phe Ile Ala Ser
180 185 190
Asp Gln Leu Ala Leu Leu Pro Val Thr Arg Arg Phe Ile Phe Leu Glu
195 200 205
Glu Gly Asp Ile Ala Glu Ile Thr Arg Arg Ser Val Asn Ile Phe Asp
210 215 220
Lys Thr Gly Ala Glu Val Lys Arg Gln Asp Ile Glu Ser Asn Leu Gln
225 230 235 240
Tyr Asp Ala Gly Asp Lys Gly Ile Tyr Arg His Tyr Met Gln Lys Glu
245 250 255
Ile Tyr Glu Gln Pro Asn Ala Ile Lys Asn Thr Leu Thr Gly Arg Ile
260 265 270
Ser His Gly Gln Val Asp Leu Ser Glu Leu Gly Pro Asn Ala Asp Glu
275 280 285
Leu Leu Ser Lys Val Glu His Ile Gln Ile Leu Ala Cys Gly Thr Ser
290 295 300
Tyr Asn Ser Gly Met Val Ser Arg Tyr Trp Phe Glu Ser Leu Ala Gly
305 310 315 320
Ile Pro Cys Asp Val Glu Ile Ala Ser Glu Phe Arg Tyr Arg Lys Ser
325 330 335
Ala Val Arg Arg Asn Ser Leu Met Ile Thr Leu Ser Gln Ser Gly Glu
340 345 350
Thr Ala Asp Thr Leu Ala Gly Leu Arg Leu Ser Lys Glu Leu Gly Tyr
355 360 365
Leu Gly Ser Leu Ala Ile Cys Asn Val Pro Gly Ser Ser Leu Val Arg
370 375 380
Glu Ser Val Leu Ala Leu Met Thr Asn Ala Gly Thr Glu Ile Gly Val
385 390 395 400
Ala Ser Thr Lys Ala Phe Thr Thr Gln Leu Thr Val Leu Leu Met Leu
405 410 415
Val Ala Lys Leu Ser Arg Leu Lys Gly Leu Asp Ala Ser Ile Glu His
420 425 430
Asp Ile Val His Gly Leu Gln Ala Leu Pro Ser Arg Ile Glu Gln Met
435 440 445
Leu Pro Gln Asp Lys Arg Ile Glu Ala Leu Ala Glu Asp Phe Ser Asp
450 455 460
Lys His His Ala Leu Phe Leu Gly Arg Gly Asp Gln Tyr Pro Ile Ala
465 470 475 480
Leu Glu Gly Ala Leu Lys Leu Lys Glu Ile Ser Tyr Ile His Ala Glu
485 490 495
Ala Tyr Ala Ala Gly Glu Leu Lys His Gly Pro Leu Ala Leu Ile Asp
500 505 510
Ala Asp Met Pro Val Ile Val Val Ala Pro Asn Asn Gly Leu Leu Glu
515 520 525
Lys Leu Lys Ser Asn Ile Glu Glu Val Arg Ala Arg Gly Gly Gln Leu
530 535 540
Tyr Val Phe Ala Asp Gln Asp Ala Gly Phe Val Ser Ser Asp Asn Met
545 550 555 560
His Ile Ile Glu Met Pro His Val Glu Glu Val Ile Ala Pro Ile Phe
565 570 575
Tyr Thr Val Pro Leu Gln Leu Leu Ala Tyr His Val Ala Leu Ile Lys
580 585 590
Gly Thr Asp Val Asp Gln Pro Arg Asn Leu Ala Lys Ser Val Thr Val
595 600 605
Glu
<210> 14
<211> 480
<212> DNA
<213> 酿酒酵母(Saccharomyces cerevisiae)
<400> 14
atgagcttac ccgatggatt ttatataagg cgaatggaag agggggattt ggaacaggtc 60
actgagacgc taaaggtttt gaccaccgtg ggcactatta cccccgaatc cttcagcaaa 120
ctcataaaat actggaatga agccacagta tggaatgata acgaagataa aaaaataatg 180
caatataacc ccatggtgat tgtggacaag cgcaccgaga cggttgccgc tacggggaat 240
atcatcatcg aaagaaagat cattcatgaa ctggggctat gtggccacat cgaggacatt 300
gcagtaaact ccaagtatca gggccaaggt ttgggcaagc tcttgattga tcaattggta 360
actatcggct ttgactacgg ttgttataag attattttag attgcgatga gaaaaatgtc 420
aaattctatg aaaaatgtgg gtttagcaac gcaggcgtgg aaatgcaaat tagaaaatag 480
<210> 15
<211> 159
<212> PRT
<213> 酿酒酵母
<400> 15
Met Ser Leu Pro Asp Gly Phe Tyr Ile Arg Arg Met Glu Glu Gly Asp
1 5 10 15
Leu Glu Gln Val Thr Glu Thr Leu Lys Val Leu Thr Thr Val Gly Thr
20 25 30
Ile Thr Pro Glu Ser Phe Ser Lys Leu Ile Lys Tyr Trp Asn Glu Ala
35 40 45
Thr Val Trp Asn Asp Asn Glu Asp Lys Lys Ile Met Gln Tyr Asn Pro
50 55 60
Met Val Ile Val Asp Lys Arg Thr Glu Thr Val Ala Ala Thr Gly Asn
65 70 75 80
Ile Ile Ile Glu Arg Lys Ile Ile His Glu Leu Gly Leu Cys Gly His
85 90 95
Ile Glu Asp Ile Ala Val Asn Ser Lys Tyr Gln Gly Gln Gly Leu Gly
100 105 110
Lys Leu Leu Ile Asp Gln Leu Val Thr Ile Gly Phe Asp Tyr Gly Cys
115 120 125
Tyr Lys Ile Ile Leu Asp Cys Asp Glu Lys Asn Val Lys Phe Tyr Glu
130 135 140
Lys Cys Gly Phe Ser Asn Ala Gly Val Glu Met Gln Ile Arg Lys
145 150 155
<210> 16
<211> 567
<212> DNA
<213> 大肠杆菌
<400> 16
atgtacgagc gttatgcagg tttaattttt gatatggatg gcacaatcct ggatacggag 60
cctacgcacc gtaaagcgtg gcgcgaagta ttagggcact acggtcttca gtacgatatt 120
caggcgatga ttgcgcttaa tggatcgccc acctggcgta ttgctcaggc aattattgag 180
ctgaatcagg ccgatctcga cccgcatgcg ttagcgcgtg aaaaaacaga agcagtaaga 240
agtatgctgc tggatagcgt cgaaccgctt cctcttgttg atgtggtgaa aagttggcat 300
ggtcgtcgcc caatggctgt aggaacgggg agtgaaagcg ccatcgctga ggcattgctg 360
gcgcacctgg gattacgcca ttattttgac gccgtcgtcg ctgccgatca cgtcaaacac 420
cataaacccg cgccagacac atttttgttg tgcgcgcagc gtatgggcgt gcaaccgacg 480
cagtgtgtgg tctttgaaga tgccgatttc ggtattcagg cggcccgtgc agcaggcatg 540
gacgccgtgg atgttcgctt gctgtga 567
<210> 17
<211> 188
<212> PRT
<213> 大肠杆菌
<400> 17
Met Tyr Glu Arg Tyr Ala Gly Leu Ile Phe Asp Met Asp Gly Thr Ile
1 5 10 15
Leu Asp Thr Glu Pro Thr His Arg Lys Ala Trp Arg Glu Val Leu Gly
20 25 30
His Tyr Gly Leu Gln Tyr Asp Ile Gln Ala Met Ile Ala Leu Asn Gly
35 40 45
Ser Pro Thr Trp Arg Ile Ala Gln Ala Ile Ile Glu Leu Asn Gln Ala
50 55 60
Asp Leu Asp Pro His Ala Leu Ala Arg Glu Lys Thr Glu Ala Val Arg
65 70 75 80
Ser Met Leu Leu Asp Ser Val Glu Pro Leu Pro Leu Val Asp Val Val
85 90 95
Lys Ser Trp His Gly Arg Arg Pro Met Ala Val Gly Thr Gly Ser Glu
100 105 110
Ser Ala Ile Ala Glu Ala Leu Leu Ala His Leu Gly Leu Arg His Tyr
115 120 125
Phe Asp Ala Val Val Ala Ala Asp His Val Lys His His Lys Pro Ala
130 135 140
Pro Asp Thr Phe Leu Leu Cys Ala Gln Arg Met Gly Val Gln Pro Thr
145 150 155 160
Gln Cys Val Val Phe Glu Asp Ala Asp Phe Gly Ile Gln Ala Ala Arg
165 170 175
Ala Ala Gly Met Asp Ala Val Asp Val Arg Leu Leu
180 185
<210> 18
<211> 600
<212> DNA
<213> 大肠杆菌
<400> 18
atgctctata tctttgattt aggtaatgtg attgtcgata tcgactttaa ccgtgtgctg 60
ggagcctgga gcgatttaac gcgtattccg ctggcatcgc ttaagaagag ttttcatatg 120
ggggaggcgt ttcatcagca tgagcgtggg gaaattagcg acgaagcgtt cgcagaggcg 180
ctgtgtcatg agatggctct accgctaagc tacgagcagt tctctcacgg ctggcaggcg 240
gtgtttgttg cgctgcgccc ggaagtgatc gccatcatgc ataaactgcg tgagcagggg 300
catcgcgtgg tggtgctttc caataccaac cgcctgcata ccaccttctg gccggaagaa 360
tacccggaaa ttcgtgatgc tgctgaccat atctatctgt cgcaagatct ggggatgcgc 420
aaacctgaag cacgaattta ccagcatgtt ttgcaggcgg aaggtttttc acccagcgat 480
acggtctttt tcgacgataa cgccgataat atagaaggag ccaatcagct gggcattacc 540
agtattctgg tgaaagataa aaccaccatc ccggactatt tcgcgaaggt gttatgctaa 600
<210> 19
<211> 199
<212> PRT
<213> 大肠杆菌
<400> 19
Met Leu Tyr Ile Phe Asp Leu Gly Asn Val Ile Val Asp Ile Asp Phe
1 5 10 15
Asn Arg Val Leu Gly Ala Trp Ser Asp Leu Thr Arg Ile Pro Leu Ala
20 25 30
Ser Leu Lys Lys Ser Phe His Met Gly Glu Ala Phe His Gln His Glu
35 40 45
Arg Gly Glu Ile Ser Asp Glu Ala Phe Ala Glu Ala Leu Cys His Glu
50 55 60
Met Ala Leu Pro Leu Ser Tyr Glu Gln Phe Ser His Gly Trp Gln Ala
65 70 75 80
Val Phe Val Ala Leu Arg Pro Glu Val Ile Ala Ile Met His Lys Leu
85 90 95
Arg Glu Gln Gly His Arg Val Val Val Leu Ser Asn Thr Asn Arg Leu
100 105 110
His Thr Thr Phe Trp Pro Glu Glu Tyr Pro Glu Ile Arg Asp Ala Ala
115 120 125
Asp His Ile Tyr Leu Ser Gln Asp Leu Gly Met Arg Lys Pro Glu Ala
130 135 140
Arg Ile Tyr Gln His Val Leu Gln Ala Glu Gly Phe Ser Pro Ser Asp
145 150 155 160
Thr Val Phe Phe Asp Asp Asn Ala Asp Asn Ile Glu Gly Ala Asn Gln
165 170 175
Leu Gly Ile Thr Ser Ile Leu Val Lys Asp Lys Thr Thr Ile Pro Asp
180 185 190
Tyr Phe Ala Lys Val Leu Cys
195
<210> 20
<211> 1266
<212> DNA
<213> 卵形拟杆菌(Bacteroides ovatus)
<400> 20
atggatagta agaataacat tggtcattca gcagacatct ctttaactgc tgaattaccc 60
ataccaatct ataatggaaa tacgattatg gatttcaaaa aactggcaag tctgtacaag 120
gatgagctcc tggacaacgt ccttcctttc tggcttgaac attcacaaga ccatgagtat 180
ggtggttact tcacctgtct ggaccgtgaa ggaaaagtat tcgatacgga taagtttatt 240
tggctgcaaa gtcgtgaggt atggatgttc tccatgcttt acaacaaagt ggagaaacgt 300
caggaatggc tagactgtgc cattcagggt ggcgaatttc taaaaaaata tggacatgac 360
ggcaattata actggtattt ttccctcgac cgttcgggta gaccattggt agaaccgtac 420
aatatattct cgtatacatt cgctaccatg gctttcggac agttgagcct tacaaccggt 480
aatcaggaat atgcggacat tgccaagaaa actttcgata taatcctttc caaagtggat 540
aatccgaaag ggagatggaa taagcttcat ccgggtaccc gtaatctgaa gaactttgcc 600
ttgccaatga tcctctgtaa cttggcactg gagatagagc atttattgga tgaaacgtat 660
ctgcgggaaa caatggatac ttgtatccat gaagtgatgg aagttttcta tcgtcctgaa 720
ctcggaggta tcattgttga aaacgtggac atagacggta atttggtcga ttgttttgaa 780
ggccgtcagg tgaccccggg acatgccatt gaagcgatgt ggtttatcat ggatctaggc 840
aagcgtctga atcgtccgga attgatagag aaagccaaag agactactct cacgatgctt 900
aattatggct gggacaagca atatggaggt atctactatt ttatggatcg taacggttgt 960
cctccccaac aattggagtg ggaccagaaa ctctggtggg tccatatcga aacgcttatt 1020
tccctgctga aaggctatca attgacggga gacaaaaaat gcttggaatg gtttgaaaag 1080
gtacatgact acacttggga gcatttcaag gataaagaat atcctgaatg gtatggctac 1140
ttgaaccgaa gaggcgaagt attgctacca ctcaaaggag gaaaatggaa aggatgcttc 1200
catgtgccaa gaggactgta tcagtgctgg aaaacattag aagaaataaa aaatatagta 1260
tcctaa 1266
<210> 21
<211> 421
<212> PRT
<213> 卵形拟杆菌
<400> 21
Met Asp Ser Lys Asn Asn Ile Gly His Ser Ala Asp Ile Ser Leu Thr
1 5 10 15
Ala Glu Leu Pro Ile Pro Ile Tyr Asn Gly Asn Thr Ile Met Asp Phe
20 25 30
Lys Lys Leu Ala Ser Leu Tyr Lys Asp Glu Leu Leu Asp Asn Val Leu
35 40 45
Pro Phe Trp Leu Glu His Ser Gln Asp His Glu Tyr Gly Gly Tyr Phe
50 55 60
Thr Cys Leu Asp Arg Glu Gly Lys Val Phe Asp Thr Asp Lys Phe Ile
65 70 75 80
Trp Leu Gln Ser Arg Glu Val Trp Met Phe Ser Met Leu Tyr Asn Lys
85 90 95
Val Glu Lys Arg Gln Glu Trp Leu Asp Cys Ala Ile Gln Gly Gly Glu
100 105 110
Phe Leu Lys Lys Tyr Gly His Asp Gly Asn Tyr Asn Trp Tyr Phe Ser
115 120 125
Leu Asp Arg Ser Gly Arg Pro Leu Val Glu Pro Tyr Asn Ile Phe Ser
130 135 140
Tyr Thr Phe Ala Thr Met Ala Phe Gly Gln Leu Ser Leu Thr Thr Gly
145 150 155 160
Asn Gln Glu Tyr Ala Asp Ile Ala Lys Lys Thr Phe Asp Ile Ile Leu
165 170 175
Ser Lys Val Asp Asn Pro Lys Gly Arg Trp Asn Lys Leu His Pro Gly
180 185 190
Thr Arg Asn Leu Lys Asn Phe Ala Leu Pro Met Ile Leu Cys Asn Leu
195 200 205
Ala Leu Glu Ile Glu His Leu Leu Asp Glu Thr Tyr Leu Arg Glu Thr
210 215 220
Met Asp Thr Cys Ile His Glu Val Met Glu Val Phe Tyr Arg Pro Glu
225 230 235 240
Leu Gly Gly Ile Ile Val Glu Asn Val Asp Ile Asp Gly Asn Leu Val
245 250 255
Asp Cys Phe Glu Gly Arg Gln Val Thr Pro Gly His Ala Ile Glu Ala
260 265 270
Met Trp Phe Ile Met Asp Leu Gly Lys Arg Leu Asn Arg Pro Glu Leu
275 280 285
Ile Glu Lys Ala Lys Glu Thr Thr Leu Thr Met Leu Asn Tyr Gly Trp
290 295 300
Asp Lys Gln Tyr Gly Gly Ile Tyr Tyr Phe Met Asp Arg Asn Gly Cys
305 310 315 320
Pro Pro Gln Gln Leu Glu Trp Asp Gln Lys Leu Trp Trp Val His Ile
325 330 335
Glu Thr Leu Ile Ser Leu Leu Lys Gly Tyr Gln Leu Thr Gly Asp Lys
340 345 350
Lys Cys Leu Glu Trp Phe Glu Lys Val His Asp Tyr Thr Trp Glu His
355 360 365
Phe Lys Asp Lys Glu Tyr Pro Glu Trp Tyr Gly Tyr Leu Asn Arg Arg
370 375 380
Gly Glu Val Leu Leu Pro Leu Lys Gly Gly Lys Trp Lys Gly Cys Phe
385 390 395 400
His Val Pro Arg Gly Leu Tyr Gln Cys Trp Lys Thr Leu Glu Glu Ile
405 410 415
Lys Asn Ile Val Ser
420
<210> 22
<211> 1176
<212> DNA
<213> 集胞藻属(Synechocystis)PCC6803
<400> 22
atgattgccc atcgccgtca ggagttagcc cagcaatatt accaggcttt acaccaggac 60
gtattgccct tttgggaaaa atattccctc gatcgccagg ggggcggtta ctttacctgc 120
ttagaccgta aaggccaggt ttttgacaca gataaattca tttggttaca aaaccgtcag 180
gtatggcagt ttgccgtttt ctacaaccgt ttggaaccaa aaccccaatg gttagaaatt 240
gcccgccatg gtgctgattt tttagctcgc cacggccgag atcaagacgg taattggtat 300
tttgctttgg atcaggaagg caaacccctg cgtcaaccct ataacgtttt ttccgattgc 360
ttcgccgcca tggcctttag tcaatatgcc ttagccagtg gggcgcagga agctaaagcc 420
attgccctgc aggcctacaa taacgtccta cgccgtcagc acaatcccaa aggtcaatac 480
gagaagtcct atccaggtac tagacccctc aaatccctgg cggtgccgat gattttagcc 540
aacctcaccc tggagatgga atggttatta ccgcctacta ccgtggaaga ggtgttggcc 600
caaaccgtca gagaagtgat gacggatttc ctcgacccag aaataggatt aatgcgggaa 660
gcggtgaccc ccacaggaga atttgttgat agttttgaag ggcggttgct caacccagga 720
cacggcattg aagccatgtg gttcatgatg gacattgccc aacgctccgg cgatcgccag 780
ttacaggagc aagccattgc agtggtgttg aacaccctgg aatatgcctg ggatgaagaa 840
tttggtggca tattttattt ccttgatcgc cagggccacc ctccccaaca actggaatgg 900
gaccaaaagc tctggtgggt acatttggaa accctggttg ccctagccaa gggccaccaa 960
gccactggcc aagaaaaatg ttggcaatgg tttgagcggg tccatgatta cgcctggagt 1020
catttcgccg atcctgagta tggggaatgg tttggctacc tgaatcgccg gggagaggtg 1080
ttactcaacc taaaaggggg gaaatggaaa gggtgcttcc acgtgccccg agctctgtgg 1140
ctctgtgcgg aaactctcca acttccggtt agttaa 1176
<210> 23
<211> 391
<212> PRT
<213> 集胞藻属PCC6803
<400> 23
Met Ile Ala His Arg Arg Gln Glu Leu Ala Gln Gln Tyr Tyr Gln Ala
1 5 10 15
Leu His Gln Asp Val Leu Pro Phe Trp Glu Lys Tyr Ser Leu Asp Arg
20 25 30
Gln Gly Gly Gly Tyr Phe Thr Cys Leu Asp Arg Lys Gly Gln Val Phe
35 40 45
Asp Thr Asp Lys Phe Ile Trp Leu Gln Asn Arg Gln Val Trp Gln Phe
50 55 60
Ala Val Phe Tyr Asn Arg Leu Glu Pro Lys Pro Gln Trp Leu Glu Ile
65 70 75 80
Ala Arg His Gly Ala Asp Phe Leu Ala Arg His Gly Arg Asp Gln Asp
85 90 95
Gly Asn Trp Tyr Phe Ala Leu Asp Gln Glu Gly Lys Pro Leu Arg Gln
100 105 110
Pro Tyr Asn Val Phe Ser Asp Cys Phe Ala Ala Met Ala Phe Ser Gln
115 120 125
Tyr Ala Leu Ala Ser Gly Ala Gln Glu Ala Lys Ala Ile Ala Leu Gln
130 135 140
Ala Tyr Asn Asn Val Leu Arg Arg Gln His Asn Pro Lys Gly Gln Tyr
145 150 155 160
Glu Lys Ser Tyr Pro Gly Thr Arg Pro Leu Lys Ser Leu Ala Val Pro
165 170 175
Met Ile Leu Ala Asn Leu Thr Leu Glu Met Glu Trp Leu Leu Pro Pro
180 185 190
Thr Thr Val Glu Glu Val Leu Ala Gln Thr Val Arg Glu Val Met Thr
195 200 205
Asp Phe Leu Asp Pro Glu Ile Gly Leu Met Arg Glu Ala Val Thr Pro
210 215 220
Thr Gly Glu Phe Val Asp Ser Phe Glu Gly Arg Leu Leu Asn Pro Gly
225 230 235 240
His Gly Ile Glu Ala Met Trp Phe Met Met Asp Ile Ala Gln Arg Ser
245 250 255
Gly Asp Arg Gln Leu Gln Glu Gln Ala Ile Ala Val Val Leu Asn Thr
260 265 270
Leu Glu Tyr Ala Trp Asp Glu Glu Phe Gly Gly Ile Phe Tyr Phe Leu
275 280 285
Asp Arg Gln Gly His Pro Pro Gln Gln Leu Glu Trp Asp Gln Lys Leu
290 295 300
Trp Trp Val His Leu Glu Thr Leu Val Ala Leu Ala Lys Gly His Gln
305 310 315 320
Ala Thr Gly Gln Glu Lys Cys Trp Gln Trp Phe Glu Arg Val His Asp
325 330 335
Tyr Ala Trp Ser His Phe Ala Asp Pro Glu Tyr Gly Glu Trp Phe Gly
340 345 350
Tyr Leu Asn Arg Arg Gly Glu Val Leu Leu Asn Leu Lys Gly Gly Lys
355 360 365
Trp Lys Gly Cys Phe His Val Pro Arg Ala Leu Trp Leu Cys Ala Glu
370 375 380
Thr Leu Gln Leu Pro Val Ser
385 390
<210> 24
<211> 708
<212> DNA
<213> 阴沟肠杆菌(Enterobacter cloacae)
<400> 24
atgaaaactg tactggatac cctgaaggga agactggtcg tctcctgtca ggcgcttgag 60
aacgaaccgt tgcatagccc gtttattatg tcgcggatgg cgctggcggc gcgtcaggga 120
ggggctgcgg ccatccgtgc caacagcgtg gtggatattg aggcgatcaa agagcaggtt 180
acgctgccgg ttattggcat catcaagcgg gagtaccccg acagcgaggt gtttatcacc 240
gcaacgatga aagaggtgga tgaactgatg accgtctccc cggcgatcat tgcgcttgat 300
gcgaccgaca gggcgcggcc tggcggggaa tctctggcaa tgctggttac gcgcattcgt 360
acccgttatc cctcggtgct gcttatggct gatatagcca ctgttgatga ggccgtcacg 420
gcgcaggcgc tggggtttga ttgtgtcggg accacgcttt acggctacac cgcgcagacc 480
gtcggccacg ccttacccga tgatgactgt cagtttctga aagcggtact ggcagccgtc 540
acggtaccgg tggtggccga aggtaacgtg gacaccccgg aacgcgccgc cagatgtctg 600
gcgttggggg cgcatatggt ggtggtgggc ggggcaatca cccgcccgca gcagattacg 660
gaacgcttta tggcggcaat tgacgcgcaa agcaccgatc gagcatga 708
<210> 25
<211> 235
<212> PRT
<213> 阴沟肠杆菌
<400> 25
Met Lys Thr Val Leu Asp Thr Leu Lys Gly Arg Leu Val Val Ser Cys
1 5 10 15
Gln Ala Leu Glu Asn Glu Pro Leu His Ser Pro Phe Ile Met Ser Arg
20 25 30
Met Ala Leu Ala Ala Arg Gln Gly Gly Ala Ala Ala Ile Arg Ala Asn
35 40 45
Ser Val Val Asp Ile Glu Ala Ile Lys Glu Gln Val Thr Leu Pro Val
50 55 60
Ile Gly Ile Ile Lys Arg Glu Tyr Pro Asp Ser Glu Val Phe Ile Thr
65 70 75 80
Ala Thr Met Lys Glu Val Asp Glu Leu Met Thr Val Ser Pro Ala Ile
85 90 95
Ile Ala Leu Asp Ala Thr Asp Arg Ala Arg Pro Gly Gly Glu Ser Leu
100 105 110
Ala Met Leu Val Thr Arg Ile Arg Thr Arg Tyr Pro Ser Val Leu Leu
115 120 125
Met Ala Asp Ile Ala Thr Val Asp Glu Ala Val Thr Ala Gln Ala Leu
130 135 140
Gly Phe Asp Cys Val Gly Thr Thr Leu Tyr Gly Tyr Thr Ala Gln Thr
145 150 155 160
Val Gly His Ala Leu Pro Asp Asp Asp Cys Gln Phe Leu Lys Ala Val
165 170 175
Leu Ala Ala Val Thr Val Pro Val Val Ala Glu Gly Asn Val Asp Thr
180 185 190
Pro Glu Arg Ala Ala Arg Cys Leu Ala Leu Gly Ala His Met Val Val
195 200 205
Val Gly Gly Ala Ile Thr Arg Pro Gln Gln Ile Thr Glu Arg Phe Met
210 215 220
Ala Ala Ile Asp Ala Gln Ser Thr Asp Arg Ala
225 230 235
<210> 26
<211> 2652
<212> DNA
<213> 大肠杆菌
<400> 26
atgaacgaac aatattccgc attgcgtagt aatgtcagta tgctcggcaa agtgctggga 60
gaaaccatca aggatgcgtt gggagaacac attcttgaac gcgtagaaac tatccgtaag 120
ttgtcgaaat cttcacgcgc tggcaatgat gctaaccgcc aggagttgct caccacctta 180
caaaatttgt cgaacgacga gctgctgccc gttgcgcgtg cgtttagtca gttcctgaac 240
ctggccaaca ccgccgagca ataccacagc atttcgccga aaggcgaagc tgccagcaac 300
ccggaagtga tcgcccgcac cctgcgtaaa ctgaaaaacc agccggaact gagcgaagac 360
accatcaaaa aagcagtgga atcgctgtcg ctggaactgg tcctcacggc tcacccaacc 420
gaaattaccc gtcgtacact gatccacaaa atggtggaag tgaacgcctg tttaaaacag 480
ctcgataaca aagatatcgc tgactacgaa cacaaccagc tgatgcgtcg cctgcgccag 540
ttgatcgccc agtcatggca taccgatgaa atccgtaagc tgcgtccaag cccggtagat 600
gaagccaaat ggggctttgc cgtagtggaa aacagcctgt ggcaaggcgt accaaattac 660
ctgcgcgaac tgaacgaaca actggaagag aacctcggct acaaactgcc cgtcgaattt 720
gttccggtcc gttttacttc gtggatgggc ggcgaccgcg acggcaaccc gaacgtcact 780
gccgatatca cccgccacgt cctgctactc agccgctgga aagccaccga tttgttcctg 840
aaagatattc aggtgctggt ttctgaactg tcgatggttg aagcgacccc tgaactgctg 900
gcgctggttg gcgaagaagg tgccgcagaa ccgtatcgct atctgatgaa aaacctgcgt 960
tctcgcctga tggcgacaca ggcatggctg gaagcgcgcc tgaaaggcga agaactgcca 1020
aaaccagaag gcctgctgac acaaaacgaa gaactgtggg aaccgctcta cgcttgctac 1080
cagtcacttc aggcgtgtgg catgggtatt atcgccaacg gcgatctgct cgacaccctg 1140
cgccgcgtga aatgtttcgg cgtaccgctg gtccgtattg atatccgtca ggagagcacg 1200
cgtcataccg aagcgctggg cgagctgacc cgctacctcg gtatcggcga ctacgaaagc 1260
tggtcagagg ccgacaaaca ggcgttcctg atccgcgaac tgaactccaa acgtccgctt 1320
ctgccgcgca actggcaacc aagcgccgaa acgcgcgaag tgctcgatac ctgccaggtg 1380
attgccgaag caccgcaagg ctccattgcc gcctacgtga tctcgatggc gaaaacgccg 1440
tccgacgtac tggctgtcca cctgctgctg aaagaagcgg gtatcgggtt tgcgatgccg 1500
gttgctccgc tgtttgaaac cctcgatgat ctgaacaacg ccaacgatgt catgacccag 1560
ctgctcaata ttgactggta tcgtggcctg attcagggca aacagatggt gatgattggc 1620
tattccgact cagcaaaaga tgcgggagtg atggcagctt cctgggcgca atatcaggca 1680
caggatgcat taatcaaaac ctgcgaaaaa gcgggtattg agctgacgtt gttccacggt 1740
cgcggcggtt ccattggtcg cggcggcgca cctgctcatg cggcgctgct gtcacaaccg 1800
ccaggaagcc tgaaaggcgg cctgcgcgta accgaacagg gcgagatgat ccgctttaaa 1860
tatggtctgc cagaaatcac cgtcagcagc ctgtcgcttt ataccggggc gattctggaa 1920
gccaacctgc tgccaccgcc ggagccgaaa gagagctggc gtcgcattat ggatgaactg 1980
tcagtcatct cctgcgatgt ctaccgcggc tacgtacgtg aaaacaaaga ttttgtgcct 2040
tacttccgct ccgctacgcc ggaacaagaa ctgggcaaac tgccgttggg ttcacgtccg 2100
gcgaaacgtc gcccaaccgg cggcgtcgag tcactacgcg ccattccgtg gatcttcgcc 2160
tggacgcaaa accgtctgat gctccccgcc tggctgggtg caggtacggc gctgcaaaaa 2220
gtggtcgaag acggcaaaca gagcgagctg gaggctatgt gccgcgattg gccattcttc 2280
tcgacgcgtc tcggcatgct ggagatggtc ttcgccaaag cagacctgtg gctggcggaa 2340
tactatgacc aacgcctggt agacaaagca ctgtggccgt taggtaaaga gttacgcaac 2400
ctgcaagaag aagacatcaa agtggtgctg gcgattgcca acgattccca tctgatggcc 2460
gatctgccgt ggattgcaga gtctattcag ctacggaata tttacaccga cccgctgaac 2520
gtattgcagg ccgagttgct gcaccgctcc cgccaggcag aaaaagaagg ccaggaaccg 2580
gatcctcgcg tcgaacaagc gttaatggtc actattgccg ggattgcggc aggtatgcgt 2640
aataccggct aa 2652
<210> 27
<211> 883
<212> PRT
<213> 大肠杆菌
<400> 27
Met Asn Glu Gln Tyr Ser Ala Leu Arg Ser Asn Val Ser Met Leu Gly
1 5 10 15
Lys Val Leu Gly Glu Thr Ile Lys Asp Ala Leu Gly Glu His Ile Leu
20 25 30
Glu Arg Val Glu Thr Ile Arg Lys Leu Ser Lys Ser Ser Arg Ala Gly
35 40 45
Asn Asp Ala Asn Arg Gln Glu Leu Leu Thr Thr Leu Gln Asn Leu Ser
50 55 60
Asn Asp Glu Leu Leu Pro Val Ala Arg Ala Phe Ser Gln Phe Leu Asn
65 70 75 80
Leu Ala Asn Thr Ala Glu Gln Tyr His Ser Ile Ser Pro Lys Gly Glu
85 90 95
Ala Ala Ser Asn Pro Glu Val Ile Ala Arg Thr Leu Arg Lys Leu Lys
100 105 110
Asn Gln Pro Glu Leu Ser Glu Asp Thr Ile Lys Lys Ala Val Glu Ser
115 120 125
Leu Ser Leu Glu Leu Val Leu Thr Ala His Pro Thr Glu Ile Thr Arg
130 135 140
Arg Thr Leu Ile His Lys Met Val Glu Val Asn Ala Cys Leu Lys Gln
145 150 155 160
Leu Asp Asn Lys Asp Ile Ala Asp Tyr Glu His Asn Gln Leu Met Arg
165 170 175
Arg Leu Arg Gln Leu Ile Ala Gln Ser Trp His Thr Asp Glu Ile Arg
180 185 190
Lys Leu Arg Pro Ser Pro Val Asp Glu Ala Lys Trp Gly Phe Ala Val
195 200 205
Val Glu Asn Ser Leu Trp Gln Gly Val Pro Asn Tyr Leu Arg Glu Leu
210 215 220
Asn Glu Gln Leu Glu Glu Asn Leu Gly Tyr Lys Leu Pro Val Glu Phe
225 230 235 240
Val Pro Val Arg Phe Thr Ser Trp Met Gly Gly Asp Arg Asp Gly Asn
245 250 255
Pro Asn Val Thr Ala Asp Ile Thr Arg His Val Leu Leu Leu Ser Arg
260 265 270
Trp Lys Ala Thr Asp Leu Phe Leu Lys Asp Ile Gln Val Leu Val Ser
275 280 285
Glu Leu Ser Met Val Glu Ala Thr Pro Glu Leu Leu Ala Leu Val Gly
290 295 300
Glu Glu Gly Ala Ala Glu Pro Tyr Arg Tyr Leu Met Lys Asn Leu Arg
305 310 315 320
Ser Arg Leu Met Ala Thr Gln Ala Trp Leu Glu Ala Arg Leu Lys Gly
325 330 335
Glu Glu Leu Pro Lys Pro Glu Gly Leu Leu Thr Gln Asn Glu Glu Leu
340 345 350
Trp Glu Pro Leu Tyr Ala Cys Tyr Gln Ser Leu Gln Ala Cys Gly Met
355 360 365
Gly Ile Ile Ala Asn Gly Asp Leu Leu Asp Thr Leu Arg Arg Val Lys
370 375 380
Cys Phe Gly Val Pro Leu Val Arg Ile Asp Ile Arg Gln Glu Ser Thr
385 390 395 400
Arg His Thr Glu Ala Leu Gly Glu Leu Thr Arg Tyr Leu Gly Ile Gly
405 410 415
Asp Tyr Glu Ser Trp Ser Glu Ala Asp Lys Gln Ala Phe Leu Ile Arg
420 425 430
Glu Leu Asn Ser Lys Arg Pro Leu Leu Pro Arg Asn Trp Gln Pro Ser
435 440 445
Ala Glu Thr Arg Glu Val Leu Asp Thr Cys Gln Val Ile Ala Glu Ala
450 455 460
Pro Gln Gly Ser Ile Ala Ala Tyr Val Ile Ser Met Ala Lys Thr Pro
465 470 475 480
Ser Asp Val Leu Ala Val His Leu Leu Leu Lys Glu Ala Gly Ile Gly
485 490 495
Phe Ala Met Pro Val Ala Pro Leu Phe Glu Thr Leu Asp Asp Leu Asn
500 505 510
Asn Ala Asn Asp Val Met Thr Gln Leu Leu Asn Ile Asp Trp Tyr Arg
515 520 525
Gly Leu Ile Gln Gly Lys Gln Met Val Met Ile Gly Tyr Ser Asp Ser
530 535 540
Ala Lys Asp Ala Gly Val Met Ala Ala Ser Trp Ala Gln Tyr Gln Ala
545 550 555 560
Gln Asp Ala Leu Ile Lys Thr Cys Glu Lys Ala Gly Ile Glu Leu Thr
565 570 575
Leu Phe His Gly Arg Gly Gly Ser Ile Gly Arg Gly Gly Ala Pro Ala
580 585 590
His Ala Ala Leu Leu Ser Gln Pro Pro Gly Ser Leu Lys Gly Gly Leu
595 600 605
Arg Val Thr Glu Gln Gly Glu Met Ile Arg Phe Lys Tyr Gly Leu Pro
610 615 620
Glu Ile Thr Val Ser Ser Leu Ser Leu Tyr Thr Gly Ala Ile Leu Glu
625 630 635 640
Ala Asn Leu Leu Pro Pro Pro Glu Pro Lys Glu Ser Trp Arg Arg Ile
645 650 655
Met Asp Glu Leu Ser Val Ile Ser Cys Asp Val Tyr Arg Gly Tyr Val
660 665 670
Arg Glu Asn Lys Asp Phe Val Pro Tyr Phe Arg Ser Ala Thr Pro Glu
675 680 685
Gln Glu Leu Gly Lys Leu Pro Leu Gly Ser Arg Pro Ala Lys Arg Arg
690 695 700
Pro Thr Gly Gly Val Glu Ser Leu Arg Ala Ile Pro Trp Ile Phe Ala
705 710 715 720
Trp Thr Gln Asn Arg Leu Met Leu Pro Ala Trp Leu Gly Ala Gly Thr
725 730 735
Ala Leu Gln Lys Val Val Glu Asp Gly Lys Gln Ser Glu Leu Glu Ala
740 745 750
Met Cys Arg Asp Trp Pro Phe Phe Ser Thr Arg Leu Gly Met Leu Glu
755 760 765
Met Val Phe Ala Lys Ala Asp Leu Trp Leu Ala Glu Tyr Tyr Asp Gln
770 775 780
Arg Leu Val Asp Lys Ala Leu Trp Pro Leu Gly Lys Glu Leu Arg Asn
785 790 795 800
Leu Gln Glu Glu Asp Ile Lys Val Val Leu Ala Ile Ala Asn Asp Ser
805 810 815
His Leu Met Ala Asp Leu Pro Trp Ile Ala Glu Ser Ile Gln Leu Arg
820 825 830
Asn Ile Tyr Thr Asp Pro Leu Asn Val Leu Gln Ala Glu Leu Leu His
835 840 845
Arg Ser Arg Gln Ala Glu Lys Glu Gly Gln Glu Pro Asp Pro Arg Val
850 855 860
Glu Gln Ala Leu Met Val Thr Ile Ala Gly Ile Ala Ala Gly Met Arg
865 870 875 880
Asn Thr Gly
<210> 28
<211> 1041
<212> DNA
<213> 空肠弯曲杆菌(Campylobacter jejuni)
<400> 28
atgaaagaaa taaaaataca aaatataatc ataagtgaag aaaaagcacc cttagtcgtg 60
cctgaaatag gcattaatca taatggcagt ttagaactag ctaaaattat ggtagatgca 120
gcctttagcg caggtgctaa gattataaag catcaaaccc acatcgttga agatgagatg 180
agtaaggccg ctaaaaaagt aattcctggt aatgcaaaaa taagcattta tgagattatg 240
caaaaatgtg ctttagatta taaagatgag ctagcactta aagaatacac agaaaaatta 300
ggtcttgttt atcttagcac acctttttct cgtgcaggtg caaaccgctt agaagatatg 360
ggagttagtg cttttaagat tggttcaggt gagtgtaata attatccgct tattaaacac 420
atagcagcct ttaaaaagcc tatgatagtt agcacaggaa tgaatagtat tgaaagtata 480
aaaccaactg taaaaatctt attagacaat gaaattccct ttgttttaat gcactcgacc 540
aatctttacc caaccccgca taatcttgta agattaaacg ctatgcttga attaaaaaaa 600
gaattttctt gcatggtagg cttaagcgac cacacaacag ataatcttgc gtgtttaggt 660
gcggttgcac ttggtgcttg tgtgcttgaa agacatttta ctgatagtat gcatagaagt 720
ggccctgata tagtttgttc tatggataca aaggctttaa aagagctaat tatccaaagt 780
gagcaaatgg ctataatgaa aggaaataat gaaagcaaaa aagcagctaa gcaagaacaa 840
gttacaattg attttgcctt tgcaagcgta gttagcatta aagatattaa aaaaggcgaa 900
gttttatcta tggacaatat ctgggttaaa agacctggac ttggtggaat tagtgcggct 960
gaatttgaaa atattttagg caaaaaagca ttaagagata tagaaaatga tactcagtta 1020
agctatgagg attttgcgtg a 1041
<210> 29
<211> 346
<212> PRT
<213> 空肠弯曲杆菌
<400> 29
Met Lys Glu Ile Lys Ile Gln Asn Ile Ile Ile Ser Glu Glu Lys Ala
1 5 10 15
Pro Leu Val Val Pro Glu Ile Gly Ile Asn His Asn Gly Ser Leu Glu
20 25 30
Leu Ala Lys Ile Met Val Asp Ala Ala Phe Ser Ala Gly Ala Lys Ile
35 40 45
Ile Lys His Gln Thr His Ile Val Glu Asp Glu Met Ser Lys Ala Ala
50 55 60
Lys Lys Val Ile Pro Gly Asn Ala Lys Ile Ser Ile Tyr Glu Ile Met
65 70 75 80
Gln Lys Cys Ala Leu Asp Tyr Lys Asp Glu Leu Ala Leu Lys Glu Tyr
85 90 95
Thr Glu Lys Leu Gly Leu Val Tyr Leu Ser Thr Pro Phe Ser Arg Ala
100 105 110
Gly Ala Asn Arg Leu Glu Asp Met Gly Val Ser Ala Phe Lys Ile Gly
115 120 125
Ser Gly Glu Cys Asn Asn Tyr Pro Leu Ile Lys His Ile Ala Ala Phe
130 135 140
Lys Lys Pro Met Ile Val Ser Thr Gly Met Asn Ser Ile Glu Ser Ile
145 150 155 160
Lys Pro Thr Val Lys Ile Leu Leu Asp Asn Glu Ile Pro Phe Val Leu
165 170 175
Met His Ser Thr Asn Leu Tyr Pro Thr Pro His Asn Leu Val Arg Leu
180 185 190
Asn Ala Met Leu Glu Leu Lys Lys Glu Phe Ser Cys Met Val Gly Leu
195 200 205
Ser Asp His Thr Thr Asp Asn Leu Ala Cys Leu Gly Ala Val Ala Leu
210 215 220
Gly Ala Cys Val Leu Glu Arg His Phe Thr Asp Ser Met His Arg Ser
225 230 235 240
Gly Pro Asp Ile Val Cys Ser Met Asp Thr Lys Ala Leu Lys Glu Leu
245 250 255
Ile Ile Gln Ser Glu Gln Met Ala Ile Met Lys Gly Asn Asn Glu Ser
260 265 270
Lys Lys Ala Ala Lys Gln Glu Gln Val Thr Ile Asp Phe Ala Phe Ala
275 280 285
Ser Val Val Ser Ile Lys Asp Ile Lys Lys Gly Glu Val Leu Ser Met
290 295 300
Asp Asn Ile Trp Val Lys Arg Pro Gly Leu Gly Gly Ile Ser Ala Ala
305 310 315 320
Glu Phe Glu Asn Ile Leu Gly Lys Lys Ala Leu Arg Asp Ile Glu Asn
325 330 335
Asp Thr Gln Leu Ser Tyr Glu Asp Phe Ala
340 345
<210> 30
<211> 876
<212> DNA
<213> 大肠杆菌
<400> 30
atgaccacac tggcgattga tatcggcggt actaaacttg ccgccgcgct gattggcgct 60
gacgggcaga tccgcgatcg tcgtgaactt cctacgccag ccagccagac accagaagcc 120
ttgcgtgatg ccttatccgc attagtctct ccgttgcaag ctcatgcgca gcgggttgcc 180
atcgcttcga ccgggataat ccgtgacggc agcttgctgg cgcttaatcc gcataatctt 240
ggtggattgc tacactttcc gttagtcaaa acgctggaac aacttaccaa tttgccgacc 300
attgccatta acgacgcgca ggccgcagca tgggcggagt ttcaggcgct ggatggcgat 360
ataaccgata tggtctttat caccgtttcc accggcgttg gcggcggtgt agtgagcggc 420
tgcaaactgc ttaccggccc tggcggtctg gcggggcata tcgggcatac gcttgccgat 480
ccacacggcc cagtctgcgg ctgtggacgc acaggttgcg tggaagcgat tgcttctggt 540
cgcggcattg cagcggcagc gcagggggag ttggctggcg cggatgcgaa aactattttc 600
acgcgcgccg ggcagggtga cgagcaggcg cagcagctga ttcaccgctc cgcacgtacg 660
cttgcaaggc tgatcgctga tattaaagcc acaactgatt gccagtgcgt ggtggtcggt 720
ggcagcgttg gtctggcaga agggtatctg gcgctggtgg aaacgtatct ggcgcaggag 780
ccagcggcat ttcatgttga tttactggcg gcgcattacc gccatgatgc aggtttactt 840
ggggctgcgc tgttggccca gggagaaaaa ttatga 876
<210> 31
<211> 291
<212> PRT
<213> 大肠杆菌
<400> 31
Met Thr Thr Leu Ala Ile Asp Ile Gly Gly Thr Lys Leu Ala Ala Ala
1 5 10 15
Leu Ile Gly Ala Asp Gly Gln Ile Arg Asp Arg Arg Glu Leu Pro Thr
20 25 30
Pro Ala Ser Gln Thr Pro Glu Ala Leu Arg Asp Ala Leu Ser Ala Leu
35 40 45
Val Ser Pro Leu Gln Ala His Ala Gln Arg Val Ala Ile Ala Ser Thr
50 55 60
Gly Ile Ile Arg Asp Gly Ser Leu Leu Ala Leu Asn Pro His Asn Leu
65 70 75 80
Gly Gly Leu Leu His Phe Pro Leu Val Lys Thr Leu Glu Gln Leu Thr
85 90 95
Asn Leu Pro Thr Ile Ala Ile Asn Asp Ala Gln Ala Ala Ala Trp Ala
100 105 110
Glu Phe Gln Ala Leu Asp Gly Asp Ile Thr Asp Met Val Phe Ile Thr
115 120 125
Val Ser Thr Gly Val Gly Gly Gly Val Val Ser Gly Cys Lys Leu Leu
130 135 140
Thr Gly Pro Gly Gly Leu Ala Gly His Ile Gly His Thr Leu Ala Asp
145 150 155 160
Pro His Gly Pro Val Cys Gly Cys Gly Arg Thr Gly Cys Val Glu Ala
165 170 175
Ile Ala Ser Gly Arg Gly Ile Ala Ala Ala Ala Gln Gly Glu Leu Ala
180 185 190
Gly Ala Asp Ala Lys Thr Ile Phe Thr Arg Ala Gly Gln Gly Asp Glu
195 200 205
Gln Ala Gln Gln Leu Ile His Arg Ser Ala Arg Thr Leu Ala Arg Leu
210 215 220
Ile Ala Asp Ile Lys Ala Thr Thr Asp Cys Gln Cys Val Val Val Gly
225 230 235 240
Gly Ser Val Gly Leu Ala Glu Gly Tyr Leu Ala Leu Val Glu Thr Tyr
245 250 255
Leu Ala Gln Glu Pro Ala Ala Phe His Val Asp Leu Leu Ala Ala His
260 265 270
Tyr Arg His Asp Ala Gly Leu Leu Gly Ala Ala Leu Leu Ala Gln Gly
275 280 285
Glu Lys Leu
290
<210> 32
<211> 690
<212> DNA
<213> 大肠杆菌
<400> 32
atgtcgttac ttgcacaact ggatcaaaaa atcgctgcta acggtggcct gattgtctcc 60
tgccagccgg ttccggacag cccgctcgat aaacccgaaa tcgtcgccgc catggcatta 120
gcggcagaac aggcgggcgc ggttgccatt cgcattgaag gtgtggcaaa tctgcaagcc 180
acgcgtgcgg tggtgagcgt gccgattatt ggaattgtga aacgcgatct ggaggattct 240
ccggtacgca tcacggccta tattgaagat gttgatgcgc tggcgcaggc gggcgcggac 300
attatcgcca ttgacggcac cgaccgcccg cgtccggtgc ctgttgaaac gctgctggca 360
cgtattcacc atcacggttt actggcgatg accgactgct caacgccgga agacggcctg 420
gcatgccaaa agctgggagc cgaaattatt ggcactacgc tttctggcta taccacgcct 480
gaaacgccag aagagccgga tctggcgctg gtgaaaacgt tgagcgacgc cggatgtcgg 540
gtgattgccg aagggcgtta caacacgcct gctcaggcgg cggatgcgat gcgccacggc 600
gcgtgggcgg tgacggtcgg ttctgcaatc acgcgtcttg agcacatttg tcagtggtac 660
aacacagcga tgaaaaaggc ggtgctatga 690
<210> 33
<211> 229
<212> PRT
<213> 大肠杆菌
<400> 33
Met Ser Leu Leu Ala Gln Leu Asp Gln Lys Ile Ala Ala Asn Gly Gly
1 5 10 15
Leu Ile Val Ser Cys Gln Pro Val Pro Asp Ser Pro Leu Asp Lys Pro
20 25 30
Glu Ile Val Ala Ala Met Ala Leu Ala Ala Glu Gln Ala Gly Ala Val
35 40 45
Ala Ile Arg Ile Glu Gly Val Ala Asn Leu Gln Ala Thr Arg Ala Val
50 55 60
Val Ser Val Pro Ile Ile Gly Ile Val Lys Arg Asp Leu Glu Asp Ser
65 70 75 80
Pro Val Arg Ile Thr Ala Tyr Ile Glu Asp Val Asp Ala Leu Ala Gln
85 90 95
Ala Gly Ala Asp Ile Ile Ala Ile Asp Gly Thr Asp Arg Pro Arg Pro
100 105 110
Val Pro Val Glu Thr Leu Leu Ala Arg Ile His His His Gly Leu Leu
115 120 125
Ala Met Thr Asp Cys Ser Thr Pro Glu Asp Gly Leu Ala Cys Gln Lys
130 135 140
Leu Gly Ala Glu Ile Ile Gly Thr Thr Leu Ser Gly Tyr Thr Thr Pro
145 150 155 160
Glu Thr Pro Glu Glu Pro Asp Leu Ala Leu Val Lys Thr Leu Ser Asp
165 170 175
Ala Gly Cys Arg Val Ile Ala Glu Gly Arg Tyr Asn Thr Pro Ala Gln
180 185 190
Ala Ala Asp Ala Met Arg His Gly Ala Trp Ala Val Thr Val Gly Ser
195 200 205
Ala Ile Thr Arg Leu Glu His Ile Cys Gln Trp Tyr Asn Thr Ala Met
210 215 220
Lys Lys Ala Val Leu
225
<210> 34
<211> 894
<212> DNA
<213> 大肠杆菌
<400> 34
atggcaacga atttacgtgg cgtaatggct gcactcctga ctccttttga ccaacaacaa 60
gcactggata aagcgagtct gcgtcgcctg gttcagttca atattcagca gggcatcgac 120
ggtttatacg tgggtggttc gaccggcgag gcctttgtac aaagcctttc cgagcgtgaa 180
caggtactgg aaatcgtcgc cgaagaggcg aaaggtaaga ttaaactcat cgcccacgtc 240
ggttgcgtca gcaccgccga aagccaacaa cttgcggcat cggctaaacg ttatggcttc 300
gatgccgtct ccgccgtcac gccgttctac tatcctttca gctttgaaga acactgcgat 360
cactatcggg caattattga ttcggcggat ggtttgccga tggtggtgta caacattcca 420
gccctgagtg gggtaaaact gaccctggat cagatcaaca cacttgttac attgcctggc 480
gtaggtgcgc tgaaacagac ctctggcgat ctctatcaga tggagcagat ccgtcgtgaa 540
catcctgatc ttgtgctcta taacggttac gacgaaatct tcgcctctgg tctgctggcg 600
ggcgctgatg gtggtatcgg cagtacctac aacatcatgg gctggcgcta tcaggggatc 660
gttaaggcgc tgaaagaagg cgatatccag accgcgcaga aactgcaaac tgaatgcaat 720
aaagtcattg atttactgat caaaacgggc gtattccgcg gcctgaaaac tgtcctccat 780
tatatggatg tcgtttctgt gccgctgtgc cgcaaaccgt ttggaccggt agatgaaaaa 840
tatctgccag aactgaaggc gctggcccag cagttgatgc aagagcgcgg gtga 894
<210> 35
<211> 297
<212> PRT
<213> 大肠杆菌
<400> 35
Met Ala Thr Asn Leu Arg Gly Val Met Ala Ala Leu Leu Thr Pro Phe
1 5 10 15
Asp Gln Gln Gln Ala Leu Asp Lys Ala Ser Leu Arg Arg Leu Val Gln
20 25 30
Phe Asn Ile Gln Gln Gly Ile Asp Gly Leu Tyr Val Gly Gly Ser Thr
35 40 45
Gly Glu Ala Phe Val Gln Ser Leu Ser Glu Arg Glu Gln Val Leu Glu
50 55 60
Ile Val Ala Glu Glu Ala Lys Gly Lys Ile Lys Leu Ile Ala His Val
65 70 75 80
Gly Cys Val Ser Thr Ala Glu Ser Gln Gln Leu Ala Ala Ser Ala Lys
85 90 95
Arg Tyr Gly Phe Asp Ala Val Ser Ala Val Thr Pro Phe Tyr Tyr Pro
100 105 110
Phe Ser Phe Glu Glu His Cys Asp His Tyr Arg Ala Ile Ile Asp Ser
115 120 125
Ala Asp Gly Leu Pro Met Val Val Tyr Asn Ile Pro Ala Leu Ser Gly
130 135 140
Val Lys Leu Thr Leu Asp Gln Ile Asn Thr Leu Val Thr Leu Pro Gly
145 150 155 160
Val Gly Ala Leu Lys Gln Thr Ser Gly Asp Leu Tyr Gln Met Glu Gln
165 170 175
Ile Arg Arg Glu His Pro Asp Leu Val Leu Tyr Asn Gly Tyr Asp Glu
180 185 190
Ile Phe Ala Ser Gly Leu Leu Ala Gly Ala Asp Gly Gly Ile Gly Ser
195 200 205
Thr Tyr Asn Ile Met Gly Trp Arg Tyr Gln Gly Ile Val Lys Ala Leu
210 215 220
Lys Glu Gly Asp Ile Gln Thr Ala Gln Lys Leu Gln Thr Glu Cys Asn
225 230 235 240
Lys Val Ile Asp Leu Leu Ile Lys Thr Gly Val Phe Arg Gly Leu Lys
245 250 255
Thr Val Leu His Tyr Met Asp Val Val Ser Val Pro Leu Cys Arg Lys
260 265 270
Pro Phe Gly Pro Val Asp Glu Lys Tyr Leu Pro Glu Leu Lys Ala Leu
275 280 285
Ala Gln Gln Leu Met Gln Glu Arg Gly
290 295
<210> 36
<211> 1491
<212> DNA
<213> 大肠杆菌
<400> 36
atgagtacta caacccagaa tatcccgtgg tatcgccatc tcaaccgtgc acaatggcgc 60
gcattttccg ctgcctggtt gggatatctg cttgacggtt ttgatttcgt tttaatcgcc 120
ctggtactca ccgaagtaca aggtgaattc gggctgacga cggtgcaggc ggcaagtctg 180
atctctgcag cctttatctc tcgctggttc ggcggcctga tgctcggcgc tatgggtgac 240
cgctacgggc gtcgtctggc aatggtcacc agcatcgttc tcttctcggc cgggacgctg 300
gcctgcggct ttgcgccagg ctacatcacc atgtttatcg ctcgtctggt catcggcatg 360
gggatggcgg gtgaatacgg ttccagcgcc acctatgtca ttgaaagctg gccaaaacat 420
ctgcgtaaca aagccagtgg ttttttgatt tcaggcttct ctgtgggggc cgtcgttgcc 480
gctcaggtct atagcctggt ggttccggtc tggggctggc gtgcgctgtt ctttatcggc 540
attttgccaa tcatctttgc tctctggctg cgtaaaaaca tcccggaagc ggaagactgg 600
aaagagaaac acgcaggtaa agcaccagta cgcacaatgg tggatattct ctaccgtggt 660
gaacatcgca ttgccaatat cgtaatgaca ctggcggcgg ctactgcgct gtggttctgc 720
ttcgccggta acctgcaaaa tgccgcgatc gtcgctgttc ttgggctgtt atgcgccgca 780
atctttatca gctttatggt gcagagtgca ggcaaacgct ggccaacggg cgtaatgctg 840
atggtggtcg tgttgtttgc tttcctctac tcatggccga ttcaggcgct gctgccaacg 900
tatctgaaaa ccgatctggc ttataacccg catactgtag ccaatgtgct gttctttagt 960
ggctttggcg cggcggtggg atgctgcgta ggtggcttcc tcggtgactg gctgggaacc 1020
cgcaaagcgt acgtttgtag cctgctggcc tcgcagctgc tgattattcc ggtatttgcg 1080
attggcggcg caaacgtctg ggtgctcggt ctgttactgt tcttccagca aatgcttgga 1140
caagggatcg ccgggatctt accaaaactg attggcggtt atttcgatac cgaccagcgt 1200
gcagcgggcc tgggctttac ctacaacgtt ggcgcattgg gcggtgcact ggccccaatc 1260
atcggcgcgt tgatcgctca acgtctggat ctgggtactg cgctggcatc gctctcgttc 1320
agtctgacgt tcgtggtgat cctgctgatt gggctggata tgccttctcg cgttcagcgt 1380
tggttgcgcc cggaagcgtt gcgtactcat gacgctatcg acggtaaacc attcagcggt 1440
gccgtgccgt ttggcagcgc caaaaacgat ttagtcaaaa ccaaaagtta a 1491
<210> 37
<211> 496
<212> PRT
<213> 大肠杆菌
<400> 37
Met Ser Thr Thr Thr Gln Asn Ile Pro Trp Tyr Arg His Leu Asn Arg
1 5 10 15
Ala Gln Trp Arg Ala Phe Ser Ala Ala Trp Leu Gly Tyr Leu Leu Asp
20 25 30
Gly Phe Asp Phe Val Leu Ile Ala Leu Val Leu Thr Glu Val Gln Gly
35 40 45
Glu Phe Gly Leu Thr Thr Val Gln Ala Ala Ser Leu Ile Ser Ala Ala
50 55 60
Phe Ile Ser Arg Trp Phe Gly Gly Leu Met Leu Gly Ala Met Gly Asp
65 70 75 80
Arg Tyr Gly Arg Arg Leu Ala Met Val Thr Ser Ile Val Leu Phe Ser
85 90 95
Ala Gly Thr Leu Ala Cys Gly Phe Ala Pro Gly Tyr Ile Thr Met Phe
100 105 110
Ile Ala Arg Leu Val Ile Gly Met Gly Met Ala Gly Glu Tyr Gly Ser
115 120 125
Ser Ala Thr Tyr Val Ile Glu Ser Trp Pro Lys His Leu Arg Asn Lys
130 135 140
Ala Ser Gly Phe Leu Ile Ser Gly Phe Ser Val Gly Ala Val Val Ala
145 150 155 160
Ala Gln Val Tyr Ser Leu Val Val Pro Val Trp Gly Trp Arg Ala Leu
165 170 175
Phe Phe Ile Gly Ile Leu Pro Ile Ile Phe Ala Leu Trp Leu Arg Lys
180 185 190
Asn Ile Pro Glu Ala Glu Asp Trp Lys Glu Lys His Ala Gly Lys Ala
195 200 205
Pro Val Arg Thr Met Val Asp Ile Leu Tyr Arg Gly Glu His Arg Ile
210 215 220
Ala Asn Ile Val Met Thr Leu Ala Ala Ala Thr Ala Leu Trp Phe Cys
225 230 235 240
Phe Ala Gly Asn Leu Gln Asn Ala Ala Ile Val Ala Val Leu Gly Leu
245 250 255
Leu Cys Ala Ala Ile Phe Ile Ser Phe Met Val Gln Ser Ala Gly Lys
260 265 270
Arg Trp Pro Thr Gly Val Met Leu Met Val Val Val Leu Phe Ala Phe
275 280 285
Leu Tyr Ser Trp Pro Ile Gln Ala Leu Leu Pro Thr Tyr Leu Lys Thr
290 295 300
Asp Leu Ala Tyr Asn Pro His Thr Val Ala Asn Val Leu Phe Phe Ser
305 310 315 320
Gly Phe Gly Ala Ala Val Gly Cys Cys Val Gly Gly Phe Leu Gly Asp
325 330 335
Trp Leu Gly Thr Arg Lys Ala Tyr Val Cys Ser Leu Leu Ala Ser Gln
340 345 350
Leu Leu Ile Ile Pro Val Phe Ala Ile Gly Gly Ala Asn Val Trp Val
355 360 365
Leu Gly Leu Leu Leu Phe Phe Gln Gln Met Leu Gly Gln Gly Ile Ala
370 375 380
Gly Ile Leu Pro Lys Leu Ile Gly Gly Tyr Phe Asp Thr Asp Gln Arg
385 390 395 400
Ala Ala Gly Leu Gly Phe Thr Tyr Asn Val Gly Ala Leu Gly Gly Ala
405 410 415
Leu Ala Pro Ile Ile Gly Ala Leu Ile Ala Gln Arg Leu Asp Leu Gly
420 425 430
Thr Ala Leu Ala Ser Leu Ser Phe Ser Leu Thr Phe Val Val Ile Leu
435 440 445
Leu Ile Gly Leu Asp Met Pro Ser Arg Val Gln Arg Trp Leu Arg Pro
450 455 460
Glu Ala Leu Arg Thr His Asp Ala Ile Asp Gly Lys Pro Phe Ser Gly
465 470 475 480
Ala Val Pro Phe Gly Ser Ala Lys Asn Asp Leu Val Lys Thr Lys Ser
485 490 495
<210> 38
<211> 1149
<212> DNA
<213> 大肠杆菌
<400> 38
atgtatgcat taacccaggg ccggatcttt accggccacg aatttcttga tgaccacgcg 60
gttgttatcg ctgatggcct gattaaaagc gtctgtccgg tagcggaact gccgccagag 120
atcgaacaac gttcactgaa cggggccatt ctctcccccg gttttatcga tgtgcagtta 180
aacggctgcg gcggcgtaca gtttaacgac accgctgaag cggtcagcgt ggaaacgctg 240
gaaatcatgc agaaagccaa tgagaaatca ggctgtacta actatctgcc gacgcttatc 300
accaccagcg atgagctgat gaaacagggc gtgcgcgtta tgcgcgagta cctggcaaaa 360
catccgaatc aggcgttagg tctgcatctg gaaggtccgt ggctgaatct ggtaaaaaaa 420
ggcacccata atccgaattt tgtgcgtaag cctgatgccg cgctggtcga tttcctgtgt 480
gaaaacgccg acgtcattac caaagtgacc ctggcaccgg aaatggttcc tgcggaagtc 540
atcagcaaac tggcaaatgc cgggattgtg gtttctgccg gtcactccaa cgcgacgttg 600
aaagaagcaa aagccggttt ccgcgcgggg attacctttg ccacccatct gtacaacgcg 660
atgccgtata ttaccggtcg tgaacctggc ctggcgggcg cgatcctcga cgaagctgac 720
atttattgcg gtattattgc tgatggcctg catgttgatt acgccaacat tcgcaacgct 780
aaacgtctga aaggcgacaa actgtgtctg gttactgacg ccaccgcgcc agcaggtgcc 840
aacattgaac agttcatttt tgcgggtaaa acaatatact accgtaacgg actttgtgtg 900
gatgagaacg gtacgttaag cggttcatcc ttaaccatga ttgaaggcgt gcgtaatctg 960
gtcgaacatt gcggtatcgc actggatgaa gtgctacgta tggcgacgct ctatccggcg 1020
cgtgcgattg gcgttgagaa acgtctcggc acactcgccg caggtaaagt agccaacctg 1080
actgcattca cacctgattt taaaatcacc aagaccatcg ttaacggtaa cgaggtcgta 1140
actcaataa 1149
<210> 39
<211> 382
<212> PRT
<213> 大肠杆菌
<400> 39
Met Tyr Ala Leu Thr Gln Gly Arg Ile Phe Thr Gly His Glu Phe Leu
1 5 10 15
Asp Asp His Ala Val Val Ile Ala Asp Gly Leu Ile Lys Ser Val Cys
20 25 30
Pro Val Ala Glu Leu Pro Pro Glu Ile Glu Gln Arg Ser Leu Asn Gly
35 40 45
Ala Ile Leu Ser Pro Gly Phe Ile Asp Val Gln Leu Asn Gly Cys Gly
50 55 60
Gly Val Gln Phe Asn Asp Thr Ala Glu Ala Val Ser Val Glu Thr Leu
65 70 75 80
Glu Ile Met Gln Lys Ala Asn Glu Lys Ser Gly Cys Thr Asn Tyr Leu
85 90 95
Pro Thr Leu Ile Thr Thr Ser Asp Glu Leu Met Lys Gln Gly Val Arg
100 105 110
Val Met Arg Glu Tyr Leu Ala Lys His Pro Asn Gln Ala Leu Gly Leu
115 120 125
His Leu Glu Gly Pro Trp Leu Asn Leu Val Lys Lys Gly Thr His Asn
130 135 140
Pro Asn Phe Val Arg Lys Pro Asp Ala Ala Leu Val Asp Phe Leu Cys
145 150 155 160
Glu Asn Ala Asp Val Ile Thr Lys Val Thr Leu Ala Pro Glu Met Val
165 170 175
Pro Ala Glu Val Ile Ser Lys Leu Ala Asn Ala Gly Ile Val Val Ser
180 185 190
Ala Gly His Ser Asn Ala Thr Leu Lys Glu Ala Lys Ala Gly Phe Arg
195 200 205
Ala Gly Ile Thr Phe Ala Thr His Leu Tyr Asn Ala Met Pro Tyr Ile
210 215 220
Thr Gly Arg Glu Pro Gly Leu Ala Gly Ala Ile Leu Asp Glu Ala Asp
225 230 235 240
Ile Tyr Cys Gly Ile Ile Ala Asp Gly Leu His Val Asp Tyr Ala Asn
245 250 255
Ile Arg Asn Ala Lys Arg Leu Lys Gly Asp Lys Leu Cys Leu Val Thr
260 265 270
Asp Ala Thr Ala Pro Ala Gly Ala Asn Ile Glu Gln Phe Ile Phe Ala
275 280 285
Gly Lys Thr Ile Tyr Tyr Arg Asn Gly Leu Cys Val Asp Glu Asn Gly
290 295 300
Thr Leu Ser Gly Ser Ser Leu Thr Met Ile Glu Gly Val Arg Asn Leu
305 310 315 320
Val Glu His Cys Gly Ile Ala Leu Asp Glu Val Leu Arg Met Ala Thr
325 330 335
Leu Tyr Pro Ala Arg Ala Ile Gly Val Glu Lys Arg Leu Gly Thr Leu
340 345 350
Ala Ala Gly Lys Val Ala Asn Leu Thr Ala Phe Thr Pro Asp Phe Lys
355 360 365
Ile Thr Lys Thr Ile Val Asn Gly Asn Glu Val Val Thr Gln
370 375 380
<210> 40
<211> 801
<212> DNA
<213> 大肠杆菌
<400> 40
atgagactga tccccctgac taccgctgaa caggtcggca aatgggctgc tcgccatatc 60
gtcaatcgta tcaatgcgtt caaaccgact gccgatcgtc cgtttgtact gggcctgccg 120
actggcggca cgccgatgac cacctataaa gcgttagtcg aaatgcataa agcaggccag 180
gtcagcttta agcacgttgt caccttcaac atggacgaat atgtcggtct gccgaaagag 240
catccggaaa gctactacag ctttatgcac cgtaatttct tcgatcacgt tgatattcca 300
gcagaaaaca tcaaccttct caacggcaac gccccggata tcgacgccga gtgccgccag 360
tatgaagaaa aaatccgttc ttacggaaaa attcatctgt ttatgggcgg tgtaggtaac 420
gacggtcata ttgcatttaa cgaaccggcg tcttctctgg cttctcgtac tcgtatcaaa 480
accctgactc atgacactcg cgtcgcaaac tctcgtttct ttgataacga tgttaatcag 540
gtgccaaaat atgccctgac tgtcggtgtt ggtacactgc tggatgccga agaagtgatg 600
attctggtgc tgggtagcca gaaagcactg gcgctgcagg ccgccgttga aggttgcgtg 660
aaccatatgt ggaccatcag ctgtctgcaa ctgcatccga aagcgatcat ggtgtgcgat 720
gaaccttcca ccatggagct gaaagttaag actttaagat atttcaatga attagaagca 780
gaaaatatca aaggtctgta a 801
<210> 41
<211> 266
<212> PRT
<213> 大肠杆菌
<400> 41
Met Arg Leu Ile Pro Leu Thr Thr Ala Glu Gln Val Gly Lys Trp Ala
1 5 10 15
Ala Arg His Ile Val Asn Arg Ile Asn Ala Phe Lys Pro Thr Ala Asp
20 25 30
Arg Pro Phe Val Leu Gly Leu Pro Thr Gly Gly Thr Pro Met Thr Thr
35 40 45
Tyr Lys Ala Leu Val Glu Met His Lys Ala Gly Gln Val Ser Phe Lys
50 55 60
His Val Val Thr Phe Asn Met Asp Glu Tyr Val Gly Leu Pro Lys Glu
65 70 75 80
His Pro Glu Ser Tyr Tyr Ser Phe Met His Arg Asn Phe Phe Asp His
85 90 95
Val Asp Ile Pro Ala Glu Asn Ile Asn Leu Leu Asn Gly Asn Ala Pro
100 105 110
Asp Ile Asp Ala Glu Cys Arg Gln Tyr Glu Glu Lys Ile Arg Ser Tyr
115 120 125
Gly Lys Ile His Leu Phe Met Gly Gly Val Gly Asn Asp Gly His Ile
130 135 140
Ala Phe Asn Glu Pro Ala Ser Ser Leu Ala Ser Arg Thr Arg Ile Lys
145 150 155 160
Thr Leu Thr His Asp Thr Arg Val Ala Asn Ser Arg Phe Phe Asp Asn
165 170 175
Asp Val Asn Gln Val Pro Lys Tyr Ala Leu Thr Val Gly Val Gly Thr
180 185 190
Leu Leu Asp Ala Glu Glu Val Met Ile Leu Val Leu Gly Ser Gln Lys
195 200 205
Ala Leu Ala Leu Gln Ala Ala Val Glu Gly Cys Val Asn His Met Trp
210 215 220
Thr Ile Ser Cys Leu Gln Leu His Pro Lys Ala Ile Met Val Cys Asp
225 230 235 240
Glu Pro Ser Thr Met Glu Leu Lys Val Lys Thr Leu Arg Tyr Phe Asn
245 250 255
Glu Leu Glu Ala Glu Asn Ile Lys Gly Leu
260 265
<210> 42
<211> 1947
<212> DNA
<213> 大肠杆菌
<400> 42
atgaatattt taggtttttt ccagcgactc ggtagggcgt tacagctccc tatcgcggtg 60
ctgccggtgg cggcactgtt gctgcgattc ggtcagccag atttacttaa cgttgcgttt 120
attgcccagg cgggcggtgc gatttttgat aacctcgcat taatcttcgc catcggtgtg 180
gcatccagct ggtcgaaaga cagcgctggt gcggcggcgc tggcgggtgc ggtaggttac 240
tttgtgttaa ccaaagcgat ggtgaccatc aacccagaaa ttaacatggg tgtactggcg 300
ggtatcatta ccggtctggt tggtggcgca gcctataacc gttggtccga tattaaactg 360
ccggacttcc tgagcttctt cggcggcaaa cgctttgtgc cgattgccac cggattcttc 420
tgcctggtgc tggcggccat ttttggttac gtctggccgc cggtacagca cgctatccat 480
gcaggcggcg agtggatcgt ttctgcgggc gcgctgggtt ccggtatctt tggtttcatc 540
aaccgtctgc tgatcccaac cggtctgcat caggtactga acaccatcgc ctggttccag 600
attggtgaat tcaccaacgc ggcgggtacg gttttccacg gtgacattaa ccgcttctat 660
gccggtgacg gcaccgcggg gatgttcatg tccggcttct tcccgatcat gatgttcggt 720
ctgccgggtg cggcgctggc gatgtacttc gcagcaccga aagagcgtcg tccgatggtt 780
ggcggtatgc tgctttctgt tgctgttact gcgttcctga ccggtgtgac tgagccgctg 840
gaattcctgt tcatgttcct tgctccgctg ctgtacctcc tgcacgcact gctgaccggt 900
atcagcctgt ttgtggcaac gctgctgggt atccacgcgg gcttctcttt ctctgcgggg 960
gctatcgact acgcgttgat gtataacctg ccggccgcca gccagaacgt ctggatgctg 1020
ctggtgatgg gcgttatctt cttcgctatc tacttcgtgg tgttcagttt ggttatccgc 1080
atgttcaacc tgaaaacgcc gggtcgtgaa gataaagaag acgagatcgt tactgaagaa 1140
gccaacagca acactgaaga aggtctgact caactggcaa ccaactatat tgctgcggtt 1200
ggcggcactg acaacctgaa agcgattgac gcctgtatca cccgtctgcg ccttacagtg 1260
gctgactctg cccgcgttaa cgatacgatg tgtaaacgtc tgggtgcttc tggggtagtg 1320
aaactgaaca aacagactat tcaggtgatt gttggcgcga aagcagaatc catcggcgat 1380
gcgatgaaga aagtcgttgc ccgtggtccg gtagccgctg cgtcagctga agcaactccg 1440
gcaactgccg cgcctgtagc aaaaccgcag gctgtaccaa acgcggtatc tatcgcggag 1500
ctggtatcgc cgattaccgg tgatgtcgtg gcactggatc aggttcctga cgaagcattc 1560
gccagcaaag cggtgggtga cggtgtggcg gtgaaaccga cagataaaat cgtcgtatca 1620
ccagccgcag ggacaatcgt gaaaatcttc aacaccaacc acgcgttctg cctggaaacc 1680
gaaaaaggcg cggagatcgt cgtccatatg ggtatcgaca ccgtagcgct ggaaggtaaa 1740
ggctttaaac gtctggtgga agagggtgcg caggtaagcg cagggcaacc gattctggaa 1800
atggatctgg attacctgaa cgctaacgcc cgctcgatga ttagcccggt ggtttgcagc 1860
aatatcgacg atttcagtgg cttgatcatt aaagctcagg gccatattgt ggcgggtcaa 1920
acaccgctgt atgaaatcaa aaagtaa 1947
<210> 43
<211> 648
<212> PRT
<213> 大肠杆菌
<400> 43
Met Asn Ile Leu Gly Phe Phe Gln Arg Leu Gly Arg Ala Leu Gln Leu
1 5 10 15
Pro Ile Ala Val Leu Pro Val Ala Ala Leu Leu Leu Arg Phe Gly Gln
20 25 30
Pro Asp Leu Leu Asn Val Ala Phe Ile Ala Gln Ala Gly Gly Ala Ile
35 40 45
Phe Asp Asn Leu Ala Leu Ile Phe Ala Ile Gly Val Ala Ser Ser Trp
50 55 60
Ser Lys Asp Ser Ala Gly Ala Ala Ala Leu Ala Gly Ala Val Gly Tyr
65 70 75 80
Phe Val Leu Thr Lys Ala Met Val Thr Ile Asn Pro Glu Ile Asn Met
85 90 95
Gly Val Leu Ala Gly Ile Ile Thr Gly Leu Val Gly Gly Ala Ala Tyr
100 105 110
Asn Arg Trp Ser Asp Ile Lys Leu Pro Asp Phe Leu Ser Phe Phe Gly
115 120 125
Gly Lys Arg Phe Val Pro Ile Ala Thr Gly Phe Phe Cys Leu Val Leu
130 135 140
Ala Ala Ile Phe Gly Tyr Val Trp Pro Pro Val Gln His Ala Ile His
145 150 155 160
Ala Gly Gly Glu Trp Ile Val Ser Ala Gly Ala Leu Gly Ser Gly Ile
165 170 175
Phe Gly Phe Ile Asn Arg Leu Leu Ile Pro Thr Gly Leu His Gln Val
180 185 190
Leu Asn Thr Ile Ala Trp Phe Gln Ile Gly Glu Phe Thr Asn Ala Ala
195 200 205
Gly Thr Val Phe His Gly Asp Ile Asn Arg Phe Tyr Ala Gly Asp Gly
210 215 220
Thr Ala Gly Met Phe Met Ser Gly Phe Phe Pro Ile Met Met Phe Gly
225 230 235 240
Leu Pro Gly Ala Ala Leu Ala Met Tyr Phe Ala Ala Pro Lys Glu Arg
245 250 255
Arg Pro Met Val Gly Gly Met Leu Leu Ser Val Ala Val Thr Ala Phe
260 265 270
Leu Thr Gly Val Thr Glu Pro Leu Glu Phe Leu Phe Met Phe Leu Ala
275 280 285
Pro Leu Leu Tyr Leu Leu His Ala Leu Leu Thr Gly Ile Ser Leu Phe
290 295 300
Val Ala Thr Leu Leu Gly Ile His Ala Gly Phe Ser Phe Ser Ala Gly
305 310 315 320
Ala Ile Asp Tyr Ala Leu Met Tyr Asn Leu Pro Ala Ala Ser Gln Asn
325 330 335
Val Trp Met Leu Leu Val Met Gly Val Ile Phe Phe Ala Ile Tyr Phe
340 345 350
Val Val Phe Ser Leu Val Ile Arg Met Phe Asn Leu Lys Thr Pro Gly
355 360 365
Arg Glu Asp Lys Glu Asp Glu Ile Val Thr Glu Glu Ala Asn Ser Asn
370 375 380
Thr Glu Glu Gly Leu Thr Gln Leu Ala Thr Asn Tyr Ile Ala Ala Val
385 390 395 400
Gly Gly Thr Asp Asn Leu Lys Ala Ile Asp Ala Cys Ile Thr Arg Leu
405 410 415
Arg Leu Thr Val Ala Asp Ser Ala Arg Val Asn Asp Thr Met Cys Lys
420 425 430
Arg Leu Gly Ala Ser Gly Val Val Lys Leu Asn Lys Gln Thr Ile Gln
435 440 445
Val Ile Val Gly Ala Lys Ala Glu Ser Ile Gly Asp Ala Met Lys Lys
450 455 460
Val Val Ala Arg Gly Pro Val Ala Ala Ala Ser Ala Glu Ala Thr Pro
465 470 475 480
Ala Thr Ala Ala Pro Val Ala Lys Pro Gln Ala Val Pro Asn Ala Val
485 490 495
Ser Ile Ala Glu Leu Val Ser Pro Ile Thr Gly Asp Val Val Ala Leu
500 505 510
Asp Gln Val Pro Asp Glu Ala Phe Ala Ser Lys Ala Val Gly Asp Gly
515 520 525
Val Ala Val Lys Pro Thr Asp Lys Ile Val Val Ser Pro Ala Ala Gly
530 535 540
Thr Ile Val Lys Ile Phe Asn Thr Asn His Ala Phe Cys Leu Glu Thr
545 550 555 560
Glu Lys Gly Ala Glu Ile Val Val His Met Gly Ile Asp Thr Val Ala
565 570 575
Leu Glu Gly Lys Gly Phe Lys Arg Leu Val Glu Glu Gly Ala Gln Val
580 585 590
Ser Ala Gly Gln Pro Ile Leu Glu Met Asp Leu Asp Tyr Leu Asn Ala
595 600 605
Asn Ala Arg Ser Met Ile Ser Pro Val Val Cys Ser Asn Ile Asp Asp
610 615 620
Phe Ser Gly Leu Ile Ile Lys Ala Gln Gly His Ile Val Ala Gly Gln
625 630 635 640
Thr Pro Leu Tyr Glu Ile Lys Lys
645
<210> 44
<211> 972
<212> DNA
<213> 大肠杆菌
<400> 44
atgaccattg ctattgttat aggcacacat ggttgggctg cagagcagtt gcttaaaacg 60
gcagaaatgc tgttaggcga gcaggaaaac gtcggctgga tcgatttcgt tccaggtgaa 120
aatgccgaaa cgctgattga aaagtacaac gctcagttgg caaaactcga caccactaaa 180
ggcgtgctgt ttctcgttga tacatgggga ggcagcccgt tcaatgctgc cagccgcatt 240
gtcgtcgaca aagagcatta tgaagtcatt gcaggcgtta acattccaat gctcgtggaa 300
acgttaatgg cccgtgatga tgacccaagc tttgatgaac tggtggcact ggcagtagaa 360
acaggccgtg aaggcgtgaa agcactgaaa gccaaaccgg ttgaaaaagc cgcgccagca 420
cccgctgccg cagcaccaaa agcggctcca actccggcaa aaccaatggg gccaaacgac 480
tacatggtta ttggccttgc gcgtatcgac gaccgtctga ttcacggtca ggtcgccacc 540
cgctggacca aagaaaccaa tgtctcccgt attattgttg ttagtgatga agtggctgcg 600
gataccgttc gtaagacact gctcacccag gttgcacctc cgggcgtaac agcacacgta 660
gttgatgttg ccaaaatgat tcgcgtctac aacaacccga aatatgctgg cgaacgcgta 720
atgctgttat ttaccaaccc aacagatgta gagcgtctcg ttgaaggcgg cgtgaaaatc 780
acctctgtta acgtcggtgg tatggcattc cgtcagggta aaacccaggt gaataacgcg 840
gtttcggttg atgaaaaaga tatcgaggcg ttcaagaaac tgaatgcgcg cggtattgag 900
ctggaagtcc gtaaggtttc caccgatccg aaactgaaaa tgatggatct gatcagcaaa 960
atcgataagt aa 972
<210> 45
<211> 323
<212> PRT
<213> 大肠杆菌
<400> 45
Met Thr Ile Ala Ile Val Ile Gly Thr His Gly Trp Ala Ala Glu Gln
1 5 10 15
Leu Leu Lys Thr Ala Glu Met Leu Leu Gly Glu Gln Glu Asn Val Gly
20 25 30
Trp Ile Asp Phe Val Pro Gly Glu Asn Ala Glu Thr Leu Ile Glu Lys
35 40 45
Tyr Asn Ala Gln Leu Ala Lys Leu Asp Thr Thr Lys Gly Val Leu Phe
50 55 60
Leu Val Asp Thr Trp Gly Gly Ser Pro Phe Asn Ala Ala Ser Arg Ile
65 70 75 80
Val Val Asp Lys Glu His Tyr Glu Val Ile Ala Gly Val Asn Ile Pro
85 90 95
Met Leu Val Glu Thr Leu Met Ala Arg Asp Asp Asp Pro Ser Phe Asp
100 105 110
Glu Leu Val Ala Leu Ala Val Glu Thr Gly Arg Glu Gly Val Lys Ala
115 120 125
Leu Lys Ala Lys Pro Val Glu Lys Ala Ala Pro Ala Pro Ala Ala Ala
130 135 140
Ala Pro Lys Ala Ala Pro Thr Pro Ala Lys Pro Met Gly Pro Asn Asp
145 150 155 160
Tyr Met Val Ile Gly Leu Ala Arg Ile Asp Asp Arg Leu Ile His Gly
165 170 175
Gln Val Ala Thr Arg Trp Thr Lys Glu Thr Asn Val Ser Arg Ile Ile
180 185 190
Val Val Ser Asp Glu Val Ala Ala Asp Thr Val Arg Lys Thr Leu Leu
195 200 205
Thr Gln Val Ala Pro Pro Gly Val Thr Ala His Val Val Asp Val Ala
210 215 220
Lys Met Ile Arg Val Tyr Asn Asn Pro Lys Tyr Ala Gly Glu Arg Val
225 230 235 240
Met Leu Leu Phe Thr Asn Pro Thr Asp Val Glu Arg Leu Val Glu Gly
245 250 255
Gly Val Lys Ile Thr Ser Val Asn Val Gly Gly Met Ala Phe Arg Gln
260 265 270
Gly Lys Thr Gln Val Asn Asn Ala Val Ser Val Asp Glu Lys Asp Ile
275 280 285
Glu Ala Phe Lys Lys Leu Asn Ala Arg Gly Ile Glu Leu Glu Val Arg
290 295 300
Lys Val Ser Thr Asp Pro Lys Leu Lys Met Met Asp Leu Ile Ser Lys
305 310 315 320
Ile Asp Lys
<210> 46
<211> 801
<212> DNA
<213> 大肠杆菌
<400> 46
atggagatta ccactcttca aattgtgctg gtatttatcg tagcctgtat cgcaggtatg 60
ggatcaatcc tcgatgaatt tcagtttcac cgtccgctaa tcgcgtgtac cctggtgggt 120
atcgttcttg gggatatgaa aaccggtatt attatcggtg gtacgctgga aatgatcgcg 180
ctgggctgga tgaacatcgg tgctgcagtt gcgcctgacg ccgctctggc ttctatcatt 240
tctaccattc tggttatcgc aggtcatcag agcattggtg caggtatcgc actggcaatc 300
cctctggccg ctgcgggcca ggtactgacc atcatcgttc gtactattac cgttgctttc 360
cagcacgctg cggataaggc tgctgataac ggcaacctga cagcgatttc ctggatccac 420
gtttcttctc tgttcctgca agcaatgcgt gtggctattc cggccgtcat cgttgcgctg 480
tctgttggta ccagcgaagt acagaacatg ctgaatgcga ttccggaagt ggtgaccaat 540
ggtctgaata tcgccggtgg catgatcgtg gtggttggtt atgcgatggt tatcaacatg 600
atgcgtgctg gctacctgat gccgttcttc tacctcggct tcgtaaccgc agcattcacc 660
aactttaacc tggttgctct gggtgtgatt ggtactgtta tggcagtgct ctacatccaa 720
cttagcccga aatacaaccg cgtagccggt gcgcctgctc aggcagctgg taacaacgat 780
ctcgataacg aactggacta a 801
<210> 47
<211> 266
<212> PRT
<213> 大肠杆菌
<400> 47
Met Glu Ile Thr Thr Leu Gln Ile Val Leu Val Phe Ile Val Ala Cys
1 5 10 15
Ile Ala Gly Met Gly Ser Ile Leu Asp Glu Phe Gln Phe His Arg Pro
20 25 30
Leu Ile Ala Cys Thr Leu Val Gly Ile Val Leu Gly Asp Met Lys Thr
35 40 45
Gly Ile Ile Ile Gly Gly Thr Leu Glu Met Ile Ala Leu Gly Trp Met
50 55 60
Asn Ile Gly Ala Ala Val Ala Pro Asp Ala Ala Leu Ala Ser Ile Ile
65 70 75 80
Ser Thr Ile Leu Val Ile Ala Gly His Gln Ser Ile Gly Ala Gly Ile
85 90 95
Ala Leu Ala Ile Pro Leu Ala Ala Ala Gly Gln Val Leu Thr Ile Ile
100 105 110
Val Arg Thr Ile Thr Val Ala Phe Gln His Ala Ala Asp Lys Ala Ala
115 120 125
Asp Asn Gly Asn Leu Thr Ala Ile Ser Trp Ile His Val Ser Ser Leu
130 135 140
Phe Leu Gln Ala Met Arg Val Ala Ile Pro Ala Val Ile Val Ala Leu
145 150 155 160
Ser Val Gly Thr Ser Glu Val Gln Asn Met Leu Asn Ala Ile Pro Glu
165 170 175
Val Val Thr Asn Gly Leu Asn Ile Ala Gly Gly Met Ile Val Val Val
180 185 190
Gly Tyr Ala Met Val Ile Asn Met Met Arg Ala Gly Tyr Leu Met Pro
195 200 205
Phe Phe Tyr Leu Gly Phe Val Thr Ala Ala Phe Thr Asn Phe Asn Leu
210 215 220
Val Ala Leu Gly Val Ile Gly Thr Val Met Ala Val Leu Tyr Ile Gln
225 230 235 240
Leu Ser Pro Lys Tyr Asn Arg Val Ala Gly Ala Pro Ala Gln Ala Ala
245 250 255
Gly Asn Asn Asp Leu Asp Asn Glu Leu Asp
260 265
<210> 48
<211> 852
<212> DNA
<213> 大肠杆菌
<400> 48
atggttgata caactcaaac taccaccgag aaaaaactca ctcaaagtga tattcgtggc 60
gtcttcctgc gttctaacct cttccagggt tcatggaact tcgaacgtat gcaggcactg 120
ggtttctgct tctctatggt accggcaatt cgtcgcctct accctgagaa caacgaagct 180
cgtaaacaag ctattcgccg tcacctggag ttctttaaca cccagccgtt cgtggctgcg 240
ccgattctcg gcgtaaccct ggcgctggaa gaacagcgtg ctaatggcgc agagatcgac 300
gacggtgcta tcaacggtat caaagtcggt ttgatggggc cactggctgg tgtaggcgac 360
ccgatcttct ggggaaccgt acgtccggta tttgcagcac tgggtgccgg tatcgcgatg 420
agcggcagcc tgttaggtcc gctgctgttc ttcatcctgt ttaacctggt gcgtctggca 480
acccgttact acggcgtagc gtatggttac tccaaaggta tcgatatcgt taaagatatg 540
ggtggtggct tcctgcaaaa actgacggaa ggggcgtcta tcctcggcct gtttgtcatg 600
ggggcattgg ttaacaagtg gacacatgtc aacatcccgc tggttgtctc tcgcattact 660
gaccagacgg gcaaagaaca cgttactact gtccagacta ttctggacca gttaatgcca 720
ggcctggtac cactgctgct gacctttgct tgtatgtggc tactgcgcaa aaaagttaac 780
ccgctgtgga tcatcgttgg cttcttcgtc atcggtatcg ctggttacgc ttgcggcctg 840
ctgggactgt aa 852
<210> 49
<211> 283
<212> PRT
<213> 大肠杆菌
<400> 49
Met Val Asp Thr Thr Gln Thr Thr Thr Glu Lys Lys Leu Thr Gln Ser
1 5 10 15
Asp Ile Arg Gly Val Phe Leu Arg Ser Asn Leu Phe Gln Gly Ser Trp
20 25 30
Asn Phe Glu Arg Met Gln Ala Leu Gly Phe Cys Phe Ser Met Val Pro
35 40 45
Ala Ile Arg Arg Leu Tyr Pro Glu Asn Asn Glu Ala Arg Lys Gln Ala
50 55 60
Ile Arg Arg His Leu Glu Phe Phe Asn Thr Gln Pro Phe Val Ala Ala
65 70 75 80
Pro Ile Leu Gly Val Thr Leu Ala Leu Glu Glu Gln Arg Ala Asn Gly
85 90 95
Ala Glu Ile Asp Asp Gly Ala Ile Asn Gly Ile Lys Val Gly Leu Met
100 105 110
Gly Pro Leu Ala Gly Val Gly Asp Pro Ile Phe Trp Gly Thr Val Arg
115 120 125
Pro Val Phe Ala Ala Leu Gly Ala Gly Ile Ala Met Ser Gly Ser Leu
130 135 140
Leu Gly Pro Leu Leu Phe Phe Ile Leu Phe Asn Leu Val Arg Leu Ala
145 150 155 160
Thr Arg Tyr Tyr Gly Val Ala Tyr Gly Tyr Ser Lys Gly Ile Asp Ile
165 170 175
Val Lys Asp Met Gly Gly Gly Phe Leu Gln Lys Leu Thr Glu Gly Ala
180 185 190
Ser Ile Leu Gly Leu Phe Val Met Gly Ala Leu Val Asn Lys Trp Thr
195 200 205
His Val Asn Ile Pro Leu Val Val Ser Arg Ile Thr Asp Gln Thr Gly
210 215 220
Lys Glu His Val Thr Thr Val Gln Thr Ile Leu Asp Gln Leu Met Pro
225 230 235 240
Gly Leu Val Pro Leu Leu Leu Thr Phe Ala Cys Met Trp Leu Leu Arg
245 250 255
Lys Lys Val Asn Pro Leu Trp Ile Ile Val Gly Phe Phe Val Ile Gly
260 265 270
Ile Ala Gly Tyr Ala Cys Gly Leu Leu Gly Leu
275 280
<210> 50
<211> 1434
<212> DNA
<213> 大肠杆菌
<400> 50
atgtttaaga atgcatttgc taacctgcaa aaggtcggta aatcgctgat gctgccggta 60
tccgtactgc ctatcgcagg tattctgctg ggcgtcggtt ccgcgaattt cagctggctg 120
cccgccgttg tatcgcatgt tatggcagaa gcaggcggtt ccgtctttgc aaacatgcca 180
ctgatttttg cgatcggtgt cgccctcggc tttaccaata acgatggcgt atccgcgctg 240
gccgcagttg ttgcctatgg catcatggtt aaaaccatgg ccgtggttgc gccactggta 300
ctgcatttac ctgctgaaga aatcgcctct aaacacctgg cggatactgg cgtactcgga 360
gggattatct ccggtgcgat cgcagcgtac atgtttaacc gtttctaccg tattaagctg 420
cctgagtatc ttggcttctt tgccggtaaa cgctttgtgc cgatcatttc tggcctggct 480
gccatcttta ctggcgttgt gctgtccttc atttggccgc cgattggttc tgcaatccag 540
accttctctc agtgggctgc ttaccagaac ccggtagttg cgtttggcat ttacggtttc 600
atcgaacgtt gcctggtacc gtttggtctg caccacatct ggaacgtacc tttccagatg 660
cagattggtg aatacaccaa cgcagcaggt caggttttcc acggcgacat tccgcgttat 720
atggcgggtg acccgactgc gggtaaactg tctggtggct tcctgttcaa aatgtacggt 780
ctgccagctg ccgcaattgc tatctggcac tctgctaaac cagaaaaccg cgcgaaagtg 840
ggcggtatta tgatctccgc ggcgctgacc tcgttcctga ccggtatcac cgagccgatc 900
gagttctcct tcatgttcgt tgcgccgatc ctgtacatca tccacgcgat tctggcaggc 960
ctggcattcc caatctgtat tcttctgggg atgcgtgacg gtacgtcgtt ctcgcacggt 1020
ctgatcgact tcatcgttct gtctggtaac agcagcaaac tgtggctgtt cccgatcgtc 1080
ggtatcggtt atgcgattgt ttactacacc atcttccgcg tgctgattaa agcactggat 1140
ctgaaaacgc cgggtcgtga agacgcgact gaagatgcaa aagcgacagg taccagcgaa 1200
atggcaccgg ctctggttgc tgcatttggt ggtaaagaaa acattactaa cctcgacgca 1260
tgtattaccc gtctgcgcgt cagcgttgct gatgtgtcta aagtggatca ggccggcctg 1320
aagaaactgg gcgcagcggg cgtagtggtt gctggttctg gtgttcaggc gattttcggt 1380
actaaatccg ataacctgaa aaccgagatg gatgagtaca tccgtaacca ctaa 1434
<210> 51
<211> 477
<212> PRT
<213> 大肠杆菌
<400> 51
Met Phe Lys Asn Ala Phe Ala Asn Leu Gln Lys Val Gly Lys Ser Leu
1 5 10 15
Met Leu Pro Val Ser Val Leu Pro Ile Ala Gly Ile Leu Leu Gly Val
20 25 30
Gly Ser Ala Asn Phe Ser Trp Leu Pro Ala Val Val Ser His Val Met
35 40 45
Ala Glu Ala Gly Gly Ser Val Phe Ala Asn Met Pro Leu Ile Phe Ala
50 55 60
Ile Gly Val Ala Leu Gly Phe Thr Asn Asn Asp Gly Val Ser Ala Leu
65 70 75 80
Ala Ala Val Val Ala Tyr Gly Ile Met Val Lys Thr Met Ala Val Val
85 90 95
Ala Pro Leu Val Leu His Leu Pro Ala Glu Glu Ile Ala Ser Lys His
100 105 110
Leu Ala Asp Thr Gly Val Leu Gly Gly Ile Ile Ser Gly Ala Ile Ala
115 120 125
Ala Tyr Met Phe Asn Arg Phe Tyr Arg Ile Lys Leu Pro Glu Tyr Leu
130 135 140
Gly Phe Phe Ala Gly Lys Arg Phe Val Pro Ile Ile Ser Gly Leu Ala
145 150 155 160
Ala Ile Phe Thr Gly Val Val Leu Ser Phe Ile Trp Pro Pro Ile Gly
165 170 175
Ser Ala Ile Gln Thr Phe Ser Gln Trp Ala Ala Tyr Gln Asn Pro Val
180 185 190
Val Ala Phe Gly Ile Tyr Gly Phe Ile Glu Arg Cys Leu Val Pro Phe
195 200 205
Gly Leu His His Ile Trp Asn Val Pro Phe Gln Met Gln Ile Gly Glu
210 215 220
Tyr Thr Asn Ala Ala Gly Gln Val Phe His Gly Asp Ile Pro Arg Tyr
225 230 235 240
Met Ala Gly Asp Pro Thr Ala Gly Lys Leu Ser Gly Gly Phe Leu Phe
245 250 255
Lys Met Tyr Gly Leu Pro Ala Ala Ala Ile Ala Ile Trp His Ser Ala
260 265 270
Lys Pro Glu Asn Arg Ala Lys Val Gly Gly Ile Met Ile Ser Ala Ala
275 280 285
Leu Thr Ser Phe Leu Thr Gly Ile Thr Glu Pro Ile Glu Phe Ser Phe
290 295 300
Met Phe Val Ala Pro Ile Leu Tyr Ile Ile His Ala Ile Leu Ala Gly
305 310 315 320
Leu Ala Phe Pro Ile Cys Ile Leu Leu Gly Met Arg Asp Gly Thr Ser
325 330 335
Phe Ser His Gly Leu Ile Asp Phe Ile Val Leu Ser Gly Asn Ser Ser
340 345 350
Lys Leu Trp Leu Phe Pro Ile Val Gly Ile Gly Tyr Ala Ile Val Tyr
355 360 365
Tyr Thr Ile Phe Arg Val Leu Ile Lys Ala Leu Asp Leu Lys Thr Pro
370 375 380
Gly Arg Glu Asp Ala Thr Glu Asp Ala Lys Ala Thr Gly Thr Ser Glu
385 390 395 400
Met Ala Pro Ala Leu Val Ala Ala Phe Gly Gly Lys Glu Asn Ile Thr
405 410 415
Asn Leu Asp Ala Cys Ile Thr Arg Leu Arg Val Ser Val Ala Asp Val
420 425 430
Ser Lys Val Asp Gln Ala Gly Leu Lys Lys Leu Gly Ala Ala Gly Val
435 440 445
Val Val Ala Gly Ser Gly Val Gln Ala Ile Phe Gly Thr Lys Ser Asp
450 455 460
Asn Leu Lys Thr Glu Met Asp Glu Tyr Ile Arg Asn His
465 470 475
<210> 52
<211> 510
<212> DNA
<213> 大肠杆菌
<400> 52
atgggtttgt tcgataaact gaaatctctg gtttccgacg acaagaagga taccggaact 60
attgagatca ttgctccgct ctctggcgag atcgtcaata tcgaagacgt gccggatgtc 120
gtttttgcgg aaaaaatcgt tggtgatggt attgctatca aaccaacggg taacaaaatg 180
gtcgcgccag tagacggcac cattggtaaa atctttgaaa ccaaccacgc attctctatc 240
gaatctgata gcggcgttga actgttcgtc cacttcggta tcgacaccgt tgaactgaaa 300
ggcgaaggct tcaagcgtat tgctgaagaa ggtcagcgcg tgaaagttgg cgatactgtc 360
attgaatttg atctgccgct gctggaagag aaagccaagt ctaccctgac tccggttgtt 420
atctccaaca tggacgaaat caaagaactg atcaaactgt ccggtagcgt aaccgtgggt 480
gaaaccccgg ttatccgcat caagaagtaa 510
<210> 53
<211> 169
<212> PRT
<213> 大肠杆菌
<400> 53
Met Gly Leu Phe Asp Lys Leu Lys Ser Leu Val Ser Asp Asp Lys Lys
1 5 10 15
Asp Thr Gly Thr Ile Glu Ile Ile Ala Pro Leu Ser Gly Glu Ile Val
20 25 30
Asn Ile Glu Asp Val Pro Asp Val Val Phe Ala Glu Lys Ile Val Gly
35 40 45
Asp Gly Ile Ala Ile Lys Pro Thr Gly Asn Lys Met Val Ala Pro Val
50 55 60
Asp Gly Thr Ile Gly Lys Ile Phe Glu Thr Asn His Ala Phe Ser Ile
65 70 75 80
Glu Ser Asp Ser Gly Val Glu Leu Phe Val His Phe Gly Ile Asp Thr
85 90 95
Val Glu Leu Lys Gly Glu Gly Phe Lys Arg Ile Ala Glu Glu Gly Gln
100 105 110
Arg Val Lys Val Gly Asp Thr Val Ile Glu Phe Asp Leu Pro Leu Leu
115 120 125
Glu Glu Lys Ala Lys Ser Thr Leu Thr Pro Val Val Ile Ser Asn Met
130 135 140
Asp Glu Ile Lys Glu Leu Ile Lys Leu Ser Gly Ser Val Thr Val Gly
145 150 155 160
Glu Thr Pro Val Ile Arg Ile Lys Lys
165
<210> 54
<211> 1248
<212> DNA
<213> 大肠杆菌
<400> 54
atggcactga atattccatt cagaaatgcg tactatcgtt ttgcatccag ttactcattt 60
ctctttttta tttcctggtc gctgtggtgg tcgttatacg ctatttggct gaaaggacat 120
ctaggattaa cagggacgga attaggtaca ctttattcgg tcaaccagtt taccagcatt 180
ctatttatga tgttctacgg catcgttcag gataaactcg gtctgaagaa accgctcatc 240
tggtgtatga gtttcattct ggtcttgacc ggaccgttta tgatttacgt ttatgaaccg 300
ttactgcaaa gcaatttttc tgtaggtcta attctggggg cgctcttttt tggcctgggg 360
tatctggcgg gatgcggttt gcttgacagc ttcaccgaaa aaatggcgcg aaattttcat 420
ttcgaatatg gaacagcgcg cgcctgggga tcttttggct atgctattgg cgcgttcttt 480
gccggtatat tttttagtat cagtccccat atcaacttct ggttggtctc gctatttggc 540
gctgtattta tgatgatcaa catgcgtttt aaagataagg atcaccagtg catagcggcg 600
gatgcgggag gggtaaaaaa agaggatttt atcgcagttt tcaaggatcg aaacttctgg 660
gttttcgtca tatttattgt ggggacgtgg tctttctata acatttttga tcaacaactc 720
tttcctgtct tttatgcagg tttattcgaa tcacacgatg taggaacgcg cctgtatggt 780
tatctcaact cattccaggt ggtactcgaa gcgctgtgca tggcgattat tcctttcttt 840
gtgaatcggg tagggccaaa aaatgcatta cttatcggtg ttgtgattat ggcgttgcgt 900
atcctttcct gcgcgttgtt cgttaacccc tggattattt cattagtgaa gctgttacat 960
gccattgagg ttccactttg tgtcatatcc gtcttcaaat acagcgtggc aaactttgat 1020
aagcgcctgt cgtcgacgat ctttctgatt ggttttcaaa ttgccagttc gcttgggatt 1080
gtgctgcttt caacgccgac tgggatactc tttgaccacg caggctacca gacagttttc 1140
ttcgcaattt cgggtattgt ctgcctgatg ttgctatttg gcattttctt cctgagtaaa 1200
aaacgcgagc aaatagttat ggaaacgcct gtaccttcag caatatag 1248
<210> 55
<211> 415
<212> PRT
<213> 大肠杆菌
<400> 55
Met Ala Leu Asn Ile Pro Phe Arg Asn Ala Tyr Tyr Arg Phe Ala Ser
1 5 10 15
Ser Tyr Ser Phe Leu Phe Phe Ile Ser Trp Ser Leu Trp Trp Ser Leu
20 25 30
Tyr Ala Ile Trp Leu Lys Gly His Leu Gly Leu Thr Gly Thr Glu Leu
35 40 45
Gly Thr Leu Tyr Ser Val Asn Gln Phe Thr Ser Ile Leu Phe Met Met
50 55 60
Phe Tyr Gly Ile Val Gln Asp Lys Leu Gly Leu Lys Lys Pro Leu Ile
65 70 75 80
Trp Cys Met Ser Phe Ile Leu Val Leu Thr Gly Pro Phe Met Ile Tyr
85 90 95
Val Tyr Glu Pro Leu Leu Gln Ser Asn Phe Ser Val Gly Leu Ile Leu
100 105 110
Gly Ala Leu Phe Phe Gly Leu Gly Tyr Leu Ala Gly Cys Gly Leu Leu
115 120 125
Asp Ser Phe Thr Glu Lys Met Ala Arg Asn Phe His Phe Glu Tyr Gly
130 135 140
Thr Ala Arg Ala Trp Gly Ser Phe Gly Tyr Ala Ile Gly Ala Phe Phe
145 150 155 160
Ala Gly Ile Phe Phe Ser Ile Ser Pro His Ile Asn Phe Trp Leu Val
165 170 175
Ser Leu Phe Gly Ala Val Phe Met Met Ile Asn Met Arg Phe Lys Asp
180 185 190
Lys Asp His Gln Cys Ile Ala Ala Asp Ala Gly Gly Val Lys Lys Glu
195 200 205
Asp Phe Ile Ala Val Phe Lys Asp Arg Asn Phe Trp Val Phe Val Ile
210 215 220
Phe Ile Val Gly Thr Trp Ser Phe Tyr Asn Ile Phe Asp Gln Gln Leu
225 230 235 240
Phe Pro Val Phe Tyr Ala Gly Leu Phe Glu Ser His Asp Val Gly Thr
245 250 255
Arg Leu Tyr Gly Tyr Leu Asn Ser Phe Gln Val Val Leu Glu Ala Leu
260 265 270
Cys Met Ala Ile Ile Pro Phe Phe Val Asn Arg Val Gly Pro Lys Asn
275 280 285
Ala Leu Leu Ile Gly Val Val Ile Met Ala Leu Arg Ile Leu Ser Cys
290 295 300
Ala Leu Phe Val Asn Pro Trp Ile Ile Ser Leu Val Lys Leu Leu His
305 310 315 320
Ala Ile Glu Val Pro Leu Cys Val Ile Ser Val Phe Lys Tyr Ser Val
325 330 335
Ala Asn Phe Asp Lys Arg Leu Ser Ser Thr Ile Phe Leu Ile Gly Phe
340 345 350
Gln Ile Ala Ser Ser Leu Gly Ile Val Leu Leu Ser Thr Pro Thr Gly
355 360 365
Ile Leu Phe Asp His Ala Gly Tyr Gln Thr Val Phe Phe Ala Ile Ser
370 375 380
Gly Ile Val Cys Leu Met Leu Leu Phe Gly Ile Phe Phe Leu Ser Lys
385 390 395 400
Lys Arg Glu Gln Ile Val Met Glu Thr Pro Val Pro Ser Ala Ile
405 410 415
<210> 56
<211> 924
<212> DNA
<213> 大肠杆菌
<400> 56
atgtcagcca aagtatgggt tttaggggat gcggtcgtag atctcttgcc agaatcagac 60
gggcggctac tgccttgtcc tggcggcgcg ccagctaacg ttgcggtggg aatcgccaga 120
ttaggcggaa caagtgggtt tataggtcgg gtcggtgatg atccttttgg tgcgttaatg 180
caaagaacgc tgctaactga gggtgtcgat atcacgtatc tgaagcaaga tgaatggcac 240
cggacatcca cggtgcttgt cgatctgaac gatcaaggag aacgttcatt tacgtttatg 300
gtccgcccca gtgccgatct ttttttagag acgacagact tgccctgctg gcgacatggc 360
gaatggttac atctctgttc aattgcgttg tctgccgagc cttcgcgtac cagcgcattt 420
actgcgatga cggcgatccg gcatgccgga ggttttgtca gcttcgatcc caatattcgt 480
gaagatctat ggcaagacga gcatttgctc cgcttgtgtt tgcggcaggc gctacaactg 540
gcggatgtcg tcaagctctc ggaagaagaa tggcgactta tcagtggaaa aacacagaac 600
gatcgggata tatgcgccct ggcaaaagag tatgagatcg ccatgctgtt ggtgactaaa 660
ggtgcagaag gggtggtggt ctgttatcga ggacaagtcc accattttgc tggaatgtct 720
gtgaattgtg tcgatagcac tggggcggga gatgcgttcg ttgccgggtt actcacaggt 780
ctgtcctcta cgggattatc tacagatgag agagaaatgc gacgaattat cgatctcgct 840
caacgttgcg gagcgcttgc agtaacagcg aaaggggcaa tgacagcgct gccatgtcga 900
caagaactgg aaagtgagaa gtaa 924
<210> 57
<211> 307
<212> PRT
<213> 大肠杆菌
<400> 57
Met Ser Ala Lys Val Trp Val Leu Gly Asp Ala Val Val Asp Leu Leu
1 5 10 15
Pro Glu Ser Asp Gly Arg Leu Leu Pro Cys Pro Gly Gly Ala Pro Ala
20 25 30
Asn Val Ala Val Gly Ile Ala Arg Leu Gly Gly Thr Ser Gly Phe Ile
35 40 45
Gly Arg Val Gly Asp Asp Pro Phe Gly Ala Leu Met Gln Arg Thr Leu
50 55 60
Leu Thr Glu Gly Val Asp Ile Thr Tyr Leu Lys Gln Asp Glu Trp His
65 70 75 80
Arg Thr Ser Thr Val Leu Val Asp Leu Asn Asp Gln Gly Glu Arg Ser
85 90 95
Phe Thr Phe Met Val Arg Pro Ser Ala Asp Leu Phe Leu Glu Thr Thr
100 105 110
Asp Leu Pro Cys Trp Arg His Gly Glu Trp Leu His Leu Cys Ser Ile
115 120 125
Ala Leu Ser Ala Glu Pro Ser Arg Thr Ser Ala Phe Thr Ala Met Thr
130 135 140
Ala Ile Arg His Ala Gly Gly Phe Val Ser Phe Asp Pro Asn Ile Arg
145 150 155 160
Glu Asp Leu Trp Gln Asp Glu His Leu Leu Arg Leu Cys Leu Arg Gln
165 170 175
Ala Leu Gln Leu Ala Asp Val Val Lys Leu Ser Glu Glu Glu Trp Arg
180 185 190
Leu Ile Ser Gly Lys Thr Gln Asn Asp Arg Asp Ile Cys Ala Leu Ala
195 200 205
Lys Glu Tyr Glu Ile Ala Met Leu Leu Val Thr Lys Gly Ala Glu Gly
210 215 220
Val Val Val Cys Tyr Arg Gly Gln Val His His Phe Ala Gly Met Ser
225 230 235 240
Val Asn Cys Val Asp Ser Thr Gly Ala Gly Asp Ala Phe Val Ala Gly
245 250 255
Leu Leu Thr Gly Leu Ser Ser Thr Gly Leu Ser Thr Asp Glu Arg Glu
260 265 270
Met Arg Arg Ile Ile Asp Leu Ala Gln Arg Cys Gly Ala Leu Ala Val
275 280 285
Thr Ala Lys Gly Ala Met Thr Ala Leu Pro Cys Arg Gln Glu Leu Glu
290 295 300
Ser Glu Lys
305
<210> 58
<211> 1434
<212> DNA
<213> 大肠杆菌
<400> 58
atgacgcaat ctcgattgca tgcggcgcaa aacgccctag caaaacttca tgagcaccgg 60
ggtaacactt tctatcccca ttttcacctc gcgcctcctg ccgggtggat gaacgatcca 120
aacggcctga tctggtttaa cgatcgttat cacgcgtttt atcaacatca tccgatgagc 180
gaacactggg ggccaatgca ctggggacat gccaccagcg acgatatgat ccactggcag 240
catgagccta ttgcgctagc gccaggagac gataatgaca aagacgggtg tttttcaggt 300
agtgctgtcg atgacaatgg tgtcctctca cttatctaca ccggacacgt ctggctcgat 360
ggtgcaggta atgacgatgc aattcgcgaa gtacaatgtc tggctaccag tcgggatggt 420
attcatttcg agaaacaggg tgtgatcctc actccaccag aaggaatcat gcacttccgc 480
gatcctaaag tgtggcgtga agccgacaca tggtggatgg tagtcggggc gaaagatcca 540
ggcaacacgg ggcagatcct gctttatcgc ggcagttcat tgcgtgaatg gaccttcgat 600
cgcgtactgg cccacgctga tgcgggtgaa agctatatgt gggaatgtcc ggactttttc 660
agccttggcg atcagcatta tctgatgttt tccccgcagg gaatgaatgc cgagggatac 720
agttaccgaa atcgctttca aagtggcgta atacccggaa tgtggtcgcc aggacgactt 780
tttgcacaat ccgggcattt tactgaactt gataacgggc atgactttta tgcaccacaa 840
agctttttag cgaaggatgg tcggcgtatt gttatcggat ggatggatat gtgggaatcg 900
ccaatgccct caaaacgtga aggctgggca ggctgcatga cgctggcgcg cgagctatca 960
gagagcaatg gcaaacttct acaacgcccg gttcacgaag ctgagtcgtt acgccagcag 1020
catcaatctg tctctccccg cacaatcagc aataaatatg ttttgcagga aaacgcgcaa 1080
gcagttgaga ttcagttgca gtgggcgctg aagaacagtg atgccgaaca ttacggatta 1140
cagctcggca ctggaatgcg gctgtatatt gataaccaat ctgagcgact tgttttgtgg 1200
cggtattacc cacacgagaa tttagacggc taccgtagta ttcccctccc gcagcgtgac 1260
acgctcgccc taaggatatt tatcgataca tcatccgtgg aagtatttat taacgacggg 1320
gaagcggtga tgagtagtcg aatctatccg cagccagaag aacgggaact gtcgctttat 1380
gcctcccacg gagtggctgt gctgcaacat ggagcactct ggctactggg ttaa 1434
<210> 59
<211> 480
<212> PRT
<213> 大肠杆菌
<400> 59
Met Ile Lys Met Thr Gln Ser Arg Leu His Ala Ala Gln Asn Ala Leu
1 5 10 15
Ala Lys Leu His Glu His Arg Gly Asn Thr Phe Tyr Pro His Phe His
20 25 30
Leu Ala Pro Pro Ala Gly Trp Met Asn Asp Pro Asn Gly Leu Ile Trp
35 40 45
Phe Asn Asp Arg Tyr His Ala Phe Tyr Gln His His Pro Met Ser Glu
50 55 60
His Trp Gly Pro Met His Trp Gly His Ala Thr Ser Asp Asp Met Ile
65 70 75 80
His Trp Gln His Glu Pro Ile Ala Leu Ala Pro Gly Asp Asp Asn Asp
85 90 95
Lys Asp Gly Cys Phe Ser Gly Ser Ala Val Asp Asp Asn Gly Val Leu
100 105 110
Ser Leu Ile Tyr Thr Gly His Val Trp Leu Asp Gly Ala Gly Asn Asp
115 120 125
Asp Ala Ile Arg Glu Val Gln Cys Leu Ala Thr Ser Arg Asp Gly Ile
130 135 140
His Phe Glu Lys Gln Gly Val Ile Leu Thr Pro Pro Glu Gly Ile Met
145 150 155 160
His Phe Arg Asp Pro Lys Val Trp Arg Glu Ala Asp Thr Trp Trp Met
165 170 175
Val Val Gly Ala Lys Asp Pro Gly Asn Thr Gly Gln Ile Leu Leu Tyr
180 185 190
Arg Gly Ser Ser Leu Arg Glu Trp Thr Phe Asp Arg Val Leu Ala His
195 200 205
Ala Asp Ala Gly Glu Ser Tyr Met Trp Glu Cys Pro Asp Phe Phe Ser
210 215 220
Leu Gly Asp Gln His Tyr Leu Met Phe Ser Pro Gln Gly Met Asn Ala
225 230 235 240
Glu Gly Tyr Ser Tyr Arg Asn Arg Phe Gln Ser Gly Val Ile Pro Gly
245 250 255
Met Trp Ser Pro Gly Arg Leu Phe Ala Gln Ser Gly His Phe Thr Glu
260 265 270
Leu Asp Asn Gly His Asp Phe Tyr Ala Pro Gln Ser Phe Leu Ala Lys
275 280 285
Asp Gly Arg Arg Ile Val Ile Gly Trp Met Asp Met Trp Glu Ser Pro
290 295 300
Met Pro Ser Lys Arg Glu Gly Trp Ala Gly Cys Met Thr Leu Ala Arg
305 310 315 320
Glu Leu Ser Glu Ser Asn Gly Lys Leu Leu Gln Arg Pro Val His Glu
325 330 335
Ala Glu Ser Leu Arg Gln Gln His Gln Ser Val Ser Pro Arg Thr Ile
340 345 350
Ser Asn Lys Tyr Val Leu Gln Glu Asn Ala Gln Ala Val Glu Ile Gln
355 360 365
Leu Gln Trp Ala Leu Lys Asn Ser Asp Ala Glu His Tyr Gly Leu Gln
370 375 380
Leu Gly Thr Gly Met Arg Leu Tyr Ile Asp Asn Gln Ser Glu Arg Leu
385 390 395 400
Val Leu Trp Arg Tyr Tyr Pro His Glu Asn Leu Asp Gly Tyr Arg Ser
405 410 415
Ile Pro Leu Pro Gln Arg Asp Thr Leu Ala Leu Arg Ile Phe Ile Asp
420 425 430
Thr Ser Ser Val Glu Val Phe Ile Asn Asp Gly Glu Ala Val Met Ser
435 440 445
Ser Arg Ile Tyr Pro Gln Pro Glu Glu Arg Glu Leu Ser Leu Tyr Ala
450 455 460
Ser His Gly Val Ala Val Leu Gln His Gly Ala Leu Trp Leu Leu Gly
465 470 475 480
<210> 60
<211> 954
<212> DNA
<213> 大肠杆菌
<400> 60
atgatgacag tctcccgggt gatgcataat gcagaatctg tgcgtcctgc aacgcgtgac 60
cgcgtattgc aggcaatcca gaccctgaat tatgttcctg atctttccgc ccgtaagatg 120
cgcgctcaag gacgtaagcc gtcgactctc gccgtgctgg cgcaggacac ggctaccact 180
cctttctctg ttgatattct gcttgccatt gagcaaaccg ccagcgagtt cggctggaat 240
agttttttaa tcaatatttt ttctgaagat gacgctgccc gtgctgcacg tcagctgctt 300
gcccaccgtc cggatggcat tatctatact acaatggggc tgcgacatat cacgctgcct 360
gagtctctgt atggtgaaaa tattgtattg gcgaactgtg tggcggatga cccagcgtta 420
cccagttata tccctgatga ttacactgca caatatgaat caacacagca tttgctcgcg 480
gcgggctatc gtcaaccgtt atgcttctgg ctaccggaaa gtgcgttggc aacagggtat 540
cgtcggcagg gatttgagca ggcctggcgt gatgctggac gagatctggc tgaggtgaaa 600
caatttcaca tggcaacagg tgatgatcac tacaccgatc tcgcaagttt actcaatgcc 660
cacttcaaat cgggcaaacc agattttgat gttctgatat gtggtaacga tcgcgcagct 720
tttgtggctt atcaggttct tttggcgaag ggggtacgta tcccgcagga tgtcgccgta 780
atgggctttg ataatctggt tggcgtcggg catctgtttt taccgccgct gaccacaatt 840
cagcttccac atgacattat cgggcgggaa gctgcattgc atattattga aggtcgtgaa 900
gggggaagag tgacccggat cccttgcccg ctgttgatcc gttgttccac ctga 954
<210> 61
<211> 317
<212> PRT
<213> 大肠杆菌
<400> 61
Met Met Thr Val Ser Arg Val Met His Asn Ala Glu Ser Val Arg Pro
1 5 10 15
Ala Thr Arg Asp Arg Val Leu Gln Ala Ile Gln Thr Leu Asn Tyr Val
20 25 30
Pro Asp Leu Ser Ala Arg Lys Met Arg Ala Gln Gly Arg Lys Pro Ser
35 40 45
Thr Leu Ala Val Leu Ala Gln Asp Thr Ala Thr Thr Pro Phe Ser Val
50 55 60
Asp Ile Leu Leu Ala Ile Glu Gln Thr Ala Ser Glu Phe Gly Trp Asn
65 70 75 80
Ser Phe Leu Ile Asn Ile Phe Ser Glu Asp Asp Ala Ala Arg Ala Ala
85 90 95
Arg Gln Leu Leu Ala His Arg Pro Asp Gly Ile Ile Tyr Thr Thr Met
100 105 110
Gly Leu Arg His Ile Thr Leu Pro Glu Ser Leu Tyr Gly Glu Asn Ile
115 120 125
Val Leu Ala Asn Cys Val Ala Asp Asp Pro Ala Leu Pro Ser Tyr Ile
130 135 140
Pro Asp Asp Tyr Thr Ala Gln Tyr Glu Ser Thr Gln His Leu Leu Ala
145 150 155 160
Ala Gly Tyr Arg Gln Pro Leu Cys Phe Trp Leu Pro Glu Ser Ala Leu
165 170 175
Ala Thr Gly Tyr Arg Arg Gln Gly Phe Glu Gln Ala Trp Arg Asp Ala
180 185 190
Gly Arg Asp Leu Ala Glu Val Lys Gln Phe His Met Ala Thr Gly Asp
195 200 205
Asp His Tyr Thr Asp Leu Ala Ser Leu Leu Asn Ala His Phe Lys Ser
210 215 220
Gly Lys Pro Asp Phe Asp Val Leu Ile Cys Gly Asn Asp Arg Ala Ala
225 230 235 240
Phe Val Ala Tyr Gln Val Leu Leu Ala Lys Gly Val Arg Ile Pro Gln
245 250 255
Asp Val Ala Val Met Gly Phe Asp Asn Leu Val Gly Val Gly His Leu
260 265 270
Phe Leu Pro Pro Leu Thr Thr Ile Gln Leu Pro His Asp Ile Ile Gly
275 280 285
Arg Glu Ala Ala Leu His Ile Ile Glu Gly Arg Glu Gly Gly Arg Val
290 295 300
Thr Arg Ile Pro Cys Pro Leu Leu Ile Arg Cys Ser Thr
305 310 315
<210> 62
<211> 1254
<212> DNA
<213> 大肠杆菌
<400> 62
atgtactatt taaaaaacac aaacttttgg atgttcggtt tattcttttt cttttacttt 60
tttatcatgg gagcctactt cccgtttttc ccgatttggc tacatgacat caaccatatc 120
agcaaaagtg atacgggtat tatttttgcc gctatttctc tgttctcgct attattccaa 180
ccgctgtttg gtctgctttc tgacaaactc gggctgcgca aatacctgct gtggattatt 240
accggcatgt tagtgatgtt tgcgccgttc tttattttta tcttcgggcc actgttacaa 300
tacaacattt tagtaggatc gattgttggt ggtatttatc taggcttttg ttttaacgcc 360
ggtgcgccag cagtagaggc atttattgag aaagtcagcc gtcgcagtaa tttcgaattt 420
ggtcgcgcgc ggatgtttgg ctgtgttggc tgggcgctgt gtgcctcgat tgtcggcatc 480
atgttcacca tcaataatca gtttgttttc tggctgggct ctggctgtgc actcatcctc 540
gccgttttac tctttttcgc caaaacggat gcgccctctt ctgccacggt tgccaatgcg 600
gtaggtgcca accattcggc atttagcctt aagctggcac tggaactgtt cagacagcca 660
aaactgtggt ttttgtcact gtatgttatt ggcgtttcct gcacctacga tgtttttgac 720
caacagtttg ctaatttctt tacttcgttc tttgctaccg gtgaacaggg tacgcgggta 780
tttggctacg taacgacaat gggcgaatta cttaacgcct cgattatgtt ctttgcgcca 840
ctgatcatta atcgcatcgg tgggaaaaac gccctgctgc tggctggcac tattatgtct 900
gtacgtatta ttggctcatc gttcgccacc tcagcgctgg aagtggttat tctgaaaacg 960
ctgcatatgt ttgaagtacc gttcctgctg gtgggctgct ttaaatatat taccagccag 1020
tttgaagtgc gtttttcagc gacgatttat ctggtctgtt tctgcttctt taagcaactg 1080
gcgatgattt ttatgtctgt actggcgggc aatatgtatg aaagcatcgg tttccagggc 1140
gcttatctgg tgctgggtct ggtggcgctg ggcttcacct taatttccgt gttcacgctt 1200
agcggccccg gcccgctttc cctgctgcgt cgtcaggtga atgaagtcgc ttaa 1254
<210> 63
<211> 417
<212> PRT
<213> 大肠杆菌
<400> 63
Met Tyr Tyr Leu Lys Asn Thr Asn Phe Trp Met Phe Gly Leu Phe Phe
1 5 10 15
Phe Phe Tyr Phe Phe Ile Met Gly Ala Tyr Phe Pro Phe Phe Pro Ile
20 25 30
Trp Leu His Asp Ile Asn His Ile Ser Lys Ser Asp Thr Gly Ile Ile
35 40 45
Phe Ala Ala Ile Ser Leu Phe Ser Leu Leu Phe Gln Pro Leu Phe Gly
50 55 60
Leu Leu Ser Asp Lys Leu Gly Leu Arg Lys Tyr Leu Leu Trp Ile Ile
65 70 75 80
Thr Gly Met Leu Val Met Phe Ala Pro Phe Phe Ile Phe Ile Phe Gly
85 90 95
Pro Leu Leu Gln Tyr Asn Ile Leu Val Gly Ser Ile Val Gly Gly Ile
100 105 110
Tyr Leu Gly Phe Cys Phe Asn Ala Gly Ala Pro Ala Val Glu Ala Phe
115 120 125
Ile Glu Lys Val Ser Arg Arg Ser Asn Phe Glu Phe Gly Arg Ala Arg
130 135 140
Met Phe Gly Cys Val Gly Trp Ala Leu Cys Ala Ser Ile Val Gly Ile
145 150 155 160
Met Phe Thr Ile Asn Asn Gln Phe Val Phe Trp Leu Gly Ser Gly Cys
165 170 175
Ala Leu Ile Leu Ala Val Leu Leu Phe Phe Ala Lys Thr Asp Ala Pro
180 185 190
Ser Ser Ala Thr Val Ala Asn Ala Val Gly Ala Asn His Ser Ala Phe
195 200 205
Ser Leu Lys Leu Ala Leu Glu Leu Phe Arg Gln Pro Lys Leu Trp Phe
210 215 220
Leu Ser Leu Tyr Val Ile Gly Val Ser Cys Thr Tyr Asp Val Phe Asp
225 230 235 240
Gln Gln Phe Ala Asn Phe Phe Thr Ser Phe Phe Ala Thr Gly Glu Gln
245 250 255
Gly Thr Arg Val Phe Gly Tyr Val Thr Thr Met Gly Glu Leu Leu Asn
260 265 270
Ala Ser Ile Met Phe Phe Ala Pro Leu Ile Ile Asn Arg Ile Gly Gly
275 280 285
Lys Asn Ala Leu Leu Leu Ala Gly Thr Ile Met Ser Val Arg Ile Ile
290 295 300
Gly Ser Ser Phe Ala Thr Ser Ala Leu Glu Val Val Ile Leu Lys Thr
305 310 315 320
Leu His Met Phe Glu Val Pro Phe Leu Leu Val Gly Cys Phe Lys Tyr
325 330 335
Ile Thr Ser Gln Phe Glu Val Arg Phe Ser Ala Thr Ile Tyr Leu Val
340 345 350
Cys Phe Cys Phe Phe Lys Gln Leu Ala Met Ile Phe Met Ser Val Leu
355 360 365
Ala Gly Asn Met Tyr Glu Ser Ile Gly Phe Gln Gly Ala Tyr Leu Val
370 375 380
Leu Gly Leu Val Ala Leu Gly Phe Thr Leu Ile Ser Val Phe Thr Leu
385 390 395 400
Ser Gly Pro Gly Pro Leu Ser Leu Leu Arg Arg Gln Val Asn Glu Val
405 410 415
Ala
<210> 64
<211> 3075
<212> DNA
<213> 大肠杆菌
<400> 64
atgaccatga ttacggattc actggccgtc gttttacaac gtcgtgactg ggaaaaccct 60
ggcgttaccc aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc 120
gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatggcgc 180
tttgcctggt ttccggcacc agaagcggtg ccggaaagct ggctggagtg cgatcttcct 240
gaggccgata ctgtcgtcgt cccctcaaac tggcagatgc acggttacga tgcgcccatc 300
tacaccaacg tgacctatcc cattacggtc aatccgccgt ttgttcccac ggagaatccg 360
acgggttgtt actcgctcac atttaatgtt gatgaaagct ggctacagga aggccagacg 420
cgaattattt ttgatggcgt taactcggcg tttcatctgt ggtgcaacgg gcgctgggtc 480
ggttacggcc aggacagtcg tttgccgtct gaatttgacc tgagcgcatt tttacgcgcc 540
ggagaaaacc gcctcgcggt gatggtgctg cgctggagtg acggcagtta tctggaagat 600
caggatatgt ggcggatgag cggcattttc cgtgacgtct cgttgctgca taaaccgact 660
acacaaatca gcgatttcca tgttgccact cgctttaatg atgatttcag ccgcgctgta 720
ctggaggctg aagttcagat gtgcggcgag ttgcgtgact acctacgggt aacagtttct 780
ttatggcagg gtgaaacgca ggtcgccagc ggcaccgcgc ctttcggcgg tgaaattatc 840
gatgagcgtg gtggttatgc cgatcgcgtc acactacgtc tgaacgtcga aaacccgaaa 900
ctgtggagcg ccgaaatccc gaatctctat cgtgcggtgg ttgaactgca caccgccgac 960
ggcacgctga ttgaagcaga agcctgcgat gtcggtttcc gcgaggtgcg gattgaaaat 1020
ggtctgctgc tgctgaacgg caagccgttg ctgattcgag gcgttaaccg tcacgagcat 1080
catcctctgc atggtcaggt catggatgag cagacgatgg tgcaggatat cctgctgatg 1140
aagcagaaca actttaacgc cgtgcgctgt tcgcattatc cgaaccatcc gctgtggtac 1200
acgctgtgcg accgctacgg cctgtatgtg gtggatgaag ccaatattga aacccacggc 1260
atggtgccaa tgaatcgtct gaccgatgat ccgcgctggc taccggcgat gagcgaacgc 1320
gtaacgcgaa tggtgcagcg cgatcgtaat cacccgagtg tgatcatctg gtcgctgggg 1380
aatgaatcag gccacggcgc taatcacgac gcgctgtatc gctggatcaa atctgtcgat 1440
ccttcccgcc cggtgcagta tgaaggcggc ggagccgaca ccacggccac cgatattatt 1500
tgcccgatgt acgcgcgcgt ggatgaagac cagcccttcc cggctgtgcc gaaatggtcc 1560
atcaaaaaat ggctttcgct acctggagag acgcgcccgc tgatcctttg cgaatacgcc 1620
cacgcgatgg gtaacagtct tggcggtttc gctaaatact ggcaggcgtt tcgtcagtat 1680
ccccgtttac agggcggctt cgtctgggac tgggtggatc agtcgctgat taaatatgat 1740
gaaaacggca acccgtggtc ggcttacggc ggtgattttg gcgatacgcc gaacgatcgc 1800
cagttctgta tgaacggtct ggtctttgcc gaccgcacgc cgcatccagc gctgacggaa 1860
gcaaaacacc agcagcagtt tttccagttc cgtttatccg ggcaaaccat cgaagtgacc 1920
agcgaatacc tgttccgtca tagcgataac gagctcctgc actggatggt ggcgctggat 1980
ggtaagccgc tggcaagcgg tgaagtgcct ctggatgtcg ctccacaagg taaacagttg 2040
attgaactgc ctgaactacc gcagccggag agcgccgggc aactctggct cacagtacgc 2100
gtagtgcaac cgaacgcgac cgcatggtca gaagccgggc acatcagcgc ctggcagcag 2160
tggcgtctgg cggaaaacct cagtgtgacg ctccccgccg cgtcccacgc catcccgcat 2220
ctgaccacca gcgaaatgga tttttgcatc gagctgggta ataagcgttg gcaatttaac 2280
cgccagtcag gctttctttc acagatgtgg attggcgata aaaaacaact gctgacgccg 2340
ctgcgcgatc agttcacccg tgcaccgctg gataacgaca ttggcgtaag tgaagcgacc 2400
cgcattgacc ctaacgcctg ggtcgaacgc tggaaggcgg cgggccatta ccaggccgaa 2460
gcagcgttgt tgcagtgcac ggcagataca cttgctgatg cggtgctgat tacgaccgct 2520
cacgcgtggc agcatcaggg gaaaacctta tttatcagcc ggaaaaccta ccggattgat 2580
ggtagtggtc aaatggcgat taccgttgat gttgaagtgg cgagcgatac accgcatccg 2640
gcgcggattg gcctgaactg ccagctggcg caggtagcag agcgggtaaa ctggctcgga 2700
ttagggccgc aagaaaacta tcccgaccgc cttactgccg cctgttttga ccgctgggat 2760
ctgccattgt cagacatgta taccccgtac gtcttcccga gcgaaaacgg tctgcgctgc 2820
gggacgcgcg aattgaatta tggcccacac cagtggcgcg gcgacttcca gttcaacatc 2880
agccgctaca gtcaacagca actgatggaa accagccatc gccatctgct gcacgcggaa 2940
gaaggcacat ggctgaatat cgacggtttc catatgggga ttggtggcga cgactcctgg 3000
agcccgtcag tatcggcgga attccagctg agcgccggtc gctaccatta ccagttggtc 3060
tggtgtcaaa aataa 3075
<210> 65
<211> 1024
<212> PRT
<213> 大肠杆菌
<400> 65
Met Thr Met Ile Thr Asp Ser Leu Ala Val Val Leu Gln Arg Arg Asp
1 5 10 15
Trp Glu Asn Pro Gly Val Thr Gln Leu Asn Arg Leu Ala Ala His Pro
20 25 30
Pro Phe Ala Ser Trp Arg Asn Ser Glu Glu Ala Arg Thr Asp Arg Pro
35 40 45
Ser Gln Gln Leu Arg Ser Leu Asn Gly Glu Trp Arg Phe Ala Trp Phe
50 55 60
Pro Ala Pro Glu Ala Val Pro Glu Ser Trp Leu Glu Cys Asp Leu Pro
65 70 75 80
Glu Ala Asp Thr Val Val Val Pro Ser Asn Trp Gln Met His Gly Tyr
85 90 95
Asp Ala Pro Ile Tyr Thr Asn Val Thr Tyr Pro Ile Thr Val Asn Pro
100 105 110
Pro Phe Val Pro Thr Glu Asn Pro Thr Gly Cys Tyr Ser Leu Thr Phe
115 120 125
Asn Val Asp Glu Ser Trp Leu Gln Glu Gly Gln Thr Arg Ile Ile Phe
130 135 140
Asp Gly Val Asn Ser Ala Phe His Leu Trp Cys Asn Gly Arg Trp Val
145 150 155 160
Gly Tyr Gly Gln Asp Ser Arg Leu Pro Ser Glu Phe Asp Leu Ser Ala
165 170 175
Phe Leu Arg Ala Gly Glu Asn Arg Leu Ala Val Met Val Leu Arg Trp
180 185 190
Ser Asp Gly Ser Tyr Leu Glu Asp Gln Asp Met Trp Arg Met Ser Gly
195 200 205
Ile Phe Arg Asp Val Ser Leu Leu His Lys Pro Thr Thr Gln Ile Ser
210 215 220
Asp Phe His Val Ala Thr Arg Phe Asn Asp Asp Phe Ser Arg Ala Val
225 230 235 240
Leu Glu Ala Glu Val Gln Met Cys Gly Glu Leu Arg Asp Tyr Leu Arg
245 250 255
Val Thr Val Ser Leu Trp Gln Gly Glu Thr Gln Val Ala Ser Gly Thr
260 265 270
Ala Pro Phe Gly Gly Glu Ile Ile Asp Glu Arg Gly Gly Tyr Ala Asp
275 280 285
Arg Val Thr Leu Arg Leu Asn Val Glu Asn Pro Lys Leu Trp Ser Ala
290 295 300
Glu Ile Pro Asn Leu Tyr Arg Ala Val Val Glu Leu His Thr Ala Asp
305 310 315 320
Gly Thr Leu Ile Glu Ala Glu Ala Cys Asp Val Gly Phe Arg Glu Val
325 330 335
Arg Ile Glu Asn Gly Leu Leu Leu Leu Asn Gly Lys Pro Leu Leu Ile
340 345 350
Arg Gly Val Asn Arg His Glu His His Pro Leu His Gly Gln Val Met
355 360 365
Asp Glu Gln Thr Met Val Gln Asp Ile Leu Leu Met Lys Gln Asn Asn
370 375 380
Phe Asn Ala Val Arg Cys Ser His Tyr Pro Asn His Pro Leu Trp Tyr
385 390 395 400
Thr Leu Cys Asp Arg Tyr Gly Leu Tyr Val Val Asp Glu Ala Asn Ile
405 410 415
Glu Thr His Gly Met Val Pro Met Asn Arg Leu Thr Asp Asp Pro Arg
420 425 430
Trp Leu Pro Ala Met Ser Glu Arg Val Thr Arg Met Val Gln Arg Asp
435 440 445
Arg Asn His Pro Ser Val Ile Ile Trp Ser Leu Gly Asn Glu Ser Gly
450 455 460
His Gly Ala Asn His Asp Ala Leu Tyr Arg Trp Ile Lys Ser Val Asp
465 470 475 480
Pro Ser Arg Pro Val Gln Tyr Glu Gly Gly Gly Ala Asp Thr Thr Ala
485 490 495
Thr Asp Ile Ile Cys Pro Met Tyr Ala Arg Val Asp Glu Asp Gln Pro
500 505 510
Phe Pro Ala Val Pro Lys Trp Ser Ile Lys Lys Trp Leu Ser Leu Pro
515 520 525
Gly Glu Thr Arg Pro Leu Ile Leu Cys Glu Tyr Ala His Ala Met Gly
530 535 540
Asn Ser Leu Gly Gly Phe Ala Lys Tyr Trp Gln Ala Phe Arg Gln Tyr
545 550 555 560
Pro Arg Leu Gln Gly Gly Phe Val Trp Asp Trp Val Asp Gln Ser Leu
565 570 575
Ile Lys Tyr Asp Glu Asn Gly Asn Pro Trp Ser Ala Tyr Gly Gly Asp
580 585 590
Phe Gly Asp Thr Pro Asn Asp Arg Gln Phe Cys Met Asn Gly Leu Val
595 600 605
Phe Ala Asp Arg Thr Pro His Pro Ala Leu Thr Glu Ala Lys His Gln
610 615 620
Gln Gln Phe Phe Gln Phe Arg Leu Ser Gly Gln Thr Ile Glu Val Thr
625 630 635 640
Ser Glu Tyr Leu Phe Arg His Ser Asp Asn Glu Leu Leu His Trp Met
645 650 655
Val Ala Leu Asp Gly Lys Pro Leu Ala Ser Gly Glu Val Pro Leu Asp
660 665 670
Val Ala Pro Gln Gly Lys Gln Leu Ile Glu Leu Pro Glu Leu Pro Gln
675 680 685
Pro Glu Ser Ala Gly Gln Leu Trp Leu Thr Val Arg Val Val Gln Pro
690 695 700
Asn Ala Thr Ala Trp Ser Glu Ala Gly His Ile Ser Ala Trp Gln Gln
705 710 715 720
Trp Arg Leu Ala Glu Asn Leu Ser Val Thr Leu Pro Ala Ala Ser His
725 730 735
Ala Ile Pro His Leu Thr Thr Ser Glu Met Asp Phe Cys Ile Glu Leu
740 745 750
Gly Asn Lys Arg Trp Gln Phe Asn Arg Gln Ser Gly Phe Leu Ser Gln
755 760 765
Met Trp Ile Gly Asp Lys Lys Gln Leu Leu Thr Pro Leu Arg Asp Gln
770 775 780
Phe Thr Arg Ala Pro Leu Asp Asn Asp Ile Gly Val Ser Glu Ala Thr
785 790 795 800
Arg Ile Asp Pro Asn Ala Trp Val Glu Arg Trp Lys Ala Ala Gly His
805 810 815
Tyr Gln Ala Glu Ala Ala Leu Leu Gln Cys Thr Ala Asp Thr Leu Ala
820 825 830
Asp Ala Val Leu Ile Thr Thr Ala His Ala Trp Gln His Gln Gly Lys
835 840 845
Thr Leu Phe Ile Ser Arg Lys Thr Tyr Arg Ile Asp Gly Ser Gly Gln
850 855 860
Met Ala Ile Thr Val Asp Val Glu Val Ala Ser Asp Thr Pro His Pro
865 870 875 880
Ala Arg Ile Gly Leu Asn Cys Gln Leu Ala Gln Val Ala Glu Arg Val
885 890 895
Asn Trp Leu Gly Leu Gly Pro Gln Glu Asn Tyr Pro Asp Arg Leu Thr
900 905 910
Ala Ala Cys Phe Asp Arg Trp Asp Leu Pro Leu Ser Asp Met Tyr Thr
915 920 925
Pro Tyr Val Phe Pro Ser Glu Asn Gly Leu Arg Cys Gly Thr Arg Glu
930 935 940
Leu Asn Tyr Gly Pro His Gln Trp Arg Gly Asp Phe Gln Phe Asn Ile
945 950 955 960
Ser Arg Tyr Ser Gln Gln Gln Leu Met Glu Thr Ser His Arg His Leu
965 970 975
Leu His Ala Glu Glu Gly Thr Trp Leu Asn Ile Asp Gly Phe His Met
980 985 990
Gly Ile Gly Gly Asp Asp Ser Trp Ser Pro Ser Val Ser Ala Glu Phe
995 1000 1005
Gln Leu Ser Ala Gly Arg Tyr His Tyr Gln Leu Val Trp Cys Gln
1010 1015 1020
Lys
<210> 66
<211> 1119
<212> DNA
<213> 表皮葡萄球菌(Staphylococcus epidermidis)
<400> 66
atgactaaac tccatatctt ttacttttct ttaatgtact ttcttattgg catgatacac 60
acttttgtcg gttcatttaa tcaattctta aaaatagaac ttaatatgaa tcaatcagat 120
gtatcaaatc taattagtat tcagttcata acatttatga ttggagtatt ttattctacc 180
ttcttagtta ataaagacat aaagaatttt ttaaaaataa tacatttatt tattctatta 240
attactacta cttttattat atttgaacat tacttaataa tatatctgat tgtagctata 300
ttaggttttt gcgctggatt tattgaatca tctatcgcat catatatttt taatagtaag 360
tttgagtccg ctaaaacttt tggatatata gaatcatttt ttgcagtagg gtcatttttg 420
ctccctgtaa ttgtgaaagt gtttgaatac cattcagata caaaacacgc tattatcttt 480
atacttataa taaatataat cttattttta attatctatt cattagagtt tgaagtaagt 540
agcagcgata gaaataaaat acccatactg tcttttaata aaaagtcaat gctagttatg 600
attattttta catggtgttt cttttatatt agtatagaaa caaatttttc aaatttacta 660
ccatatatca acttagtttc tgaaaaatat agctatatta ctgtaagtat attttgggtt 720
ggaataatta taggaaggtt tttatatacg ctaatattga cgttaattag attcaggcta 780
gaatcattac tattaacata tacagtgaca tctttttttc tatatattat tttaatatat 840
ttgaatactc aagatgaagt caaattaata attttgtttt tacttactct attcttagca 900
cctatgttcc ctttaggtgt tagtatcatt aatcaacata gtagtaataa gaacttacta 960
actagtattt ttattgctgt agctggatgt ggtggtgcag ttggtgcagt aattataaaa 1020
tcagctttat acattcatat tcccgtacat ttatctattt tattaatatt aatgacttgc 1080
ttatttttaa gtactgtgat tttaaaaata aagctttaa 1119
<210> 67
<211> 372
<212> PRT
<213> 表皮葡萄球菌
<400> 67
Met Thr Lys Leu His Ile Phe Tyr Phe Ser Leu Met Tyr Phe Leu Ile
1 5 10 15
Gly Met Ile His Thr Phe Val Gly Ser Phe Asn Gln Phe Leu Lys Ile
20 25 30
Glu Leu Asn Met Asn Gln Ser Asp Val Ser Asn Leu Ile Ser Ile Gln
35 40 45
Phe Ile Thr Phe Met Ile Gly Val Phe Tyr Ser Thr Phe Leu Val Asn
50 55 60
Lys Asp Ile Lys Asn Phe Leu Lys Ile Ile His Leu Phe Ile Leu Leu
65 70 75 80
Ile Thr Thr Thr Phe Ile Ile Phe Glu His Tyr Leu Ile Ile Tyr Leu
85 90 95
Ile Val Ala Ile Leu Gly Phe Cys Ala Gly Phe Ile Glu Ser Ser Ile
100 105 110
Ala Ser Tyr Ile Phe Asn Ser Lys Phe Glu Ser Ala Lys Thr Phe Gly
115 120 125
Tyr Ile Glu Ser Phe Phe Ala Val Gly Ser Phe Leu Leu Pro Val Ile
130 135 140
Val Lys Val Phe Glu Tyr His Ser Asp Thr Lys His Ala Ile Ile Phe
145 150 155 160
Ile Leu Ile Ile Asn Ile Ile Leu Phe Leu Ile Ile Tyr Ser Leu Glu
165 170 175
Phe Glu Val Ser Ser Ser Asp Arg Asn Lys Ile Pro Ile Leu Ser Phe
180 185 190
Asn Lys Lys Ser Met Leu Val Met Ile Ile Phe Thr Trp Cys Phe Phe
195 200 205
Tyr Ile Ser Ile Glu Thr Asn Phe Ser Asn Leu Leu Pro Tyr Ile Asn
210 215 220
Leu Val Ser Glu Lys Tyr Ser Tyr Ile Thr Val Ser Ile Phe Trp Val
225 230 235 240
Gly Ile Ile Ile Gly Arg Phe Leu Tyr Thr Leu Ile Leu Thr Leu Ile
245 250 255
Arg Phe Arg Leu Glu Ser Leu Leu Leu Thr Tyr Thr Val Thr Ser Phe
260 265 270
Phe Leu Tyr Ile Ile Leu Ile Tyr Leu Asn Thr Gln Asp Glu Val Lys
275 280 285
Leu Ile Ile Leu Phe Leu Leu Thr Leu Phe Leu Ala Pro Met Phe Pro
290 295 300
Leu Gly Val Ser Ile Ile Asn Gln His Ser Ser Asn Lys Asn Leu Leu
305 310 315 320
Thr Ser Ile Phe Ile Ala Val Ala Gly Cys Gly Gly Ala Val Gly Ala
325 330 335
Val Ile Ile Lys Ser Ala Leu Tyr Ile His Ile Pro Val His Leu Ser
340 345 350
Ile Leu Leu Ile Leu Met Thr Cys Leu Phe Leu Ser Thr Val Ile Leu
355 360 365
Lys Ile Lys Leu
370
<210> 68
<211> 1350
<212> DNA
<213> 短乳杆菌(Lactobacillus brevis)
<400> 68
atgcaagcaa ctgaaaccaa gcacggctgg acccaattag cggacggcta cctcagcaaa 60
acgccattat ttcaatttat tttagtttca ttgatttttc cactgtgggg aactgcggca 120
agtttaaatg atattttgat tacgcagttc aagacagtct ttcaacttaa cgatgccgcg 180
acggcctttg ttcaaagtgc cttctatggt gggtatttct taattgccat tccggcatcc 240
ctgattatta agaagaacag ttataaattt gccatcatga ccgggttgat cttttatatc 300
atcgggtgtg ggctgttttt cccggcctca catctcgcaa cttacagtat gttcctggtg 360
gccatctttg ccattgccat tggtctgagc ttcttggaaa catcatgtga tacgtatagt 420
tcaatgctgg gaccgaagca acacgccacg atgcgcttga acttttccca gacattaatt 480
ccgttaggcg acatcatggg aattgtttta gggaagtact taatttttgg ttctgtaggt 540
aatttatctg aaaagatgag ccatatgcac ggcgcagcac gcattgctta cggcgaacag 600
atgttacaat tgacgttacg gccttacaaa tatatcttaa tcgtgttact cgtgatgctg 660
attatctttg ccgtaacgcc tatgccacgg gctaaggcga cgaaggaaat tggtggggaa 720
caacaagaag aacgtcctag tcttggggaa actctgaagt atctatcaca caacaagcac 780
tatattaaag gggtagtaac ccagttcttt tatgcgggtc tgcaaacaac cgtctggtcc 840
tttacgattc gtttggtatt aaacttgaac catcaaatta ccgacagcgg tgcatcaacc 900
tttatgattt atagttatgt ggcgtggttc gttggtaagc tggttgccaa tacctttatg 960
agtcgcttct caattacgaa ggtgctgacg tggtactcct tattggggac attagcatta 1020
gttgtgacct ttacggttcc gaatatgatt gcggtctacg cagccatctt aacgagtttc 1080
ttctttggtc cagaatggcc aacaatttat gcgcacacgt tggatgccgt tacggagaag 1140
aaatacactg aaacggctgg ggcaattatc gtgatggccc tgatcggtgg tgcagtcatt 1200
ccagccattc aaggcctggt ttctgatgcg accggttcaa tgcagttctc attcgttgta 1260
ccaatgctct gctacgcttt aattacaggg tactttttct tcgaacatcg ttttgagaaa 1320
gctcacccta acgaagttca agaacattaa 1350
<210> 69
<211> 449
<212> PRT
<213> 短乳杆菌
<400> 69
Met Gln Ala Thr Glu Thr Lys His Gly Trp Thr Gln Leu Ala Asp Gly
1 5 10 15
Tyr Leu Ser Lys Thr Pro Leu Phe Gln Phe Ile Leu Val Ser Leu Ile
20 25 30
Phe Pro Leu Trp Gly Thr Ala Ala Ser Leu Asn Asp Ile Leu Ile Thr
35 40 45
Gln Phe Lys Thr Val Phe Gln Leu Asn Asp Ala Ala Thr Ala Phe Val
50 55 60
Gln Ser Ala Phe Tyr Gly Gly Tyr Phe Leu Ile Ala Ile Pro Ala Ser
65 70 75 80
Leu Ile Ile Lys Lys Asn Ser Tyr Lys Phe Ala Ile Met Thr Gly Leu
85 90 95
Ile Phe Tyr Ile Ile Gly Cys Gly Leu Phe Phe Pro Ala Ser His Leu
100 105 110
Ala Thr Tyr Ser Met Phe Leu Val Ala Ile Phe Ala Ile Ala Ile Gly
115 120 125
Leu Ser Phe Leu Glu Thr Ser Cys Asp Thr Tyr Ser Ser Met Leu Gly
130 135 140
Pro Lys Gln His Ala Thr Met Arg Leu Asn Phe Ser Gln Thr Leu Ile
145 150 155 160
Pro Leu Gly Asp Ile Met Gly Ile Val Leu Gly Lys Tyr Leu Ile Phe
165 170 175
Gly Ser Val Gly Asn Leu Ser Glu Lys Met Ser His Met His Gly Ala
180 185 190
Ala Arg Ile Ala Tyr Gly Glu Gln Met Leu Gln Leu Thr Leu Arg Pro
195 200 205
Tyr Lys Tyr Ile Leu Ile Val Leu Leu Val Met Leu Ile Ile Phe Ala
210 215 220
Val Thr Pro Met Pro Arg Ala Lys Ala Thr Lys Glu Ile Gly Gly Glu
225 230 235 240
Gln Gln Glu Glu Arg Pro Ser Leu Gly Glu Thr Leu Lys Tyr Leu Ser
245 250 255
His Asn Lys His Tyr Ile Lys Gly Val Val Thr Gln Phe Phe Tyr Ala
260 265 270
Gly Leu Gln Thr Thr Val Trp Ser Phe Thr Ile Arg Leu Val Leu Asn
275 280 285
Leu Asn His Gln Ile Thr Asp Ser Gly Ala Ser Thr Phe Met Ile Tyr
290 295 300
Ser Tyr Val Ala Trp Phe Val Gly Lys Leu Val Ala Asn Thr Phe Met
305 310 315 320
Ser Arg Phe Ser Ile Thr Lys Val Leu Thr Trp Tyr Ser Leu Leu Gly
325 330 335
Thr Leu Ala Leu Val Val Thr Phe Thr Val Pro Asn Met Ile Ala Val
340 345 350
Tyr Ala Ala Ile Leu Thr Ser Phe Phe Phe Gly Pro Glu Trp Pro Thr
355 360 365
Ile Tyr Ala His Thr Leu Asp Ala Val Thr Glu Lys Lys Tyr Thr Glu
370 375 380
Thr Ala Gly Ala Ile Ile Val Met Ala Leu Ile Gly Gly Ala Val Ile
385 390 395 400
Pro Ala Ile Gln Gly Leu Val Ser Asp Ala Thr Gly Ser Met Gln Phe
405 410 415
Ser Phe Val Val Pro Met Leu Cys Tyr Ala Leu Ile Thr Gly Tyr Phe
420 425 430
Phe Phe Glu His Arg Phe Glu Lys Ala His Pro Asn Glu Val Gln Glu
435 440 445
His
<210> 70
<211> 2379
<212> DNA
<213> 大肠杆菌
<400> 70
atgtccaaca atggctcgtc accgctggtg ctttggtata accaactcgg catgaatgat 60
gtagacaggg ttgggggcaa aaatgcctcc ctgggtgaaa tgattactaa tctttccgga 120
atgggtgttt ccgttccgaa tggtttcgcc acaaccgccg acgcgtttaa ccagtttctg 180
gaccaaagcg gcgtaaacca gcgcatttat gaactgctgg ataaaacgga tattgacgat 240
gttactcagc ttgcgaaagc gggcgcgcaa atccgccagt ggattatcga cactcccttc 300
cagcctgagc tggaaaacgc catccgcgaa gcctatgcac agctttccgc cgatgacgaa 360
aacgcctctt ttgcggtgcg ctcctccgcc accgcagaag atatgccgga cgcttctttt 420
gccggtcagc aggaaacctt cctcaacgtt cagggttttg acgccgttct cgtggcagtg 480
aaacatgtat ttgcttctct gtttaacgat cgcgccatct cttatcgtgt gcaccagggt 540
tacgatcacc gtggtgtggc gctctccgcc ggtgttcaac ggatggtgcg ctctgacctc 600
gcatcatctg gcgtgatgtt ctccattgat accgaatccg gctttgacca ggtggtgttt 660
atcacttccg catggggcct tggtgagatg gtcgtgcagg gtgcggttaa cccggatgag 720
ttttacgtgc ataaaccgac actggcggcg aatcgcccgg ctatcgtgcg ccgcaccatg 780
gggtcgaaaa aaatccgcat ggtttacgcg ccgacccagg agcacggcaa gcaggttaaa 840
atcgaagacg taccgcagga acagcgtgac atcttctcgc tgaccaacga agaagtgcag 900
gaactggcaa aacaggccgt acaaattgag aaacactacg gtcgcccgat ggatattgag 960
tgggcgaaag atggccacac cggtaaactg ttcattgtgc aggcgcgtcc ggaaaccgtg 1020
cgctcacgcg gtcaggtcat ggagcgttat acgctgcatt cacagggtaa gattatcgcc 1080
gaaggccgtg ctatcggtca tcgcatcggt gcgggtccgg tgaaagtcat ccatgacatc 1140
agcgaaatga accgcatcga acctggcgac gtgctggtta ctgacatgac cgacccggac 1200
tgggaaccga tcatgaagaa agcatctgcc atcgtcacca accgtggcgg tcgtacctgt 1260
cacgcggcga tcatcgctcg tgaactgggc attccggcgg tagtgggctg tggagatgca 1320
acagaacgga tgaaagacgg tgagaacgtc actgtttctt gtgccgaagg tgataccggt 1380
tacgtctatg cggagttgct ggaatttagc gtgaaaagct ccagcgtaga aacgatgccg 1440
gatctgccgt tgaaagtgat gatgaacgtc ggtaacccgg accgtgcttt cgacttcgcc 1500
tgcctaccga acgaaggcgt gggccttgcg cgtctggaat ttatcatcaa ccgtatgatt 1560
ggcgtccacc cacgcgcact gcttgagttt gacgatcagg aaccgcagtt gcaaaacgaa 1620
atccgcgaga tgatgaaagg ttttgattct ccgcgtgaat tttacgttgg tcgtctgact 1680
gaagggatcg cgacgctggg tgccgcgttt tatccgaagc gcgtcattgt ccgtctctct 1740
gattttaaat cgaacgaata tgccaacctg gtcggtggtg agcgttacga gccagatgaa 1800
gagaacccga tgctcggctt ccgtggcgcg ggccgctatg tttccgacag cttccgcgac 1860
tgtttcgcgc tggagtgtga agcagtgaaa cgtgtgcgca acgacatggg actgaccaac 1920
gttgagatca tgatcccgtt cgtgcgtacc gtagatcagg cgaaagcggt ggttgaagaa 1980
ctggcgcgtc aggggctgaa acgtggcgag aacgggctga aaatcatcat gatgtgtgaa 2040
atcccgtcca acgccttgct ggccgagcag ttcctcgaat atttcgacgg cttctcaatt 2100
ggctcaaacg atatgacgca gctggcgctc ggtctggacc gtgactccgg cgtggtgtct 2160
gaattgttcg atgagcgcaa cgatgcggtg aaagcactgc tgtcgatggc tatccgtgcc 2220
gcgaagaaac agggcaaata tgtcgggatt tgcggtcagg gtccgtccga ccacgaagac 2280
tttgccgcat ggttgatgga agaggggatc gatagcctgt ctctgaaccc ggacaccgtg 2340
gtgcaaacct ggttaagcct ggctgaactg aagaaataa 2379
<210> 71
<211> 792
<212> PRT
<213> 大肠杆菌
<400> 71
Met Ser Asn Asn Gly Ser Ser Pro Leu Val Leu Trp Tyr Asn Gln Leu
1 5 10 15
Gly Met Asn Asp Val Asp Arg Val Gly Gly Lys Asn Ala Ser Leu Gly
20 25 30
Glu Met Ile Thr Asn Leu Ser Gly Met Gly Val Ser Val Pro Asn Gly
35 40 45
Phe Ala Thr Thr Ala Asp Ala Phe Asn Gln Phe Leu Asp Gln Ser Gly
50 55 60
Val Asn Gln Arg Ile Tyr Glu Leu Leu Asp Lys Thr Asp Ile Asp Asp
65 70 75 80
Val Thr Gln Leu Ala Lys Ala Gly Ala Gln Ile Arg Gln Trp Ile Ile
85 90 95
Asp Thr Pro Phe Gln Pro Glu Leu Glu Asn Ala Ile Arg Glu Ala Tyr
100 105 110
Ala Gln Leu Ser Ala Asp Asp Glu Asn Ala Ser Phe Ala Val Arg Ser
115 120 125
Ser Ala Thr Ala Glu Asp Met Pro Asp Ala Ser Phe Ala Gly Gln Gln
130 135 140
Glu Thr Phe Leu Asn Val Gln Gly Phe Asp Ala Val Leu Val Ala Val
145 150 155 160
Lys His Val Phe Ala Ser Leu Phe Asn Asp Arg Ala Ile Ser Tyr Arg
165 170 175
Val His Gln Gly Tyr Asp His Arg Gly Val Ala Leu Ser Ala Gly Val
180 185 190
Gln Arg Met Val Arg Ser Asp Leu Ala Ser Ser Gly Val Met Phe Ser
195 200 205
Ile Asp Thr Glu Ser Gly Phe Asp Gln Val Val Phe Ile Thr Ser Ala
210 215 220
Trp Gly Leu Gly Glu Met Val Val Gln Gly Ala Val Asn Pro Asp Glu
225 230 235 240
Phe Tyr Val His Lys Pro Thr Leu Ala Ala Asn Arg Pro Ala Ile Val
245 250 255
Arg Arg Thr Met Gly Ser Lys Lys Ile Arg Met Val Tyr Ala Pro Thr
260 265 270
Gln Glu His Gly Lys Gln Val Lys Ile Glu Asp Val Pro Gln Glu Gln
275 280 285
Arg Asp Ile Phe Ser Leu Thr Asn Glu Glu Val Gln Glu Leu Ala Lys
290 295 300
Gln Ala Val Gln Ile Glu Lys His Tyr Gly Arg Pro Met Asp Ile Glu
305 310 315 320
Trp Ala Lys Asp Gly His Thr Gly Lys Leu Phe Ile Val Gln Ala Arg
325 330 335
Pro Glu Thr Val Arg Ser Arg Gly Gln Val Met Glu Arg Tyr Thr Leu
340 345 350
His Ser Gln Gly Lys Ile Ile Ala Glu Gly Arg Ala Ile Gly His Arg
355 360 365
Ile Gly Ala Gly Pro Val Lys Val Ile His Asp Ile Ser Glu Met Asn
370 375 380
Arg Ile Glu Pro Gly Asp Val Leu Val Thr Asp Met Thr Asp Pro Asp
385 390 395 400
Trp Glu Pro Ile Met Lys Lys Ala Ser Ala Ile Val Thr Asn Arg Gly
405 410 415
Gly Arg Thr Cys His Ala Ala Ile Ile Ala Arg Glu Leu Gly Ile Pro
420 425 430
Ala Val Val Gly Cys Gly Asp Ala Thr Glu Arg Met Lys Asp Gly Glu
435 440 445
Asn Val Thr Val Ser Cys Ala Glu Gly Asp Thr Gly Tyr Val Tyr Ala
450 455 460
Glu Leu Leu Glu Phe Ser Val Lys Ser Ser Ser Val Glu Thr Met Pro
465 470 475 480
Asp Leu Pro Leu Lys Val Met Met Asn Val Gly Asn Pro Asp Arg Ala
485 490 495
Phe Asp Phe Ala Cys Leu Pro Asn Glu Gly Val Gly Leu Ala Arg Leu
500 505 510
Glu Phe Ile Ile Asn Arg Met Ile Gly Val His Pro Arg Ala Leu Leu
515 520 525
Glu Phe Asp Asp Gln Glu Pro Gln Leu Gln Asn Glu Ile Arg Glu Met
530 535 540
Met Lys Gly Phe Asp Ser Pro Arg Glu Phe Tyr Val Gly Arg Leu Thr
545 550 555 560
Glu Gly Ile Ala Thr Leu Gly Ala Ala Phe Tyr Pro Lys Arg Val Ile
565 570 575
Val Arg Leu Ser Asp Phe Lys Ser Asn Glu Tyr Ala Asn Leu Val Gly
580 585 590
Gly Glu Arg Tyr Glu Pro Asp Glu Glu Asn Pro Met Leu Gly Phe Arg
595 600 605
Gly Ala Gly Arg Tyr Val Ser Asp Ser Phe Arg Asp Cys Phe Ala Leu
610 615 620
Glu Cys Glu Ala Val Lys Arg Val Arg Asn Asp Met Gly Leu Thr Asn
625 630 635 640
Val Glu Ile Met Ile Pro Phe Val Arg Thr Val Asp Gln Ala Lys Ala
645 650 655
Val Val Glu Glu Leu Ala Arg Gln Gly Leu Lys Arg Gly Glu Asn Gly
660 665 670
Leu Lys Ile Ile Met Met Cys Glu Ile Pro Ser Asn Ala Leu Leu Ala
675 680 685
Glu Gln Phe Leu Glu Tyr Phe Asp Gly Phe Ser Ile Gly Ser Asn Asp
690 695 700
Met Thr Gln Leu Ala Leu Gly Leu Asp Arg Asp Ser Gly Val Val Ser
705 710 715 720
Glu Leu Phe Asp Glu Arg Asn Asp Ala Val Lys Ala Leu Leu Ser Met
725 730 735
Ala Ile Arg Ala Ala Lys Lys Gln Gly Lys Tyr Val Gly Ile Cys Gly
740 745 750
Gln Gly Pro Ser Asp His Glu Asp Phe Ala Ala Trp Leu Met Glu Glu
755 760 765
Gly Ile Asp Ser Leu Ser Leu Asn Pro Asp Thr Val Val Gln Thr Trp
770 775 780
Leu Ser Leu Ala Glu Leu Lys Lys
785 790
<210> 72
<211> 1623
<212> DNA
<213> 大肠杆菌
<400> 72
atgcgcgtta acaatggttt gaccccgcaa gaactcgagg cttatggtat cagtgacgta 60
catgatatcg tttacaaccc aagctacgac ctgctgtatc aggaagagct cgatccgagc 120
ctgacaggtt atgagcgcgg ggtgttaact aatctgggtg ccgttgccgt cgataccggg 180
atcttcaccg gtcgttcacc aaaagataag tatatcgtcc gtgacgatac cactcgcgat 240
actttctggt gggcagacaa aggcaaaggt aagaacgaca acaaacctct ctctccggaa 300
acctggcagc atctgaaagg cctggtgacc aggcagcttt ccggcaaacg tctgttcgtt 360
gtcgacgctt tctgtggtgc gaacccggat actcgtcttt ccgtccgttt catcaccgaa 420
gtggcctggc aggcgcattt tgtcaaaaac atgtttattc gcccgagcga tgaagaactg 480
gcaggtttca aaccagactt tatcgttatg aacggcgcga agtgcactaa cccgcagtgg 540
aaagaacagg gtctcaactc cgaaaacttc gtggcgttta acctgaccga gcgcatgcag 600
ctgattggcg gcacctggta cggcggcgaa atgaagaaag ggatgttctc gatgatgaac 660
tacctgctgc cgctgaaagg tatcgcttct atgcactgct ccgccaacgt tggtgagaaa 720
ggcgatgttg cggtgttctt cggcctttcc ggcaccggta aaaccaccct ttccaccgac 780
ccgaaacgtc gcctgattgg cgatgacgaa cacggctggg acgatgacgg cgtgtttaac 840
ttcgaaggcg gctgctacgc aaaaactatc aagctgtcga aagaagcgga acctgaaatc 900
tacaacgcta tccgtcgtga tgcgttgctg gaaaacgtca ccgtgcgtga agatggcact 960
atcgactttg atgatggttc aaaaaccgag aacacccgcg tttcttatcc gatctatcac 1020
atcgataaca ttgttaagcc ggtttccaaa gcgggccacg cgactaaggt tatcttcctg 1080
actgctgatg ctttcggcgt gttgccgccg gtttctcgcc tgactgccga tcaaacccag 1140
tatcacttcc tctctggctt caccgccaaa ctggccggta ctgagcgtgg catcaccgaa 1200
ccgacgccaa ccttctccgc ttgcttcggc gcggcattcc tgtcgctgca cccgactcag 1260
tacgcagaag tgctggtgaa acgtatgcag gcggcgggcg cgcaggctta tctggttaac 1320
actggctgga acggcactgg caaacgtatc tcgattaaag atacccgcgc cattatcgac 1380
gccatcctca acggttcgct ggataatgca gaaaccttca ctctgccgat gtttaacctg 1440
gcgatcccaa ccgaactgcc gggcgtagac acgaagattc tcgatccgcg taacacctac 1500
gcttctccgg aacagtggca ggaaaaagcc gaaaccctgg cgaaactgtt tatcgacaac 1560
ttcgataaat acaccgacac ccctgcgggt gccgcgctgg tagcggctgg tccgaaactg 1620
taa 1623
<210> 73
<211> 540
<212> PRT
<213> 大肠杆菌
<400> 73
Met Arg Val Asn Asn Gly Leu Thr Pro Gln Glu Leu Glu Ala Tyr Gly
1 5 10 15
Ile Ser Asp Val His Asp Ile Val Tyr Asn Pro Ser Tyr Asp Leu Leu
20 25 30
Tyr Gln Glu Glu Leu Asp Pro Ser Leu Thr Gly Tyr Glu Arg Gly Val
35 40 45
Leu Thr Asn Leu Gly Ala Val Ala Val Asp Thr Gly Ile Phe Thr Gly
50 55 60
Arg Ser Pro Lys Asp Lys Tyr Ile Val Arg Asp Asp Thr Thr Arg Asp
65 70 75 80
Thr Phe Trp Trp Ala Asp Lys Gly Lys Gly Lys Asn Asp Asn Lys Pro
85 90 95
Leu Ser Pro Glu Thr Trp Gln His Leu Lys Gly Leu Val Thr Arg Gln
100 105 110
Leu Ser Gly Lys Arg Leu Phe Val Val Asp Ala Phe Cys Gly Ala Asn
115 120 125
Pro Asp Thr Arg Leu Ser Val Arg Phe Ile Thr Glu Val Ala Trp Gln
130 135 140
Ala His Phe Val Lys Asn Met Phe Ile Arg Pro Ser Asp Glu Glu Leu
145 150 155 160
Ala Gly Phe Lys Pro Asp Phe Ile Val Met Asn Gly Ala Lys Cys Thr
165 170 175
Asn Pro Gln Trp Lys Glu Gln Gly Leu Asn Ser Glu Asn Phe Val Ala
180 185 190
Phe Asn Leu Thr Glu Arg Met Gln Leu Ile Gly Gly Thr Trp Tyr Gly
195 200 205
Gly Glu Met Lys Lys Gly Met Phe Ser Met Met Asn Tyr Leu Leu Pro
210 215 220
Leu Lys Gly Ile Ala Ser Met His Cys Ser Ala Asn Val Gly Glu Lys
225 230 235 240
Gly Asp Val Ala Val Phe Phe Gly Leu Ser Gly Thr Gly Lys Thr Thr
245 250 255
Leu Ser Thr Asp Pro Lys Arg Arg Leu Ile Gly Asp Asp Glu His Gly
260 265 270
Trp Asp Asp Asp Gly Val Phe Asn Phe Glu Gly Gly Cys Tyr Ala Lys
275 280 285
Thr Ile Lys Leu Ser Lys Glu Ala Glu Pro Glu Ile Tyr Asn Ala Ile
290 295 300
Arg Arg Asp Ala Leu Leu Glu Asn Val Thr Val Arg Glu Asp Gly Thr
305 310 315 320
Ile Asp Phe Asp Asp Gly Ser Lys Thr Glu Asn Thr Arg Val Ser Tyr
325 330 335
Pro Ile Tyr His Ile Asp Asn Ile Val Lys Pro Val Ser Lys Ala Gly
340 345 350
His Ala Thr Lys Val Ile Phe Leu Thr Ala Asp Ala Phe Gly Val Leu
355 360 365
Pro Pro Val Ser Arg Leu Thr Ala Asp Gln Thr Gln Tyr His Phe Leu
370 375 380
Ser Gly Phe Thr Ala Lys Leu Ala Gly Thr Glu Arg Gly Ile Thr Glu
385 390 395 400
Pro Thr Pro Thr Phe Ser Ala Cys Phe Gly Ala Ala Phe Leu Ser Leu
405 410 415
His Pro Thr Gln Tyr Ala Glu Val Leu Val Lys Arg Met Gln Ala Ala
420 425 430
Gly Ala Gln Ala Tyr Leu Val Asn Thr Gly Trp Asn Gly Thr Gly Lys
435 440 445
Arg Ile Ser Ile Lys Asp Thr Arg Ala Ile Ile Asp Ala Ile Leu Asn
450 455 460
Gly Ser Leu Asp Asn Ala Glu Thr Phe Thr Leu Pro Met Phe Asn Leu
465 470 475 480
Ala Ile Pro Thr Glu Leu Pro Gly Val Asp Thr Lys Ile Leu Asp Pro
485 490 495
Arg Asn Thr Tyr Ala Ser Pro Glu Gln Trp Gln Glu Lys Ala Glu Thr
500 505 510
Leu Ala Lys Leu Phe Ile Asp Asn Phe Asp Lys Tyr Thr Asp Thr Pro
515 520 525
Ala Gly Ala Ala Leu Val Ala Ala Gly Pro Lys Leu
530 535 540
<210> 74
<211> 1443
<212> DNA
<213> 大肠杆菌
<400> 74
atgtccagaa ggcttcgcag aacaaaaatc gttaccacgt taggcccagc aacagatcgc 60
gataataatc ttgaaaaagt tatcgcggcg ggtgccaacg ttgtacgtat gaacttttct 120
cacggctcgc ctgaagatca caaaatgcgc gcggataaag ttcgtgagat tgccgcaaaa 180
ctggggcgtc atgtggctat tctgggtgac ctccaggggc ccaaaatccg tgtatccacc 240
tttaaagaag gcaaagtttt cctcaatatt ggggataaat tcctgctcga cgccaacctg 300
ggtaaaggtg aaggcgacaa agaaaaagtc ggtatcgact acaaaggcct gcctgctgac 360
gtcgtgcctg gtgacatcct gctgctggac gatggtcgcg tccagttaaa agtactggaa 420
gttcagggca tgaaagtgtt caccgaagtc accgtcggtg gtcccctctc caacaataaa 480
ggtatcaaca aacttggcgg cggtttgtcg gctgaagcgc tgaccgaaaa agacaaagca 540
gacattaaga ctgcggcgtt gattggcgta gattacctgg ctgtctcctt cccacgctgt 600
ggcgaagatc tgaactatgc ccgtcgcctg gcacgcgatg caggatgtga tgcgaaaatt 660
gttgccaagg ttgaacgtgc ggaagccgtt tgcagccagg atgcaatgga tgacatcatc 720
ctcgcctctg acgtggtaat ggttgcacgt ggcgacctcg gtgtggaaat tggcgacccg 780
gaactggtcg gcattcagaa agcgttgatc cgtcgtgcgc gtcagctaaa ccgagcggta 840
atcacggcga cccagatgat ggagtcaatg attactaacc cgatgccgac gcgtgcagaa 900
gtcatggacg tagcaaacgc cgttctggat ggtactgacg ctgtgatgct gtctgcagaa 960
actgccgctg ggcagtatcc gtcagaaacc gttgcagcca tggcgcgcgt ttgcctgggt 1020
gcggaaaaaa tcccgagcat caacgtttct aaacaccgtc tggacgttca gttcgacaat 1080
gtggaagaag ctattgccat gtcagcaatg tacgcagcta accacctgaa aggcgttacg 1140
gcgatcatca ccatgaccga atcgggtcgt accgcgctga tgacctcccg tatcagctct 1200
ggtctgccaa ttttcgccat gtcgcgccat gaacgtacgc tgaacctgac tgctctctat 1260
cgtggcgtta cgccggtgca ctttgatagc gctaatgacg gcgtagcagc tgccagcgaa 1320
gcggttaatc tgctgcgcga taaaggttac ttgatgtctg gtgacctggt gattgtcacc 1380
cagggcgacg tgatgagtac cgtgggttct actaatacca cgcgtatttt aacggtagag 1440
taa 1443
<210> 75
<211> 480
<212> PRT
<213> 大肠杆菌
<400> 75
Met Ser Arg Arg Leu Arg Arg Thr Lys Ile Val Thr Thr Leu Gly Pro
1 5 10 15
Ala Thr Asp Arg Asp Asn Asn Leu Glu Lys Val Ile Ala Ala Gly Ala
20 25 30
Asn Val Val Arg Met Asn Phe Ser His Gly Ser Pro Glu Asp His Lys
35 40 45
Met Arg Ala Asp Lys Val Arg Glu Ile Ala Ala Lys Leu Gly Arg His
50 55 60
Val Ala Ile Leu Gly Asp Leu Gln Gly Pro Lys Ile Arg Val Ser Thr
65 70 75 80
Phe Lys Glu Gly Lys Val Phe Leu Asn Ile Gly Asp Lys Phe Leu Leu
85 90 95
Asp Ala Asn Leu Gly Lys Gly Glu Gly Asp Lys Glu Lys Val Gly Ile
100 105 110
Asp Tyr Lys Gly Leu Pro Ala Asp Val Val Pro Gly Asp Ile Leu Leu
115 120 125
Leu Asp Asp Gly Arg Val Gln Leu Lys Val Leu Glu Val Gln Gly Met
130 135 140
Lys Val Phe Thr Glu Val Thr Val Gly Gly Pro Leu Ser Asn Asn Lys
145 150 155 160
Gly Ile Asn Lys Leu Gly Gly Gly Leu Ser Ala Glu Ala Leu Thr Glu
165 170 175
Lys Asp Lys Ala Asp Ile Lys Thr Ala Ala Leu Ile Gly Val Asp Tyr
180 185 190
Leu Ala Val Ser Phe Pro Arg Cys Gly Glu Asp Leu Asn Tyr Ala Arg
195 200 205
Arg Leu Ala Arg Asp Ala Gly Cys Asp Ala Lys Ile Val Ala Lys Val
210 215 220
Glu Arg Ala Glu Ala Val Cys Ser Gln Asp Ala Met Asp Asp Ile Ile
225 230 235 240
Leu Ala Ser Asp Val Val Met Val Ala Arg Gly Asp Leu Gly Val Glu
245 250 255
Ile Gly Asp Pro Glu Leu Val Gly Ile Gln Lys Ala Leu Ile Arg Arg
260 265 270
Ala Arg Gln Leu Asn Arg Ala Val Ile Thr Ala Thr Gln Met Met Glu
275 280 285
Ser Met Ile Thr Asn Pro Met Pro Thr Arg Ala Glu Val Met Asp Val
290 295 300
Ala Asn Ala Val Leu Asp Gly Thr Asp Ala Val Met Leu Ser Ala Glu
305 310 315 320
Thr Ala Ala Gly Gln Tyr Pro Ser Glu Thr Val Ala Ala Met Ala Arg
325 330 335
Val Cys Leu Gly Ala Glu Lys Ile Pro Ser Ile Asn Val Ser Lys His
340 345 350
Arg Leu Asp Val Gln Phe Asp Asn Val Glu Glu Ala Ile Ala Met Ser
355 360 365
Ala Met Tyr Ala Ala Asn His Leu Lys Gly Val Thr Ala Ile Ile Thr
370 375 380
Met Thr Glu Ser Gly Arg Thr Ala Leu Met Thr Ser Arg Ile Ser Ser
385 390 395 400
Gly Leu Pro Ile Phe Ala Met Ser Arg His Glu Arg Thr Leu Asn Leu
405 410 415
Thr Ala Leu Tyr Arg Gly Val Thr Pro Val His Phe Asp Ser Ala Asn
420 425 430
Asp Gly Val Ala Ala Ala Ser Glu Ala Val Asn Leu Leu Arg Asp Lys
435 440 445
Gly Tyr Leu Met Ser Gly Asp Leu Val Ile Val Thr Gln Gly Asp Val
450 455 460
Met Ser Thr Val Gly Ser Thr Asn Thr Thr Arg Ile Leu Thr Val Glu
465 470 475 480
<210> 76
<211> 1413
<212> DNA
<213> 大肠杆菌
<400> 76
atgaaaaaga ccaaaattgt ttgcaccatc ggaccgaaaa ccgaatctga agagatgtta 60
gctaaaatgc tggacgctgg catgaacgtt atgcgtctga acttctctca tggtgactat 120
gcagaacacg gtcagcgcat tcagaatctg cgcaacgtga tgagcaaaac tggtaaaacc 180
gccgctatcc tgcttgatac caaaggtccg gaaatccgca ccatgaaact ggaaggcggt 240
aacgacgttt ctctgaaagc tggtcagacc tttactttca ccactgataa atctgttatc 300
ggcaacagcg aaatggttgc ggtaacgtat gaaggtttca ctactgacct gtctgttggc 360
aacaccgtac tggttgacga tggtctgatc ggtatggaag ttaccgccat tgaaggtaac 420
aaagttatct gtaaagtgct gaacaacggt gacctgggcg aaaacaaagg tgtgaacctg 480
cctggcgttt ccattgctct gccagcactg gctgaaaaag acaaacagga cctgatcttt 540
ggttgcgaac aaggcgtaga ctttgttgct gcttccttta ttcgtaagcg ttctgacgtt 600
atcgaaatcc gtgagcacct gaaagcgcac ggcggcgaaa acatccacat catctccaaa 660
atcgaaaacc aggaaggcct caacaacttc gacgaaatcc tcgaagcctc tgacggcatc 720
atggttgcgc gtggcgacct gggtgtagaa atcccggtag aagaagttat cttcgcccag 780
aagatgatga tcgaaaaatg tatccgtgca cgtaaagtcg ttatcactgc gacccagatg 840
ctggattcca tgatcaaaaa cccacgcccg actcgcgcag aagccggtga cgttgcaaac 900
gccatcctcg acggtactga cgcagtgatg ctgtctggtg aatccgcaaa aggtaaatac 960
ccgctggaag cggtttctat catggcgacc atctgcgaac gtaccgaccg cgtgatgaac 1020
agccgtctcg agttcaacaa tgacaaccgt aaactgcgca ttaccgaagc ggtatgccgt 1080
ggtgccgttg aaactgctga aaaactggat gctccgctga tcgtggttgc tactcagggc 1140
ggtaaatctg ctcgcgcagt acgtaaatac ttcccggatg ccaccatcct ggcactgacc 1200
accaacgaaa aaacggctca tcagttggta ctgagcaaag gcgttgtgcc gcagcttgtt 1260
aaagagatca cttctactga tgatttctac cgtctgggta aagaactggc tctgcagagc 1320
ggtctggcac acaaaggtga cgttgtagtt atggtttctg gtgcactggt accgagcggc 1380
actactaaca ccgcatctgt tcacgtcctg taa 1413
<210> 77
<211> 470
<212> PRT
<213> 大肠杆菌
<400> 77
Met Lys Lys Thr Lys Ile Val Cys Thr Ile Gly Pro Lys Thr Glu Ser
1 5 10 15
Glu Glu Met Leu Ala Lys Met Leu Asp Ala Gly Met Asn Val Met Arg
20 25 30
Leu Asn Phe Ser His Gly Asp Tyr Ala Glu His Gly Gln Arg Ile Gln
35 40 45
Asn Leu Arg Asn Val Met Ser Lys Thr Gly Lys Thr Ala Ala Ile Leu
50 55 60
Leu Asp Thr Lys Gly Pro Glu Ile Arg Thr Met Lys Leu Glu Gly Gly
65 70 75 80
Asn Asp Val Ser Leu Lys Ala Gly Gln Thr Phe Thr Phe Thr Thr Asp
85 90 95
Lys Ser Val Ile Gly Asn Ser Glu Met Val Ala Val Thr Tyr Glu Gly
100 105 110
Phe Thr Thr Asp Leu Ser Val Gly Asn Thr Val Leu Val Asp Asp Gly
115 120 125
Leu Ile Gly Met Glu Val Thr Ala Ile Glu Gly Asn Lys Val Ile Cys
130 135 140
Lys Val Leu Asn Asn Gly Asp Leu Gly Glu Asn Lys Gly Val Asn Leu
145 150 155 160
Pro Gly Val Ser Ile Ala Leu Pro Ala Leu Ala Glu Lys Asp Lys Gln
165 170 175
Asp Leu Ile Phe Gly Cys Glu Gln Gly Val Asp Phe Val Ala Ala Ser
180 185 190
Phe Ile Arg Lys Arg Ser Asp Val Ile Glu Ile Arg Glu His Leu Lys
195 200 205
Ala His Gly Gly Glu Asn Ile His Ile Ile Ser Lys Ile Glu Asn Gln
210 215 220
Glu Gly Leu Asn Asn Phe Asp Glu Ile Leu Glu Ala Ser Asp Gly Ile
225 230 235 240
Met Val Ala Arg Gly Asp Leu Gly Val Glu Ile Pro Val Glu Glu Val
245 250 255
Ile Phe Ala Gln Lys Met Met Ile Glu Lys Cys Ile Arg Ala Arg Lys
260 265 270
Val Val Ile Thr Ala Thr Gln Met Leu Asp Ser Met Ile Lys Asn Pro
275 280 285
Arg Pro Thr Arg Ala Glu Ala Gly Asp Val Ala Asn Ala Ile Leu Asp
290 295 300
Gly Thr Asp Ala Val Met Leu Ser Gly Glu Ser Ala Lys Gly Lys Tyr
305 310 315 320
Pro Leu Glu Ala Val Ser Ile Met Ala Thr Ile Cys Glu Arg Thr Asp
325 330 335
Arg Val Met Asn Ser Arg Leu Glu Phe Asn Asn Asp Asn Arg Lys Leu
340 345 350
Arg Ile Thr Glu Ala Val Cys Arg Gly Ala Val Glu Thr Ala Glu Lys
355 360 365
Leu Asp Ala Pro Leu Ile Val Val Ala Thr Gln Gly Gly Lys Ser Ala
370 375 380
Arg Ala Val Arg Lys Tyr Phe Pro Asp Ala Thr Ile Leu Ala Leu Thr
385 390 395 400
Thr Asn Glu Lys Thr Ala His Gln Leu Val Leu Ser Lys Gly Val Val
405 410 415
Pro Gln Leu Val Lys Glu Ile Thr Ser Thr Asp Asp Phe Tyr Arg Leu
420 425 430
Gly Lys Glu Leu Ala Leu Gln Ser Gly Leu Ala His Lys Gly Asp Val
435 440 445
Val Val Met Val Ser Gly Ala Leu Val Pro Ser Gly Thr Thr Asn Thr
450 455 460
Ala Ser Val His Val Leu
465 470
<210> 78
<211> 1410
<212> DNA
<213> 大肠杆菌
<400> 78
atgtccgctg aacacgtact gacgatgctg aacgagcacg aagtgaagtt tgttgatttg 60
cgcttcaccg atactaaagg taaagaacag cacgtcacta tccctgctca tcaggtgaat 120
gctgaattct tcgaagaagg caaaatgttt gacggctcct cgattggcgg ctggaaaggc 180
attaacgagt ccgacatggt gctgatgcca gacgcatcca ccgcagtgat tgacccgttc 240
ttcgccgact ccaccctgat tatccgttgc gacatccttg aacctggcac cctgcaaggc 300
tatgaccgtg acccgcgctc cattgcgaag cgcgccgaag attacctgcg ttccactggc 360
attgccgaca ccgtactgtt cgggccagaa cctgaattct tcctgttcga tgacatccgt 420
ttcggatcat ctatctccgg ttcccacgtt gctatcgacg atatcgaagg cgcatggaac 480
tcctccaccc aatacgaagg tggtaacaaa ggtcaccgtc cggcagtgaa aggcggttac 540
ttcccggttc caccggtaga ctcggctcag gatattcgtt ctgaaatgtg tctggtgatg 600
gaacagatgg gtctggtggt tgaagcccat caccacgaag tagcgactgc tggtcagaac 660
gaagtggcta cccgcttcaa taccatgacc aaaaaagctg acgaaattca gatctacaaa 720
tatgttgtgc acaacgtagc gcaccgcttc ggtaaaaccg cgacctttat gccaaaaccg 780
atgttcggtg ataacggctc cggtatgcac tgccacatgt ctctgtctaa aaacggcgtt 840
aacctgttcg caggcgacaa atacgcaggt ctgtctgagc aggcgctgta ctacattggc 900
ggcgtaatca aacacgctaa agcgattaac gccctggcaa acccgaccac caactcttat 960
aagcgtctgg tcccgggcta tgaagcaccg gtaatgctgg cttactctgc gcgtaaccgt 1020
tctgcgtcta tccgtattcc ggtggtttct tctccgaaag cacgtcgtat cgaagtacgt 1080
ttcccggatc cggcagctaa cccgtacctg tgctttgctg ccctgctgat ggccggtctt 1140
gatggtatca agaacaagat ccatccgggc gaagccatgg acaaaaacct gtatgacctg 1200
ccgccagaag aagcgaaaga gatcccacag gttgcaggct ctctggaaga agcactgaac 1260
gaactggatc tggaccgcga gttcctgaaa gccggtggcg tgttcactga cgaagcaatt 1320
gatgcgtaca tcgctctgcg tcgcgaagaa gatgaccgcg tgcgtatgac tccgcatccg 1380
gtagagtttg agctgtacta cagcgtctaa 1410
<210> 79
<211> 469
<212> PRT
<213> 大肠杆菌
<400> 79
Met Ser Ala Glu His Val Leu Thr Met Leu Asn Glu His Glu Val Lys
1 5 10 15
Phe Val Asp Leu Arg Phe Thr Asp Thr Lys Gly Lys Glu Gln His Val
20 25 30
Thr Ile Pro Ala His Gln Val Asn Ala Glu Phe Phe Glu Glu Gly Lys
35 40 45
Met Phe Asp Gly Ser Ser Ile Gly Gly Trp Lys Gly Ile Asn Glu Ser
50 55 60
Asp Met Val Leu Met Pro Asp Ala Ser Thr Ala Val Ile Asp Pro Phe
65 70 75 80
Phe Ala Asp Ser Thr Leu Ile Ile Arg Cys Asp Ile Leu Glu Pro Gly
85 90 95
Thr Leu Gln Gly Tyr Asp Arg Asp Pro Arg Ser Ile Ala Lys Arg Ala
100 105 110
Glu Asp Tyr Leu Arg Ser Thr Gly Ile Ala Asp Thr Val Leu Phe Gly
115 120 125
Pro Glu Pro Glu Phe Phe Leu Phe Asp Asp Ile Arg Phe Gly Ser Ser
130 135 140
Ile Ser Gly Ser His Val Ala Ile Asp Asp Ile Glu Gly Ala Trp Asn
145 150 155 160
Ser Ser Thr Gln Tyr Glu Gly Gly Asn Lys Gly His Arg Pro Ala Val
165 170 175
Lys Gly Gly Tyr Phe Pro Val Pro Pro Val Asp Ser Ala Gln Asp Ile
180 185 190
Arg Ser Glu Met Cys Leu Val Met Glu Gln Met Gly Leu Val Val Glu
195 200 205
Ala His His His Glu Val Ala Thr Ala Gly Gln Asn Glu Val Ala Thr
210 215 220
Arg Phe Asn Thr Met Thr Lys Lys Ala Asp Glu Ile Gln Ile Tyr Lys
225 230 235 240
Tyr Val Val His Asn Val Ala His Arg Phe Gly Lys Thr Ala Thr Phe
245 250 255
Met Pro Lys Pro Met Phe Gly Asp Asn Gly Ser Gly Met His Cys His
260 265 270
Met Ser Leu Ser Lys Asn Gly Val Asn Leu Phe Ala Gly Asp Lys Tyr
275 280 285
Ala Gly Leu Ser Glu Gln Ala Leu Tyr Tyr Ile Gly Gly Val Ile Lys
290 295 300
His Ala Lys Ala Ile Asn Ala Leu Ala Asn Pro Thr Thr Asn Ser Tyr
305 310 315 320
Lys Arg Leu Val Pro Gly Tyr Glu Ala Pro Val Met Leu Ala Tyr Ser
325 330 335
Ala Arg Asn Arg Ser Ala Ser Ile Arg Ile Pro Val Val Ser Ser Pro
340 345 350
Lys Ala Arg Arg Ile Glu Val Arg Phe Pro Asp Pro Ala Ala Asn Pro
355 360 365
Tyr Leu Cys Phe Ala Ala Leu Leu Met Ala Gly Leu Asp Gly Ile Lys
370 375 380
Asn Lys Ile His Pro Gly Glu Ala Met Asp Lys Asn Leu Tyr Asp Leu
385 390 395 400
Pro Pro Glu Glu Ala Lys Glu Ile Pro Gln Val Ala Gly Ser Leu Glu
405 410 415
Glu Ala Leu Asn Glu Leu Asp Leu Asp Arg Glu Phe Leu Lys Ala Gly
420 425 430
Gly Val Phe Thr Asp Glu Ala Ile Asp Ala Tyr Ile Ala Leu Arg Arg
435 440 445
Glu Glu Asp Asp Arg Val Arg Met Thr Pro His Pro Val Glu Phe Glu
450 455 460
Leu Tyr Tyr Ser Val
465
<210> 80
<211> 4461
<212> DNA
<213> 大肠杆菌
<400> 80
atgttgtacg ataaatccct tgagagggat aactgtggtt tcggcctgat cgcccacata 60
gaaggcgaac ctagccacaa ggtagtgcgt actgcaatac acgcactggc ccgcatgcag 120
caccgtggcg cgattctcgc cgatggtaaa accggcgacg gttgcggctt gctgttacaa 180
aaaccggatc gcttttttcg catcgttgcg caggagcgcg gctggcgttt agcaaaaaac 240
tacgctgtcg ggatgctctt cctgaataaa gatcctgaac tcgccgctgc cgcacgccgc 300
atcgttgaag aagaactgca acgcgaaacc ttgtcgattg tgggctggcg tgatgtcccc 360
actaacgaag gcgtgctggg tgaaatcgcc ctctcctctc tgccacgcat tgagcaaatt 420
tttgtgaacg ccccggcagg ctggcgtcca cgcgatatgg agcgccgtct gtttatcgcc 480
cgccgccgca ttgaaaagcg tctcgaagcc gacaaagact tctacgtctg tagcctgtcg 540
aatctggtga acatctataa aggtctgtgt atgccgacgg atctgccgcg cttttatctg 600
gatcttgcgg acctgcgtct ggaatcggcc atttgcctgt tccaccagcg cttctccact 660
aacaccgtac cgcgctggcc gctggcgcaa ccgttccgct atctggcgca taacggtgaa 720
atcaacacca tcaccggtaa ccgccaatgg gcgcgtgcgc gtacttataa attccagaca 780
ccgcttatcc ctgacctgca cgacgccgca ccgttcgtca acgaaaccgg ctctgactcc 840
agttcgatgg ataacatgct ggaactgctg ctggcaggcg ggatggatat catccgcgcc 900
atgcgtctat tagtaccacc cgcctggcag aacaacccgg atatggaccc ggaactgcgt 960
gccttctttg actttaactc catgcatatg gagccgtggg atggcccggc gggcatcgtg 1020
atgtccgacg gtcgttttgc cgcctgtaac ctcgaccgta acggtctgcg tccggcgcgc 1080
tacgtcatca ccaaagataa gctcatcacc tgcgcctctg aagtcggtat ctgggattac 1140
cagcctgacg aagtggtcga aaaaggccgc gtcgggccag gcgaactgat ggttatcgac 1200
acccgcagtg ggcgtattct gcactcggca gaaaccgatg acgatctgaa aagccgccat 1260
ccatataaag agtggatgga gaaaaacgtc cgccgactgg taccgtttga agatctgccc 1320
gatgaagaag tgggtagccg cgaactggac gacgacacgc ttgccagcta ccagaaacag 1380
tttaactaca gcgcggaaga gctggactcc gtaattcgcg tactgggcga aaacggtcag 1440
gaagcggtcg gttcgatggg cgatgatacc ccattcgccg tgctctccag tcagccgcgc 1500
attatttacg actacttccg ccagcagttt gcccaggtga ctaacccgcc aatcgacccg 1560
ctgcgtgaag cgcatgttat gtcgctcgcc accagtatcg gtcgtgaaat gaacgtcttt 1620
tgcgaagcag agggccaggc gcaccgttta agctttaaat cgccgattct gctctactcc 1680
gatttcaaac agctcacgac gatgaaagag gagcactacc gcgcagatac gctggatatc 1740
acctttgacg tcactaaaac cacgctcgaa gcgacagtca aagagctgtg cgacaaagcc 1800
gaaaaaatgg tacgtagcgg caccgtgctg ctggtgctct ccgaccggaa tatcgctaaa 1860
gatcgcctgc cggttccagc cccgatggcg gttggcgcga tccagacccg tctggtcgat 1920
caaagcctgc gttgcgatgc caacatcatc gtcgaaaccg ccagcgcccg cgatccgcac 1980
cacttcgccg tgttgctggg cttcggcgcg acggctattt atccatacct tgcctatgaa 2040
acgctgggcc gcctggtaga cacccatgcg attgccaaag attatcgtac cgtgatgctc 2100
aactaccgta acggcatcaa caaaggcttg tacaaaatca tgtccaaaat gggcatctcc 2160
accatcgcct cttaccgctg ctcgaaactg tttgaagcgg tcggtctaca cgatgatgta 2220
gtgggcctgt gcttccaggg ggcggtcagc cgcattggtg gagcaagctt tgaagacttc 2280
cagcaggatc tgctgaatct gtcgaaacgt gcctggctgg cgcgtaagcc catcagccag 2340
ggcggtctgc tgaaatacgt ccacggcggc gaataccacg cctacaaccc ggacgtggtg 2400
cgcacgctgc aacaagcggt acaaagcggc gagtacagcg actatcagga atacgcgaag 2460
ctggttaatg agcgtccggc aaccacgctg cgcgatctgc tggcaattac gccgggtgaa 2520
aacgcggtca acattgctga tgttgaaccg gcaagcgaac tgtttaaacg ctttgatacc 2580
gccgcgatgt ctatcggcgc gttaagcccg gaagcccacg aggcgctggc ggaagcgatg 2640
aacagcatcg gcggtaattc gaactccggt gaaggcggcg aagacccggc gcgctacggc 2700
accaacaaag tgtcgcgcat caagcaggtg gcttccggtc gctttggggt tactccggcg 2760
tatctggtca atgccgacgt cattcagatt aaagtcgccc agggcgcgaa gccaggcgaa 2820
ggcggtcagt tgccgggtga taaagtcacg ccttacatcg ccaaactgcg ctattcggtg 2880
cccggagtga cgctgatctc cccgccgccg caccacgata tctactctat cgaggactta 2940
gcgcagctca ttttcgacct caagcaggtt aacccgaaag cgatgatctc cgtgaagctg 3000
gtttccgaac cgggagtagg caccatcgcg actggcgtgg caaaagctta tgcggacttg 3060
atcaccatcg caggctatga cggcggcacc ggcgcaagtc cgctttcatc ggtgaaatac 3120
gcaggctgtc cgtgggagct ggggcttgtt gaaacccagc aggcgctggt tgctaacggc 3180
ttgcgtcaca agatccgttt gcaggtcgat ggcggcctga aaacgggtgt cgatatcatc 3240
aaggcggcga ttctcggcgc agaaagcttc ggcttcggca ctggcccgat ggtggcgctc 3300
ggctgtaaat atctacgtat ttgccatctg aacaactgcg caacgggtgt agcaactcag 3360
gatgacaaac tgcgtaagaa ccactatcac ggcctgccat tcaaggtgac gaattacttt 3420
gagtttatcg cccgtgaaac ccgcgagctg atggcacagc ttggcgtaac acgtctggtg 3480
gatctgattg gtcgcaccga cctgctgaaa gagctggacg gtttcaccgc caaacagcag 3540
aagctggcgc tgtcgaagct gctggagact gccgaaccgc atccaggtaa ggcactctac 3600
tgcaccgaaa acaacccgcc gtttgataac ggcctgctga acgcgcagtt gctgcaacag 3660
gcgaaaccgt ttgtcgatga gcgccagagc aaaaccttct ggttcgatat tcgcaacacc 3720
gaccgttctg tcggcgcgtc gctttcaggc tatatcgccc agacgcacgg cgatcagggg 3780
ctggcagccg atcctatcaa agcgtacttc aacggcaccg caggccagag cttcggcgtg 3840
tggaacgcgg gcggcgtgga actgtacctg accggtgatg ccaacgacta tgtcggtaaa 3900
ggcatggcgg gcggcttaat cgccattcgt cctccggttg gttccgcctt ccgcagccat 3960
gaagcaagca ttatcggcaa cacctgcctg tatggcgcga ccggtggtcg tctgtatgcc 4020
gcaggccgcg cgggtgaacg tttcggcgtg cgtaactccg gtgctatcac cgtggtagaa 4080
ggcattggcg acaacggttg tgaatatatg acgggtggta tcgtctgcat tctgggtaaa 4140
accggcgtta acttcggtgc gggcatgacc ggcggtttcg cttacgttct cgatgaaagc 4200
ggcgatttcc gcaaacgcgt taacccggaa ctggtcgagg tcttaagcgt tgacgctctg 4260
gcgatccatg aagagcatct gcgtggtctt atcaccgagc atgtgcagca taccggctct 4320
cagcgcggtg aagagattct ggcgaactgg tcaaccttcg ccactaaatt tgcgctggtt 4380
aaaccgaagt ccagtgatgt aaaagcactg ctgggtcacc gtagtcgtag cgcagctgag 4440
ttgcgcgtgc aggcgcagta a 4461
<210> 81
<211> 1486
<212> PRT
<213> 大肠杆菌
<400> 81
Met Leu Tyr Asp Lys Ser Leu Glu Arg Asp Asn Cys Gly Phe Gly Leu
1 5 10 15
Ile Ala His Ile Glu Gly Glu Pro Ser His Lys Val Val Arg Thr Ala
20 25 30
Ile His Ala Leu Ala Arg Met Gln His Arg Gly Ala Ile Leu Ala Asp
35 40 45
Gly Lys Thr Gly Asp Gly Cys Gly Leu Leu Leu Gln Lys Pro Asp Arg
50 55 60
Phe Phe Arg Ile Val Ala Gln Glu Arg Gly Trp Arg Leu Ala Lys Asn
65 70 75 80
Tyr Ala Val Gly Met Leu Phe Leu Asn Lys Asp Pro Glu Leu Ala Ala
85 90 95
Ala Ala Arg Arg Ile Val Glu Glu Glu Leu Gln Arg Glu Thr Leu Ser
100 105 110
Ile Val Gly Trp Arg Asp Val Pro Thr Asn Glu Gly Val Leu Gly Glu
115 120 125
Ile Ala Leu Ser Ser Leu Pro Arg Ile Glu Gln Ile Phe Val Asn Ala
130 135 140
Pro Ala Gly Trp Arg Pro Arg Asp Met Glu Arg Arg Leu Phe Ile Ala
145 150 155 160
Arg Arg Arg Ile Glu Lys Arg Leu Glu Ala Asp Lys Asp Phe Tyr Val
165 170 175
Cys Ser Leu Ser Asn Leu Val Asn Ile Tyr Lys Gly Leu Cys Met Pro
180 185 190
Thr Asp Leu Pro Arg Phe Tyr Leu Asp Leu Ala Asp Leu Arg Leu Glu
195 200 205
Ser Ala Ile Cys Leu Phe His Gln Arg Phe Ser Thr Asn Thr Val Pro
210 215 220
Arg Trp Pro Leu Ala Gln Pro Phe Arg Tyr Leu Ala His Asn Gly Glu
225 230 235 240
Ile Asn Thr Ile Thr Gly Asn Arg Gln Trp Ala Arg Ala Arg Thr Tyr
245 250 255
Lys Phe Gln Thr Pro Leu Ile Pro Asp Leu His Asp Ala Ala Pro Phe
260 265 270
Val Asn Glu Thr Gly Ser Asp Ser Ser Ser Met Asp Asn Met Leu Glu
275 280 285
Leu Leu Leu Ala Gly Gly Met Asp Ile Ile Arg Ala Met Arg Leu Leu
290 295 300
Val Pro Pro Ala Trp Gln Asn Asn Pro Asp Met Asp Pro Glu Leu Arg
305 310 315 320
Ala Phe Phe Asp Phe Asn Ser Met His Met Glu Pro Trp Asp Gly Pro
325 330 335
Ala Gly Ile Val Met Ser Asp Gly Arg Phe Ala Ala Cys Asn Leu Asp
340 345 350
Arg Asn Gly Leu Arg Pro Ala Arg Tyr Val Ile Thr Lys Asp Lys Leu
355 360 365
Ile Thr Cys Ala Ser Glu Val Gly Ile Trp Asp Tyr Gln Pro Asp Glu
370 375 380
Val Val Glu Lys Gly Arg Val Gly Pro Gly Glu Leu Met Val Ile Asp
385 390 395 400
Thr Arg Ser Gly Arg Ile Leu His Ser Ala Glu Thr Asp Asp Asp Leu
405 410 415
Lys Ser Arg His Pro Tyr Lys Glu Trp Met Glu Lys Asn Val Arg Arg
420 425 430
Leu Val Pro Phe Glu Asp Leu Pro Asp Glu Glu Val Gly Ser Arg Glu
435 440 445
Leu Asp Asp Asp Thr Leu Ala Ser Tyr Gln Lys Gln Phe Asn Tyr Ser
450 455 460
Ala Glu Glu Leu Asp Ser Val Ile Arg Val Leu Gly Glu Asn Gly Gln
465 470 475 480
Glu Ala Val Gly Ser Met Gly Asp Asp Thr Pro Phe Ala Val Leu Ser
485 490 495
Ser Gln Pro Arg Ile Ile Tyr Asp Tyr Phe Arg Gln Gln Phe Ala Gln
500 505 510
Val Thr Asn Pro Pro Ile Asp Pro Leu Arg Glu Ala His Val Met Ser
515 520 525
Leu Ala Thr Ser Ile Gly Arg Glu Met Asn Val Phe Cys Glu Ala Glu
530 535 540
Gly Gln Ala His Arg Leu Ser Phe Lys Ser Pro Ile Leu Leu Tyr Ser
545 550 555 560
Asp Phe Lys Gln Leu Thr Thr Met Lys Glu Glu His Tyr Arg Ala Asp
565 570 575
Thr Leu Asp Ile Thr Phe Asp Val Thr Lys Thr Thr Leu Glu Ala Thr
580 585 590
Val Lys Glu Leu Cys Asp Lys Ala Glu Lys Met Val Arg Ser Gly Thr
595 600 605
Val Leu Leu Val Leu Ser Asp Arg Asn Ile Ala Lys Asp Arg Leu Pro
610 615 620
Val Pro Ala Pro Met Ala Val Gly Ala Ile Gln Thr Arg Leu Val Asp
625 630 635 640
Gln Ser Leu Arg Cys Asp Ala Asn Ile Ile Val Glu Thr Ala Ser Ala
645 650 655
Arg Asp Pro His His Phe Ala Val Leu Leu Gly Phe Gly Ala Thr Ala
660 665 670
Ile Tyr Pro Tyr Leu Ala Tyr Glu Thr Leu Gly Arg Leu Val Asp Thr
675 680 685
His Ala Ile Ala Lys Asp Tyr Arg Thr Val Met Leu Asn Tyr Arg Asn
690 695 700
Gly Ile Asn Lys Gly Leu Tyr Lys Ile Met Ser Lys Met Gly Ile Ser
705 710 715 720
Thr Ile Ala Ser Tyr Arg Cys Ser Lys Leu Phe Glu Ala Val Gly Leu
725 730 735
His Asp Asp Val Val Gly Leu Cys Phe Gln Gly Ala Val Ser Arg Ile
740 745 750
Gly Gly Ala Ser Phe Glu Asp Phe Gln Gln Asp Leu Leu Asn Leu Ser
755 760 765
Lys Arg Ala Trp Leu Ala Arg Lys Pro Ile Ser Gln Gly Gly Leu Leu
770 775 780
Lys Tyr Val His Gly Gly Glu Tyr His Ala Tyr Asn Pro Asp Val Val
785 790 795 800
Arg Thr Leu Gln Gln Ala Val Gln Ser Gly Glu Tyr Ser Asp Tyr Gln
805 810 815
Glu Tyr Ala Lys Leu Val Asn Glu Arg Pro Ala Thr Thr Leu Arg Asp
820 825 830
Leu Leu Ala Ile Thr Pro Gly Glu Asn Ala Val Asn Ile Ala Asp Val
835 840 845
Glu Pro Ala Ser Glu Leu Phe Lys Arg Phe Asp Thr Ala Ala Met Ser
850 855 860
Ile Gly Ala Leu Ser Pro Glu Ala His Glu Ala Leu Ala Glu Ala Met
865 870 875 880
Asn Ser Ile Gly Gly Asn Ser Asn Ser Gly Glu Gly Gly Glu Asp Pro
885 890 895
Ala Arg Tyr Gly Thr Asn Lys Val Ser Arg Ile Lys Gln Val Ala Ser
900 905 910
Gly Arg Phe Gly Val Thr Pro Ala Tyr Leu Val Asn Ala Asp Val Ile
915 920 925
Gln Ile Lys Val Ala Gln Gly Ala Lys Pro Gly Glu Gly Gly Gln Leu
930 935 940
Pro Gly Asp Lys Val Thr Pro Tyr Ile Ala Lys Leu Arg Tyr Ser Val
945 950 955 960
Pro Gly Val Thr Leu Ile Ser Pro Pro Pro His His Asp Ile Tyr Ser
965 970 975
Ile Glu Asp Leu Ala Gln Leu Ile Phe Asp Leu Lys Gln Val Asn Pro
980 985 990
Lys Ala Met Ile Ser Val Lys Leu Val Ser Glu Pro Gly Val Gly Thr
995 1000 1005
Ile Ala Thr Gly Val Ala Lys Ala Tyr Ala Asp Leu Ile Thr Ile
1010 1015 1020
Ala Gly Tyr Asp Gly Gly Thr Gly Ala Ser Pro Leu Ser Ser Val
1025 1030 1035
Lys Tyr Ala Gly Cys Pro Trp Glu Leu Gly Leu Val Glu Thr Gln
1040 1045 1050
Gln Ala Leu Val Ala Asn Gly Leu Arg His Lys Ile Arg Leu Gln
1055 1060 1065
Val Asp Gly Gly Leu Lys Thr Gly Val Asp Ile Ile Lys Ala Ala
1070 1075 1080
Ile Leu Gly Ala Glu Ser Phe Gly Phe Gly Thr Gly Pro Met Val
1085 1090 1095
Ala Leu Gly Cys Lys Tyr Leu Arg Ile Cys His Leu Asn Asn Cys
1100 1105 1110
Ala Thr Gly Val Ala Thr Gln Asp Asp Lys Leu Arg Lys Asn His
1115 1120 1125
Tyr His Gly Leu Pro Phe Lys Val Thr Asn Tyr Phe Glu Phe Ile
1130 1135 1140
Ala Arg Glu Thr Arg Glu Leu Met Ala Gln Leu Gly Val Thr Arg
1145 1150 1155
Leu Val Asp Leu Ile Gly Arg Thr Asp Leu Leu Lys Glu Leu Asp
1160 1165 1170
Gly Phe Thr Ala Lys Gln Gln Lys Leu Ala Leu Ser Lys Leu Leu
1175 1180 1185
Glu Thr Ala Glu Pro His Pro Gly Lys Ala Leu Tyr Cys Thr Glu
1190 1195 1200
Asn Asn Pro Pro Phe Asp Asn Gly Leu Leu Asn Ala Gln Leu Leu
1205 1210 1215
Gln Gln Ala Lys Pro Phe Val Asp Glu Arg Gln Ser Lys Thr Phe
1220 1225 1230
Trp Phe Asp Ile Arg Asn Thr Asp Arg Ser Val Gly Ala Ser Leu
1235 1240 1245
Ser Gly Tyr Ile Ala Gln Thr His Gly Asp Gln Gly Leu Ala Ala
1250 1255 1260
Asp Pro Ile Lys Ala Tyr Phe Asn Gly Thr Ala Gly Gln Ser Phe
1265 1270 1275
Gly Val Trp Asn Ala Gly Gly Val Glu Leu Tyr Leu Thr Gly Asp
1280 1285 1290
Ala Asn Asp Tyr Val Gly Lys Gly Met Ala Gly Gly Leu Ile Ala
1295 1300 1305
Ile Arg Pro Pro Val Gly Ser Ala Phe Arg Ser His Glu Ala Ser
1310 1315 1320
Ile Ile Gly Asn Thr Cys Leu Tyr Gly Ala Thr Gly Gly Arg Leu
1325 1330 1335
Tyr Ala Ala Gly Arg Ala Gly Glu Arg Phe Gly Val Arg Asn Ser
1340 1345 1350
Gly Ala Ile Thr Val Val Glu Gly Ile Gly Asp Asn Gly Cys Glu
1355 1360 1365
Tyr Met Thr Gly Gly Ile Val Cys Ile Leu Gly Lys Thr Gly Val
1370 1375 1380
Asn Phe Gly Ala Gly Met Thr Gly Gly Phe Ala Tyr Val Leu Asp
1385 1390 1395
Glu Ser Gly Asp Phe Arg Lys Arg Val Asn Pro Glu Leu Val Glu
1400 1405 1410
Val Leu Ser Val Asp Ala Leu Ala Ile His Glu Glu His Leu Arg
1415 1420 1425
Gly Leu Ile Thr Glu His Val Gln His Thr Gly Ser Gln Arg Gly
1430 1435 1440
Glu Glu Ile Leu Ala Asn Trp Ser Thr Phe Ala Thr Lys Phe Ala
1445 1450 1455
Leu Val Lys Pro Lys Ser Ser Asp Val Lys Ala Leu Leu Gly His
1460 1465 1470
Arg Ser Arg Ser Ala Ala Glu Leu Arg Val Gln Ala Gln
1475 1480 1485
<210> 82
<211> 1419
<212> DNA
<213> 大肠杆菌
<400> 82
atgagtcaga atgtttatca atttatcgac ctgcagcgcg ttgatccgcc aaagaaaccg 60
ctgaagatcc gcaaaattga gtttgttgaa atttacgagc cgttttccga aggccaggcc 120
aaagcgcagg ctgaccgctg cctgtcgtgc ggcaacccat actgcgagtg gaaatgcccg 180
gtacacaact acatcccgaa ctggctgaag ctcgccaacg aggggcgtat ttttgaagcg 240
gcggaactgt cgcaccagac caacaccctg ccggaagttt gcggacgagt ctgcccgcaa 300
gaccgtctgt gcgaaggttc ctgcactctg aacgatgagt ttggcgcggt gaccatcggc 360
aacattgagc gctatatcaa cgataaagcg ttcgagatgg gctggcgtcc ggatatgtct 420
ggtgtgaaac agaccggtaa aaaagtggcg attatcggcg caggcccggc aggtctggcg 480
tgtgcggatg tcctgacgcg taacggcgta aaagccgttg tcttcgaccg tcatccagaa 540
attggcgggc tgctgacctt cggtattccg gccttcaagc tggaaaaaga ggtaatgacg 600
cgtcgccgtg aaatcttcac cggcatgggt attgaattca aactcaatac cgaagtgggc 660
cgcgacgtac agctggacga tctgctgagt gattacgatg ccgtgttcct tggcgtcggg 720
acttatcagt caatgcgcgg cgggctggaa aacgaagacg ccgatggcgt gtacgcagcg 780
ctgccgttcc tcatcgccaa caccaaacag ttaatgggct ttggtgaaac ccgcgacgaa 840
ccgttcgtca gcatggaagg caaacgcgtg gtggtccttg gcggtggcga cactgcgatg 900
gactgcgtgc gtacgtccgt gcgccaggga gcgaagcacg ttacctgtgc ctatcgtcgt 960
gatgaagaga acatgccggg ttcccgccgc gaagtgaaaa acgcgcggga agaaggcgta 1020
gagttcaaat tcaacgtcca gccgctgggt attgaagtga acggtaacgg caaagtcagc 1080
ggcgtaaaaa tggtgcgtac cgaaatgggc gaaccggacg ccaaaggccg tcgccgcgcg 1140
gagatcgttg caggttccga acatatcgtt ccggcagatg cggtgatcat ggcgtttggt 1200
ttccgtccac acaacatgga atggctggca aaacacagcg tcgagctgga ttcacaaggc 1260
cgcatcatcg ccccggaagg cagcgacaac gccttccaga ccagcaaccc gaaaatcttt 1320
gctggcggcg atatcgtccg tggttccgat ctggtggtga ccgctattgc cgaaggtcgt 1380
aaggcggcag acggtattat gaactggctg gaagtttaa 1419
<210> 83
<211> 472
<212> PRT
<213> 大肠杆菌
<400> 83
Met Ser Gln Asn Val Tyr Gln Phe Ile Asp Leu Gln Arg Val Asp Pro
1 5 10 15
Pro Lys Lys Pro Leu Lys Ile Arg Lys Ile Glu Phe Val Glu Ile Tyr
20 25 30
Glu Pro Phe Ser Glu Gly Gln Ala Lys Ala Gln Ala Asp Arg Cys Leu
35 40 45
Ser Cys Gly Asn Pro Tyr Cys Glu Trp Lys Cys Pro Val His Asn Tyr
50 55 60
Ile Pro Asn Trp Leu Lys Leu Ala Asn Glu Gly Arg Ile Phe Glu Ala
65 70 75 80
Ala Glu Leu Ser His Gln Thr Asn Thr Leu Pro Glu Val Cys Gly Arg
85 90 95
Val Cys Pro Gln Asp Arg Leu Cys Glu Gly Ser Cys Thr Leu Asn Asp
100 105 110
Glu Phe Gly Ala Val Thr Ile Gly Asn Ile Glu Arg Tyr Ile Asn Asp
115 120 125
Lys Ala Phe Glu Met Gly Trp Arg Pro Asp Met Ser Gly Val Lys Gln
130 135 140
Thr Gly Lys Lys Val Ala Ile Ile Gly Ala Gly Pro Ala Gly Leu Ala
145 150 155 160
Cys Ala Asp Val Leu Thr Arg Asn Gly Val Lys Ala Val Val Phe Asp
165 170 175
Arg His Pro Glu Ile Gly Gly Leu Leu Thr Phe Gly Ile Pro Ala Phe
180 185 190
Lys Leu Glu Lys Glu Val Met Thr Arg Arg Arg Glu Ile Phe Thr Gly
195 200 205
Met Gly Ile Glu Phe Lys Leu Asn Thr Glu Val Gly Arg Asp Val Gln
210 215 220
Leu Asp Asp Leu Leu Ser Asp Tyr Asp Ala Val Phe Leu Gly Val Gly
225 230 235 240
Thr Tyr Gln Ser Met Arg Gly Gly Leu Glu Asn Glu Asp Ala Asp Gly
245 250 255
Val Tyr Ala Ala Leu Pro Phe Leu Ile Ala Asn Thr Lys Gln Leu Met
260 265 270
Gly Phe Gly Glu Thr Arg Asp Glu Pro Phe Val Ser Met Glu Gly Lys
275 280 285
Arg Val Val Val Leu Gly Gly Gly Asp Thr Ala Met Asp Cys Val Arg
290 295 300
Thr Ser Val Arg Gln Gly Ala Lys His Val Thr Cys Ala Tyr Arg Arg
305 310 315 320
Asp Glu Glu Asn Met Pro Gly Ser Arg Arg Glu Val Lys Asn Ala Arg
325 330 335
Glu Glu Gly Val Glu Phe Lys Phe Asn Val Gln Pro Leu Gly Ile Glu
340 345 350
Val Asn Gly Asn Gly Lys Val Ser Gly Val Lys Met Val Arg Thr Glu
355 360 365
Met Gly Glu Pro Asp Ala Lys Gly Arg Arg Arg Ala Glu Ile Val Ala
370 375 380
Gly Ser Glu His Ile Val Pro Ala Asp Ala Val Ile Met Ala Phe Gly
385 390 395 400
Phe Arg Pro His Asn Met Glu Trp Leu Ala Lys His Ser Val Glu Leu
405 410 415
Asp Ser Gln Gly Arg Ile Ile Ala Pro Glu Gly Ser Asp Asn Ala Phe
420 425 430
Gln Thr Ser Asn Pro Lys Ile Phe Ala Gly Gly Asp Ile Val Arg Gly
435 440 445
Ser Asp Leu Val Val Thr Ala Ile Ala Glu Gly Arg Lys Ala Ala Asp
450 455 460
Gly Ile Met Asn Trp Leu Glu Val
465 470
<210> 84
<211> 1665
<212> DNA
<213> 大肠杆菌
<400> 84
atgtgttcaa tttttggcgt attcgatatc aaaacagacg cagttgagct gcgtaagaaa 60
gccctcgagc tgtcacgcct gatgcgtcat cgtggcccgg actggtccgg tatttatgcc 120
agcgataacg ccattctcgc ccacgaacgt ctgtcaattg ttgacgttaa cgcgggggcg 180
caacctctct acaaccaaca aaaaacccac gtactggcgg taaacggtga aatctacaac 240
caccaggcat tgcgcgccga atatggcgat cgttaccagt tccagaccgg gtctgactgt 300
gaagtgatcc tcgcgctgta tcaggaaaaa gggccggaat ttctcgacga cttgcagggc 360
atgtttgcct ttgcactgta cgacagcgaa aaagatgcct acctgattgg tcgcgaccat 420
ctggggatca tcccactgta tatggggtat gacgaacacg gtcagctgta tgtggcctca 480
gaaatgaaag cgctggtgcc agtttgccgc acgattaaag agttcccggc ggggagctat 540
ttgtggagcc aggacggcga aatccgttct tactatcatc gcgactggtt cgactacgat 600
gcggtgaaag ataacgtgac cgacaaaaac gagctgcgtc aggcactgga agattcagtt 660
aaaagccatc tgatgtctga tgtgccttac ggtgtgctgc tttctggtgg tctggattcc 720
tcaattattt ccgctatcac caagaaatac gcagcccgtc gcgtggaaga tcaggaacgc 780
tctgaagcct ggtggccgca gttacactcc tttgctgtag gtctgccggg ttcaccggat 840
ctgaaagcag cccaggaagt ggcaaaccat ctgggcacgg tgcatcacga aattcacttc 900
actgtacagg aaggtctgga tgccatccgc gacgtgattt accacatcga aacttatgat 960
gtgaccacta ttcgcgcttc aacaccgatg tatttaatgt cgcgtaagat caaggcgatg 1020
ggcattaaaa tggtgctgtc cggtgaaggt tctgatgaag tgttcggcgg ttatctttac 1080
ttccacaaag caccgaatgc caaagaactg catgaagaga cggtgcgtaa actgctggcc 1140
ctgcatatgt atgactgcgc gcgtgccaac aaagcgatgt cagcctgggg cgtggaagca 1200
cgcgttccgt tcctcgacaa aaaattcctt gatgtggcga tgcgtattaa cccacaggat 1260
aaaatgtgcg gtaacggcaa aatggaaaaa cacatcctgc gtgaatgttt tgaagcgtat 1320
ctgcctgcaa gcgtggcctg gcggcagaaa gagcagttct ccgatggcgt cggttacagt 1380
tggatcgaca ccctgaaaga agtggctgcg cagcaggttt ctgatcagca actggaaact 1440
gcccgcttcc gcttcccgta caacacgcca acctctaaag aagcgtactt gtatcgggag 1500
atctttgaag aactattccc gcttccgagc gccgctgagt gcgtgccggg cggtccttcc 1560
gtcgcttgtt cttccgctaa agcgatcgaa tgggatgaag cgttcaagaa aatggacgat 1620
ccgtctggtc gcgcggttgg tgttcaccag tcggcgtata agtaa 1665
<210> 85
<211> 554
<212> PRT
<213> 大肠杆菌
<400> 85
Met Cys Ser Ile Phe Gly Val Phe Asp Ile Lys Thr Asp Ala Val Glu
1 5 10 15
Leu Arg Lys Lys Ala Leu Glu Leu Ser Arg Leu Met Arg His Arg Gly
20 25 30
Pro Asp Trp Ser Gly Ile Tyr Ala Ser Asp Asn Ala Ile Leu Ala His
35 40 45
Glu Arg Leu Ser Ile Val Asp Val Asn Ala Gly Ala Gln Pro Leu Tyr
50 55 60
Asn Gln Gln Lys Thr His Val Leu Ala Val Asn Gly Glu Ile Tyr Asn
65 70 75 80
His Gln Ala Leu Arg Ala Glu Tyr Gly Asp Arg Tyr Gln Phe Gln Thr
85 90 95
Gly Ser Asp Cys Glu Val Ile Leu Ala Leu Tyr Gln Glu Lys Gly Pro
100 105 110
Glu Phe Leu Asp Asp Leu Gln Gly Met Phe Ala Phe Ala Leu Tyr Asp
115 120 125
Ser Glu Lys Asp Ala Tyr Leu Ile Gly Arg Asp His Leu Gly Ile Ile
130 135 140
Pro Leu Tyr Met Gly Tyr Asp Glu His Gly Gln Leu Tyr Val Ala Ser
145 150 155 160
Glu Met Lys Ala Leu Val Pro Val Cys Arg Thr Ile Lys Glu Phe Pro
165 170 175
Ala Gly Ser Tyr Leu Trp Ser Gln Asp Gly Glu Ile Arg Ser Tyr Tyr
180 185 190
His Arg Asp Trp Phe Asp Tyr Asp Ala Val Lys Asp Asn Val Thr Asp
195 200 205
Lys Asn Glu Leu Arg Gln Ala Leu Glu Asp Ser Val Lys Ser His Leu
210 215 220
Met Ser Asp Val Pro Tyr Gly Val Leu Leu Ser Gly Gly Leu Asp Ser
225 230 235 240
Ser Ile Ile Ser Ala Ile Thr Lys Lys Tyr Ala Ala Arg Arg Val Glu
245 250 255
Asp Gln Glu Arg Ser Glu Ala Trp Trp Pro Gln Leu His Ser Phe Ala
260 265 270
Val Gly Leu Pro Gly Ser Pro Asp Leu Lys Ala Ala Gln Glu Val Ala
275 280 285
Asn His Leu Gly Thr Val His His Glu Ile His Phe Thr Val Gln Glu
290 295 300
Gly Leu Asp Ala Ile Arg Asp Val Ile Tyr His Ile Glu Thr Tyr Asp
305 310 315 320
Val Thr Thr Ile Arg Ala Ser Thr Pro Met Tyr Leu Met Ser Arg Lys
325 330 335
Ile Lys Ala Met Gly Ile Lys Met Val Leu Ser Gly Glu Gly Ser Asp
340 345 350
Glu Val Phe Gly Gly Tyr Leu Tyr Phe His Lys Ala Pro Asn Ala Lys
355 360 365
Glu Leu His Glu Glu Thr Val Arg Lys Leu Leu Ala Leu His Met Tyr
370 375 380
Asp Cys Ala Arg Ala Asn Lys Ala Met Ser Ala Trp Gly Val Glu Ala
385 390 395 400
Arg Val Pro Phe Leu Asp Lys Lys Phe Leu Asp Val Ala Met Arg Ile
405 410 415
Asn Pro Gln Asp Lys Met Cys Gly Asn Gly Lys Met Glu Lys His Ile
420 425 430
Leu Arg Glu Cys Phe Glu Ala Tyr Leu Pro Ala Ser Val Ala Trp Arg
435 440 445
Gln Lys Glu Gln Phe Ser Asp Gly Val Gly Tyr Ser Trp Ile Asp Thr
450 455 460
Leu Lys Glu Val Ala Ala Gln Gln Val Ser Asp Gln Gln Leu Glu Thr
465 470 475 480
Ala Arg Phe Arg Phe Pro Tyr Asn Thr Pro Thr Ser Lys Glu Ala Tyr
485 490 495
Leu Tyr Arg Glu Ile Phe Glu Glu Leu Phe Pro Leu Pro Ser Ala Ala
500 505 510
Glu Cys Val Pro Gly Gly Pro Ser Val Ala Cys Ser Ser Ala Lys Ala
515 520 525
Ile Glu Trp Asp Glu Ala Phe Lys Lys Met Asp Asp Pro Ser Gly Arg
530 535 540
Ala Val Gly Val His Gln Ser Ala Tyr Lys
545 550
<210> 86
<211> 933
<212> DNA
<213> 大肠杆菌
<400> 86
atgttagatg caaacaaatt acagcaggca gtggatcagg cttacaccca atttcactca 60
cttaacggcg gacaaaatgc cgattacatt ccctttctgg cgaatgtacc aggtcaactg 120
gcggcagtgg ctatcgtgac ctgcgatggc aacgtctata gtgcgggtga cagtgattac 180
cgctttgcac tggaatccat ctcgaaagtc tgtacgttag cccttgcgtt agaagatgtc 240
ggcccgcagg cggtacagga caaaattggc gctgacccga ccggattgcc ctttaactca 300
gttatcgcct tagagttgca tggcggcaaa ccgctttcgc cactggtaaa tgctggcgct 360
attgccacca ccagcctgat taacgctgaa aatgttgaac aacgctggca gcgaatttta 420
catatccaac agcaactggc tggcgagcag gtagcgctct ctgacgaagt caaccagtcg 480
gaacaaacaa ccaacttcca taaccgggcc atagcctggc tgctgtactc cgccggatat 540
ctctattgtg atgcaatgga agcctgtgac gtgtataccc gtcagtgctc cacgctcctc 600
aatactattg aactggcaac gcttggcgcg acgctggcgg caggtggtgt gaatccgttg 660
acgcataaac gcgttcttca ggccgacaac gtgccgtaca ttctggccga aatgatgatg 720
gaagggctgt atggtcgctc cggtgactgg gcgtatcgtg ttggtttacc gggcaaaagc 780
ggtgtaggtg gcggtattct ggcggtcgtc cctggagtga tgggaattgc cgcgttctca 840
ccaccgctgg acgaagatgg caacagtgtt cgcggtcaaa aaatggtggc atcggtcgct 900
aagcaactcg gctataacgt gtttaagggc tga 933
<210> 87
<211> 310
<212> PRT
<213> 大肠杆菌
<400> 87
Met Leu Asp Ala Asn Lys Leu Gln Gln Ala Val Asp Gln Ala Tyr Thr
1 5 10 15
Gln Phe His Ser Leu Asn Gly Gly Gln Asn Ala Asp Tyr Ile Pro Phe
20 25 30
Leu Ala Asn Val Pro Gly Gln Leu Ala Ala Val Ala Ile Val Thr Cys
35 40 45
Asp Gly Asn Val Tyr Ser Ala Gly Asp Ser Asp Tyr Arg Phe Ala Leu
50 55 60
Glu Ser Ile Ser Lys Val Cys Thr Leu Ala Leu Ala Leu Glu Asp Val
65 70 75 80
Gly Pro Gln Ala Val Gln Asp Lys Ile Gly Ala Asp Pro Thr Gly Leu
85 90 95
Pro Phe Asn Ser Val Ile Ala Leu Glu Leu His Gly Gly Lys Pro Leu
100 105 110
Ser Pro Leu Val Asn Ala Gly Ala Ile Ala Thr Thr Ser Leu Ile Asn
115 120 125
Ala Glu Asn Val Glu Gln Arg Trp Gln Arg Ile Leu His Ile Gln Gln
130 135 140
Gln Leu Ala Gly Glu Gln Val Ala Leu Ser Asp Glu Val Asn Gln Ser
145 150 155 160
Glu Gln Thr Thr Asn Phe His Asn Arg Ala Ile Ala Trp Leu Leu Tyr
165 170 175
Ser Ala Gly Tyr Leu Tyr Cys Asp Ala Met Glu Ala Cys Asp Val Tyr
180 185 190
Thr Arg Gln Cys Ser Thr Leu Leu Asn Thr Ile Glu Leu Ala Thr Leu
195 200 205
Gly Ala Thr Leu Ala Ala Gly Gly Val Asn Pro Leu Thr His Lys Arg
210 215 220
Val Leu Gln Ala Asp Asn Val Pro Tyr Ile Leu Ala Glu Met Met Met
225 230 235 240
Glu Gly Leu Tyr Gly Arg Ser Gly Asp Trp Ala Tyr Arg Val Gly Leu
245 250 255
Pro Gly Lys Ser Gly Val Gly Gly Gly Ile Leu Ala Val Val Pro Gly
260 265 270
Val Met Gly Ile Ala Ala Phe Ser Pro Pro Leu Asp Glu Asp Gly Asn
275 280 285
Ser Val Arg Gly Gln Lys Met Val Ala Ser Val Ala Lys Gln Leu Gly
290 295 300
Tyr Asn Val Phe Lys Gly
305 310
<210> 88
<211> 927
<212> DNA
<213> 大肠杆菌
<400> 88
atggcagtcg ccatggataa tgcaatttta gaaaacatct tgcggcaagt gcggccgctc 60
attggtcagg gtaaagtcgc ggattatatt ccggcgctgg ctacagtaga cggttcccga 120
ttggggattg ctatctgtac cgttgacgga cagctttttc aggccggaga cgcgcaagaa 180
cgtttttcca ttcagtctat ttccaaagtg ctgagtctcg ttgtcgccat gcgtcattac 240
tccgaagagg aaatctggca acgcgtcggc aaagatccgt ctggatcacc gttcaattcc 300
ttagtgcaac tggaaatgga gcagggtata ccgcgtaatc cgttcattaa tgccggtgcg 360
ctggtggtct gcgatatgtt gcaagggcga ttaagcgcac cacggcaacg tatgctggaa 420
gtcgtgcgcg gcttaagcgg tgtgtctgat atttcctacg atacggtggt agcgcgttcc 480
gaatttgaac attccgcgcg aaatgcggct atcgcctggc tgatgaagtc gtttggcaat 540
ttccatcatg acgtgacaac cgttctgcaa aactactttc attactgcgc tctgaaaatg 600
agctgtgtag agctggcccg gacgtttgtc tttctggcta atcaggggaa agctattcat 660
attgatgaac cagtggtgac gccaatgcag gcgcggcaaa ttaacgcgct gatggcgacc 720
agtggtatgt accagaacgc gggggagttt gcctggcggg tggggctacc ggcgaaatct 780
ggcgttggtg gcggtattgt ggcgattgtt ccgcatgaaa tggccatcgc tgtctggagt 840
ccggaactgg atgatgcagg taactcgctt gcgggtattg ccgttcttga acaattgacg 900
aaacagttag ggcgttcggt ttattaa 927
<210> 89
<211> 927
<212> PRT
<213> 大肠杆菌
<400> 89
Gly Thr Gly Gly Cys Ala Gly Thr Cys Gly Cys Cys Ala Thr Gly Gly
1 5 10 15
Ala Thr Ala Ala Thr Gly Cys Ala Ala Thr Thr Thr Thr Ala Gly Ala
20 25 30
Ala Ala Ala Cys Ala Thr Cys Thr Thr Gly Cys Gly Gly Cys Ala Ala
35 40 45
Gly Thr Gly Cys Gly Gly Cys Cys Gly Cys Thr Cys Ala Thr Thr Gly
50 55 60
Gly Thr Cys Ala Gly Gly Gly Thr Ala Ala Ala Gly Thr Cys Gly Cys
65 70 75 80
Gly Gly Ala Thr Thr Ala Thr Ala Thr Thr Cys Cys Gly Gly Cys Gly
85 90 95
Cys Thr Gly Gly Cys Thr Ala Cys Ala Gly Thr Ala Gly Ala Cys Gly
100 105 110
Gly Thr Thr Cys Cys Cys Gly Ala Thr Thr Gly Gly Gly Gly Ala Thr
115 120 125
Thr Gly Cys Thr Ala Thr Cys Thr Gly Thr Ala Cys Cys Gly Thr Thr
130 135 140
Gly Ala Cys Gly Gly Ala Cys Ala Gly Cys Thr Thr Thr Thr Thr Cys
145 150 155 160
Ala Gly Gly Cys Cys Gly Gly Ala Gly Ala Cys Gly Cys Gly Cys Ala
165 170 175
Ala Gly Ala Ala Cys Gly Thr Thr Thr Thr Thr Cys Cys Ala Thr Thr
180 185 190
Cys Ala Gly Thr Cys Thr Ala Thr Thr Thr Cys Cys Ala Ala Ala Gly
195 200 205
Thr Gly Cys Thr Gly Ala Gly Thr Cys Thr Cys Gly Thr Thr Gly Thr
210 215 220
Cys Gly Cys Cys Ala Thr Gly Cys Gly Thr Cys Ala Thr Thr Ala Cys
225 230 235 240
Thr Cys Cys Gly Ala Ala Gly Ala Gly Gly Ala Ala Ala Thr Cys Thr
245 250 255
Gly Gly Cys Ala Ala Cys Gly Cys Gly Thr Cys Gly Gly Cys Ala Ala
260 265 270
Ala Gly Ala Thr Cys Cys Gly Thr Cys Thr Gly Gly Ala Thr Cys Ala
275 280 285
Cys Cys Gly Thr Thr Cys Ala Ala Thr Thr Cys Cys Thr Thr Ala Gly
290 295 300
Thr Gly Cys Ala Ala Cys Thr Gly Gly Ala Ala Ala Thr Gly Gly Ala
305 310 315 320
Gly Cys Ala Gly Gly Gly Thr Ala Thr Ala Cys Cys Gly Cys Gly Thr
325 330 335
Ala Ala Thr Cys Cys Gly Thr Thr Cys Ala Thr Thr Ala Ala Thr Gly
340 345 350
Cys Cys Gly Gly Thr Gly Cys Gly Cys Thr Gly Gly Thr Gly Gly Thr
355 360 365
Cys Thr Gly Cys Gly Ala Thr Ala Thr Gly Thr Thr Gly Cys Ala Ala
370 375 380
Gly Gly Gly Cys Gly Ala Thr Thr Ala Ala Gly Cys Gly Cys Ala Cys
385 390 395 400
Cys Ala Cys Gly Gly Cys Ala Ala Cys Gly Thr Ala Thr Gly Cys Thr
405 410 415
Gly Gly Ala Ala Gly Thr Cys Gly Thr Gly Cys Gly Cys Gly Gly Cys
420 425 430
Thr Thr Ala Ala Gly Cys Gly Gly Thr Gly Thr Gly Thr Cys Thr Gly
435 440 445
Ala Thr Ala Thr Thr Thr Cys Cys Thr Ala Cys Gly Ala Thr Ala Cys
450 455 460
Gly Gly Thr Gly Gly Thr Ala Gly Cys Gly Cys Gly Thr Thr Cys Cys
465 470 475 480
Gly Ala Ala Thr Thr Thr Gly Ala Ala Cys Ala Thr Thr Cys Cys Gly
485 490 495
Cys Gly Cys Gly Ala Ala Ala Thr Gly Cys Gly Gly Cys Thr Ala Thr
500 505 510
Cys Gly Cys Cys Thr Gly Gly Cys Thr Gly Ala Thr Gly Ala Ala Gly
515 520 525
Thr Cys Gly Thr Thr Thr Gly Gly Cys Ala Ala Thr Thr Thr Cys Cys
530 535 540
Ala Thr Cys Ala Thr Gly Ala Cys Gly Thr Gly Ala Cys Ala Ala Cys
545 550 555 560
Cys Gly Thr Thr Cys Thr Gly Cys Ala Ala Ala Ala Cys Thr Ala Cys
565 570 575
Thr Thr Thr Cys Ala Thr Thr Ala Cys Thr Gly Cys Gly Cys Thr Cys
580 585 590
Thr Gly Ala Ala Ala Ala Thr Gly Ala Gly Cys Thr Gly Thr Gly Thr
595 600 605
Ala Gly Ala Gly Cys Thr Gly Gly Cys Cys Cys Gly Gly Ala Cys Gly
610 615 620
Thr Thr Thr Gly Thr Cys Thr Thr Thr Cys Thr Gly Gly Cys Thr Ala
625 630 635 640
Ala Thr Cys Ala Gly Gly Gly Gly Ala Ala Ala Gly Cys Thr Ala Thr
645 650 655
Thr Cys Ala Thr Ala Thr Thr Gly Ala Thr Gly Ala Ala Cys Cys Ala
660 665 670
Gly Thr Gly Gly Thr Gly Ala Cys Gly Cys Cys Ala Ala Thr Gly Cys
675 680 685
Ala Gly Gly Cys Gly Cys Gly Gly Cys Ala Ala Ala Thr Thr Ala Ala
690 695 700
Cys Gly Cys Gly Cys Thr Gly Ala Thr Gly Gly Cys Gly Ala Cys Cys
705 710 715 720
Ala Gly Thr Gly Gly Thr Ala Thr Gly Thr Ala Cys Cys Ala Gly Ala
725 730 735
Ala Cys Gly Cys Gly Gly Gly Gly Gly Ala Gly Thr Thr Thr Gly Cys
740 745 750
Cys Thr Gly Gly Cys Gly Gly Gly Thr Gly Gly Gly Gly Cys Thr Ala
755 760 765
Cys Cys Gly Gly Cys Gly Ala Ala Ala Thr Cys Thr Gly Gly Cys Gly
770 775 780
Thr Thr Gly Gly Thr Gly Gly Cys Gly Gly Thr Ala Thr Thr Gly Thr
785 790 795 800
Gly Gly Cys Gly Ala Thr Thr Gly Thr Thr Cys Cys Gly Cys Ala Thr
805 810 815
Gly Ala Ala Ala Thr Gly Gly Cys Cys Ala Thr Cys Gly Cys Thr Gly
820 825 830
Thr Cys Thr Gly Gly Ala Gly Thr Cys Cys Gly Gly Ala Ala Cys Thr
835 840 845
Gly Gly Ala Thr Gly Ala Thr Gly Cys Ala Gly Gly Thr Ala Ala Cys
850 855 860
Thr Cys Gly Cys Thr Thr Gly Cys Gly Gly Gly Thr Ala Thr Thr Gly
865 870 875 880
Cys Cys Gly Thr Thr Cys Thr Thr Gly Ala Ala Cys Ala Ala Thr Thr
885 890 895
Gly Ala Cys Gly Ala Ala Ala Cys Ala Gly Thr Thr Ala Gly Gly Gly
900 905 910
Cys Gly Thr Thr Cys Gly Gly Thr Thr Thr Ala Thr Thr Ala Ala
915 920 925
<210> 90
<211> 1344
<212> DNA
<213> 大肠杆菌
<400> 90
atggatcaga catattctct ggagtcattc ctcaaccatg tccaaaagcg cgacccgaat 60
caaaccgagt tcgcgcaagc cgttcgtgaa gtaatgacca cactctggcc ttttcttgaa 120
caaaatccaa aatatcgcca gatgtcatta ctggagcgtc tggttgaacc ggagcgcgtg 180
atccagtttc gcgtggtatg ggttgatgat cgcaaccaga tacaggtcaa ccgtgcatgg 240
cgtgtgcagt tcagctctgc catcggcccg tacaaaggcg gtatgcgctt ccatccgtca 300
gttaaccttt ccattctcaa attcctcggc tttgaacaaa ccttcaaaaa tgccctgact 360
actctgccga tgggcggtgg taaaggcggc agcgatttcg atccgaaagg aaaaagcgaa 420
ggtgaagtga tgcgtttttg ccaggcgctg atgactgaac tgtatcgcca cctgggcgcg 480
gataccgacg ttccggcagg tgatatcggg gttggtggtc gtgaagtcgg ctttatggcg 540
gggatgatga aaaagctctc caacaatacc gcctgcgtct tcaccggtaa gggcctttca 600
tttggcggca gtcttattcg cccggaagct accggctacg gtctggttta tttcacagaa 660
gcaatgctaa aacgccacgg tatgggtttt gaagggatgc gcgtttccgt ttctggctcc 720
ggcaacgtcg cccagtacgc tatcgaaaaa gcgatggaat ttggtgctcg tgtgatcact 780
gcgtcagact ccagcggcac tgtagttgat gaaagcggat tcacgaaaga gaaactggca 840
cgtcttatcg aaatcaaagc cagccgcgat ggtcgagtgg cagattacgc caaagaattt 900
ggtctggtct atctcgaagg ccaacagccg tggtctctac cggttgatat cgccctgcct 960
tgcgccaccc agaatgaact ggatgttgac gccgcgcatc agcttatcgc taatggcgtt 1020
aaagccgtcg ccgaaggggc aaatatgccg accaccatcg aagcgactga actgttccag 1080
caggcaggcg tactatttgc accgggtaaa gcggctaatg ctggtggcgt cgctacatcg 1140
ggcctggaaa tggcacaaaa cgctgcgcgc ctgggctgga aagccgagaa agttgacgca 1200
cgtttgcatc acatcatgct ggatatccac catgcctgtg ttgagcatgg tggtgaaggt 1260
gagcaaacca actacgtgca gggcgcgaac attgccggtt ttgtgaaggt tgccgatgcg 1320
atgctggcgc agggtgtgat ttaa 1344
<210> 91
<211> 447
<212> PRT
<213> 大肠杆菌
<400> 91
Met Asp Gln Thr Tyr Ser Leu Glu Ser Phe Leu Asn His Val Gln Lys
1 5 10 15
Arg Asp Pro Asn Gln Thr Glu Phe Ala Gln Ala Val Arg Glu Val Met
20 25 30
Thr Thr Leu Trp Pro Phe Leu Glu Gln Asn Pro Lys Tyr Arg Gln Met
35 40 45
Ser Leu Leu Glu Arg Leu Val Glu Pro Glu Arg Val Ile Gln Phe Arg
50 55 60
Val Val Trp Val Asp Asp Arg Asn Gln Ile Gln Val Asn Arg Ala Trp
65 70 75 80
Arg Val Gln Phe Ser Ser Ala Ile Gly Pro Tyr Lys Gly Gly Met Arg
85 90 95
Phe His Pro Ser Val Asn Leu Ser Ile Leu Lys Phe Leu Gly Phe Glu
100 105 110
Gln Thr Phe Lys Asn Ala Leu Thr Thr Leu Pro Met Gly Gly Gly Lys
115 120 125
Gly Gly Ser Asp Phe Asp Pro Lys Gly Lys Ser Glu Gly Glu Val Met
130 135 140
Arg Phe Cys Gln Ala Leu Met Thr Glu Leu Tyr Arg His Leu Gly Ala
145 150 155 160
Asp Thr Asp Val Pro Ala Gly Asp Ile Gly Val Gly Gly Arg Glu Val
165 170 175
Gly Phe Met Ala Gly Met Met Lys Lys Leu Ser Asn Asn Thr Ala Cys
180 185 190
Val Phe Thr Gly Lys Gly Leu Ser Phe Gly Gly Ser Leu Ile Arg Pro
195 200 205
Glu Ala Thr Gly Tyr Gly Leu Val Tyr Phe Thr Glu Ala Met Leu Lys
210 215 220
Arg His Gly Met Gly Phe Glu Gly Met Arg Val Ser Val Ser Gly Ser
225 230 235 240
Gly Asn Val Ala Gln Tyr Ala Ile Glu Lys Ala Met Glu Phe Gly Ala
245 250 255
Arg Val Ile Thr Ala Ser Asp Ser Ser Gly Thr Val Val Asp Glu Ser
260 265 270
Gly Phe Thr Lys Glu Lys Leu Ala Arg Leu Ile Glu Ile Lys Ala Ser
275 280 285
Arg Asp Gly Arg Val Ala Asp Tyr Ala Lys Glu Phe Gly Leu Val Tyr
290 295 300
Leu Glu Gly Gln Gln Pro Trp Ser Leu Pro Val Asp Ile Ala Leu Pro
305 310 315 320
Cys Ala Thr Gln Asn Glu Leu Asp Val Asp Ala Ala His Gln Leu Ile
325 330 335
Ala Asn Gly Val Lys Ala Val Ala Glu Gly Ala Asn Met Pro Thr Thr
340 345 350
Ile Glu Ala Thr Glu Leu Phe Gln Gln Ala Gly Val Leu Phe Ala Pro
355 360 365
Gly Lys Ala Ala Asn Ala Gly Gly Val Ala Thr Ser Gly Leu Glu Met
370 375 380
Ala Gln Asn Ala Ala Arg Leu Gly Trp Lys Ala Glu Lys Val Asp Ala
385 390 395 400
Arg Leu His His Ile Met Leu Asp Ile His His Ala Cys Val Glu His
405 410 415
Gly Gly Glu Gly Glu Gln Thr Asn Tyr Val Gln Gly Ala Asn Ile Ala
420 425 430
Gly Phe Val Lys Val Ala Asp Ala Met Leu Ala Gln Gly Val Ile
435 440 445
<210> 92
<211> 1479
<212> DNA
<213> 大肠杆菌
<400> 92
atgagcttac gtgaaaaaac catcagcggc gcgaagtggt cggcgattgc cacggtgatc 60
atcatcggcc tcgggctggt gcagatgacc gtgctggcgc ggattatcga caaccaccag 120
ttcggcctgc ttaccgtgtc gctggtgatt atcgcgctgg cagatacgct ttctgacttc 180
ggtatcgcta actcgattat tcagcgaaaa gaaatcagtc accttgaact caccacgttg 240
tactggctga acgtcgggct ggggatcgtg gtgtgcgtgg cggtgttttt gttgagtgat 300
ctcatcggcg acgtgctgaa taacccggac ctggcaccgt tgattaaaac attatcgctg 360
gcgtttgtgg taatccccca cgggcaacag ttccgcgcgt tgatgcaaaa agagctggag 420
ttcaacaaaa tcggcatgat cgaaaccagc gcggtgctgg cgggcttcac ttgtacggtg 480
gttagcgccc atttctggcc gctggcgatg accgcgatcc tcggttatct ggtcaatagt 540
gcggtgagaa cgctgctgtt tggctacttt ggccgcaaaa tttatcgccc cggtctgcat 600
ttctcgctgg cgtcggtggc accgaactta cgctttggtg cctggctgac ggcggacagc 660
atcatcaact atctcaatac caacctttca acgctcgtgc tggcgcgtat tctcggcgcg 720
ggcgtggcag ggggatacaa cctggcgtac aacgtggccg ttgtgccacc gatgaagctg 780
aacccaatca tcacccgcgt gttgtttccg gcattcgcca aaattcagga cgataccgaa 840
aagctgcgtg ttaacttcta caagctgctg tcggtagtgg ggattatcaa ctttccggcg 900
ctgctcgggc taatggtggt gtcgaataac tttgtaccgc tggtctttgg tgagaagtgg 960
aacagcatta ttccggtgct gcaattgctg tgtgtggtgg gtctgctgcg ctccgtaggt 1020
aacccgattg gttcgctgct gatggcgaaa gcgcgggtcg atatcagctt taaattcaac 1080
gtattcaaaa catttctgtt tattccggcg attgttatag gtgggcagat ggcgggcgcg 1140
atcggcgtca cgcttggctt cctgctggtg caaattatca acaccattct gagttacttc 1200
gtgatgatta aaccggttct tggttccagt tatcgccagt acatcctgag tttatggctg 1260
ccgttttatc tctcgctgcc gacgctggtg gtcagttatg cgctgggcat tgtgctgaaa 1320
gggcaactgg cgctggggat gctgctggcg gtgcaaatag ccacgggggt gctggcgttt 1380
gtggtgatga ttgtgctgtc gcgccatccg ctggtggtgg aagtgaagcg tcagttttgt 1440
cgcagcgaaa aaatgaaaat gcttttacgg gcggggtga 1479
<210> 93
<211> 492
<212> PRT
<213> 大肠杆菌
<400> 93
Met Ser Leu Arg Glu Lys Thr Ile Ser Gly Ala Lys Trp Ser Ala Ile
1 5 10 15
Ala Thr Val Ile Ile Ile Gly Leu Gly Leu Val Gln Met Thr Val Leu
20 25 30
Ala Arg Ile Ile Asp Asn His Gln Phe Gly Leu Leu Thr Val Ser Leu
35 40 45
Val Ile Ile Ala Leu Ala Asp Thr Leu Ser Asp Phe Gly Ile Ala Asn
50 55 60
Ser Ile Ile Gln Arg Lys Glu Ile Ser His Leu Glu Leu Thr Thr Leu
65 70 75 80
Tyr Trp Leu Asn Val Gly Leu Gly Ile Val Val Cys Val Ala Val Phe
85 90 95
Leu Leu Ser Asp Leu Ile Gly Asp Val Leu Asn Asn Pro Asp Leu Ala
100 105 110
Pro Leu Ile Lys Thr Leu Ser Leu Ala Phe Val Val Ile Pro His Gly
115 120 125
Gln Gln Phe Arg Ala Leu Met Gln Lys Glu Leu Glu Phe Asn Lys Ile
130 135 140
Gly Met Ile Glu Thr Ser Ala Val Leu Ala Gly Phe Thr Cys Thr Val
145 150 155 160
Val Ser Ala His Phe Trp Pro Leu Ala Met Thr Ala Ile Leu Gly Tyr
165 170 175
Leu Val Asn Ser Ala Val Arg Thr Leu Leu Phe Gly Tyr Phe Gly Arg
180 185 190
Lys Ile Tyr Arg Pro Gly Leu His Phe Ser Leu Ala Ser Val Ala Pro
195 200 205
Asn Leu Arg Phe Gly Ala Trp Leu Thr Ala Asp Ser Ile Ile Asn Tyr
210 215 220
Leu Asn Thr Asn Leu Ser Thr Leu Val Leu Ala Arg Ile Leu Gly Ala
225 230 235 240
Gly Val Ala Gly Gly Tyr Asn Leu Ala Tyr Asn Val Ala Val Val Pro
245 250 255
Pro Met Lys Leu Asn Pro Ile Ile Thr Arg Val Leu Phe Pro Ala Phe
260 265 270
Ala Lys Ile Gln Asp Asp Thr Glu Lys Leu Arg Val Asn Phe Tyr Lys
275 280 285
Leu Leu Ser Val Val Gly Ile Ile Asn Phe Pro Ala Leu Leu Gly Leu
290 295 300
Met Val Val Ser Asn Asn Phe Val Pro Leu Val Phe Gly Glu Lys Trp
305 310 315 320
Asn Ser Ile Ile Pro Val Leu Gln Leu Leu Cys Val Val Gly Leu Leu
325 330 335
Arg Ser Val Gly Asn Pro Ile Gly Ser Leu Leu Met Ala Lys Ala Arg
340 345 350
Val Asp Ile Ser Phe Lys Phe Asn Val Phe Lys Thr Phe Leu Phe Ile
355 360 365
Pro Ala Ile Val Ile Gly Gly Gln Met Ala Gly Ala Ile Gly Val Thr
370 375 380
Leu Gly Phe Leu Leu Val Gln Ile Ile Asn Thr Ile Leu Ser Tyr Phe
385 390 395 400
Val Met Ile Lys Pro Val Leu Gly Ser Ser Tyr Arg Gln Tyr Ile Leu
405 410 415
Ser Leu Trp Leu Pro Phe Tyr Leu Ser Leu Pro Thr Leu Val Val Ser
420 425 430
Tyr Ala Leu Gly Ile Val Leu Lys Gly Gln Leu Ala Leu Gly Met Leu
435 440 445
Leu Ala Val Gln Ile Ala Thr Gly Val Leu Ala Phe Val Val Met Ile
450 455 460
Val Leu Ser Arg His Pro Leu Val Val Glu Val Lys Arg Gln Phe Cys
465 470 475 480
Arg Ser Glu Lys Met Lys Met Leu Leu Arg Ala Gly
485 490
<210> 94
<211> 1395
<212> DNA
<213> 大肠杆菌
<400> 94
atgacaaatc taaaaaagcg cgagcgagcg aaaaccaatg catcgttaat ctctatggtg 60
caacgctttt cagatatcac catcatgttt gccggactat ggctggtttg cgaagtcagc 120
ggactgtcat tcctctacat gcacctgttg gtggcgctga ttacgctggt ggtgttccag 180
atgctgggcg gcatcaccga tttttatcgc tcatggcgcg gtgttcgggc agcgacagaa 240
tttgccctgt tgctacaaaa ctggacctta agcgtgattt tcagcgccgg actggtggcg 300
ttcaacaatg atttcgacac gcaactgaaa atctggctgg cgtggtatgc gctgaccagc 360
atcggactgg tggtttgccg ttcgtgtatt cgcattgggg cgggctggct gcgtaatcat 420
ggctataaca agcgcatggt cgcggtggcg ggggatttag ccgccgggca aatgctgatg 480
gagagcttcc gtaaccagcc gtggttaggg tttgaagtgg tgggcgttta ccacgacccg 540
aaaccgggcg gcgtttctaa cgactgggcg ggtaacctgc aacagctggt cgaggacgcg 600
aaagcgggca agattcataa cgtctatatc gcgatgcaaa tgtgcgacgg cgcgcgagtg 660
aaaaaactgg tccatcaact ggcggacacc acctgttcgg tgctgctgat ccccgacgtc 720
tttaccttca acattctcca ttcacgcctc gaagagatga acggcgtacc ggtggtgccg 780
ctttacgaca cgccgctttc cggggttaac cgcctgctca aacgtgcgga agacattgtg 840
ctggcgacgc ttattctgct gctgatctcc ccggtgctgt gctgtattgc gctggcggtg 900
aaactcagtt caccagggcc ggttattttc cgccagactc gctacggcat ggatggcaag 960
ccgatcaaag tgtggaagtt ccgttccatg aaagtgatgg agaacgacaa agtggtgacc 1020
caggcgacgc agaacgatcc gcgcgtcacc aaagtgggga actttctgcg ccgtacctcg 1080
ctggatgaat tgccgcagtt tatcaatgtg ctgaccgggg ggatgtcgat tgtcggtcca 1140
cgtccgcacg cagtagcgca taacgaacag tatcgacagc tcattgaagg ctacatgctg 1200
cgccataagg tgaaaccggg cattaccggc tgggcgcaga ttaacggctg gcgcggcgaa 1260
accgacacgc tggagaaaat ggaaaaacgc gtcgagttcg accttgagta catccgcgaa 1320
tggagcgtct ggttcgatat caaaatcgtt ttcctgacgg tgttcaaagg tttcgttaac 1380
aaagcggcat attga 1395
<210> 95
<211> 464
<212> PRT
<213> 大肠杆菌
<400> 95
Met Thr Asn Leu Lys Lys Arg Glu Arg Ala Lys Thr Asn Ala Ser Leu
1 5 10 15
Ile Ser Met Val Gln Arg Phe Ser Asp Ile Thr Ile Met Phe Ala Gly
20 25 30
Leu Trp Leu Val Cys Glu Val Ser Gly Leu Ser Phe Leu Tyr Met His
35 40 45
Leu Leu Val Ala Leu Ile Thr Leu Val Val Phe Gln Met Leu Gly Gly
50 55 60
Ile Thr Asp Phe Tyr Arg Ser Trp Arg Gly Val Arg Ala Ala Thr Glu
65 70 75 80
Phe Ala Leu Leu Leu Gln Asn Trp Thr Leu Ser Val Ile Phe Ser Ala
85 90 95
Gly Leu Val Ala Phe Asn Asn Asp Phe Asp Thr Gln Leu Lys Ile Trp
100 105 110
Leu Ala Trp Tyr Ala Leu Thr Ser Ile Gly Leu Val Val Cys Arg Ser
115 120 125
Cys Ile Arg Ile Gly Ala Gly Trp Leu Arg Asn His Gly Tyr Asn Lys
130 135 140
Arg Met Val Ala Val Ala Gly Asp Leu Ala Ala Gly Gln Met Leu Met
145 150 155 160
Glu Ser Phe Arg Asn Gln Pro Trp Leu Gly Phe Glu Val Val Gly Val
165 170 175
Tyr His Asp Pro Lys Pro Gly Gly Val Ser Asn Asp Trp Ala Gly Asn
180 185 190
Leu Gln Gln Leu Val Glu Asp Ala Lys Ala Gly Lys Ile His Asn Val
195 200 205
Tyr Ile Ala Met Gln Met Cys Asp Gly Ala Arg Val Lys Lys Leu Val
210 215 220
His Gln Leu Ala Asp Thr Thr Cys Ser Val Leu Leu Ile Pro Asp Val
225 230 235 240
Phe Thr Phe Asn Ile Leu His Ser Arg Leu Glu Glu Met Asn Gly Val
245 250 255
Pro Val Val Pro Leu Tyr Asp Thr Pro Leu Ser Gly Val Asn Arg Leu
260 265 270
Leu Lys Arg Ala Glu Asp Ile Val Leu Ala Thr Leu Ile Leu Leu Leu
275 280 285
Ile Ser Pro Val Leu Cys Cys Ile Ala Leu Ala Val Lys Leu Ser Ser
290 295 300
Pro Gly Pro Val Ile Phe Arg Gln Thr Arg Tyr Gly Met Asp Gly Lys
305 310 315 320
Pro Ile Lys Val Trp Lys Phe Arg Ser Met Lys Val Met Glu Asn Asp
325 330 335
Lys Val Val Thr Gln Ala Thr Gln Asn Asp Pro Arg Val Thr Lys Val
340 345 350
Gly Asn Phe Leu Arg Arg Thr Ser Leu Asp Glu Leu Pro Gln Phe Ile
355 360 365
Asn Val Leu Thr Gly Gly Met Ser Ile Val Gly Pro Arg Pro His Ala
370 375 380
Val Ala His Asn Glu Gln Tyr Arg Gln Leu Ile Glu Gly Tyr Met Leu
385 390 395 400
Arg His Lys Val Lys Pro Gly Ile Thr Gly Trp Ala Gln Ile Asn Gly
405 410 415
Trp Arg Gly Glu Thr Asp Thr Leu Glu Lys Met Glu Lys Arg Val Glu
420 425 430
Phe Asp Leu Glu Tyr Ile Arg Glu Trp Ser Val Trp Phe Asp Ile Lys
435 440 445
Ile Val Phe Leu Thr Val Phe Lys Gly Phe Val Asn Lys Ala Ala Tyr
450 455 460
<210> 96
<211> 1119
<212> DNA
<213> 大肠杆菌
<400> 96
atgattaatt atggcgttgt tggtgttgga tactttggcg ctgaattagc tcgttttatg 60
aatatgcatg ataatgcaaa aattacatgt gtatacgatc ctgaaaatgg agaaaatatt 120
gcccgtgaat tgcagtgtat caatatgtca agcttggatg ctttagtctc aagtaaatta 180
gtcgattgcg tgatcgtagc caccccaaat tatctgcata aagaaccagt aattaaagca 240
gcaaagaata agaagcatgt tttttgtgaa aaaccaattg cattaagtta tgaagattgt 300
gtggatatgg tcaaagcgtg taaagaagct ggtgtgacct ttatggccgg gcatattatg 360
aattttttca atggggttca atatgcacgg aagttaatta aagaaggtgt tatcggcgaa 420
atattatcat gtcatactaa gagaaatggc tgggaaaaca aacaagagag actttcctgg 480
aaaaagatga aagaacaatc tggtggacat ctatatcatc atatacatga gttagattgt 540
gttcagcatt tacttggaga aataccagag acggttacta tgattggtgg aaatttggcc 600
cattctggtc caggatttgg caatgaagat gatatgttat ttatgacctt ggaattcccg 660
tcaggaaaac tagcaacctt agagtggggg agtgcattta actggccgga acattatgtc 720
atcatcaatg gaactaaagg ctctattaaa attgatatgc aagaaacagc agggtcactt 780
aggattggcg gtcagacaaa gcattttttg gtccatgaaa cacaagaaga agatgatgat 840
cgtcggaaag gcaatatgac ctcagaaatg gatggcgcta tagcatatgg tcatccagga 900
aaaaaaacac cattatggct tgccagttta ataagaaagg agacgttatt cctccataat 960
atcctctgtg gtgcaaaacc tgaagaagat tatattgacc ttctcaatgg tgaggcggcc 1020
atgtcggcga ttgctactgc tgatgctgcc actctttcaa gatcgcagga caggaaagtg 1080
aaaatcagtg agatcattaa acatacatca gtaatgtaa 1119
<210> 97
<211> 372
<212> PRT
<213> 大肠杆菌
<400> 97
Met Ile Asn Tyr Gly Val Val Gly Val Gly Tyr Phe Gly Ala Glu Leu
1 5 10 15
Ala Arg Phe Met Asn Met His Asp Asn Ala Lys Ile Thr Cys Val Tyr
20 25 30
Asp Pro Glu Asn Gly Glu Asn Ile Ala Arg Glu Leu Gln Cys Ile Asn
35 40 45
Met Ser Ser Leu Asp Ala Leu Val Ser Ser Lys Leu Val Asp Cys Val
50 55 60
Ile Val Ala Thr Pro Asn Tyr Leu His Lys Glu Pro Val Ile Lys Ala
65 70 75 80
Ala Lys Asn Lys Lys His Val Phe Cys Glu Lys Pro Ile Ala Leu Ser
85 90 95
Tyr Glu Asp Cys Val Asp Met Val Lys Ala Cys Lys Glu Ala Gly Val
100 105 110
Thr Phe Met Ala Gly His Ile Met Asn Phe Phe Asn Gly Val Gln Tyr
115 120 125
Ala Arg Lys Leu Ile Lys Glu Gly Val Ile Gly Glu Ile Leu Ser Cys
130 135 140
His Thr Lys Arg Asn Gly Trp Glu Asn Lys Gln Glu Arg Leu Ser Trp
145 150 155 160
Lys Lys Met Lys Glu Gln Ser Gly Gly His Leu Tyr His His Ile His
165 170 175
Glu Leu Asp Cys Val Gln His Leu Leu Gly Glu Ile Pro Glu Thr Val
180 185 190
Thr Met Ile Gly Gly Asn Leu Ala His Ser Gly Pro Gly Phe Gly Asn
195 200 205
Glu Asp Asp Met Leu Phe Met Thr Leu Glu Phe Pro Ser Gly Lys Leu
210 215 220
Ala Thr Leu Glu Trp Gly Ser Ala Phe Asn Trp Pro Glu His Tyr Val
225 230 235 240
Ile Ile Asn Gly Thr Lys Gly Ser Ile Lys Ile Asp Met Gln Glu Thr
245 250 255
Ala Gly Ser Leu Arg Ile Gly Gly Gln Thr Lys His Phe Leu Val His
260 265 270
Glu Thr Gln Glu Glu Asp Asp Asp Arg Arg Lys Gly Asn Met Thr Ser
275 280 285
Glu Met Asp Gly Ala Ile Ala Tyr Gly His Pro Gly Lys Lys Thr Pro
290 295 300
Leu Trp Leu Ala Ser Leu Ile Arg Lys Glu Thr Leu Phe Leu His Asn
305 310 315 320
Ile Leu Cys Gly Ala Lys Pro Glu Glu Asp Tyr Ile Asp Leu Leu Asn
325 330 335
Gly Glu Ala Ala Met Ser Ala Ile Ala Thr Ala Asp Ala Ala Thr Leu
340 345 350
Ser Arg Ser Gln Asp Arg Lys Val Lys Ile Ser Glu Ile Ile Lys His
355 360 365
Thr Ser Val Met
370
<210> 98
<211> 1776
<212> DNA
<213> 大肠杆菌
<400> 98
atgaaaaaaa tcagcttacc gaaaattggt atccgcccgg ttattgacgg tcgtcgcatg 60
ggtgttcgtg agtcgcttga agaacaaaca atgaatatgg cgaaagctac ggccgcactg 120
ctgaccgaga aactgcgcca tgcctgcgga gctgccgtcg agtgtgtcat ttccgatacc 180
tgtatcgcgg gtatggctga agccgctgct tgcgaagaaa aattcagcag tcagaatgta 240
ggcctcacca ttacggtaac gccttgctgg tgctatggca gtgaaaccat cgacatggat 300
ccaacccgcc cgaaggccat ttggggcttt aacggcactg aacgccccgg cgctgtttac 360
ctggcagcgg ctctggcagc tcacagccag aaaggcatcc cagcattctc catttacggt 420
catgacgttc aggatgccga tgacacatcg attcctgccg atgttgaaga aaaactgctg 480
cgctttgccc gcgccggttt ggccgtcgcc agcatgaaag gtaaaagcta tctgtcgctg 540
ggcggcgttt cgatgggtat cgccggttcc attgttgatc acaacttctt tgaatcctgg 600
ctgggaatga aagtccaggc ggtggatatg accgaactgc gtcgccgtat cgatcagaag 660
atttacgacg aagccgaatt ggaaatggca ctggcctggg ctgataaaaa cttccgctat 720
ggcgaagatg aaaataacaa acagtatcaa cgtaatgccg agcaaagccg cgcagttctg 780
cgcgaaagtt tactgatggc gatgtgtatc cgcgacatga tgcaaggcaa cagcaaactg 840
gccgatattg gtcgcgtgga agaatcactt ggctacaacg ccatcgctgc gggcttccag 900
gggcaacgtc actggaccga tcaatatccc aatggtgaca ccgccgaagc gatcctcaac 960
agttcatttg actggaatgg cgtgcgcgaa ccctttgtcg tggcgaccga aaacgacagt 1020
cttaacggcg tggcaatgct aatgggtcac cagctcaccg gcaccgctca ggtatttgcc 1080
gatgtgcgta cctactggtc accagaagca attgagcgtg taacggggca taaactggat 1140
ggactggcag aacacggcat catccatttg atcaactccg gttctgctgc gctggacggt 1200
tcctgtaaac aacgcgacag cgaaggtaac ccgacgatga agccacactg ggaaatctct 1260
cagcaagagg ctgacgcttg cctcgccgct accgaatggt gcccggcgat ccacgaatac 1320
ttccgtggcg gcggttactc ttcccgcttc cttaccgaag gcggcgtccc gttcaccatg 1380
actcgtgtca acatcatcaa aggcctggga ccggtactgc aaatcgcgga aggctggagc 1440
gtggaattgc cgaaggatgt gcatgacatc ctcaacaaac gcaccaactc aacctggcca 1500
accacctggt ttgcaccgcg cctcaccggt aaagggccgt ttacggatgt gtactcggta 1560
atggcgaact ggggcgctaa ccatggggtt ctgaccatcg gccacgttgg cgcagacttt 1620
atcactctcg cctccatgct gcgtatcccg gtatgtatgc acaacgttga agagaccaaa 1680
gtgtatcgtc cttctgcctg ggctgcgcac ggcatggata ttgaaggcca ggattaccgc 1740
gcttgccaga actacggtcc gttgtacaag cgttaa 1776
<210> 99
<211> 591
<212> PRT
<213> 大肠杆菌
<400> 99
Met Lys Lys Ile Ser Leu Pro Lys Ile Gly Ile Arg Pro Val Ile Asp
1 5 10 15
Gly Arg Arg Met Gly Val Arg Glu Ser Leu Glu Glu Gln Thr Met Asn
20 25 30
Met Ala Lys Ala Thr Ala Ala Leu Leu Thr Glu Lys Leu Arg His Ala
35 40 45
Cys Gly Ala Ala Val Glu Cys Val Ile Ser Asp Thr Cys Ile Ala Gly
50 55 60
Met Ala Glu Ala Ala Ala Cys Glu Glu Lys Phe Ser Ser Gln Asn Val
65 70 75 80
Gly Leu Thr Ile Thr Val Thr Pro Cys Trp Cys Tyr Gly Ser Glu Thr
85 90 95
Ile Asp Met Asp Pro Thr Arg Pro Lys Ala Ile Trp Gly Phe Asn Gly
100 105 110
Thr Glu Arg Pro Gly Ala Val Tyr Leu Ala Ala Ala Leu Ala Ala His
115 120 125
Ser Gln Lys Gly Ile Pro Ala Phe Ser Ile Tyr Gly His Asp Val Gln
130 135 140
Asp Ala Asp Asp Thr Ser Ile Pro Ala Asp Val Glu Glu Lys Leu Leu
145 150 155 160
Arg Phe Ala Arg Ala Gly Leu Ala Val Ala Ser Met Lys Gly Lys Ser
165 170 175
Tyr Leu Ser Leu Gly Gly Val Ser Met Gly Ile Ala Gly Ser Ile Val
180 185 190
Asp His Asn Phe Phe Glu Ser Trp Leu Gly Met Lys Val Gln Ala Val
195 200 205
Asp Met Thr Glu Leu Arg Arg Arg Ile Asp Gln Lys Ile Tyr Asp Glu
210 215 220
Ala Glu Leu Glu Met Ala Leu Ala Trp Ala Asp Lys Asn Phe Arg Tyr
225 230 235 240
Gly Glu Asp Glu Asn Asn Lys Gln Tyr Gln Arg Asn Ala Glu Gln Ser
245 250 255
Arg Ala Val Leu Arg Glu Ser Leu Leu Met Ala Met Cys Ile Arg Asp
260 265 270
Met Met Gln Gly Asn Ser Lys Leu Ala Asp Ile Gly Arg Val Glu Glu
275 280 285
Ser Leu Gly Tyr Asn Ala Ile Ala Ala Gly Phe Gln Gly Gln Arg His
290 295 300
Trp Thr Asp Gln Tyr Pro Asn Gly Asp Thr Ala Glu Ala Ile Leu Asn
305 310 315 320
Ser Ser Phe Asp Trp Asn Gly Val Arg Glu Pro Phe Val Val Ala Thr
325 330 335
Glu Asn Asp Ser Leu Asn Gly Val Ala Met Leu Met Gly His Gln Leu
340 345 350
Thr Gly Thr Ala Gln Val Phe Ala Asp Val Arg Thr Tyr Trp Ser Pro
355 360 365
Glu Ala Ile Glu Arg Val Thr Gly His Lys Leu Asp Gly Leu Ala Glu
370 375 380
His Gly Ile Ile His Leu Ile Asn Ser Gly Ser Ala Ala Leu Asp Gly
385 390 395 400
Ser Cys Lys Gln Arg Asp Ser Glu Gly Asn Pro Thr Met Lys Pro His
405 410 415
Trp Glu Ile Ser Gln Gln Glu Ala Asp Ala Cys Leu Ala Ala Thr Glu
420 425 430
Trp Cys Pro Ala Ile His Glu Tyr Phe Arg Gly Gly Gly Tyr Ser Ser
435 440 445
Arg Phe Leu Thr Glu Gly Gly Val Pro Phe Thr Met Thr Arg Val Asn
450 455 460
Ile Ile Lys Gly Leu Gly Pro Val Leu Gln Ile Ala Glu Gly Trp Ser
465 470 475 480
Val Glu Leu Pro Lys Asp Val His Asp Ile Leu Asn Lys Arg Thr Asn
485 490 495
Ser Thr Trp Pro Thr Thr Trp Phe Ala Pro Arg Leu Thr Gly Lys Gly
500 505 510
Pro Phe Thr Asp Val Tyr Ser Val Met Ala Asn Trp Gly Ala Asn His
515 520 525
Gly Val Leu Thr Ile Gly His Val Gly Ala Asp Phe Ile Thr Leu Ala
530 535 540
Ser Met Leu Arg Ile Pro Val Cys Met His Asn Val Glu Glu Thr Lys
545 550 555 560
Val Tyr Arg Pro Ser Ala Trp Ala Ala His Gly Met Asp Ile Glu Gly
565 570 575
Gln Asp Tyr Arg Ala Cys Gln Asn Tyr Gly Pro Leu Tyr Lys Arg
580 585 590
<210> 100
<211> 1419
<212> DNA
<213> 大肠杆菌
<400> 100
atgaaacaag aagttatcct ggtactcgac tgtggcgcga ccaatgtcag ggccatcgcg 60
gttaatcggc agggcaaaat tgttgcccgc gcctcaacgc ctaatgccag cgatatcgcg 120
atggaaaaca acacctggca ccagtggtct ttagacgcca ttttgcaacg ctttgctgat 180
tgctgtcggc aaatcaatag tgaactgact gaatgccaca tccgcggtat cgccgtcacc 240
acctttggtg tggatggcgc tctggtagat aagcaaggca atctgctcta tccgattatt 300
agctggaaat gtccgcgaac agcagcggtt atggacaata ttgaacggtt aatctccgca 360
cagcggttgc aggctatttc tggcgtcgga gcctttagtt tcaatacgtt atataagttg 420
gtgtggttga aagaaaatca tccacaactg ctggaacgcg cgcacgcctg gctctttatt 480
tcgtcgctga ttaaccaccg tttaaccggc gaattcacta ctgatatcac gatggccgga 540
accagccaga tgctggatat ccagcaacgc gatttcagtc cgcaaatttt acaagccacc 600
ggtattccac gccgactctt ccctcgtctg gtggaagcgg gtgaacagat tggtacgcta 660
cagaacagcg ccgcagcaat gctcggctta cccgttggca taccggtgat ttccgcaggt 720
cacgataccc agttcgccct ttttggcgct ggtgctgaac aaaatgaacc cgtgctctct 780
tccggtacat gggaaatttt aatggttcgc agcgcccagg ttgatacttc gctgttaagt 840
cagtacgccg gttccacctg cgaactggat agccaggcag ggttgtataa cccaggtatg 900
caatggctgg catccggcgt gctggaatgg gtgagaaaac tgttctggac ggctgaaaca 960
ccctggcaaa tgttgattga agaagctcgt ctgatcgcgc ctggcgcgga tggcgtaaaa 1020
atgcagtgtg atttattgtc gtgtcagaac gctggctggc aaggagtgac gcttaatacc 1080
acgcgggggc atttctatcg cgcggcgctg gaagggttaa ctgcgcaatt acagcgcaat 1140
ctacagatgc tggaaaaaat cgggcacttt aaggcctctg aattattgtt agtcggtgga 1200
ggaagtcgca acacattgtg gaatcagatt aaagccaata tgcttgatat tccggtaaaa 1260
gttctcgacg acgccgaaac gaccgtcgca ggagctgcgc tgttcggttg gtatggcgta 1320
ggggaattta acagcccgga agaagcccgc gcacagattc attatcagta ccgttatttc 1380
tacccgcaaa ctgaacctga atttatagag gaagtgtga 1419
<210> 101
<211> 472
<212> PRT
<213> 大肠杆菌
<400> 101
Met Lys Gln Glu Val Ile Leu Val Leu Asp Cys Gly Ala Thr Asn Val
1 5 10 15
Arg Ala Ile Ala Val Asn Arg Gln Gly Lys Ile Val Ala Arg Ala Ser
20 25 30
Thr Pro Asn Ala Ser Asp Ile Ala Met Glu Asn Asn Thr Trp His Gln
35 40 45
Trp Ser Leu Asp Ala Ile Leu Gln Arg Phe Ala Asp Cys Cys Arg Gln
50 55 60
Ile Asn Ser Glu Leu Thr Glu Cys His Ile Arg Gly Ile Ala Val Thr
65 70 75 80
Thr Phe Gly Val Asp Gly Ala Leu Val Asp Lys Gln Gly Asn Leu Leu
85 90 95
Tyr Pro Ile Ile Ser Trp Lys Cys Pro Arg Thr Ala Ala Val Met Asp
100 105 110
Asn Ile Glu Arg Leu Ile Ser Ala Gln Arg Leu Gln Ala Ile Ser Gly
115 120 125
Val Gly Ala Phe Ser Phe Asn Thr Leu Tyr Lys Leu Val Trp Leu Lys
130 135 140
Glu Asn His Pro Gln Leu Leu Glu Arg Ala His Ala Trp Leu Phe Ile
145 150 155 160
Ser Ser Leu Ile Asn His Arg Leu Thr Gly Glu Phe Thr Thr Asp Ile
165 170 175
Thr Met Ala Gly Thr Ser Gln Met Leu Asp Ile Gln Gln Arg Asp Phe
180 185 190
Ser Pro Gln Ile Leu Gln Ala Thr Gly Ile Pro Arg Arg Leu Phe Pro
195 200 205
Arg Leu Val Glu Ala Gly Glu Gln Ile Gly Thr Leu Gln Asn Ser Ala
210 215 220
Ala Ala Met Leu Gly Leu Pro Val Gly Ile Pro Val Ile Ser Ala Gly
225 230 235 240
His Asp Thr Gln Phe Ala Leu Phe Gly Ala Gly Ala Glu Gln Asn Glu
245 250 255
Pro Val Leu Ser Ser Gly Thr Trp Glu Ile Leu Met Val Arg Ser Ala
260 265 270
Gln Val Asp Thr Ser Leu Leu Ser Gln Tyr Ala Gly Ser Thr Cys Glu
275 280 285
Leu Asp Ser Gln Ala Gly Leu Tyr Asn Pro Gly Met Gln Trp Leu Ala
290 295 300
Ser Gly Val Leu Glu Trp Val Arg Lys Leu Phe Trp Thr Ala Glu Thr
305 310 315 320
Pro Trp Gln Met Leu Ile Glu Glu Ala Arg Leu Ile Ala Pro Gly Ala
325 330 335
Asp Gly Val Lys Met Gln Cys Asp Leu Leu Ser Cys Gln Asn Ala Gly
340 345 350
Trp Gln Gly Val Thr Leu Asn Thr Thr Arg Gly His Phe Tyr Arg Ala
355 360 365
Ala Leu Glu Gly Leu Thr Ala Gln Leu Gln Arg Asn Leu Gln Met Leu
370 375 380
Glu Lys Ile Gly His Phe Lys Ala Ser Glu Leu Leu Leu Val Gly Gly
385 390 395 400
Gly Ser Arg Asn Thr Leu Trp Asn Gln Ile Lys Ala Asn Met Leu Asp
405 410 415
Ile Pro Val Lys Val Leu Asp Asp Ala Glu Thr Thr Val Ala Gly Ala
420 425 430
Ala Leu Phe Gly Trp Tyr Gly Val Gly Glu Phe Asn Ser Pro Glu Glu
435 440 445
Ala Arg Ala Gln Ile His Tyr Gln Tyr Arg Tyr Phe Tyr Pro Gln Thr
450 455 460
Glu Pro Glu Phe Ile Glu Glu Val
465 470
<210> 102
<211> 492
<212> DNA
<213> 大肠杆菌
<400> 102
atgcattgct ataacgggat gacaggttta catcaccgcg aaccgggaat ggttggcgcg 60
ggattaacgg acaagcgcgc ctggctggaa ctgatagccg atggtcatca tgtgcatccg 120
gcggcaatgt cgctgtgttg ttgctgtgcg aaagagagaa tcgtactgat caccgacgcg 180
atgcaggcag ctgggatgcc ggatggtcgc tatacgttat gtggtgaaga agtgcagatg 240
cacggtggcg ttgtccgtac cgcgtctggt gggctggcgg gcagtacgct gtctgttgat 300
gcggcagtgc gcaatatggt cgagttgacg ggcgtaacgc tgcggaagcc atccatatgg 360
cgtcgctgca tccggcgcga atgctgggtg ttgatggtgt tctgggatcg cttaaaccgg 420
gcaaacgcgc cagagtcgtt gcgctggata gcgggctaca tgtgcaacaa atctggattc 480
agggtcaatt ag 492
<210> 103
<211> 167
<212> PRT
<213> 大肠杆菌
<400> 103
Met His Cys Tyr Asn Gly Met Thr Gly Leu His His Arg Glu Pro Gly
1 5 10 15
Met Val Gly Ala Gly Leu Thr Asp Lys Arg Ala Trp Leu Glu Leu Ile
20 25 30
Ala Asp Gly His His Val His Pro Ala Ala Met Ser Leu Cys Cys Cys
35 40 45
Cys Ala Lys Glu Arg Ile Val Leu Ile Thr Asp Ala Met Gln Ala Ala
50 55 60
Gly Met Pro Asp Gly Arg Tyr Thr Leu Cys Gly Glu Glu Val Gln Met
65 70 75 80
His Gly Gly Val Val Arg Thr Ala Ser Gly Gly Leu Ala Gly Ser Thr
85 90 95
Leu Ser Val Asp Ala Ala Val Arg Asn Met Val Glu Leu Thr Gly Val
100 105 110
Thr Pro Ala Glu Ala Ile His Met Ala Ser Leu His Pro Ala Arg Met
115 120 125
Leu Gly Val Asp Gly Val Leu Gly Ser Leu Lys Pro Gly Lys Arg Ala
130 135 140
Ser Val Val Ala Leu Asp Ser Gly Leu His Val Gln Gln Ile Trp Ile
145 150 155 160
Gln Gly Gln Leu Ala Ser Phe
165

Claims (23)

1.一种非天然存在的微生物,其能够产生Neu5Ac,其中所述非天然存在的微生物具有包括至少一种异源酶的唾液酸生物合成途径,其中所述微生物的天然存在的唾液酸分解代谢途径已被失效,其中至少一种用于输入在发酵生产Neu5Ac期间不用作碳源的糖的磷酸烯醇丙酮酸:糖磷酸转移酶系统已被失效,其中所述微生物可以利用发酵液中存在的外源碳源作为唯一的碳源,而不使用磷酸烯醇丙酮酸:糖磷酸转移酶系统来获取所述外源碳源。
2.根据权利要求1所述的非天然存在的微生物,其中所述唾液酸生物合成途径包括谷氨酰胺-果糖-6-磷酸转氨酶、葡糖胺-6-磷酸N-乙酰转移酶、N-乙酰葡糖胺-2-差向异构酶、N-乙酰神经氨酸合酶和类HAD超家族的糖磷酸酶。
3.根据权利要求1或2所述的非天然存在的微生物,其中所述唾液酸生物合成途径的至少一种酶为异源酶。
4.根据权利要求1至3中任一项所述的非天然存在的微生物,其中天然存在的唾液酸分解代谢途径已被失效。
5.根据权利要求1至4中任一项所述的非天然存在的微生物,其中一种或多种编码参与唾液酸分解途代谢径的酶的基因被从非天然存在微生物的基因组中缺失,其中一种或多种编码参与唾液酸分解代谢途径的酶的基因的表达被削弱,或者其中至少一种编码参与唾液酸分解代谢途径的酶的基因的蛋白质编码区的核苷酸序列被改变,使得由所述改变的蛋白质编码核苷酸序列编码的多肽不具有由未改变的核苷酸序列编码的酶的酶活性。
6.根据权利要求1至5中任一项所述的非天然存在的微生物,其中所述非天然存在的微生物已被遗传工程化,以缺失一个或多个选自以下的基因:编码N-乙酰甘露糖胺激酶、N-乙酰甘露糖胺-6-磷酸差向异构酶、N-乙酰神经氨酸醛缩酶和唾液酸通透酶的基因,其中这些基因中一个或多个的表达被削弱,或者其中这些基因中至少一个的蛋白质编码区的核苷酸序列被改变,使得由所述改变的核苷酸序列编码的多肽不具有由未改变的核苷酸序列编码的酶的酶活性。
7.根据权利要求1至6中任一项所述的非天然存在的微生物,其中一种或多种选自N-乙酰葡糖胺-6-磷酸脱乙酰酶和N-乙酰葡糖胺-6-磷酸脱氨酶的酶的活性已被消除。
8.根据权利要求1至7中任一项所述的非天然存在的微生物,其中所述非天然存在的微生物已被遗传工程化,以通过以下方式消除N-乙酰葡糖胺-6-磷酸脱乙酰酶和/或N-乙酰葡糖胺-6-磷酸脱氨酶的活性:通过缺失一个或两个编码这些酶的基因,通过削弱这些基因中一个或两个的表达,或通过突变一个或两个基因的蛋白质编码区,使得由每个改变的核苷酸序列编码的多肽不具有由未改变的核苷酸序列编码的酶的酶活性。
9.根据权利要求1至8中任一项所述的非天然存在的微生物,其中至少一种用于输入碳水化合物的磷酸烯醇丙酮酸(PEP)依赖的糖转运磷酸转移酶系统已被失效。
10.根据权利要求1至9中任一项所述的非天然存在的微生物,其中所述非天然存在的微生物包含糖/H+-同向转运体,优选地糖/H+-同向转运体选自蔗糖质子同向转运体、乳糖质子同向转运体和葡萄糖质子同向转运体。
11.根据权利要求1至10中任一项所述的非天然存在的微生物,其中与野生型微生物相比,所述非天然存在的微生物具有增强的PEP生物合成,优选地由于PEP合酶的过表达。
12.根据权利要求1至11中任一项所述的非天然存在的微生物,其中所述非天然存在的微生物确实缺乏一种或多种选自以下的酶:功能性PEP羧化酶、功能性谷氨酸合酶、功能性WzxC蛋白、功能性UDP-葡萄糖:十一异戊烯基磷酸葡萄糖-1-磷酸转移酶、功能性β-半乳糖苷通透酶、功能性β-半乳糖苷酶、功能性YjhC蛋白、功能性岩藻糖异构酶、功能性墨角藻糖激酶和功能性N-乙酰谷氨酰胺氨基酰化酶。
13.根据权利要求1至12中任一项所述的非天然存在的微生物,其中与野生型微生物相比,所述非天然存在的微生物具有增强的谷氨酰胺合成,优选地由于谷氨酰胺合酶的过表达。
14.根据权利要求1至13中任一项所述的非天然存在的微生物用于生产Neu5Ac的用途。
15.一种通过使用能够生产Neu5Ac的非天然存在的微生物的发酵来生产Neu5Ac的方法,该方法包括以下步骤:
a)提供权利要求1至13中任一项所述的非天然存在的微生物;
b)在发酵液中并在允许非天然存在的微生物产生Neu5Ac的条件下培养所述非天然存在的微生物;和
c)任选地,从发酵液中回收Neu5Ac。
16.根据权利要求15的方法,其中所述发酵液含有用于生长非天然存在的微生物的碳源,所述碳源优选地选自葡萄糖、木糖、蔗糖、果糖、乳糖、甘油、合成气及其组合。
17.通过根据权利要求15或16的方法生产的Neu5Ac用于制备营养组合物的用途。
18.一种含有通过根据权利要求15或16的方法生产的Neu5Ac的营养组合物。
19.根据权利要求18所述的营养组合物,其中所述的组合物还包含至少一种HMO,优选至少一种中性HMO和/或至少一种唾液酸化HMO。
20.根据权利要求18或19所述的营养组合物,其中所述营养组合物还含有益生菌微生物。
21.根据权利要求18至20中任一项所述的营养组合物,其中所述营养组合物选自药物制剂、婴儿配方物和膳食补充剂。
22.根据权利要求18至21中任一项所述的营养组合物,其中所述营养组合物以液体形式存在,优选地以浓缩物或即饮饮品的形式;或以固体形式存在,优选地以粉末剂、颗粒剂、薄片和丸剂的形式。
23.一种含有通过权利要求1至13中任一项的方法生产的Neu5Ac的婴儿配方物。
CN201880081537.1A 2017-10-17 2018-10-17 N-乙酰神经氨酸的发酵生产 Pending CN111556873A (zh)

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