CN117396217A - 无催化活性的梭菌神经毒素用于疼痛和炎性疾病的治疗 - Google Patents

无催化活性的梭菌神经毒素用于疼痛和炎性疾病的治疗 Download PDF

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CN117396217A
CN117396217A CN202180098695.XA CN202180098695A CN117396217A CN 117396217 A CN117396217 A CN 117396217A CN 202180098695 A CN202180098695 A CN 202180098695A CN 117396217 A CN117396217 A CN 117396217A
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M·加里尼契夫
C·佩里埃
C·法夫尔
S·科尔内
J·克虏伯
M·埃利奥特
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Abstract

本发明涉及用于治疗疼痛或炎性疾病的多肽，其中该多肽包含梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)，其中当该多肽包含梭菌神经毒素L链时，该L链无催化活性。还提供相应的治疗方法和用途。

Description

无催化活性的梭菌神经毒素用于疼痛和炎性疾病的治疗
技术领域
本发明涉及多肽用于治疗，例如多肽在治疗疼痛或炎性疾病中的用途。
背景技术
梭菌属(Clostridia)的细菌产生强效和特异性的蛋白质毒素，这些毒素可以使它们所递送到的神经元及其他细胞中毒。这类梭菌毒素的实例包括由破伤风梭菌(C.tetani)(TeNT)和肉毒梭菌(C.botulinum)(BoNT)血清型A-G和X产生的神经毒素(参见WO 2018/009903 A2)，以及由巴氏梭菌(C.baratii)和丁酸梭菌(C.butyricum)产生的神经毒素。破伤风毒素和肉毒毒素都通过抑制受影响神经元的功能、特别是神经递质的释放发挥作用。肉毒毒素作用于神经肌肉接头并抑制外周神经系统中的胆碱能传递，而破伤风毒素作用于中枢神经系统。
在自然界中，梭菌神经毒素作为单链多肽合成，该单链多肽在翻译后通过蛋白水解切割事件进行修饰，形成通过二硫键连接在一起的两条多肽链。切割发生在位于提供链间二硫键的半胱氨酸残基之间的通常称为活化位点的特异性切割位点处。正是这种双链形式是毒素的活性形式。这两条链称为重链(H链)和轻链(L链)，重链的分子量约为100kDa，轻链的分子量约为50kDa。H链包含N端易位组分(HN结构域)和C端靶向组分(HC结构域)。切割位点位于L链和转运结构域(translocation domain)组分之间。在HC结构域与其靶神经元结合并通过内体将结合的毒素内化到细胞中之后，HN结构域使L链易位跨过内体膜并进入到胞质溶胶中，并且L链提供蛋白酶功能(也称为非细胞毒性蛋白酶)。
非细胞毒性蛋白酶通过蛋白水解切割称为SNARE蛋白的细胞内转运蛋白(例如SNAP-25、VAMP或突触融合蛋白)发挥作用。首字母缩写SNARE来源于术语可溶性NSF附着受体(Soluble NSF Attachment Receptor)，其中NSF是指N-乙基马来酰亚胺敏感因子(N-ethylmaleimide-Sensitive Factor)。SNARE蛋白是细胞内囊泡融合的组成部分，从而通过囊泡运输自细胞分泌分子。该蛋白酶功能是锌依赖性内肽酶活性，并对SNARE蛋白表现出高底物特异性。因此，一旦递送到所希望的靶细胞，非细胞毒性蛋白酶就能够抑制自靶细胞的细胞分泌。梭菌神经毒素的L链蛋白酶是切割SNARE蛋白的非细胞毒性蛋白酶。
鉴于SNARE蛋白的遍在特性，梭菌神经毒素如肉毒毒素已成功应用于广泛的治疗中。
梭菌神经毒素是已知的最强效的一些毒素。例如，肉毒神经毒素对小鼠的半数致死剂量(LD50)值在0.5至5ng/kg的范围内，这取决于血清型。因此，使用该毒素并非没有风险。认为毒素从施用部位扩散到周围组织或全身循环中造成了梭菌神经毒素治疗的不良副作用，在极端情况下可能会危及生命。在使用高剂量、高浓度和/或高注射体积的梭菌神经毒素时，这可以是特别令人担忧的问题。据报道，商业化BoNT/A疗法的不良反应包括乏力、全身肌无力、复视、上睑下垂、吞咽困难、发音困难、构音障碍、尿失禁和呼吸困难。吞咽和呼吸困难可以危及生命，与毒素效应扩散相关的死亡已有报道。
本发明克服了上述问题中的一个或多个问题。
发明概述
本发明人已经发现无催化活性的梭菌神经毒素在治疗疼痛方面有效。这一发现尤其令人惊讶，因为认为导致SNARE蛋白切割的催化活性是梭菌神经毒素治疗的必要作用机制。因此，本发明的多肽避免了与常规催化活性梭菌神经毒素治疗相关的毒性副作用，并且构成了更安全(基本无毒)的治疗剂。有利地，与常规的催化活性梭菌神经毒素治疗剂相比，本发明的多肽可以以更大的量给药。此外，本发明多肽降低的毒性允许在整个产品生命周期中易于制造和处理，并消除了医生进行复杂(例如个性化)给药方案计算以避免受试者的毒性的需要。
同样令人惊讶的是，本发明人已经发现无催化活性梭菌神经毒素在治疗炎性疾病方面有效。
发明详述
在一个方面，本发明提供用于治疗疼痛的多肽(例如镇痛多肽)，其中该多肽包含梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)，其中当该多肽包含梭菌神经毒素L链时，该L链无催化活性。
在一个相关方面，提供用于治疗疼痛的方法，该方法包括对受试者施用多肽(例如镇痛多肽)，其中该多肽包含梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)，其中当该多肽包含梭菌神经毒素L链时，该L链无催化活性。
在另一个相关方面，本发明提供多肽(例如镇痛多肽)在制备用于治疗疼痛的药物中的用途，其中该多肽包含梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)，其中当该多肽包含梭菌神经毒素L链时，该L链无催化活性。
本发明的多肽优选具有镇痛特性。换言之，本发明的多肽优选为镇痛多肽。
优选地，本发明的多肽既不促进神经元生长也不促进神经元修复以治疗疼痛。换言之，优选地，该多肽不通过以下任何方式治疗疼痛：不通过促进神经元生长、不通过促进神经元修复或不通过促进神经元生长和修复。
在一个方面，本发明提供用于治疗炎性疾病的多肽，其中该多肽包含梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)，其中当该多肽包含梭菌神经毒素L链时，该L链无催化活性。
在一个相关方面，提供用于治疗炎性疾病的方法，该方法包括对受试者施用多肽，其中该多肽包含梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)，其中当该多肽包含梭菌神经毒素L链时，该L链无催化活性。
在另一个相关方面，本发明提供多肽在制备用于治疗炎性疾病的药物中的用途，其中该多肽包含梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)，其中当该多肽包含梭菌神经毒素L链时，该L链无催化活性。
本发明的多肽可以具有抗炎特性。换言之，本发明的多肽可以是抗炎多肽。
在多肽用于治疗本文所述的炎性疾病时，该多肽可以包含肉毒神经毒素血清型X(BoNT/X)L链、BoNT/X HN结构域和/或BoNT/X HC结构域，其中当该多肽包含梭菌神经毒素L链时，该L链无催化活性。例如，该多肽可以是嵌合肉毒神经毒素(BoNT)，其包含无催化活性BoNT/X轻链和转运结构域，以及来自不同(即，非BoNT/X)梭菌神经毒素的受体结合结构域(HC结构域)。因此，在一个方面，本发明提供用于治疗炎性疾病的多肽，其中该多肽包含无催化活性BoNT/X轻链和转运结构域，以及来自不同(即，非BoNT/X)梭菌神经毒素的受体结合结构域(HC结构域)(优选BoNT/B HC结构域)。本发明还提供相应的治疗方法和用途。
优选地，本发明的多肽既不促进神经元生长也不促进神经元修复以治疗炎性病症。换言之，优选地，该多肽不通过以下任何方式治疗炎性病症：不通过促进神经元生长、不通过促进神经元修复或不通过促进神经元生长和修复。
术语“促进神经元生长和/或神经元修复”涵盖神经元生长和/或神经元修复速率的增加。术语“神经元生长和/或神经元修复”涵盖重建受损的神经元回路，从而恢复神经元网络或群体中的活动和/或神经元通讯。因此，本文所用的术语“神经元修复”涵盖具体神经元的修复以及神经元回路的修复。该术语还涵盖神经元可塑性。本文所用的术语“神经元可塑性”涵盖轴突发芽、树突发芽、神经发生(例如产生新神经元)、成熟、分化和/或突触可塑性(例如包括突触强度、活动、解剖结构和/或连接性的变化)。术语“促进神经元生长和/或神经元修复”还涵盖促进功能性突触的建立(例如在损伤部位或损伤部位附近)。本文所用的术语“神经元生长”涵盖神经元任何部分的生长，包括轴突和/或树突的生长。该术语涵盖增加轴突长度、轴突数量(例如每个细胞的轴突数量)和/或增加神经元的细胞体或细胞膜的突起的长度和/或数量，例如神经元的轴突生长和/或轴突发芽，例如受试者中的神经元。该轴突生长可以促进神经元之间的连接和/或化学通讯。
优选地，本发明的多肽不促进神经免疫反应来治疗疼痛或炎性疾病。在这种情况下，神经免疫反应涵盖小胶质细胞反应。因此，在一个实施方案中，本发明的多肽不促进小胶质细胞反应来治疗疼痛或炎性病症。
在一个优选实施方案中，该疼痛不是与大脑病症相关或由大脑病症引起的疼痛。在另一个优选实施方案中，该炎性疾病不是炎性大脑病症。在这种情况下使用的术语“大脑病症”可以与“大脑疾病”互换。本文所用的“大脑病症”涵盖源自大脑内部或外部的疾病，且包括与导致大脑组织损伤的身体损伤相关的疾病。本文中所涵盖的大脑病症的实例包括创伤性脑损伤、癌症(例如脑瘤)、感染性疾病(例如脑炎、脑膜炎、脑脓肿和脑炎)、中风、神经退行性障碍(例如阿尔茨海默病、帕金森病、帕金森病相关疾病、运动神经元疾病(例如肌萎缩性侧索硬化症)、朊病毒病、亨廷顿病、脊髓小脑共济失调、共济失调、哈勒沃登-施帕茨病和额颞叶变性)、脑动脉瘤、多发性硬化、缺氧性损伤、毒性损伤和代谢性损伤中的任何一种(或多种)。脑障碍可以由创伤性脑损伤、癌症、感染性疾病(例如脑炎、脑膜炎、脑脓肿和脑炎)、中风、神经退行性障碍(例如阿尔茨海默病、帕金森病、帕金森病相关疾病、运动神经元病(如肌萎缩性侧索硬化症)、朊病毒病、亨廷顿病、脊髓小脑共济失调、共济失调、哈勒沃登-施帕茨病和额颞叶变性)、脑动脉瘤、多发性硬化、缺氧性损伤、毒性损伤和/或代谢性损伤引起。
活性梭菌神经毒素L链具有非细胞毒性蛋白酶活性。具体而言，活性梭菌神经毒素L链具有内肽酶活性，并且能够在靶细胞中切割外细胞融合器的蛋白质。外细胞融合器的蛋白质优选SNARE蛋白，例如SNAP25、小突触小泡蛋白/VAMP或突触融合蛋白。
本文所用的关于梭菌神经毒素L链的术语“无催化活性”是指该L链基本上不表现出非细胞毒性蛋白酶活性，优选地，本文所用的关于梭菌神经毒素L链的术语“无催化活性”是指该L链不表现出非细胞毒性蛋白酶活性。在一个实施方案中，无催化活性梭菌神经毒素L链是不切割靶细胞中的外细胞融合器的蛋白质的L链。术语“基本上没有非细胞毒性蛋白酶活性”是指梭菌神经毒素L链具有小于5％的催化活性梭菌神经毒素L链的非细胞毒性蛋白酶活性，例如小于2％、1％或优选小于0.1％的催化活性梭菌神经毒素L链的非细胞毒性蛋白酶活性。非细胞毒性蛋白酶活性可通过在相同条件下将测试梭菌神经毒素L链与SNARE蛋白孵育，并将受试梭菌神经毒素L链切割的SNARE蛋白的量与催化活性梭菌神经毒素L链切割的SNARE蛋白的量相比较来在体外进行测定。常规技术，如SDS-PAGE和蛋白质印迹，可用于定量所切割的SNARE蛋白的量。适宜的体外测定描述于WO 2019/145577 A1中，其在此引入作为参考。
基于细胞的测定法和体内测定法也可用于确定包含L链及功能性细胞结合和转运结构域的梭菌神经毒素是否具有非细胞毒性蛋白酶活性。诸如趾外展评分(DAS)测定法、背根神经节(DRG)测定法、脊髓神经元(SCN)测定法和小鼠膈神经半膈肌(PNHD)测定法的测定法是本领域的常规测定法。用于测定非细胞毒性蛋白酶活性的适宜测定法可以是在AokiKR,Toxicon 39:1815-1820；2001或Donald等(2018),Pharmacol Res Perspect,e00446,1-14中所述的测定法，其在此引入作为参考。
无催化活性L链可以具有一个或多个使该催化活性失活的突变。因此，催化活性L链(例如本文所述)可以经修饰以引入一个或多个使L链的催化活性失活的突变。例如，无催化活性L链可以包含活性位点残基的突变。突变可以是取代或缺失，但是优选取代，尤其是用化学相似的氨基酸取代。谷氨酸可以用谷氨酰胺取代，组氨酸可以用酪氨酸取代，精氨酸可以用谷氨酰胺取代，和/或酪氨酸可以用苯丙氨酸取代。备选地，任何残基都可以用丙氨酸取代。
无催化活性BoNT/A L链可以在H223、E224、H227、E262、R363和/或Y366处，优选至少在E224和H227处包含突变。优选地，无催化活性BoNT/AL链可以包含在E224处用谷氨酰胺取代(E224Q)和在H227处用酪氨酸取代(H227Y)。位置编号对应于SEQ ID NO:60的氨基酸位置，并且可以通过将多肽与SEQ ID NO:60比对来确定。由于在SEQ ID NO:60的位置1处的甲硫氨酸残基的存在是任选的，本领域技术人员在确定氨基酸残基编号时将考虑甲硫氨酸残留的存在/不存在。例如，在SEQ ID NO:60包含甲硫氨酸的情况下，位置编号将如上所定义(例如His223将是SEQ ID NO:60的His223)。备选地，在SEQ ID NO:60中不存在甲硫氨酸的情况下，氨基酸残基编号应改为-1(例如His223将是SEQ ID NO:60的His222)。在本文所述的其他多肽序列的位置1处的甲硫氨酸存在/不存在时，类似的考虑适用，并且本领域技术人员将使用本领域常规技术容易地确定正确的氨基酸残基编号。
无催化活性BoNT/B L链可以在E231和/或H234处，优选在E231和H234处包含突变。优选地，无催化活性BoNT/B L链包含在E231处用谷氨酰胺取代(E231Q)和在H234处用酪氨酸取代(H234Y)。位置编号对应于SEQ ID NO:52的氨基酸位置，并且可以通过将多肽与SEQID NO:52比对来确定。由于在SEQ ID NO:52的位置1处甲硫氨酸残基的存在是任选的，本领域技术人员在确定氨基酸残基编号时将考虑甲硫氨酸残基的存在/不存在。
无催化活性BoNT/C L链可以在H229、E230和/或H233处，优选在H229、E230和H233处包含突变。优选地，无催化活性BoNT/C L链包含在H229处用甘氨酸取代(H229G)，在E230处用苏氨酸取代(E230T)，以及在H233处用天冬酰胺取代(H233N)。位置编号对应于SEQ IDNO:53的氨基酸位置，并且可以通过将多肽与SEQ ID NO:53比对来确定。由于在SEQ ID NO:53的位置1处甲硫氨酸残基的存在是任选的，本领域技术人员在确定氨基酸残基编号时将考虑甲硫氨酸残基的存在/不存在。
无催化活性BoNT/D L链可以在H229、E230、H233和/或H236处，优选至少在E230和H236处包含突变。优选地，无催化活性BoNT/D L链至少包含在E230处用谷氨酰胺取代(E230Q)和在H236处用酪氨酸取代(H236Y)。位置编号对应于SEQ ID NO:54的氨基酸位置，并且可以通过将多肽与SEQ ID NO:54比对来确定。由于在SEQ ID NO:54的位置1处甲硫氨酸残基的存在是任选的，本领域技术人员在确定氨基酸残基编号时将考虑甲硫氨酸残基的存在/不存在。
无催化活性BoNT/E L链可以在E213和/或H216处，优选在E213和H216处包含突变。优选地，无催化活性BoNT/E L链包含在E213处用谷氨酰胺取代(E213Q)和在H216处用酪氨酸取代(H216Y)。位置编号对应于SEQ ID NO:55的氨基酸位置，并且可以通过将多肽与SEQID NO:55比对来确定。由于在SEQ ID NO:55的位置1处甲硫氨酸残基的存在是任选的，本领域技术人员在确定氨基酸残基编号时将考虑甲硫氨酸残基的存在/不存在。
无催化活性BoNT/F L链可以在E228和/或H231处，优选在E228和H231处包含突变。优选地，无催化活性BoNT/F L链包含在E228处用谷氨酰胺取代(E228Q)和在H231处用酪氨酸取代(H231Y)。位置编号对应于SEQ ID NO:56的氨基酸位置，并且可以通过将多肽与SEQID NO:66比对来确定。由于在SEQ ID NO:56的位置1处甲硫氨酸残基的存在是任选的，本领域技术人员在确定氨基酸残基编号时将考虑甲硫氨酸残基的存在/不存在。
无催化活性BoNT/G L链可以在E231和/或H234处，优选在E231和H234处包含突变。优选地，无催化活性BoNT/G L链包含在E231处用谷氨酰胺取代(E231Q)和在H234处用酪氨酸取代(H234Y)。位置编号对应于SEQ ID NO:57的氨基酸位置，并且可以通过将多肽与SEQID NO:57比对来确定。由于在SEQ ID NO:57的位置1处甲硫氨酸残基的存在是任选的，本领域技术人员在确定氨基酸残基编号时将考虑甲硫氨酸残基的存在/不存在。
无催化活性BoNT/X L链可以在E228和/或H231处，优选在E228和H231处包含突变。优选地，无催化活性BoNT/X L链包含在E228处用谷氨酰胺取代(E228Q)和在H231处用酪氨酸取代(H231Y)。位置编号对应于SEQ ID NO:59的氨基酸位置，并且可以通过将多肽与SEQID NO:59比对来确定。由于在SEQ ID NO:59的位置1处甲硫氨酸残基的存在是任选的，本领域技术人员在确定氨基酸残基编号时将考虑甲硫氨酸残基的存在/不存在。
无催化活性TeNT L链可以在E234、R372和/或Y375处，优选至少在R372和Y375处(例如在E234、R372和Y375处)包含突变。优选地，无催化活性TeNT L链包含在R372处用谷氨酰胺或丙氨酸取代(R372Q或R372A)，更优选用丙氨酸取代，以及在Y375处用苯丙氨酸取代(Y375F)。位置编号对应于SEQ ID NO:58的氨基酸位置，并且可以通过将多肽与SEQ ID NO:58比对来确定。由于在SEQ ID NO:58的位置1处甲硫氨酸残基的存在是任选的，本领域技术人员在确定氨基酸残基编号时将考虑甲硫氨酸残基的存在/不存在。
本发明的多肽可以包含全长梭菌神经毒素(条件是L链无催化活性)或不具有非细胞毒性蛋白酶活性的梭菌神经毒素片段(例如HN结构域和/或HC结构域)。换言之，本发明的多肽不具有非细胞毒性蛋白酶活性。
术语“梭菌神经毒素”包括由肉毒梭菌(C.botulinum)(肉毒神经毒素血清型A、B、C1、D、E、F、G和X)、破伤风梭菌(C.tetani)(破伤风神经毒素)、丁酸梭菌(C.butyricum)(肉毒神经毒素血清型E)和巴氏梭菌(C.baratii)(肉毒神经毒素血清型F)产生的毒素。参考BoNT/A序列显示为SEQ ID NO:51。参考BoNT/B序列显示为SEQ ID NO:52。参考BoNT/C序列显示为SEQ ID NO:53。参考BoNT/D序列显示为SEQ ID NO:54。参考BoNT/E序列显示为SEQID NO:55。参考BoNT/F序列显示为SEQ ID NO:56。参考BoNT/G序列显示为SEQ ID NO:57。参考TeNT序列显示为SEQ ID NO:58。参考BoNT/X序列显示为SEQ ID NO:59。术语“梭菌神经毒素”也可以包括新发现的由非梭菌微生物表达的肉毒神经毒素蛋白家族成员，如与BoNT/X具有最接近序列同一性的肠球菌属(Enterococcus)编码的毒素，称为BoNT/Wo的米魏斯氏菌(Weissella oryzae)编码的毒素(NCBI参考序列:WP_027699549.1)，其在W89-W90处切割VAMP2，粪肠球菌(Enterococcus faecium)编码的毒素(GenBank:OTO22244.1)，其切割VAMP2和SNAP25，以及胡椒金黄杆菌(Chryseobacterium pipero)编码的毒素(NCBI参考序列:WP_034687872.1)。
因此，梭菌神经毒素可以选自BoNT/A、BoNT/B、BoNT/C、BoNT/D、BoNT/E、BoNT/F、BoNT/G、BoNT/X和TeNT(破伤风神经毒素)。优选地，梭菌神经毒素是肉毒神经毒素，如选自BoNT/A、BoNT/B、BoNT/C、BoNT/D、BoNT/E、BoNT/F、BoNT/G和BoNT/X的肉毒神经毒素。例如，梭菌神经毒素HN结构域可以是来自BoNT A、B、C1、D、E、F、G、X或TeNT的HN结构域。类似地，L链可以是来自BoNT A、B、C1、D、E、F、G、X或TeNT的L链，条件是该L链无催化活性(例如已修饰以使其无催化活性)。更优选地，该梭菌神经毒素是BoNT/A。
如上所述，(全长)梭菌神经毒素由两条多肽链形成，重链(H链)具有约100kDa的分子量，轻链(L链)具有约50kDa的分子量。H链包含C端靶向组分(受体结合结构域或HC结构域)和N端易位组分(HN结构域)。肉毒神经毒素(BoNT)由肉毒梭菌以大型蛋白质复合物的形式产生，由BoNT本身与许多辅助蛋白质复合而成。目前有八类不同的肉毒神经毒素，即：肉毒神经毒素血清型A、B、C1、D、E、F、G和X，所有这些都具有相似的结构和作用模式。不同的BoNT血清型可以根据特异性中和抗血清的失活来区分，这种按血清型的分类与氨基酸水平的百分比序列同一性相关。给定血清型的BoNT蛋白根据氨基酸百分比序列同一性进一步分为不同的亚型。
常规(催化活性)BoNT在胃肠道中被吸收，进入总循环后，与胆碱能神经末梢的突触前膜结合，并阻止其神经递质乙酰胆碱的释放。BoNT/B、BoNT/D、BoNT/F和BoNT/G切割小突触小泡蛋白/小泡相关膜蛋白(VAMP)；BoNT/C1、BoNT/A和BoNT/E切割25kDa的突触体相关蛋白(SNAP-25)；BoNT/C1切割突触融合蛋白。已发现BoNT/X切割SNAP-25、VAMP1、VAMP2、VAMP3、VAMP4、VAMP5、Ykt6和突触融合蛋白1。破伤风毒素由破伤风梭菌以单一血清型产生。丁酸梭菌产生BoNT/E，而巴氏梭菌产生BoNT/F。
在一个实施方案中，本发明的多肽可以由与SEQ ID NO:1、3、5、7、9、11、13、15、17、19、21、23、25、27、29、31、33、35、37、39、41、43、45、47或49中的任一个具有至少70％序列同一性的核苷酸序列编码，条件是在该多肽包含梭菌神经毒素L链时，该L链无催化活性。在一个实施方案中，本发明的多肽可以由与SEQ ID NO:1、3、5、7、9、11、13、15、17、19、21、23、25、27、29、31、33、35、37、39、41、43、45、47或49中的任一个具有至少80％、90％、95％或98％序列同一性的核苷酸序列编码，条件是在该多肽包含梭菌神经毒素L链时，该L链无催化活性。优选地，本发明的多肽可以由包含SEQ ID NO:1、7、9、11、13、15、17、21、25、29、33、37、41、43、45、47或49中的任一个的核苷酸序列编码。
在一个实施方案中，本发明的多肽可以包含与SEQ ID NO:2、4、6、8、10、12、14、16、18、20、22、24、26、28、30、32、34、36、38、40、42、44、46、48、50、51、52、53、54、55、56、57、58、59、60、61、62、63、64、65、66、67、68、69、70、74、75或76中的任一个具有至少70％序列同一性的多肽序列，条件是在该多肽包含梭菌神经毒素L链时，该L链无催化活性。在一个实施方案中，本发明的多肽可以包含与SEQ ID NO:2、4、6、8、10、12、14、16、18、20、22、24、26、28、30、32、34、36、38、40、42、44、46、48、50、51、52、53、54、55、56、57、58、59、60、61、62、63、64、65、66、67、68、69、70、74、75或76中的任一个具有至少80％、90％、95％或98％序列同一性的多肽序列，条件是在该多肽包含梭菌神经毒素L链时，该L链无催化活性。优选地，本发明的多肽可以包含SEQ ID NO:2、8、10、12、14、16、18、22、26、30、34、38、42、44、46、48、50、61、62、63、64、65、66、67、68、69、70、74、75或76中的任一个的多肽序列。
在一个实施方案中，本发明的多肽可以包含与SEQ ID NO:2,10,12,14,16,18,26,34,51,52,53,54,55,56,57,58,59,60,61,62,63,64,65,66,67,68,69或70中的任一个具有至少70％序列同一性的多肽序列的片段，条件是在该多肽包含梭菌神经毒素L链时，该L链无催化活性。在一个实施方案中，本发明的多肽可以包含与SEQ ID NO:2,10,12,14,16,18,26,34,51,52,53,54,55,56,57,58,59,60,61,62,63,64,65,66,67,68,69或70中的任一个具有至少80％、90％、95％或98％序列同一性的多肽序列的片段，条件是在该多肽包含梭菌神经毒素L链时，该L链无催化活性。优选地，本发明的多肽可以包含含有SEQ ID NO:2,10,12,14,16,18,26,34,51,52,53,54,55,56,57,58,59,60,61,62,63,64,65,66,67,68,69或70中的任一个的多肽序列的片段，条件是在该多肽包含梭菌神经毒素L链时，该L链无催化活性。该片段可以是任一所述SEQ ID NO中的无催化活性L链、HN结构域或HC结构域。
优选地，本发明的多肽包含无催化活性梭菌神经毒素L链(或由其组成)。本文中提及的无催化活性梭菌神经毒素还涵盖梭菌神经毒素L链的片段。梭菌神经毒素L链的片段可以具有梭菌神经毒素L链的≤400、≤350、≤300、≤250、≤200、≤150、≤100或≤50个氨基酸残基。在一个实施方案中，梭菌神经毒素L链的片段具有梭菌神经毒素L链的至少20、30、40、50、60、70、80、90、100、120、150或200个氨基酸残基。例如，梭菌神经毒素L链的片段可以具有梭菌神经毒素L链的20-400、50-300或100-200个氨基酸残基。然而，优选地，提及无催化活性梭菌神经毒素是指全长无催化活性梭菌神经毒素L链。
L链参考序列的实例包括：
肉毒A型神经毒素：氨基酸残基1-448
肉毒B型神经毒素：氨基酸残基1-440
肉毒C1型神经毒素：氨基酸残基1-441
肉毒D型神经毒素：氨基酸残基1-445
肉毒E型神经毒素：氨基酸残基1-422
肉毒F型神经毒素：氨基酸残基1-439
肉毒G型神经毒素：氨基酸残基1-441
破伤风神经毒素：氨基酸残基1-457
对于最近鉴定的BoNT/X，已报道L链对应于其氨基酸1-439，L链边界可能变化约25个氨基酸(例如1-414或1-464)。
上述确定的参考序列应视为指南，因为根据亚血清型可以存在轻微的变化。作为实例，US2007/0166332(在此以其整体引入作为参考)引用了略微不同的梭菌序列：
肉毒A型神经毒素：氨基酸残基M1-K448
肉毒B型神经毒素：氨基酸残基M1-K441
肉毒C1型神经毒素：氨基酸残基M1-K449
肉毒D型神经毒素：氨基酸残基M1-R445
肉毒E型神经毒素：氨基酸残基M1-R422
肉毒F型神经毒素：氨基酸残基M1-K439
肉毒G型神经毒素：氨基酸残基M1-K446
破伤风神经毒素：氨基酸残基M1-A457
本文描述了适宜的梭菌神经毒素L链。
梭菌神经毒素L链可以包含与SEQ ID NO:6、24、32、40、74或76中的任一个具有至少70％序列同一性的多肽序列，条件是该L链无催化活性(例如该L链已通过修饰失活)。在一个实施方案中，梭菌神经毒素L链包含与SEQ ID NO:6、24、32、40、74或76中的任一个具有至少80％、90％、95％或98％序列同一性的多肽序列，条件是该L链无催化活性(例如该L链已通过修饰失活)。优选地，梭菌神经毒素L链包含含有SEQ ID NO:6、24、32或40中的任一个的多肽序列(更优选由其组成)，其已修饰以催化失活L链，例如SEQ ID NO:74或76。
梭菌神经毒素L链可以是由与SEQ ID NO:5、23、31或39中的任一个具有至少70％序列同一性的核苷酸序列编码的梭菌神经毒素L链，条件是该L链无催化活性(例如该L链已通过修饰失活)。在一个实施方案中，梭菌神经毒素L链是由与SEQ ID NO:5、23、31或39中的任一个具有至少80％、90％、95％或98％序列同一性的核苷酸序列编码的梭菌神经毒素L链，条件是该L链无催化活性(例如该L链已通过修饰失活)。优选地，梭菌神经毒素L链是由包含SEQ ID NO:5、23、31或39中的任一个的核苷酸序列编码的梭菌神经毒素L链，其已修饰以催化失活所编码的L链。
鉴于轻链的催化活性对于治疗疼痛的功效不是必需的，因此不包含L链的多肽(或仅包含L链的片段)可以治疗疼痛是可信的。出于类似的原因，不包含L链的多肽(或仅包含L链的片段)可以治疗炎性病症是可信的。因此，在一个实施方案中，该多肽可以包含梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)。在一个实施方案中，本发明的多肽不包含梭菌神经毒素转运结构域(HN结构域)和梭菌神经毒素受体结合结构域(HC结构域)二者。
在一个实施方案中，本发明的多肽包含梭菌神经毒素重链(H链)(或由其组成)。该H链包含梭菌神经毒素转运结构域(HN结构域)和受体结合结构域(HC结构域)。本文中提及的梭菌神经毒素H链也涵盖梭菌神经毒素H链的片段。梭菌神经毒素H链的片段可以具有梭菌神经毒素H链的≤800、≤700、≤600、≤500、≤400、≤350、≤300、≤250、≤200、≤150、≤100或≤50个氨基酸残基。在一个实施方案中，梭菌神经毒素H链的片段具有梭菌神经毒素H链的至少20、30、40、50、60、70、80、90、100、120、150或200个氨基酸残基。例如，梭菌神经毒素H链的片段可以具有梭菌神经毒素H链的20-800、30-600、40-400、50-300或100-200个氨基酸残基。然而，优选地，提及H链是指全长H链。
在一个实施方案中，本发明的多肽包含梭菌神经毒素转运结构域(HN结构域)(或由其组成)。本文中提及的梭菌神经毒素转运结构域也涵盖转运结构域的片段。梭菌神经毒素转运结构域的片段可以具有≤400、≤350、≤300、≤250、≤200、≤150、≤100或≤50个氨基酸残基。在一个实施方案中，梭菌神经毒素转运结构域的片段具有梭菌神经毒素转运结构域的至少20、30、40、50、60、70、80、90、100、120、150或200个氨基酸残基。例如，梭菌神经毒素转运结构域的片段可以具有梭菌神经毒素转运结构域的20-400、50-300或100-200个氨基酸残基。然而，优选地，提及转运结构域是指全长转运结构域。
转运结构域是梭菌神经毒素H链的片段，大致相当于H链的氨基端一半，或与完整H链中的该片段相对应的结构域。在一个实施方案中，H链的HC功能可以通过HC氨基酸序列的缺失来去除(在DNA合成水平，或在合成后水平，通过核酸酶或蛋白酶处理)。备选地，HC功能可以通过化学或生物学处理来失活。因此，在一些实施方案中，H链可以不能与天然梭菌神经毒素(即全毒素)结合的靶细胞上的结合位点结合。
适宜的(参考)转运结构域的实例包括：
肉毒A型神经毒素-氨基酸残基(449-871)
肉毒B型神经毒素-氨基酸残基(441-858)
肉毒C型神经毒素-氨基酸残基(442-866)
肉毒D型神经毒素-氨基酸残基(446-862)
肉毒E型神经毒素-氨基酸残基(423-845)
肉毒F型神经毒素-氨基酸残基(440-864)
肉毒G型神经毒素-氨基酸残基(442-863)
破伤风神经毒素-氨基酸残基(458-879)
上述确定的参考序列应视为指南，因为根据亚血清型可以发生轻微的变化。作为实例，US2007/0166332(在此引入作为参考)引用了略微不同的梭菌序列：
肉毒A型神经毒素-氨基酸残基(A449-K871)
肉毒B型神经毒素-氨基酸残基(A442-S858)
肉毒C型神经毒素-氨基酸残基(T450-N866)
肉毒D型神经毒素-氨基酸残基(D446-N862)
肉毒E型神经毒素-氨基酸残基(K423-K845)
肉毒F型神经毒素-氨基酸残基(A440-K864)
肉毒G型神经毒素-氨基酸残基(S447-S863)
破伤风神经毒素-氨基酸残基(S458-V879)
在本发明的上下文中，包含转运结构域的多种梭菌神经毒素HN区可用于本发明的各方面。梭菌神经毒素重链的HN区长度约为410-430个氨基酸，并包含转运结构域。研究表明，梭菌神经毒素重链的HN区的整个长度对于转运结构域的转运活性不是必需的。因此，该实施方案的各方面可以包括这样的梭菌神经毒素HN区，其包含长度为例如至少350个氨基酸、至少375个氨基酸、至少400个氨基酸和至少425个氨基酸的转运结构域。该实施方案的其他方面可以包括这样的梭菌神经毒素HN区，其包含长度为例如至多350个氨基酸、至多375个氨基酸、至多400个氨基酸和至多425个氨基酸的转运结构域。
关于肉毒梭菌和破伤风梭菌中毒素产生的遗传基础的更多细节，参见Henderson等(1997)，The Clostridia:Molecular Biology and Pathogenesis,Academic press。
术语HN包括天然存在的神经毒素HN部分，以及具有在自然界中不存在的氨基酸序列和/或合成氨基酸残基的修饰HN部分。在一个实施方案中，该修饰HN部分仍然表现出上述转运功能。
在一个实施方案中，本发明的多肽包含梭菌神经毒素受体结合结构域(HC结构域)(或由其组成)。本文中提及的梭菌神经毒素受体结合结构域(HC)还涵盖梭菌神经毒素受体结合结构域(HC)的片段。梭菌神经毒素受体结合结构域(HC)的片段可以具有≤350、≤300、≤250、≤200、≤150、≤100或≤50个梭菌神经毒素受体结合结构域的氨基酸残基。在一个实施方案中，梭菌神经毒素受体结合结构域(HC)的片段具有梭菌神经毒素受体结合结构域的至少20、30、40、50、60、70、80、90、100、120、150或200个氨基酸残基。例如，梭菌神经毒素受体结合结构域(HC)的片段可以具有梭菌神经毒素受体结合结构域的20-350、50-300或100-200个氨基酸残基。然而，优选地，提及梭菌神经毒素受体结合结构域(HC)是指全长梭菌神经毒素受体结合结构域(HC)。
梭菌神经毒素受体结合结构域(HC)参考序列的实例包括：
BoNT/A-N872-L1296
BoNT/B-E859-E1291
BoNT/C1-N867-E1291
BoNT/D-S863-E1276
BoNT/E-R846-K1252
BoNT/F-K865-E1274
BoNT/G-N864-E1297
TeNT-I880-D1315
对于最近鉴定的BoNT/X，已报道HC结构域对应于其氨基酸893-1306，结构域边界可能变化约25个氨基酸(例如868-1306或918-1306)。
梭菌神经毒素H链(例如HC结构域部分)可进一步包含转运促进结构域(或其片段可以是转运促进结构域片段)。该结构域促进L链递送到靶细胞的胞质溶胶中，并且在例如WO 08/008803和WO 08/008805中进行了描述，其中每一个在此引入作为参考。
作为实例，转运促进结构域可以包含梭菌神经毒素HCN结构域或其片段或变体。更详细地，梭菌神经毒素HCN转运促进结构域可以具有至少200个氨基酸、至少225个氨基酸、至少250个氨基酸、至少275个氨基酸的长度。就此而言，梭菌神经毒素HCN转运促进结构域优选具有至多200个氨基酸、至多225个氨基酸、至多250个氨基酸或至多275个氨基酸的长度。具体(参考)实例包括：
肉毒A型神经毒素-氨基酸残基(872-1110)
肉毒B型神经毒素-氨基酸残基(859-1097)
肉毒C型神经毒素-氨基酸残基(867-1111)
肉毒D型神经毒素-氨基酸残基(863-1098)
肉毒E型神经毒素-氨基酸残基(846-1085)
肉毒F型神经毒素-氨基酸残基(865-1105)
肉毒G型神经毒素-氨基酸残基(864-1105)
破伤风神经毒素-氨基酸残基(880-1127)
上述序列位置可以根据血清型/亚型略有变化，适宜的(参考)梭菌神经毒素HCN结构域的其他实例包括：
肉毒A型神经毒素-氨基酸残基(874-1110)
肉毒B型神经毒素-氨基酸残基(861-1097)
肉毒C型神经毒素-氨基酸残基(869-1111)
肉毒D型神经毒素-氨基酸残基(865-1098)
肉毒E型神经毒素-氨基酸残基(848-1085)
肉毒F型神经毒素-氨基酸残基(867-1105)
肉毒G型神经毒素-氨基酸残基(866-1105)
破伤风神经毒素-氨基酸残基(882-1127)
本文描述了适宜的梭菌神经毒素HC结构域。
梭菌神经毒素HC结构域可以包含与SEQ ID NO:8、22、30、38、42、44、46、48或50中的任一个具有至少70％序列同一性的多肽序列。在一个实施方案中，梭菌神经毒素HC结构域包含与SEQ ID NO:8、22、30、38、42、44、46、48或50中的任一个具有至少80％、90％、95％或98％序列同一性的多肽序列。优选地，梭菌神经毒素HC结构域包含含有SEQ ID NO:8、22、30、38、42、44、46、48或50中的任一个的多肽序列(更优选由其组成)。
梭菌神经毒素HC结构域可以是由与SEQ ID NO:7、21、29、37、41、43、45、47或49中的任一个具有至少70％序列同一性的核苷酸序列编码的梭菌神经毒素HC结构域。在一个实施方案中，梭菌神经毒素HC结构域是由与SEQ ID NO:7、21、29、37、41、43、45、47或49中的任一个具有至少80％、90％、95％或98％序列同一性的核苷酸序列编码的梭菌神经毒素HC结构域。优选地，梭菌神经毒素HC结构域是由包含SEQ ID NO:7、21、29、37、41、43、45、47或49中的任一个的核苷酸序列编码的梭菌神经毒素HC结构域。
任何上述促进结构域都可以与任何适用于本发明的前述转运结构域肽组合。因此，作为实例，非梭菌促进结构域可以与非梭菌转运结构域肽或与梭菌转运结构域肽组合。备选地，梭菌神经毒素HCN转运促进结构域可以与非梭菌转运结构域肽组合。备选地，梭菌神经毒素HCN促进结构域可以与梭菌转运结构域肽组合，其实例包括：
肉毒A型神经毒素-氨基酸残基(449-1110)
肉毒B型神经毒素-氨基酸残基(442-1097)
肉毒C型神经毒素-氨基酸残基(450-1111)
肉毒D型神经毒素-氨基酸残基(446-1098)
肉毒E型神经毒素-氨基酸残基(423-1085)
肉毒F型神经毒素-氨基酸残基(440-1105)
肉毒G型神经毒素-氨基酸残基(447-1105)
破伤风神经毒素-氨基酸残基(458-1127)
在一些实施方案中，本发明的多肽可以缺乏梭菌神经毒素的功能性HC结构域。在一个实施方案中，该多肽优选缺乏梭菌神经毒素全毒素的最后50个C端氨基酸。在另一个实施方案中，该多肽优选缺乏梭菌神经毒素全毒素的最后100个、优选最后150个、更优选最后200个、尤其优选最后250个、最优选最后300个C端氨基酸残基。备选地，HC结合活性可以通过诱变来废除/降低－作为实例，为了方便起见，参考BoNT/A，神经节苷脂结合袋中一个或两个氨基酸残基突变(W1266到L和Y1267到F)的修饰导致HC区失去其受体结合功能。可以对非血清型A梭菌肽组分进行类似的突变，例如基于具有突变的肉毒B(W1262至L和Y1263至F)或肉毒E(W1224至L和Y1225至F)的构建体。对活性位点的其他突变实现了HC受体结合活性的相同废除，例如肉毒A型毒素中的Y1267S和其他梭菌神经毒素中相应的高度保守残基。此突变和其它突变的细节描述于Rummel等(2004)(Molecular Microbiol.51:631-634)中，其在此引入作为参考。
天然梭菌神经毒素的HC肽包含约400-440个氨基酸残基，并由两个功能不同的结构域组成，每个结构域约25kDa，即N端区(通常称为HCN肽或结构域)和C端区(通常称为HCC肽或结构域)。以下出版物确认了这一事实，每一出版物在此以其整体引入作为参考：UmlandTC(1997)Nat.Struct.Biol.4:788-792；Herreros J(2000)Biochem.J.347:199-204；Halpern J(1993)J.Biol.Chem.268:15,11188-11192页；Rummel A(2007)PNAS104:359-364；Lacey DB(1998)Nat.Struct.Biol.5:898-902；Knapp(1998)Am.Cryst.Assoc.Abstract Papers 25:90；Swaminathan和Eswaramoorthy(2000)Nat.Struct.Biol.7:1751-1759；及Rummel A(2004)Mol.Microbiol.51(3),631-643。此外，已经有充分的证据表明，构成C端160-200个氨基酸残基的C端区(HCC)负责梭菌神经毒素与其天然细胞受体的结合，即与神经肌肉接头处的神经末梢的结合－这一事实也得到了上述出版物的确认。因此，在本说明书中，谈及缺乏功能性重链HC肽(或结构域)的梭菌重链来使得该重链不能与天然梭菌神经毒素结合的细胞表面受体结合，意味着该梭菌重链只是缺乏功能性HCC肽。换言之，HCC肽区可以部分或全部缺失，或以其他方式修饰(例如通过常规化学或蛋白水解处理)，以降低其对神经肌肉接头处神经末梢的天然结合能力。
因此，在一个实施方案中，本发明的梭菌神经毒素HN肽缺乏梭菌神经毒素的C端肽部分(HCC)的一部分，因此缺乏天然梭菌神经毒素的HC结合功能。作为实例，在一个实施方案中，C端延伸的梭菌HN肽缺乏梭菌神经毒素重链的C端40个氨基酸残基、或C端60个氨基酸残基、或C端80个氨基酸残基、或C端100个氨基酸残基、或C端120个氨基酸残基、或C端140个氨基酸残基、或C端150个氨基酸残基、或C端160个氨基酸残基。在另一个实施方案中，本发明的梭菌HN肽缺乏梭菌神经毒素的整个C端肽部分(HCC)，因此缺乏天然梭菌神经毒素的HC结合功能。作为实例，在一个实施方案中，梭菌HN肽缺乏梭菌神经毒素重链的C端165个氨基酸残基、或C端170个氨基酸残基、或C端175个氨基酸残基、或C端180个氨基酸残基、或C端185个氨基酸残基、或C端190个氨基酸残基、或C端195个氨基酸残基。作为另一个实例，本发明的梭菌HN肽缺乏选自以下的梭菌HCC参考序列：
肉毒A型神经毒素-氨基酸残基(Y1111-L1296)
肉毒B型神经毒素-氨基酸残基(Y1098-E1291)
肉毒C型神经毒素-氨基酸残基(Y1112-E1291)
肉毒D型神经毒素-氨基酸残基(Y1099-E1276)
肉毒E型神经毒素-氨基酸残基(Y1086-K1252)
肉毒F型神经毒素-氨基酸残基(Y1106-E1274)
肉毒G型神经毒素-氨基酸残基(Y1106-E1297)
破伤风神经毒素-氨基酸残基(Y1128-D1315)。
上述确定的参考序列应视为指南，因为根据亚血清型可以发生轻微的变化。
在其他实施方案中，HC结构域的片段可以包含本文所述的HCC肽。
本发明的多肽可以包含无催化活性梭菌神经毒素L链和梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)。例如，多肽可以包含无催化活性梭菌神经毒素L链和梭菌神经毒素转运结构域(HN)。
本文描述了包含无催化活性梭菌神经毒素L链和转运结构域的适宜多肽。
包含梭菌神经毒素L链和转运结构域的多肽可以包含与SEQ ID NO:4、20、28、36或75中的任一个具有至少70％序列同一性的多肽序列，条件是该L链无催化活性(例如该L链已经通过修饰失活)。在一个实施方案中，包含梭菌神经毒素L链和转运结构域的多肽包含与SEQ ID NO:4、20、28、36或75中的任一个具有至少80％、90％、95％或98％序列同一性的多肽序列，条件是该L链无催化活性(例如该L链已通过修饰失活)。优选地，包含梭菌神经毒素L链和转运结构域的多肽包含含有SEQ ID NO:4、20、28、36或75中的任一个的多肽序列(更优选由其组成)，其已修饰以催化失活L链，如SEQ ID NO:75。
包含梭菌神经毒素L链和转运结构域(或由其组成)的多肽可以是由与SEQ ID NO:3、19、27或35中的任一个具有至少70％序列同一性的核苷酸序列编码的多肽，条件是该L链无催化活性(例如该L链已通过修饰失活)。在一个实施方案中，包含梭菌神经毒素L链和转运结构域(或由其组成)的多肽是由与SEQ ID NO:3、19、27或35中的任一个具有至少80％、90％、95％或98％序列同一性的核苷酸序列编码的多肽，条件是该L链无催化活性(例如该L链已通过修饰失活)。优选地，包含梭菌神经毒素L链和转运结构域(或由其组成)的多肽是由包含SEQ ID NO:3、19、27或35中的任一个的核苷酸序列编码的多肽，其已修饰以催化失活所编码的L链。
优选地，该多肽包含无催化活性梭菌神经毒素L链、梭菌神经毒素转运结构域(HN结构域)和梭菌神经毒素受体结合结构域(HC结构域)。
在一个实施方案中，本发明的多肽不包含梭菌神经毒素受体结合结构域(HC)或不包含至少梭菌神经毒素受体结合结构域的C端部分(HCC)。因此，在一个实施方案中，本发明的多肽缺乏梭菌神经毒素受体结合结构域的C端部分(HCC)。有利地，这类多肽缺乏内源性梭菌神经毒素受体结合能力，因此在施用该多肽的受试者中可以表现出较少的脱靶作用。
本发明的多肽可以基本上由无催化活性梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)组成。例如，多肽可以基本上由无催化活性梭菌神经毒素L链和梭菌神经毒素转运结构域(HN)组成。
优选地，该多肽基本上由无催化活性梭菌神经毒素L链、梭菌神经毒素转运结构域(HN结构域)和梭菌神经毒素受体结合结构域(HC结构域)组成。
本文所用的术语“基本上由…组成”是指多肽不进一步包含赋予该多肽额外的功能性的一个或多个氨基酸残基，例如在对受试者施用时。换言之，基本上由无催化活性梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)组成的多肽可以进一步包含一个或多个氨基酸残基(相对于无催化活性梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)而言)，但该一个或多个其他氨基酸残基不赋予该多肽额外的功能性，例如在对受试者施用时。额外的功能性可以包括酶活性、结合活性和/或任何生理活性。
本发明的多肽可以由无催化活性梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)组成。例如，多肽可以由无催化活性梭菌神经毒素L链和梭菌神经毒素转运结构域(HN)组成。
优选地，该多肽由无催化活性梭菌神经毒素L链、梭菌神经毒素转运结构域(HN结构域)和梭菌神经毒素受体结合结构域(HC结构域)组成。
在一个实施方案中，除了任何梭菌神经毒素序列之外，多肽还可以包含非梭菌神经毒素序列，只要该非梭菌神经毒素序列不破坏多肽达到其治疗效果(例如治疗疼痛)的能力。优选地，该非梭菌神经毒素序列不是具有催化活性例如酶活性的序列。在一个实施方案中，本发明的多肽不包含催化活性结构域(例如，不包含非梭菌催化活性结构域)。在一个实施方案中，该非梭菌序列不是与细胞受体结合的序列。换言之，在一个实施方案中，该非梭菌序列不是细胞受体的配体。细胞受体可以是蛋白质细胞受体，如整合膜蛋白。细胞受体的实例可见于可在https://www.guidetopharmacology.org/download.jsp#db_reports上获得的IUPHAR Guide to Pharmacology Database,2019.4版中。非梭菌神经毒素序列可以包括有助于纯化的标签，如His标签。在一个实施方案中，本发明的多肽不包含标记或用于添加标记的位点，如分选酶受体或供体位点。
在一个优选实施方案中，本发明的多肽不包含无催化活性梭菌神经毒素L链、HN结构域和/或HC结构域之外的额外治疗或诊断剂(例如核酸、蛋白质、肽或小分子治疗或诊断剂)。例如，在一个实施方案中，该多肽可以不包含共价或非共价结合的治疗或诊断剂。因此，本发明的多肽优选不作为另一种治疗或诊断剂的递送载体发挥作用。
本发明的多肽可以包含经修饰的梭菌神经毒素或其衍生物或经修饰的梭菌神经毒素片段或衍生物片段(或由其组成)，包括但不限于下文所述的那些，条件是存在的任何L链都无催化活性。经修饰的梭菌神经毒素或衍生物(或经修饰的梭菌神经毒素片段或衍生物片段)可以包含一个或多个与天然(未修饰)形式的梭菌神经毒素(或梭菌神经毒素片段)相比已经修饰的氨基酸，或者可以包含一个或多个不存在于天然(未修饰)形式的梭菌神经毒素(或梭菌神经毒素片段)中的插入氨基酸。作为实例，相对于天然(未修饰)梭菌神经毒素序列(或梭菌神经毒素片段)，经修饰的梭菌神经毒素(或梭菌神经毒素片段)可以在一个或多个结构域中具有经修饰的氨基酸序列。这类修饰可以修饰毒素(或毒素片段)的功能方面。因此，在一个实施方案中，本发明的多肽是或包含经修饰的梭菌神经毒素或经修饰的梭菌神经毒素衍生物，或梭菌神经毒素衍生物或经修饰的梭菌神经毒素片段或衍生物片段(例如无催化活性L链、HN结构域和/或HC结构域)，条件是存在的任何L链都无催化活性。
本发明的多肽可以包含在重链的氨基酸序列中具有一个或多个修饰的经修饰的梭菌神经毒素或梭菌神经毒素片段(例如HC结构域)(或由其组成)(如经修饰的HC结构域)，其中该经修饰的重链以比天然(未修饰)梭菌神经毒素或梭菌神经毒素片段更高或更低的亲和力结合靶神经细胞，条件是存在的任何L链都无催化活性。HC结构域中的这类修饰可以包括修饰HC结构域的神经节苷脂结合位点中或蛋白质(SV2或突触结合蛋白(synaptotagmin))结合位点中的残基，其改变与靶神经细胞的神经节苷酯受体和/或蛋白质受体的结合。这类经修饰的梭菌神经毒素的实例描述于WO 2006/027207和WO 2006/114308中，二者在此以其整体引入作为参考。
经修饰的梭菌神经毒素(或梭菌神经毒素片段)可以是这样的，与缺乏该一种或多种修饰的等同的未修饰梭菌神经毒素(或梭菌神经毒素片段)相比，其包含一种或多种提高该梭菌神经毒素的等电点的修饰，条件是存在的任何L链都无催化活性。适宜的经修饰的梭菌神经毒素(条件是存在的任何L链已修饰为无催化活性)描述于下文以及WO 2015/004461A1和WO 2016/110662 A1中，其在此引入作为参考。示例性序列包括本文所述的SEQ ID NO:42和62。
在一个实施方案中，本发明的多肽可以包含经修饰的BoNT/A或其片段(例如BoNT/A HC结构域或其片段)。该经修饰的BoNT/A或其片段可以是在选自以下的一个或多个氨基酸残基处包含修饰的经修饰的BoNT/A或其片段：ASN 886、ASN 905、GLN 915、ASN 918、GLU920、ASN 930、ASN 954、SER 955、GLN 991、GLU 992、GLN 995、ASN 1006、ASN 1025、ASN1026、ASN 1032、ASN 1043、ASN 1046、ASN 1052、ASP 1058、HIS1064、ASN 1080、GLU 1081、GLU 1083、ASP 1086、ASN 1188、ASP 1213、GLY 1215、ASN 1216、GLN 1229、ASN1242、ASN1243、SER 1274和THR 1277。
该修饰可以是与SEQ ID NO:2所示的无催化活性BoNT/A相比的修饰，其中氨基酸残基编号通过与SEQ ID NO：2比对来确定。由于在SEQ ID NO:2(以及对应于本文所述的经修饰的BoNT/A多肽或其片段的SEQ ID NO)的位置1处甲硫氨酸残基的存在是任选的，本领域技术人员在确定氨基酸残基编号时将考虑甲硫氨酸残基的存在/不存在。例如，在SEQ IDNO:2包括甲硫氨酸的情况下，该位置编号将如上所定义(例如ASN 886将是SEQ ID NO:2的ASN 886)。备选地，在SEQ ID NO:2中不存在甲硫氨酸的情况下，该氨基酸残基编号应改为-1(例如ASN 886将是SEQ ID NO:2的ASN 885)。在本文所述的其他多肽序列的位置1处的甲硫氨酸存在/不存在时，类似的考虑适用，并且本领域技术人员将使用本领域常规技术容易地确定正确的氨基酸残基编号。
本文所述用于确定氨基酸残基编号的比对可以使用本文所述的用于确定序列同源性和/或％序列同一性的任何方法进行。
上文所示的用于修饰的氨基酸残基是表面暴露的氨基酸残基。
经修饰的BoNT/A或其片段可以在选自以下的一个或多个氨基酸残基处包含修饰：ASN 886、ASN 930、ASN 954、SER 955、GLN 991、ASN 1025、ASN 1026、ASN 1052、ASN 1188、ASP 1213、GLY 1215、ASN 1216、GLN 1229、ASN 1242、ASN 1243、SER 1274和THR 1277。
在经修饰的BoNT/A或其片段的上下文中使用时，术语“一个或多个氨基酸残基”优选指至少2、3、4、5、6或7个所示的氨基酸残基。因此，经修饰的BoNT/A或其片段可以在所示的氨基酸残基处包含至少2、3、4、5、6或7个(优选7个)修饰。经修饰的BoNT/A或其片段可以包含1-30、3-20或5-10个氨基酸修饰。更优选地，在经修饰的BoNT/A或其片段的上下文中使用时，术语“一个或多个氨基酸残基”是指所有所示的氨基酸残基。
优选地，除了所示的氨基酸残基处的一个或多个氨基酸修饰之外，与SEQ ID NO:2相比时，该经修饰的BoNT/A或其片段不包含任何其他氨基酸修饰。
该修饰可以选自：
i.用碱性氨基酸残基取代表面暴露的酸性氨基酸残基；
ii.用不带电荷的氨基酸残基取代表面暴露的酸性氨基酸残基；
iii.用碱性氨基酸残基取代表面暴露的不带电荷的氨基酸残基；
iv.插入碱性氨基酸残基；和
v.缺失表面暴露的酸性氨基酸残基。
如上所述的修饰导致经修饰的BoNT/A或其片段，其与相应的未修饰BoNT/A或其片段相比具有增加的表面正电荷和提高的等电点。
等电点(pI)是给定蛋白质的一种特定性质。如本领域中众所周知，蛋白质由氨基酸(在蛋白质中也称为氨基酸残基)的特定序列组成。二十种标准组的每种氨基酸都具有不同的侧链(或R基团)，这意味着蛋白质中的每个氨基酸残基都显示出不同的化学性质，如电荷和疏水性。这些性质可以受到周围化学环境的影响，如温度和pH。蛋白质的整体化学特征将取决于这些多种因素的总和。
某些氨基酸残基(详见下文)具有可电离的侧链，其可以取决于周围pH显示电荷。这种侧链在给定pH下是否带电取决于相关可电离部分的pKa，其中pKa是来自共轭碱的特定质子的酸离解常数(Ka)的负对数。
例如，酸性残基如天冬氨酸和谷氨酸具有pKa值约为4.1的侧链羧酸基团(精确的pKa值可以取决于温度、离子强度和可电离基团的微环境)。因此，这些侧链在7.4的pH(通常称为“生理pH”)下显示出负电荷。在低pH值下，这些侧链将质子化并失去其电荷。
相反，碱性残基如赖氨酸和精氨酸具有pKa值约为10-12的含氮侧链基团。因此，这些侧链在7.4的pH下显示出正电荷。在高pH值下，这些侧链将去质子化并失去其电荷。
因此，蛋白质分子的总(净)电荷取决于蛋白质中存在的酸性和碱性残基的数量(及其表面暴露程度)以及周围的pH。改变周围的pH会改变蛋白质的总电荷。因此，对于每种蛋白质，都存在一个给定的pH，在该pH下正电荷和负电荷的数量相等，并且蛋白质不显示出总净电荷。这个点称为等电点(pI)。等电点是本领域技术人员熟悉的蛋白质生物化学中的标准概念。
因此，等电点(pI)定义为蛋白质显示零净电荷的pH值。pI的增加意味着蛋白质需要更高的pH值才能显示零净电荷。因此，pI的增加表示在给定pH下蛋白质的净正电荷的增加。相反，pI的降低意味着蛋白质需要更低的pH值才能显示零净电荷。因此，pI的降低表示在给定pH下蛋白质的净正电荷的减少。
测定蛋白质pI的方法是本领域已知的，并且对于技术人员来说是熟悉的。作为实例，蛋白质的pI可以根据蛋白质中存在的每个氨基酸的平均pKa值来计算(“计算pI”)。这类计算可以使用本领域已知的计算机程序来进行，例如来自ExPASy的Compute pI/MW Tool(https://web.expasy.org/compute_pi/)，这是根据本发明计算pI的优选方法。应使用相同的计算技术/程序对不同分子之间的pI值进行比较。
根据需要，可以使用等电聚焦技术(“观测pI”)通过实验来确认蛋白质的计算等电点。该技术使用电泳根据蛋白质的pI来分离蛋白质。等电聚焦通常使用具有固定pH梯度的凝胶进行。在施加电场时，蛋白质通过pH梯度迁移，直到达到净电荷为零的pH，这一点是蛋白质的pI。由等电聚焦提供的结果本质上通常是相对低的分辨率，因此本发明人认为由计算pI(如上所述)提供的结果更适合于使用。
在本说明书中，除非另有说明，否则“pI”是指“计算pI”。
蛋白质的pI可以通过改变其表面上展示的碱性和/或酸性基团的数量来增加或减少。这可以通过修饰蛋白质的一个或多个氨基酸来达到。例如，pI的增加可以通过减少酸性残基的数量或通过增加碱性残基的数量来提供。
本发明的经修饰的BoNT/A或其片段的pI值可以比无催化活性的BoNT/A(例如SEQID NO:2)或其片段的pI值高至少0.2、0.4、0.5或1个pI单位。优选地，经修饰的BoNT/A或其片段可以具有至少6.6，例如至少6.8的pI。
20种标准氨基酸的性质如下表所示：
氨基酸 侧链
天冬氨酸 Asp D 带电荷(酸性)
谷氨酸 Glu E 带电荷(酸性)
精氨酸 Arg R 带电荷(碱性)
赖氨酸 Lys K 带电荷(碱性)
组氨酸 His H 不带电荷(极性)
天冬酰胺 Asn N 不带电荷(极性)
谷氨酰胺 Gln Q 不带电荷(极性)
丝氨酸 Ser S 不带电荷(极性)
苏氨酸 Thr T 不带电荷(极性)
酪氨酸 Tyr Y 不带电荷(极性)
甲硫氨酸 Met M 不带电荷(极性)
色氨酸 Trp W 不带电荷(极性)
半胱氨酸 Cys C 不带电荷(极性)
丙氨酸 Ala A 不带电荷(疏水)
甘氨酸 Gly G 不带电荷(疏水)
缬氨酸 Val V 不带电荷(疏水)
亮氨酸 Leu L 不带电荷(疏水)
异亮氨酸 Ile I 不带电荷(疏水)
脯氨酸 Pro P 不带电荷(疏水)
苯丙氨酸 Phe F 不带电荷(疏水)
认为以下氨基酸是带电荷氨基酸：天冬氨酸(负)、谷氨酸(负)、精氨酸(正)和赖氨酸(正)。
在7.4的pH下，天冬氨酸(pKa 3.1)和谷氨酸(pKa 4.1)的侧链具有负电荷，而精氨酸(pKa 12.5)和赖氨酸(pKa 10.8)的侧链具有正电荷。将天冬氨酸和谷氨酸称为酸性氨基酸残基。将精氨酸和赖氨酸称为碱性氨基酸残基。
认为以下氨基酸是不带电荷的极性氨基酸(意味着它们可以参与氢键)：天冬酰胺、谷氨酰胺、组氨酸、丝氨酸、苏氨酸、酪氨酸、半胱氨酸、甲硫氨酸和色氨酸。
认为以下氨基酸是不带电荷的疏水性氨基酸：丙氨酸、缬氨酸、亮氨酸、异亮氨酸、苯丙氨酸、脯氨酸和甘氨酸。
在氨基酸插入中，将额外氨基酸残基(通常不存在的氨基酸残基)掺入BoNT/A多肽序列或其片段中，从而增加该序列中氨基酸残基的总数。在氨基酸缺失中，从梭菌毒素氨基酸序列中去除氨基酸残基，从而减少该序列中氨基酸残基的总数。
优选地，该修饰是取代，其有利地在经修饰的BoNT/A或其片段中保持相同数量的氨基酸残基。在氨基酸取代中，用不同的氨基酸残基替换形成BoNT/A多肽序列或其片段的一部分的氨基酸残基。替换氨基酸残基可以是上述20种标准氨基酸之一。备选地，氨基酸取代中的替换氨基酸可以是非标准氨基酸(不属于上述20种标准组的一部分的氨基酸)。作为实例，替换氨基酸可以是碱性非标准氨基酸，例如L-鸟氨酸、L-2-氨基-3-胍丙酸、或赖氨酸、精氨酸和鸟氨酸的D-异构体)。用于将非标准氨基酸引入蛋白质的方法是本领域已知的，并且包括使用大肠杆菌(E.coli)营养缺陷型表达宿主的重组蛋白质合成。
在一个实施方案中，该取代选自：用碱性氨基酸残基取代酸性氨基酸残基，用不带电荷的氨基酸残基取代酸性氨基酸残基，以及用碱性氨基酸残基取代不带电荷的氨基酸残基。在一个实施方案中，其中该取代是用不带电荷的氨基酸残基取代酸性氨基酸残基，用其相应的不带电荷的酰胺氨基酸残基替换酸性氨基酸残基(即用天冬酰胺替换天冬氨酸，用谷氨酰胺替换谷氨酸)。
优选地，该碱性氨基酸残基是赖氨酸残基或精氨酸残基。换言之，该取代是用赖氨酸或精氨酸取代。最优选地，该修饰是用赖氨酸取代。
优选地，用于本发明的经修饰的BoNT/A或其片段包含位于梭菌毒素HCN结构域中的4至40个氨基酸修饰。该经修饰的BoNT/A或其片段优选也具有至少6.6的pI。该经修饰的BoNT/A优选包含选自ASN 886、ASN 930、ASN 954、SER 955、GLN 991、ASN 1025、ASN 1026和ASN 1052的至少4个氨基酸的修饰，其中该修饰包括用赖氨酸残基或精氨酸残基取代该氨基酸。例如，该经修饰的BoNT/A或其片段可以包含选自ASN 886、ASN 930、ASN 954、SER955、GLN 991、ASN 1025、ASN 1026、ASN 1052和GLN 1229的至少5个氨基酸的修饰，其中该修饰包含用赖氨酸残基或精氨酸残基取代氨基酸。
用于通过氨基酸残基的取代、插入或缺失来修饰蛋白质的方法是本领域已知的。作为实例，可以通过修饰编码多肽(例如编码未修饰的BoNT/A或其片段)的DNA序列来引入氨基酸修饰。这可以使用标准分子克隆技术来达到，例如通过定点诱变，其中使用编码所希望的氨基酸的短链DNA(寡核苷酸)通过聚合酶来替换原始编码序列，或者通过用多种酶(例如连接酶和限制性内切核酸酶)插入/缺失基因的部分来实现修饰。备选地，可以化学合成经修饰的基因序列。
在一个实施方案中，根据本发明使用的多肽包含与SEQ ID NO:42具有至少70％序列同一性的多肽序列和/或由与SEQ ID NO:41具有至少70％序列同一性的核苷酸序列编码的多肽序列。在一个实施方案中，根据本发明使用的多肽包含与SEQ ID NO:42具有至少80％、90％、95％或98％序列同一性的多肽序列。优选地，根据本发明使用的多肽包含如SEQID NO:42所示的多肽序列。在一个实施方案中，根据本发明使用的多肽包含由与SEQ IDNO:41具有至少80％、90％、95％或98％序列同一性的核苷酸序列编码的多肽序列。优选地，根据本发明使用的多肽包含由SEQ ID NO:41所示的核苷酸序列编码的多肽序列。
在一个实施方案中，根据本发明使用的多肽包含与SEQ ID NO:62具有至少70％序列同一性的多肽序列。在一个实施方案中，根据本发明使用的多肽包含与SEQ ID NO:62具有至少80％、90％、95％或98％序列同一性的多肽序列。优选地，根据本发明使用的多肽包含如SEQ ID NO:62所示的多肽序列(更优选由其组成)。
SEQ ID NO:42是经修饰的BoNT/A片段的实例，SEQ ID NO:62是无催化活性的经修饰的BoNT/A多肽的实例。这类经修饰的BoNT/A多肽和片段尤其优选用于本发明。与野生型BoNT/A相比，SEQ ID NO:42和62所示的多肽具有许多氨基酸修饰(例如取代)，其增加了多肽的等电点。不希望受限于理论，认为增加的净正电荷促进多肽和阴离子细胞外组分之间的静电相互作用，从而促进多肽和细胞表面之间的结合，从而增加在施用部位的保留和/或作用持续时间。因此，可以设想，与缺乏该修饰的等同多肽相比，使用SEQ ID NO:42和62将改善治疗。
在一个实施方案中，包含与SEQ ID NO:42或62具有至少70％序列同一性的多肽序列和/或包含由与SEQ ID NO:41具有至少70％序列同一性的核苷酸序列编码的多肽序列的多肽在位置930、955、991、1026、1052、1229和886中的一个或多个位置(优选两个或多个、三个或多个、四个或更多、五个或多个或六个或多个，更优选全部)处包含取代。
优选地，包含与SEQ ID NO:42或62具有至少70％序列同一性的多肽序列和/或包含由与SEQ ID NO:41具有至少70％序列同一性的核苷酸序列编码的多肽序列的多肽在位置930、955、991、1026、1052、1229和886中的一个或多个位置处包含赖氨酸或精氨酸(更优选赖氨酸)。在一个实施方案中，该多肽在位置930、955、991、1026、1052、1229和886中的至少两个、三个、四个、五个、六个或全部位置处包含赖氨酸或精氨酸(更优选赖氨酸)。最优选地，该多肽在位置930、955、991、1026、1052、1229和886中的所有位置处包含赖氨酸或精氨酸(更优选赖氨酸)。
在一个实施方案中，用于本发明的梭菌神经毒素HC结构域是经修饰的BoNT/A HC结构域，其包含选自Y1117、F1252、H1253和L1278的一个或多个氨基酸残基的修饰。例如，经修饰的BoNT/A HC结构域可以包含一种或多种(优选两种或多种)以下修饰Y1117V、F1252Y、H1253K和L1278F或L1278H。
在一个实施方案中，经修饰的BoNT/A HC结构域包含以下修饰：Y1117V和H1253K；或Y1117V、F1252Y、H1253K和L1278F；或Y1117V、F1252Y、H1253K和L1278H。
优选地，经修饰的BoNT/A HC结构域包含以下修饰：Y1117V和H1253K；或Y1117V、F1252Y、H1253K和L1278H。
该修饰可以是与SEQ ID NO:2所示的无催化活性BoNT/A相比的修饰，其中氨基酸残基编号通过与SEQ ID NO:2比对来确定。由于在SEQ ID NO:2的位置1处甲硫氨酸残基的存在是任选的，本领域技术人员在确定氨基酸残基编号时将考虑甲硫氨酸残基的存在/不存在。例如，在SEQ ID NO:2包含甲硫氨酸的情况下，位置编号将如上所定义(例如Y1117将与SEQ ID NO:2的Y1117比对)。备选地，在SEQ ID NO:2中不存在甲硫氨酸的情况下，氨基酸残基编号应改为-1(例如Y1117将与SEQ ID NO:2的Y1116比对)。在本文所述的其他多肽序列的位置1处的甲硫氨酸存在/不存在时，类似的考虑适用，并且本领域技术人员将使用本领域常规技术容易地确定正确的氨基酸残基编号。
经修饰的BoNT/A HC结构域可以包含与SEQ ID NO:46、48或50中的任一个具有至少70％序列同一性的多肽序列，条件是该经修饰的BoNT/A HC结构域包含如上所述的修饰。在一个实施方案中，经修饰的BoNT/A HC结构域包含与SEQ ID NO:46、48或50中的任一个具有至少80％、90％、95％或98％序列同一性的多肽序列，条件是该经修饰的BoNT/A HC结构域包含如上所述的修饰。在一个实施方案中，经修饰的BoNT/A HC结构域包含与SEQ ID NO:46、48或50中的任一个具有至少99％或99.9％序列同一性的多肽序列，条件是该经修饰的BoNT/A HC结构域包含如上所述的修饰。优选地，修饰的BoNT/A HC结构域包含(更优选由)多肽序列，所述多肽序列包含SEQ ID NO:46、48或50中的任一个。
修饰的BoNT/A HC结构域可以包含与SEQ ID NO:46或50中的任一个具有至少70％序列同一性的多肽序列，条件是修饰的BoNT/A HC结构域包含如上所述的修饰。在一个实施方案中，修饰的BoNT/A HC结构域包含与SEQ ID NO:46或50中的任一个具有至少80％、90％、95％或98％序列同一性的多肽序列，条件是修饰的BoNT/A HC结构域包含如上所述的修饰。在一个实施方案中，修饰的BoNT/A HC结构域包含与SEQ ID NO:46或50中的任一个具有至少99％或99.9％序列同一性的多肽序列，条件是该经修饰的BoNT/A HC结构域包含如上所述的修饰。优选地，经修饰的BoNT/A HC结构域包含含有SEQ ID NO:46或50中的任一个的多肽序列(更优选由其组成)。
经修饰的BoNT/A HC结构域可以是由与SEQ ID NO:45、47或49中的任一个具有至少70％序列同一性的核苷酸序列编码的结构域，条件是该经修饰的BoNT/A HC结构域包含如上所述的修饰。在一个实施方案中，经修饰的BoNT/A HC结构域是由与SEQ ID NO:45、47或49中的任一个具有至少80％、90％、95％或98％序列同一性的核苷酸序列编码的结构域，条件是该经修饰的BoNT/A HC结构域包含如上所述的修饰。在一个实施方案中，经修饰的BoNT/A HC结构域是由与SEQ ID NO:45、47或49中的任一个具有至少99％或99.9％序列同一性的核苷酸序列编码的结构域，条件是该经修饰的BoNT/A HC结构域包含如上所述的修饰。优选地，经修饰的BoNT/A HC结构域是由SEQ ID NO:45、47或49中的任一个编码的结构域。
经修饰的BoNT/A HC结构域可以是由与SEQ ID NO:45或49中的任一个具有至少70％序列同一性的核苷酸序列编码的结构域，条件是该经修饰的BoNT/A HC结构域包含如上所述的修饰。在一个实施方案中，经修饰的BoNT/A HC结构域是由与SEQ ID NO:45或49中的任一个具有至少80％、90％、95％或98％序列同一性的核苷酸序列编码的结构域，条件是该经修饰的BoNT/A HC结构域包含如上所述的修饰。在一个实施方案中，经修饰的BoNT/A HC结构域是由与SEQ ID NO:45或49中的任一个具有至少99％或99.9％序列同一性的核苷酸序列编码的结构域，条件是该经修饰的BoNT/A HC结构域包含如上所述的修饰。优选地，经修饰的BoNT/A HC结构域是由SEQ ID NO:45或49中的任一个编码的结构域。
本发明的多肽可以包含杂合或嵌合梭菌神经毒素(或杂合或嵌合梭菌神经毒素的片段)(或由其组成)，条件是该L链无催化活性(在存在L链时)。一种杂合梭菌神经毒素包含来自一种梭菌神经毒素或其亚型的轻链的至少一部分，以及来自另一种梭菌神经毒素或梭菌神经毒素亚型的重链的至少一部分。在一个实施方案中，该杂合梭菌神经毒素可以包含来自一种梭菌神经毒素亚型的轻链的整个轻链和来自另一种梭菌神经毒素亚型的重链，条件是该L链无催化活性(在存在L链时)。在另一个实施方案中，嵌合梭菌神经毒素可以包含一种梭菌神经毒素亚型的重链的一部分(例如结合结构域)，重链的另一部分来自另一种梭菌神经毒素亚型。类似地或备选地，治疗元件可以包含来自不同梭菌神经毒素的轻链部分，条件是该L链无催化活性(在存在L链时)。这种杂合或嵌合梭菌神经毒素是有用的，例如，作为将这种梭菌神经毒素的治疗益处递送给对给定的梭菌神经毒素亚型具有免疫抗性的受试者、对给定的梭菌神经毒素重链结合结构域可以具有低于平均值的受体浓度的受试者、或可以具有膜或小泡毒素底物(例如SNAP-25、VAMP和突触融合蛋白)的蛋白酶抗性变体的受试者的手段。杂合和嵌合梭菌神经毒素描述于US 8071110中，该出版物在此以其整体引入作为参考。因此，在一个实施方案中，本发明的多肽是或包含杂合梭菌神经毒素或嵌合梭菌神经毒素，条件是该L链无催化活性。
在一个尤其优选的实施方案中，本发明的多肽可以是嵌合梭菌神经毒素，其包含无催化活性BoNT/A轻链和转运结构域(LHN结构域)以及BoNT/B受体结合结构域(HC结构域)或其一部分(优选由其组成)。适宜的嵌合和/或杂合梭菌神经毒素可以是WO 2017/191315A1中教导的梭菌神经毒素，其在此引入作为参考，条件是该L链无催化活性(例如已通过修饰失活)。这类优选序列包括SEQ ID NO:44和61。
无催化活性BoNT/A LHN结构域可以共价连接到BoNT/B HC结构域。该嵌合BoNT/A在本文中也称为“BoNT/AB”或“BoNT/AB嵌合体”。
LHN结构域的C端氨基酸残基可以对应于将BoNT/A的LHN和HC结构域分开的310螺旋的第一氨基酸残基，并且HC结构域的N端氨基酸残基可对应于在BoNT/B中将LHN与HC结构域分开的310螺旋的第二氨基酸残基。
本文中提及的“将BoNT/A的LHN和HC结构域分开的310螺旋的第一氨基酸残基”是指将LHN和HC结构域分开的310螺旋的N端残基。
本文中提及的“将BoNT/B的LHN和HC结构域分开的310螺旋的第二氨基酸残基”是指在将LHN和HC结构域分开的310螺旋的N端残基之后的氨基酸残基。
“310螺旋”是连同α-螺旋、β-片层和反转(reverse turn)一起见于蛋白质和多肽中，是一类二级结构。310螺旋中的氨基酸排列成右手螺旋结构，其中每一整转由三个残基和十个原子形成，这些残基和原子分开它们之间的分子内氢键。每个氨基酸对应于螺旋中的120°转弯(即，螺旋每转具有三个残基)，沿着螺旋轴平移(＝0.2nm)，并且在通过产生氢键形成的环中有10个原子。最重要的是，氨基酸的N-H基团与之前的三个残基的氨基酸的C＝O基团形成氢键；这个重复的i+3→i氢键定义了310螺旋。310螺旋是本领域技术人员熟悉的结构生物学中的标准概念。
该310螺旋对应于形成实际螺旋的四个残基和两个帽(或过渡)残基，在这四个残基每端有一个所述帽残基。本文所用的术语“将LHN和HC结构域分开的310螺旋”由这6个残基组成。
通过进行结构分析和序列比对，鉴定出将LHN和HC结构域分开的310螺旋。该310螺旋在其N端(即在LHN结构域的C端部分)由α-螺旋包围，在其C端(即在HC结构域的N端部分)由β-链包围。310螺旋的第一(N端)残基(帽或过渡残基)也对应于该α-螺旋的C端残基。
将LHN和HC结构域分开的310螺旋可以例如从肉毒神经毒素的公开可获得的晶体结构确定，例如3BTA(http://www.rcsb.org/pdb/explore/explore.do？structureId＝3BTA)和1EPW(http://www.rcsb.org/pdb/explore/explore.do？structureId＝1EPW)分别用于肉毒神经毒素A1和B1。
公开可用的计算机建模和比对工具也可用于确定将其他神经毒素中的LHN和HC结构域分开的310螺旋的位置，例如同源建模服务器LOOPP(Learning,Observing andOutputting Protein Patterns,http://loopp.org)、PHYRE(Protein Homology/analogYRecognition Engine,http://www.sbg.bio.ic.ac.uk/phyre2/)和Rosetta(https://www.rosettacommons.org/)，蛋白质叠加服务器SuperPose(http://wishart.biology.ualberta.ca/superpose/)，比对程序Clustal Omega(http://www.clustal.org/omega/)，以及分子和细胞生物学家互联网资源(Internet Resourcesfor Molecular and Cell Biologists)上列出的许多其他工具/服务(http://molbiol-tools.ca/)。特别地，“HN/HCN”连接处周围的区域在结构上高度保守，这使其成为叠加不同血清型的理想区域。
例如，以下方法可用于确定其他神经毒素中该310螺旋的序列：
1.用结构同源性建模工具LOOP(http://loopp.org)来获得基于BoNT/A1晶体结构(3BTA.pdb)的其他BoNT血清型的预测结构；
2.编辑如此获得的结构(pdb)文件以仅包括HCN结构域的N端及其之前的约80个残基(它们是HN结构域的一部分)，从而保留结构上高度保守的“HN/HCN”区域；
3.用蛋白质叠加服务器SuperPose(http://wishart.biology.ualberta.ca/superpose/)来将每个血清型叠加到3BTA.pdb结构上；
4.检查叠加的pdb文件以定位BoNT/A1 HC结构域起始处的310螺旋，然后鉴定其他血清型中的相应残基；
5.用Clustal Omega比对其他BoNT血清型序列，以检查相应的残基是否正确。
通过该方法确定的LHN、HC和310螺旋结构域的实例如下：
使用结构分析和序列比对，发现将LHN和HC结构域分开的310螺旋之后的β-链是所有肉毒和破伤风神经毒素中的保守结构，并且在从将LHN与HC结构域分开的310螺旋的第1个残基开始计数时，在第8个残基处开始(例如，在BoNT/A1的残基879处)。
BoNT/AB嵌合体可以包含与来自BoNT/B的HC结构域共价连接的来自BoNT/A的LHN结构域(具有无催化活性的L链)，
·其中LHN结构域的C端氨基酸残基对应于位于BoNT/A HC结构域的起始(N端)处的β-链N端第八氨基酸残基，以及
·其中HC结构域的N端氨基酸残基对应于位于BoNT/B的HC结构域起始(N端)处的β-链N端第七氨基酸残基。
BoNT/AB嵌合体可以包含与来自BoNT/B的HC结构域共价连接的来自BoNT/A的LHN结构域(具有无催化活性的L链)，
·其中LHN结构域的C端氨基酸残基对应于位于BoNT/A LHN结构域结束(C端)处的α-螺旋的C端氨基酸残基，以及
·其中HC结构域的N端氨基酸残基对应于位于BoNT/B LHN结构域结束(C端)处的α-螺旋的C端氨基酸残基的紧挨C端的氨基酸残基。
BoNT/AB嵌合体设计过程的基本原理是试图确保二级结构不受影响，从而最大限度地减少三级结构的任何变化。不希望受限于理论，假定通过不破坏BoNT/AB嵌合体中310螺旋的四个中心氨基酸残基，确保了嵌合神经毒素的最佳构象。
来自BoNT/A的无催化活性LHN结构域可以对应于SEQ ID NO:2或61的氨基酸残基1-872，或与之具有至少70％序列同一性的多肽序列。来自BoNT/A的无催化活性LHN结构域可以对应于SEQ ID NO:2或61的氨基酸残基1-872，或与之具有至少80％、90％或95％序列同一性的多肽序列。优选地，来自BoNT/A的无催化活性LHN结构域对应于SEQ ID NO:2或61的氨基酸残基1-872。
来自BoNT/B的HC结构域可以对应于SEQ ID NO:52的氨基酸残基860至1291，或与之具有至少70％序列同一性的多肽序列。来自BoNT/B的HC结构域可以对应于SEQ ID NO:52的氨基酸残基860至1291，或与之具有至少80％、90％或95％序列同一性的多肽序列。优选地，来自BoNT/B的HC结构域对应于SEQ ID NO:52的氨基酸残基860-1291。
优选地，无催化活性LHN结构域对应于BoNT/A(SEQ ID NO:2或61)的氨基酸残基1-872，HC结构域对应于BoNT/B(SEQ ID NO:52)的氨基酸残基860-1291。
优选地，BoNT/B HC结构域在HCC亚结构域中进一步包含至少一个氨基酸残基取代、添加或缺失，与天然BoNT/B序列相比，其具有增加BoNT/B神经毒素对人Syt II的结合亲和力的作用。BoNT/B HCC亚结构域中适宜的氨基酸残基取代、添加或缺失已在WO 2013/180799和WO 2016/154534中公开(均在此引入作为参考)。
BoNT/B HCC亚结构域中适宜的氨基酸残基取代、添加或缺失包括选自以下的取代突变：V1118M；Y1183M；E1191M；E1191I；E1191Q；E1191T；S1199Y；S1199F；S1199L；S1201V；E1191C、E1191V、E1191L、E1191Y、S1199W、S1199E、S1199H、W1178Y、W1178Q、W1178A、W1178S、Y1183C、Y1183P及其组合。
BoNT/B HCC亚结构域中适宜的氨基酸残基取代、添加或缺失还包括选自以下的两个取代突变的组合：E1191M和S1199L、E1191M和S1199Y、E1191M和S1199F、E1191Q和S1199L、E1191Q和S1199Y、E1191Q和S1199F、E1191M和S1199W、E1191M和W1178Q、E1191C和S1199W、E1191C和S1199Y、E1191C和W1178Q、E1191Q和S1199W、E1191V和S1199W、E1191V和S1199Y、或E1191V和W1178Q。
BoNT/B HCC亚结构域中适宜的氨基酸残基取代、添加或缺失还包括E1191M、S1199W和W1178Q三个取代突变的组合。
优选地，BoNT/B HCC亚结构域中适宜的氨基酸残基取代、添加或缺失包括E1191M和S1199Y两个取代突变的组合。
该修饰可以是与SEQ ID NO:52所示的未修饰BoNT/B相比的修饰，其中氨基酸残基编号通过与SEQ ID NO:52比对来确定。由于在SEQ ID NO:52的位置1处甲硫氨酸残基的存在是任选的，本领域技术人员在确定氨基酸残基编号时将考虑甲硫氨酸残基的存在/不存在。例如，在SEQ ID NO:52包含甲硫氨酸的情况下，位置编号将如上所定义(例如E1191将是SEQ ID NO:52的E1191)。备选地，在SEQ ID NO:52中不存在甲硫氨酸的情况下，氨基酸残基编号应改为-1(例如E1191将是SEQ ID NO:52的E1190)。在本文所述的其他多肽序列的位置1处的甲硫氨酸存在/不存在时，类似的考虑适用，并且本领域技术人员将使用本领域常规技术容易地确定正确的氨基酸残基编号。
在一个实施方案中，根据本发明使用的多肽包含与SEQ ID NO:61具有至少70％序列同一性的多肽序列。在一个实施方案中，根据本发明使用的多肽包含与SEQ ID NO:61具有至少80％、90％、95％或98％序列同一性的多肽序列。优选地，根据本发明使用的多肽包含如SEQ ID NO:61所示的多肽序列(更优选由其组成)。
用于本发明的嵌合和/或杂合梭菌神经毒素可以包含BoNT/A多肽的一部分和BoNT/B多肽的一部分，其实例包括本文中描述为SEQ ID NO:44的多肽。
在一个实施方案中，根据本发明使用的多肽包含与SEQ ID NO:44具有至少70％序列同一性的多肽序列和/或由与SEQ ID NO:43具有至少70％序列同一性的核苷酸序列编码的多肽序列。在一个实施方案中，根据本发明使用的多肽包含与SEQ ID NO:44具有至少80％、90％、95％或98％序列同一性的多肽序列。优选地，根据本发明使用的多肽包含如SEQID NO:44所示的多肽序列。在一个实施方案中，根据本发明使用的多肽包含由与SEQ IDNO:43具有至少80％、90％、95％或98％序列同一性的核苷酸序列编码的多肽序列。优选地，根据本发明使用的多肽包含由SEQ ID NO:43所示的核苷酸序列编码的多肽序列。
适宜的嵌合梭菌神经毒素可以包括BoNT/FA，条件是存在的任何L链都无催化活性。因此，本发明的多肽可以包含BoNT/FA或其片段，条件是存在的任何L链都无催化活性。无催化活性形式的BoNT/FA在本文中描述为SEQ ID NO:26和34。BoNT/FA的适宜片段在本文中也描述为SEQ ID NO:28、30和32。
在另一个优选实施方案中，本发明的多肽可以是嵌合梭菌神经毒素，其包含无催化活性BoNT/X轻链和转运结构域(LHN结构域)以及来自不同(即，非BoNT/X)梭菌神经毒素的受体结合结构域(HC结构域)或其一部分。适宜的嵌合和/或杂合梭菌神经毒素可以是WO2020/065336A1中教导的神经毒素，其在此引入作为参考，条件是该L链无催化活性(例如已经通过修饰失活)。这类优选序列包括本文所述的SEQ ID NO:63-70。
嵌合梭菌神经毒素可以包含无催化活性BoNT/X轻链和转运结构域(LHN结构域)，以及：
(i)BoNT/A受体结合结构域(HC结构域)或其一部分；或
(ii)BoNT/B受体结合结构域(HC结构域)或其一部分；或
(iii)BoNT/C受体结合结构域(HC结构域)或其一部分；或
(iv)BoNT/D受体结合结构域(HC结构域)或其一部分；或
(v)BoNT/E受体结合结构域(HC结构域)或其一部分；或
(vi)BoNT/F受体结合结构域(HC结构域)或其一部分；或
(vii)BoNT/G受体结合结构域(HC结构域)或其一部分；或
(viii)TeNT受体结合结构域(HC结构域)或其一部分。
在一个实施方案中，来自不同(即，非BoNT/X)梭菌神经毒素结构域的受体结合结构域(HC结构域)或其部分可以是结合突触结合蛋白I和/或II(Syt I/II)的结构域或其部分。
优选地，嵌合梭菌神经毒素可以包含无催化活性BoNT/X轻链和转运结构域(LHN结构域)以及BoNT/B受体结合结构域(HC结构域)或其一部分。
包含无催化活性BoNT/X轻链和转运结构域(LHN结构域)以及来自不同(即，非BoNT/X)梭菌神经毒素的受体结合结构域(HC结构域)或其一部分的多肽可以包含与SEQ IDNO:63-70中的任一个具有至少70％序列同一性的多肽序列。在一个实施方案中，包含无催化活性BoNT/X轻链和转运结构域(LHN结构域)以及来自不同(即，非BoNT/X)梭菌神经毒素的受体结合结构域(HC结构域)或其一部分的多肽包含与SEQ ID NO:63-70中的任一个具有至少80％、90％、95％或98％序列同一性的多肽序列。优选地，包含无催化活性BoNT/X轻链和转运结构域(LHN结构域)以及来自不同(即，非BoNT/X)梭菌神经毒素的受体结合结构域(HC结构域)或其一部分的多肽包含SEQ ID NO:63-70中的任一个(更优选地由其组成)。在这些多肽中，SEQ ID NO:63-66是最优选的。
包含无催化活性BoNT/X轻链和转运结构域(LHN结构域)以及来自不同(即，非BoNT/X)梭菌神经毒素的受体结合结构域(HC结构域)或其一部分的多肽可以包含以下N端氨基酸序列MGS。其中SEQ ID NO:63-66包含该N端氨基酸序列，该序列是任选的。在一个实施方案中，SEQ ID NO:63-66缺乏显示为MGS的N端氨基酸序列。在一个实施方案中，SEQ IDNO:63-66包含显示为MGS的N端氨基酸序列。
来自BoNT/X的无催化活性LHN结构域可以对应于SEQ ID NO:63的氨基酸残基1-899，或与之具有至少70％序列同一性的多肽序列。来自BoNT/X的无催化活性LHN结构域可以对应于SEQ ID NO:63的氨基酸残基1-899，或与之具有至少80％、90％或95％序列同一性的多肽序列。优选地，来自BoNT/X的无催化活性LHN结构域可以对应于SEQ ID NO:63的氨基酸残基1-899。
来自BoNT/X的无催化活性LHN结构域可以对应于SEQ ID NO:63的氨基酸残基4-899，或与之具有至少70％序列同一性的多肽序列。来自BoNT/X的无催化活性LHN结构域可以对应于SEQ ID NO:63的氨基酸残基4-899，或与之具有至少80％、90％或95％序列同一性的多肽序列。优选地，来自BoNT/X的无催化活性LHN结构域可以对应于SEQ ID NO:63的氨基酸残基4-899。
来自BoNT/X的无催化活性LHN结构域可以对应于SEQ ID NO:65的氨基酸残基1-866，或与之具有至少70％序列同一性的多肽序列。来自BoNT/X的无催化活性LHN结构域可以对应于SEQ ID NO:65的氨基酸残基1-866，或与之具有至少80％、90％或95％序列同一性的多肽序列。优选地，来自BoNT/X的无催化活性LHN结构域可以对应于SEQ ID NO:65的氨基酸残基1-866。
来自BoNT/X的无催化活性LHN结构域可以对应于SEQ ID NO:65的氨基酸残基4-866，或与之具有至少70％序列同一性的多肽序列。来自BoNT/X的无催化活性LHN结构域可以对应于SEQ ID NO:65的氨基酸残基4-866，或与之具有至少80％、90％或95％序列同一性的多肽序列。优选地，来自BoNT/X的无催化活性LHN结构域可以对应于SEQ ID NO:65的氨基酸残基4-866。
来自BoNT/B的HC结构域可以对应于SEQ ID NO:52的氨基酸残基860至1291，或与之具有至少70％序列同一性的多肽序列。来自BoNT/B的HC结构域可以对应于SEQ ID NO:52的氨基酸残基860至1291，或与之具有至少80％、90％或95％序列同一性的多肽序列。优选地，来自BoNT/B的HC结构域对应于SEQ ID NO:52的氨基酸残基860-1291。
优选地，BoNT/B HC结构域在HCC亚结构域中进一步包含至少一个氨基酸残基取代、添加或缺失，与天然BoNT/B序列相比，其具有增加BoNT/B神经毒素对人Syt II的结合亲和力的作用。BoNT/B HCC亚结构域中适宜的氨基酸残基取代、添加或缺失已在WO 2013/180799和WO 2016/154534中公开(均在此引入作为参考)。
BoNT/B HCC亚结构域中适宜的氨基酸残基取代、添加或缺失包括选自以下的取代突变：V1118M；Y1183M；E1191M；E1191I；E1191Q；E1191T；S1199Y；S1199F；S1199L；S1201V；E1191C、E1191V、E1191L、E1191Y、S1199W、S1199E、S1199H、W1178Y、W1178Q、W1178A、W1178S、Y1183C、Y1183P及其组合。
BoNT/B HCC亚结构域中适宜的氨基酸残基取代、添加或缺失还包括选自以下的两个取代突变的组合：E1191M和S1199L、E1191M和S1199Y、E1191M和S1199F、E1191Q和S1199L、E1191Q和S1199Y、E1191Q和S1199F、E1191M和S1199W、E1191M和W1178Q、E1191C和S1199W、E1191C和S1199Y、E1191C和W1178Q、E1191Q和S1199W、E1191V和S1199W、E1191V和S1199Y、或E1191V和W1178Q。
BoNT/B HCC亚结构域中适宜的氨基酸残基取代、添加或缺失还包括E1191M、S1199W和W1178Q三个取代突变的组合。
优选地，BoNT/B HCC亚结构域中适宜的氨基酸残基取代、添加或缺失包括E1191M和S1199Y两个取代突变的组合。
该修饰可以是与SEQ ID NO:52所示的未修饰BoNT/B相比的修饰，其中氨基酸残基编号通过与SEQ ID NO:52比对来确定。由于在SEQ ID NO:52的位置1处甲硫氨酸残基的存在是任选的，本领域技术人员在确定氨基酸残基编号时将考虑甲硫氨酸残基的存在/不存在。例如，在SEQ ID NO:52包含甲硫氨酸的情况下，位置编号将如上所定义(例如E1191将是SEQ ID NO:52的E1191)。备选地，在SEQ ID NO:52中不存在甲硫氨酸的情况下，氨基酸残基编号应改为-1(例如E1191将是SEQ ID NO.52的E1190)。在本文所述的其他多肽序列的位置1处的甲硫氨酸存在/不存在时，类似的考虑适用，并且本领域技术人员将使用本领域常规技术容易地确定正确的氨基酸残基编号。
来自BoNT/A的HC结构域可以对应于SEQ ID NO:60的氨基酸残基873至1296，或与之具有至少70％序列同一性的多肽序列。来自BoNT/A的HC结构域可以对应于SEQ ID NO:60的氨基酸残基873至1296，或与之具有至少80％、90％或95％序列同一性的多肽序列。优选地，来自BoNT/B的HC结构域对应于SEQ ID NO:60的氨基酸残基873至1296。
在一个实施方案中，在多肽用于治疗炎性疾病的情况下，所使用的多肽不包含无催化活性BoNT/X L链、BoNT/X转运结构域(HN结构域)和BoNT/A受体结合结构域(HC结构域)。因此，在一个实施方案中，在多肽用于治疗炎性疾病的情况下，该多肽可以包含无催化活性BoNT/X L链、BoNT/X转运结构域(HN结构域)和：(i)BoNT/B受体结合结构域(HC结构域)；(ii)BoNT/D受体结合结构域(HC结构域)；或(iii)BoNT/F受体结合结构域(HC结构域)。
类似地，在一个实施方案中，在多肽用于治疗疼痛的情况下，所使用的多肽不包含无催化活性BoNT/X L链、BoNT/X转运结构域(HN结构域)和BoNT/A受体结合结构域(HC结构域)。因此，在一个实施方案中，在多肽用于治疗疼痛的情况下，该多肽可以包含无催化活性BoNT/X L链、BoNT/X转运结构域(HN结构域)和：(i)BoNT/B受体结合结构域(HC结构域)；(ii)BoNT/D受体结合结构域(HC结构域)；或(iii)BoNT/F受体结合结构域(HC结构域)。
来自BoNT/D的HC结构域可以对应于SEQ ID NO:54的氨基酸残基865-1276，或与之具有至少70％序列同一性的多肽序列。来自BoNT/D的HC结构域可以对应于SEQ ID NO:54的氨基酸残基865-1276，或与之具有至少80％、90％或95％序列同一性的多肽序列。优选地，来自BoNT/D的HC结构域对应于SEQ ID NO:54的氨基酸残基865-1276。
来自BoNT/F的HC结构域可以对应于SEQ ID NO:56的氨基酸残基866-1278，或与之具有至少70％序列同一性的多肽序列。来自BoNT/F的HC结构域可以对应于SEQ ID NO:56的氨基酸残基866-1278，或与之具有至少80％、90％或95％序列同一性的多肽序列。优选地，来自BoNT/F的HC结构域对应于SEQ ID NO:56的氨基酸残基866-1278，其在位置1241处具有组氨酸至赖氨酸的取代(H1241K)。
优选地，该嵌合梭菌神经毒素可以包含无催化活性BoNT/X轻链和转运结构域(LHN结构域)以及来自不同(即，非BoNT/X)梭菌神经毒素的受体结合结构域(HC结构域)或其一部分和Cys-(Xaa)a-Ile-Asp/Glu-Gly-Arg-(Yaa)b-Cys(SEQ ID NO:71)(优选由其组成)，其中a＝1-10，b＝4-15。SEQ ID NO:71是基于BoNT/C1激活环的激活环共有序列。
此外，本发明的任何多肽可以包含Cys-(Xaa)a-Ile-Asp/Glu-Gly-Arg-(Yaa)b-Cys(SEQ ID NO:71)，其中a＝1-10，b＝4-15。该激活环可以适宜地取代存在于本文所述多肽中存在的梭菌神经毒素L链和/或转运结构域(HN结构域)中的任何激活环序列。
在SEQ ID NO:71的上下文中使用时，Xaa或Yaa可以是任何氨基酸。Xaa和Yaa位置处的氨基酸数量分别用字母“a”和“b”表示。在一个实施方案中，“a”和“b”可以是允许激活环的蛋白水解切割并产生活性双链梭菌神经毒素的任何整数。在一个实施方案中，“a”是至少1、2、3、4、5、6、7、8、9或10。在一个实施方案中，“b”是至少1、2、3、4、5、6、7、8、9、10、11、12、13、14或15。在一个实施方案中，“a”是≤12、≤11、≤10、≤9、≤8、≤7、≤6、≤5或≤4。在一个实施方案中，“b”是≤20、≤19、≤18、≤17、≤16、≤15、≤14、≤13、≤12、≤11、≤10或≤9。
在一个实施方案中，“a”是1-12，例如1-10。优选地，“a”是1-7，如2-4。更优选地，“a”是3。在一个实施方案中，“b”是1-20，例如4-15。优选地，“b”是6-10。更优选地，“b”是8。
Xaa或Yaa并非仅限于一种类型的氨基酸。因此，存在于位置Xaa处的一个或多个残基可以独立地选自标准氨基酸：天冬氨酸、谷氨酸、精氨酸、赖氨酸、组氨酸、天冬酰胺、谷氨酰胺、丝氨酸、苏氨酸、酪氨酸、甲硫氨酸、色氨酸、半胱氨酸、丙氨酸、甘氨酸、缬氨酸、亮氨酸、异亮氨酸、脯氨酸和苯丙氨酸。存在于位置Yaa处的一个或多个残基可以独立地选自标准氨基酸：天冬氨酸、谷氨酸、精氨酸、赖氨酸、组氨酸、天冬酰胺、谷氨酰胺、丝氨酸、苏氨酸、酪氨酸、甲硫氨酸、色氨酸、半胱氨酸、丙氨酸、甘氨酸、缬氨酸、亮氨酸、异亮氨酸、脯氨酸和苯丙氨酸。优选地，位置Yaa(更优选紧邻SEQ ID NO:71的Arg残基的C端)处的氨基酸不是脯氨酸。
备选地/额外地，存在于位置Xaa或Yaa处的一个或多个残基可以独立地选自非标准氨基酸(不是上述20种标准组的一部分的氨基酸)。作为实例，非标准氨基酸可以包括4-羟基脯氨酸、6-N-甲基赖氨酸、2-氨基异丁酸、异缬氨酸、α-甲基丝氨酸、反式-3-甲基脯氨酸、2,4-甲基脯氨酸、顺式-4-羟基脯氨酸、反式-4-羟基-脯氨酸、N-甲基甘氨酸、别苏氨酸、甲基苏氨酸、羟乙基半胱氨酸、羟乙基同型半胱氨酸、硝基谷氨酰胺、同型谷氨酰胺、六氢吡啶羧酸、叔亮氨酸、正缬氨酸、2-氮杂苯丙氨酸、3-氮杂苯丙氨酸、4-氮杂苯丙氨酸、L-鸟氨酸、L-2-氨基-3-胍丙酸或赖氨酸、精氨酸和/或鸟氨酸的D-异构体，以及4-氟苯丙氨酸。用于将非标准氨基酸引入蛋白质的方法是本领域已知的，并且包括使用大肠杆菌营养缺陷型表达宿主的重组蛋白质合成。
包含在SEQ ID NO:71中的序列Ile-Asp/Glu-Gly-Arg是指在WO 2020/065336 A1中意外发现的由肠激酶(以及因子Xa)识别的位点。该文献描述了用于在Ile-Asp/Glu-Gly-Arg处切割从而产生双链多肽的适宜方法。优选该序列是Ile-Asp-Gly-Arg，例如Cys-(Xaa)a-Ile-Asp-Gly-Arg-(Yaa)b-Cys。认为肠激酶和因子Xa水解SEQ ID NO:71的Arg C端紧邻的肽键(即Arg和Yaa之间的肽键)。
在一个实施方案中，SEQ ID NO:71Ile的N端紧邻的Xaa处的氨基酸残基是不带电荷的疏水性氨基酸，优选丙氨酸。在一些实施方案中，“a”是至少2，并且Xaa包含至少一个C端不带电荷的极性氨基酸和紧邻其N端的带电荷的碱性氨基酸。带电荷的碱性氨基酸优选为赖氨酸。因此，在“a”为至少2的实施方案中，Xaa可以包含至少Lys-Ala，其中Ala紧邻SEQID NO:71Ile的N端。
在一个实施方案中，Xaa包含序列HKA或由序列HKA组成。
在一个实施方案中，紧邻SEQ ID NO:71Arg C端的Yaa处的氨基酸残基是不带电荷的极性氨基酸，优选丝氨酸。在一些实施方案中，“b”是至少2，并且Yaa包含至少一个N端不带电荷的极性氨基酸和紧邻其C端的不带电荷的疏水性氨基酸。不带电荷的疏水性氨基酸优选为亮氨酸。因此，在“b”为至少2的实施方案中，Yaa可以包含至少Ser-Leu，其中Ser紧邻SEQ ID NO:71Arg的C端。
在一个实施方案中，Yaa包含序列SLYNKTLDC或由序列SLYNKTLDC组成。
在一些实施方案中，本文的多肽包含与SEQ ID NO:72具有至少70％序列同一性的激活环。在一个实施方案中，本文的多肽包含与SEQ ID NO:72具有至少80％、85％或90％序列同一性的激活环。优选地，本文的多肽包含与SEQ ID NO:72具有至少95％序列同一性的激活环。更优选地，本文的多肽包含与SEQ ID NO:72具有至少99％序列同一性的激活环。
在一个尤其优选的实施方案中，本文的多肽包含含有SEQ ID NO:72的激活环，更优选地包含由SEQ ID NO:72组成的激活环。
激活环也可以是SEQ ID NO:72的变体，如SEQ ID NO:73或与之具有至少70％序列同一性的序列。SEQ ID NO:73是SEQ ID NO:72的变体，其中肠激酶识别位点IDGR已突变为IEGR。在一个实施方案中，本文的多肽包含与SEQ ID NO:73具有至少70％序列同一性的激活环。在一个实施方案中，本文的多肽包含与SEQ ID NO:73具有至少80％、85％或90％序列同一性的激活环。优选地，本文的多肽包含与SEQ ID NO:73具有至少95％序列同一性的激活环。更优选地，本文的多肽包含与SEQ ID NO:73具有至少99％序列同一性的激活环。
在一个尤其优选的实施方案中，本文的多肽包含含有SEQ ID NO:73的激活环，更优选由SEQ ID NO:73组成。
在一个实施方案中，可以修饰本文所述的激活环(例如SEQ ID NO:71、72或73)以包含额外或备选的蛋白酶位点。例如，如SEQ ID NO:77所示的蛋白酶位点。这种经修饰的激活环的一个实例显示为SEQ ID NO:78。因此，在一个实施方案中，本文的多肽包含与SEQ IDNO:78具有至少70％序列同一性的激活环。在一个实施方案中，本文的多肽包含与SEQ IDNO:78具有至少80％、85％或90％序列同一性的激活环。优选地，本文的多肽包含与SEQ IDNO:78具有至少95％序列同一性的激活环。更优选地，本文的多肽包含与SEQ ID NO:78具有至少99％序列同一性的激活环。在尤其优选的实施方案中，本文的多肽包含含有SEQ IDNO:78的激活环，更优选地包含由SEQ ID NO:78组成的激活环。
在一个实施方案中，本发明的多肽可以包含重靶向梭菌神经毒素(或由其组成)，条件是存在的任何L链都无催化活性。在重靶向梭菌神经毒素中，修饰梭菌神经毒素以包含称为靶向部分(Targeting Moiety，TM)的外源配体。选择TM以提供对所希望的靶细胞的结合特异性，并且作为重靶向过程的一部分，可以去除梭菌神经毒素的天然结合部分(例如HC结构域或HCC结构域)。重靶向技术描述于例如EP-B-0689459、WO 1994/021300、EP-B-0939818、US 6,461,617、US 7,192,596、WO 1998/007864、EP-B-0826051、US 5,989,545、US6,395,513、US 6,962,703、WO 1996/033273、EP-B-0996468、US 7,052,702、WO 1999/017806、EP-B-1107794、US 6,632,440、WO 2000/010598、WO 2001/21213、WO 2006/059093、WO 2000/62814、WO 2000/04926、WO 1993/15766、WO 2000/61192和WO 1999/58571中，所有这些文献在此以其整体引入作为参考。因此，在一个实施方案中，本发明的多肽是重靶向梭菌神经毒素，条件是存在的任何L链都无催化活性。本发明的多肽可以缺乏梭菌神经毒素的功能性HC结构域，并且也缺乏任何功能等同的TM。
在本文所述的多肽具有用于纯化的标签(例如His标签)和/或接头的实施方案中，该标签和/或接头是任选的。
本发明的多肽可以不含存在于天然存在的梭菌神经毒素复合物中的复合蛋白质。
本发明的多肽可以使用重组核酸技术产生。因此，在一个实施方案中，(如上所述的)多肽是重组多肽。
在一个实施方案中，提供包含编码多肽的核酸序列的核酸(例如DNA)。在一个实施方案中，该核酸序列作为包含启动子和终止子的DNA载体的一部分制备。该核酸序列可以选自本文所述的任何核酸序列。
在一个优选实施方案中，该载体具有选自以下的启动子：
在另一个优选实施方案中，该载体具有选自以下的启动子：
该核酸分子可以使用本领域已知的任何适宜的方法制备。因此，该核酸分子可以使用化学合成技术制备。备选地，本发明的核酸分子可以使用分子生物学技术制备。
本发明的DNA构建体优选在计算机中设计，然后通过常规DNA合成技术合成。
根据要使用的最终宿主细胞(例如大肠杆菌)表达系统，任选地针对密码子偏好修饰上述核酸序列信息。
术语“核苷酸序列”和“核酸”在本文中用作同义词。优选地，该核苷酸序列是DNA序列。
本发明的多肽(尤其是其任何梭菌神经毒素部分)可以作为单链或双链存在。然而，优选的是，该多肽作为双链存在，其中无催化活性L链通过二硫键与H链(或其组分，例如HN结构域)连接。
本发明提供产生具有无催化活性轻链和重链的单链多肽的方法，该方法包括在表达宿主中表达本文所述的核酸，裂解宿主细胞以提供含有该单链多肽的宿主细胞匀浆，并分离该单链多肽。在一个方面，本发明提供蛋白水解处理本文所述多肽的方法，该方法包括使该多肽与水解多肽激活环中肽键的蛋白酶接触，从而将(单链)多肽转化为相应的双链多肽(例如其中无催化活性轻链和重链通过二硫键连接在一起)。
因此，本发明提供可通过本发明的方法获得的双链多肽。
本文中所用的“受试者”可以是哺乳动物，如人类或其他哺乳动物。优选地，“受试者”意指人受试者。
本文所用的术语“障碍(disorder)”也涵盖“疾病”。在一个实施方案中，该障碍是疾病。
本文所用的术语“治疗”涵盖预防性治疗(例如以预防疾病[例如疼痛]的发作)以及矫正性治疗(治疗已经患有疾病[例如疼痛]的受试者)。优选地，本文所用的“治疗”意指矫正性治疗。
本文所用的术语“治疗”是指疾病(例如疼痛)和/或其症状。
因此，本发明的多肽可以以治疗有效量或预防有效量对受试者施用。优选地，本发明的多肽以治疗有效量对受试者施用。
“治疗有效量”是指任何量的多肽，在单独或与另一种药物组合对受试者施用以治疗该疾病(如疼痛)(或其症状)时，其足以对该疾病或其症状进行这种治疗。
“预防有效量”是指任何量的多肽，在单独或与另一种药物组合对受试者施用时，其抑制或延迟疾病(例如疼痛)(或其症状)的发作或复发。在一些实施方案中，预防有效量完全防止疾病(例如疼痛)的发作或复发。“抑制”发作意味着降低疾病发作的可能性(例如，在该疾病是疼痛的情况下)(或其症状)，或完全防止发作。
本发明的多肽可以以任何适宜的方式配制用于对受试者施用，例如作为药物组合物的一部分。因此，在一个方面，本发明提供药物组合物，其包含本发明的多肽和可药用载体、赋形剂、佐剂、推进剂和/或盐。
本发明的多肽可配制用于口服、胃肠外、连续输注、吸入或局部应用。适合用于注射的组合物可以是溶液、悬液或乳液或在使用前溶解或悬浮在适宜溶媒中的干粉的形式。
在局部递送的多肽的情况下，多肽可以配制成膏剂(例如用于局部应用)或用于皮下注射。
局部递送方式可以包括气雾剂或其他喷雾(例如喷雾剂)。在这方面，多肽的气雾剂制剂能够递送到肺部和/或其他的鼻腔和/或支气管或气道。
本发明的多肽可以通过脊柱中的鞘内或硬膜外注射在涉及受影响器官的神经支配的脊柱节段水平对受试者施用。
给药途径可以是通过腹腔镜和/或局部注射。在一个实施方案中，本发明的多肽在待治疗的部位或附近施用，优选在待治疗的部位施用。例如，多肽可以鞘内或脊柱内施用。在一个实施方案中，本发明多肽的给药途径可以是神经周围、神经内、脊柱内和/或鞘内。
在一个实施方案中，本发明的多肽可以通过外周施用。在一个实施方案中，多肽可以皮内、皮下或肌内施用。优选地，本发明的多肽皮内施用。
本发明多肽的施用剂量范围是产生所希望的治疗和/或预防效果的剂量范围。应理解，所需的剂量范围取决于梭菌神经毒素或组合物的确切性质、给药途径、制剂的性质、受试者的年龄、受试者病症的性质、程度或严重程度、禁忌症(如果有的话)以及主治医师的判断。这些剂量水平的变化可以使用标准经验程序进行调整以进行优化。
在一个实施方案中，多肽的剂量是固定剂量(flat dose)。固定剂量可以在50pg至250μg、优选100pg至100μg的范围内。在一个实施方案中，固定剂量可以是至少50pg、100pg、500pg、1ng、50ng、100ng、500ng、1μg或50μg。该剂量可以是单个固定剂量。
在一个优选实施方案中，多肽可以按大于250μg的量给药。在一个实施方案中，本发明多肽可以按大于500μg、1mg、10mg、100mg、500mg、1g或5g的量给药。在一个实施方案中，本发明的多肽可以按等于或小于10g、5g、1g、500mg、100mg、10mg或1mg的量给药。优选地，本发明的多肽按251μg至10g、251μg至5g、251μg至1g、251μg到500mg、251μg至100mg、251μg至10mg或251μg至1000μg、例如251μg至500μg的量给药。在一个实施方案中，本发明的多肽按500μg至5g、例如1mg至1g或1g至3g的量施用。这是由于本发明多肽的无毒(例如基本无毒)性质使所述剂量成为可能。
流体剂型通常利用多肽和无热原无菌溶媒制备。取决于所使用的溶媒和浓度，梭菌神经毒素可以溶解或悬浮在溶媒中。在制备溶液时，多肽可以溶解在溶媒中，根据需要，通过添加氯化钠使溶液等渗，并使用无菌技术通过无菌过滤器过滤进行灭菌，然后灌装到适宜的无菌小瓶或安瓿中并密封。备选地，如果溶液稳定性足够，可以通过高压灭菌对密封容器中的溶液进行灭菌。有利地，添加剂如缓冲剂、增溶剂、稳定剂、防腐剂或杀菌剂、悬浮剂或乳化剂和/或局部麻醉剂可以溶解在溶媒中。
在使用前溶解或悬浮在适宜溶媒中的干粉可以通过在无菌区域使用无菌技术将预灭菌成分灌装到无菌容器中来制备。备选地，可以在无菌区域使用无菌技术将成分溶解到适宜的容器中。然后将产品冷冻干燥，并对容器进行无菌密封。
适用于本文所述给药途径的胃肠外悬浮液以基本相同的方式制备，不同之处在于无菌成分悬浮在无菌溶媒中，而不是溶解，并且不能通过过滤完成灭菌。成分可以在无菌状态下分离，或者备选地，可以在分离后进行灭菌，例如通过伽马辐射。
有利地，将悬浮剂例如聚乙烯吡咯烷酮包含在组合物中以便于组分的均匀分布。
根据本发明的施用可以利用多种递送技术，包括微粒封装(microparticleencapsulation)或高压气溶胶冲击。
本发明的多肽优选作为治疗方案的一部分反复施用(例如，多至5、10、15或20次)。反复施用是指至少施用两次，例如至少5、10、15或20次。因此，在一个实施方案中，本发明的多肽可以施用两次或多次以治疗受试者(例如治疗受试者的疼痛)。这对于慢性病的治疗尤其重要，如慢性疼痛，在这种情况下通常需要持续治疗。在一个实施方案中，本发明的多肽可以每周一次、每月两次、每月一次、每两个月一次、每六个月一次或每年一次施用，优选每年至少两次或每年一次。在一个实施方案中，本发明的多肽在10年、5年、2年或1年的时间内施用两次或多次。优选地，本发明的多肽在一年的时间内施用两次或多次。治疗可持续至少6个月、1年、2年、3年、5年、10年、15年、20年、25年或30年。
优选的是，多肽不与除了无催化活性梭菌神经毒素L链、HN结构域和/或HC结构域之外的另一种治疗或诊断剂(例如核酸、蛋白质、肽或小分子治疗或诊断剂)一起施用。例如，在一个实施方案中，多肽不与另一种镇痛剂和/或抗炎剂一起施用。在一个实施方案中，本发明的多肽不与共价结合的治疗剂一起施用。在一个实施方案中，本发明的多肽不与非共价结合的治疗剂一起施用。
本文所述的多肽可用于治疗患有一种或多种类型疼痛的受试者。疼痛可以是慢性或急性疼痛。疼痛可以是选自以下四类疼痛中的一种或多种：伤害性疼痛；神经性疼痛(neuropathic pain)；混合性疼痛；以及来源不明的疼痛。伤害性疼痛可以由已知的对伤害感受器(疼痛受体)的伤害性刺激引起，并且可以是躯体的或内脏的。神经性疼痛可以是由神经系统的原发性损伤或功能障碍起始或引起的疼痛。混合性疼痛可以是伤害性疼痛和神经性疼痛的组合。
本发明治疗的疼痛(例如慢性疼痛)的实例包括神经性疼痛、炎性疼痛、头痛、躯体疼痛、内脏疼痛、牵涉性疼痛、异常性疼痛、混合性疼痛和手术后疼痛。
此处使用的术语“疼痛”是指任何不愉快的感官体验，通常与身体疾病有关。对于临床医生来说，身体疾病可以是显而易见的，也可以不是显而易见的。疼痛有两种类型：慢性和急性。“急性疼痛”是指突然发作的短时间疼痛。例如，一种类型的急性疼痛是皮肤或其他浅表组织受伤时感觉到的皮肤疼痛，如由割伤或烧伤引起的疼痛。皮肤伤害感受器终止于皮肤下方，由于神经末梢浓度高，产生明确的、局部的短时间疼痛。“慢性疼痛”是指急性疼痛以外的疼痛。
本发明的多肽可以用于治疗由以下任何神经性疼痛病症引起或与之相关的疼痛。“神经性疼痛”是指来自外周神经系统、中枢神经系统或二者的导致不适的异常感觉输入。神经性疼痛的症状可以涉及持续的自发疼痛，以及异常性疼痛(对通常不疼痛的刺激的疼痛反应)、痛觉过敏(对通常只引起轻微不适的疼痛刺激(如针刺)的加重反应)或过度疼痛(hyperpathia)(短暂的不适变为长期的剧烈疼痛)。神经性疼痛可以由以下任何一种引起：
1.创伤性损伤，例如神经压迫性损伤(例如神经挤压、神经拉伸、神经卡压或不完全神经横断)；脊髓损伤(例如脊髓半截)；截肢；挫伤；炎症(例如脊髓炎症)；或外科手术。
2.缺血性事件，包括例如中风和心脏病发作。
3.感染性物质。
4.暴露于毒性物质，包括例如药物、酒精、重金属(如铅、砷、汞)、工业物质(例如溶剂、胶水产生的烟雾)或一氧化二氮。
5.疾病，包括例如炎性疾病、肿瘤、获得性免疫缺陷综合征(AIDS)、莱姆病、麻风病、代谢性疾病、外周神经疾病，如神经瘤、单神经病或多发性神经病。
神经性疼痛的类型包括以下几种：
1.神经痛(neuralgia)
神经痛是一种沿着一条或多条特定神经传导的疼痛，通常神经结构没有任何明显的病理变化。神经痛的病因多种多样。化学刺激、炎症、创伤(包括手术)、附近结构(例如肿瘤)的压迫和感染都可以导致神经痛。然而，在许多情况下，原因未知或无法识别。神经痛在老年人中最常见，但它可以发生在任何年龄。神经痛包括但不限于三叉神经痛、疱疹后神经痛、带状疱疹后神经疼痛、舌咽神经痛、坐骨神经痛和非典型面部疼痛。
神经痛是一种或多种神经分布的疼痛。实例有三叉神经痛、非典型面部疼痛和带状疱疹后神经痛(由带状疱疹或疱疹引起)。受影响的神经负责感应从下巴到前额的面部区域的触摸、温度和压力。这种障碍通常导致短暂的剧烈疼痛，通常不到两分钟，而且只在一侧面部。疼痛可以用多种方式描述，如“刺痛”、“尖锐”、“像闪电一样”、“灼热”和甚至“发痒”。在非典型形式的TN中，疼痛也可以表现为严重疼痛或仅仅疼痛，并持续很长时间。与TN相关的疼痛被认为是可以经历的最剧烈的疼痛之一。
简单的刺激，如吃饭、说话、洗脸，或任何轻微的触摸或感觉都可以引发攻击(甚至是微风的感觉)。攻击可以集群发生，也可以作为孤立攻击发生。
症状包括剧烈刺痛或持续灼痛，其位于任何地方，通常在体表或体表附近，每次发作都在同一位置；沿着特定神经路径的疼痛；受影响身体部位因疼痛而功能受损，或因伴随的运动神经损伤而肌肉无力；皮肤的敏感性增加或受影响皮肤区域的麻木(感觉类似于局部麻醉剂，如奴佛卡因(Novocaine)注射)；并且任何触摸或压力都被解释为疼痛。运动也可以是疼痛的。
三叉神经痛是最常见的神经痛形式。它影响面部的主要感觉神经，三叉神经(“三叉神经”的字面意思是“三个起源”，指的是将神经分为3个分支)。这种病症涉及面部一侧沿着由该侧的三叉神经供应的区域突然出现短暂的剧烈疼痛。疼痛攻击的严重程度可足以导致脸部扭曲，这通常称为疼痛性抽搐(tic douloureux)。有时，三叉神经痛的病因是压迫神经的血管或小肿瘤。多发性硬化(一种影响大脑和脊髓的炎性疾病)、某些形式的关节炎和糖尿病(高血糖)等疾病也可以导致三叉神经痛，但病因并不总是明确的。在这种病症中，某些动作，如咀嚼、说话、吞咽或触摸面部区域可以引发剧烈疼痛的痉挛。
一种相关但相当罕见的神经痛影响舌咽神经，舌咽神经为喉咙提供感觉。这种神经痛的症状是位于喉咙的短暂的、类似休克的疼痛发作。
带状疱疹等感染后可以出现神经痛，带状疱疹是由水痘-带状疱疹病毒(一种疱疹病毒)引起的。带状疱疹皮疹痊愈后，这种神经痛产生持续的灼痛。疼痛因受影响区域的移动或接触而加重。并非所有诊断为带状疱疹的人都会经历带状疱疹后神经痛，这可以比带状疱疹更痛苦。疼痛和敏感可以持续数月甚至数年。疼痛通常表现为对任何触摸都无法忍受的敏感，尤其是轻微触摸。带状疱疹后神经痛并不局限于面部；它可以发生在身体的任何地方，但通常发生在带状疱疹皮疹的位置。由于疾病期间的疼痛和社交孤立，抑郁症并不罕见。
带状疱疹后神经痛可以在原始疱疹感染症状消失后很长一段时间内使人衰弱。其他可以引起神经痛的传染病有梅毒和莱姆病。
糖尿病是神经痛的另一个常见原因。这种非常常见的医疗问题在成年期间几乎每20个美国人中就有1个受到影响。糖尿病损害为神经提供循环的微小动脉，导致神经纤维功能紊乱，有时还会导致神经损失。糖尿病几乎可以产生任何神经痛，包括三叉神经痛、腕管综合征(手和手腕的疼痛和麻木)和感觉异常性股痛(股外侧皮神经损伤引起的股麻木和疼痛)。严格控制血糖可以预防糖尿病神经损伤，并可以加速发生了神经痛的受试者的康复。
其他可以与神经痛有关的病症包括慢性肾功能不全和卟啉症－一种遗传性疾病，在这种疾病中，身体无法清除体内血液正常分解后产生的某些物质。某些药物也可以导致这个问题。
2.传入神经阻滞
传入神经阻滞(Deafferentation)表示身体某一部分的感觉输入丧失，可以由外周感觉纤维或中枢神经系统的神经中断引起。传入神经阻滞疼痛综合征包括但不限于大脑或脊髓损伤、中风后疼痛、幻像疼痛、截瘫、臂丛神经撕脱伤、腰椎神经根病。
3.复杂性区域疼痛综合征(CRPS)
CRPS是一种由交感神经维持的疼痛引起的慢性疼痛综合征，有两种形式。CRPS1目前取代了术语“反射性交感神经营养不良综合征”。它是一种慢性神经疾病，最常见于轻伤或重伤后的手臂或腿部。CRPS1与严重疼痛有关；指甲、骨骼和皮肤的变化；以及受影响肢体对触摸的敏感性增加。CRPS2取代了术语灼痛(causalgia)，是由已确定的神经损伤引起的。CRPS包括但不限于CRPS I型(反射性交感神经营养不良)和CRPS II型(灼痛)。
4.神经病
神经病(neuropathy)是神经的功能或病理变化，临床特征为感觉或运动神经元异常。
中枢神经病是中枢神经系统的功能或病理变化。
外周神经病是一种或多种外周神经的功能或病理变化。外周神经将信息从中枢神经系统(大脑和脊髓)传递到肌肉和其他器官，并从皮肤、关节和其他器官传递回大脑。在这些神经无法将信息传至和传出大脑和脊髓时，发生外周神经病，导致疼痛、感觉丧失或无法控制肌肉。在某些情况下，控制血管、肠道和其他器官的神经失效导致血压异常、消化问题和其他身体基本过程的丧失。神经病的风险因素包括糖尿病、大量饮酒以及接触某些化学物质和药物。有些人有神经病的遗传倾向。对神经的长期压力是神经损伤的另一个风险。压力损伤可以由长时间的不动(如长时间的手术或长期疾病)或石膏、夹板、支架、拐杖或其他装置压迫神经引起。多发性神经病意味着一个广泛的过程，通常对身体两侧产生同等影响。症状取决于受影响的神经类型。三种主要类型的神经是感觉神经、运动神经和自主神经。神经病可以影响任何一种神经或所有三种神经的组合。症状还取决于这种病症是影响全身还是只影响一条神经(如受伤)。慢性炎性多发性神经病的病因是异常免疫反应。特异性抗原、免疫过程和触发因素是可变的，且在许多情况下是未知的。它可以与其他病症有关，如HIV、炎性肠病、红斑狼疮、慢性活动性肝炎和血细胞异常。
外周神经病可以涉及单个神经或神经组的功能或病理变化(单神经病)或影响多个神经的功能或病理变化(多发性神经病)。
外周神经病可以包括以下：
遗传性疾病
Charcot-Marie-Tooth病
弗里德赖希共济失调
系统性或代谢性疾病
糖尿病(糖尿病神经病)
饮食缺乏(尤其是维生素B-12)
过度饮酒(酒精性神经病)
尿毒症(肾衰竭)
癌症
感染性或炎性疾病
艾滋病
肝炎
科罗拉多蜱热
白喉
吉兰-巴雷综合征
未发展为AIDS的HIV感染
麻风病
莱姆病
结节性多动脉炎
类风湿性关节炎
结节病
干燥综合征
梅毒
系统性红斑狼疮
淀粉样蛋白
接触有毒化合物
嗅胶或其他毒性化合物
一氧化二氮
工业物质，尤其是溶剂
重金属(铅、砷、汞等)
药物如镇痛剂肾病继发的神经病
其他原因
局部缺血(氧气减少/血流量减少)
长时间暴露于低温
a.多发性神经病
多发性神经病(Polyneuropathy)是一种外周神经病，涉及多个外周神经受损或破坏导致某个区域的运动或感觉丧失。多发性神经病性疼痛，包括但不限于脊髓灰质炎后综合征、乳房切除术后综合征、糖尿病性神经病、酒精性神经病、淀粉样蛋白、毒素、AIDS、甲状腺功能减退、尿毒症、维生素缺乏、化疗引起的疼痛、2',3'-双脱氧胞苷(ddC)治疗、吉兰-巴雷综合征或法布里病。
b.单神经病
单神经病(Mononeuropathy)是一种外周神经病，涉及由单个外周神经或神经组的损伤或破坏引起的某个区域的运动或感觉丧失。单神经病通常由损伤或创伤对局部区域造成的损伤引起，尽管偶尔系统性疾病可以导致孤立的神经损伤(如多发性单神经炎)。常见的原因是直接创伤、对神经的长期压迫以及附近身体结构的肿胀或损伤对神经的压迫。损伤包括神经髓鞘(覆盖物)或部分神经细胞(轴突)的破坏。这种损伤减缓或阻止冲动通过神经传导。单神经病可以涉及身体的任何部位。单神经病性疼痛，包括但不限于坐骨神经功能障碍、腓总神经功能障碍、桡神经功能障碍、尺神经功能障碍、颅单神经病VI、颅单神经病VII、颅单神经病III(压迫型)、颅单神经病III(糖尿病型)、腋神经功能障碍、腕管综合征、股神经功能障碍、胫神经功能障碍、贝尔麻痹、胸廓出口综合征、腕管综合征和第六(外展)神经麻痹。
c.全身性外周神经病
全身性外周神经病是对称的，通常是由于多种系统性病痛和疾病过程影响整个外周神经系统。它们被进一步细分为几个类别：
i.远端轴突病变是神经元某些代谢性或毒性紊乱的结果。它们可以由代谢性疾病引起，如糖尿病、肾衰竭、营养不良和酗酒等缺乏综合征、或毒素或药物的影响。远端轴突病变(又称死亡性背部神经病)是一种由外周神经系统(PNS)神经元的某些代谢性或毒性紊乱引起的外周神经病。它是神经对代谢性或毒性紊乱最常见的反应，因此可以由糖尿病、肾衰竭、营养不良和酗酒等代谢性疾病或毒素或药物的影响引起。远端轴突病变最常见的病因是糖尿病，最常见的远端轴突病变是糖尿病性神经病。
ii.髓鞘质病是由于髓磷脂的原发性攻击导致冲动传导的急性衰竭。最常见的病因是急性炎性脱髓鞘性多发性神经病(AIDP；又称吉兰-巴雷综合征)，尽管其他病因包括慢性炎性脱髓鞘综合征(CIDP)、遗传性代谢性疾病(例如白细胞营养不良)或毒素。髓鞘质病是由于髓鞘或有髓鞘的施旺细胞的原发性破坏，使轴突完好无损，但导致冲动传导的急性衰竭。这种脱髓鞘减缓或完全阻断了电冲动通过神经传导。最常见的病因是急性炎性脱髓鞘性多发性神经病(AIDP，更为人所知的是吉兰-巴雷综合征)，尽管其他病因包括慢性炎性脱髓鞘性多发性神经病(CIDP)、遗传性代谢性疾病(例如白细胞营养不良或Charcot-Marie-Tooth病)或毒素。
iii.神经元病(Neuronopathies)是外周神经系统(PNS)神经元破坏的结果。它们可以由运动神经元病、感觉神经元病(例如带状疱疹)、毒素或自主神经功能障碍引起。神经毒素可以引起神经元病，如化疗剂长春新碱。神经元病是由外周神经系统(PNS)的神经元受损引起的功能障碍，导致外周神经病。它可以由运动神经元病、感觉神经元病(如带状疱疹)、有毒物质或自主神经功能障碍引起。神经元病患者可以以不同的方式表现，这取决于病因、影响神经细胞的方式以及受影响最大的神经细胞类型。
iv.局灶性卡压性神经病(例如腕管综合征)。
在疼痛是神经病性疼痛的情况下，在一个实施方案中，所使用的多肽不包含无催化活性BoNT/X L链、BoNT/X转运结构域(HN结构域)和/或BoNT/X受体结合结构域(HC结构域)。在一个实施方案中，在疼痛是神经病性疼痛的情况下，所使用的多肽不包含无催化活性BoNT/X L链和转运结构域(HN结构域)，任选地与来自不同(即，非BoNT/X)梭菌神经毒素的HC结构域(例如来自BoNT/B的HC结构域)组合。优选地，在疼痛是神经病性疼痛的情况下，所使用的多肽不包含无催化活性BoNT/X L链和转运结构域(HN结构域)以及BoNT/BHC结构域。
本发明的多肽可用于治疗由以下任何炎性病症引起或与之相关的疼痛。类似地，本发明的多肽可用于治疗一种或多种以下炎性病症。
A.关节炎
关节疾病包括例如类风湿性关节炎；幼年类风湿性关节炎；系统性红斑狼疮(SLE)；痛风性关节炎；硬皮病；骨关节炎；银屑病关节炎；强直性脊柱炎；Reiter综合征(反应性关节炎)；成人斯蒂尔病；由病毒感染引起的关节炎；细菌感染引起的关节炎，例如淋球菌性关节炎和非淋球菌性细菌性关节炎(败血症性关节炎)；第三期莱姆病；结核性关节炎；以及真菌感染引起的关节炎，例如芽生霉菌病。
B.自身免疫性疾病
自身免疫性疾病包括例如吉兰-巴雷综合征、桥本甲状腺炎、恶性贫血、艾迪生病、I型糖尿病、系统性红斑狼疮、皮肌炎、干燥综合征、红斑狼疮、多发性硬化、重症肌无力、赖特综合征和格雷夫病。
C.结缔组织疾病
结缔组织疾病包括例如脊柱关节炎、皮肌炎和纤维肌痛。
D.损伤
损伤(包括例如挤压、穿刺、拉伸组织或关节)引起的炎症可以引起慢性炎性疼痛。
E.感染
感染(包括肺结核或间质性角膜炎)引起的炎症可以引起慢性炎性疼痛。
F.神经炎
神经炎是一种影响神经或神经组的炎症过程。症状取决于所涉及的神经，但可以包括疼痛、感觉异常、麻痹或感觉迟钝(麻木)。
实例包括：
a.臂丛神经炎
b.眼球后神经病，一种影响眼球正后方视神经部分的炎症过程。
c.视神经病，一种影响视神经的炎症过程，导致受影响眼睛的视力突然下降。视神经炎的病因尚不清楚。视神经(连接眼睛和大脑的神经)的突然炎症导致髓鞘肿胀和破坏。炎症有时可以是病毒感染的结果，或者它也可以是由自身免疫性疾病如多发性硬化引起。风险因素与可能的病因有关。
d.前庭神经炎，一种引起影响前庭神经的炎症过程的病毒感染。
G.关节炎症
关节炎症，如由滑囊炎或肌腱炎引起的关节炎症，可以引起慢性炎性疼痛。
H.晒伤和/或UV引起的损伤
本发明的多肽可用于治疗由以下任何头痛病症引起或与之相关的疼痛。头疼痛(医学上称为头痛(cephalgia))是一种头部轻度至重度疼痛的情况；有时颈部或上背部疼痛也可以解释为头痛。它可以指示潜在的局部或系统性疾病，或者本身就是一种疾病。
A.肌肉性/肌源性头痛
肌肉性/肌源性头痛似乎涉及面部和颈部肌肉的收紧或紧张；它们可以辐射到前额。紧张性头痛是肌源性头痛最常见的形式。
紧张性头痛是一种涉及头部、头皮或颈部疼痛或不适的病症，通常与这些区域中的肌肉紧张有关。紧张性头痛由颈部和头皮肌肉的收缩引起。这种肌肉收缩的一个原因是对压力、抑郁或焦虑的反应。任何导致头部长时间保持在一个位置而不动的活动都可以导致头痛。这些活动包括打字或使用电脑、手工精细作业和使用显微镜。在寒冷的房间里睡觉或脖子处于异常位置睡觉也可以引发这种类型的头痛。紧张型头痛包括但不限于发作性紧张性头痛和慢性紧张性头痛。
B.血管性头痛
血管性头痛最常见的类型是偏头痛。其他类型的血管性头痛包括引起反复剧烈疼痛的丛集性头痛，以及由高血压引起的头痛。
1.偏头痛
偏头痛(migraine)是一种异质性疾病，通常涉及反复发作的头痛。偏头痛与其他头痛不同，因为偏头痛伴有其他症状，如恶心、呕吐或对光敏感。对大多数人来说，抽痛只在一侧头部感觉到。临床特征，如先兆症状的类型、前驱症状的存在或眩晕等相关症状，可以出现在具有不同潜在病理生理和遗传机制的受试者亚组中。偏头痛包括但不限于无先兆偏头痛(普通偏头痛)、有先兆偏头痛(经典偏头痛)、月经期偏头痛、偏头痛等位发作(头痛)、复杂型偏头痛、腹部偏头痛和混合紧张型偏头痛。
2.丛集性头痛
丛集性头痛(Cluster headaches)影响一侧头部(单侧)，可以与眼睛流泪和鼻塞有关。它们呈集群状发生，每天在同一时间重复发生数周，然后消退。
D.高血压性头痛
E.牵引性和炎性头痛
牵引性和炎性头痛通常是从中风到鼻窦感染的其他障碍的症状。
F.激素性头痛
G.反弹性头痛
反弹性头痛，也称为药物过度使用性头痛，发生在药物服用过于频繁而无法缓解头痛的情况下。反弹性头痛每天都会频繁发生，而且可以非常痛苦。
H.慢性鼻窦炎性头痛
鼻窦炎是鼻旁窦的细菌性、真菌性、病毒性、过敏性或自身免疫性炎症。慢性鼻窦炎是普通感冒最常见的并发症之一。症状包括：鼻腔充血；面部疼痛；头痛；发烧；全身不适；浓绿色或黄色分泌物；弯腰后面部“饱胀感”加重。在少数情况下，慢性上颌窦炎也可以由牙齿感染引起的细菌传播引起。慢性增生性嗜酸性鼻窦炎是一种非感染性的慢性鼻窦炎。
I.器质性头痛
J.发作性头痛
发作性头痛是与癫痫活动相关的头痛。
本发明的多肽可用于治疗由以下任何躯体疼痛病症引起或以其他方式与之相关的疼痛。躯体疼痛源于韧带、肌腱、骨骼、血管、甚至神经本身。它是用体细胞伤害感受器检测到的。这些区域中疼痛受体的缺乏产生比皮肤疼痛持续时间更长的钝性、局部性差的疼痛；例如包括扭伤和骨折。其他实例包括以下。
A.肌肉过度紧张
肌肉过度紧张可以由例如扭伤或拉伤引起。
B.重复性运动障碍
重复性运动障碍可以由过度使用手、手腕、肘部、肩部、颈部、背部、臀部、膝盖、脚部、腿部或脚踝造成。
C.肌肉障碍
引起躯体疼痛的肌肉障碍包括例如多发性肌炎、皮肌炎、狼疮、纤维肌痛、风湿性多肌痛和横纹肌溶解。
D.肌痛
肌痛是肌肉疼痛，是许多疾病和障碍的症状。肌痛最常见的病因是过度使用或过度拉伸肌肉或肌肉群。没有外伤史的肌痛通常由病毒感染引起。长期肌痛可以指示代谢性肌病、某些营养缺乏或慢性疲劳综合征。
E.感染
感染可以引起身体疼痛。这种感染的实例包括例如肌肉脓肿、旋毛虫病、流感、莱姆病、疟疾、落基山斑疹热、禽流感、普通感冒、社区获得性肺炎、脑膜炎、猴痘、严重急性呼吸综合征、中毒性休克综合征、旋毛虫病、伤寒和上呼吸道感染。
F.药物
药物可以引起身体疼痛。这类药物包括例如可卡因、用于降低胆固醇的他汀类药物(如阿托伐他汀、辛伐他汀和洛伐他汀)和用于降低血压的ACE抑制剂(如依那普利和卡托普利)
本发明的多肽可用于治疗由以下任何内脏疼痛病症引起或与之相关的疼痛。内脏疼痛源于身体的内脏或器官。内脏伤害感受器位于身体器官和内腔内。这些区域中的伤害感受器更加稀缺，产生的疼痛通常比身体疼痛更疼痛，持续时间更长。内脏疼痛极难定位，内脏组织的几种损伤表现出“牵涉性”疼痛，其中感觉定位在与损伤部位完全无关的区域。内脏疼痛的实例包括以下。
A.功能性内脏疼痛
功能性内脏疼痛包括例如肠易激综合征和慢性功能性腹痛(CFAP)、功能性便秘和功能性消化不良、非心脏性胸痛(NCCP)和慢性腹痛。
B.慢性胃肠道炎症
慢性胃肠道炎症包括例如胃炎、炎性肠病，例如克罗恩病、溃疡性结肠炎、微小性结肠炎、憩室炎和肠胃炎；间质性膀胱炎；肠缺血；胆囊炎；阑尾炎；胃食管反流；溃疡、肾结石、尿路感染、胰腺炎和疝气。
C.自身免疫性疼痛
自身免疫性疼痛包括例如结节病和血管炎。
D.器质性内脏疼痛
器质性内脏疼痛包括例如由肠道的创伤性、炎症性或退行性病变引起或由肿瘤碰撞感觉神经支配产生的疼痛。
E.治疗引起的内脏疼痛
治疗引起的内脏疼痛包括例如化疗引起的疼痛或放射治疗引起的疼痛。
本发明的多肽可用于治疗由以下提及的任何疼痛病症引起或与之相关的疼痛。
牵涉性疼痛(referred pain)源自定位至与疼痛刺激部位分离的疼痛。通常，在神经在其起源处或附近受到压迫或损伤时出现牵涉性疼痛。在这种情况下，疼痛感通常会在神经所服务的区域感受到，即使损伤来自其他地方。一个常见的实例发生在椎间盘突出症中，其中源自脊髓的神经根被相邻的椎间盘材料压迫。虽然疼痛可以源自由受损的椎间盘本身，但在被压迫的神经所服务的区域(例如大腿、膝盖或脚)也会感觉到疼痛。只要没有发生永久性神经损伤，减轻神经根上的压力可以减轻牵涉性疼痛。心肌缺血(心肌组织的一部分的血流量损失)可能是牵涉性疼痛最著名的实例；这种感觉可以作为一种受限的感觉出现在上胸部，或者作为疼痛出现在左肩、手臂或甚至手部。
本发明的多肽可以用于治疗术后疼痛。
术后(例如手术后)疼痛是由外科手术引起的一种不适感觉。术后疼痛可以由切口、手术本身、伤口闭合以及手术过程中施加的任何力对组织造成的损伤引起。术后疼痛(例如手术后疼痛)也可以源于伴随手术的因素。例如，受试者可以由于其在手术台上的定位方式而遭受背痛，或者可以由于胸部区域的切口而遭受胸痛。全身麻醉后也可以出现喉咙痛，因为插入呼吸管可以引起刺激。然而，最常见的是手术切口切入皮肤和肌肉引起的术后疼痛。
例如，手术过程(或者更具体而言，手术切口)可以代表引起疼痛的“伤害性刺激”。伤害性刺激(可引起组织损伤的刺激)可激活伤害性传入末梢神经递质的释放，以及感觉末梢神经肽如P物质和降钙素基因相关肽(CGRP)的释放。然后，伤害性信息从外周神经系统传导到中枢神经系统，在那里疼痛被该个体感知。
术后疼痛可以由炎症和神经组织损伤共同引起。例如，响应组织损伤的活化肥大细胞的脱颗粒可以导致多种物质的释放，包括蛋白酶、细胞因子、血清素和细胞外空间。这些物质可以使初级传入神经元敏感(在较低阈值下激活)，以产生疼痛超敏反应。由于组织受到广泛的神经支配，身体的任何部位都容易受到手术引起的神经损伤。
手术是指涉及治疗受试者的损伤或疾病的医疗程序，包括对身体的一部分进行切口(任选地移除或修复身体的损伤部分)。尽管侵入性水平(例如所需手术切口的水平)可以因手术类型而异，但一旦手术完成，具有导致受试者疼痛的侵入性水平的手术也旨在包括在内。
手术可以包括对皮肤和/或筋膜和/或肌肉的切口。优选地，该手术包括对皮肤的切口。
手术不限于可以由医生进行的手术，还包括例如牙科手术。手术的非限制性实例包括阑尾切除术、乳房活检、隆胸或缩小、整容、胆囊切除术、冠状动脉搭桥术、清创术(例如伤口、烧伤或感染)、皮肤移植、器官移植和扁桃体切除术。
优选地，“术后”可以指手术后(例如手术后)至多一天开始的时间段。换言之，术语“术后”可以指手术后不超过一天开始的时间段。例如，术语“术后”可以指手术后1-20小时、任选地手术后2-15小时、任选地手术后5-10小时开始的时间点。这种时间可以代表在对受试者施用的外科麻醉剂的镇痛作用减弱(例如逐渐减弱)，从而受试者开始感知疼痛的时间界面开始的时间段。
此外，术语“术后”可以与术语“手术后”互换使用，因为“手术”在本文中的含义是“手术”。
类似地，术语“术后疼痛”可以指在手术后(例如手术后)至多一天开始的时间段内感知到(或更具体而言，开始感知到)的疼痛。换言之，术语“术后疼痛”可以指受试者在手术后不超过一天开始的时间段内感知到的疼痛。例如，术语“术后疼痛”可以指在手术后1-20小时、任选地在手术后2-15小时、任选地在手术后5-10小时开始的时间段内感知到的疼痛。
该时间段可以是手术后1-50周、例如5-45周、10-40周或10-35周。
这与术语“围手术期”形成了对比，该术语可以指例如受试者正在接受手术时或手术前后的时间段(例如受试者在手术室时的时间)，适宜地，是指从手术前至少1小时开始和/或在手术后不到1小时结束的时间段。
本发明涉及广泛的疼痛病症，例如慢性疼痛病症。在一些实施方案中，本发明的多肽用于治疗癌性和非癌性疼痛。
优选地，本发明的多肽用于治疗神经病性疼痛。神经病性疼痛可以是急性的或慢性的。在一个实施方案中，神经病性疼痛是损伤诱导的神经病性疼痛(与损伤相关的神经病性疼痛)。在一个实施方案中，神经病性疼痛是化疗诱导的神经病性疼痛(与化疗相关的神经病性疼痛)。
优选地，本发明的多肽用于治疗炎性疼痛。炎性疼痛可以是急性的或慢性的。在一个实施方案中，炎性疼痛可以是烧伤。例如，炎性疼痛可以由UV损伤引起(例如UV-B损伤)。
最优选地，本发明的多肽用于治疗膀胱疼痛综合征、幻肢疼痛或偏头痛。膀胱疼痛综合征可以由间质性膀胱炎引起或与之相关。
治疗疼痛优选意指减轻疼痛。换言之，在一个实施方案中，施用本发明的多肽减轻了受试者的疼痛。
更详细地说，(就疼痛而言)提及“减轻”优选指与受试者在施用之前(给药前)感知到的疼痛水平相比，受试者在施用本发明的多肽后(施用后)感知到的疼痛水平较低。例如，相对于施用前，施用后感知到的疼痛水平可以降低至少15％、25％、35％、45％、55％、65％、75％、85％或95％。例如，施用后感知到的疼痛水平可以降低至少75％、优选至少85％、更优选至少95％。
本领域技术人员已知用于评估疼痛感知的多种方法。例如，如Pogatzki-Zahn等(Pain Rep.2017年3月；2(2):e588)所述，在人类疼痛研究中常规使用机械性异常性疼痛的评价(静态或动态)，其在此引入作为参考。
用于评估受试者疼痛感知的适宜(但不是限制性的)方法包括以下：数值评定量表(NRS)评分；尽管本领域技术人员知道可以额外地或备选地使用的其他方法，如感觉阈值、疼痛感知阈值、静态机械性异常性疼痛、动态机械性异常性疼痛、时间总和、压力疼痛阈值、条件性疼痛调节和温度阈值。
疼痛感知测量的其他非限制性实例包括：在每个预定时间点SF-36评分与基线相比的变化；研究期间服用的抢救药物量以及首次服用抢救药物的时间。这些可以视为“探索性”终点或疼痛感知评估措施。
因此，在一个优选实施方案中，在施用本发明的多肽之后，疼痛感知可以通过以下中的一个或多个来评估：(a)数值评定量表(NRS)；(b)刺激诱发的NRS；(c)疼痛区域的温度；(d)疼痛区域的大小；(e)镇痛作用开始的时间；(f)峰值镇痛作用；(g)至镇痛作用达到峰值的时间；(h)镇痛作用持续时间；以及(i)SF-36生活质量。
本领域技术人员知道这类用于评估疼痛感知的方法。为了方便起见，下面提供数值评定评分和生活质量问卷简表-36的进一步描述。
数值评定量表(NRS)：根据本发明的疼痛感知通常使用数值评定量表(NR)。NRS是一个评估受试者疼痛感知的11分量表。受试者被要求给出一个0到10之间的数字，这个数字最适合他们的疼痛强度。零表示“完全没有疼痛”，而上限10表示“最严重的疼痛”。
NRS可用于评估疼痛的许多方面，包括自发平均疼痛、自发最严重疼痛和自发当前疼痛。通过要求受试者选择最能描述受试者在一段时间内(例如至少6小时、12小时、24小时或至少48小时)的平均疼痛(例如感知的疼痛)的数字来评估自发平均疼痛。通过要求受试者选择最能描述受试者在指定时间段(例如至少之前6小时、12小时、24小时或之前48小时)内最严重的疼痛的数字来评估自发最严重疼痛。通过要求受试者选择最能描述受试者在评估时疼痛程度的数字来评估自发当前疼痛。
NRS还可用于评估受试者响应多种不同刺激的疼痛感知。为了评估响应刺激的疼痛感知，受试者将接受施加在疼痛区域的多种性质的刺激。受试者将被问及他们在给药前和给药后的当前NRS评分。
所使用的刺激的实例包括：(i)轻度触摸(其可以通过在应用本文所述的冯-弗雷纤维丝之后测量径向辐条上疼痛区域表面上的疼痛来评估)；(ii)压力(压力疼痛阈值)，其可以通过要求受试者在使用压痛计施加越来越大的压力时给出NRS评分来评估；以及(iii)温度(可以通过使用施加到疼痛区域的热电极向受试者询问温暖、寒冷和热刺激的NRS评分来评估)。
优选地，与施用前受试者的NRS评分相比，本发明多肽的施用降低了受试者施用后的NRS评分(例如从≥7的评分降低到≤6的评分)。
生活质量问卷简表-36(SF-36)：SF-36生活质量问卷可用于评估个体的疼痛感知。SF-36是一个36项的受试者报告的受试者健康调查。SF-36由八个量表评分组成(活力、身体功能、身体疼痛、一般健康感知、身体角色功能、情感角色功能、社会角色功能和心理健康)。假设每个问题的权重相等，则将每个量表直接转换为0-100量表。SF-36评分越高，残疾程度越低。
在用于治疗疼痛的临床试验中通常测试的相关参数是本领域已知的，并且可以由本领域普通技术人员容易地选择。这些参数的实例包括但不限于NRS；刺激诱发NRS；疼痛区域的温度；疼痛区域的大小；镇痛作用开始的时间；峰值镇痛作用；至镇痛作用达到峰值的时间；镇痛作用持续时间；和/或如本文所述的SF-36生活质量。用于评估这些参数的方法也是本领域已知的，并且可以由普通技术人员使用常规方法和程序来进行。
优选地，与施用前受试者的SF-36评分相比，施用本发明的多肽增加了施用后受试者的SF-36评分(例如从≤50的评分增加到≥50的评分)。
通过本发明的多肽治疗的炎性疾病可以是神经系统、心血管系统、呼吸系统、消化系统、表皮系统、肌肉骨骼系统、泌尿系统、生殖系统、内分泌系统或淋巴系统的炎性疾病。
神经系统的炎性疾病可以是选自以下的一种或多种：中枢神经系统炎症(例如脑炎、脊髓炎、脑膜炎或蛛网膜炎)、外周神经系统炎症(例如神经炎)、眼部炎症(例如泪腺炎、巩膜炎、表层巩膜炎、角膜炎、视网膜炎、脉络膜视网膜炎、睑炎、结膜炎或葡萄膜炎)和耳朵炎症(例如外耳炎、中耳炎、迷路炎和乳突炎)。
心血管系统的炎性疾病可以是选自以下的一种或多种：心脏炎(例如心内膜炎、心肌炎或心包炎)和血管炎(例如动脉炎、静脉炎或毛细血管炎)。
呼吸系统的炎性疾病可以是选自以下的一种或多种：上呼吸系统炎性疾病(例如鼻窦炎、鼻炎、咽炎或喉炎)、下呼吸系统炎性疾病(例如气管炎、支气管炎、细支气管炎、肺炎或胸膜炎)和纵隔炎。
消化系统的炎性疾病可以是选自以下的一种或多种：口腔炎症(例如口炎、龈炎、龈口炎、舌炎、扁桃体炎、涎腺炎/腮腺炎、唇炎、牙髓炎或颌炎)、胃肠道炎症(例如食道炎、胃炎、肠胃炎、肠炎、结肠炎、小肠结肠炎、十二指肠炎、回肠炎、盲肠炎、阑尾炎或直肠炎)和副消化器官炎症(例如肝炎、上行性胆管炎、胆囊炎、胰腺炎或腹膜炎)。
表皮系统的炎性疾病可以选自以下一种或多种：皮炎(例如毛囊炎)、蜂窝组织炎和汗腺炎。
肌肉骨骼系统的炎性疾病可以是选自以下的一种或多种：关节炎、皮肌炎、软组织炎症(例如肌炎、滑膜炎/腱鞘炎、滑囊炎、起止点炎、筋膜炎、囊炎、上髁炎、腱炎或脂膜炎)、骨软骨炎、骨炎/骨髓炎、脊椎炎、骨膜炎和软骨炎。
泌尿系统的炎性疾病可以是选自以下的一种或多种：肾炎(例如肾小球肾炎或肾盂肾炎)、输尿管炎、膀胱炎和尿道炎。
生殖系统的炎性疾病可以是选自以下的一种或多种：女性生殖系统炎症(例如卵巢炎、输卵管炎、子宫内膜炎、子宫旁炎、宫颈炎、阴道炎、外阴炎或乳腺炎)、男性生殖系统炎症(例如睾丸炎、附睾炎、前列腺炎、精囊炎、龟头炎、包皮炎或龟头包皮炎)以及与妊娠、分娩和/或新生儿相关的炎症(例如绒毛膜羊膜炎、脐带炎或脐炎)。
内分泌系统的炎性疾病可以是选自以下的一种或多种：胰岛炎、垂体炎、甲状腺炎、甲状旁腺炎和肾上腺炎。
淋巴系统的炎性疾病可以是选自以下的一种或多种：淋巴管炎和淋巴结炎。
优选地，炎性疾病是选自以下的一种或多种：复杂性区域疼痛综合征、子宫内膜异位症、类风湿性关节炎、膀胱炎和神经炎。膀胱炎优选间质性膀胱炎。神经炎优选外周神经炎。
与本发明的多种治疗用途相关的实施方案旨在同等地应用于治疗方法，反之亦然。
序列同源性
多种序列比对方法中的任何一种都可以用于确定百分比同一性，包括但不限于全局方法、局部方法和混合方法，例如分段方法。确定百分比同一性的方案是本领域技术人员范围内的常规程序。全局方法从分子的开始到结束比对序列，并通过将单个残基对的分数相加和施加缺口罚分来确定最佳比对。非限制性方法包括例如CLUSTAL W，参见例如JulieD.Thompson等,CLUSTAL W:Improving the Sensitivity of Progressive MultipleSequence Alignment Through Sequence Weighting,Position-Specific Gap Penaltiesand Weight Matrix Choice,22(22)Nucleic Acids Research 4673-4680(1994)；以及迭代细化，参见例如Osamu Gotoh,Significant Improvement in Accuracy of MultipleProtein.Sequence Alignments by Iterative Refinement as Assessed by Referenceto Structural Alignments,264(4)J.MoI.Biol.823-838(1996)。局部方法通过鉴定所有输入序列共享的一个或多个保守基序来比对序列。非限制性方法包括例如Match box，参见例如Eric Depiereux和Ernest Feytmans,Match-Box:A Fundamentally New Algorithmfor the Simultaneous Alignment of Several Protein Sequences,8(5)CABIOS 501 -509(1992)；Gibbs采样，参见例如C.E.Lawrence等,Detecting Subtle Sequence Signals:A Gibbs Sampling Strategy for Multiple Alignment,262(5131)Science 208-214(1993)；Align-M，参见例如Ivo Van WaIIe等,Align-M-A New Algorithm for MultipleAlignment of Highly Divergent Sequences,20(9)Bioinformatics:1428-1435(2004)。
因此，百分比序列同一性是通过常规方法确定的。参见例如Altschul等,Bull.Math.Bio.48:603-16,1986以及Henikoff和Henikoff,Proc.Natl.Acad.Sci.USA 89:10915-19,1992。简言之，使用缺口开放罚分10、缺口延伸罚分1以及Henikoff和Henikoff(同上)的“blosum 62”评分矩阵对两个氨基酸序列进行比对，以优化比对得分，如下所示(氨基酸表示为标准单字母代码)；优选该方法用于将序列与本文所述的SEQ ID NO比对以定义本文所述氨基酸位置编号。
两个或多个核酸或氨基酸序列之间的“百分比序列同一性”是序列共享的相同位置数量的函数。因此，％同一性可以计算为相同核苷酸/氨基酸的数量除以核苷酸/氨基酸的总数，再乘以100。％序列同一性的计算还可以考虑缺口的数量，以及需要引入以优化两个或多个序列的比对的每个缺口的长度。两个或多个序列之间的序列比较和百分比同一性确定可以使用特定的数学算法来进行，如BLAST，这对于技术人员来说是熟悉的。
用于确定序列同一性的比对分值
然后，将百分比同一性计算为：
基本上同源的多肽的特征在于具有一个或多个氨基酸取代、缺失或添加。这些变化优选具有较小的性质，即，保守氨基酸取代(见下文)和不显著影响多肽折叠或活性的其他取代；小的缺失，通常为1至约30个氨基酸；以及小的氨基或羧基端延伸，如氨基端甲硫氨酸残基、多至约20-25个残基的小接头肽或亲和标签。
保守氨基酸取代
碱性氨基酸：精氨酸
赖氨酸
组氨酸
酸性氨基酸：谷氨酸
天冬氨酸
极性氨基酸：谷氨酰胺
天冬酰胺
疏水性氨基酸：亮氨酸
异亮氨酸
缬氨酸
芳香族氨基酸：苯丙氨酸
色氨酸
酪氨酸
小氨基酸：甘氨酸
丙氨酸
丝氨酸
苏氨酸
甲硫氨酸
除了20种标准氨基酸之外，非标准氨基酸(如4-羟基脯氨酸、6-N-甲基赖氨酸、2-氨基异丁酸、异缬氨酸和α-甲基丝氨酸)可以取代本发明的多肽的氨基酸残基。有限数量的非保守氨基酸、非遗传密码编码的氨基酸和非天然氨基酸可以取代多肽氨基酸残基。本发明的多肽还可以包含非天然存在的氨基酸残基。
非天然存在的氨基酸包括但不限于反式-3-甲基脯氨酸、2,4-甲基脯氨酸、顺式-4-羟基脯氨酸、反式-4-羟基脯氨酸、N-甲基甘氨酸、别苏氨酸、甲基苏氨酸、羟乙基半胱氨酸、羟乙基同型半胱氨酸、硝基谷氨酰胺、同型谷氨酰胺、六氢吡啶羧酸、叔亮氨酸、正缬氨酸、2-氮杂苯丙氨酸、3-氮杂苯丙氨酸、4-氮杂苯丙氨酸和4-氟苯丙氨酸。本领域已知几种将非天然存在的氨基酸残基掺入蛋白质中的方法。例如，可以使用体外系统，其中使用化学氨酰化的抑制tRNA抑制无义突变。用于合成氨基酸和氨酰化tRNA的方法是本领域已知的。含有无义突变的质粒的转录和翻译在包含大肠杆菌S30提取物以及市售酶和其他试剂的无细胞系统中进行。蛋白质通过层析纯化。参见例如Robertson等,J.Am.Chem.Soc.113:2722,1991；Ellman等,Methods Enzymol.202:301,1991；Chung等,Science 259:806-9,1993；以及Chung等,Proc.Natl.Acad.Sci.USA90:10145-9,1993)。在第二种方法中，通过显微注射突变的mRNA和化学氨酰化的抑制tRNA在非洲爪蟾卵母细胞中进行翻译(Turcatti等,J.Biol.Chem.271:19991-8,1996)。在第三种方法中，将大肠杆菌细胞在不存在要取代的天然氨基酸(例如苯丙氨酸)和存在所希望的非天然存在的氨基酸(例如2-氮杂苯丙氨酸、3-氮杂苯丙氨酸、4-氮杂苯丙氨酸或4-氟苯丙氨酸)的情况下培养。非天然存在的氨基酸代替其天然对应物掺入到多肽中。参见Koide等,Biochem.33:7470-6,1994。天然存在的氨基酸残基可以通过体外化学修饰转化为非天然存在的类别。化学修饰可以与定点诱变相结合，以进一步扩大取代的范围(Wynn和Richards,Protein Sci.2:395-403,1993)。
有限数量的非保守氨基酸、非遗传密码编码的氨基酸、非天然存在的氨基酸和非天然氨基酸可以取代本发明的多肽的氨基酸残基。
本发明的多肽中的必需氨基酸可以根据本领域已知的操作确定，例如定点诱变或丙氨酸扫描诱变(Cunningham和Wells,Science 244:1081-5,1989)。生物相互作用的位点也可以通过结构的物理分析来确定，如通过核磁共振、晶体学、电子衍射或光亲和标记等技术，结合假定的接触位点氨基酸的突变来确定。参见例如de Vos等,Science 255:306-12,1992；Smith等,J.Mol.Biol.224:899-904,1992；Wlodaver等,FEBS Lett.309:59-64,1992。还可以通过分析与本发明的多肽的相关组分(例如转运组分或蛋白酶组分)的同源性来推断必需氨基酸的身份。
可以使用已知的诱变和筛选方法，例如Reidhaar-Olson和Sauer(Science 241:53-7,1988)或Bowie和Sauer(Proc.Natl.Acad.Sci.USA 86:2152-6,1989)中所公开的那些方法来制备和测试多个氨基酸取代。简言之，这些作者公开了同时随机化多肽中的两个或多个位置，筛选功能性多肽，然后对诱变的多肽进行测序以确定每个位置上可允许的取代谱的方法。可以使用的其他方法包括噬菌体展示(例如Lowman等,Biochem.30:10832-7,1991；Ladner等,美国专利号5,223,409；Huse,WIPO公开号WO 92/06204)和区域定向诱变(Derbyshire等,Gene 46:145,1986；Ner等,DNA 7:127,1988)。
除非另有定义，否则本文中使用的所有技术和科学术语具有与本公开所属领域的普通技术人员通常理解的含义相同的含义。Singleton等,DICTIONARY OF MICROBIOLOGYAND MOLECULAR BIOLOGY,20ED.,John Wiley and Sons,New York(1994)以及Hale&Marham,THE HARPER COLLINSDICTIONARY OF BIOLOGY,Harper Perennial,NY(1991)为本领域技术人员提供了本公开中使用的许多术语的通用词典。
本公开不受本文公开的示例性方法和材料的限制，并且与本文描述的方法和材料类似或等同的任何方法和材料都可以用于本公开的实施例的实施或测试。数值范围包括定义该范围的数字。除非另有说明，否则任何核酸序列都以5'到3'的方向从左到右书写；氨基酸序列以氨基到羧基的方向从左到右书写。
本文提供的标题不是对本公开的多个方面或实施方案的限制。
在本文中使用氨基酸的名称、三字母缩写或单字母缩写来指代氨基酸。本文所用的术语“蛋白质”包括蛋白质、多肽和肽。本文所用的术语“氨基酸序列”与术语“多肽”和/或术语“蛋白质”同义。在某些情况下，术语“氨基酸序列”与术语“肽”同义。在一些情况下，术语“氨基酸序列”与术语“酶”同义。术语“蛋白质”和“多肽”在本文中可互换使用。在本公开和权利要求书中，可以使用常规的氨基酸残基的单字母和三字母代码。根据IUPACIUB生物化学命名联合委员会(JCBN)定义的氨基酸的三字母代码。还应理解，由于遗传密码的简并性，多肽可以由一个以上的核苷酸序列编码。
术语的其他定义可以出现在整个说明书中。在更详细地描述示例性实施方案之前，应理解，本公开不限于所描述的具体实施方案，并且因此可以变化。还应理解，此处使用的术语仅用于描述具体实施方案的目的，而不旨在限制，因为本公开的范围将仅由所附权利要求书限定。
在提供值的范围的情况下，应当理解，除非上下文另有明确规定，否则还具体公开了该范围的上限和下限之间的每个中间值，至下限单位的十分之一。在所述范围内的任何所述值或中间值与所述范围内的任何其他所述或中间值之间的每个较小范围都包含在本公开内。这些较小范围的上限和下限可以独立地包括在该范围内或在该范围内排除，并且在较小范围中包括任一个，没有一个或者两个极限的每个范围也包括在本公开内容中，属于规定的范围中任何特别排除的极限。在所述范围包括一个或两个极限的情况下，排除那些包括的极限之一或两者的范围也包括在本公开中。
必须注意的是，本文和所附权利要求中所使用的单数形式“一”、“一个”和“该”包括复数指称，除非上下文另有明确说明。因此，例如，提及“一种梭菌神经毒素”包括多种这样的候选物质，并且提及“该梭菌神经毒素”包括提及一种或多种梭菌神经毒素及其本领域技术人员已知的等同物，等。
提供本文所讨论的出版物仅为了其在本申请的提交日期之前的公开内容。本文中的任何内容都不应解释为承认此类出版物构成所附权利要求的现有技术。
附图简述
现在将参考以下附图和实施例，仅通过举例的方式描述本发明的实施方案。
图1显示施用BoNT/A或BoNT/A(0)的人神经元细胞中的％SNAP25切割。
图2显示施用BoNT/A或BoNT/A(0)的大鼠神经元细胞中的％SNAP25切割。
图3(A)显示尾巴悬吊的小鼠的特征性惊吓反应。(B)显示在趾外展评分(DigitAbduction Score，DAS)测定法中使用的评分。
图4(A)显示一项使用成年雄性Sprague-Dawley大鼠(220-250g)慢性神经病性疼痛的慢性挤压损伤(CCI)模型进行研究的实验示意图。显示BoNT/A(0)(60pg/kg足底内施用)、BoNT/A(30pg/kg足底内(i.pl.)施用)、BoNT/A(60pg/kg足底内施用)、溶媒(明胶磷酸盐缓冲液(GPB)足底内施用-阴性对照)或加巴喷丁(gabapentin)(100mg/kg口服(p.o.)施用-阳性对照)施用前和施用后的天数(D)。施用发生在第0天(D0)，CCI外科手术在D-14进行。vF表示在指定的日期进行冯-弗雷试验(von Frey test)。(B)显示对于如(A)中所述施用的动物，同侧爪(即，施用对照组合物、BoNT/A或BoNT/A(0)的爪)随时间的机械敏感性(通过冯-弗雷试验测量)。(C)显示对于如(A)中所述施用的动物，对侧爪(即，未施用对照组合物、BoNT/A或BoNT/A(0)的爪)随时间的机械敏感性(通过冯-弗雷试验测量)。(D)显示如(A)中所述施用的大鼠的体重随时间的变化。
图5(A)显示一项使用成年雄性Sprague-Dawley大鼠急性奥沙利铂(oxaliplatin)诱导的神经病性疼痛模型进行的研究的实验示意图。在第0天(D0)施用奥沙利铂(10mg/kg腹膜内(i.p.)施用)和BoNT/A(0)(1000pg/kg足底内施用)、BoNT/A(50pg/kg足底内施用)、BoNT/A(100pg/kg足底内施用)、BoNT/A(160pg/kg足底内施用)或溶媒(GPB足底内施用-阴性对照)。作为奥沙利铂处理的另一阴性对照，对一亚组大鼠腹膜内施用5％葡萄糖和GBP(足底内施用)。在D3试验前1小时施用度洛西汀(Duloxetine)(100mg/kg口服施用-阳性对照)。显示施用后的天数和小时数。PI表示爪浸泡(冷)试验在所指定的日期进行。(B)显示对于如(A)中所述施用的动物，同侧爪(即施用对照组合物、BoNT/A或BoNT/A(0)的爪)随时间的冷敏感性(通过爪浸泡试验测量)。(C)显示对于如(A)中所述施用的动物，对侧爪(即未施用对照组合物、BoNT/A或BoNT/A(0)的爪)随时间的冷敏感性(通过爪浸泡试验测量)。
图6(A)显示一项使用成年雄性Sprague-Dawley大鼠(180-210g)的慢性奥沙利铂诱导的神经病性疼痛模型进行的研究的实验示意图。奥沙利铂在第-2天(D-2)施用。在第0天(D0)施用BoNT/A(0)(100pg/kg足底内施用)、BoNT/A(100pg/kg足底内施用)或溶媒(GPB足底内施用-阴性对照)。第3天施用普瑞巴林(Pregabalin)(30mg/kg口服施用-阳性对照)。显示施用前和施用后的天数。vF和CP表明，冯-弗雷和冷板试验(分别)在所示日期进行。(B)显示对于如(A)中所述施用的动物，同侧爪(即施用对照组合物、BoNT/A或BoNT/A(0)的爪)随时间的机械敏感性(通过冯-弗雷试验测量)。(C)显示对于如(A)中所述施用的动物，对侧爪(即未施用对照组合物、BoNT/A或BoNT/A(0)的爪)随时间的机械敏感性(通过冯-弗雷试验测量)。(D)显示对于如(A)中所述施用的动物，随时间的热敏性(通过冷板试验测量)。
图7(A)显示一项使用成年雄性Wistar大鼠(180-210g)急性紫外线-B(UV-B)灼伤诱导的炎性疼痛模型进行的研究的实验示意图。在第0天(D0)施用BoNT/A(0)(100pg/kg足底内施用)、BoNT/A(100pg/kg足底内施用)或溶媒(GPB足底内施用-阴性对照)。在D3读数前1h施用吲哚美辛(Indomethacin)(5mg/kg口服施用-阳性对照)。大鼠在第1天(D1)暴露于UV-B(500mJ/cm2)。vF指示冯-弗雷试验在所示日期进行。(B)显示如(A)中所述施用的动物的机械敏感性(通过冯-弗雷试验测量)。
图8显示施用溶媒、催化活性的嵌合BoNT/XB(0.3ng/kg,n＝10)、催化活性的嵌合BoNT/XB(30ng/kg,n＝10)、无催化活性的嵌合BoNT/XB(0)(0.3ng/kg,n＝10)、无催化活性的嵌合BoNT/XB(0)(30ng/kg,n＝10)、BoNT/A(160pg/kg,n＝10)或吲哚美辛(10mg/kg,n＝9)的小鼠的机械敏感性(通过冯-弗雷试验测量)。显示未处理的动物(基线)、施用BoNT或溶媒后2天和施用完全弗氏佐剂(CFA)前(第0天CFA，第2天)以及施用CFA后1天(第1天CFA，第3天)的敏感性。**P＜0.1，***P＜0.01(重复测量双因素方差分析后与溶媒的Dunnett多重比较)。
序列表
当以下任何SEQ ID NO中示出起始Met氨基酸残基或相应的起始密码子的情况下，该残基/密码子是任选的。
SEQ ID NO:1-重组无催化活性BoNT/A(rBoNT/A(0))的核苷酸序列
SEQ ID NO:2-rBoNT/A(0)的多肽序列
SEQ ID NO:3-rLHN/A(仅轻链加转运结构域)的核苷酸序列
SEQ ID NO:4-rLHN/A的多肽序列
SEQ ID NO:5-rL/A(仅轻链)的核苷酸序列
SEQ ID NO:6-rL/A的多肽序列
SEQ ID NO:7-rHC/A的核苷酸序列
SEQ ID NO:8-rHC/A的多肽序列
SEQ ID NO:9-rBoNT/B(0)的核苷酸序列
SEQ ID NO:10-rBoNT/B(0)的多肽序列
SEQ ID NO:11-rBoNT/C(0)的核苷酸序列
SEQ ID NO:12-rBoNT/C(0)的多肽序列
SEQ ID NO:13-rBoNT/E(0)的核苷酸序列
SEQ ID NO:14-rBoNT/E(0)的多肽序列
SEQ ID NO:15-rBoNT/F(0)的核苷酸序列
SEQ ID NO:16-rBoNT/F(0)的多肽序列
SEQ ID NO:17-rBoNT/A(0)的核苷酸序列(His标签)
SEQ ID NO:18-rBoNT/A(0)的多肽序列(His标签)
SEQ ID NO:19-rLHN/A的核苷酸序列(His标签)
SEQ ID NO:20-rLHN/A的多肽序列(His标签)
SEQ ID NO:21-rHC/A的核苷酸序列(His标签)
SEQ ID NO:22-rHC/A的多肽序列(His标签)
SEQ ID NO:23-rLC/A的核苷酸序列(His标签)
SEQ ID NO:24-rLC/A的多肽序列(His标签)
SEQ ID NO:25-rBoNT/FA(0)的核苷酸序列(His标签)
SEQ ID NO:26-rBoNT/FA(0)的多肽序列(His标签)
SEQ ID NO:27-rLHN/FA的核苷酸序列(His标签)
SEQ ID NO:28-rLHN/FA的多肽序列(His标签)
SEQ ID NO:29-rHC/FA的核苷酸序列(His标签)
SEQ ID NO:30-rHC/FA的多肽序列(His标签)
SEQ ID NO:31-rLC/FA的核苷酸序列(His标签)
SEQ ID NO:32-rLC/FA的多肽序列(His标签)
SEQ ID NO:33-rBoNT/F(0)的核苷酸序列(His标签)
SEQ ID NO:34-rBoNT/F(0)的多肽序列(His标签)
SEQ ID NO:35-rLHN/F的核苷酸序列(His标签)
SEQ ID NO:36-rLHN/F的多肽序列(His标签)
SEQ ID NO:37-rHC/F的核苷酸序列(His标签)
SEQ ID NO:38-rHC/F的多肽序列(His标签)
SEQ ID NO:39-rLC/F的核苷酸序列(His标签)
SEQ ID NO:40-rLC/F的多肽序列(His标签)
SEQ ID NO:41-阳离子rHC/A的核苷酸序列(His标签)
SEQ ID NO:42-阳离子rHC/A的多肽序列(His标签)
SEQ ID NO:43-rHC/AB的核苷酸序列(His标签)
SEQ ID NO:44-rHC/AB的多肽序列(His标签)
SEQ ID NO:45-rHC/A变体Y1117V H1253K的核苷酸序列(His标签)
SEQ ID NO:46-rHC/A变体Y1117V H1253K的多肽序列(His标签)
SEQ ID NO:47-rHC/A变体Y1117V F1252Y H1253K L1278F的核苷酸序列(His标签)
SEQ ID NO:48-rHC/A变体Y1117V F1252Y H1253K L1278F的多肽序列(His标签)
SEQ ID NO:49-rHC/A变体Y1117V F1252Y H1253K L1278H的核苷酸序列(His标签)
SEQ ID NO:50-rHC/A变体Y1117V F1252Y H1253K L1278H的多肽序列(His标签)
SEQ ID NO:51-BoNT/A的多肽序列-UniProt P10845
SEQ ID NO:52-BoNT/B的多肽序列-UniProt P10844
SEQ ID NO:53-BoNT/C的多肽序列-UniProt P18640
SEQ ID NO:54-BoNT/D的多肽序列-UniProt P19321
SEQ ID NO:55-BoNT/E的多肽序列-UniProt Q00496
SEQ ID NO:56-BoNT/F的多肽序列-UniProt A7GBG3
SEQ ID NO:57-BoNT/G的多肽序列-UniProt Q60393
SEQ ID NO:58-TeNT的多肽序列-UniProt P04958
SEQ ID NO:59-BoNT/X的多肽序列
SEQ ID NO:60-未修饰BoNT/A1的多肽序列
SEQ ID NO:61-mrBoNT/AB(0)的多肽序列
SEQ ID NO:62-mrBoNT/A(0)的多肽序列
SEQ ID NO:63-BoNT/XB(0)的多肽序列(His标签)
SEQ ID NO:64-BoNT/XB(0)的多肽序列
SEQ ID NO:65-BoNT/XB(0)变体的多肽序列(His标签)
SEQ ID NO:66-BoNT/XB(0)变体的多肽序列
SEQ ID NO:67-BoNT/XA(0)的多肽序列
SEQ ID NO:68-BoNT/XA(0)变体的多肽序列
SEQ ID NO:69-BoNT/XD(0)的多肽序列
SEQ ID NO:70-BoNT/XF(0)的多肽序列
SEQ ID NO:71-C1激活环共有序列
SEQ ID NO:72-C1激活环
SEQ ID NO:73-C1激活环变体
SEQ ID NO:74-rLC/A(0)的多肽序列(His标签)
SEQ ID NO:75-rLHN/A(0)的多肽序列(His标签)
SEQ ID NO:76-rLC/X(0)的多肽序列
SEQ ID NO:77-前切割(PreScission)蛋白酶位点
SEQ ID NO:78-C1激活环变体2
SEQ ID NO:1-rBoNT/A(0)的核苷酸序列
ATGCCATTCGTCAACAAGCAATTCAACTACAAAGACCCAGTCAACGGCGTCGACATCGCATACATCAAGATTCCGAACGCCGGTCAAATGCAGCCGGTTAAGGCTTTTAAGATCCACAACAAGATTTGGGTTATCCCGGAGCGTGACACCTTCACGAACCCGGAAGAAGGCGATCTGAACCCGCCACCGGAAGCGAAGCAAGTCCCTGTCAGCTACTACGATTCGACGTACCTGAGCACGGATAACGAAAAAGATAACTACCTGAAAGGTGTGACCAAGCTGTTCGAACGTATCTACAGCACGGATCTGGGTCGCATGCTGCTGACTAGCATTGTTCGCGGTATCCCGTTCTGGGGTGGTAGCACGATTGACACCGAACTGAAGGTTATCGACACTAACTGCATTAACGTTATTCAACCGGATGGTAGCTATCGTAGCGAAGAGCTGAATCTGGTCATCATTGGCCCGAGCGCAGACATTATCCAATTCGAGTGCAAGAGCTTTGGTCACGAGGTTCTGAATCTGACCCGCAATGGCTATGGTAGCACCCAGTACATTCGTTTTTCGCCGGATTTTACCTTCGGCTTTGAAGAGAGCCTGGAGGTTGATACCAATCCGTTGCTGGGTGCGGGCAAATTCGCTACCGATCCGGCTGTCACGCTGGCCCATcAACTGATCtACGCAGGCCACCGCCTGTACGGCATTGCCATCAACCCAAACCGTGTGTTCAAGGTTAATACGAATGCATACTACGAGATGAGCGGCCTGGAAGTCAGCTTCGAAGAACTGCGCACCTTCGGTGGCCATGACGCTAAATTCATTGACAGCTTGCAAGAGAATGAGTTCCGTCTGTACTACTATAACAAATTCAAAGACATTGCAAGCACGTTGAACAAGGCCAAAAGCATCGTTGGTACTACCGCGTCGTTGCAGTATATGAAGAATGTGTTTAAAGAGAAGTACCTGCTGTCCGAGGATACCTCCGGCAAGTTTAGCGTTGATAAGCTGAAGTTTGACAAACTGTACAAGATGCTGACCGAGATTTACACCGAGGACAACTTTGTGAAATTCTTCAAAGTGTTGAATCGTAAAACCTATCTGAATTTTGACAAAGCGGTTTTCAAGATTAACATCGTGCCGAAGGTGAACTACACCATCTATGACGGTTTTAACCTGCGTAACACCAACCTGGCGGCGAACTTTAACGGTCAGAATACGGAAATCAACAACATGAATTTCACGAAGTTGAAGAACTTCACGGGTCTGTTCGAGTTCTATAAGCTGCTGTGCGTGCGCGGTATCATCACCAGCAAAACCAAAAGCCTGGACAAAGGCTACAACAAGGCGCTGAATGACCTGTGCATTAAGGTAAACAATTGGGATCTGTTCTTTTCGCCATCCGAAGATAATTTTACCAACGACCTGAACAAGGGTGAAGAAATCACCAGCGATACGAATATTGAAGCAGCGGAAGAGAATATCAGCCTGGATCTGATCCAGCAGTACTATCTGACCTTTAACTTCGACAATGAACCGGAGAACATTAGCATTGAGAATCTGAGCAGCGACATTATCGGTCAGCTGGAACTGATGCCGAATATCGAACGTTTCCCGAACGGCAAAAAGTACGAGCTGGACAAGTACACTATGTTCCATTACCTGCGTGCACAGGAGTTTGAACACGGTAAAAGCCGTATCGCGCTGACCAACAGCGTTAACGAGGCCCTGCTGAACCCGAGCCGTGTCTATACCTTCTTCAGCAGCGACTATGTTAAGAAAGTGAACAAAGCCACTGAGGCCGCGATGTTCCTGGGCTGGGTGGAACAGCTGGTATATGACTTCACGGACGAGACGAGCGAAGTGAGCACTACCGACAAAATTGCTGATATTACCATCATTATCCCGTATATTGGTCCGGCACTGAACATTGGCAACATGCTGTACAAAGACGATTTTGTGGGTGCCCTGATCTTCTCCGGTGCCGTGATTCTGCTGGAGTTCATTCCGGAGATTGCGATCCCGGTGTTGGGTACCTTCGCGCTGGTGTCCTACATCGCGAATAAGGTTCTGACGGTTCAGACCATCGATAACGCGCTGTCGAAACGTAATGAAAAATGGGACGAGGTTTACAAATACATTGTTACGAATTGGCTGGCGAAAGTCAATACCCAGATCGACCTGATCCGTAAGAAAATGAAAGAGGCGCTGGAGAATCAGGCGGAGGCCACCAAAGCAATTATCAACTACCAATACAACCAGTACACGGAAGAAGAGAAGAATAACATTAACTTCAATATCGATGATTTGAGCAGCAAGCTGAATGAATCTATCAACAAAGCGATGATCAATATCAACAAGTTTTTGAATCAGTGTAGCGTTTCGTACCTGATGAATAGCATGATTCCGTATGGCGTCAAACGTCTGGAGGACTTCGACGCCAGCCTGAAAGATGCGTTGCTGAAATACATTTACGACAATCGTGGTACGCTGATTGGCCAAGTTGACCGCTTGAAAGACAAAGTTAACAATACCCTGAGCACCGACATCCCATTTCAACTGAGCAAGTATGTTGATAATCAACGTCTGTTGAGCACTTTCACCGAGTATATCAAAAACATCATCAATACTAGCATTCTGAACCTGCGTTACGAGAGCAATCATCTGATTGATCTGAGCCGTTATGCAAGCAAGATCAACATCGGTAGCAAGGTCAATTTTGACCCGATCGATAAGAACCAGATCCAGCTGTTTAATCTGGAATCGAGCAAAATTGAGGTTATCCTGAAAAACGCCATTGTCTACAACTCCATGTACGAGAATTTCTCCACCAGCTTCTGGATTCGCATCCCGAAATACTTCAACAGCATTAGCCTGAACAACGAGTATACTATCATCAACTGTATGGAGAACAACAGCGGTTGGAAGGTGTCTCTGAACTATGGTGAGATCATTTGGACCTTGCAGGACACCCAAGAGATCAAGCAGCGCGTCGTGTTCAAGTACTCTCAAATGATCAACATTTCCGATTACATTAATCGTTGGATCTTCGTGACCATTACGAATAACCGTCTGAATAACAGCAAGATTTACATCAATGGTCGCTTGATCGATCAGAAACCGATTAGCAACCTGGGTAATATCCACGCAAGCAACAACATTATGTTCAAATTGGACGGTTGCCGCGATACCCATCGTTATATCTGGATCAAGTATTTCAACCTGTTTGATAAAGAACTGAATGAGAAGGAGATCAAAGATTTGTATGACAACCAATCTAACAGCGGCATTTTGAAGGACTTCTGGGGCGATTATCTGCAATACGATAAGCCGTACTATATGCTGAACCTGTATGATCCGAACAAATATGTGGATGTCAATAATGTGGGTATTCGTGGTTACATGTATTTGAAGGGTCCGCGTGGCAGCGTTATGACGACCAACATTTACCTGAACTCTAGCCTGTACCGTGGTACGAAATTCATCATTAAGAAATATGCCAGCGGCAACAAAGATAACATTGTGCGTAATAACGATCGTGTCTACATCAACGTGGTCGTGAAGAATAAAGAGTACCGTCTGGCGACCAACGCTTCGCAGGCGGGTGTTGAGAAAATTCTGAGCGCGTTGGAGATCCCTGATGTCGGTAATCTGAGCCAAGTCGTGGTTATGAAGAGCAAGAACGACCAGGGTATCACTAACAAGTGCAAGATGAACCTGCAAGACAACAATGGTAACGACATCGGCTTTATTGGTTTCCACCAGTTCAACAATATTGCTAAACTGGTAGCGAGCAATTGGTACAATCGTCAGATTGAGCGCAGCAGCCGTACTTTGGGCTGTAGCTGGGAGTTTATCCCGGTCGATGATGGTTGGGGCGAACGTCCGCTG
SEQ ID NO:2-rBoNT/A(0)的多肽序列
MPFVNKQFNYKDPVNGVDIAYIKIPNAGQMQPVKAFKIHNKIWVIPERDTFTNPEEGDLNPPPEAKQVPVSYYDSTYLSTDNEKDNYLKGVTKLFERIYSTDLGRMLLTSIVRGIPFWGGSTIDTELKVIDTNCINVIQPDGSYRSEELNLVIIGPSADIIQFECKSFGHEVLNLTRNGYGSTQYIRFSPDFTFGFEESLEVDTNPLLGAGKFATDPAVTLAHQLIYAGHRLYGIAINPNRVFKVNTNAYYEMSGLEVSFEELRTFGGHDAKFIDSLQENEFRLYYYNKFKDIASTLNKAKSIVGTTASLQYMKNVFKEKYLLSEDTSGKFSVDKLKFDKLYKMLTEIYTEDNFVKFFKVLNRKTYLNFDKAVFKINIVPKVNYTIYDGFNLRNTNLAANFNGQNTEINNMNFTKLKNFTGLFEFYKLLCVRGIITSKTKSLDKGYNKALNDLCIKVNNWDLFFSPSEDNFTNDLNKGEEITSDTNIEAAEENISLDLIQQYYLTFNFDNEPENISIENLSSDIIGQLELMPNIERFPNGKKYELDKYTMFHYLRAQEFEHGKSRIALTNSVNEALLNPSRVYTFFSSDYVKKVNKATEAAMFLGWVEQLVYDFTDETSEVSTTDKIADITIIIPYIGPALNIGNMLYKDDFVGALIFSGAVILLEFIPEIAIPVLGTFALVSYIANKVLTVQTIDNALSKRNEKWDEVYKYIVTNWLAKVNTQIDLIRKKMKEALENQAEATKAIINYQYNQYTEEEKNNINFNIDDLSSKLNESINKAMININKFLNQCSVSYLMNSMIPYGVKRLEDFDASLKDALLKYIYDNRGTLIGQVDRLKDKVNNTLSTDIPFQLSKYVDNQRLLSTFTEYIKNIINTSILNLRYESNHLIDLSRYASKINIGSKVNFDPIDKNQIQLFNLESSKIEVILKNAIVYNSMYENFSTSFWIRIPKYFNSISLNNEYTIINCMENNSGWKVSLNYGEIIWTLQDTQEIKQRVVFKYSQMINISDYINRWIFVTITNNRLNNSKIYINGRLIDQKPISNLGNIHASNNIMFKLDGCRDTHRYIWIKYFNLFDKELNEKEIKDLYDNQSNSGILKDFWGDYLQYDKPYYMLNLYDPNKYVDVNNVGIRGYMYLKGPRGSVMTTNIYLNSSLYRGTKFIIKKYASGNKDNIVRNNDRVYINVVVKNKEYRLATNASQAGVEKILSALEIPDVGNLSQVVVMKSKNDQGITNKCKMNLQDNNGNDIGFIGFHQFNNIAKLVASNWYNRQIERSSRTLGCSWEFIPVDDGWGERPL
SEQ ID NO:3-rLHN/A的核苷酸序列
atggagttcgttaacaaacagttcaactataaagacccagttaacggtgttgacattgcttacatcaaaatcccgaacgctggccagatgcagccggtaaaggcattcaaaatccacaacaaaatctgggttatcccggaacgtgatacctttactaacccggaagaaggtgacctgaacccgccaccggaagcgaaacaggtgccggtatcttactatgactccacctacctgtctaccgataacgaaaaggacaactacctgaaaggtgttactaaactgttcgagcgtatttactccaccgacctgggccgtatgctgctgactagcatcgttcgcggtatcccgttctggggcggttctaccatcgataccgaactgaaagtaatcgacactaactgcatcaacgttattcagccggacggttcctatcgttccgaagaactgaacctggtgatcatcggcccgtctgctgatatcatccagttcgagtgtaagagctttggtcacgaagttctgaacctcacccgtaacggctacggttccactcagtacatccgtttctctccggacttcaccttcggttttgaagaatccctggaagtagacacgaacccactgctgggcgctggtaaattcgcaactgatcctgcggttaccctggctcacgaactgattcatgcaggccaccgcctgtacggtatcgccatcaatccgaaccgtgtcttcaaagttaacaccaacgcgtattacgagatgtccggtctggaagttagcttcgaagaactgcgtacttttggcggtcacgacgctaaattcatcgactctctgcaagaaaacgagttccgtctgtactactataacaagttcaaagatatcgcatccaccctgaacaaagcgaaatccatcgtgggtaccactgcttctctccagtacatgaagaacgtttttaaagaaaaatacctgctcagcgaagacacctccggcaaattctctgtagacaagttgaaattcgataaactttacaaaatgctgactgaaatttacaccgaagacaacttcgttaagttctttaaagttctgaaccgcaaaacctatctgaacttcgacaaggcagtattcaaaatcaacatcgtgccgaaagttaactacactatctacgatggtttcaacctgcgtaacaccaacctggctgctaattttaacggccagaacacggaaatcaacaacatgaacttcacaaaactgaaaaacttcactggtctgttcgagttttacaagctgctgtgcGTCGACGGCATCATTACCTCCAAAACTAAATCTGACGATGACGATAAAAACAAAGCGCTGAACCTGCAGtgtatcaaggttaacaactgggatttattcttcagcccgagtgaagacaacttcaccaacgacctgaacaaaggtgaagaaatcacctcagatactaacatcgaagcagccgaagaaaacatctcgctggacctgatccagcagtactacctgacctttaatttcgacaacgagccggaaaacatttctatcgaaaacctgagctctgatatcatcggccagctggaactgatgccgaacatcgaacgtttcccaaacggtaaaaagtacgagctggacaaatataccatgttccactacctgcgcgcgcaggaatttgaacacggcaaatcccgtatcgcactgactaactccgttaacgaagctctgctcaacccgtcccgtgtatacaccttcttctctagcgactacgtgaaaaaggtcaacaaagcgactgaagctgcaatgttcttgggttgggttgaacagcttgtttatgattttaccgacgagacgtccgaagtatctactaccgacaaaattgcggatatcactatcatcatcccgtacatcggtccggctctgaacattggcaacatgctgtacaaagacgacttcgttggcgcactgatcttctccggtgcggtgatcctgctggagttcatcccggaaatcgccatcccggtactgggcacctttgctctggtttcttacattgcaaacaaggttctgactgtacaaaccatcgacaacgcgctgagcaaacgtaacgaaaaatgggatgaagtttacaaatatatcgtgaccaactggctggctaaggttaatactcagatcgacctcatccgcaaaaaaatgaaagaagcactggaaaaccaggcggaagctaccaaggcaatcattaactaccagtacaaccagtacaccgaggaagaaaaaaacaacatcaacttcaacatcgacgatctgtcctctaaactgaacgaatccatcaacaaagctatgatcaacatcaacaagttcctgaaccagtgctctgtaagctatctgatgaactccatgatcccgtacggtgttaaacgtctggaggacttcgatgcgtctctgaaagacgccctgctgaaatacatttacgacaaccgtggcactctgatcggtcaggttgatcgtctgaaggacaaagtgaacaataccttatcgaccgacatcccttttcagctcagtaaatatgtcgataaccaacgccttttgtccactctagaagcaCACCATCATCACcaccatcaccatcaccat
SEQ ID NO:4-rLHN/A的多肽序列
MEFVNKQFNYKDPVNGVDIAYIKIPNAGQMQPVKAFKIHNKIWVIPERDTFTNPEEGDLNPPPEAKQVPVSYYDSTYLSTDNEKDNYLKGVTKLFERIYSTDLGRMLLTSIVRGIPFWGGSTIDTELKVIDTNCINVIQPDGSYRSEELNLVIIGPSADIIQFECKSFGHEVLNLTRNGYGSTQYIRFSPDFTFGFEESLEVDTNPLLGAGKFATDPAVTLAHELIHAGHRLYGIAINPNRVFKVNTNAYYEMSGLEVSFEELRTFGGHDAKFIDSLQENEFRLYYYNKFKDIASTLNKAKSIVGTTASLQYMKNVFKEKYLLSEDTSGKFSVDKLKFDKLYKMLTEIYTEDNFVKFFKVLNRKTYLNFDKAVFKINIVPKVNYTIYDGFNLRNTNLAANFNGQNTEINNMNFTKLKNFTGLFEFYKLLCVDGIITSKTKSDDDDKNKALNLQCIKVNNWDLFFSPSEDNFTNDLNKGEEITSDTNIEAAEENISLDLIQQYYLTFNFDNEPENISIENLSSDIIGQLELMPNIERFPNGKKYELDKYTMFHYLRAQEFEHGKSRIALTNSVNEALLNPSRVYTFFSSDYVKKVNKATEAAMFLGWVEQLVYDFTDETSEVSTTDKIADITIIIPYIGPALNIGNMLYKDDFVGALIFSGAVILLEFIPEIAIPVLGTFALVSYIANKVLTVQTIDNALSKRNEKWDEVYKYIVTNWLAKVNTQIDLIRKKMKEALENQAEATKAIINYQYNQYTEEEKNNINFNIDDLSSKLNESINKAMININKFLNQCSVSYLMNSMIPYGVKRLEDFDASLKDALLKYIYDNRGTLIGQVDRLKDKVNNTLSTDIPFQLSKYVDNQRLLSTLEAHHHHHHHHHH
SEQ ID NO:5-rL/A的核苷酸序列
ATGCCATTCGTCAACAAGCAATTCAACTACAAAGACCCAGTCAACGGCGTCGACATCGCATACATCAAGATTCCGAACGCCGGTCAAATGCAGCCGGTTAAGGCTTTTAAGATCCACAACAAGATTTGGGTTATCCCGGAGCGTGACACCTTCACGAACCCGGAAGAAGGCGATCTGAACCCGCCACCGGAAGCGAAGCAAGTCCCTGTCAGCTACTACGATTCGACGTACCTGAGCACGGATAACGAAAAAGATAACTACCTGAAAGGTGTGACCAAGCTGTTCGAACGTATCTACAGCACGGATCTGGGTCGCATGCTGCTGACTAGCATTGTTCGCGGTATCCCGTTCTGGGGTGGTAGCACGATTGACACCGAACTGAAGGTTATCGACACTAACTGCATTAACGTTATTCAACCGGATGGTAGCTATCGTAGCGAAGAGCTGAATCTGGTCATCATTGGCCCGAGCGCAGACATTATCCAATTCGAGTGCAAGAGCTTTGGTCACGAGGTTCTGAATCTGACCCGCAATGGCTATGGTAGCACCCAGTACATTCGTTTTTCGCCGGATTTTACCTTCGGCTTTGAAGAGAGCCTGGAGGTTGATACCAATCCGTTGCTGGGTGCGGGCAAATTCGCTACCGATCCGGCTGTCACGCTGGCCCATGAACTGATCCACGCAGGCCACCGCCTGTACGGCATTGCCATCAACCCAAACCGTGTGTTCAAGGTTAATACGAATGCATACTACGAGATGAGCGGCCTGGAAGTCAGCTTCGAAGAACTGCGCACCTTCGGTGGCCATGACGCTAAATTCATTGACAGCTTGCAAGAGAATGAGTTCCGTCTGTACTACTATAACAAATTCAAAGACATTGCAAGCACGTTGAACAAGGCCAAAAGCATCGTTGGTACTACCGCGTCGTTGCAGTATATGAAGAATGTGTTTAAAGAGAAGTACCTGCTGTCCGAGGATACCTCCGGCAAGTTTAGCGTTGATAAGCTGAAGTTTGACAAACTGTACAAGATGCTGACCGAGATTTACACCGAGGACAACTTTGTGAAATTCTTCAAAGTGTTGAATCGTAAAACCTATCTGAATTTTGACAAAGCGGTTTTCAAGATTAACATCGTGCCGAAGGTGAACTACACCATCTATGACGGTTTTAACCTGCGTAACACCAACCTGGCGGCGAACTTTAACGGTCAGAATACGGAAATCAACAACATGAATTTCACGAAGTTGAAGAACTTCACGGGTCTGTTCGAGTTCTATAAGCTGCTGggtctagaagcaCACCATCATCACcaccatcaccatcaccat
SEQ ID NO:6-rL/A的多肽序列
MPFVNKQFNYKDPVNGVDIAYIKIPNAGQMQPVKAFKIHNKIWVIPERDTFTNPEEGDLNPPPEAKQVPVSYYDSTYLSTDNEKDNYLKGVTKLFERIYSTDLGRMLLTSIVRGIPFWGGSTIDTELKVIDTNCINVIQPDGSYRSEELNLVIIGPSADIIQFECKSFGHEVLNLTRNGYGSTQYIRFSPDFTFGFEESLEVDTNPLLGAGKFATDPAVTLAHELIHAGHRLYGIAINPNRVFKVNTNAYYEMSGLEVSFEELRTFGGHDAKFIDSLQENEFRLYYYNKFKDIASTLNKAKSIVGTTASLQYMKNVFKEKYLLSEDTSGKFSVDKLKFDKLYKMLTEIYTEDNFVKFFKVLNRKTYLNFDKAVFKINIVPKVNYTIYDGFNLRNTNLAANFNGQNTEINNMNFTKLKNFTGLFEFYKLLGLEAHHHHHHHHHH
SEQ ID NO:7-rHC/A的核苷酸序列
ATGCATCATCACCATCACCACAAAAACATCATCAATACTAGCATTCTGAACCTGCGTTACGAGAGCAATCATCTGATTGATCTGAGCCGTTATGCAAGCAAGATCAACATCGGTAGCAAGGTCAATTTTGACCCGATCGATAAGAACCAGATCCAGCTGTTTAATCTGGAATCGAGCAAAATTGAGGTTATCCTGAAAAACGCCATTGTCTACAACTCCATGTACGAGAATTTCTCCACCAGCTTCTGGATTCGCATCCCGAAATACTTCAACAGCATTAGCCTGAACAACGAGTATACTATCATCAACTGTATGGAGAACAACAGCGGTTGGAAGGTGTCTCTGAACTATGGTGAGATCATTTGGACCTTGCAGGACACCCAAGAGATCAAGCAGCGCGTCGTGTTCAAGTACTCTCAAATGATCAACATTTCCGATTACATTAATCGTTGGATCTTCGTGACCATTACGAATAACCGTCTGAATAACAGCAAGATTTACATCAATGGTCGCTTGATCGATCAGAAACCGATTAGCAACCTGGGTAATATCCACGCAAGCAACAACATTATGTTCAAATTGGACGGTTGCCGCGATACCCATCGTTATATCTGGATCAAGTATTTCAACCTGTTTGATAAAGAACTGAATGAGAAGGAGATCAAAGATTTGTATGACAACCAATCTAACAGCGGCATTTTGAAGGACTTCTGGGGCGATTATCTGCAATACGATAAGCCGTACTATATGCTGAACCTGTATGATCCGAACAAATATGTGGATGTCAATAATGTGGGTATTCGTGGTTACATGTATTTGAAGGGTCCGCGTGGCAGCGTTATGACGACCAACATTTACCTGAACTCTAGCCTGTACCGTGGTACGAAATTCATCATTAAGAAATATGCCAGCGGCAACAAAGATAACATTGTGCGTAATAACGATCGTGTCTACATCAACGTGGTCGTGAAGAATAAAGAGTACCGTCTGGCGACCAACGCTTCGCAGGCGGGTGTTGAGAAAATTCTGAGCGCGTTGGAGATCCCTGATGTCGGTAATCTGAGCCAAGTCGTGGTTATGAAGAGCAAGAACGACCAGGGTATCACTAACAAGTGCAAGATGAACCTGCAAGACAACAATGGTAACGACATCGGCTTTATTGGTTTCCACCAGTTCAACAATATTGCTAAACTGGTAGCGAGCAATTGGTACAATCGTCAGATTGAGCGCAGCAGCCGTACTTTGGGCTGTAGCTGGGAGTTTATCCCGGTCGATGATGGTTGGGGCGAACGTCCGCTG
SEQ ID NO:8-rHC/A的多肽序列
MHHHHHHKNIINTSILNLRYESNHLIDLSRYASKINIGSKVNFDPIDKNQIQLFNLESSKIEVILKNAIVYNSMYENFSTSFWIRIPKYFNSISLNNEYTIINCMENNSGWKVSLNYGEIIWTLQDTQEIKQRVVFKYSQMINISDYINRWIFVTITNNRLNNSKIYINGRLIDQKPISNLGNIHASNNIMFKLDGCRDTHRYIWIKYFNLFDKELNEKEIKDLYDNQSNSGILKDFWGDYLQYDKPYYMLNLYDPNKYVDVNNVGIRGYMYLKGPRGSVMTTNIYLNSSLYRGTKFIIKKYASGNKDNIVRNNDRVYINVVVKNKEYRLATNASQAGVEKILSALEIPDVGNLSQVVVMKSKNDQGITNKCKMNLQDNNGNDIGFIGFHQFNNIAKLVASNWYNRQIERSSRTLGCSWEFIPVDDGWGERPL
SEQ ID NO:9-rBoNT/B(0)的核苷酸序列
ATGCCGGTGACGATTAACAACTTCAACTACAACGACCCGATTGACAACAACAACATTATCATGATGGAACCGCCGTTTGCACGCGGCACGGGCCGTTATTACAAAGCGTTTAAAATCACCGATCGTATTTGGATTATCCCGGAACGCTACACGTTTGGTTATAAACCGGAAGACTTCAACAAAAGCTCTGGCATCTTCAACCGTGATGTTTGCGAATACTACGATCCGGACTACCTGAACACCAACGATAAGAAAAACATTTTTCTGCAAACGATGATCAAACTGTTCAATCGCATTAAAAGCAAACCGCTGGGTGAAAAACTGCTGGAAATGATTATCAATGGCATTCCGTATCTGGGTGATCGTCGCGTGCCGCTGGAAGAATTTAACACCAATATCGCGAGTGTTACGGTCAACAAACTGATTTCCAATCCGGGTGAAGTCGAACGTAAAAAAGGCATCTTCGCCAACCTGATCATCTTCGGCCCGGGTCCGGTGCTGAACGAAAATGAAACCATTGATATCGGTATTCAGAACCATTTTGCCTCACGCGAAGGCTTCGGCGGTATTATGCAAATGAAATTTTGCCCGGAATATGTGTCGGTTTTCAACAATGTTCAGGAAAACAAAGGTGCAAGCATCTTTAATCGTCGCGGCTATTTCTCTGATCCGGCTCTGATCCTGATGCACcAACTGATTtATGTGCTGCACGGCCTGTATGGTATCAAAGTGGATGACCTGCCGATCGTTCCGAACGAGAAAAAATTTTTCATGCAGAGCACCGACGCAATTCAAGCTGAAGAACTGTATACGTTTGGCGGTCAGGACCCGTCTATTATCACCCCGAGCACCGACAAAAGCATCTACGATAAAGTGCTGCAAAACTTTCGTGGCATTGTTGACCGCCTGAATAAAGTCCTGGTGTGTATCTCTGATCCGAACATCAACATCAACATCTACAAAAACAAATTCAAAGACAAATACAAATTCGTTGAAGATTCTGAAGGCAAATATAGTATTGACGTCGAATCCTTTGATAAACTGTACAAAAGTCTGATGTTCGGTTTCACCGAAACGAACATCGCGGAAAACTACAAAATCAAAACCCGCGCCTCCTATTTCAGCGACTCTCTGCCGCCGGTTAAAATCAAAAATCTGCTGGATAACGAAATTTATACGATCGAAGAAGG
TTTCAACATCAGCGATAAAGACATGGAAAAAGAATACCGTGGCCAGAATAAAGCAATCA
ACAAACAGGCGTATGAAGAAATTAGTAAAGAACATCTGGCGGTCTACAAAATTCAGATG
TGCAAATCCGTGAAAGCCCCGGGTATTTGTATCGATGTTGACAATGAAGACCTGTTTTTC
ATCGCCGATAAAAACAGTTTTTCCGATGACCTGTCAAAAAATGAACGCATCGAATACAA
CACCCAATCGAACTACATCGAAAACGATTTCCCGATCAACGAACTGATTCTGGATACGG
ACCTGATTAGTAAAATCGAACTGCCGTCAGAAAACACCGAATCGCTGACGGACTTTAAT
GTTGATGTCCCGGTGTATGAAAAACAGCCGGCAATTAAGAAAATTTTTACCGATGAAAA
CACGATCTTCCAGTACCTGTACAGCCAAACCTTTCCGCTGGACATTCGCGATATCTCTCT
GACGAGTTCCTTTGATGACGCACTGCTGTTCAGCAACAAAGTGTACTCCTTTTTCTCAAT
GGATTACATCAAAACCGCTAACAAAGTGGTTGAAGCGGGCCTGTTTGCCGGTTGGGTGA
AACAGATCGTTAACGATTTCGTCATCGAAGCCAACAAAAGTAACACGATGGATAAAATT
GCTGATATCTCCCTGATTGTCCCGTATATTGGCCTGGCACTGAATGTGGGTAACGAAACG
GCGAAAGGCAATTTTGAAAACGCCTTCGAAATTGCAGGCGCTTCAATCCTGCTGGAATT
TATTCCGGAACTGCTGATCCCGGTCGTGGGTGCGTTCCTGCTGGAATCTTACATCGACAA
CAAAAACAAAATCATCAAAACCATTGATAACGCGCTGACGAAACGTAACGAAAAATGG
TCAGATATGTACGGCCTGATTGTTGCCCAGTGGCTGAGCACCGTCAACACGCAATTTTAC
ACCATCAAAGAAGGTATGTACAAAGCGCTGAATTATCAGGCGCAAGCCCTGGAAGAAAT
CATCAAATACCGCTACAACATCTACAGCGAAAAAGAAAAATCTAACATCAACATCGACT
TTAATGATATCAACAGCAAACTGAACGAAGGTATCAACCAGGCAATCGATAACATCAAC
AACTTCATCAACGGCTGCTCAGTGTCGTATCTGATGAAGAAAATGATCCCGCTGGCTGTT
GAAAAACTGCTGGATTTTGACAACACCCTGAAGAAAAACCTGCTGAACTACATCGATGA
AAACAAACTGTACCTGATCGGCTCAGCCGAATACGAAAAATCGAAAGTGAACAAATACC
TGAAAACCATCATGCCGTTTGACCTGAGTATTTACACCAACGATACGATCCTGATCGAAA
TGTTCAACAAATACAACTCCGAAATTCTGAACAATATTATCCTGAACCTGCGTTACAAAG
ACAACAATCTGATCGATCTGAGCGGCTATGGTGCAAAAGTTGAAGTCTACGACGGTGTC
GAACTGAACGATAAAAACCAGTTCAAACTGACCTCATCGGCTAACTCAAAAATTCGTGT
GACGCAGAACCAAAACATCATCTTCAACTCGGTCTTTCTGGACTTCAGCGTGTCTTTCT
GGATTCGCATCCCGAAATATAAAAATGATGGCATCCAGAACTACATCCATAACGAATACA
CCATCATCAACTGTATGAAAAACAACAGTGGTTGGAAAATTTCCATCCGTGGCAACCGC
ATTATCTGGACCCTGATTGATATCAATGGTAAAACGAAAAGCGTGTTTTTCGAATACAAC
ATCCGTGAAGATATCTCTGAATACATCAATCGCTGGTTTTTCGTGACCATTACGAACAATC
TGAACAATGCGAAAATCTATATCAACGGCAAACTGGAAAGTAATACCGACATCAAAGAT
ATTCGTGAAGTTATCGCCAACGGTGAAATCATCTTCAAACTGGATGGCGACATCGATCGC
ACCCAGTTCATTTGGATGAAATACTTCTCCATCTTCAACACGGAACTGAGTCAGTCCAAT
ATCGAAGAACGCTACAAAATCCAATCATACTCGGAATACCTGAAAGATTTCTGGGGTAA
CCCGCTGATGTACAACAAAGAATACTACATGTTCAACGCGGGCAACAAAAACTCATACA
TCAAACTGAAAAAAGATTCGCCGGTGGGTGAAATCCTGACCCGTAGCAAATACAACCA
GAACTCTAAATACATCAACTATCGCGATCTGTACATTGGCGAAAAATTTATTATCCGTCGC
AAAAGCAACTCTCAGAGTATTAATGATGACATCGTGCGTAAAGAAGACTACATCTATCTG
GATTTCTTTAATCTGAACCAAGAATGGCGCGTTTATACCTACAAATACTTCAAAAAAGAA
GAAGAGAAACTGTTCCTGGCCCCGATTAGCGACAGCGATGAATTTTACAACACCATCCA
GATCAAAGAATACGATGAACAGCCGACGTATAGTTGCCAACTGCTGTTCAAAAAAGACG
AAGAATCCACCGATGAAATTGGCCTGATTGGTATCCACCGTTTCTATGAAAGCGGTATCG
TTTTCGAAGAATACAAAGATTACTTCTGTATCTCTAAATGGTATCTGAAAGAAGTCAAAC
GCAAACCGTACAACCTGAAACTGGGCTGCAACTGGCAATTTATCCCGAAAGACGAAGG
CTGGACCGAA
SEQ ID NO:10-rBoNT/B(0)的多肽序列
MPVTINNFNYNDPIDNNNIIMMEPPFARGTGRYYKAFKITDRIWIIPERYTFGYKPEDFNKSSGIFNRDVCEYYDPDYLNTNDKKNIFLQTMIKLFNRIKSKPLGEKLLEMIINGIPYLGDRRVPLEEFNTNIASVTVNKLISNPGEVERKKGIFANLIIFGPGPVLNENETIDIGIQNHFASREGFGGIMQMKFCPEYVSVFNNVQENKGASIFNRRGYFSDPALILMHQLIYVLHGLYGIKVDDLPIVPNEKKFFMQSTDAIQAEELYTFGGQDPSIITPSTDKSIYDKVLQNFRGIVDRLNKVLVCISDPNININIYKNKFKDKYKFVEDSEGKYSIDVESFDKLYKSLMFGFTETNIAENYKIKTRASYFSDSLPPVKIKNLLDNEIYTIEEGFNISDKDMEKEYRGQNKAINKQAYEEISKEHLAVYKIQMCKSVKAPGICIDVDNEDLFFIADKNSFSDDLSKNERIEYNTQSNYIENDFPINELILDTDLISKIELPSENTESLTDFNVDVPVYEKQPAIKKIFTDENTIFQYLYSQTFPLDIRDISLTSSFDDALLFSNKVYSFFSMDYIKTANKVVEAGLFAGWVKQIVNDFVIEANKSNTMDKIADISLIVPYIGLALNVGNETAKGNFENAFEIAGASILLEFIPELLIPVVGAFLLESYIDNKNKIIKTIDNALTKRNEKWSDMYGLIVAQWLSTVNTQFYTIKEGMYKALNYQAQALEEIIKYRYNIYSEKEKSNINIDFNDINSKLNEGINQAIDNINNFINGCSVSYLMKKMIPLAVEKLLDFDNTLKKNLLNYIDENKLYLIGSAEYEKSKVNKYLKTIMPFDLSIYTNDTILIEMFNKYNSEILNNIILNLRYKDNNLIDLSGYGAKVEVYDGVELNDKNQFKLTSSANSKIRVTQNQNIIFNSVFLDFSVSFWIRIPKYKNDGIQNYIHNEYTIINCMKNNSGWKISIRGNRIIWTLIDINGKTKSVFFEYNIREDISEYINRWFFVTITNNLNNAKIYINGKLESNTDIKDIREVIANGEIIFKLDGDIDRTQFIWMKYFSIFNTELSQSNIEERYKIQSYSEYLKDFWGNPLMYNKEYYMFNAGNKNSYIKLKKDSPVGEILTRSKYNQNSKYINYRDLYIGEKFIIRRKSNSQSINDDIVRKEDYIYLDFFNLNQEWRVYTYKYFKKEEEKLFLAPISDSDEFYNTIQIKEYDEQPTYSCQLLFKKDEESTDEIGLIGIHRFYESGIVFEEYKDYFCISKWYLKEVKRKPYNLKLGCNWQFIPKDEGWTE
SEQ ID NO:11-rBoNT/C(0)的核苷酸序列
ATGCCGATCACGATTAATAATTTCAACTATAGCGATCCGGTGGACAATAAGAATATTCTGTATCTGGATACTCATCTGAATACGCTGGCTAACGAACCGGAGAAAGCGTTCCGCATCACAGGCAACATCTGGGTTATTCCCGATCGCTTTTCACGCAACAGCAACCCTAATCTGAACAAACCTCCTCGTGTCACCAGTCCTAAATCCGGTTATTACGACCCAAACTATCTGAGTACGGATAGCGATAAAGATCCCTTTCTGAAAGAGATCATTAAGCTGTTCAAACGCATTAACTCTCGCGAAATTGGGGAAGAGCTGATCTATCGGCTTTCGACAGATATCCCGTTCCCAGGTAACAATAATACCCCGATTAATACTTTCGACTTTGATGTTGATTTCAATTCTGTGGATGTGAAAACGCGTCAAGGCAATAATTGGGTGAAAACTGGTAGCATTAACCCGAGTGTAATTATCACAGGTCCCCGTGAGAACATCATCGACCCGGAAACCTCTACCTTCAAGCTGACGAACAACACGTTTGCTGCACAGGAAGGGTTTGGTGCCCTGTCAATCATTTCCATCTCACCGCGTTTCATGTTAACCTACTCCAATGCCACAAATGATGTTGGCGAAGGACGTTTTAGCAAATCAGAATTTTGCATGGACCCAATTCTCATTCTGATGggCacGCTGAACaATGCGATGCACAACTTGTATGGCATTGCTATTCCAAACGATCAAACCATTAGCTCCGTTACCAGTAATATCTTCTATAGCCAGTATAATGTCAAATTGGAGTATGCCGAAATTTACGCCTTTGGAGGCCCGACCATTGACCTGATTCCGAAATCTGCACGCAAATACTTCGAAGAAAAGGCGTTAGATTACTATCGCAGCATCGCGAAACGCCTGAACTCGATTACCACGGCCAATCCGTCGTCGTTCAACAAATACATTGGTGAATATAAACAGAAACTGATTCGCAAATATCGGTTTGTCGTAGAAAGCTCTGGTGAAGTGACTGTAAACCGCAACAAATTTGTCGAACTCTACAACGAGTTGACCCAAATCTTTACCGAGTTTAACTACGCAAAGATCTATAACGTACAGAACCGCAAGATTTATCTTAGCAATGTATACACACCGGTTACTGCGAACATCTTAGACGACAATGTGTATGATATTCAGAATGGCTTTAACATCCCGAAATCAAATCTGAACGTTCTGTTTATGGGCCAGAACCTGAGTCGTAATCCAGCACTGCGTAAAGTGAACCCGGAAAATATGCTCTACTTGTTTACCAAATTTTGCCACAAAGCGATTGATGGCCGCTCTCTCTATAACAAAACGCTGGATTGTCGTGAGTTACTTGTGAAGAACACTGATTTACCGTTCATTGGGGATATCTCCGACGTGAAAACCGATATCTTCCTGCGCAAAGACATTAATGAAGAAACGGAAGTCATCTATTACCCCGACAATGTGAGCGTTGATCAGGTCATTTTATCGAAGAACACCTCCGAACATGGTCAGTTGGATTTGCTGTACCCTAGCATTGACTCGGAGAGTGAAATCCTTCCGGGCGAAAATCAAGTGTTTTACGACAACCGTACCCAAAATGTTGATTATTTGAATTCTTATTACTACCTGGAATCTCAGAAATTGAGCGACAATGTGGAAGATTTCACGTTCACACGCTCCATTGAGGAAGCGCTGGATAATAGCGCGAAAGTGTATACGTATTTCCCTACCTTGGCGAATAAAGTAAATGCTGGTGTCCAGGGAGGCTTATTTCTGATGTGGGCGAATGATGTGGTAGAAGATTTTACGACCAATATTTTGCGTAAGGACACCTTAGATAAAATTAGCGATGTTAGCGCCATCATCCCCTATATTGGCCCAGCACTGAATATCTCGAACTCTGTGCGTCGCGGAAACTTCACCGAAGCATTTGCGGTGACCGGGGTTACTATTCTGTTGGAAGCCTTTCCGGAGTTTACTATTCCGGCGCTGGGTGCGTTTGTGATTTATTCGAAAGTACAAGAACGCAATGAAATTATCAAAACCATCGATAATTGCCTGGAACAACGCATTAAACGCTGGAAGGATTCTTATGAATGGATGATGGGCACCTGGTTATCCCGTATTATCACACAGTTTAACAACATCTCGTATCAGATGTACGATTCACTGAACTACCAAGCAGGGGCGATCAAAGCCAAGATCGACTTAGAATACAAGAAATATTCAGGTAGCGATAAAGAGAATATTAAAAGCCAGGTTGAAAACCTGAAGAACTCTCTGGATGTCAAAATTTCAGAGGCTATGAACAACATTAACAAATTTATCCGCGAATGTAGCGTCACGTATCTGTTTAAAAACATGCTCCCGAAAGTGATTGATGAGCTCAACGAGTTTGATCGCAACACAAAGGCCAAACTGATTAACCTGATTGATAGTCACAATATTATTTTAGTCGGTGAAGTTGACAAGCTGAAGGCTAAGGTCAATAACAGCTTTCAGAACACTATTCCGTTTAATATTTTCTCCTATACGAACAATAGTCTGCTGAAAGACATTATCAACGAATACTTCAACAATATTAATGACAGCAAAATTCTGAGCCTGCAGAATCGTAAGAATACGCTGGTAGATACCAGTGGATATAATGCGGAAGTCTCAGAAGAGGGTGATGTACAGCTGAACCCGATCTTTCCGTTCGACTTTAAACTGGGGTCTAGTGGTGAAGATCGCGGTAAAGTGATCGTTACCCAAAACGAGAACATTGTGTATAACAGCATGTACGAGAGTTTCTCAATTTCTTTCTGGATTCGCATCAATAAATGGGTTTCTAATTTGCCTGGCTATACCATCATTGATAGCGTCAAAAACAACTCGGGCTGGTCGATTGGCATTATTAGCAACTTTCTGGTGTTTACCCTGAAACAGAATGAGGATTCGGAACAGAGCATTAACTTCTCCTACGACATCAGCAACAATGCACCAGGGTATAACAAATGGTTCTTCGTAACGGTGACGAACAATATGATGGGCAATATGAAAATCTACATTAACGGGAAACTTATCGACACCATTAAAGTGAAAGAGCTTACTGGGATCAATTTTAGTAAAACCATTACCTTTGAGATCAACAAAATTCCGGACACGGGTCTGATTACCTCCGATTCGGATAATATCAATATGTGGATTCGCGACTTTTATATCTTCGCCAAAGAACTTGATGGCAAAGATATCAACATTTTGTTTAATTCCCTGCAGTATACCAATGTCGTTAAGGACTATTGGGGCAATGATCTCCGCTACAATAAAGAATACTACATGGTTAACATCGACTATCTCAATCGCTACATGTATGCTAACTCGCGTCAAATTGTGTTTAACACACGTCGTAACAACAACGATTTTAACGAAGGTTATAAAATCATTATCAAACGGATCCGCGGCAATACGAACGATACTCGTGTTCGTGGCGGTGACATTCTGTATTTCGACATGACGATTAATAATAAAGCGTACAATCTGTTCATGAAGAACGAAACCATGTACGCCGATAACCATTCCACTGAAGATATCTACGCAATCGGACTTCGCGAACAGACCAAAGACATTAACGACAACATCATCTTTCAGATTCAACCGATGAATAATACCTACTACTATGCCTCCCAGATCTTCAAAAGTAATTTCAACGGCGAAAACATTTCAGGCATTTGCTCAATCGGCACTTATCGGTTCCGGTTAGGTGGTGATTGGTATCGTCACAACTACCTTGTTCCCACAGTGAAACAAGGCAACTATGCATCGCTCTTAGAAAGCACATCTACGCATTGGGGTTTTGTGCCAGTCAGTGAA
SEQ ID NO:12-rBoNT/C(0)的多肽序列
MPITINNFNYSDPVDNKNILYLDTHLNTLANEPEKAFRITGNIWVIPDRFSRNSNPNLNKPPRVTSPKSGYYDPNYLSTDSDKDPFLKEIIKLFKRINSREIGEELIYRLSTDIPFPGNNNTPINTFDFDVDFNSVDVKTRQGNNWVKTGSINPSVIITGPRENIIDPETSTFKLTNNTFAAQEGFGALSIISISPRFMLTYSNATNDVGEGRFSKSEFCMDPILILMGTLNNAMHNLYGIAIPNDQTISSVTSNIFYSQYNVKLEYAEIYAFGGPTIDLIPKSARKYFEEKALDYYRSIAKRLNSITTANPSSFNKYIGEYKQKLIRKYRFVVESSGEVTVNRNKFVELYNELTQIFTEFNYAKIYNVQNRKIYLSNVYTPVTANILDDNVYDIQNGFNIPKSNLNVLFMGQNLSRNPALRKVNPENMLYLFTKFCHKAIDGRSLYNKTLDCRELLVKNTDLPFIGDISDVKTDIFLRKDINEETEVIYYPDNVSVDQVILSKNTSEHGQLDLLYPSIDSESEILPGENQVFYDNRTQNVDYLNSYYYLESQKLSDNVEDFTFTRSIEEALDNSAKVYTYFPTLANKVNAGVQGGLFLMWANDVVEDFTTNILRKDTLDKISDVSAIIPYIGPALNISNSVRRGNFTEAFAVTGVTILLEAFPEFTIPALGAFVIYSKVQERNEIIKTIDNCLEQRIKRWKDSYEWMMGTWLSRIITQFNNISYQMYDSLNYQAGAIKAKIDLEYKKYSGSDKENIKSQVENLKNSLDVKISEAMNNINKFIRECSVTYLFKNMLPKVIDELNEFDRNTKAKLINLIDSHNIILVGEVDKLKAKVNNSFQNTIPFNIFSYTNNSLLKDIINEYFNNINDSKILSLQNRKNTLVDTSGYNAEVSEEGDVQLNPIFPFDFKLGSSGEDRGKVIVTQNENIVYNSMYESFSISFWIRINKWVSNLPGYTIIDSVKNNSGWSIGIISNFLVFTLKQNEDSEQSINFSYDISNNAPGYNKWFFVTVTNNMMGNMKIYINGKLIDTIKVKELTGINFSKTITFEINKIPDTGLITSDSDNINMWIRDFYIFAKELDGKDINILFNSLQYTNVVKDYWGNDLRYNKEYYMVNIDYLNRYMYANSRQIVFNTRRNNNDFNEGYKIIIKRIRGNTNDTRVRGGDILYFDMTINNKAYNLFMKNETMYADNHSTEDIYAIGLREQTKDINDNIIFQIQPMNNTYYYASQIFKSNFNGENISGICSIGTYRFRLGGDWYRHNYLVPTVKQGNYASLLESTSTHWGFVPVSE
SEQ ID NO:13-rBoNT/E(0)的核苷酸序列
atgccgaaaatcaactctttcaactacaacgacccggttaacgaccgtaccatcctgtatatcaaaccgggtggttgccaggagttctacaaatctttcaacatcatgaaaaacatctggatcatcccggaacgtaacgttatcggtaccaccccgcaggacttccacccgccgacctctctgaaaaacggtgactcttcttactacgacccgaactacctccagtctgacgaagaaaaagaccgtttcctgaaaatcgttaccaaaatcttcaaccgtatcaacaacaacctgtctggtggtatcctgctggaagaactgtctaaagctaacccgtacctgggtaacgacaacaccccggacaaccagttccacatcggtgacgcttctgctgttgaaatcaaattctctaacggttctcaggacatcctgctgccgaacgttatcatcatgggtgctgaaccggacctgttcgaaaccaactcttctaacatctctctgcgtaacaactacatgccgtctaaccacggtttcggttctatcgctatcgttaccttctctccggaatactctttccgtttcaacgacaacagcatgaacgagttcatccaggacccggctctgaccctgatgcaccaactgatctactctctgcacggtctgtacggtgctaaaggtatcaccaccaaatacaccatcacccagaaacagaacccgctgatcaccaacatccgtggtaccaacatcgaagagttcctgaccttcggtggtaccgacctgaacatcatcacctctgctcagtctaacgacatctacaccaacctgctggctgactacaaaaaaatcgcttctaaactgtctaaagttcaggtttctaacccgctgctgaacccgtacaaagacgttttcgaagctaaatacggtctggacaaagacgcttctggtatctactctgttaacatcaacaaattcaacgacatcttcaaaaaactgtactctttcaccgagttcgacctggcgaccaaattccaggttaaatgccgtcagacctacatcggtcagtacaaatacttcaaactgtctaacctgctgaacgactctatctacaacatctctgaaggttacaacatcaacaacctgaaagttaacttccgtggtcagaacgctaacctgaacccgcgtatcatcaccccgatcaccggtcgtggtctggttaaaaaaatcatccgtttctgcAAGAATATTGTAAGCGTTAAAGGAATAAGAAAAAGTATCtgcatcgaaatcaacaacggtgaactgttcttcgttgcttctgaaaactcttacaacgacgacaacatcaacaccccgaaagaaatcgacgacaccgttacctctaacaacaactacgaaaacgacctggaccaggttatcctgaacttcaactctgaatctgctccgggtctgtctgacgaaaaactgaacctgaccatccagaacgacgcttacatcccgaaatacgactctaacggtacctctgacatcgaacagcacgacgttaacgaactgaacgttttcttctacctggacgctcagaaagttccggaaggtgaaaacaacgttaacctgacctcttctatcgacaccgctctgctggaacagccgaaaatctacaccttcttctcttctgagttcatcaacaacgttaacaaaccggttcaggctgctctgttcgtttcttggattcagcaggttctggttgacttcaccaccgaagctaaccagaaatctaccgttgacaaaatcgctgacatctctatcgttgttccgtacatcggtctggctctgaacatcggtaacgaagctcagaaaggtaacttcaaagacgctctggaactgctgggtgctggtatcctgctggagttcgaaccggaactgctgatcccgaccatcctggttttcaccatcaaatctttcctgggttcttctgacaacaaaaacaaagttatcaaagctatcaacaacgctctgaaagaacgtgacgaaaaatggaaagaagtttactctttcatcgtttctaactggatgaccaaaatcaacacccagttcaacaaacgtaaagaacagatgtaccaggctctccagaaccaggttaacgctatcaaaaccatcatcgaatctaaatacaactcttacaccctggaagaaaaaaacgaactgaccaacaaatacgacatcaaacagatcgaaaacgaactgaaccagaaagtttctatcgctatgaacaacatcgaccgtttcctgaccgaatcttctatctcttacctgatgaaactcatcaacgaagttaaaatcaacaaactgcgtgaatacgacgaaaacgttaaaacctacctgctgaactacatcatccagcacggttctatcctgggtgaatctcagcaggaactgaactctatggttaccgacaccctgaacaactctatcccgttcaaactgtcttcttacaccgacgacaaaatcctGATCTCTTACTTCAACAAATTCTTTAAAcgcATTAAGAGTTCATCGGTTctgaatATGCGGTACAAAAATGATAAAtatGTCGATACTTCTGGATATgatAGCAATATCAACATTAACGGCGACGTGTATAAATATccgACAAATAAAAACCAGTTTGGGATATATAACGACAAGctgTCGGAGGTCAATattTCTCAAAACGACtatATCattTACGATAATaaaTATAAAAACTTTAGCATTAGTtttTGGGTTcgtATACCTAATtatGACAATaaaattGTAAATGTGAATAACGAGTATACCATTATAAACTGTATGcgcGACAATAACAGTGGTTGGAAGGTATCGctgAACCATAATGAGATTATCTGGACCctgcagGATAATgcaGGTATAAACCAGAAACTGGCTTTTAACTATGGAAACGCAAATGGGATCTCAGATTACATTaataaaTGGatttttGTTaccATTACGAACGATcgcTTAGGCGACTCAAAACTTTATATTAATggcAATctgATAGATCAGAAATCAATCTTAAATTTGGGCAATATTCATGTCTCTgatAACATCTTGTTCAAGATCGTTAATTGCAGTTACACTcgtTATATTGGCATTCGTTACTTTAATATCTTCgataaaGAActgGACGAGACGGAAATCcagACTCTGTATTCAAACGAGCCCAATACTAATATATTGAAAGATTTTTGGGGTAACTATCTTTTATATGATAAAGAATACTATCTCCTGaatGTATTGAAGCCAAACAATTTCATAGATAGACGCAAGGATAGCACATTAAGTATCAACAATATCAGATCTACTATActgttaGCAAATCGCCTcTACTCCggtATTAAAGTGAAGATTcagCGGGTTAATAACTCCAGTACCAATGATAATCTGGTCCGTAAGAACGATCAGGTATACATCaatTTCGTCGCGAGCAAAACTcatCTCTTCCCGCTTTACGCCgatACAGCTACGACAAACAAGGAAAAAACCATAAAAATTTCCAGCTCCGGAAACAGATTCAATCAAGTAGTTGTAATGAACTCTGTGGGTaatAATTGTACGATGAACTTTaagAATAACAATGGGAACAATattGGACTTTTGGGCTTcAAAGCCGACACAGTGGTGGCGTCCACCTGGTATTACACGcacATGcggGACCATACGAATTCGAACGGTTGCTTCTGGAACTTTATCTCGGAAgaaCACGGGTGGCAAGAAAAASEQ ID NO:14-rBoNT/E(0)的多肽序列
MPKINSFNYNDPVNDRTILYIKPGGCQEFYKSFNIMKNIWIIPERNVIGTTPQDFHPPTSLKNGDSSYYDPNYLQSDEEKDRFLKIVTKIFNRINNNLSGGILLEELSKANPYLGNDNTPDNQFHIGDASAVEIKFSNGSQDILLPNVIIMGAEPDLFETNSSNISLRNNYMPSNHGFGSIAIVTFSPEYSFRFNDNSMNEFIQDPALTLMHQLIYSLHGLYGAKGITTKYTITQKQNPLITNIRGTNIEEFLTFGGTDLNIITSAQSNDIYTNLLADYKKIASKLSKVQVSNPLLNPYKDVFEAKYGLDKDASGIYSVNINKFNDIFKKLYSFTEFDLATKFQVKCRQTYIGQYKYFKLSNLLNDSIYNISEGYNINNLKVNFRGQNANLNPRIITPITGRGLVKKIIRFCKNIVSVKGIRKSICIEINNGELFFVASENSYNDDNINTPKEIDDTVTSNNNYENDLDQVILNFNSESAPGLSDEKLNLTIQNDAYIPKYDSNGTSDIEQHDVNELNVFFYLDAQKVPEGENNVNLTSSIDTALLEQPKIYTFFSSEFINNVNKPVQAALFVSWIQQVLVDFTTEANQKSTVDKIADISIVVPYIGLALNIGNEAQKGNFKDALELLGAGILLEFEPELLIPTILVFTIKSFLGSSDNKNKVIKAINNALKERDEKWKEVYSFIVSNWMTKINTQFNKRKEQMYQALQNQVNAIKTIIESKYNSYTLEEKNELTNKYDIKQIENELNQKVSIAMNNIDRFLTESSISYLMKLINEVKINKLREYDENVKTYLLNYIIQHGSILGESQQELNSMVTDTLNNSIPFKLSSYTDDKILISYFNKFFKRIKSSSVLNMRYKNDKYVDTSGYDSNININGDVYKYPTNKNQFGIYNDKLSEVNISQNDYIIYDNKYKNFSISFWVRIPNYDNKIVNVNNEYTIINCMRDNNSGWKVSLNHNEIIWTLQDNAGINQKLAFNYGNANGISDYINKWIFVTITNDRLGDSKLYINGNLIDQKSILNLGNIHVSDNILFKIVNCSYTRYIGIRYFNIFDKELDETEIQTLYSNEPNTNILKDFWGNYLLYDKEYYLLNVLKPNNFIDRRKDSTLSINNIRSTILLANRLYSGIKVKIQRVNNSSTNDNLVRKNDQVYINFVASKTHLFPLYADTATTNKEKTIKISSSGNRFNQVVVMNSVGNNCTMNFKNNNGNNIGLLGFKADTVVASTWYYTHMRDHTNSNGCFWNFISEEHGWQEK
SEQ ID NO:15-rBoNT/F(0)的核苷酸序列
ATGCCGGTGGTCATCAACAGCTTCAACTACAACGACCCAGTAAACGACGACACGATCCTGTATATGCAAATCCCGTATGAAGAGAAGAGCAAGAAGTACTATAAGGCCTTTGAAATCATGCGCAATGTGTGGATTATTCCGGAGCGTAATACGATTGGTACTGACCCAAGCGACTT
CGATCCACCTGCGTCTTTGGAAAACGGCTCGTCCGCATATTACGACCCGAATTACCTGAC
CACCGATGCGGAGAAAGATCGTTATTTGAAAACCACCATCAAGCTGTTCAAACGCATTA
ACAGCAATCCGGCAGGTGAGGTCCTGCTGCAAGAGATTAGCTACGCAAAGCCTTATCTG
GGTAATGAGCATACGCCTATTAACGAGTTTCACCCGGTTACCCGCACTACCAGCGTTAAC
ATCAAGTCCTCGACCAACGTGAAGTCTAGCATTATCCTGAACCTGCTGGTTCTGGGTGCC
GGTCCGGACATCTTCGAAAACTCTAGCTACCCGGTGCGTAAACTGATGGATAGCGGCGG
TGTTTATGACCCGAGCAATGACGGTTTTGGCAGCATCAATATCGTGACGTTTAGCCCGGA
GTACGAGTACACCTTCAATGATATCAGCGGTGGTTACAATTCTTCTACCGAGAGCTTCAT
CGCCGACCCGGCGATCAGCCTGGCACACCAACTGATCTATGCATTGCATGGCTTGTACG
GTGCCCGTGGTGTGACGTATAAAGAGACTATCAAGGTTAAGCAGGCACCTCTGATGATT
GCGGAAAAGCCGATTCGCCTGGAAGAGTTCCTGACCTTCGGCGGTCAAGATTTGAACAT
CATTACCTCGGCCATGAAAGAGAAAATCTATAACAATTTGCTGGCCAACTATGAAAAGAT
TGCAACGCGCTTGTCTCGTGTTAACTCCGCTCCGCCGGAATACGACATTAATGAGTACAA
AGACTACTTTCAATGGAAATATGGCCTGGACAAAAATGCGGATGGTTCTTATACCGTGAA
TGAAAACAAATTCAATGAAATCTACAAGAAACTGTACAGCTTCACCGAAATCGATCTGG
CGAACAAGTTCAAAGTCAAATGTCGTAATACCTACTTCATCAAATATGGCTTCCTGAAAG
TCCCGAACCTGCTGGACGATGACATCTATACCGTCAGCGAAGGCTTCAACATCGGCAAT
CTGGCCGTGAATAATCGTGGTCAGAACATCAAACTGAATCCGAAAATCATTGACTCCATC
CCAGACAAGGGCCTGGTTGAGAAAATCGTGAAGTTCTGCAAAAGCGTTATTCCGCGTAA
AGGTACGAAAGCACCGCCTCGCCTGTGCATTCGCGTTAACAACCGTGAGTTGTTCTTTG
TGGCATCTGAAAGCAGCTACAACGAGAACGACATCAACACCCCTAAAGAAATTGATGAT
ACCACGAACCTGAATAACAATTATCGCAACAATCTGGACGAGGTGATCCTGGATTACAAT
TCGGAAACCATTCCGCAAATTAGCAATCAGACGCTGAACACCCTGGTTCAGGACGATAG
CTACGTTCCGCGTTACGACTCCAATGGTACTAGCGAGATTGAAGAACACAACGTAGTGG
ACTTGAACGTTTTCTTTTATCTGCACGCCCAGAAGGTTCCGGAGGGCGAAACCAATATTA
GCCTGACCAGCTCGATCGACACCGCGCTGTCTGAGGAGAGCCAAGTCTACACCTTTTTC
AGCAGCGAGTTTATCAACACTATTAACAAGCCAGTTCATGCTGCATTGTTTATCTCTTGGA
TTAACCAGGTGATTCGCGACTTTACGACGGAGGCGACCCAGAAGTCTACCTTCGACAAA
ATTGCAGACATCTCCCTGGTCGTCCCATACGTCGGCCTGGCGTTGAATATTGGCAATGAA
GTTCAAAAAGAGAACTTCAAAGAAGCGTTCGAGCTGCTGGGTGCAGGCATCCTGCTGG
AGTTCGTGCCGGAACTGTTGATCCCGACCATCCTGGTGTTCACCATTAAGAGCTTCATTG
GATCCTCCGAGAATAAGAACAAGATCATCAAGGCGATCAATAACAGCCTGATGGAGCGT
GAAACGAAGTGGAAAGAAATCTATAGCTGGATTGTTAGCAATTGGCTGACTCGTATTAAC
ACGCAATTCAACAAGCGTAAAGAGCAAATGTACCAAGCCCTGCAAAACCAAGTTGACG
CCATCAAAACGGTAATTGAATACAAGTACAACAATTACACGAGCGATGAGCGCAACCGC
CTGGAAAGCGAATACAACATCAACAACATTCGCGAAGAATTGAACAAGAAAGTGAGCC
TGGCGATGGAGAACATTGAGCGTTTTATCACCGAAAGCAGCATCTTTTACCTGATGAAAT
TGATTAATGAGGCGAAAGTCTCGAAACTGCGTGAGTACGACGAAGGTGTGAAAGAGTA
TCTGCTGGATTACATTAGCGAGCACCGTAGCATCTTGGGTAACTCGGTTCAGGAGCTGA
ACGATCTGGTGACCTCTACCCTGAACAATAGCATCCCGTTCGAACTGAGCAGCTATACCA
ATGACAAGATTCTGATTCTGTATTTCAATAAACTGTATAAGAAGATCAAGGATAACAGCA
TTCTGGATATGCGTTACGAAAACAATAAGTTTATCGACATTTCTGGTTACGGCAGCAACA
TTTCCATCAATGGCGATGTCTACATCTACAGCACCAATCGCAACCAGTTCGGCATCTACT
CTAGCAAACCGAGCGAAGTTAACATCGCACAGAACAATGATATTATTTATAACGGTCGTT
ATCAAAACTTCTCTATCAGCTTTTGGGTCCGTATCCCGAAGTACTTCAATAAAGTCAATCT
GAATAATGAATACACGATCATCGACTGCATTCGCAATAACAACAGCGGTTGGAAAATCA
GCCTGAATTACAACAAAATTATTTGGACCCTGCAAGATACGGCGGGTAACAATCAGAAACTGGTGTTTAACTACACGCAAATGATCAGCATTTCTGACTATATCAACAAGTGGATCTTTGTTACCATCACCAATAATCGTCTGGGCAATAGCCGTATTTACATCAACGGTAACCTGATTGATGAGAAAAGCATCAGCAACCTGGGCGATATTCACGTCAGCGACAACATTCTGTTCAAAATTGTTGGTTGTAACGATACCCGTTACGTCGGCATCCGTTATTTCAAGGTTTTCGATACGGAGCTGGGTAAAACGGAAATCGAAACGTTGTACTCCGATGAACCAGATCCGAGCATTCTGAAGGACTTTTGGGGTAACTACTTGCTGTACAATAAACGTTACTATCTGCTGAATCTGTTGCGCACCGACAAGAGCATTACCCAAAACAGCAATTTCCTGAACATTAATCAGCAACGCGGCGTATACCAAAAACCGAACATCTTCAGCAATACGCGCCTGTATACTGGTGTTGAAGTGATCATTCGTAAGAACGGTAGCACCGACATTAGCAACACGGACAATTTCGTCCGTAAGAATGACCTGGCGTACATTAACGTCGTGGACCGTGATGTCGAGTATCGTCTGTACGCAGACATCAGCATTGCGAAACCGGAAAAGATTATCAAGCTGATCCGTACCAGCAACAGCAACAACAGCCTGGGTCAGATCATTGTGATGGACAGCATTGGTAATAACTGCACGATGAACTTCCAGAACAACAATGGTGGTAATATCGGTCTGCTGGGTTTTCACAGCAATAATCTGGTTGCTTCCAGCTGGTACTACAATAACATTCGTAAAAACACGTCTAGCAATGGTTGTTTTTGGAGCTTTATCAGCAAAGAGCACGGCTGGCAAGAAAAT
SEQ ID NO:16-rBoNT/F(0)的多肽序列
MPVVINSFNYNDPVNDDTILYMQIPYEEKSKKYYKAFEIMRNVWIIPERNTIGTDPSDFDPPASLENGSSAYYDPNYLTTDAEKDRYLKTTIKLFKRINSNPAGEVLLQEISYAKPYLGNEHTPINEFHPVTRTTSVNIKSSTNVKSSIILNLLVLGAGPDIFENSSYPVRKLMDSGGVYDPSNDGFGSINIVTFSPEYEYTFNDISGGYNSSTESFIADPAISLAHQLIYALHGLYGARGVTYKETIKVKQAPLMIAEKPIRLEEFLTFGGQDLNIITSAMKEKIYNNLLANYEKIATRLSRVNSAPPEYDINEYKDYFQWKYGLDKNADGSYTVNENKFNEIYKKLYSFTEIDLANKFKVKCRNTYFIKYGFLKVPNLLDDDIYTVSEGFNIGNLAVNNRGQNIKLNPKIIDSIPDKGLVEKIVKFCKSVIPRKGTKAPPRLCIRVNNRELFFVASESSYNENDINTPKEIDDTTNLNNNYRNNLDEVILDYNSETIPQISNQTLNTLVQDDSYVPRYDSNGTSEIEEHNVVDLNVFFYLHAQKVPEGETNISLTSSIDTALSEESQVYTFFSSEFINTINKPVHAALFISWINQVIRDFTTEATQKSTFDKIADISLVVPYVGLALNIGNEVQKENFKEAFELLGAGILLEFVPELLIPTILVFTIKSFIGSSENKNKIIKAINNSLMERETKWKEIYSWIVSNWLTRINTQFNKRKEQMYQALQNQVDAIKTVIEYKYNNYTSDERNRLESEYNINNIREELNKKVSLAMENIERFITESSIFYLMKLINEAKVSKLREYDEGVKEYLLDYISEHRSILGNSVQELNDLVTSTLNNSIPFELSSYTNDKILILYFNKLYKKIKDNSILDMRYENNKFIDISGYGSNISINGDVYIYSTNRNQFGIYSSKPSEVNIAQNNDIIYNGRYQNFSISFWVRIPKYFNKVNLNNEYTIIDCIRNNNSGWKISLNYNKIIWTLQDTAGNNQKLVFNYTQMISISDYINKWIFVTITNNRLGNSRIYINGNLIDEKSISNLGDIHVSDNILFKIVGCNDTRYVGIRYFKVFDTELGKTEIETLYSDEPDPSILKDFWGNYLLYNKRYYLLNLLRTDKSITQNSNFLNINQQRGVYQKPNIFSNTRLYTGVEVIIRKNGSTDISNTDNFVRKNDLAYINVVDRDVEYRLYADISIAKPEKIIKLIRTSNSNNSLGQIIVMDSIGNNCTMNFQNNNGGNIGLLGFHSNNLVASSWYYNNIRKNTSSNGCFWSFISKEHGWQEN
SEQ ID NO:17-rBoNT/A(0)的核苷酸序列(His标签)
ATGCCGTTTGTGAACAAGCAGTTCAACTATAAAGATCCGGTTAATGGTGTGGATATCGCCTATATCAAAATTCCGAATGCAGGTCAGATGCAGCCGGTTAAAGCCTTTAAAATCCATAACAAAATTTGGGTGATTCCGGAACGTGATACCTTTACCAATCCGGAAGAAGGTGATCTGAATCCGCCTCCGGAAGCAAAACAGGTTCCGGTTAGCTATTATGATAGCACCTATCTGAGCACCGATAACGAGAAAGATAACTATCTGAAAGGTGTGACCAAACTGTTTGAACGCATTTATAGTACCGATCTGGGTCGTATGCTGCTGACCAGCATTGTTCGTGGTATTCCGTTTTGGGGTGGTAGCACCATTGATACCGAACTGAAAGTTATTGACACCAACTGCATTAATGTGATTC
AGCCGGATGGTAGCTATCGTAGCGAAGAACTGAATCTGGTTATTATTGGTCCGAGCGCAG
ATATCATTCAGTTTGAATGTAAAAGCTTTGGCCACGAAGTTCTGAATCTGACCCGTAATG
GTTATGGTAGTACCCAGTATATTCGTTTCAGTCCGGATTTTACCTTTGGCTTTGAAGAAAG
CCTGGAAGTTGATACAAATCCGCTGTTAGGTGCAGGTAAATTTGCAACCGATCCGGCAG
TTACCCTGGCACACCAGCTGATTTATGCCGGTCATCGTCTGTATGGTATTGCCATTAATCC
GAATCGTGTGTTCAAAGTGAATACCAACGCCTATTATGAAATGAGCGGTCTGGAAGTGA
GTTTTGAAGAACTGCGTACCTTTGGTGGTCATGATGCCAAATTTATCGATAGCCTGCAAG
AAAATGAATTTCGCCTGTACTACTATAACAAATTCAAGGATATTGCGAGCACCCTGAATA
AAGCCAAAAGCATTGTTGGCACCACCGCAAGCCTGCAGTATATGAAAAATGTGTTTAAA
GAAAAATATCTGCTGAGCGAAGATACCAGCGGTAAATTTAGCGTTGACAAACTGAAATT
CGATAAACTGTACAAGATGCTGACCGAGATTTATACCGAAGATAACTTCGTGAAGTTTTT
CAAAGTGCTGAACCGCAAAACCTACCTGAACTTTGATAAAGCCGTGTTCAAAATCAACA
TCGTGCCGAAAGTGAACTATACCATCTATGATGGTTTTAACCTGCGCAATACCAATCTGG
CAGCAAACTTTAATGGTCAGAACACCGAAATCAACAACATGAACTTTACCAAACTGAAG
AACTTCACCGGTCTGTTCGAATTTTACAAACTGCTGTGTGTTCGTGGCATTATTACCAGC
AAAACCAAAAGTCTGGATAAAGGCTACAATAAAGCCCTGAATGATCTGTGCATTAAGGT
GAATAATTGGGACCTGTTTTTTAGCCCGAGCGAGGATAATTTCACCAACGATCTGAACAA
AGGCGAAGAAATTACCAGCGATACCAATATTGAAGCAGCCGAAGAAAACATTAGCCTGG
ATCTGATTCAGCAGTATTATCTGACCTTCAACTTCGATAATGAGCCGGAAAATATCAGCAT
TGAAAACCTGAGCAGCGATATTATTGGCCAGCTGGAACTGATGCCGAATATTGAACGTTT
TCCGAACGGCAAAAAATACGAGCTGGATAAATACACCATGTTCCATTATCTGCGTGCCCA
AGAATTTGAACATGGTAAAAGCCGTATTGCACTGACCAATAGCGTTAATGAAGCACTGC
TGAACCCGAGCCGTGTTTATACCTTTTTTAGCAGCGATTACGTGAAAAAGGTTAACAAA
GCAACCGAAGCAGCCATGTTTTTAGGTTGGGTTGAACAGCTGGTTTATGATTTCACCGAT
GAAACCAGCGAAGTTAGCACCACCGATAAAATTGCAGATATTACCATCATCATCCCGTAT
ATCGGTCCGGCACTGAATATTGGCAATATGCTGTATAAAGACGATTTTGTGGGTGCCCTG
ATTTTTAGCGGTGCAGTTATTCTGCTGGAATTTATTCCGGAAATTGCCATTCCGGTTCTGG
GCACCTTTGCACTGGTGAGCTATATTGCAAATAAAGTTCTGACCGTGCAGACCATCGATA
ATGCACTGAGCAAACGTAACGAAAAATGGGATGAAGTGTACAAGTATATCGTGACCAAT
TGGCTGGCAAAAGTTAACACCCAGATTGACCTGATTCGCAAGAAGATGAAAGAAGCAC
TGGAAAATCAGGCAGAAGCAACCAAAGCCATTATCAACTATCAGTATAACCAGTACACC
GAAGAAGAGAAAAATAACATCAACTTCAACATCGACGATCTGTCCAGCAAACTGAACG
AAAGCATCAACAAAGCCATGATTAACATTAACAAATTTCTGAACCAGTGCAGCGTGAGC
TATCTGATGAATAGCATGATTCCGTATGGTGTGAAACGTCTGGAAGATTTTGATGCAAGC
CTGAAAGATGCCCTGCTGAAATATATCTATGATAATCGTGGCACCCTGATTGGTCAGGTT
GATCGTCTGAAAGATAAAGTGAACAACACCCTGAGTACCGATATTCCTTTTCAGCTGAG
CAAATATGTGGATAATCAGCGTCTGCTGTCAACCTTTACCGAATACATTAAGAACATCATC
AACACCAGCATTCTGAACCTGCGTTATGAAAGCAATCATCTGATTGATCTGAGCCGTTAT
GCCAGCAAAATCAATATAGGCAGCAAGGTTAACTTCGACCCGATTGACAAAAATCAGAT
ACAGCTGTTTAATCTGGAAAGCAGCAAAATTGAGGTGATCCTGAAAAACGCCATTGTGT
ATAATAGCATGTACGAGAATTTCTCGACCAGCTTTTGGATTCGTATCCCGAAATACTTTAA
TAGCATCAGCCTGAACAACGAGTACACCATTATTAACTGCATGGAAAACAATAGCGGCT
GGAAAGTTAGCCTGAATTATGGCGAAATTATCTGGACCCTGCAGGATACCCAAGAAATC
AAACAGCGTGTGGTTTTCAAATACAGCCAGATGATTAATATCAGCGACTATATCAACCGC
TGGATTTTTGTGACCATTACCAATAATCGCCTGAATAACAGCAAGATCTATATTAACGGTC
GTCTGATTGACCAGAAACCGATTAGTAATCTGGGTAATATTCATGCGAGCAACAACATCA
TGTTTAAACTGGATGGTTGTCGTGATACCCATCGTTATATTTGGATCAAGTACTTCAACCTGTTCGATAAAGAGTTGAACGAAAAAGAAATTAAAGACCTGTATGATAACCAGAGCAACAGCGGTATTCTGAAGGATTTTTGGGGAGATTATCTGCAGTATGACAAACCGTATTATATGCTGAATCTGTACGACCCGAATAAATACGTGGATGTGAATAATGTTGGCATCCGTGGTTATATGTACCTGAAAGGTCCGCGTGGTAGCGTTATGACCACAAACATTTATCTGAATAGCAGCCTGTATCGCGGAACCAAATTCATCATTAAAAAGTATGCCAGCGGCAACAAGGATAATATTGTGCGTAATAATGATCGCGTGTACATTAACGTTGTGGTGAAGAATAAAGAATATCGCCTGGCAACCAATGCAAGCCAGGCAGGCGTTGAAAAAATTCTGAGTGCCCTGGAAATTCCGGATGTTGGTAATCTGAGCCAGGTTGTTGTGATGAAAAGCAAAAATGATCAGGGCATCACCAACAAGTGCAAAATGAATCTGCAGGACAATAACGGCAACGATATTGGTTTTATTGGCTTCCACCAGTTCAACAATATTGCGAAACTGGTTGCAAGCAATTGGTATAATCGTCAGATTGAACGTAGCAGTCGTACCCTGGGTTGTAGCTGGGAATTTATCCCTGTGGATGATGGTTGGGGTGAACGTCCGCTGGAAAACCTGTATTTTCAAGGTGCAAGTCATCATCACCATCACCACCATCATTAA
SEQ ID NO:18-rBoNT/A(0)的多肽序列(His标签)
MPFVNKQFNYKDPVNGVDIAYIKIPNAGQMQPVKAFKIHNKIWVIPERDTFTNPEEGDLNPPPEAKQVPVSYYDSTYLSTDNEKDNYLKGVTKLFERIYSTDLGRMLLTSIVRGIPFWGGSTIDTELKVIDTNCINVIQPDGSYRSEELNLVIIGPSADIIQFECKSFGHEVLNLTRNGYGSTQYIRFSPDFTFGFEESLEVDTNPLLGAGKFATDPAVTLAHQLIYAGHRLYGIAINPNRVFKVNTNAYYEMSGLEVSFEELRTFGGHDAKFIDSLQENEFRLYYYNKFKDIASTLNKAKSIVGTTASLQYMKNVFKEKYLLSEDTSGKFSVDKLKFDKLYKMLTEIYTEDNFVKFFKVLNRKTYLNFDKAVFKINIVPKVNYTIYDGFNLRNTNLAANFNGQNTEINNMNFTKLKNFTGLFEFYKLLCVRGIITSKTKSLDKGYNKALNDLCIKVNNWDLFFSPSEDNFTNDLNKGEEITSDTNIEAAEENISLDLIQQYYLTFNFDNEPENISIENLSSDIIGQLELMPNIERFPNGKKYELDKYTMFHYLRAQEFEHGKSRIALTNSVNEALLNPSRVYTFFSSDYVKKVNKATEAAMFLGWVEQLVYDFTDETSEVSTTDKIADITIIIPYIGPALNIGNMLYKDDFVGALIFSGAVILLEFIPEIAIPVLGTFALVSYIANKVLTVQTIDNALSKRNEKWDEVYKYIVTNWLAKVNTQIDLIRKKMKEALENQAEATKAIINYQYNQYTEEEKNNINFNIDDLSSKLNESINKAMININKFLNQCSVSYLMNSMIPYGVKRLEDFDASLKDALLKYIYDNRGTLIGQVDRLKDKVNNTLSTDIPFQLSKYVDNQRLLSTFTEYIKNIINTSILNLRYESNHLIDLSRYASKINIGSKVNFDPIDKNQIQLFNLESSKIEVILKNAIVYNSMYENFSTSFWIRIPKYFNSISLNNEYTIINCMENNSGWKVSLNYGEIIWTLQDTQEIKQRVVFKYSQMINISDYINRWIFVTITNNRLNNSKIYINGRLIDQKPISNLGNIHASNNIMFKLDGCRDTHRYIWIKYFNLFDKELNEKEIKDLYDNQSNSGILKDFWGDYLQYDKPYYMLNLYDPNKYVDVNNVGIRGYMYLKGPRGSVMTTNIYLNSSLYRGTKFIIKKYASGNKDNIVRNNDRVYINVVVKNKEYRLATNASQAGVEKILSALEIPDVGNLSQVVVMKSKNDQGITNKCKMNLQDNNGNDIGFIGFHQFNNIAKLVASNWYNRQIERSSRTLGCSWEFIPVDDGWGERPLENLYFQGASHHHHHHHH
SEQ ID NO:19-rLHN/A的核苷酸序列(His标签)
ATGCCGTTTGTGAACAAGCAGTTCAACTATAAAGATCCGGTTAATGGTGTGGATATCGCCTATATCAAAATTCCGAATGCAGGTCAGATGCAGCCGGTTAAAGCCTTTAAAATCCATAACAAAATTTGGGTGATTCCGGAACGTGATACCTTTACCAATCCGGAAGAAGGTGATCTGAATCCGCCTCCGGAAGCAAAACAGGTTCCGGTTAGCTATTATGATAGCACCTATCTGAGCACCGATAACGAGAAAGATAACTATCTGAAAGGTGTGACCAAACTGTTTGAACGCATTTATAGTACCGATCTGGGTCGTATGCTGCTGACCAGCATTGTTCGTGGTATTCCGTTTTGGGGTGGTAGCACCATTGATACCGAACTGAAAGTTATTGACACCAACTGCATTAATGTGATTCAGCCGGATGGTAGCTATCGTAGCGAAGAACTGAATCTGGTTATTATTGGTCCGAGCGCAGATATCATTCAGTTTGAATGTAAATCCTTTGGCCACGAAGTTCTGAATCTGACCCGTAATGGTTATGGTAGTACCCAGTATATTCGTTTCAGTCCGGATTTTACCTTTGGCTTTGAAGAAAGCCTGGAAGTTGATACAAATCCGCTGTTAGGTGCAGGTAAATTTGCAACCGATCCGGCAGTTACCCTGGCACATGAACTGATTCATGCCGGTCATCGTCTGTATGGTATTGCAATTAATCCGAACCGTGTGTTCAAAGTGAATACCAACGCATATTATGAAATGAGCGGTCTGGAAGTGTCATTTGAAGAACTGCGTACCTTTGGTGGTCATGATGCCAAATTTATCGATAGCCTGCAAGAAAATGAATTTCGCCTGTACTACTATAACAAATTCAAGGATATTGCGAGCACCCTGAATAAAGCCAAAAGCATTGTTGGCACCACCGCAAGCCTGCAGTATATGAAAAATGTGTTTAAAGAAAAATATCTGCTGAGCGAAGATACCAGCGGTAAATTTAGCGTTGACAAACTGAAATTCGATAAACTGTACAAGATGCTGACCGAGATTTATACCGAAGATAACTTCGTGAAGTTTTTCAAAGTGCTGAACCGCAAAACCTACCTGAACTTTGATAAAGCCGTGTTCAAAATCAACATCGTGCCGAAAGTGAACTATACCATCTATGATGGTTTTAACCTGCGCAATACCAATCTGGCAGCAAACTTTAATGGTCAGAACACCGAAATCAACAACATGAACTTTACCAAACTGAAGAACTTCACCGGTCTGTTCGAATTTTACAAACTGCTGTGTGTTCGTGGCATTATTACCAGCAAAACCAAAAGTCTGGATAAAGGCTACAATAAAGCCCTGAATGATCTGTGCATTAAGGTGAATAATTGGGACCTGTTTTTTAGCCCGAGCGAGGATAATTTCACCAACGATCTGAACAAAGGCGAAGAAATTACCAGCGATACCAATATTGAAGCAGCCGAAGAAAACATTAGCCTGGATCTGATTCAGCAGTATTATCTGACCTTCAACTTCGATAATGAGCCGGAAAATATCAGCATTGAAAACCTGAGCAGCGATATTATTGGCCAGCTGGAACTGATGCCGAATATTGAACGTTTTCCGAACGGCAAAAAATACGAGCTGGATAAATACACCATGTTCCATTATCTGCGTGCCCAAGAATTTGAACATGGTAAAAGCCGTATTGCACTGACCAATAGCGTTAATGAAGCACTGCTGAACCCGAGCCGTGTTTATACCTTTTTTAGCAGCGATTACGTGAAAAAGGTTAACAAAGCAACCGAAGCAGCCATGTTTTTAGGTTGGGTTGAACAGCTGGTTTATGATTTCACCGATGAAACCAGCGAAGTTAGCACCACCGATAAAATTGCAGATATTACCATCATCATCCCGTATATCGGTCCGGCACTGAATATTGGCAATATGCTGTATAAAGACGATTTTGTGGGTGCCCTGATTTTTAGCGGTGCAGTTATTCTGCTGGAATTTATTCCGGAAATTGCCATTCCGGTTCTGGGCACCTTTGCACTGGTGAGCTATATTGCAAATAAAGTTCTGACCGTGCAGACCATCGATAATGCACTGAGCAAACGTAACGAAAAATGGGATGAAGTGTACAAGTATATCGTGACCAATTGGCTGGCAAAAGTTAACACCCAGATTGACCTGATTCGCAAGAAGATGAAAGAAGCACTGGAAAATCAGGCAGAAGCAACCAAAGCCATTATCAACTATCAGTATAACCAGTACACCGAAGAAGAGAAAAATAACATCAACTTCAACATCGACGATCTGTCCAGCAAACTGAACGAAAGCATCAACAAAGCCATGATTAACATTAACAAATTTCTGAACCAGTGCAGCGTGAGCTATCTGATGAATAGCATGATTCCGTATGGTGTGAAACGTCTGGAAGATTTTGATGCAAGCCTGAAAGATGCCCTGCTGAAATATATCTATGATAATCGTGGCACCCTGATTGGTCAGGTTGATCGTCTGAAAGATAAAGTGAACAACACCCTGAGTACCGATATTCCTTTTCAGCTGAGCAAATATGTGGATAATCAGCGTCTGCTGTCAACCGAAAATCTGTATTTCCAGGGTGCAAGTCATCATCACCATCACCACCATCATTAA
SEQ ID NO:20-rLHN/A的多肽序列(His标签)
MPFVNKQFNYKDPVNGVDIAYIKIPNAGQMQPVKAFKIHNKIWVIPERDTFTNPEEGDLNPPPEAKQVPVSYYDSTYLSTDNEKDNYLKGVTKLFERIYSTDLGRMLLTSIVRGIPFWGGSTIDTELKVIDTNCINVIQPDGSYRSEELNLVIIGPSADIIQFECKSFGHEVLNLTRNGYGSTQYIRFSPDFTFGFEESLEVDTNPLLGAGKFATDPAVTLAHELIHAGHRLYGIAINPNRVFKVNTNAYYEMSGLEVSFEELRTFGGHDAKFIDSLQENEFRLYYYNKFKDIASTLNKAKSIVGTTASLQYMKNVFKEKYLLSEDTSGKFSVDKLKFDKLYKMLTEIYTEDNFVKFFKVLNRKTYLNFDKAVFKINIVPKVNYTIYDGFNLRNTNLAANFNGQNTEINNMNFTKLKNFTGLFEFYKLLCVRGIITSKTKSLDKGYNKALNDLCIKVNNWDLFFSPSEDNFTNDLNKGEEITSDTNIEAAEENISLDLIQQYYLTFNFDNEPENISIENLSSDIIGQLELMPNIERFPNGKKYELDKYTMFHYLRAQEFEHGKSRIALTNSVNEALLNPSRVYTFFSSDYVKKVNKATEAAMFLGWVEQLVYDFTDETSEVSTTDKIADITIIIPYIGPALNIGNMLYKDDFVGALIFSGAVILLEFIPEIAIPVLGTFALVSYIANKVLTVQTIDNALSKRNEKWDEVYKYIVTNWLAKVNTQIDLIRKKMKEALENQAEATKAIINYQYNQYTEEEKNNINFNIDDLSSKLNESINKAMININKFLNQCSVSYLMNSMIPYGVKRLEDFDASLKDALLKYIYDNRGTLIGQVDRLKDKVNNTLSTDIPFQLSKYVDNQRLLSTENLYFQGASHHHHHHHH
SEQ ID NO:21-rHC/A的核苷酸序列(His标签)
atgcatcatcaccatcaccacGAAAATCTATACTTCCAAGGAaaaaacatcatcaatactagcattctgaacctgcgttacgagagcaatcatctgattgatctgagccgttatgcaagcaagatcaacatcggtagcaaggtcaattttgacccgatcgataagaaccagatccagctgtttaatctggaatcgagcaaaattgaggttatcctgaaaaacgccattgtctacaactccatgtacgagaatttctccaccagcttctggattcgcatcccgaaatacttcaacagcattagcctgaacaacgagtatactatcatcaactgtatggagaacaacagcggttggaaggtgtctctgaactatggtgagatcatttggaccttgcaggacacccaagagatcaagcagcgcgtcgtgttcaagtactctcaaatgatcaacatttccgattacattaatcgttggatcttcgtgaccattacgaataaccgtctgaataacagcaagatttacatcaatggtcgcttgatcgatcagaaaccgattagcaacctgggtaatatccacgcaagcaacaacattatgttcaaattggacggttgccgcgatacccatcgttatatctggatcaagtatttcaacctgtttgataaagaactgaatgagaaggagatcaaagatttgtatgacaaccaatctaacagcggcattttgaaggacttctggggcgattatctgcaatacgataagccgtactatatgctgaacctgtatgatccgaacaaatatgtggatgtcaataatgtgggtattcgtggttacatgtatttgaagggtccgcgtggcagcgttatgacgaccaacatttacctgaactctagcctgtaccgtggtacgaaattcatcattaagaaatatgccagcggcaacaaagataacattgtgcgtaataacgatcgtgtctacatcaacgtggtcgtgaagaataaagagtaccgtctggcgaccaacgcttcgcaggcgggtgttgagaaaattctgagcgcgttggagatccctgatgtcggtaatctgagccaagtcgtggttatgaagagcaagaacgaccagggtatcactaacaagtgcaagatgaacctgcaagacaacaatggtaacgacatcggctttattggtttccaccagttcaacaatattgctaaactggtagcgagcaattggtacaatcgtcagattgagcgcagcagccgtactttgggctgtagctgggagtttatcccggtcgatgatggttggggcgaacgtccgctgtaa
SEQ ID NO:22-rHC/A的多肽序列(His标签)
MHHHHHHENLYFQGKNIINTSILNLRYESNHLIDLSRYASKINIGSKVNFDPIDKNQIQLFNLESSKIEVILKNAIVYNSMYENFSTSFWIRIPKYFNSISLNNEYTIINCMENNSGWKVSLNYGEIIWTLQDTQEIKQRVVFKYSQMINISDYINRWIFVTITNNRLNNSKIYINGRLIDQKPISNLGNIHASNNIMFKLDGCRDTHRYIWIKYFNLFDKELNEKEIKDLYDNQSNSGILKDFWGDYLQYDKPYYMLNLYDPNKYVDVNNVGIRGYMYLKGPRGSVMTTNIYLNSSLYRGTKFIIKKYASGNKDNIVRNNDRVYINVVVKNKEYRLATNASQAGVEKILSALEIPDVGNLSQVVVMKSKNDQGITNKCKMNLQDNNGNDIGFIGFHQFNNIAKLVASNWYNRQIERSSRTLGCSWEFIPVDDGWGERPL
SEQ ID NO:23-rLC/A的核苷酸序列(His标签)
ATGCCGTTTGTGAACAAGCAGTTCAACTATAAAGATCCGGTTAATGGTGTGGATATCGCCTATATCAAAATTCCGAATGCAGGTCAGATGCAGCCGGTTAAAGCCTTTAAAATCCATAACAAAATTTGGGTGATTCCGGAACGTGATACCTTTACCAATCCGGAAGAAGGTGATCTGAATCCGCCTCCGGAAGCAAAACAGGTTCCGGTTAGCTATTATGATAGCACCTATCTGAGCACCGATAACGAGAAAGATAACTATCTGAAAGGTGTGACCAAACTGTTTGAACGCATTTATAGTACCGATCTGGGTCGTATGCTGCTGACCAGCATTGTTCGTGGTATTCCGTTTTGGGGTGGTAGCACCATTGATACCGAACTGAAAGTTATTGACACCAACTGCATTAATGTGATTCAGCCGGATGGTAGCTATCGTAGCGAAGAACTGAATCTGGTTATTATTGGTCCGAGCGCAGATATCATTCAGTTTGAATGTAAATCCTTTGGCCACGAAGTTCTGAATCTGACCCGTAATGGTTATGGTAGTACCCAGTATATTCGTTTCAGTCCGGATTTTACCTTTGGCTTTGAAGAAAGCCTGGAAGTTGATACAAATCCGCTGTTAGGTGCAGGTAAATTTGCAACCGATCCGGCAGTTACCCTGGCACATGAACTGATTCATGCCGGTCATCGTCTGTATGGTATTGCAATTAATCCGAACCGTGTGTTCAAAGTGAATACCAACGCATATTATGAAATGAGCGGTCTGGAAGTGTCATTTGAAGAACTGCGTACCTTTGGTGGTCATGATGCCAAATTTATCGATAGCCTGCAAGAAAATGAATTTCGCCTGTACTACTATAACAAATTCAAGGATATTGCGAGCACCCTGAATAAAGCCAAAAGCATTGTTGGCACCACCGCAAGCCTGCAGTATATGAAAAATGTGTTTAAAGAAAAATATCTGCTGAGCGAAGATACCAGCGGTAAATTTAGCGTTGACAAACTGAAATTCGATAAACTGTACAAGATGCTGACCGAGATTTATACCGAAGATAACTTCGTGAAGTTTTTCAAAGTGCTGAACCGCAAAACCTACCTGAACTTTGATAAAGCCGTGTTCAAAATCAACATCGTGCCGAAAGTGAACTATACCATCTATGATGGTTTTAACCTGCGCAATACCAATCTGGCAGCAAACTTTAATGGTCAGAACACCGAAATCAACAACATGAACTTTACCAAACTGAAGAACTTCACCGGTCTGTTTGAAGAGAATCTGTATTTCCAGGGTGCAAGTCATCATCACCATCACCACCATCATTAA
SEQ ID NO:24-rLC/A的多肽序列(His标签)
MPFVNKQFNYKDPVNGVDIAYIKIPNAGQMQPVKAFKIHNKIWVIPERDTFTNPEEGDLNPPPEAKQVPVSYYDSTYLSTDNEKDNYLKGVTKLFERIYSTDLGRMLLTSIVRGIPFWGGSTIDTELKVIDTNCINVIQPDGSYRSEELNLVIIGPSADIIQFECKSFGHEVLNLTRNGYGSTQYIRFSPDFTFGFEESLEVDTNPLLGAGKFATDPAVTLAHELIHAGHRLYGIAINPNRVFKVNTNAYYEMSGLEVSFEELRTFGGHDAKFIDSLQENEFRLYYYNKFKDIASTLNKAKSIVGTTASLQYMKNVFKEKYLLSEDTSGKFSVDKLKFDKLYKMLTEIYTEDNFVKFFKVLNRKTYLNFDKAVFKINIVPKVNYTIYDGFNLRNTNLAANFNGQNTEINNMNFTKLKNFTGLFEENLYFQGASHHHHHHHH
SEQ ID NO:25-rBoNT/FA(0)的核苷酸序列(His标签)
ATGCCGGTTGTGATTAACAGCTTCAATTATGATGATCCGGTGAACGATAACACCATCATTTATATCCGTCCGCCTTATTATGAAACCAGCAACACCTATTTCAAAGCCTTCCAGATTATGGATAACGTGTGGATTATTCCGGAACGTTATCGTCTGGGTATTGATCCGAGCCTGTTTAATCCGCCTGTTAGCCTGAAAGCAGGTAGTGATGGTTATTTTGATCCGAATTATCTGAGCACCAACACCGAGAAAAACAAATACCTGCAGATTATGATCAAGCTGTTCAAACGCATTAATAGCAAACCGGCAGGTCAGATTCTGCTGGAAGAAATCAAAAATGCAATTCCGTATCTGGGCAACAGCTATACCCAAGAAGAACAGTTTACCACCAATAATCGTACCGTGAGCTTTAATGTTAAACTGGCCAATGGTAATATCGTTCAGCAGATGGCAAATCTGATTATTTGGGGTCCGGGTCCTGATCTGACCACAAATAAAACCGGTGGTATCATCTATAGCCCGTATCAGAGCATGGAAGCAACCCCGTATAAAGATGGTTTTGGTAGCATTATGACCGTGGAATTTAGTCCGGAATATGCAACCGCCTTTAACGATATTTCAATTGCAAGCCATAGTCCGTCGCTGTTTATCAAAGATCCGGCACTGATTCTGATGCACCAGCTGATTTATGTTCTGCATGGTCTGTATGGCACCTATATCACCGAATACAAAATTACCCCGAATGTGGTTCAGAGCTATATGAAAGTTACCAAACCGATTACCAGCGCAGAATTTCTGACCTTTGGTGGTCGTGATCGCAATATTGTTCCGCAGAGCATTCAGAGCCAGCTGTATAACAAAGTTCTGAGCGATTATAAACGTATTGCCAGCCGTCTGAATAAAGTTAATACCGCAACCGCACTGATCAACATCGATGAATTCAAAAACCTGTACGAGTGGAAATACCAGTTTGCCAAAGATAGCAATGGTGTGTATAGCGTGGATCTGAACAAATTTGAGCAGCTGTACAAAAAAATCTATAGCTTCACCGAATTCAACCTGGCCTATGAGTTTAAAATCAAAACCCGTCTGGGTTATCTGGCCGAAAATTTTGGTCCGTTTTATCTGCCGAATCTGCTGGATGATAGCATTTATACCGAAGTGGATGGTTTTAACATTGGTGCACTGAGCATTAACTATCAGGGTCAGAATATTGGCAGCGATATCAACAGCATCAAAAAACTGCAAGGTCAGGGTGTTGTTAGCCGTGTTGTTCGTCTGTGTAGCAATAGCAATACCAAAAACAGCCTGTGCATTACCGTTAATAATCGCGACCTGTTTTTTATCGCAAGCCAAGAAAGCTATGGCGAGAATACCATTAACACCTATAAAGAGATTGACGATACCACCACACTGGATCCGAGCTTTGAAGATATTCTGGATAAAGTGATCCTGAACTTCAACGAACAGGTTATTCCGCAGATGCCGAATCGTAATGTTAGCACCGATATTCAGAAAGACAACTACATCCCGAAATACGATTATAACCGCACCGACATTATCGATAGCTATGAAGTTGGTCGCAACTACAACACCTTTTTCTATCTGAATGCCCAGAAATTTAGCCCGAACGAAAGCAATATTACCCTGACCAGCAGCTTTGATACAGGTCTGTTAGAAGGTAGCAAAGTGTATACCTTTTTCAGCAGCGATTTCATTAACAACATCAACAAACCGGTTCAGGCCCTGCTGTTTATTGAATGGGTTAAACAGGTGATTCGCGATTTTACCACCGAAGCAACCAAAACCTCAACCGTTGATAAACTGAAAGATATTAGCCTGGTGGTGCCGTATATTGGTCTGGCACTGAATATTGGTGATGAGATCTACAAACAGCATTTTGCAGAAGCAGTTGAACTGGTTGGTGCAGGTCTGCTGCTGGAATTTTCACCGGAATTTCTTATTCCGACGCTGCTGATTTTTACCATCAAAGGTTATCTGACCGGTAGCATTCGCGATAAAGACAAAATCATTAAAACCCTGGATAACGCCCTGAATGTTCGTGATCAGAAATGGAAAGAACTGTATCGTTGGGTTGTTAGCAAATGGCTGACCACCATTAATACGCAGTTCAACAAACGCAAAGAACAAATGTACAAAGCCCTGAAAAATCAGGCCACCGCCATTAAAAAGATCATCGAGAACAAATATAACAACTATACCACCGATGAAAAAAGCAAGATCGATAGCAGCTATAACATCAACGAAATTGAACGCACCCTGAACGAAAAAATCAATCTGGCCATGAAAAACATCGAGCAGTTTATTACCGAAAGCAGCATTGCCTATCTGATCAATATCATCAACAACGAAACGATCCAGAAACTGAAAAGCTATGATGACCTGGTTCGTCGTTATCTGCTGGGTTATATTCGTAATCATAGCAGCATTCTGGGCAATAGCGTTGAAGAACTGAATTCCAAAGTGAACAACCATCTGGATAATGGCATTCCGTTTGAACTGAGCAGTTATACCAATGATAGCCTGCTGATCCGCTACTTCAATAAAAACTATGGCGAACTGAAGTACAACTGCATTCTGAACATCAAATATGAGATGGATCGTGACAAACTGGTTGATAGCAGCGGTTATCGTAGCCGTATCAATATTGGTACAGGCGTCAAATTTAGCGAGATCGATAAAAATCAAGTGCAGCTGAGCAATCTGGAATCCAGCAAAATTGAAGTCATTCTGAATAACGGCGTCATCTATAACAGCATGTATGAAAACTTTTCGACCAGCTTTTGGATTCGCATTCCGAAATACTTTCGCAACATCAATAACGAGTACAAGATCATCAGCTGTATGCAGAATAATAGCGGTTGGGAAGTGAGCCTGAATTTTAGCAATATGAACTCGAAAATCATCTGGACCCTGCAGGATACCGAAGGTATCAAAAAAACCGTTGTGTTTCAGTACACCCAGAACATTAACATTAGCGACTATATCAACCGCTGGATCTTTGTGACCATTACAAATAATCGTCTGAGCAACAGCAAAATCTACATTAATGGTCGCCTGATCAACGAAGAAAGCATTAGCGATCTGGGTAATATCCATGCCAGCAACAACATTATGTTTAAACTGGATGGTTGCCGTGATCCGCATCGTTATATCTGGATTAAATACTTTAACCTGTTTGACAAAGAGCTGAACAAGAAAGAAATTAAAGATCTGTACGACAACCAGAGCAATAGCGGTATTCTGAAAGATTTCTGGGGTGATTATCTGCAGTATGACAAACCGTATTATATGCTGAATCTGTATGACCCGAATAAGTATCTGGATGTGAATAATGTTGGCATCCGTGGCTATATGTATCTGAAAGGTCCGCGTGGTCGTATTGTGACCACCAACATTTATCTGAATAGCACCCTGTATATGGGCACCAAATTCATCATTAAGAAATATGCCAGCGGCAACAAAGATAACATTGTGCGTAATAATGATCGCGTGTATATTAACGTGGTGGTGAAGAATAAAGAATATCGCCTGGCAACCAATGCAAGCCAGGCAGGCGTTGAAAAAATTCTGAGCGCAGTTGAAATCCCGGATGTTGGTAATCTGAGCCAGGTTGTTGTGATGAAAAGCGAAAATGATCAGGGCATTCGCAACAAGTGTAAAATGAATCTGCAAGACAATAACGGCAACGATATTGGCTTTATCGGCTTTCACCAGTTTAATAACATTGCAAAACTGGTGGCCAGCAACTGGTATAACCGTCAGATTGGTAAAGCAAGCCGTACCTTTGGTTGTAGCTGGGAATTTATCCCGGTTGATGATGGTTGGGGTGAAAGCAGCCTGGAAAATCTGTATTTCCAGGGTGCCAGTCATCATCACCACCATCACCATCACTGA
SEQ ID NO:26-rBoNT/FA(0)的多肽序列(His标签)
MPVVINSFNYDDPVNDNTIIYIRPPYYETSNTYFKAFQIMDNVWIIPERYRLGIDPSLFNPPVSLKAGSDGYFDPNYLSTNTEKNKYLQIMIKLFKRINSKPAGQILLEEIKNAIPYLGNSYTQEEQFTTNNRTVSFNVKLANGNIVQQMANLIIWGPGPDLTTNKTGGIIYSPYQSMEATPYKDGFGSIMTVEFSPEYATAFNDISIASHSPSLFIKDPALILMHQLIYVLHGLYGTYITEYKITPNVVQSYMKVTKPITSAEFLTFGGRDRNIVPQSIQSQLYNKVLSDYKRIASRLNKVNTATALINIDEFKNLYEWKYQFAKDSNGVYSVDLNKFEQLYKKIYSFTEFNLAYEFKIKTRLGYLAENFGPFYLPNLLDDSIYTEVDGFNIGALSINYQGQNIGSDINSIKKLQGQGVVSRVVRLCSNSNTKNSLCITVNNRDLFFIASQESYGENTINTYKEIDDTTTLDPSFEDILDKVILNFNEQVIPQMPNRNVSTDIQKDNYIPKYDYNRTDIIDSYEVGRNYNTFFYLNAQKFSPNESNITLTSSFDTGLLEGSKVYTFFSSDFINNINKPVQALLFIEWVKQVIRDFTTEATKTSTVDKLKDISLVVPYIGLALNIGDEIYKQHFAEAVELVGAGLLLEFSPEFLIPTLLIFTIKGYLTGSIRDKDKIIKTLDNALNVRDQKWKELYRWVVSKWLTTINTQFNKRKEQMYKALKNQATAIKKIIENKYNNYTTDEKSKIDSSYNINEIERTLNEKINLAMKNIEQFITESSIAYLINIINNETIQKLKSYDDLVRRYLLGYIRNHSSILGNSVEELNSKVNNHLDNGIPFELSSYTNDSLLIRYFNKNYGELKYNCILNIKYEMDRDKLVDSSGYRSRINIGTGVKFSEIDKNQVQLSNLESSKIEVILNNGVIYNSMYENFSTSFWIRIPKYFRNINNEYKIISCMQNNSGWEVSLNFSNMNSKIIWTLQDTEGIKKTVVFQYTQNINISDYINRWIFVTITNNRLSNSKIYINGRLINEESISDLGNIHASNNIMFKLDGCRDPHRYIWIKYFNLFDKELNKKEIKDLYDNQSNSGILKDFWGDYLQYDKPYYMLNLYDPNKYLDVNNVGIRGYMYLKGPRGRIVTTNIYLNSTLYMGTKFIIKKYASGNKDNIVRNNDRVYINVVVKNKEYRLATNASQAGVEKILSAVEIPDVGNLSQVVVMKSENDQGIRNKCKMNLQDNNGNDIGFIGFHQFNNIAKLVASNWYNRQIGKASRTFGCSWEFIPVDDGWGESSLENLYFQGASHHHHHHHH
SEQ ID NO:27-rLHN/FA的核苷酸序列(His标签)
ATGCCGGTTGTGATTAACAGCTTCAATTATGATGATCCGGTGAACGATAACACCATCATTTATATCCGTCCGCCTTATTATGAAACCAGCAACACCTATTTCAAAGCCTTCCAGATTATGGATAACGTGTGGATTATTCCGGAACGTTATCGTCTGGGTATTGATCCGAGCCTGTTTAATCCGCCTGTTAGCCTGAAAGCAGGTAGTGATGGTTATTTTGATCCGAATTATCTGAGCACCAACACCGAGAAAAACAAATACCTGCAGATTATGATCAAGCTGTTCAAACGCATTAATAGCAAACCGGCAGGTCAGATTCTGCTGGAAGAAATCAAAAATGCAATTCCGTATCTGGGCAACAGCTATACCCAAGAAGAACAGTTTACCACCAATAATCGTACCGTGAGCTTTAATGTTAAACTGGCCAATGGTAATATCGTTCAGCAGATGGCAAATCTGATTATTTGGGGTCCGGGTCCTGATCTGACCACAAATAAAACCGGTGGTATCATCTATAGCCCGTATCAGAGCATGGAAGCAACCCCGTATAAAGATGGTTTTGGTAGCATTATGACCGTGGAATTTAGTCCGGAATATGCAACCGCCTTTAACGATATTTCAATTGCAAGCCATAGTCCGTCGCTGTTTATCAAAGATCCGGCACTGATTCTGATGCATGAACTGATTCATGTTCTGCATGGTCTGTATGGCACCTATATTACCGAATACAAAATTACCCCGAATGTGGTGCAGAGCTATATGAAAGTTACCAAACCGATTACCAGCGCAGAATTTCTGACCTTTGGTGGTCGTGATCGCAATATTGTTCCGCAGAGCATTCAGAGCCAGCTGTATAACAAAGTTCTGAGCGATTATAAACGTATTGCCAGCCGTCTGAATAAAGTTAATACCGCAACCGCACTGATCAACATCGATGAATTCAAAAACCTGTACGAGTGGAAATACCAGTTTGCCAAAGATAGCAATGGTGTGTATAGCGTGGATCTGAACAAATTTGAGCAGCTGTACAAAAAAATCTATAGCTTCACCGAATTCAACCTGGCCTATGAGTTTAAAATCAAAACCCGTCTGGGTTATCTGGCCGAAAATTTTGGTCCGTTTTATCTGCCGAATCTGCTGGATGATAGCATTTATACCGAAGTGGATGGTTTTAACATTGGTGCACTGAGCATTAACTATCAGGGTCAGAATATTGGCAGCGATATCAACAGCATCAAAAAACTGCAAGGTCAGGGTGTTGTTAGCCGTGTTGTTCGTCTGTGTAGCAATAGCAATACCAAAAACAGCCTGTGCATTACCGTTAATAATCGCGACCTGTTTTTTATCGCAAGCCAAGAAAGCTATGGCGAGAATACCATTAACACCTATAAAGAGATTGACGATACCACCACACTGGATCCGAGCTTTGAAGATATTCTGGATAAAGTGATCCTGAACTTCAACGAACAGGTTATTCCGCAGATGCCGAATCGTAATGTTAGCACCGATATTCAGAAAGACAACTACATCCCGAAATACGATTATAACCGCACCGACATTATCGATAGCTATGAAGTTGGTCGCAACTACAACACCTTTTTCTATCTGAATGCCCAGAAATTTAGCCCGAACGAAAGCAATATTACCCTGACCAGCAGCTTTGATACAGGTCTGTTAGAAGGTAGCAAAGTGTATACCTTTTTCAGCAGCGATTTCATTAACAACATCAACAAACCGGTTCAGGCCCTGCTGTTTATTGAATGGGTTAAACAGGTGATTCGCGATTTTACCACCGAAGCAACCAAAACCTCAACCGTTGATAAACTGAAAGATATTAGCCTGGTGGTGCCGTATATTGGTCTGGCACTGAATATTGGTGATGAGATCTACAAACAGCATTTTGCAGAAGCAGTTGAACTGGTTGGTGCAGGTCTGCTGCTGGAATTTTCACCGGAATTTCTTATTCCGACGCTGCTGATTTTTACCATCAAAGGTTATCTGACCGGTAGCATTCGCGATAAAGACAAAATCATTAAAACCCTGGATAACGCCCTGAATGTTCGTGATCAGAAATGGAAAGAACTGTATCGTTGGGTTGTTAGCAAATGGCTGACCACCATTAATACGCAGTTCAACAAACGCAAAGAACAAATGTACAAAGCCCTGAAAAATCAGGCCACCGCCATTAAAAAGATCATCGAGAACAAATATAACAACTATACCACCGATGAAAAAAGCAAGATCGATAGCAGCTATAACATCAACGAAATTGAACGCACCCTGAACGAAAAAATCAATCTGGCCATGAAAAACATCGAGCAGTTTATTACAGAAAGCAGCATTGCCTACCTGATCAATATCATCAACAACGAAACCATTCAGAAACTGAAAAGCTATGATGACCTGGTTCGTCGTTATCTGCTGGGTTATATTCGTAATCATAGCAGCATTCTGGGCAATAGCGTTGAAGAACTGAATTCCAAAGTGAACAACCATCTGGATAATGGCATTCCGTTTGAACTGAGCAGTTATACCAATGATAGCCTGCTGATCCGCTACTTCAATAAAAACTATGGCGAAGAGAACCTGTATTTCCAGGGTGCCAGTCATCATCACCACCATCACCATCACTGA
SEQ ID NO:28-rLHN/FA的多肽序列(His标签)
MPVVINSFNYDDPVNDNTIIYIRPPYYETSNTYFKAFQIMDNVWIIPERYRLGIDPSLFNPPVSLKAGSDGYFDPNYLSTNTEKNKYLQIMIKLFKRINSKPAGQILLEEIKNAIPYLGNSYTQEEQFTTNNRTVSFNVKLANGNIVQQMANLIIWGPGPDLTTNKTGGIIYSPYQSMEATPYKDGFGSIMTVEFSPEYATAFNDISIASHSPSLFIKDPALILMHELIHVLHGLYGTYITEYKITPNVVQSYMKVTKPITSAEFLTFGGRDRNIVPQSIQSQLYNKVLSDYKRIASRLNKVNTATALINIDEFKNLYEWKYQFAKDSNGVYSVDLNKFEQLYKKIYSFTEFNLAYEFKIKTRLGYLAENFGPFYLPNLLDDSIYTEVDGFNIGALSINYQGQNIGSDINSIKKLQGQGVVSRVVRLCSNSNTKNSLCITVNNRDLFFIASQESYGENTINTYKEIDDTTTLDPSFEDILDKVILNFNEQVIPQMPNRNVSTDIQKDNYIPKYDYNRTDIIDSYEVGRNYNTFFYLNAQKFSPNESNITLTSSFDTGLLEGSKVYTFFSSDFINNINKPVQALLFIEWVKQVIRDFTTEATKTSTVDKLKDISLVVPYIGLALNIGDEIYKQHFAEAVELVGAGLLLEFSPEFLIPTLLIFTIKGYLTGSIRDKDKIIKTLDNALNVRDQKWKELYRWVVSKWLTTINTQFNKRKEQMYKALKNQATAIKKIIENKYNNYTTDEKSKIDSSYNINEIERTLNEKINLAMKNIEQFITESSIAYLINIINNETIQKLKSYDDLVRRYLLGYIRNHSSILGNSVEELNSKVNNHLDNGIPFELSSYTNDSLLIRYFNKNYGEENLYFQGASHHHHHHHH
SEQ ID NO:29-rHC/FA的核苷酸序列(His标签)
ATGCTGAAGTATAACTGCATCCTGAACATCAAATATGAGATGGATCGTGATAAACTGGTTGATAGCAGCGGTTATCGTAGCCGTATCAATATTGGCACCGGTGTGAAATTTAGCGAGATCGATAAAAATCAGGTGCAGCTGAGCAATCTGGAAAGCAGCAAAATTGAAGTGATTCTGAATAACGGCGTGATCTACAATAGCATGTATGAAAACTTTTCGACCAGCTTCTGGATTCGCATTCCGAAATACTTTCGCAACATCAACAACGAGTACAAGATTATCAGCTGTATGCAGAATAATAGCGGTTGGGAAGTTAGCCTGAATTTCAGCAATATGAACAGCAAAATCATTTGGACCCTGCAGGATACCGAAGGTATCAAAAAAACCGTTGTGTTTCAGTACACCCAGAACATTAACATCAGCGATTACATTAACCGCTGGATCTTTGTGACCATTACCAATAATCGTCTGAGCAACAGCAAGATCTATATTAACGGTCGCCTGATTAACGAAGAGAGCATTAGCGATCTGGGTAATATTCATGCCAGCAACAACATCATGTTTAAACTGGATGGTTGTCGTGATCCGCATCGTTATATTTGGATCAAATACTTCAACCTGTTTGATAAAGAACTGAACAAAAAAGAAATCAAAGACCTGTATGATAACCAGAGCAATAGCGGCATTCTGAAAGATTTTTGGGGTGATTATCTGCAGTATGACAAACCGTATTACATGCTGAATCTGTACGATCCGAACAAATATCTGGATGTGAATAATGTGGGTATCCGTGGCTATATGTATCTGAAAGGTCCGCGTGGTCGTATTGTTACCACCAACATTTATCTGAATAGCACCCTGTATATGGGCACCAAATTCATCATTAAAAAGTATGCCAGCGGCAACAAAGATAACATTGTGCGTAATAATGATCGCGTGTATATCAATGTGGTGGTGAAGAATAAAGAATATCGTCTGGCCACCAATGCAAGCCAGGCAGGCGTTGAAAAAATTCTGAGCGCAGTTGAAATTCCGGATGTTGGTAATCTGAGCCAGGTTGTTGTTATGAAAAGCGAAAATGATCAGGGCATTCGCAACAAATGCAAAATGAATCTGCAGGACAATAACGGCAACGATATTGGTTTTATTGGCTTCCACCAGTTCAACAACATTGCAAAACTGGTGGCGAGCAATTGGTATAATCGTCAGATTGGTAAAGCAAGCCGTACCTTTGGTTGTAGCTGGGAATTTATTCCGGTTGATGATGGTTGGGGTGAAAGCAGCCTGGAAAATCTGTATTTTCAGGGTGCAAGTCATCATCACCACCATCACCATCATTAA
SEQ ID NO:30-rHC/FA的多肽序列(His标签)
MLKYNCILNIKYEMDRDKLVDSSGYRSRINIGTGVKFSEIDKNQVQLSNLESSKIEVILNNGVIYNSMYENFSTSFWIRIPKYFRNINNEYKIISCMQNNSGWEVSLNFSNMNSKIIWTLQDTEGIKKTVVFQYTQNINISDYINRWIFVTITNNRLSNSKIYINGRLINEESISDLGNIHASNNIMFKLDGCRDPHRYIWIKYFNLFDKELNKKEIKDLYDNQSNSGILKDFWGDYLQYDKPYYMLNLYDPNKYLDVNNVGIRGYMYLKGPRGRIVTTNIYLNSTLYMGTKFIIKKYASGNKDNIVRNNDRVYINVVVKNKEYRLATNASQAGVEKILSAVEIPDVGNLSQVVVMKSENDQGIRNKCKMNLQDNNGNDIGFIGFHQFNNIAKLVASNWYNRQIGKASRTFGCSWEFIPVDDGWGESSLENLYFQGASHHHHHHHH
SEQ ID NO:31-rLC/FA的核苷酸序列(His标签)
ATGCCGGTTGTGATTAACAGCTTCAATTATGATGATCCGGTGAACGATAACACCATCATTTATATCCGTCCGCCTTATTATGAAACCAGCAACACCTATTTCAAAGCCTTCCAGATTATGGATAACGTGTGGATTATTCCGGAACGTTATCGTCTGGGTATTGATCCGAGCCTGTTTAATCCGCCTGTTAGCCTGAAAGCAGGTAGTGATGGTTATTTTGATCCGAATTATCTGAGCACCAACACCGAGAAAAACAAATACCTGCAGATTATGATCAAGCTGTTCAAACGCATTAATAGCAAACCGGCAGGTCAGATTCTGCTGGAAGAAATCAAAAATGCAATTCCGTATCTGGGCAACAGCTATACCCAAGAAGAACAGTTTACCACCAATAATCGTACCGTGAGCTTTAATGTTAAACTGGCCAATGGTAATATCGTTCAGCAGATGGCAAATCTGATTATTTGGGGTCCGGGTCCTGATCTGACCACAAATAAAACCGGTGGTATCATCTATAGCCCGTATCAGAGCATGGAAGCAACCCCGTATAAAGATGGTTTTGGTAGCATTATGACCGTGGAATTTAGTCCGGAATATGCAACCGCCTTTAACGATATTTCAATTGCAAGCCATAGTCCGTCGCTGTTTATCAAAGATCCGGCACTGATTCTGATGCATGAACTGATTCATGTTCTGCATGGTCTGTATGGCACCTATATTACCGAATACAAAATTACCCCGAATGTGGTGCAGAGCTATATGAAAGTTACCAAACCGATTACCAGCGCAGAATTTCTGACCTTTGGTGGTCGTGATCGCAATATTGTTCCGCAGAGCATTCAGAGCCAGCTGTATAACAAAGTTCTGAGCGATTATAAACGTATTGCCAGCCGTCTGAATAAAGTTAATACCGCAACCGCACTGATCAACATCGATGAATTCAAAAACCTGTACGAGTGGAAATACCAGTTTGCCAAAGATAGCAATGGTGTGTATAGCGTGGATCTGAACAAATTTGAGCAGCTGTACAAAAAAATCTATAGCTTCACCGAATTCAACCTGGCCTATGAGTTTAAAATCAAAACCCGTCTGGGTTATCTGGCCGAAAATTTTGGTCCGTTTTATCTGCCGAATCTGCTGGATGATAGCATTTATACCGAAGTGGATGGTTTTAACATTGGTGCACTGAGCATTAACTATCAGGGTCAGAATATTGGCAGCGATATCAACAGCATCAAAAAACTGCAAGGTCAGGGTGTTGTTAGCCGTGTTGTTCGTCTGTGTAGCAATAGCGAAAATCTGTATTTTCAGGGTGCCAGTCATCATCACCACCATCACCATCACTGA
SEQ ID NO:32-rLC/FA的多肽序列(His标签)
MPVVINSFNYDDPVNDNTIIYIRPPYYETSNTYFKAFQIMDNVWIIPERYRLGIDPSLFNPPVSLKAGSDGYFDPNYLSTNTEKNKYLQIMIKLFKRINSKPAGQILLEEIKNAIPYLGNSYTQEEQFTTNNRTVSFNVKLANGNIVQQMANLIIWGPGPDLTTNKTGGIIYSPYQSMEATPYKDGFGSIMTVEFSPEYATAFNDISIASHSPSLFIKDPALILMHELIHVLHGLYGTYITEYKITPNVVQSYMKVTKPITSAEFLTFGGRDRNIVPQSIQSQLYNKVLSDYKRIASRLNKVNTATALINIDEFKNLYEWKYQFAKDSNGVYSVDLNKFEQLYKKIYSFTEFNLAYEFKIKTRLGYLAENFGPFYLPNLLDDSIYTEVDGFNIGALSINYQGQNIGSDINSIKKLQGQGVVSRVVRLCSNSENLYFQGASHHHHHHHH
SEQ ID NO:33-rBoNT/F(0)的核苷酸序列(His标签)
ATGCCGGTTGTGATTAACAGCTTCAATTATAACGATCCGGTGAACGATGATACCATCCTGTATATGCAGATTCCGTATGAAGAGAAAAGCAAAAAGTACTACAAAGCCTTTGAGATCATGCGCAACGTTTGGATTATTCCGGAACGTAATACCATTGGCACCGATCCGAGCGATTTTGATCCGCCTGCAAGCCTGGAAAATGGTAGCAGCGCATATTATGATCCGAATTATCTGACCACCGATGCCGAAAAAGATCGTTATCTGAAAACCACCATCAAACTGTTCAAACGCATTAATAGCAATCCGGCAGGCGAAGTTCTGCTGCAAGAAATTAGCTATGCAAAACCGTATCTGGGCAATGAACATACCCCGATTAATGAATTTCATCCGGTTACACGTACCACGAGCGTTAACATTAAAAGCAGCACCAATGTGAAGTCCAGCATTATTCTGAATCTGCTGGTTTTAGGTGCAGGTCCGGATATTTTTGAAAATTCAAGCTATCCGGTGCGCAAACTGATGGATAGCGGTGGTGTGTATGATCCGTCAAATGATGGTTTTGGCAGCATTAACATTGTGACCTTTAGTCCGGAATATGAATACACCTTCAACGATATTAGCGGTGGCTATAATAGCAGCACCGAAAGTTTTATTGCAGATCCGGCAATTAGCCTGGCACACCAGCTGATTTATGCACTGCATGGTCTGTATGGTGCACGTGGTGTTACCTATAAAGAAACCATTAAAGTTAAACAGGCACCGCTGATGATTGCGGAAAAACCGATTCGTCTGGAAGAATTTCTGACCTTTGGTGGTCAGGATCTGAACATTATTACCAGCGCAATGAAAGAGAAAATCTATAATAACCTGCTGGCCAACTATGAGAAAATTGCAACCCGTCTGAGCCGTGTTAATAGCGCACCTCCTGAATATGATATCAACGAGTATAAAGACTATTTTCAGTGGAAATACGGCCTGGATAAAAATGCAGATGGTAGCTATACCGTGAACGAGAACAAATTTAACGAGATCTACAAAAAACTGTATAGCTTCACCGAAATCGATCTGGCCAACAAATTCAAAGTGAAATGCCGCAACACCTACTTCATCAAATATGGCTTTCTGAAAGTTCCGAACCTGCTTGATGATGATATCTATACCGTTAGCGAAGGCTTTAACATTGGTAATCTGGCCGTTAATAATCGCGGTCAGAACATTAAACTGAACCCGAAAATTATCGATAGCATCCCGGATAAAGGCCTGGTTGAAAAAATTGTGAAATTCTGCAAAAGCGTGATTCCGCGTAAAGGCACCAAAGCACCGCCTCGTCTGTGTATTCGTGTGAATAATCGTGAACTGTTTTTTGTTGCAAGCGAGAGCAGCTATAACGAGAATGATATTAACACCCCGAAAGAGATTGACGATACCACCAATCTGAATAACAACTATCGCAACAATCTGGATGAAGTGATCCTGGATTATAACAGCGAAACCATTCCGCAGATTAGCAATCAGACCCTGAATACCCTGGTTCAGGATGATAGCTATGTTCCGCGTTATGATAGCAATGGCACCAGCGAAATTGAAGAACATAATGTGGTTGATCTGAACGTGTTCTTTTATCTGCATGCACAGAAAGTGCCGGAAGGTGAAACCAATATTAGCCTGACCAGCAGCATTGATACCGCACTGAGCGAAGAAAGCCAGGTTTATACCTTTTTTAGCAGCGAATTCATCAACACCATTAACAAACCGGTTCATGCAGCACTGTTTATTAGCTGGATTAATCAGGTGATTCGCGATTTTACCACCGAAGCAACCCAGAAAAGCACCTTTGATAAAATTGCCGATATTAGTCTGGTGGTGCCGTATGTTGGTCTGGCACTGAATATTGGTAATGAAGTGCAGAAAGAGAACTTTAAAGAAGCCTTCGAACTGTTAGGTGCCGGTATTCTGCTGGAATTTGTGCCGGAACTGCTGATTCCGACCATTCTGGTTTTTACCATTAAGAGCTTTATTGGCAGCAGCGAGAACAAGAACAAAATCATTAAAGCCATCAACAACAGCCTGATGGAACGCGAAACCAAATGGAAAGAAATTTACAGCTGGATTGTGAGCAATTGGCTGACCCGTATCAATACCCAGTTTAACAAACGCAAAGAACAAATGTATCAGGCCCTGCAGAATCAGGTTGATGCAATTAAAACCGTGATCGAATACAAATACAACAACTATACCAGCGACGAACGTAATCGCCTGGAAAGCGAATACAACATTAATAACATTCGCGAAGAACTGAACAAAAAAGTGAGCCTGGCAATGGAAAACATCGAACGTTTTATTACCGAAAGCAGCATCTTCTACCTGATGAAACTGATTAACGAAGCCAAAGTTAGCAAACTGCGCGAATATGATGAAGGCGTTAAAGAATATCTGCTGGACTATATTAGCGAACATCGTAGCATTCTGGGTAATAGCGTTCAAGAGCTGAATGATCTGGTTACCAGCACACTGAATAATAGCATTCCGTTTGAACTGAGCAGCTACACCAACGATAAAATCCTGATCCTGTACTTCAACAAACTGTACAAGAAGATCAAGGACAACAGCATACTGGATATGCGCTATGAAAACAACAAGTTCATTGATATCAGCGGCTATGGTAGCAACATTAGCATTAATGGTGATGTGTATATCTACAGCACCAACCGCAATCAGTTTGGTATTTATAGCAGCAAACCGAGCGAAGTTAATATTGCGCAGAATAACGATATCATCTACAACGGTCGCTATCAGAACTTTAGCATTAGCTTTTGGGTTCGCATTCCGAAATACTTTAACAAGGTGAACCTGAACAACGAGTACACCATTATTGATTGCATTCGCAATAATAACAGCGGCTGGAAAATCAGCCTGAACTATAACAAAATTATCTGGACCCTGCAGGATACCGCAGGTAATAATCAGAAACTGGTGTTTAACTACACCCAGATGATTAGCATCAGCGACTATATCAACAAATGGATCTTTGTGACCATTACCAACAATCGTCTGGGTAACAGCCGCATTTATATCAATGGCAATCTGATCGACGAAAAAAGCATTTCAAATCTGGGCGATATTCACGTGAGCGATAACATTCTGTTCAAAATTGTTGGCTGCAACGATACCCGTTATGTTGGTATTCGTTACTTCAAAGTGTTTGATACGGAACTGGGCAAAACGGAAATTGAAACCCTGTATAGTGATGAACCGGATCCGAGCATTCTGAAAGATTTTTGGGGTAATTATCTGCTGTACAACAAACGCTACTATCTGCTGAACCTGCTGCGTACCGATAAAAGCATTACACAGAATAGCAACTTTCTGAACATCAATCAGCAGCGTGGTGTTTATCAGAAACCGAACATTTTTAGCAACACCCGTCTGTATACCGGTGTGGAAGTTATTATTCGTAAAAACGGTAGCACCGATATCAGCAACACCGATAACTTTGTGCGTAAAAATGACCTGGCCTATATTAACGTTGTTGATCGTGATGTTGAGTATCGTCTGTATGCGGATATTAGCATTGCCAAACCGGAAAAGATTATCAAACTGATCCGTACCAGCAACAGCAATAATTCACTGGGTCAGATTATCGTGATGGACAGCATTGGTAACAATTGCACCATGAATTTCCAGAACAATAACGGTGGTAATATTGGCCTGCTGGGCTTTCATAGCAATAATCTGGTTGCAAGCAGCTGGTATTACAACAACATCCGTAAAAATACCAGCAGTAATGGTTGCTTTTGGAGCTTTATCAGTAAAGAACATGGCTGGCAAGAAAACGAGAACCTGTATTTTCAGGGTGCAAGTCATCATCACCATCACCACCATCATTAA
SEQ ID NO:34-rBoNT/F(0)的多肽序列(His标签)
MPVVINSFNYNDPVNDDTILYMQIPYEEKSKKYYKAFEIMRNVWIIPERNTIGTDPSDFDPPASLENGSSAYYDPNYLTTDAEKDRYLKTTIKLFKRINSNPAGEVLLQEISYAKPYLGNEHTPINEFHPVTRTTSVNIKSSTNVKSSIILNLLVLGAGPDIFENSSYPVRKLMDSGGVYDPSNDGFGSINIVTFSPEYEYTFNDISGGYNSSTESFIADPAISLAHQLIYALHGLYGARGVTYKETIKVKQAPLMIAEKPIRLEEFLTFGGQDLNIITSAMKEKIYNNLLANYEKIATRLSRVNSAPPEYDINEYKDYFQWKYGLDKNADGSYTVNENKFNEIYKKLYSFTEIDLANKFKVKCRNTYFIKYGFLKVPNLLDDDIYTVSEGFNIGNLAVNNRGQNIKLNPKIIDSIPDKGLVEKIVKFCKSVIPRKGTKAPPRLCIRVNNRELFFVASESSYNENDINTPKEIDDTTNLNNNYRNNLDEVILDYNSETIPQISNQTLNTLVQDDSYVPRYDSNGTSEIEEHNVVDLNVFFYLHAQKVPEGETNISLTSSIDTALSEESQVYTFFSSEFINTINKPVHAALFISWINQVIRDFTTEATQKSTFDKIADISLVVPYVGLALNIGNEVQKENFKEAFELLGAGILLEFVPELLIPTILVFTIKSFIGSSENKNKIIKAINNSLMERETKWKEIYSWIVSNWLTRINTQFNKRKEQMYQALQNQVDAIKTVIEYKYNNYTSDERNRLESEYNINNIREELNKKVSLAMENIERFITESSIFYLMKLINEAKVSKLREYDEGVKEYLLDYISEHRSILGNSVQELNDLVTSTLNNSIPFELSSYTNDKILILYFNKLYKKIKDNSILDMRYENNKFIDISGYGSNISINGDVYIYSTNRNQFGIYSSKPSEVNIAQNNDIIYNGRYQNFSISFWVRIPKYFNKVNLNNEYTIIDCIRNNNSGWKISLNYNKIIWTLQDTAGNNQKLVFNYTQMISISDYINKWIFVTITNNRLGNSRIYINGNLIDEKSISNLGDIHVSDNILFKIVGCNDTRYVGIRYFKVFDTELGKTEIETLYSDEPDPSILKDFWGNYLLYNKRYYLLNLLRTDKSITQNSNFLNINQQRGVYQKPNIFSNTRLYTGVEVIIRKNGSTDISNTDNFVRKNDLAYINVVDRDVEYRLYADISIAKPEKIIKLIRTSNSNNSLGQIIVMDSIGNNCTMNFQNNNGGNIGLLGFHSNNLVASSWYYNNIRKNTSSNGCFWSFISKEHGWQENENLYFQGASHHHHHHHH
SEQ ID NO:35-rLHN/F的核苷酸序列(His标签)
ATGCCGGTTGTGATTAACAGCTTCAATTATAACGATCCGGTGAACGATGATACCATCCTGTATATGCAGATTCCGTATGAAGAGAAAAGCAAAAAGTACTACAAAGCCTTTGAGATCATGCGCAACGTTTGGATTATTCCGGAACGTAATACCATTGGCACCGATCCGAGCGATTTTGATCCGCCTGCAAGCCTGGAAAATGGTAGCAGCGCATATTATGATCCGAATTATCTGACCACCGATGCCGAAAAAGATCGTTATCTGAAAACCACCATCAAACTGTTCAAACGCATTAATAGCAATCCGGCAGGCGAAGTTCTGCTGCAAGAAATTAGCTATGCAAAACCGTATCTGGGCAATGAACATACCCCGATTAATGAATTTCATCCGGTTACACGTACCACGAGCGTTAACATTAAAAGCAGCACCAATGTGAAGTCCAGCATTATTCTGAATCTGCTGGTTTTAGGTGCAGGTCCGGATATTTTTGAAAATTCAAGCTATCCGGTGCGCAAACTGATGGATAGCGGTGGTGTGTATGATCCGTCAAATGATGGTTTTGGCAGCATTAACATTGTGACCTTTAGTCCGGAATATGAATACACCTTCAACGATATTAGCGGTGGCTATAATAGCAGCACCGAAAGTTTTATTGCAGATCCGGCAATTAGCCTGGCACATGAACTGATTCATGCACTGCATGGTCTGTATGGTGCACGTGGTGTTACCTATAAAGAAACCATTAAAGTTAAACAGGCACCGCTGATGATTGCGGAAAAACCGATTCGTCTGGAAGAATTTCTGACCTTTGGTGGTCAGGATCTGAACATTATTACCAGCGCAATGAAAGAGAAAATCTATAATAACCTGCTGGCCAACTATGAGAAAATTGCAACCCGTCTGAGCCGTGTTAATAGCGCACCTCCTGAATATGATATCAACGAGTATAAAGACTATTTTCAGTGGAAATACGGCCTGGATAAAAATGCAGATGGTAGCTATACCGTGAACGAGAACAAATTTAACGAGATCTACAAAAAACTGTATAGCTTCACCGAAATCGATCTGGCCAACAAATTCAAAGTGAAATGCCGCAACACCTACTTCATCAAATATGGCTTTCTGAAAGTTCCGAACCTGCTTGATGATGATATCTATACCGTTAGCGAAGGCTTTAACATTGGTAATCTGGCCGTTAATAATCGCGGTCAGAACATTAAACTGAACCCGAAAATTATCGATAGCATCCCGGATAAAGGCCTGGTTGAAAAAATTGTGAAATTCTGCAAAAGCGTGATTCCGCGTAAAGGCACCAAAGCACCGCCTCGTCTGTGTATTCGTGTGAATAATCGTGAACTGTTTTTTGTTGCAAGCGAGAGCAGCTATAACGAGAATGATATTAACACCCCGAAAGAGATTGACGATACCACCAATCTGAATAACAACTATCGCAACAATCTGGATGAAGTGATCCTGGATTATAACAGCGAAACCATTCCGCAGATTAGCAATCAGACCCTGAATACCCTGGTTCAGGATGATAGCTATGTTCCGCGTTATGATAGCAATGGCACCAGCGAAATTGAAGAACATAATGTGGTTGATCTGAACGTGTTCTTTTATCTGCATGCACAGAAAGTGCCGGAAGGTGAAACCAATATTAGCCTGACCAGCAGCATTGATACCGCACTGAGCGAAGAAAGCCAGGTTTATACCTTTTTTAGCAGCGAATTCATCAACACCATTAACAAACCGGTTCATGCAGCACTGTTTATTAGCTGGATTAATCAGGTGATTCGCGATTTTACCACCGAAGCAACCCAGAAAAGCACCTTTGATAAAATTGCCGATATTAGTCTGGTGGTGCCGTATGTTGGTCTGGCACTGAATATTGGTAATGAAGTGCAGAAAGAGAACTTTAAAGAAGCCTTCGAACTGTTAGGTGCCGGTATTCTGCTGGAATTTGTGCCGGAACTGCTGATTCCGACCATTCTGGTTTTTACCATTAAGAGCTTTATTGGCAGCAGCGAGAACAAGAACAAAATCATTAAAGCCATCAACAACAGCCTGATGGAACGCGAAACCAAATGGAAAGAAATTTACAGCTGGATTGTGAGCAATTGGCTGACCCGTATCAATACCCAGTTTAACAAACGCAAAGAACAAATGTATCAGGCCCTGCAGAATCAGGTTGATGCAATTAAAACCGTGATCGAATACAAATACAACAACTATACCAGCGACGAACGTAATCGCCTGGAAAGCGAATACAACATTAATAACATTCGCGAAGAACTGAACAAAAAAGTGAGCCTGGCAATGGAAAACATCGAACGTTTTATTACCGAAAGCAGCATCTTCTACCTGATGAAACTGATTAACGAAGCCAAAGTTAGCAAACTGCGCGAATATGATGAAGGCGTTAAAGAATATCTGCTGGACTATATTAGCGAACATCGTAGCATTCTGGGTAATAGCGTTCAAGAGCTGAATGATCTGGTTACCAGCACACTGAATAATAGCATTCCGTTTGAACTGAGCAGCTACACCAACGATAAAATCCTGATCCTGTACTTCAACAAACTGTACAAGAAAGAAAACCTGTATTTTCAGGGTGCAAGCCATCATCACCACCATCACCATCATTAA
SEQ ID NO:36-rLHN/F的多肽序列(His标签)
MPVVINSFNYNDPVNDDTILYMQIPYEEKSKKYYKAFEIMRNVWIIPERNTIGTDPSDFDPPASLENGSSAYYDPNYLTTDAEKDRYLKTTIKLFKRINSNPAGEVLLQEISYAKPYLGNEHTPINEFHPVTRTTSVNIKSSTNVKSSIILNLLVLGAGPDIFENSSYPVRKLMDSGGVYDPSNDGFGSINIVTFSPEYEYTFNDISGGYNSSTESFIADPAISLAHELIHALHGLYGARGVTYKETIKVKQAPLMIAEKPIRLEEFLTFGGQDLNIITSAMKEKIYNNLLANYEKIATRLSRVNSAPPEYDINEYKDYFQWKYGLDKNADGSYTVNENKFNEIYKKLYSFTEIDLANKFKVKCRNTYFIKYGFLKVPNLLDDDIYTVSEGFNIGNLAVNNRGQNIKLNPKIIDSIPDKGLVEKIVKFCKSVIPRKGTKAPPRLCIRVNNRELFFVASESSYNENDINTPKEIDDTTNLNNNYRNNLDEVILDYNSETIPQISNQTLNTLVQDDSYVPRYDSNGTSEIEEHNVVDLNVFFYLHAQKVPEGETNISLTSSIDTALSEESQVYTFFSSEFINTINKPVHAALFISWINQVIRDFTTEATQKSTFDKIADISLVVPYVGLALNIGNEVQKENFKEAFELLGAGILLEFVPELLIPTILVFTIKSFIGSSENKNKIIKAINNSLMERETKWKEIYSWIVSNWLTRINTQFNKRKEQMYQALQNQVDAIKTVIEYKYNNYTSDERNRLESEYNINNIREELNKKVSLAMENIERFITESSIFYLMKLINEAKVSKLREYDEGVKEYLLDYISEHRSILGNSVQELNDLVTSTLNNSIPFELSSYTNDKILILYFNKLYKKENLYFQGASHHHHHHHH
SEQ ID NO:37-rHC/F的核苷酸序列(His标签)
ATGATCAAGGATAACAGCATTCTGGATATGCGCTATGAGAACAACAAATTCATTGATATTAGCGGCTATGGCAGCAACATTAGCATTAATGGTGATGTGTATATCTACAGCACCAACCGTAATCAGTTTGGCATTTATAGCAGCAAACCGAGCGAAGTTAATATTGCCCAGAACAACGATATCATCTATAACGGTCGCTATCAGAACTTCAGCATTAGCTTTTGGGTTCGCATTCCGAAATACTTCAATAAGGTGAACCTGAACAACGAGTATACCATCATTGATTGCATTCGCAATAATAACAGCGGCTGGAAAATTAGCCTGAACTACAACAAAATTATCTGGACCCTGCAGGATACCGCAGGTAATAATCAGAAACTGGTGTTTAACTACACCCAGATGATTAGCATCAGCGACTATATCAACAAATGGATCTTTGTGACCATTACCAATAATCGCCTGGGTAATAGCCGCATTTATATCAATGGTAACCTGATCGATGAGAAAAGCATTAGCAATCTGGGTGATATTCATGTGAGCGATAACATCCTGTTTAAAATCGTGGGTTGTAACGATACCCGTTATGTTGGTATTCGCTACTTCAAAGTGTTTGATACCGAACTGGGTAAAACCGAAATTGAAACCCTGTATAGTGATGAACCGGATCCGAGCATTCTGAAAGATTTTTGGGGTAATTATCTGCTGTACAACAAACGCTACTATCTGCTGAATCTGCTGCGTACCGATAAATCAATTACCCAGAATAGCAACTTCCTGAACATTAATCAGCAGCGTGGTGTTTATCAGAAACCGAACATTTTTAGCAACACCCGTCTGTATACCGGTGTGGAAGTTATTATTCGTAAAAATGGCAGCACCGATATCAGCAACACCGATAACTTTGTTCGCAAAAATGATCTGGCGTATATCAACGTTGTTGATCGTGATGTTGAATATCGTCTGTATGCCGATATTAGCATTGCCAAACCGGAAAAAATCATCAAACTGATCCGTACCAGCAACAGCAATAATTCACTGGGTCAGATTATTGTGATGGATAGCATTGGTAATAACTGCACCATGAACTTTCAGAACAATAACGGTGGTAATATTGGTCTGCTGGGCTTTCATAGTAATAATCTGGTTGCAAGCAGCTGGTATTATAACAACATCCGTAAAAATACCAGCAGCAATGGTTGCTTTTGGAGCTTTATTAGCAAAGAACATGGCTGGCAAGAAAACGAGAATCTGTATTTTCAGGGTGCAAGTCATCATCACCACCATCACCATCATTAA
SEQ ID NO:38-rHC/F的多肽序列(His标签)
MIKDNSILDMRYENNKFIDISGYGSNISINGDVYIYSTNRNQFGIYSSKPSEVNIAQNNDIIYNGRYQNFSISFWVRIPKYFNKVNLNNEYTIIDCIRNNNSGWKISLNYNKIIWTLQDTAGNNQKLVFNYTQMISISDYINKWIFVTITNNRLGNSRIYINGNLIDEKSISNLGDIHVSDNILFKIVGCNDTRYVGIRYFKVFDTELGKTEIETLYSDEPDPSILKDFWGNYLLYNKRYYLLNLLRTDKSITQNSNFLNINQQRGVYQKPNIFSNTRLYTGVEVIIRKNGSTDISNTDNFVRKNDLAYINVVDRDVEYRLYADISIAKPEKIIKLIRTSNSNNSLGQIIVMDSIGNNCTMNFQNNNGGNIGLLGFHSNNLVASSWYYNNIRKNTSSNGCFWSFISKEHGWQENENLYFQGASHHHHHHHH
SEQ ID NO:39-rLC/F的核苷酸序列(His标签)
ATGCCGGTTGTGATTAACAGCTTCAATTATAACGATCCGGTGAACGATGATACCATCCTGTATATGCAGATTCCGTATGAAGAGAAAAGCAAAAAGTACTACAAAGCCTTTGAGATCATGCGCAACGTTTGGATTATTCCGGAACGTAATACCATTGGCACCGATCCGAGCGATTTTGATCCGCCTGCAAGCCTGGAAAATGGTAGCAGCGCATATTATGATCCGAATTATCTGACCACCGATGCCGAAAAAGATCGTTATCTGAAAACCACCATCAAACTGTTCAAACGCATTAATAGCAATCCGGCAGGCGAAGTTCTGCTGCAAGAAATTAGCTATGCAAAACCGTATCTGGGCAATGAACATACCCCGATTAATGAATTTCATCCGGTTACACGTACCACGAGCGTTAACATTAAAAGCAGCACCAATGTGAAGTCCAGCATTATTCTGAATCTGCTGGTTTTAGGTGCAGGTCCGGATATTTTTGAAAATTCAAGCTATCCGGTGCGCAAACTGATGGATAGCGGTGGTGTGTATGATCCGTCAAATGATGGTTTTGGCAGCATTAACATTGTGACCTTTAGTCCGGAATATGAATACACCTTCAACGATATTAGCGGTGGCTATAATAGCAGCACCGAAAGTTTTATTGCAGATCCGGCAATTAGCCTGGCACATGAACTGATTCATGCACTGCATGGTCTGTATGGTGCACGTGGTGTTACCTATAAAGAAACCATTAAAGTTAAACAGGCACCGCTGATGATTGCGGAAAAACCGATTCGTCTGGAAGAATTTCTGACCTTTGGTGGTCAGGATCTGAACATTATTACCAGCGCAATGAAAGAGAAAATCTATAATAACCTGCTGGCCAACTATGAGAAAATTGCAACCCGTCTGAGCCGTGTTAATAGCGCACCTCCTGAATATGATATCAACGAGTATAAAGACTATTTTCAGTGGAAATACGGCCTGGATAAAAATGCAGATGGTAGCTATACCGTGAACGAGAACAAATTTAACGAGATCTACAAAAAACTGTATAGCTTCACCGAAATCGATCTGGCCAACAAATTCAAAGTGAAATGCCGCAACACCTACTTCATCAAATATGGCTTTCTGAAAGTTCCGAACCTGCTTGATGATGATATCTATACCGTTAGCGAAGGCTTTAACATTGGTAATCTGGCCGTTAATAATCGCGGTCAGAACATTAAACTGAACCCGAAAATTATCGATAGCATCCCGGATAAAGGCCTGGTTGAAAAAATTGTGAAATTCTGCAAAAGCGAGAACCTGTATTTTCAGGGTGCAAGTCATCATCACCATCACCACCATCATTAA
SEQ ID NO:40-rLC/F的多肽序列(His标签)
MPVVINSFNYNDPVNDDTILYMQIPYEEKSKKYYKAFEIMRNVWIIPERNTIGTDPSDFDPPASLENGSSAYYDPNYLTTDAEKDRYLKTTIKLFKRINSNPAGEVLLQEISYAKPYLGNEHTPINEFHPVTRTTSVNIKSSTNVKSSIILNLLVLGAGPDIFENSSYPVRKLMDSGGVYDPSNDGFGSINIVTFSPEYEYTFNDISGGYNSSTESFIADPAISLAHELIHALHGLYGARGVTYKETIKVKQAPLMIAEKPIRLEEFLTFGGQDLNIITSAMKEKIYNNLLANYEKIATRLSRVNSAPPEYDINEYKDYFQWKYGLDKNADGSYTVNENKFNEIYKKLYSFTEIDLANKFKVKCRNTYFIKYGFLKVPNLLDDDIYTVSEGFNIGNLAVNNRGQNIKLNPKIIDSIPDKGLVEKIVKFCKSENLYFQGASHHHHHHHH
SEQ ID NO:41-阳离子rHC/A的核苷酸序列(His标签)
ATGATCATCAACACCAGCATTCTGAACCTGCGTTATGAAAGCAAACATCTGATTGATCTGAGCCGTTATGCCAGCAAAATCAATATAGGCAGCAAGGTTAACTTCGACCCGATTGACAAAAATCAGATACAGCTGTTTAATCTGGAAAGCAGCAAAATTGAGGTGATCCTGAAAAAAGCGATCGTGTATAATAGCATGTACGAGAATTTTTCGACCAGCTTTTGGATTCGCATCCCGAAATACTTTAACAAGATTAGCCTGAACAACGAGTATACCATCATTAACTGCATGGAAAACAATAGCGGTTGGAAAGTCAGCCTGAATTATGGCGAAATTATCTGGACCCTGCAGGATACCAAAGAAATCAAACAGCGTGTGGTGTTCAAATACAGCCAGATGATTAATATCAGCGACTATATCAACCGCTGGATTTTTGTGACCATTACCAATAATCGGCTGAACAAGAGCAAGATCTATATTAACGGTCGTCTGATTGACCAGAAACCGATTAGTAATCTGGGTAATATTCATGCGAGCAACAAAATCATGTTTAAACTGGATGGTTGCCGTGATACCCATCGTTATATTTGGATCAAATACTTCAACCTGTTCGATAAAGAGTTGAACGAAAAAGAAATTAAAGACCTGTACGATAACCAGAGCAATAGCGGCATACTGAAAGATTTTTGGGGAGATTATCTGCAGTATGACAAACCGTATTATATGCTGAATCTGTACGACCCGAATAAATACGTGGATGTTAATAATGTGGGCATCCGTGGTTATATGTACCTGAAAGGTCCGCGTGGTAGCGTTATGACCACAAACATTTATCTGAATAGCAGCCTGTATCGCGGAACCAAATTCATCATTAAAAAGTATGCCAGCGGCAACAAGGATAATATTGTGCGTAATAATGATCGCGTGTACATTAACGTTGTGGTGAAGAATAAAGAATATCGCCTGGCAACCAATGCAAGCCAGGCAGGCGTTGAAAAAATTCTGAGTGCCCTGGAAATTCCGGATGTTGGTAATCTGAGCCAGGTTGTTGTGATGAAAAGCAAAAACGATAAAGGCATCACCAACAAATGCAAGATGAATCTGCAGGACAATAACGGCAATGATATTGGCTTCATTGGCTTTCACCAGTTTAACAACATTGCAAAACTGGTTGCGAGCAATTGGTATAATCGTCAGATTGAACGTAGCAGTCGTACCCTGGGTTGTAGCTGGGAATTTATCCCTGTGGATGATGGTTGGGGTGAACGTCCGCTGAAGCTTGCGGCCGCACTCGAGCACCACCACCACCACCACTGA
SEQ ID NO:42-阳离子rHC/A的多肽序列(His标签)
MIINTSILNLRYESKHLIDLSRYASKINIGSKVNFDPIDKNQIQLFNLESSKIEVILKKAIVYNSMYENFSTSFWIRIPKYFNKISLNNEYTIINCMENNSGWKVSLNYGEIIWTLQDTKEIKQRVVFKYSQMINISDYINRWIFVTITNNRLNKSKIYINGRLIDQKPISNLGNIHASNKIMFKLDGCRDTHRYIWIKYFNLFDKELNEKEIKDLYDNQSNSGILKDFWGDYLQYDKPYYMLNLYDPNKYVDVNNVGIRGYMYLKGPRGSVMTTNIYLNSSLYRGTKFIIKKYASGNKDNIVRNNDRVYINVVVKNKEYRLATNASQAGVEKILSALEIPDVGNLSQVVVMKSKNDKGITNKCKMNLQDNNGNDIGFIGFHQFNNIAKLVASNWYNRQIERSSRTLGCSWEFIPVDDGWGERPLKLAAALEHHHHHH
SEQ ID NO:43-rHC/AB的核苷酸序列(His标签)
ATGATTCTGAACAATATTATCCTGAACCTGCGTTACAAAGACAACAATCTGATCGATCTGAGCGGCTATGGTGCAAAAGTTGAAGTCTACGACGGTGTCGAACTGAACGATAAAAACCAGTTCAAACTGACCTCATCGGCTAACTCAAAAATTCGTGTGACGCAGAACCAAAACATCATCTTCAACTCGGTCTTTCTGGACTTCAGCGTGTCTTTCTGGATTCGCATCCCGAAATATAAAAATGATGGCATCCAGAACTACATCCATAACGAATACACCATCATCAACTGTATGAAAAACAACAGTGGTTGGAAAATTTCCATCCGTGGCAACCGCATTATCTGGACCCTGATTGATATCAATGGTAAAACGAAAAGCGTGTTTTTCGAATACAACATCCGTGAAGATATCTCTGAATACATCAATCGCTGGTTTTTCGTGACCATTACGAACAATCTGAACAATGCGAAAATCTATATCAACGGCAAACTGGAAAGTAATACCGACATCAAAGATATTCGTGAAGTTATCGCCAACGGTGAAATCATCTTCAAACTGGATGGCGACATCGATCGCACCCAGTTCATTTGGATGAAATACTTCTCCATCTTCAACACGGAACTGAGTCAGTCCAATATCGAAGAACGCTACAAAATCCAATCATACTCGGAATACCTGAAAGATTTCTGGGGTAACCCGCTGATGTACAACAAAGAATACTACATGTTCAACGCGGGCAACAAAAACTCATACATCAAACTGAAAAAAGATTCGCCGGTGGGTGAAATCCTGACCCGTAGCAAATACAACCAGAACTCTAAATACATCAACTATCGCGATCTGTACATTGGCGAAAAATTTATTATCCGTCGCAAAAGCAACTCTCAGAGTATTAATGATGACATCGTGCGTAAAGAAGACTACATCTATCTGGATTTCTTTAATCTGAACCAAGAATGGCGCGTTTATACCTACAAATACTTCAAAAAAGAAGAAATGAAACTGTTCCTGGCCCCGATTTACGACAGCGATGAATTTTACAACACCATCCAGATCAAAGAATACGATGAACAGCCGACGTATAGTTGCCAACTGCTGTTCAAAAAAGACGAAGAATCCACCGATGAAATTGGCCTGATTGGTATCCACCGTTTCTATGAAAGCGGTATCGTTTTCGAAGAATACAAAGATTACTTCTGTATCTCTAAATGGTATCTGAAAGAAGTCAAACGCAAACCGTACAACCTGAAACTGGGCTGCAACTGGCAATTTATCCCGAAAGACGAAGGCTGGACCGAAAAGCTTGCGGCCGCACTCGAGCACCACCACCACCACCACTGA
SEQ ID NO:44-rHC/AB的多肽序列(His标签)
MILNNIILNLRYKDNNLIDLSGYGAKVEVYDGVELNDKNQFKLTSSANSKIRVTQNQNIIFNSVFLDFSVSFWIRIPKYKNDGIQNYIHNEYTIINCMKNNSGWKISIRGNRIIWTLIDINGKTKSVFFEYNIREDISEYINRWFFVTITNNLNNAKIYINGKLESNTDIKDIREVIANGEIIFKLDGDIDRTQFIWMKYFSIFNTELSQSNIEERYKIQSYSEYLKDFWGNPLMYNKEYYMFNAGNKNSYIKLKKDSPVGEILTRSKYNQNSKYINYRDLYIGEKFIIRRKSNSQSINDDIVRKEDYIYLDFFNLNQEWRVYTYKYFKKEEMKLFLAPIYDSDEFYNTIQIKEYDEQPTYSCQLLFKKDEESTDEIGLIGIHRFYESGIVFEEYKDYFCISKWYLKEVKRKPYNLKLGCNWQFIPKDEGWTEKLAAALEHHHHHH
SEQ ID NO:45-rHC/A变体Y1117V H1253K的核苷酸序列(His标签)
ATGATCATCAATACTAGCATTCTGAACCTGCGTTACGAGAGCAATCATCTGATTGATCTGAGCCGTTATGCAAGCAAGATCAACATCGGTAGCAAGGTCAATTTTGACCCGATCGATAAGAACCAGATCCAGCTGTTTAATCTGGAATCGAGCAAAATTGAGGTTATCCTGAAAAACGCCATTGTCTACAACTCCATGTACGAGAATTTCTCCACCAGCTTCTGGATTCGCATCCCGAAATACTTCAACAGCATTAGCCTGAACAACGAGTATACTATCATCAACTGTATGGAGAACAACAGCGGTTGGAAGGTGTCTCTGAACTATGGTGAGATCATTTGGACCTTGCAGGACACCCAAGAGATCAAGCAGCGCGTCGTGTTCAAGTACTCTCAAATGATCAACATTTCCGATTACATTAATCGTTGGATCTTCGTGACCATTACGAATAACCGTCTGAATAACAGCAAGATTTACATCAATGGTCGCTTGATCGATCAGAAACCGATTAGCAACCTGGGTAATATCCACGCAAGCAACAACATTATGTTCAAATTGGACGGTTGCCGCGATACCCATCGTTATATCTGGATCAAGTATTTCAACCTGTTTGATAAAGAACTGAATGAGAAGGAGATCAAAGATTTGTATGACAACCAATCTAACAGCGGCATTTTGAAGGACTTCTGGGGCGATTATCTGCAATACGATAAGCCGTACTATATGCTGAACCTGgtTGATCCGAACAAATATGTGGATGTCAATAATGTGGGTATTCGTGGTTACATGTATTTGAAGGGTCCGCGTGGCAGCGTTATGACGACCAACATTTACCTGAACTCTAGCCTGTACCGTGGTACGAAATTCATCATTAAGAAATATGCCAGCGGCAACAAAGATAACATTGTGCGTAATAACGATCGTGTCTACATCAACGTGGTCGTGAAGAATAAAGAGTACCGTCTGGCGACCAACGCTTCGCAGGCGGGTGTTGAGAAAATTCTGAGCGCGTTGGAGATCCCTGATGTCGGTAATCTGAGCCAAGTCGTGGTTATGAAGAGCAAGAACGACCAGGGTATCACTAACAAGTGCAAGATGAACCTGCAAGACAACAATGGTAACGACATCGGCTTTATTGGTTTCaAaCAGTTCAACAATATTGCTAAACTGGTAGCGAGCAATTGGTACAATCGTCAGATTGAGCGCAGCAGCCGTACTTTGGGCTGTAGCTGGGAGTTTATCCCGGTCGATGATGGTTGGGGCGAACGTCCGCTGCaCcatcaccatCaCcatcaccatCaCcatT
SEQ ID NO:46-rHC/A变体Y1117V H1253K的多肽序列(His标签)
MIINTSILNLRYESNHLIDLSRYASKINIGSKVNFDPIDKNQIQLFNLESSKIEVILKNAIVYNSMYENFSTSFWIRIPKYFNSISLNNEYTIINCMENNSGWKVSLNYGEIIWTLQDTQEIKQRVVFKYSQMINISDYINRWIFVTITNNRLNNSKIYINGRLIDQKPISNLGNIHASNNIMFKLDGCRDTHRYIWIKYFNLFDKELNEKEIKDLYDNQSNSGILKDFWGDYLQYDKPYYMLNLVDPNKYVDVNNVGIRGYMYLKGPRGSVMTTNIYLNSSLYRGTKFIIKKYASGNKDNIVRNNDRVYINVVVKNKEYRLATNASQAGVEKILSALEIPDVGNLSQVVVMKSKNDQGITNKCKMNLQDNNGNDIGFIGFKQFNNIAKLVASNWYNRQIERSSRTLGCSWEFIPVDDGWGERPLHHHHHHHHHH
SEQ ID NO:47-rHC/A变体Y1117V F1252Y H1253K L1278F的核苷酸序列(His标 签)
ATGATCATCAATACTAGCATTCTGAACCTGCGTTACGAGAGCAATCATCTGATTGATCTGAGCCGTTATGCAAGCAAGATCAACATCGGTAGCAAGGTCAATTTTGACCCGATCGATAAGAACCAGATCCAGCTGTTTAATCTGGAATCGAGCAAAATTGAGGTTATCCTGAAAAACGCCATTGTCTACAACTCCATGTACGAGAATTTCTCCACCAGCTTCTGGATTCGCATCCCGAAATACTTCAACAGCATTAGCCTGAACAACGAGTATACTATCATCAACTGTATGGAGAACAACAGCGGTTGGAAGGTGTCTCTGAACTATGGTGAGATCATTTGGACCTTGCAGGACACCCAAGAGATCAAGCAGCGCGTCGTGTTCAAGTACTCTCAAATGATCAACATTTCCGATTACATTAATCGTTGGATCTTCGTGACCATTACGAATAACCGTCTGAATAACAGCAAGATTTACATCAATGGTCGCTTGATCGATCAGAAACCGATTAGCAACCTGGGTAATATCCACGCAAGCAACAACATTATGTTCAAATTGGACGGTTGCCGCGATACCCATCGTTATATCTGGATCAAGTATTTCAACCTGTTTGATAAAGAACTGAATGAGAAGGAGATCAAAGATTTGTATGACAACCAATCTAACAGCGGCATTTTGAAGGACTTCTGGGGCGATTATCTGCAATACGATAAGCCGTACTATATGCTGAACCTGgtTGATCCGAACAAATATGTGGATGTCAATAATGTGGGTATTCGTGGTTACATGTATTTGAAGGGTCCGCGTGGCAGCGTTATGACGACCAACATTTACCTGAACTCTAGCCTGTACCGTGGTACGAAATTCATCATTAAGAAATATGCCAGCGGCAACAAAGATAACATTGTGCGTAATAACGATCGTGTCTACATCAACGTGGTCGTGAAGAATAAAGAGTACCGTCTGGCGACCAACGCTTCGCAGGCGGGTGTTGAGAAAATTCTGAGCGCGTTGGAGATCCCTGATGTCGGTAATCTGAGCCAAGTCGTGGTTATGAAGAGCAAGAACGACCAGGGTATCACTAACAAGTGCAAGATGAACCTGCAAGACAACAATGGTAACGACATCGGCTTTATTGGTTaCaAaCAGTTCAACAATATTGCTAAACTGGTAGCGAGCAATTGGTACAATCGTCAGATTGAGCGCAGCAGCCGTACTTTtGGCTGTAGCTGGGAGTTTATCCCGGTCGATGATGGTTGGGGCGAACGTCCGCTGCaCcatcaccatCaCcatcaccatCaCcatTAA
SEQ ID NO:48-rHC/A变体Y1117V F1252Y H1253K L1278F的多肽序列(His标签)
MIINTSILNLRYESNHLIDLSRYASKINIGSKVNFDPIDKNQIQLFNLESSKIEVILKNAIVYNSMYENFSTSFWIRIPKYFNSISLNNEYTIINCMENNSGWKVSLNYGEIIWTLQDTQEIKQRVVFKYSQMINISDYINRWIFVTITNNRLNNSKIYINGRLIDQKPISNLGNIHASNNIMFKLDGCRDTHRYIWIKYFNLFDKELNEKEIKDLYDNQSNSGILKDFWGDYLQYDKPYYMLNLVDPNKYVDVNNVGIRGYMYLKGPRGSVMTTNIYLNSSLYRGTKFIIKKYASGNKDNIVRNNDRVYINVVVKNKEYRLATNASQAGVEKILSALEIPDVGNLSQVVVMKSKNDQGITNKCKMNLQDNNGNDIGFIGYKQFNNIAKLVASNWYNRQIERSSRTFGCSWEFIPVDDGWGERPLHHHHHHHHHH
SEQ ID NO:49-rHC/A变体Y1117V F1252Y H1253K L1278H的核苷酸序列(His标 签)
ATGATCATCAATACTAGCATTCTGAACCTGCGTTACGAGAGCAATCATCTGATTGATCTGAGCCGTTATGCAAGCAAGATCAACATCGGTAGCAAGGTCAATTTTGACCCGATCGATAAGAACCAGATCCAGCTGTTTAATCTGGAATCGAGCAAAATTGAGGTTATCCTGAAAAACGCCATTGTCTACAACTCCATGTACGAGAATTTCTCCACCAGCTTCTGGATTCGCATCCCGAAATACTTCAACAGCATTAGCCTGAACAACGAGTATACTATCATCAACTGTATGGAGAACAACAGCGGTTGGAAGGTGTCTCTGAACTATGGTGAGATCATTTGGACCTTGCAGGACACCCAAGAGATCAAGCAGCGCGTCGTGTTCAAGTACTCTCAAATGATCAACATTTCCGATTACATTAATCGTTGGATCTTCGTGACCATTACGAATAACCGTCTGAATAACAGCAAGATTTACATCAATGGTCGCTTGATCGATCAGAAACCGATTAGCAACCTGGGTAATATCCACGCAAGCAACAACATTATGTTCAAATTGGACGGTTGCCGCGATACCCATCGTTATATCTGGATCAAGTATTTCAACCTGTTTGATAAAGAACTGAATGAGAAGGAGATCAAAGATTTGTATGACAACCAATCTAACAGCGGCATTTTGAAGGACTTCTGGGGCGATTATCTGCAATACGATAAGCCGTACTATATGCTGAACCTGgtTGATCCGAACAAATATGTGGATGTCAATAATGTGGGTATTCGTGGTTACATGTATTTGAAGGGTCCGCGTGGCAGCGTTATGACGACCAACATTTACCTGAACTCTAGCCTGTACCGTGGTACGAAATTCATCATTAAGAAATATGCCAGCGGCAACAAAGATAACATTGTGCGTAATAACGATCGTGTCTACATCAACGTGGTCGTGAAGAATAAAGAGTACCGTCTGGCGACCAACGCTTCGCAGGCGGGTGTTGAGAAAATTCTGAGCGCGTTGGAGATCCCTGATGTCGGTAATCTGAGCCAAGTCGTGGTTATGAAGAGCAAGAACGACCAGGGTATCACTAACAAGTGCAAGATGAACCTGCAAGACAACAATGGTAACGACATCGGCTTTATTGGTTaCaAaCAGTTCAACAATATTGCTAAACTGGTAGCGAGCAATTGGTACAATCGTCAGATTGAGCGCAGCAGCCGTACTcatGGCTGTAGCTGGGAGTTTATCCCGGTCGATGATGGTTGGGGCGAACGTCCGCTGCaCcatcaccatCaCcat
SEQ ID NO:50-rHC/A变体Y1117V F1252Y H1253K L1278H的多肽序列(His标签)
MIINTSILNLRYESNHLIDLSRYASKINIGSKVNFDPIDKNQIQLFNLESSKIEVILKNAIVYNSMYENFSTSFWIRIPKYFNSISLNNEYTIINCMENNSGWKVSLNYGEIIWTLQDTQEIKQRVVFKYSQMINISDYINRWIFVTITNNRLNNSKIYINGRLIDQKPISNLGNIHASNNIMFKLDGCRDTHRYIWIKYFNLFDKELNEKEIKDLYDNQSNSGILKDFWGDYLQYDKPYYMLNLVDPNKYVDVNNVGIRGYMYLKGPRGSVMTTNIYLNSSLYRGTKFIIKKYASGNKDNIVRNNDRVYINVVVKNKEYRLATNASQAGVEKILSALEIPDVGNLSQVVVMKSKNDQGITNKCKMNLQDNNGNDIGFIGYKQFNNIAKLVASNWYNRQIERSSRTHGCSWEFIPVDDGWGERPLHHHHHH
SEQ ID NO:51-BoNT/A的多肽序列-UniProt P10845
MPFVNKQFNYKDPVNGVDIAYIKIPNVGQMQPVKAFKIHNKIWVIPERDTFTNPEEGDLNPPPEAKQVPVSYYDSTYLSTDNEKDNYLKGVTKLFERIYSTDLGRMLLTSIVRGIPFWGGSTIDTELKVIDTNCINVIQPDGSYRSEELNLVIIGPSADIIQFECKSFGHEVLNLTRNGYGSTQYIRFSPDFTFGFEESLEVDTNPLLGAGKFATDPAVTLAHELIHAGHRLYGIAINPNRVFKVNTNAYYEMSGLEVSFEELRTFGGHDAKFIDSLQENEFRLYYYNKFKDIASTLNKAKSIVGTTASLQYMKNVFKEKYLLSEDTSGKFSVDKLKFDKLYKMLTEIYTEDNFVKFFKVLNRKTYLNFDKAVFKINIVPKVNYTIYDGFNLRNTNLAANFNGQNTEINNMNFTKLKNFTGLFEFYKLLCVRGIITSKTKSLDKGYNKALNDLCIKVNNWDLFFSPSEDNFTNDLNKGEEITSDTNIEAAEENISLDLIQQYYLTFNFDNEPENISIENLSSDIIGQLELMPNIERFPNGKKYELDKYTMFHYLRAQEFEHGKSRIALTNSVNEALLNPSRVYTFFSSDYVKKVNKATEAAMFLGWVEQLVYDFTDETSEVSTTDKIADITIIIPYIGPALNIGNMLYKDDFVGALIFSGAVILLEFIPEIAIPVLGTFALVSYIANKVLTVQTIDNALSKRNEKWDEVYKYIVTNWLAKVNTQIDLIRKKMKEALENQAEATKAIINYQYNQYTEEEKNNINFNIDDLSSKLNESINKAMININKFLNQCSVSYLMNSMIPYGVKRLEDFDASLKDALLKYIYDNRGTLIGQVDRLKDKVNNTLSTDIPFQLSKYVDNQRLLSTFTEYIKNIINTSILNLRYESNHLIDLSRYASKINIGSKVNFDPIDKNQIQLFNLESSKIEVILKNAIVYNSMYENFSTSFWIRIPKYFNSISLNNEYTIINCMENNSGWKVSLNYGEIIWTLQDTQEIKQRVVFKYSQMINISDYINRWIFVTITNNRLNNSKIYINGRLIDQKPISNLGNIHASNNIMFKLDGCRDTHRYIWIKYFNLFDKELNEKEIKDLYDNQSNSGILKDFWGDYLQYDKPYYMLNLYDPNKYVDVNNVGIRGYMYLKGPRGSVMTTNIYLNSSLYRGTKFIIKKYASGNKDNIVRNNDRVYINVVVKNKEYRLATNASQAGVEKILSALEIPDVGNLSQVVVMKSKNDQGITNKCKMNLQDNNGNDIGFIGFHQFNNIAKLVASNWYNRQIERSSRTLGCSWEFIPVDDGWGERPL
SEQ ID NO:52-BoNT/B的多肽序列-UniProt P10844
MPVTINNFNYNDPIDNNNIIMMEPPFARGTGRYYKAFKITDRIWIIPERYTFGYKPEDFNKSSGIFNRDVCEYYDPDYLNTNDKKNIFLQTMIKLFNRIKSKPLGEKLLEMIINGIPYLGDRRVPLEEFNTNIASVTVNKLISNPGEVERKKGIFANLIIFGPGPVLNENETIDIGIQNHFASREGFGGIMQMKFCPEYVSVFNNVQENKGASIFNRRGYFSDPALILMHELIHVLHGLYGIKVDDLPIVPNEKKFFMQSTDAIQAEELYTFGGQDPSIITPSTDKSIYDKVLQNFRGIVDRLNKVLVCISDPNININIYKNKFKDKYKFVEDSEGKYSIDVESFDKLYKSLMFGFTETNIAENYKIKTRASYFSDSLPPVKIKNLLDNEIYTIEEGFNISDKDMEKEYRGQNKAINKQAYEEISKEHLAVYKIQMCKSVKAPGICIDVDNEDLFFIADKNSFSDDLSKNERIEYNTQSNYIENDFPINELILDTDLISKIELPSENTESLTDFNVDVPVYEKQPAIKKIFTDENTIFQYLYSQTFPLDIRDISLTSSFDDALLFSNKVYSFFSMDYIKTANKVVEAGLFAGWVKQIVNDFVIEANKSNTMDKIADISLIVPYIGLALNVGNETAKGNFENAFEIAGASILLEFIPELLIPVVGAFLLESYIDNKNKIIKTIDNALTKRNEKWSDMYGLIVAQWLSTVNTQFYTIKEGMYKALNYQAQALEEIIKYRYNIYSEKEKSNINIDFNDINSKLNEGINQAIDNINNFINGCSVSYLMKKMIPLAVEKLLDFDNTLKKNLLNYIDENKLYLIGSAEYEKSKVNKYLKTIMPFDLSIYTNDTILIEMFNKYNSEILNNIILNLRYKDNNLIDLSGYGAKVEVYDGVELNDKNQFKLTSSANSKIRVTQNQNIIFNSVFLDFSVSFWIRIPKYKNDGIQNYIHNEYTIINCMKNNSGWKISIRGNRIIWTLIDINGKTKSVFFEYNIREDISEYINRWFFVTITNNLNNAKIYINGKLESNTDIKDIREVIANGEIIFKLDGDIDRTQFIWMKYFSIFNTELSQSNIEERYKIQSYSEYLKDFWGNPLMYNKEYYMFNAGNKNSYIKLKKDSPVGEILTRSKYNQNSKYINYRDLYIGEKFIIRRKSNSQSINDDIVRKEDYIYLDFFNLNQEWRVYTYKYFKKEEEKLFLAPISDSDEFYNTIQIKEYDEQPTYSCQLLFKKDEESTDEIGLIGIHRFYESGIVFEEYKDYFCISKWYLKEVKRKPYNLKLGCNWQFIPKDEGWTE
SEQ ID NO:53-BoNT/C的多肽序列-UniProt P18640
MPITINNFNYSDPVDNKNILYLDTHLNTLANEPEKAFRITGNIWVIPDRFSRNSNPNLNKPPRVTSPKSGYYDPNYLSTDSDKDPFLKEIIKLFKRINSREIGEELIYRLSTDIPFPGNNNTPINTFDFDVDFNSVDVKTRQGNNWVKTGSINPSVIITGPRENIIDPETSTFKLTNNTFAAQEGFGALSIISISPRFMLTYSNATNDVGEGRFSKSEFCMDPILILMHELNHAMHNLYGIAIPNDQTISSVTSNIFYSQYNVKLEYAEIYAFGGPTIDLIPKSARKYFEEKALDYYRSIAKRLNSITTANPSSFNKYIGEYKQKLIRKYRFVVESSGEVTVNRNKFVELYNELTQIFTEFNYAKIYNVQNRKIYLSNVYTPVTANILDDNVYDIQNGFNIPKSNLNVLFMGQNLSRNPALRKVNPENMLYLFTKFCHKAIDGRSLYNKTLDCRELLVKNTDLPFIGDISDVKTDIFLRKDINEETEVIYYPDNVSVDQVILSKNTSEHGQLDLLYPSIDSESEILPGENQVFYDNRTQNVDYLNSYYYLESQKLSDNVEDFTFTRSIEEALDNSAKVYTYFPTLANKVNAGVQGGLFLMWANDVVEDFTTNILRKDTLDKISDVSAIIPYIGPALNISNSVRRGNFTEAFAVTGVTILLEAFPEFTIPALGAFVIYSKVQERNEIIKTIDNCLEQRIKRWKDSYEWMMGTWLSRIITQFNNISYQMYDSLNYQAGAIKAKIDLEYKKYSGSDKENIKSQVENLKNSLDVKISEAMNNINKFIRECSVTYLFKNMLPKVIDELNEFDRNTKAKLINLIDSHNIILVGEVDKLKAKVNNSFQNTIPFNIFSYTNNSLLKDIINEYFNNINDSKILSLQNRKNTLVDTSGYNAEVSEEGDVQLNPIFPFDFKLGSSGEDRGKVIVTQNENIVYNSMYESFSISFWIRINKWVSNLPGYTIIDSVKNNSGWSIGIISNFLVFTLKQNEDSEQSINFSYDISNNAPGYNKWFFVTVTNNMMGNMKIYINGKLIDTIKVKELTGINFSKTITFEINKIPDTGLITSDSDNINMWIRDFYIFAKELDGKDINILFNSLQYTNVVKDYWGNDLRYNKEYYMVNIDYLNRYMYANSRQIVFNTRRNNNDFNEGYKIIIKRIRGNTNDTRVRGGDILYFDMTINNKAYNLFMKNETMYADNHSTEDIYAIGLREQTKDINDNIIFQIQPMNNTYYYASQIFKSNFNGENISGICSIGTYRFRLGGDWYRHNYLVPTVKQGNYASLLESTSTHWGFVPVSE
SEQ ID NO:54-BoNT/D的多肽序列-UniProt P19321
MTWPVKDFNYSDPVNDNDILYLRIPQNKLITTPVKAFMITQNIWVIPERFSSDTNPSLSKPPRPTSKYQSYYDPSYLSTDEQKDTFLKGIIKLFKRINERDIGKKLINYLVVGSPFMGDSSTPEDTFDFTRHTTNIAVEKFENGSWKVTNIITPSVLIFGPLPNILDYTASLTLQGQQSNPSFEGFGTLSILKVAPEFLLTFSDVTSNQSSAVLGKSIFCMDPVIALMHELTHSLHQLYGINIPSDKRIRPQVSEGFFSQDGPNVQFEELYTFGGLDVEIIPQIERSQLREKALGHYKDIAKRLNNINKTIPSSWISNIDKYKKIFSEKYNFDKDNTGNFVVNIDKFNSLYSDLTNVMSEVVYSSQYNVKNRTHYFSRHYLPVFANILDDNIYTIRDGFNLTNKGFNIENSGQNIERNPALQKLSSESVVDLFTKVCLRLTKNSRDDSTCIKVKNNRLPYVADKDSISQEIFENKIITDETNVQNYSDKFSLDESILDGQVPINPEIVDPLLPNVNMEPLNLPGEEIVFYDDITKYVDYLNSYYYLESQKLSNNVENITLTTSVEEALGYSNKIYTFLPSLAEKVNKGVQAGLFLNWANEVVEDFTTNIMKKDTLDKISDVSVIIPYIGPALNIGNSALRGNFNQAFATAGVAFLLEGFPEFTIPALGVFTFYSSIQEREKIIKTIENCLEQRVKRWKDSYQWMVSNWLSRITTQFNHINYQMYDSLSYQADAIKAKIDLEYKKYSGSDKENIKSQVENLKNSLDVKISEAMNNINKFIRECSVTYLFKNMLPKVIDELNKFDLRTKTELINLIDSHNIILVGEVDRLKAKVNESFENTMPFNIFSYTNNSLLKDIINEYFNSINDSKILSLQNKKNALVDTSGYNAEVRVGDNVQLNTIYTNDFKLSSSGDKIIVNLNNNILYSAIYENSSVSFWIKISKDLTNSHNEYTIINSIEQNSGWKLCIRNGNIEWILQDVNRKYKSLIFDYSESLSHTGYTNKWFFVTITNNIMGYMKLYINGELKQSQKIEDLDEVKLDKTIVFGIDENIDENQMLWIRDFNIFSKELSNEDINIVYEGQILRNVIKDYWGNPLKFDTEYYIINDNYIDRYIAPESNVLVLVQYPDRSKLYTGNPITIKSVSDKNPYSRILNGDNIILHMLYNSRKYMIIRDTDTIYATQGGECSQNCVYALKLQSNLGNYGIGIFSIKNIVSKNKYCSQIFSSFRENTMLLADIYKPWRFSFKNAYTPVAVTNYETKLLSTSSFWKFISRDPGWVE
SEQ ID NO:55-BoNT/E的多肽序列-UniProt Q00496
MPKINSFNYNDPVNDRTILYIKPGGCQEFYKSFNIMKNIWIIPERNVIGTTPQDFHPPTSLKNGDSSYYDPNYLQSDEEKDRFLKIVTKIFNRINNNLSGGILLEELSKANPYLGNDNTPDNQFHIGDASAVEIKFSNGSQDILLPNVIIMGAEPDLFETNSSNISLRNNYMPSNHRFGSIAIVTFSPEYSFRFNDNCMNEFIQDPALTLMHELIHSLHGLYGAKGITTKYTITQKQNPLITNIRGTNIEEFLTFGGTDLNIITSAQSNDIYTNLLADYKKIASKLSKVQVSNPLLNPYKDVFEAKYGLDKDASGIYSVNINKFNDIFKKLYSFTEFDLRTKFQVKCRQTYIGQYKYFKLSNLLNDSIYNISEGYNINNLKVNFRGQNANLNPRIITPITGRGLVKKIIRFCKNIVSVKGIRKSICIEINNGELFFVASENSYNDDNINTPKEIDDTVTSNNNYENDLDQVILNFNSESAPGLSDEKLNLTIQNDAYIPKYDSNGTSDIEQHDVNELNVFFYLDAQKVPEGENNVNLTSSIDTALLEQPKIYTFFSSEFINNVNKPVQAALFVSWIQQVLVDFTTEANQKSTVDKIADISIVVPYIGLALNIGNEAQKGNFKDALELLGAGILLEFEPELLIPTILVFTIKSFLGSSDNKNKVIKAINNALKERDEKWKEVYSFIVSNWMTKINTQFNKRKEQMYQALQNQVNAIKTIIESKYNSYTLEEKNELTNKYDIKQIENELNQKVSIAMNNIDRFLTESSISYLMKIINEVKINKLREYDENVKTYLLNYIIQHGSILGESQQELNSMVTDTLNNSIPFKLSSYTDDKILISYFNKFFKRIKSSSVLNMRYKNDKYVDTSGYDSNININGDVYKYPTNKNQFGIYNDKLSEVNISQNDYIIYDNKYKNFSISFWVRIPNYDNKIVNVNNEYTIINCMRDNNSGWKVSLNHNEIIWTFEDNRGINQKLAFNYGNANGISDYINKWIFVTITNDRLGDSKLYINGNLIDQKSILNLGNIHVSDNILFKIVNCSYTRYIGIRYFNIFDKELDETEIQTLYSNEPNTNILKDFWGNYLLYDKEYYLLNVLKPNNFIDRRKDSTLSINNIRSTILLANRLYSGIKVKIQRVNNSSTNDNLVRKNDQVYINFVASKTHLFPLYADTATTNKEKTIKISSSGNRFNQVVVMNSVGNCTMNFKNNNGNNIGLLGFKADTVVASTWYYTHMRDHTNSNGCFWNFISEEHGWQEK
SEQ ID NO:56-BoNT/F的多肽序列-UniProt A7GBG3
MPVVINSFNYNDPVNDDTILYMQIPYEEKSKKYYKAFEIMRNVWIIPERNTIGTDPSDFDPPASLENGSSAYYDPNYLTTDAEKDRYLKTTIKLFKRINSNPAGEVLLQEISYAKPYLGNEHTPINEFHPVTRTTSVNIKSSTNVKSSIILNLLVLGAGPDIFENSSYPVRKLMDSGGVYDPSNDGFGSINIVTFSPEYEYTFNDISGGYNSSTESFIADPAISLAHELIHALHGLYGARGVTYKETIKVKQAPLMIAEKPIRLEEFLTFGGQDLNIITSAMKEKIYNNLLANYEKIATRLSRVNSAPPEYDINEYKDYFQWKYGLDKNADGSYTVNENKFNEIYKKLYSFTEIDLANKFKVKCRNTYFIKYGFLKVPNLLDDDIYTVSEGFNIGNLAVNNRGQNIKLNPKIIDSIPDKGLVEKIVKFCKSVIPRKGTKAPPRLCIRVNNRELFFVASESSYNENDINTPKEIDDTTNLNNNYRNNLDEVILDYNSETIPQISNQTLNTLVQDDSYVPRYDSNGTSEIEEHNVVDLNVFFYLHAQKVPEGETNISLTSSIDTALSEESQVYTFFSSEFINTINKPVHAALFISWINQVIRDFTTEATQKSTFDKIADISLVVPYVGLALNIGNEVQKENFKEAFELLGAGILLEFVPELLIPTILVFTIKSFIGSSENKNKIIKAINNSLMERETKWKEIYSWIVSNWLTRINTQFNKRKEQMYQALQNQVDAIKTVIEYKYNNYTSDERNRLESEYNINNIREELNKKVSLAMENIERFITESSIFYLMKLINEAKVSKLREYDEGVKEYLLDYISEHRSILGNSVQELNDLVTSTLNNSIPFELSSYTNDKILILYFNKLYKKIKDNSILDMRYENNKFIDISGYGSNISINGDVYIYSTNRNQFGIYSSKPSEVNIAQNNDIIYNGRYQNFSISFWVRIPKYFNKVNLNNEYTIIDCIRNNNSGWKISLNYNKIIWTLQDTAGNNQKLVFNYTQMISISDYINKWIFVTITNNRLGNSRIYINGNLIDEKSISNLGDIHVSDNILFKIVGCNDTRYVGIRYFKVFDTELGKTEIETLYSDEPDPSILKDFWGNYLLYNKRYYLLNLLRTDKSITQNSNFLNINQQRGVYQKPNIFSNTRLYTGVEVIIRKNGSTDISNTDNFVRKNDLAYINVVDRDVEYRLYADISIAKPEKIIKLIRTSNSNNSLGQIIVMDSIGNNCTMNFQNNNGGNIGLLGFHSNNLVASSWYYNNIRKNTSSNGCFWSFISKEHGWQEN
SEQ ID NO:57-BoNT/G的多肽序列-UniProt Q60393
MPVNIKXFNYNDPINNDDIIMMEPFNDPGPGTYYKAFRIIDRIWIVPERFTYGFQPDQFNASTGVFSKDVYEYYDPTYLKTDAEKDKFLKTMIKLFNRINSKPSGQRLLDMIVDAIPYLGNASTPPDKFAANVANVSINKKIIQPGAEDQIKGLMTNLIIFGPGPVLSDNFTDSMIMNGHSPISEGFGARMMIRFCPSCLNVFNNVQENKDTSIFSRRAYFADPALTLMHELIHVLHGLYGIKISNLPITPNTKEFFMQHSDPVQAEELYTFGGHDPSVISPSTDMNIYNKALQNFQDIANRLNIVSSAQGSGIDISLYKQIYKNKYDFVEDPNGKYSVDKDKFDKLYKALMFGFTETNLAGEYGIKTRYSYFSEYLPPIKTEKLLDNTIYTQNEGFNIASKNLKTEFNGQNKAVNKEAYEEISLEHLVIYRIAMCKPVMYKNTGKSEQCIIVNNEDLFFIANKDSFSKDLAKAETIAYNTQNNTIENNFSIDQLILDNDLSSGIDLPNENTEPFTNFDDIDIPVYIKQSALKKIFVDGDSLFEYLHAQTFPSNIENLQLTNSLNDALRNNNKVYTFFSTNLVEKANTVVGASLFVNWVKGVIDDFTSESTQKSTIDKVSDVSIIIPYIGPALNVGNETAKENFKNAFEIGGAAILMEFIPELIVPIVGFFTLESYVGNKGHIIMTISNALKKRDQKWTDMYGLIVSQWLSTVNTQFYTIKERMYNALNNQSQAIEKIIEDQYNRYSEEDKMNINIDFNDIDFKLNQSINLAINNIDDFINQCSISYLMNRMIPLAVKKLKDFDDNLKRDLLEYIDTNELYLLDEVNILKSKVNRHLKDSIPFDLSLYTKDTILIQVFNNYISNISSNAILSLSYRGGRLIDSSGYGATMNVGSDVIFNDIGNGQFKLNNSENSNITAHQSKFVVYDSMFDNFSINFWVRTPKYNNNDIQTYLQNEYTIISCIKNDSGWKVSIKGNRIIWTLIDVNAKSKSIFFEYSIKDNISDYINKWFSITITNDRLGNANIYINGSLKKSEKILNLDRINSSNDIDFKLINCTDTTKFVWIKDFNIFGRELNATEVSSLYWIQSSTNTLKDFWGNPLRYDTQYYLFNQGMQNIYIKYFSKASMGETAPRTNFNNAAINYQNLYLGLRFIIKKASNSRNINNDNIVREGDYIYLNIDNISDESYRVYVLVNSKEIQTQLFLAPINDDPTFYDVLQIKKYYEKTTYNCQILCEKDTKTFGLFGIGKFVKDYGYVWDTYDNYFCISQWYLRRISENINKLRLGCNWQFIPVDEGWTE
SEQ ID NO:58-TeNT的多肽序列-UniProt P04958
MPITINNFRYSDPVNNDTIIMMEPPYCKGLDIYYKAFKITDRIWIVPERYEFGTKPEDFNPPSSLIEGASEYYDPNYLRTDSDKDRFLQTMVKLFNRIKNNVAGEALLDKIINAIPYLGNSYSLLDKFDTNSNSVSFNLLEQDPSGATTKSAMLTNLIIFGPGPVLNKNEVRGIVLRVDNKNYFPCRDGFGSIMQMAFCPEYVPTFDNVIENITSLTIGKSKYFQDPALLLMHELIHVLHGLYGMQVSSHEIIPSKQEIYMQHTYPISAEELFTFGGQDANLISIDIKNDLYEKTLNDYKAIANKLSQVTSCNDPNIDIDSYKQIYQQKYQFDKDSNGQYIVNEDKFQILYNSIMYGFTEIELGKKFNIKTRLSYFSMNHDPVKIPNLLDDTIYNDTEGFNIESKDLKSEYKGQNMRVNTNAFRNVDGSGLVSKLIGLCKKIIPPTNIRENLYNRTASLTDLGGELCIKIKNEDLTFIAEKNSFSEEPFQDEIVSYNTKNKPLNFNYSLDKIIVDYNLQSKITLPNDRTTPVTKGIPYAPEYKSNAASTIEIHNIDDNTIYQYLYAQKSPTTLQRITMTNSVDDALINSTKIYSYFPSVISKVNQGAQGILFLQWVRDIIDDFTNESSQKTTIDKISDVSTIVPYIGPALNIVKQGYEGNFIGALETTGVVLLLEYIPEITLPVIAALSIAESSTQKEKIIKTIDNFLEKRYEKWIEVYKLVKAKWLGTVNTQFQKRSYQMYRSLEYQVDAIKKIIDYEYKIYSGPDKEQIADEINNLKNKLEEKANKAMININIFMRESSRSFLVNQMINEAKKQLLEFDTQSKNILMQYIKANSKFIGITELKKLESKINKVFSTPIPFSYSKNLDCWVDNEEDIDVILKKSTILNLDINNDIISDISGFNSSVITYPDAQLVPGINGKAIHLVNNESSEVIVHKAMDIEYNDMFNNFTVSFWLRVPKVSASHLEQYGTNEYSIISSMKKHSLSIGSGWSVSLKGNNLIWTLKDSAGEVRQITFRDLPDKFNAYLANKWVFITITNDRLSSANLYINGVLMGSAEITGLGAIREDNNITLKLDRCNNNNQYVSIDKFRIFCKALNPKEIEKLYTSYLSITFLRDFWGNPLRYDTEYYLIPVASSSKDVQLKNITDYMYLTNAPSYTNGKLNIYYRRLYNGLKFIIKRYTPNNEIDSFVKSGDFIKLYVSYNNNEHIVGYPKDGNAFNNLDRILRVGYNAPGIPLYKKMEAVKLRDLKTYSVQLKLYDDKNASLGLVGTHNGQIGNDPNRDILIASNWYFNHLKDKILGCDWYFVPTDEGWTND
SEQ ID NO:59-BoNT/X的多肽序列
MKLEINKFNYNDPIDGINVITMRPPRHSDKINKGKGPFKAFQVIKNIWIVPERYNFTNNTNDLNIPSEPIMEADAIYNPNYLNTPSEKDEFLQGVIKVLERIKSKPEGEKLLELISSSIPLPLVSNGALTLSDNETIAYQENNNIVSNLQANLVIYGPGPDIANNATYGLYSTPISNGEGTLSEVSFSPFYLKPFDESYGNYRSLVNIVNKFVKREFAPDPASTLMHELVHVTHNLYGISNRNFYYNFDTGKIETSRQQNSLIFEELLTFGGIDSKAISSLIIKKIIETAKNNYTTLISERLNTVTVENDLLKYIKNKIPVQGRLGNFKLDTAEFEKKLNTILFVLNESNLAQRFSILVRKHYLKERPIDPIYVNILDDNSYSTLEGFNISSQGSNDFQGQLLESSYFEKIESNALRAFIKICPRNGLLYNAIYRNSKNYLNNIDLEDKKTTSKTNVSYPCSLLNGCIEVENKDLFLISNKDSLNDINLSEEKIKPETTVFFKDKLPPQDITLSNYDFTEANSIPSISQQNILERNEELYEPIRNSLFEIKTIYVDKLTTFHFLEAQNIDESIDSSKIRVELTDSVDEALSNPNKVYSPFKNMSNTINSIETGITSTYIFYQWLRSIVKDFSDETGKIDVIDKSSDTLAIVPYIGPLLNIGNDIRHGDFVGAIELAGITALLEYVPEFTIPILVGLEVIGGELAREQVEAIVNNALDKRDQKWAEVYNITKAQWWGTIHLQINTRLAHTYKALSRQANAIKMNMEFQLANYKGNIDDKAKIKNAISETEILLNKSVEQAMKNTEKFMIKLSNSYLTKEMIPKVQDNLKNFDLETKKTLDKFIKEKEDILGTNLSSSLRRKVSIRLNKNIAFDINDIPFSEFDDLINQYKNEIEDYEVLNLGAEDGKIKDLSGTTSDINIGSDIELADGRENKAIKIKGSENSTIKIAMNKYLRFSATDNFSISFWIKHPKPTNLLNNGIEYTLVENFNQRGWKISIQDSKLIWYLRDHNNSIKIVTPDYIAFNGWNLITITNNRSKGSIVYVNGSKIEEKDISSIWNTEVDDPIIFRLKNNRDTQAFTLLDQFSIYRKELNQNEVVKLYNYYFNSNYIRDIWGNPLQYNKKYYLQTQDKPGKGLIREYWSSFGYDYVILSDSKTITFPNNIRYGALYNGSKVLIKNSKKLDGLVRNKDFIQLEIDGYNMGISADRFNEDTNYIGTTYGTTHDLTTDFEIIQRQEKYRNYCQLKTPYNIFHKSGLMSTETSKPTFHDYRDWVYSSAWYFQNYENLNLRKHTKTNWYFIPKDEGWDED
SEQ ID NO:60-未修饰的BoNT/A1的多肽序列
MPFVNKQFNYKDPVNGVDIAYIKIPNAGQMQPVKAFKIHNKIWVIPERDTFTNPEEGDLNPPPEAKQVPVSYYDSTYLSTDNEKDNYLKGVTKLFERIYSTDLGRMLLTSIVRGIPFWGGSTIDTELKVIDTNCINVIQPDGSYRSEELNLVIIGPSADIIQFECKSFGHEVLNLTRNGYGSTQYIRFSPDFTFGFEESLEVDTNPLLGAGKFATDPAVTLAHELIHAGHRLYGIAINPNRVFKVNTNAYYEMSGLEVSFEELRTFGGHDAKFIDSLQENEFRLYYYNKFKDIASTLNKAKSIVGTTASLQYMKNVFKEKYLLSEDTSGKFSVDKLKFDKLYKMLTEIYTEDNFVKFFKVLNRKTYLNFDKAVFKINIVPKVNYTIYDGFNLRNTNLAANFNGQNTEINNMNFTKLKNFTGLFEFYKLLCVRGIITSKTKSLDKGYNKALNDLCIKVNNWDLFFSPSEDNFTNDLNKGEEITSDTNIEAAEENISLDLIQQYYLTFNFDNEPENISIENLSSDIIGQLELMPNIERFPNGKKYELDKYTMFHYLRAQEFEHGKSRIALTNSVNEALLNPSRVYTFFSSDYVKKVNKATEAAMFLGWVEQLVYDFTDETSEVSTTDKIADITIIIPYIGPALNIGNMLYKDDFVGALIFSGAVILLEFIPEIAIPVLGTFALVSYIANKVLTVQTIDNALSKRNEKWDEVYKYIVTNWLAKVNTQIDLIRKKMKEALENQAEATKAIINYQYNQYTEEEKNNINFNIDDLSSKLNESINKAMININKFLNQCSVSYLMNSMIPYGVKRLEDFDASLKDALLKYIYDNRGTLIGQVDRLKDKVNNTLSTDIPFQLSKYVDNQRLLSTFTEYIKNIINTSILNLRYESNHLIDLSRYASKINIGSKVNFDPIDKNQIQLFNLESSKIEVILKNAIVYNSMYENFSTSFWIRIPKYFNSISLNNEYTIINCMENNSGWKVSLNYGEIIWTLQDTQEIKQRVVFKYSQMINISDYINRWIFVTITNNRLNNSKIYINGRLIDQKPISNLGNIHASNNIMFKLDGCRDTHRYIWIKYFNLFDKELNEKEIKDLYDNQSNSGILKDFWGDYLQYDKPYYMLNLYDPNKYVDVNNVGIRGYMYLKGPRGSVMTTNIYLNSSLYRGTKFIIKKYASGNKDNIVRNNDRVYINVVVKNKEYRLATNASQAGVEKILSALEIPDVGNLSQVVVMKSKNDQGITNKCKMNLQDNNGNDIGFIGFHQFNNIAKLVASNWYNRQIERSSRTLGCSWEFIPVDDGWGERPL
SEQ ID NO:61-mrBoNT/AB(0)的多肽序列
MPFVNKQFNYKDPVNGVDIAYIKIPNAGQMQPVKAFKIHNKIWVIPERDTFTNPEEGDLNPPPEAKQVPVSYYDSTYLSTDNEKDNYLKGVTKLFERIYSTDLGRMLLTSIVRGIPFWGGSTIDTELKVIDTNCINVIQPDGSYRSEELNLVIIGPSADIIQFECKSFGHEVLNLTRNGYGSTQYIRFSPDFTFGFEESLEVDTNPLLGAGKFATDPAVTLAHQLIYAGHRLYGIAINPNRVFKVNTNAYYEMSGLEVSFEELRTFGGHDAKFIDSLQENEFRLYYYNKFKDIASTLNKAKSIVGTTASLQYMKNVFKEKYLLSEDTSGKFSVDKLKFDKLYKMLTEIYTEDNFVKFFKVLNRKTYLNFDKAVFKINIVPKVNYTIYDGFNLRNTNLAANFNGQNTEINNMNFTKLKNFTGLFEFYKLLCVRGIITSKTKSLDKGYNKALNDLCIKVNNWDLFFSPSEDNFTNDLNKGEEITSDTNIEAAEENISLDLIQQYYLTFNFDNEPENISIENLSSDIIGQLELMPNIERFPNGKKYELDKYTMFHYLRAQEFEHGKSRIALTNSVNEALLNPSRVYTFFSSDYVKKVNKATEAAMFLGWVEQLVYDFTDETSEVSTTDKIADITIIIPYIGPALNIGNMLYKDDFVGALIFSGAVILLEFIPEIAIPVLGTFALVSYIANKVLTVQTIDNALSKRNEKWDEVYKYIVTNWLAKVNTQIDLIRKKMKEALENQAEATKAIINYQYNQYTEEEKNNINFNIDDLSSKLNESINKAMININKFLNQCSVSYLMNSMIPYGVKRLEDFDASLKDALLKYIYDNRGTLIGQVDRLKDKVNNTLSTDIPFQLSKYVDNQRLLSTFTEYIKNILNNIILNLRYKDNNLIDLSGYGAKVEVYDGVELNDKNQFKLTSSANSKIRVTQNQNIIFNSVFLDFSVSFWIRIPKYKNDGIQNYIHNEYTIINCMKNNSGWKISIRGNRIIWTLIDINGKTKSVFFEYNIREDISEYINRWFFVTITNNLNNAKIYINGKLESNTDIKDIREVIANGEIIFKLDGDIDRTQFIWMKYFSIFNTELSQSNIEERYKIQSYSEYLKDFWGNPLMYNKEYYMFNAGNKNSYIKLKKDSPVGEILTRSKYNQNSKYINYRDLYIGEKFIIRRKSNSQSINDDIVRKEDYIYLDFFNLNQEWRVYTYKYFKKEEMKLFLAPIYDSDEFYNTIQIKEYDEQPTYSCQLLFKKDEESTDEIGLIGIHRFYESGIVFEEYKDYFCISKWYLKEVKRKPYNLKLGCNWQFIPKDEGWTE
SEQ ID NO:62-mrBoNT/A(0)的多肽序列
MPFVNKQFNYKDPVNGVDIAYIKIPNAGQMQPVKAFKIHNKIWVIPERDTFTNPEEGDLNPPPEAKQVPVSYYDSTYLSTDNEKDNYLKGVTKLFERIYSTDLGRMLLTSIVRGIPFWGGSTIDTELKVIDTNCINVIQPDGSYRSEELNLVIIGPSADIIQFECKSFGHEVLNLTRNGYGSTQYIRFSPDFTFGFEESLEVDTNPLLGAGKFATDPAVTLAHQLIYAGHRLYGIAINPNRVFKVNTNAYYEMSGLEVSFEELRTFGGHDAKFIDSLQENEFRLYYYNKFKDIASTLNKAKSIVGTTASLQYMKNVFKEKYLLSEDTSGKFSVDKLKFDKLYKMLTEIYTEDNFVKFFKVLNRKTYLNFDKAVFKINIVPKVNYTIYDGFNLRNTNLAANFNGQNTEINNMNFTKLKNFTGLFEFYKLLCVRGIITSKTKSLDKGYNKALNDLCIKVNNWDLFFSPSEDNFTNDLNKGEEITSDTNIEAAEENISLDLIQQYYLTFNFDNEPENISIENLSSDIIGQLELMPNIERFPNGKKYELDKYTMFHYLRAQEFEHGKSRIALTNSVNEALLNPSRVYTFFSSDYVKKVNKATEAAMFLGWVEQLVYDFTDETSEVSTTDKIADITIIIPYIGPALNIGNMLYKDDFVGALIFSGAVILLEFIPEIAIPVLGTFALVSYIANKVLTVQTIDNALSKRNEKWDEVYKYIVTNWLAKVNTQIDLIRKKMKEALENQAEATKAIINYQYNQYTEEEKNNINFNIDDLSSKLNESINKAMININKFLNQCSVSYLMNSMIPYGVKRLEDFDASLKDALLKYIYDNRGTLIGQVDRLKDKVNNTLSTDIPFQLSKYVDNQRLLSTFTEYIKNIINTSILNLRYESKHLIDLSRYASKINIGSKVNFDPIDKNQIQLFNLESSKIEVILKKAIVYNSMYENFSTSFWIRIPKYFNKISLNNEYTIINCMENNSGWKVSLNYGEIIWTLQDTKEIKQRVVFKYSQMINISDYINRWIFVTITNNRLNKSKIYINGRLIDQKPISNLGNIHASNKIMFKLDGCRDTHRYIWIKYFNLFDKELNEKEIKDLYDNQSNSGILKDFWGDYLQYDKPYYMLNLYDPNKYVDVNNVGIRGYMYLKGPRGSVMTTNIYLNSSLYRGTKFIIKKYASGNKDNIVRNNDRVYINVVVKNKEYRLATNASQAGVEKILSALEIPDVGNLSQVVVMKSKNDKGITNKCKMNLQDNNGNDIGFIGFHQFNNIAKLVASNWYNRQIERSSRTLGCSWEFIPVDDGWGERPL
SEQ ID NO:63-BoNT/XB(0)的多肽序列(His标签)
MGSMKLEINKFNYNDPIDGINVITMRPPRH SDKINKGKGPFKAFQVIKNIWIVPERYNFTNNTNDLNIPSEPIMEADAIYNPNYLNTPSE KDEFLQGVIKVLERIKSKPEGEKLLELISSSIPLPLVSNGALTLSDNETIAYQENNNIVS NLQANLVIYGPGPDIANNATYGLYSTPISNGEGTLSEVSFSPFYLKPFDESYGNYRSLVN IVNKFVKREFAPDPASTLMHQLVYVTHNLYGISNRNFYYNFDTGKIETSRQQNSLIFEEL LTFGGIDSKAISSLIIKKIIETAKNNYTTLISERLNTVTVENDLLKYIKNKIPVQGRLGN FKLDTAEFEKKLNTILFVLNESNLAQRFSILVRKHYLKERPIDPIYVNILDDNSYSTLEG FNISSQGSNDFQGQLLESSYFEKIESNALRAFIKICHKAIDGRSLYNKTLDCIEVENKDL FLISNKDSLNDINLSEEKIKPETTVFFKDKLPPQDITLSNYDFTEANSIPSISQQNILER NEELYEPIRNSLFEIKTIYVDKLTTFHFLEAQNIDESIDSSKIRVELTDSVDEALSNPNK VYSPFKNMSNTINSIETGITSTYIFYQWLRSIVKDFSDETGKIDVIDKSSDTLAIVPYIG PLLNIGNDIRHGDFVGAIELAGITALLEYVPEFTIPILVGLEVIGGELAREQVEAIVNNA LDKRDQKWAEVYNITKAQWWGTIHLQINTRLAHTYKALSRQANAIKMNMEFQLANYKGNIDDKAKIKNAISETEILLNKSVEQAMKNTEKFMIKLSNSYLTKEMIPKVQDNLKNFDLETK KTLDKFIKEKEDILGTNLSSSLRRKVSIRLNKNIAFDINDIPFSEFDDLINQYKNEIEDY EVLNLGAEDGKIKDLSGTTSDINIGSDIEILNNIILNLRYKDNNLIDLSGYGAKVEVYDG VELNDKNQFKLTSSANSKIRVTQNQNIIFNSVFLDFSVSFWIRIPKYKNDGIQNYIHNEY TIINCMKNNSGWKISIRGNRIIWTLIDINGKTKSVFFEYNIREDISEYINRWFFVTITNN LNNAKIYINGKLESNTDIKDIREVIANGEIIFKLDGDIDRTQFIWMKYFSIFNTELSQSN IEERYKIQSYSEYLKDFWGNPLMYNKEYYMFNAGNKNSYIKLKKDSPVGEILTRSKYNQN SKYINYRDLYIGEKFIIRRKSNSQSINDDIVRKEDYIYLDFFNLNQEWRVYTYKYFKKEE MKLFLAPIYDSDEFYNTIQIKEYDEQPTYSCQLLFKKDEESTDEIGLIGIHRFYESGIVF EEYKDYFCISKWYLKEVKRKPYNLKLGCNWQFIPKDEGWTEHHHHHHHHHH
SEQ ID NO:64-BoNT/XB(0)的多肽序列
MGSMKLEINKFNYNDPIDGINVITMRPPRH SDKINKGKGPFKAFQVIKNIWIVPERYNFTNNTNDLNIPSEPIMEADAIYNPNYLNTPSE KDEFLQGVIKVLERIKSKPEGEKLLELISSSIPLPLVSNGALTLSDNETIAYQENNNIVS NLQANLVIYGPGPDIANNATYGLYSTPISNGEGTLSEVSFSPFYLKPFDESYGNYRSLVN IVNKFVKREFAPDPASTLMHQLVYVTHNLYGISNRNFYYNFDTGKIETSRQQNSLIFEEL LTFGGIDSKAISSLIIKKIIETAKNNYTTLISERLNTVTVENDLLKYIKNKIPVQGRLGN FKLDTAEFEKKLNTILFVLNESNLAQRFSILVRKHYLKERPIDPIYVNILDDNSYSTLEG FNISSQGSNDFQGQLLESSYFEKIESNALRAFIKICHKAIDGRSLYNKTLDCIEVENKDL FLISNKDSLNDINLSEEKIKPETTVFFKDKLPPQDITLSNYDFTEANSIPSISQQNILER NEELYEPIRNSLFEIKTIYVDKLTTFHFLEAQNIDESIDSSKIRVELTDSVDEALSNPNK VYSPFKNMSNTINSIETGITSTYIFYQWLRSIVKDFSDETGKIDVIDKSSDTLAIVPYIG PLLNIGNDIRHGDFVGAIELAGITALLEYVPEFTIPILVGLEVIGGELAREQVEAIVNNA LDKRDQKWAEVYNITKAQWWGTIHLQINTRLAHTYKALSRQANAIKMNMEFQLANYKGNIDDKAKIKNAISETEILLNKSVEQAMKNTEKFMIKLSNSYLTKEMIPKVQDNLKNFDLETK KTLDKFIKEKEDILGTNLSSSLRRKVSIRLNKNIAFDINDIPFSEFDDLINQYKNEIEDY EVLNLGAEDGKIKDLSGTTSDINIGSDIEILNNIILNLRYKDNNLIDLSGYGAKVEVYDG VELNDKNQFKLTSSANSKIRVTQNQNIIFNSVFLDFSVSFWIRIPKYKNDGIQNYIHNEY TIINCMKNNSGWKISIRGNRIIWTLIDINGKTKSVFFEYNIREDISEYINRWFFVTITNN LNNAKIYINGKLESNTDIKDIREVIANGEIIFKLDGDIDRTQFIWMKYFSIFNTELSQSN IEERYKIQSYSEYLKDFWGNPLMYNKEYYMFNAGNKNSYIKLKKDSPVGEILTRSKYNQN SKYINYRDLYIGEKFIIRRKSNSQSINDDIVRKEDYIYLDFFNLNQEWRVYTYKYFKKEE MKLFLAPIYDSDEFYNTIQIKEYDEQPTYSCQLLFKKDEESTDEIGLIGIHRFYESGIVF EEYKDYFCISKWYLKEVKRKPYNLKLGCNWQFIPKDEGWTE
SEQ ID NO:65-BoNT/XB(0)变体的多肽序列(His标签)
MGSMKLEINKFNYNDPIDGINVITMRPPRH SDKINKGKGPFKAFQVIKNIWIVPERYNFTNNTNDLNIPSEPIMEADAIYNPNYLNTPSE KDEFLQGVIKVLERIKSKPEGEKLLELISSSIPLPLVSNGALTLSDNETIAYQENNNIVS NLQANLVIYGPGPDIANNATYGLYSTPISNGEGTLSEVSFSPFYLKPFDESYGNYRSLVN IVNKFVKREFAPDPASTLMHQLVYVTHNLYGISNRNFYYNFDTGKIETSRQQNSLIFEEL LTFGGIDSKAISSLIIKKIIETAKNNYTTLISERLNTVTVENDLLKYIKNKIPVQGRLGN FKLDTAEFEKKLNTILFVLNESNLAQRFSILVRKHYLKERPIDPIYVNILDDNSYSTLEG FNISSQGSNDFQGQLLESSYFEKIESNALRAFIKICHKAIDGRSLYNKTLDCIEVENKDL FLISNKDSLNDINLSEEKIKPETTVFFKDKLPPQDITLSNYDFTEANSIPSISQQNILER NEELYEPIRNSLFEIKTIYVDKLTTFHFLEAQNIDESIDSSKIRVELTDSVDEALSNPNK VYSPFKNMSNTINSIETGITSTYIFYQWLRSIVKDFSDETGKIDVIDKSSDTLAIVPYIG PLLNIGNDIRHGDFVGAIELAGITALLEYVPEFTIPILVGLEVIGGELAREQVEAIVNNA LDKRDQKWAEVYNITKAQWWGTIHLQINTRLAHTYKALSRQANAIKMNMEFQLANYKGNI DDKAKIKNAISETEILLNKSVEQAMKNTEKFMIKLSNSYLTKEMIPKVQDNLKNFDLETK KTLDKFIKEKEDILGTNLSSSLRRKVSIRLNKNIAFDINDIPFSEFDDLINQYKNEILNN IILNLRYKDNNLIDLSGYGAKVEVYDGVELNDKNQFKLTSSANSKIRVTQNQNIIFNSVF LDFSVSFWIRIPKYKNDGIQNYIHNEYTIINCMKNNSGWKISIRGNRIIWTLIDINGKTK SVFFEYNIREDISEYINRWFFVTITNNLNNAKIYINGKLESNTDIKDIREVIANGEIIFK LDGDIDRTQFIWMKYFSIFNTELSQSNIEERYKIQSYSEYLKDFWGNPLMYNKEYYMFNA GNKNSYIKLKKDSPVGEILTRSKYNQNSKYINYRDLYIGEKFIIRRKSNSQSINDDIVRK EDYIYLDFFNLNQEWRVYTYKYFKKEEMKLFLAPIYDSDEFYNTIQIKEYDEQPTYSCQL LFKKDEESTDEIGLIGIHRFYESGIVFEEYKDYFCISKWYLKEVKRKPYNLKLGCNWQFI PKDEGWTEHHHHHHHHHH
SEQ ID NO:66-BoNT/XB(0)变体的多肽序列
MGSMKLEINKFNYNDPIDGINVITMRPPRH SDKINKGKGPFKAFQVIKNIWIVPERYNFTNNTNDLNIPSEPIMEADAIYNPNYLNTPSE KDEFLQGVIKVLERIKSKPEGEKLLELISSSIPLPLVSNGALTLSDNETIAYQENNNIVS NLQANLVIYGPGPDIANNATYGLYSTPISNGEGTLSEVSFSPFYLKPFDESYGNYRSLVN IVNKFVKREFAPDPASTLMHQLVYVTHNLYGISNRNFYYNFDTGKIETSRQQNSLIFEEL LTFGGIDSKAISSLIIKKIIETAKNNYTTLISERLNTVTVENDLLKYIKNKIPVQGRLGN FKLDTAEFEKKLNTILFVLNESNLAQRFSILVRKHYLKERPIDPIYVNILDDNSYSTLEG FNISSQGSNDFQGQLLESSYFEKIESNALRAFIKICHKAIDGRSLYNKTLDCIEVENKDL FLISNKDSLNDINLSEEKIKPETTVFFKDKLPPQDITLSNYDFTEANSIPSISQQNILER NEELYEPIRNSLFEIKTIYVDKLTTFHFLEAQNIDESIDSSKIRVELTDSVDEALSNPNK VYSPFKNMSNTINSIETGITSTYIFYQWLRSIVKDFSDETGKIDVIDKSSDTLAIVPYIG PLLNIGNDIRHGDFVGAIELAGITALLEYVPEFTIPILVGLEVIGGELAREQVEAIVNNA LDKRDQKWAEVYNITKAQWWGTIHLQINTRLAHTYKALSRQANAIKMNMEFQLANYKGNI DDKAKIKNAISETEILLNKSVEQAMKNTEKFMIKLSNSYLTKEMIPKVQDNLKNFDLETK KTLDKFIKEKEDILGTNLSSSLRRKVSIRLNKNIAFDINDIPFSEFDDLINQYKNEILNN IILNLRYKDNNLIDLSGYGAKVEVYDGVELNDKNQFKLTSSANSKIRVTQNQNIIFNSVF LDFSVSFWIRIPKYKNDGIQNYIHNEYTIINCMKNNSGWKISIRGNRIIWTLIDINGKTK SVFFEYNIREDISEYINRWFFVTITNNLNNAKIYINGKLESNTDIKDIREVIANGEIIFK LDGDIDRTQFIWMKYFSIFNTELSQSNIEERYKIQSYSEYLKDFWGNPLMYNKEYYMFNA GNKNSYIKLKKDSPVGEILTRSKYNQNSKYINYRDLYIGEKFIIRRKSNSQSINDDIVRK EDYIYLDFFNLNQEWRVYTYKYFKKEEMKLFLAPIYDSDEFYNTIQIKEYDEQPTYSCQL LFKKDEESTDEIGLIGIHRFYESGIVFEEYKDYFCISKWYLKEVKRKPYNLKLGCNWQFI PKDEGWTE
SEQ ID NO:67-BoNT/XA(0)的多肽序列
MGSMKLEINKFNYNDPIDGINVITMRPPRH SDKINKGKGPFKAFQVIKNIWIVPERYNFTNNTNDLNIPSEPIMEADAIYNPNYLNTPSE KDEFLQGVIKVLERIKSKPEGEKLLELISSSIPLPLVSNGALTLSDNETIAYQENNNIVS NLQANLVIYGPGPDIANNATYGLYSTPISNGEGTLSEVSFSPFYLKPFDESYGNYRSLVN IVNKFVKREFAPDPASTLMHQLVYVTHNLYGISNRNFYYNFDTGKIETSRQQNSLIFEEL LTFGGIDSKAISSLIIKKIIETAKNNYTTLISERLNTVTVENDLLKYIKNKIPVQGRLGN FKLDTAEFEKKLNTILFVLNESNLAQRFSILVRKHYLKERPIDPIYVNILDDNSYSTLEG FNISSQGSNDFQGQLLESSYFEKIESNALRAFIKICHKAIDGRSLEVLFQGPLYNKTLDC IEVENKDLFLISNKDSLNDINLSEEKIKPETTVFFKDKLPPQDITLSNYDFTEANSIPSI SQQNILERNEELYEPIRNSLFEIKTIYVDKLTTFHFLEAQNIDESIDSSKIRVELTDSVD EALSNPNKVYSPFKNMSNTINSIETGITSTYIFYQWLRSIVKDFSDETGKIDVIDKSSDT LAIVPYIGPLLNIGNDIRHGDFVGAIELAGITALLEYVPEFTIPILVGLEVIGGELAREQ VEAIVNNALDKRDQKWAEVYNITKAQWWGTIHLQINTRLAHTYKALSRQANAIKMNMEFQ LANYKGNIDDKAKIKNAISETEILLNKSVEQAMKNTEKFMIKLSNSYLTKEMIPKVQDNL KNFDLETKKTLDKFIKEKEDILGTNLSSSLRRKVSIRLNKNIAFDINDIPFSEFDDLINQ YKNEIEDYEVLNLGAEDGKIKDLSGTTSDINIGSDIEIINTSILNLRYESNHLIDLSRYA SKINIGSKVNFDPIDKNQIQLFNLESSKIEVILKNAIVYNSMYENFSTSFWIRIPKYFNS ISLNNEYTIINCMENNSGWKVSLNYGEIIWTLQDTQEIKQRVVFKYSQMINISDYINRWI FVTITNNRLNNSKIYINGRLIDQKPISNLGNIHASNNIMFKLDGCRDTHRYIWIKYFNLF DKELNEKEIKDLYDNQSNSGILKDFWGDYLQYDKPYYMLNLYDPNKYVDVNNVGIRGYMY LKGPRGSVMTTNIYLNSSLYRGTKFIIKKYASGNKDNIVRNNDRVYINVVVKNKEYRLAT NASQAGVEKILSALEIPDVGNLSQVVVMKSKNDQGITNKCKMNLQDNNGNDIGFIGFHQF NNIAKLVASNWYNRQIERSSRTLGCSWEFIPVDDGWGERPL
SEQ ID NO:68-BoNT/XA(0)变体的多肽序列
MGSMKLEINKFNYNDPIDGINVITMRPPRH SDKINKGKGPFKAFQVIKNIWIVPERYNFTNNTNDLNIPSEPIMEADAIYNPNYLNTPSE KDEFLQGVIKVLERIKSKPEGEKLLELISSSIPLPLVSNGALTLSDNETIAYQENNNIVS NLQANLVIYGPGPDIANNATYGLYSTPISNGEGTLSEVSFSPFYLKPFDESYGNYRSLVN IVNKFVKREFAPDPASTLMHQLVYVTHNLYGISNRNFYYNFDTGKIETSRQQNSLIFEEL LTFGGIDSKAISSLIIKKIIETAKNNYTTLISERLNTVTVENDLLKYIKNKIPVQGRLGN FKLDTAEFEKKLNTILFVLNESNLAQRFSILVRKHYLKERPIDPIYVNILDDNSYSTLEG FNISSQGSNDFQGQLLESSYFEKIESNALRAFIKICHKAIDGRSLEVLFQGPLYNKTLDC IEVENKDLFLISNKDSLNDINLSEEKIKPETTVFFKDKLPPQDITLSNYDFTEANSIPSI SQQNILERNEELYEPIRNSLFEIKTIYVDKLTTFHFLEAQNIDESIDSSKIRVELTDSVD EALSNPNKVYSPFKNMSNTINSIETGITSTYIFYQWLRSIVKDFSDETGKIDVIDKSSDT LAIVPYIGPLLNIGNDIRHGDFVGAIELAGITALLEYVPEFTIPILVGLEVIGGELAREQ VEAIVNNALDKRDQKWAEVYNITKAQWWGTIHLQINTRLAHTYKALSRQANAIKMNMEFQ LANYKGNIDDKAKIKNAISETEILLNKSVEQAMKNTEKFMIKLSNSYLTKEMIPKVQDNL KNFDLETKKTLDKFIKEKEDILGTNLSSSLRRKVSIRLNKNIAFDINDIPFSEFDDLINQ YKNEIINTSILNLRYESNHLIDLSRYASKINIGSKVNFDPIDKNQIQLFNLESSKIEVIL KNAIVYNSMYENFSTSFWIRIPKYFNSISLNNEYTIINCMENNSGWKVSLNYGEIIWTLQ DTQEIKQRVVFKYSQMINISDYINRWIFVTITNNRLNNSKIYINGRLIDQKPISNLGNIH ASNNIMFKLDGCRDTHRYIWIKYFNLFDKELNEKEIKDLYDNQSNSGILKDFWGDYLQYD KPYYMLNLYDPNKYVDVNNVGIRGYMYLKGPRGSVMTTNIYLNSSLYRGTKFIIKKYASG NKDNIVRNNDRVYINVVVKNKEYRLATNASQAGVEKILSALEIPDVGNLSQVVVMKSKND QGITNKCKMNLQDNNGNDIGFIGFHQFNNIAKLVASNWYNRQIERSSRTLGCSWEFIPVD DGWGERPL
SEQ ID NO:69-BoNT/XD(0)的多肽序列
MGSMKLEINKFNYNDPIDGINVITMRPPRH SDKINKGKGPFKAFQVIKNIWIVPERYNFTNNTNDLNIPSEPIMEADAIYNPNYLNTPSE KDEFLQGVIKVLERIKSKPEGEKLLELISSSIPLPLVSNGALTLSDNETIAYQENNNIVS NLQANLVIYGPGPDIANNATYGLYSTPISNGEGTLSEVSFSPFYLKPFDESYGNYRSLVN IVNKFVKREFAPDPASTLMHQLVYVTHNLYGISNRNFYYNFDTGKIETSRQQNSLIFEEL LTFGGIDSKAISSLIIKKIIETAKNNYTTLISERLNTVTVENDLLKYIKNKIPVQGRLGN FKLDTAEFEKKLNTILFVLNESNLAQRFSILVRKHYLKERPIDPIYVNILDDNSYSTLEG FNISSQGSNDFQGQLLESSYFEKIESNALRAFIKICHKAIDGRSLYNKTLDCIEVENKDL FLISNKDSLNDINLSEEKIKPETTVFFKDKLPPQDITLSNYDFTEANSIPSISQQNILER NEELYEPIRNSLFEIKTIYVDKLTTFHFLEAQNIDESIDSSKIRVELTDSVDEALSNPNK VYSPFKNMSNTINSIETGITSTYIFYQWLRSIVKDFSDETGKIDVIDKSSDTLAIVPYIG PLLNIGNDIRHGDFVGAIELAGITALLEYVPEFTIPILVGLEVIGGELAREQVEAIVNNA LDKRDQKWAEVYNITKAQWWGTIHLQINTRLAHTYKALSRQANAIKMNMEFQLANYKGNIDDKAKIKNAISETEILLNKSVEQAMKNTEKFMIKLSNSYLTKEMIPKVQDNLKNFDLETK KTLDKFIKEKEDILGTNLSSSLRRKVSIRLNKNIAFDINDIPFSEFDDLINQYKNEIEDY EVLNLGAEDGKIKDLSGTTSDINIGSDIENDSKILSLQNKKNALVDTSGYNAEVRVGDNV QLNTIYTNDFKLSSSGDKIIVNLNNNILYSAIYENSSVSFWIKISKDLTNSHNEYTIINS IEQNSGWKLCIRNGNIEWILQDVNRKYKSLIFDYSESLSHTGYTNKWFFVTITNNIMGYM KLYINGELKQSQKIEDLDEVKLDKTIVFGIDENIDENQMLWIRDFNIFSKELSNEDINIV YEGQILRNVIKDYWGNPLKFDTEYYIINDNYIDRYIAPESNVLVLVQYPDRSKLYTGNPI TIKSVSDKNPYSRILNGDNIILHMLYNSRKYMIIRDTDTIYATQGGECSQNCVYALKLQS NLGNYGIGIFSIKNIVSKNKYCSQIFSSFRENTMLLADIYKPWRFSFKNAYTPVAVTNYE TKLLSTSSFWKFISRDPGWVE
SEQ ID NO:70-BoNT/XF(0)的多肽序列
MGSMKLEINKFNYNDPIDGINVITMRPPRH SDKINKGKGPFKAFQVIKNIWIVPERYNFTNNTNDLNIPSEPIMEADAIYNPNYLNTPSE KDEFLQGVIKVLERIKSKPEGEKLLELISSSIPLPLVSNGALTLSDNETIAYQENNNIVS NLQANLVIYGPGPDIANNATYGLYSTPISNGEGTLSEVSFSPFYLKPFDESYGNYRSLVN IVNKFVKREFAPDPASTLMHQLVYVTHNLYGISNRNFYYNFDTGKIETSRQQNSLIFEEL LTFGGIDSKAISSLIIKKIIETAKNNYTTLISERLNTVTVENDLLKYIKNKIPVQGRLGN FKLDTAEFEKKLNTILFVLNESNLAQRFSILVRKHYLKERPIDPIYVNILDDNSYSTLEG FNISSQGSNDFQGQLLESSYFEKIESNALRAFIKICHKAIDGRSLYNKTLDCIEVENKDL FLISNKDSLNDINLSEEKIKPETTVFFKDKLPPQDITLSNYDFTEANSIPSISQQNILER NEELYEPIRNSLFEIKTIYVDKLTTFHFLEAQNIDESIDSSKIRVELTDSVDEALSNPNK VYSPFKNMSNTINSIETGITSTYIFYQWLRSIVKDFSDETGKIDVIDKSSDTLAIVPYIG PLLNIGNDIRHGDFVGAIELAGITALLEYVPEFTIPILVGLEVIGGELAREQVEAIVNNA LDKRDQKWAEVYNITKAQWWGTIHLQINTRLAHTYKALSRQANAIKMNMEFQLANYKGNIDDKAKIKNAISETEILLNKSVEQAMKNTEKFMIKLSNSYLTKEMIPKVQDNLKNFDLETK KTLDKFIKEKEDILGTNLSSSLRRKVSIRLNKNIAFDINDIPFSEFDDLINQYKNEIEDY EVLNLGAEDGKIKDLSGTTSDINIGSDIEIKDNSILDMRYENNKFIDISGYGSNISINGD VYIYSTNRNQFGIYSSKPSEVNIAQNNDIIYNGRYQNFSISFWVRIPKYFNKVNLNNEYT IIDCIRNNNSGWKISLNYNKIIWTLQDTAGNNQKLVFNYTQMISISDYINKWIFVTITNN RLGNSRIYINGNLIDEKSISNLGDIHVSDNILFKIVGCNDTRYVGIRYFKVFDTELGKTE IETLYSDEPDPSILKDFWGNYLLYNKRYYLLNLLRTDKSITQNSNFLNINQQRGVYQKPN IFSNTRLYTGVEVIIRKNGSTDISNTDNFVRKNDLAYINVVDRDVEYRLYADISIAKPEK IIKLIRTSNSNNSLGQIIVMDSIGNNCTMNFQNNNGGNIGLLGFKSNNLVASSWYYNNIR KNTSSNGCFWSFISKEHGWQEN
SEQ ID NO:71-C1激活环共有序列
Cys-(Xaa)a-Ile-Asp/Glu-Gly-Arg-(Yaa)b-Cys,其中a＝1-10且b＝4-15
SEQ ID NO:72-C1激活环
CHKAIDGRSLYNKTLDC
SEQ ID NO:73-C1激活环变体
CHKAIEGRSLYNKTLDC
SEQ ID NO:74-rLC/A(0)的多肽序列(His标签)
SEQ ID NO:75-rLHN/A(0)的多肽序列(His标签)
SEQ ID NO:76-rLC/X(0)的多肽序列
SEQ ID NO:77-前切割蛋白酶位点
LEVLFQGP
SEQ ID NO:78-C1激活环变体2
CHKAIDGRSLEVLFQGPLYNKTLDC
实施例
实施例1
BoNT/A(0)在体外和体内都无催化活性
在体外基于细胞的模型中测试BoNT/A(0)(SEQ ID NO:2)的催化活性，该模型测量BoNT/A靶SNARE蛋白SNAP25的切割。图1显示，与野生型BoNT/A(SEQ ID NO:60)相比，在人神经元测定法中，BoNT/A(0)不切割SNAP25。图2在大鼠神经元测定法中确认了这一结果。
作为确认，使用BoNT/A和BoNT/A(0)进行了体内DAS测定。通过将20μl配制在明胶磷酸盐缓冲液中的梭菌毒素注射到小鼠腓肠肌/比目鱼肌复合体中，然后使用Aoki的方法(Aoki KR,Toxicon 39:1815-1820；2001)评估趾外展评分来进行DAS测定。在DAS测定法中，将小鼠短暂尾巴悬吊，以引发特征性的惊吓反应(图3A)，其中小鼠伸展其后肢并外展后趾。梭菌毒素注射后，以五分制对不同程度的趾外展进行评分(0＝正常至4＝趾外展的最大减少－图3B)。这提供神经-肌肉连接处神经毒素活性诱导的麻痹的功能测量。此外，在施用7天内评估小鼠的体重变化。这提供毒素从施用部位扩散的毒性和不良影响的测量。结果如下表1所示。
表1.施用BoNT/A(0)或BoNT/A后的DAS评分(24小时后)和体重变化。
DAS评分 观察到体重变化
BoNT/A 4 是
BoNT/A(0) 0 否
结果确认BoNT/A(0)在体内无催化活性，并且不会导致任何毒性症状。因此，BoNT/A(0)是一种安全、基本无毒的治疗剂。
实施例2
使用无催化活性的BoNT治疗慢性神经病性疼痛(慢性挤压损伤(CCI)大鼠模型)
材料和方法
慢性挤压损伤(chronic constriction injury，CCI)如Bennett和Xie(1988),Pain,33(1):87-107之前所述进行。在第-14天，将成年雄性Sprague-Dawley大鼠(220-250g)麻醉，然后暴露左侧坐骨神经的一段，并在神经上放置四个松散的丝线结扎。在第0天(D0)，通过足底内(i.pl.)途径对大鼠注射BoNT/A(30pg/kg)、BoNT/A(60pg/kg)、BoNT/A(0)(60pg/kg)或溶媒(GPB)(n＝10-11只/组)，作为阳性对照，将加巴喷丁(100mg/kg)通过口服途径施用(n＝8只/组)。用加巴喷丁处理的动物在处理后1、2和4小时进行测试。BoNT/A、BoNT/A(0)或溶媒处理的动物在第3、5和9天测试。在冯-弗雷试验中评估动物的机械敏感性。
结果
实验显示，通过施用无催化活性BoNT(BoNT/A(0))，同侧爪的机械敏感性降低(图4B)。此外，与同等给药的BoNT/A相比，BoNT/A(0)在降低机械敏感性方面更有效。在以后的时间点，BoNT/A(0)也比加巴喷丁更有效。为了确认敏感性降低是施用BoNT/A(0)的结果，还测试了对侧爪的机械敏感性。结果显示，不同条件下对侧爪的机械敏感性差异不显著(图4C)。此外，作为BoNT/A(0)基本无毒性的确认，体重随时间的变化等同于施用溶媒或加巴喷丁的大鼠的变化(图4D)。这与在施用催化活性BoNT/A的大鼠中观察到的变化形成对比，其在第9天具有统计学显著差异。
总之，无催化活性梭菌神经毒素出人意料地能够减轻疼痛(例如慢性神经病性疼痛)，从而表明这类神经毒素是适宜的疼痛治疗剂。
实施例3
使用无催化活性BoNT治疗急性神经病性疼痛(奥沙利铂大鼠模型)
材料和方法
通过腹膜内注射奥沙利铂来诱导奥沙利铂诱导的周围感觉神经病的实验模型(Ling等(2007),Pain,128(3):225-234；Ling等(2007),Toxicology,20；234(3):176-84)。在第0天，成年雄性Sprague-Dawley大鼠(100-133g)接受腹膜内注射假处理(5％葡萄糖)或奥沙利铂(10mg/kg)。紧接着，假处理的动物接受溶媒的足底内注射，而奥沙利铂处理的动物则接受BoNT/A(0)(1000pg/kg)、BoNT/A(50pg/kg)、BoNT/A(100pg/kg)、BoNT/A(160pg/kg)或溶媒(GPB；n＝10/组)的足底内注射。在D3，通过口服途径施用作为阳性对照的度洛西汀(100mg/kg)。在D3和D5评价动物的热(冷)敏感性。
结果
实验显示，通过施用无催化活性BoNT(BoNT/A(0))，同侧爪的冷敏感性降低(图5B)。BoNT/A、BoNT/A(0)或溶媒处理组对侧爪的热敏感性没有差异(图5C)。
总之，无催化活性梭菌神经毒素出人意料地能够减轻急性神经病性疼痛，从而表明这类神经毒素在治疗疼痛方面具有通用应用。
实施例4
使用无催化活性BoNT治疗慢性神经病性疼痛(奥沙利铂大鼠模型)
材料和方法
在第-2天(D-2)，雄性Sprague-Dawley大鼠(180–210g)接受奥沙利铂(10mg/kg)的腹膜内注射，然后在第0天通过足底内途径用BoNT/A(100pg/kg)、BoNT/A(0)(100pg/kg)或溶媒(GPB)处理(n＝11-12/组)。在D3，施用作为阳性对照的普瑞巴林(n＝12)。在第3、6和9天对动物进行机械敏感性(冯-弗雷试验)和热(冷)敏感性(冷板试验)测试。
结果
实验显示，通过施用无催化活性BoNT(BoNT/A(0))，同侧爪的机械敏感性(图6B)和冷敏感性(图6D)降低。
总之，在不同的化疗诱导的疼痛模型中，无催化活性梭菌神经毒素出人意料地能够减轻慢性神经病性疼痛。
实施例5
使用无催化活性BoNT治疗炎性疼痛(UV-B灼伤大鼠模型)
材料和方法
在人类以及啮齿类动物模型中，紫外线(UV)-B照射导致机械和热痛觉过敏。成年雄性Wistar大鼠(180-210g)通过足底内注射施用BoNT/A(100pg/kg)、BoNT/A(0)(100pg/kg)或溶媒(GPB；n＝12/组)。24小时后，使同侧爪的足底表面暴露于紫外线-B(UVB)照射约5分钟，接受500mJ/cm2的剂量。UVB后48小时和BoNT/A、BoNT/A(0)或溶媒注射后72小时，在冯-弗雷试验中测试动物的机械敏感性。另外一组暴露于UVB的动物在48小时后注射阳性对照吲哚美辛，并在注射后1小时在冯-弗雷试验中进行测试(n＝12/组)。
结果
实验显示，通过施用无催化活性BoNT(BoNT/A(0))，机械敏感性降低(图7B)。总之，无催化活性梭菌神经毒素出人意料地能够减轻炎性疼痛(例如急性炎性疼痛)，从而确认了这类神经毒素在治疗疼痛方面具有通用应用。
无催化活性梭菌神经毒素减轻炎性疼痛这一令人惊讶的发现表明，它在治疗潜在的炎性病症(例如，包括治疗炎性病症的至少一种症状，即相关疼痛)方面是有用的。因此，认为无催化活性梭菌神经毒素可用于治疗炎性疾病是可信的。
实施例6
使用无催化活性的嵌合BoNT治疗炎性疼痛(CFA诱导的炎性疼痛模型)
材料和方法
在BoNT或溶媒给药之前，使用递增力的分级冯-弗雷纤维丝连续3天评估70只成年雄性C57/BL6小鼠(22-26g)的爪缩回阈值(PWT，g)。最后2天的平均值视为基线。在第0天，在气体麻醉下，将BoNT/XB(0.3和30ng/kg)、BoNT/XB(0)(0.3和30ng/kg)、BoNT/A(160pg/kg)或溶媒(840μl/kg)足底内注射到左后爪的足垫。第2天，在注射CFA之前，对PWT进行重新评估。然后在异氟烷麻醉下，将固定体积为20μL的CFA(1.5mg/mL)注射到同一后爪中。在第3天(CFA后第1天)，在PWT评估前1小时，用吲哚美辛(10mg/kg，n＝9)对分配给吲哚美辛组的动物进行口服给药。
结果
实验显示，包含无催化活性BoNT/X L链以及转运结构域(BoNT/X LHN)和BoNT/B受体结合结构域BoNT/X(HC结构域)的无催化活性的嵌合BoNT(BoNT/XB(0))在治疗炎性疼痛方面有效。更详细地说，图8显示，在施用30ng/kg剂量的催化活性BoNT/XB和无催化活性BoNT/XB(0)的小鼠中，CFA诱导炎性疼痛后的机械敏感性降低。敏感性的降低等同于BoNT/A或阳性对照吲哚美辛的降低。
BoNT/XB(0)降低炎性疼痛的惊人发现表明，它在治疗潜在的炎性病症(例如，包括治疗炎性病症的至少一种症状，即相关疼痛)方面是有用的。因此，认为它进一步证明了无催化活性梭菌神经毒素用于治疗炎性病症的可信度。
实施例7
使用无催化活性的嵌合BoNT治疗特应性皮炎
在暴露于钙泊三醇(Calcipotriol)前一天，在成年C57/BL6小鼠背部内侧部皮下施用溶媒或BoNT/XB(0)(40pg/小鼠、100pg/小鼠或400pg/小鼠)。然后连续5天用钙泊三醇处理小鼠。研究结束时，对动物实施安乐死，收集背部皮肤，固定并处理用于进行组织学分析。苏木精和伊红染色后，评价表皮厚度。进行免疫标记以证明CD45+细胞。
实验显示，包含无催化活性BoNT/X L链以及转运结构域(BoNT/X LHN)和BoNT/B受体结合结构域BoNT/X(HC结构域)的无催化活性的嵌合BoNT(BoNT/XB(0))对治疗特应性皮炎(一种模型炎性病症)有效。结果显示施用BoNT/XB(0)后皮肤厚度改善。皮肤厚度指示纤维化，这是对钙泊三醇的炎症反应，并且在BoNT/XB(0)治疗的动物中显示出统计学显著性降低。此外，BoNT/XB(0)的抗炎作用通过BoNT/XB(0)处理的动物中CD45阳性细胞数量的减少得到确认(CD45在炎症细胞中转导激活信号)。
因此，结论是BoNT/XB(0)具有抗炎特性，从而在治疗炎性疾病中有用。
上述说明书中提到的所有出版物在此引入作为参考。在不脱离本发明的范围和精神的情况下，所描述的本发明的方法和系统的多种修改和变化对于本领域技术人员而言显而易见。尽管已经结合具体的优选实施方案描述了本发明，但是应当理解，所要求保护的本发明不应当过度地局限于这类具体实施方案。事实上，生物化学和生物技术或相关领域的技术人员显而易见的用于实施本发明的所描述的方式的多种修改旨在落入以下权利要求书的范围之内。
序列表
<110> 益普生生物制药有限公司
<120> 无催化活性的梭菌神经毒素用于疼痛和炎性疾病的治疗
<130> P68298WO
<160> 78
<170> PatentIn版本3.5
<210> 1
<211> 3888
<212> DNA
<213> 人工序列
<220>
<223> rBoNT/A(0)的核苷酸序列
<400> 1
atgccattcg tcaacaagca attcaactac aaagacccag tcaacggcgt cgacatcgca 60
tacatcaaga ttccgaacgc cggtcaaatg cagccggtta aggcttttaa gatccacaac 120
aagatttggg ttatcccgga gcgtgacacc ttcacgaacc cggaagaagg cgatctgaac 180
ccgccaccgg aagcgaagca agtccctgtc agctactacg attcgacgta cctgagcacg 240
gataacgaaa aagataacta cctgaaaggt gtgaccaagc tgttcgaacg tatctacagc 300
acggatctgg gtcgcatgct gctgactagc attgttcgcg gtatcccgtt ctggggtggt 360
agcacgattg acaccgaact gaaggttatc gacactaact gcattaacgt tattcaaccg 420
gatggtagct atcgtagcga agagctgaat ctggtcatca ttggcccgag cgcagacatt 480
atccaattcg agtgcaagag ctttggtcac gaggttctga atctgacccg caatggctat 540
ggtagcaccc agtacattcg tttttcgccg gattttacct tcggctttga agagagcctg 600
gaggttgata ccaatccgtt gctgggtgcg ggcaaattcg ctaccgatcc ggctgtcacg 660
ctggcccatc aactgatcta cgcaggccac cgcctgtacg gcattgccat caacccaaac 720
cgtgtgttca aggttaatac gaatgcatac tacgagatga gcggcctgga agtcagcttc 780
gaagaactgc gcaccttcgg tggccatgac gctaaattca ttgacagctt gcaagagaat 840
gagttccgtc tgtactacta taacaaattc aaagacattg caagcacgtt gaacaaggcc 900
aaaagcatcg ttggtactac cgcgtcgttg cagtatatga agaatgtgtt taaagagaag 960
tacctgctgt ccgaggatac ctccggcaag tttagcgttg ataagctgaa gtttgacaaa 1020
ctgtacaaga tgctgaccga gatttacacc gaggacaact ttgtgaaatt cttcaaagtg 1080
ttgaatcgta aaacctatct gaattttgac aaagcggttt tcaagattaa catcgtgccg 1140
aaggtgaact acaccatcta tgacggtttt aacctgcgta acaccaacct ggcggcgaac 1200
tttaacggtc agaatacgga aatcaacaac atgaatttca cgaagttgaa gaacttcacg 1260
ggtctgttcg agttctataa gctgctgtgc gtgcgcggta tcatcaccag caaaaccaaa 1320
agcctggaca aaggctacaa caaggcgctg aatgacctgt gcattaaggt aaacaattgg 1380
gatctgttct tttcgccatc cgaagataat tttaccaacg acctgaacaa gggtgaagaa 1440
atcaccagcg atacgaatat tgaagcagcg gaagagaata tcagcctgga tctgatccag 1500
cagtactatc tgacctttaa cttcgacaat gaaccggaga acattagcat tgagaatctg 1560
agcagcgaca ttatcggtca gctggaactg atgccgaata tcgaacgttt cccgaacggc 1620
aaaaagtacg agctggacaa gtacactatg ttccattacc tgcgtgcaca ggagtttgaa 1680
cacggtaaaa gccgtatcgc gctgaccaac agcgttaacg aggccctgct gaacccgagc 1740
cgtgtctata ccttcttcag cagcgactat gttaagaaag tgaacaaagc cactgaggcc 1800
gcgatgttcc tgggctgggt ggaacagctg gtatatgact tcacggacga gacgagcgaa 1860
gtgagcacta ccgacaaaat tgctgatatt accatcatta tcccgtatat tggtccggca 1920
ctgaacattg gcaacatgct gtacaaagac gattttgtgg gtgccctgat cttctccggt 1980
gccgtgattc tgctggagtt cattccggag attgcgatcc cggtgttggg taccttcgcg 2040
ctggtgtcct acatcgcgaa taaggttctg acggttcaga ccatcgataa cgcgctgtcg 2100
aaacgtaatg aaaaatggga cgaggtttac aaatacattg ttacgaattg gctggcgaaa 2160
gtcaataccc agatcgacct gatccgtaag aaaatgaaag aggcgctgga gaatcaggcg 2220
gaggccacca aagcaattat caactaccaa tacaaccagt acacggaaga agagaagaat 2280
aacattaact tcaatatcga tgatttgagc agcaagctga atgaatctat caacaaagcg 2340
atgatcaata tcaacaagtt tttgaatcag tgtagcgttt cgtacctgat gaatagcatg 2400
attccgtatg gcgtcaaacg tctggaggac ttcgacgcca gcctgaaaga tgcgttgctg 2460
aaatacattt acgacaatcg tggtacgctg attggccaag ttgaccgctt gaaagacaaa 2520
gttaacaata ccctgagcac cgacatccca tttcaactga gcaagtatgt tgataatcaa 2580
cgtctgttga gcactttcac cgagtatatc aaaaacatca tcaatactag cattctgaac 2640
ctgcgttacg agagcaatca tctgattgat ctgagccgtt atgcaagcaa gatcaacatc 2700
ggtagcaagg tcaattttga cccgatcgat aagaaccaga tccagctgtt taatctggaa 2760
tcgagcaaaa ttgaggttat cctgaaaaac gccattgtct acaactccat gtacgagaat 2820
ttctccacca gcttctggat tcgcatcccg aaatacttca acagcattag cctgaacaac 2880
gagtatacta tcatcaactg tatggagaac aacagcggtt ggaaggtgtc tctgaactat 2940
ggtgagatca tttggacctt gcaggacacc caagagatca agcagcgcgt cgtgttcaag 3000
tactctcaaa tgatcaacat ttccgattac attaatcgtt ggatcttcgt gaccattacg 3060
aataaccgtc tgaataacag caagatttac atcaatggtc gcttgatcga tcagaaaccg 3120
attagcaacc tgggtaatat ccacgcaagc aacaacatta tgttcaaatt ggacggttgc 3180
cgcgataccc atcgttatat ctggatcaag tatttcaacc tgtttgataa agaactgaat 3240
gagaaggaga tcaaagattt gtatgacaac caatctaaca gcggcatttt gaaggacttc 3300
tggggcgatt atctgcaata cgataagccg tactatatgc tgaacctgta tgatccgaac 3360
aaatatgtgg atgtcaataa tgtgggtatt cgtggttaca tgtatttgaa gggtccgcgt 3420
ggcagcgtta tgacgaccaa catttacctg aactctagcc tgtaccgtgg tacgaaattc 3480
atcattaaga aatatgccag cggcaacaaa gataacattg tgcgtaataa cgatcgtgtc 3540
tacatcaacg tggtcgtgaa gaataaagag taccgtctgg cgaccaacgc ttcgcaggcg 3600
ggtgttgaga aaattctgag cgcgttggag atccctgatg tcggtaatct gagccaagtc 3660
gtggttatga agagcaagaa cgaccagggt atcactaaca agtgcaagat gaacctgcaa 3720
gacaacaatg gtaacgacat cggctttatt ggtttccacc agttcaacaa tattgctaaa 3780
ctggtagcga gcaattggta caatcgtcag attgagcgca gcagccgtac tttgggctgt 3840
agctgggagt ttatcccggt cgatgatggt tggggcgaac gtccgctg 3888
<210> 2
<211> 1296
<212> PRT
<213> 人工序列
<220>
<223> rBoNT/A(0)的多肽序列
<400> 2
Met Pro Phe Val Asn Lys Gln Phe Asn Tyr Lys Asp Pro Val Asn Gly
1 5 10 15
Val Asp Ile Ala Tyr Ile Lys Ile Pro Asn Ala Gly Gln Met Gln Pro
20 25 30
Val Lys Ala Phe Lys Ile His Asn Lys Ile Trp Val Ile Pro Glu Arg
35 40 45
Asp Thr Phe Thr Asn Pro Glu Glu Gly Asp Leu Asn Pro Pro Pro Glu
50 55 60
Ala Lys Gln Val Pro Val Ser Tyr Tyr Asp Ser Thr Tyr Leu Ser Thr
65 70 75 80
Asp Asn Glu Lys Asp Asn Tyr Leu Lys Gly Val Thr Lys Leu Phe Glu
85 90 95
Arg Ile Tyr Ser Thr Asp Leu Gly Arg Met Leu Leu Thr Ser Ile Val
100 105 110
Arg Gly Ile Pro Phe Trp Gly Gly Ser Thr Ile Asp Thr Glu Leu Lys
115 120 125
Val Ile Asp Thr Asn Cys Ile Asn Val Ile Gln Pro Asp Gly Ser Tyr
130 135 140
Arg Ser Glu Glu Leu Asn Leu Val Ile Ile Gly Pro Ser Ala Asp Ile
145 150 155 160
Ile Gln Phe Glu Cys Lys Ser Phe Gly His Glu Val Leu Asn Leu Thr
165 170 175
Arg Asn Gly Tyr Gly Ser Thr Gln Tyr Ile Arg Phe Ser Pro Asp Phe
180 185 190
Thr Phe Gly Phe Glu Glu Ser Leu Glu Val Asp Thr Asn Pro Leu Leu
195 200 205
Gly Ala Gly Lys Phe Ala Thr Asp Pro Ala Val Thr Leu Ala His Gln
210 215 220
Leu Ile Tyr Ala Gly His Arg Leu Tyr Gly Ile Ala Ile Asn Pro Asn
225 230 235 240
Arg Val Phe Lys Val Asn Thr Asn Ala Tyr Tyr Glu Met Ser Gly Leu
245 250 255
Glu Val Ser Phe Glu Glu Leu Arg Thr Phe Gly Gly His Asp Ala Lys
260 265 270
Phe Ile Asp Ser Leu Gln Glu Asn Glu Phe Arg Leu Tyr Tyr Tyr Asn
275 280 285
Lys Phe Lys Asp Ile Ala Ser Thr Leu Asn Lys Ala Lys Ser Ile Val
290 295 300
Gly Thr Thr Ala Ser Leu Gln Tyr Met Lys Asn Val Phe Lys Glu Lys
305 310 315 320
Tyr Leu Leu Ser Glu Asp Thr Ser Gly Lys Phe Ser Val Asp Lys Leu
325 330 335
Lys Phe Asp Lys Leu Tyr Lys Met Leu Thr Glu Ile Tyr Thr Glu Asp
340 345 350
Asn Phe Val Lys Phe Phe Lys Val Leu Asn Arg Lys Thr Tyr Leu Asn
355 360 365
Phe Asp Lys Ala Val Phe Lys Ile Asn Ile Val Pro Lys Val Asn Tyr
370 375 380
Thr Ile Tyr Asp Gly Phe Asn Leu Arg Asn Thr Asn Leu Ala Ala Asn
385 390 395 400
Phe Asn Gly Gln Asn Thr Glu Ile Asn Asn Met Asn Phe Thr Lys Leu
405 410 415
Lys Asn Phe Thr Gly Leu Phe Glu Phe Tyr Lys Leu Leu Cys Val Arg
420 425 430
Gly Ile Ile Thr Ser Lys Thr Lys Ser Leu Asp Lys Gly Tyr Asn Lys
435 440 445
Ala Leu Asn Asp Leu Cys Ile Lys Val Asn Asn Trp Asp Leu Phe Phe
450 455 460
Ser Pro Ser Glu Asp Asn Phe Thr Asn Asp Leu Asn Lys Gly Glu Glu
465 470 475 480
Ile Thr Ser Asp Thr Asn Ile Glu Ala Ala Glu Glu Asn Ile Ser Leu
485 490 495
Asp Leu Ile Gln Gln Tyr Tyr Leu Thr Phe Asn Phe Asp Asn Glu Pro
500 505 510
Glu Asn Ile Ser Ile Glu Asn Leu Ser Ser Asp Ile Ile Gly Gln Leu
515 520 525
Glu Leu Met Pro Asn Ile Glu Arg Phe Pro Asn Gly Lys Lys Tyr Glu
530 535 540
Leu Asp Lys Tyr Thr Met Phe His Tyr Leu Arg Ala Gln Glu Phe Glu
545 550 555 560
His Gly Lys Ser Arg Ile Ala Leu Thr Asn Ser Val Asn Glu Ala Leu
565 570 575
Leu Asn Pro Ser Arg Val Tyr Thr Phe Phe Ser Ser Asp Tyr Val Lys
580 585 590
Lys Val Asn Lys Ala Thr Glu Ala Ala Met Phe Leu Gly Trp Val Glu
595 600 605
Gln Leu Val Tyr Asp Phe Thr Asp Glu Thr Ser Glu Val Ser Thr Thr
610 615 620
Asp Lys Ile Ala Asp Ile Thr Ile Ile Ile Pro Tyr Ile Gly Pro Ala
625 630 635 640
Leu Asn Ile Gly Asn Met Leu Tyr Lys Asp Asp Phe Val Gly Ala Leu
645 650 655
Ile Phe Ser Gly Ala Val Ile Leu Leu Glu Phe Ile Pro Glu Ile Ala
660 665 670
Ile Pro Val Leu Gly Thr Phe Ala Leu Val Ser Tyr Ile Ala Asn Lys
675 680 685
Val Leu Thr Val Gln Thr Ile Asp Asn Ala Leu Ser Lys Arg Asn Glu
690 695 700
Lys Trp Asp Glu Val Tyr Lys Tyr Ile Val Thr Asn Trp Leu Ala Lys
705 710 715 720
Val Asn Thr Gln Ile Asp Leu Ile Arg Lys Lys Met Lys Glu Ala Leu
725 730 735
Glu Asn Gln Ala Glu Ala Thr Lys Ala Ile Ile Asn Tyr Gln Tyr Asn
740 745 750
Gln Tyr Thr Glu Glu Glu Lys Asn Asn Ile Asn Phe Asn Ile Asp Asp
755 760 765
Leu Ser Ser Lys Leu Asn Glu Ser Ile Asn Lys Ala Met Ile Asn Ile
770 775 780
Asn Lys Phe Leu Asn Gln Cys Ser Val Ser Tyr Leu Met Asn Ser Met
785 790 795 800
Ile Pro Tyr Gly Val Lys Arg Leu Glu Asp Phe Asp Ala Ser Leu Lys
805 810 815
Asp Ala Leu Leu Lys Tyr Ile Tyr Asp Asn Arg Gly Thr Leu Ile Gly
820 825 830
Gln Val Asp Arg Leu Lys Asp Lys Val Asn Asn Thr Leu Ser Thr Asp
835 840 845
Ile Pro Phe Gln Leu Ser Lys Tyr Val Asp Asn Gln Arg Leu Leu Ser
850 855 860
Thr Phe Thr Glu Tyr Ile Lys Asn Ile Ile Asn Thr Ser Ile Leu Asn
865 870 875 880
Leu Arg Tyr Glu Ser Asn His Leu Ile Asp Leu Ser Arg Tyr Ala Ser
885 890 895
Lys Ile Asn Ile Gly Ser Lys Val Asn Phe Asp Pro Ile Asp Lys Asn
900 905 910
Gln Ile Gln Leu Phe Asn Leu Glu Ser Ser Lys Ile Glu Val Ile Leu
915 920 925
Lys Asn Ala Ile Val Tyr Asn Ser Met Tyr Glu Asn Phe Ser Thr Ser
930 935 940
Phe Trp Ile Arg Ile Pro Lys Tyr Phe Asn Ser Ile Ser Leu Asn Asn
945 950 955 960
Glu Tyr Thr Ile Ile Asn Cys Met Glu Asn Asn Ser Gly Trp Lys Val
965 970 975
Ser Leu Asn Tyr Gly Glu Ile Ile Trp Thr Leu Gln Asp Thr Gln Glu
980 985 990
Ile Lys Gln Arg Val Val Phe Lys Tyr Ser Gln Met Ile Asn Ile Ser
995 1000 1005
Asp Tyr Ile Asn Arg Trp Ile Phe Val Thr Ile Thr Asn Asn Arg
1010 1015 1020
Leu Asn Asn Ser Lys Ile Tyr Ile Asn Gly Arg Leu Ile Asp Gln
1025 1030 1035
Lys Pro Ile Ser Asn Leu Gly Asn Ile His Ala Ser Asn Asn Ile
1040 1045 1050
Met Phe Lys Leu Asp Gly Cys Arg Asp Thr His Arg Tyr Ile Trp
1055 1060 1065
Ile Lys Tyr Phe Asn Leu Phe Asp Lys Glu Leu Asn Glu Lys Glu
1070 1075 1080
Ile Lys Asp Leu Tyr Asp Asn Gln Ser Asn Ser Gly Ile Leu Lys
1085 1090 1095
Asp Phe Trp Gly Asp Tyr Leu Gln Tyr Asp Lys Pro Tyr Tyr Met
1100 1105 1110
Leu Asn Leu Tyr Asp Pro Asn Lys Tyr Val Asp Val Asn Asn Val
1115 1120 1125
Gly Ile Arg Gly Tyr Met Tyr Leu Lys Gly Pro Arg Gly Ser Val
1130 1135 1140
Met Thr Thr Asn Ile Tyr Leu Asn Ser Ser Leu Tyr Arg Gly Thr
1145 1150 1155
Lys Phe Ile Ile Lys Lys Tyr Ala Ser Gly Asn Lys Asp Asn Ile
1160 1165 1170
Val Arg Asn Asn Asp Arg Val Tyr Ile Asn Val Val Val Lys Asn
1175 1180 1185
Lys Glu Tyr Arg Leu Ala Thr Asn Ala Ser Gln Ala Gly Val Glu
1190 1195 1200
Lys Ile Leu Ser Ala Leu Glu Ile Pro Asp Val Gly Asn Leu Ser
1205 1210 1215
Gln Val Val Val Met Lys Ser Lys Asn Asp Gln Gly Ile Thr Asn
1220 1225 1230
Lys Cys Lys Met Asn Leu Gln Asp Asn Asn Gly Asn Asp Ile Gly
1235 1240 1245
Phe Ile Gly Phe His Gln Phe Asn Asn Ile Ala Lys Leu Val Ala
1250 1255 1260
Ser Asn Trp Tyr Asn Arg Gln Ile Glu Arg Ser Ser Arg Thr Leu
1265 1270 1275
Gly Cys Ser Trp Glu Phe Ile Pro Val Asp Asp Gly Trp Gly Glu
1280 1285 1290
Arg Pro Leu
1295
<210> 3
<211> 2634
<212> DNA
<213> 人工序列
<220>
<223> rLHN/A的核苷酸序列
<400> 3
atggagttcg ttaacaaaca gttcaactat aaagacccag ttaacggtgt tgacattgct 60
tacatcaaaa tcccgaacgc tggccagatg cagccggtaa aggcattcaa aatccacaac 120
aaaatctggg ttatcccgga acgtgatacc tttactaacc cggaagaagg tgacctgaac 180
ccgccaccgg aagcgaaaca ggtgccggta tcttactatg actccaccta cctgtctacc 240
gataacgaaa aggacaacta cctgaaaggt gttactaaac tgttcgagcg tatttactcc 300
accgacctgg gccgtatgct gctgactagc atcgttcgcg gtatcccgtt ctggggcggt 360
tctaccatcg ataccgaact gaaagtaatc gacactaact gcatcaacgt tattcagccg 420
gacggttcct atcgttccga agaactgaac ctggtgatca tcggcccgtc tgctgatatc 480
atccagttcg agtgtaagag ctttggtcac gaagttctga acctcacccg taacggctac 540
ggttccactc agtacatccg tttctctccg gacttcacct tcggttttga agaatccctg 600
gaagtagaca cgaacccact gctgggcgct ggtaaattcg caactgatcc tgcggttacc 660
ctggctcacg aactgattca tgcaggccac cgcctgtacg gtatcgccat caatccgaac 720
cgtgtcttca aagttaacac caacgcgtat tacgagatgt ccggtctgga agttagcttc 780
gaagaactgc gtacttttgg cggtcacgac gctaaattca tcgactctct gcaagaaaac 840
gagttccgtc tgtactacta taacaagttc aaagatatcg catccaccct gaacaaagcg 900
aaatccatcg tgggtaccac tgcttctctc cagtacatga agaacgtttt taaagaaaaa 960
tacctgctca gcgaagacac ctccggcaaa ttctctgtag acaagttgaa attcgataaa 1020
ctttacaaaa tgctgactga aatttacacc gaagacaact tcgttaagtt ctttaaagtt 1080
ctgaaccgca aaacctatct gaacttcgac aaggcagtat tcaaaatcaa catcgtgccg 1140
aaagttaact acactatcta cgatggtttc aacctgcgta acaccaacct ggctgctaat 1200
tttaacggcc agaacacgga aatcaacaac atgaacttca caaaactgaa aaacttcact 1260
ggtctgttcg agttttacaa gctgctgtgc gtcgacggca tcattacctc caaaactaaa 1320
tctgacgatg acgataaaaa caaagcgctg aacctgcagt gtatcaaggt taacaactgg 1380
gatttattct tcagcccgag tgaagacaac ttcaccaacg acctgaacaa aggtgaagaa 1440
atcacctcag atactaacat cgaagcagcc gaagaaaaca tctcgctgga cctgatccag 1500
cagtactacc tgacctttaa tttcgacaac gagccggaaa acatttctat cgaaaacctg 1560
agctctgata tcatcggcca gctggaactg atgccgaaca tcgaacgttt cccaaacggt 1620
aaaaagtacg agctggacaa atataccatg ttccactacc tgcgcgcgca ggaatttgaa 1680
cacggcaaat cccgtatcgc actgactaac tccgttaacg aagctctgct caacccgtcc 1740
cgtgtataca ccttcttctc tagcgactac gtgaaaaagg tcaacaaagc gactgaagct 1800
gcaatgttct tgggttgggt tgaacagctt gtttatgatt ttaccgacga gacgtccgaa 1860
gtatctacta ccgacaaaat tgcggatatc actatcatca tcccgtacat cggtccggct 1920
ctgaacattg gcaacatgct gtacaaagac gacttcgttg gcgcactgat cttctccggt 1980
gcggtgatcc tgctggagtt catcccggaa atcgccatcc cggtactggg cacctttgct 2040
ctggtttctt acattgcaaa caaggttctg actgtacaaa ccatcgacaa cgcgctgagc 2100
aaacgtaacg aaaaatggga tgaagtttac aaatatatcg tgaccaactg gctggctaag 2160
gttaatactc agatcgacct catccgcaaa aaaatgaaag aagcactgga aaaccaggcg 2220
gaagctacca aggcaatcat taactaccag tacaaccagt acaccgagga agaaaaaaac 2280
aacatcaact tcaacatcga cgatctgtcc tctaaactga acgaatccat caacaaagct 2340
atgatcaaca tcaacaagtt cctgaaccag tgctctgtaa gctatctgat gaactccatg 2400
atcccgtacg gtgttaaacg tctggaggac ttcgatgcgt ctctgaaaga cgccctgctg 2460
aaatacattt acgacaaccg tggcactctg atcggtcagg ttgatcgtct gaaggacaaa 2520
gtgaacaata ccttatcgac cgacatccct tttcagctca gtaaatatgt cgataaccaa 2580
cgccttttgt ccactctaga agcacaccat catcaccacc atcaccatca ccat 2634
<210> 4
<211> 878
<212> PRT
<213> 人工序列
<220>
<223> rLHN/A的多肽序列
<400> 4
Met Glu Phe Val Asn Lys Gln Phe Asn Tyr Lys Asp Pro Val Asn Gly
1 5 10 15
Val Asp Ile Ala Tyr Ile Lys Ile Pro Asn Ala Gly Gln Met Gln Pro
20 25 30
Val Lys Ala Phe Lys Ile His Asn Lys Ile Trp Val Ile Pro Glu Arg
35 40 45
Asp Thr Phe Thr Asn Pro Glu Glu Gly Asp Leu Asn Pro Pro Pro Glu
50 55 60
Ala Lys Gln Val Pro Val Ser Tyr Tyr Asp Ser Thr Tyr Leu Ser Thr
65 70 75 80
Asp Asn Glu Lys Asp Asn Tyr Leu Lys Gly Val Thr Lys Leu Phe Glu
85 90 95
Arg Ile Tyr Ser Thr Asp Leu Gly Arg Met Leu Leu Thr Ser Ile Val
100 105 110
Arg Gly Ile Pro Phe Trp Gly Gly Ser Thr Ile Asp Thr Glu Leu Lys
115 120 125
Val Ile Asp Thr Asn Cys Ile Asn Val Ile Gln Pro Asp Gly Ser Tyr
130 135 140
Arg Ser Glu Glu Leu Asn Leu Val Ile Ile Gly Pro Ser Ala Asp Ile
145 150 155 160
Ile Gln Phe Glu Cys Lys Ser Phe Gly His Glu Val Leu Asn Leu Thr
165 170 175
Arg Asn Gly Tyr Gly Ser Thr Gln Tyr Ile Arg Phe Ser Pro Asp Phe
180 185 190
Thr Phe Gly Phe Glu Glu Ser Leu Glu Val Asp Thr Asn Pro Leu Leu
195 200 205
Gly Ala Gly Lys Phe Ala Thr Asp Pro Ala Val Thr Leu Ala His Glu
210 215 220
Leu Ile His Ala Gly His Arg Leu Tyr Gly Ile Ala Ile Asn Pro Asn
225 230 235 240
Arg Val Phe Lys Val Asn Thr Asn Ala Tyr Tyr Glu Met Ser Gly Leu
245 250 255
Glu Val Ser Phe Glu Glu Leu Arg Thr Phe Gly Gly His Asp Ala Lys
260 265 270
Phe Ile Asp Ser Leu Gln Glu Asn Glu Phe Arg Leu Tyr Tyr Tyr Asn
275 280 285
Lys Phe Lys Asp Ile Ala Ser Thr Leu Asn Lys Ala Lys Ser Ile Val
290 295 300
Gly Thr Thr Ala Ser Leu Gln Tyr Met Lys Asn Val Phe Lys Glu Lys
305 310 315 320
Tyr Leu Leu Ser Glu Asp Thr Ser Gly Lys Phe Ser Val Asp Lys Leu
325 330 335
Lys Phe Asp Lys Leu Tyr Lys Met Leu Thr Glu Ile Tyr Thr Glu Asp
340 345 350
Asn Phe Val Lys Phe Phe Lys Val Leu Asn Arg Lys Thr Tyr Leu Asn
355 360 365
Phe Asp Lys Ala Val Phe Lys Ile Asn Ile Val Pro Lys Val Asn Tyr
370 375 380
Thr Ile Tyr Asp Gly Phe Asn Leu Arg Asn Thr Asn Leu Ala Ala Asn
385 390 395 400
Phe Asn Gly Gln Asn Thr Glu Ile Asn Asn Met Asn Phe Thr Lys Leu
405 410 415
Lys Asn Phe Thr Gly Leu Phe Glu Phe Tyr Lys Leu Leu Cys Val Asp
420 425 430
Gly Ile Ile Thr Ser Lys Thr Lys Ser Asp Asp Asp Asp Lys Asn Lys
435 440 445
Ala Leu Asn Leu Gln Cys Ile Lys Val Asn Asn Trp Asp Leu Phe Phe
450 455 460
Ser Pro Ser Glu Asp Asn Phe Thr Asn Asp Leu Asn Lys Gly Glu Glu
465 470 475 480
Ile Thr Ser Asp Thr Asn Ile Glu Ala Ala Glu Glu Asn Ile Ser Leu
485 490 495
Asp Leu Ile Gln Gln Tyr Tyr Leu Thr Phe Asn Phe Asp Asn Glu Pro
500 505 510
Glu Asn Ile Ser Ile Glu Asn Leu Ser Ser Asp Ile Ile Gly Gln Leu
515 520 525
Glu Leu Met Pro Asn Ile Glu Arg Phe Pro Asn Gly Lys Lys Tyr Glu
530 535 540
Leu Asp Lys Tyr Thr Met Phe His Tyr Leu Arg Ala Gln Glu Phe Glu
545 550 555 560
His Gly Lys Ser Arg Ile Ala Leu Thr Asn Ser Val Asn Glu Ala Leu
565 570 575
Leu Asn Pro Ser Arg Val Tyr Thr Phe Phe Ser Ser Asp Tyr Val Lys
580 585 590
Lys Val Asn Lys Ala Thr Glu Ala Ala Met Phe Leu Gly Trp Val Glu
595 600 605
Gln Leu Val Tyr Asp Phe Thr Asp Glu Thr Ser Glu Val Ser Thr Thr
610 615 620
Asp Lys Ile Ala Asp Ile Thr Ile Ile Ile Pro Tyr Ile Gly Pro Ala
625 630 635 640
Leu Asn Ile Gly Asn Met Leu Tyr Lys Asp Asp Phe Val Gly Ala Leu
645 650 655
Ile Phe Ser Gly Ala Val Ile Leu Leu Glu Phe Ile Pro Glu Ile Ala
660 665 670
Ile Pro Val Leu Gly Thr Phe Ala Leu Val Ser Tyr Ile Ala Asn Lys
675 680 685
Val Leu Thr Val Gln Thr Ile Asp Asn Ala Leu Ser Lys Arg Asn Glu
690 695 700
Lys Trp Asp Glu Val Tyr Lys Tyr Ile Val Thr Asn Trp Leu Ala Lys
705 710 715 720
Val Asn Thr Gln Ile Asp Leu Ile Arg Lys Lys Met Lys Glu Ala Leu
725 730 735
Glu Asn Gln Ala Glu Ala Thr Lys Ala Ile Ile Asn Tyr Gln Tyr Asn
740 745 750
Gln Tyr Thr Glu Glu Glu Lys Asn Asn Ile Asn Phe Asn Ile Asp Asp
755 760 765
Leu Ser Ser Lys Leu Asn Glu Ser Ile Asn Lys Ala Met Ile Asn Ile
770 775 780
Asn Lys Phe Leu Asn Gln Cys Ser Val Ser Tyr Leu Met Asn Ser Met
785 790 795 800
Ile Pro Tyr Gly Val Lys Arg Leu Glu Asp Phe Asp Ala Ser Leu Lys
805 810 815
Asp Ala Leu Leu Lys Tyr Ile Tyr Asp Asn Arg Gly Thr Leu Ile Gly
820 825 830
Gln Val Asp Arg Leu Lys Asp Lys Val Asn Asn Thr Leu Ser Thr Asp
835 840 845
Ile Pro Phe Gln Leu Ser Lys Tyr Val Asp Asn Gln Arg Leu Leu Ser
850 855 860
Thr Leu Glu Ala His His His His His His His His His His
865 870 875
<210> 5
<211> 1329
<212> DNA
<213> 人工序列
<220>
<223> rL/A的核苷酸序列
<400> 5
atgccattcg tcaacaagca attcaactac aaagacccag tcaacggcgt cgacatcgca 60
tacatcaaga ttccgaacgc cggtcaaatg cagccggtta aggcttttaa gatccacaac 120
aagatttggg ttatcccgga gcgtgacacc ttcacgaacc cggaagaagg cgatctgaac 180
ccgccaccgg aagcgaagca agtccctgtc agctactacg attcgacgta cctgagcacg 240
gataacgaaa aagataacta cctgaaaggt gtgaccaagc tgttcgaacg tatctacagc 300
acggatctgg gtcgcatgct gctgactagc attgttcgcg gtatcccgtt ctggggtggt 360
agcacgattg acaccgaact gaaggttatc gacactaact gcattaacgt tattcaaccg 420
gatggtagct atcgtagcga agagctgaat ctggtcatca ttggcccgag cgcagacatt 480
atccaattcg agtgcaagag ctttggtcac gaggttctga atctgacccg caatggctat 540
ggtagcaccc agtacattcg tttttcgccg gattttacct tcggctttga agagagcctg 600
gaggttgata ccaatccgtt gctgggtgcg ggcaaattcg ctaccgatcc ggctgtcacg 660
ctggcccatg aactgatcca cgcaggccac cgcctgtacg gcattgccat caacccaaac 720
cgtgtgttca aggttaatac gaatgcatac tacgagatga gcggcctgga agtcagcttc 780
gaagaactgc gcaccttcgg tggccatgac gctaaattca ttgacagctt gcaagagaat 840
gagttccgtc tgtactacta taacaaattc aaagacattg caagcacgtt gaacaaggcc 900
aaaagcatcg ttggtactac cgcgtcgttg cagtatatga agaatgtgtt taaagagaag 960
tacctgctgt ccgaggatac ctccggcaag tttagcgttg ataagctgaa gtttgacaaa 1020
ctgtacaaga tgctgaccga gatttacacc gaggacaact ttgtgaaatt cttcaaagtg 1080
ttgaatcgta aaacctatct gaattttgac aaagcggttt tcaagattaa catcgtgccg 1140
aaggtgaact acaccatcta tgacggtttt aacctgcgta acaccaacct ggcggcgaac 1200
tttaacggtc agaatacgga aatcaacaac atgaatttca cgaagttgaa gaacttcacg 1260
ggtctgttcg agttctataa gctgctgggt ctagaagcac accatcatca ccaccatcac 1320
catcaccat 1329
<210> 6
<211> 443
<212> PRT
<213> 人工序列
<220>
<223> rL/A的多肽序列
<400> 6
Met Pro Phe Val Asn Lys Gln Phe Asn Tyr Lys Asp Pro Val Asn Gly
1 5 10 15
Val Asp Ile Ala Tyr Ile Lys Ile Pro Asn Ala Gly Gln Met Gln Pro
20 25 30
Val Lys Ala Phe Lys Ile His Asn Lys Ile Trp Val Ile Pro Glu Arg
35 40 45
Asp Thr Phe Thr Asn Pro Glu Glu Gly Asp Leu Asn Pro Pro Pro Glu
50 55 60
Ala Lys Gln Val Pro Val Ser Tyr Tyr Asp Ser Thr Tyr Leu Ser Thr
65 70 75 80
Asp Asn Glu Lys Asp Asn Tyr Leu Lys Gly Val Thr Lys Leu Phe Glu
85 90 95
Arg Ile Tyr Ser Thr Asp Leu Gly Arg Met Leu Leu Thr Ser Ile Val
100 105 110
Arg Gly Ile Pro Phe Trp Gly Gly Ser Thr Ile Asp Thr Glu Leu Lys
115 120 125
Val Ile Asp Thr Asn Cys Ile Asn Val Ile Gln Pro Asp Gly Ser Tyr
130 135 140
Arg Ser Glu Glu Leu Asn Leu Val Ile Ile Gly Pro Ser Ala Asp Ile
145 150 155 160
Ile Gln Phe Glu Cys Lys Ser Phe Gly His Glu Val Leu Asn Leu Thr
165 170 175
Arg Asn Gly Tyr Gly Ser Thr Gln Tyr Ile Arg Phe Ser Pro Asp Phe
180 185 190
Thr Phe Gly Phe Glu Glu Ser Leu Glu Val Asp Thr Asn Pro Leu Leu
195 200 205
Gly Ala Gly Lys Phe Ala Thr Asp Pro Ala Val Thr Leu Ala His Glu
210 215 220
Leu Ile His Ala Gly His Arg Leu Tyr Gly Ile Ala Ile Asn Pro Asn
225 230 235 240
Arg Val Phe Lys Val Asn Thr Asn Ala Tyr Tyr Glu Met Ser Gly Leu
245 250 255
Glu Val Ser Phe Glu Glu Leu Arg Thr Phe Gly Gly His Asp Ala Lys
260 265 270
Phe Ile Asp Ser Leu Gln Glu Asn Glu Phe Arg Leu Tyr Tyr Tyr Asn
275 280 285
Lys Phe Lys Asp Ile Ala Ser Thr Leu Asn Lys Ala Lys Ser Ile Val
290 295 300
Gly Thr Thr Ala Ser Leu Gln Tyr Met Lys Asn Val Phe Lys Glu Lys
305 310 315 320
Tyr Leu Leu Ser Glu Asp Thr Ser Gly Lys Phe Ser Val Asp Lys Leu
325 330 335
Lys Phe Asp Lys Leu Tyr Lys Met Leu Thr Glu Ile Tyr Thr Glu Asp
340 345 350
Asn Phe Val Lys Phe Phe Lys Val Leu Asn Arg Lys Thr Tyr Leu Asn
355 360 365
Phe Asp Lys Ala Val Phe Lys Ile Asn Ile Val Pro Lys Val Asn Tyr
370 375 380
Thr Ile Tyr Asp Gly Phe Asn Leu Arg Asn Thr Asn Leu Ala Ala Asn
385 390 395 400
Phe Asn Gly Gln Asn Thr Glu Ile Asn Asn Met Asn Phe Thr Lys Leu
405 410 415
Lys Asn Phe Thr Gly Leu Phe Glu Phe Tyr Lys Leu Leu Gly Leu Glu
420 425 430
Ala His His His His His His His His His His
435 440
<210> 7
<211> 1299
<212> DNA
<213> 人工序列
<220>
<223> rHC/A的核苷酸序列
<400> 7
atgcatcatc accatcacca caaaaacatc atcaatacta gcattctgaa cctgcgttac 60
gagagcaatc atctgattga tctgagccgt tatgcaagca agatcaacat cggtagcaag 120
gtcaattttg acccgatcga taagaaccag atccagctgt ttaatctgga atcgagcaaa 180
attgaggtta tcctgaaaaa cgccattgtc tacaactcca tgtacgagaa tttctccacc 240
agcttctgga ttcgcatccc gaaatacttc aacagcatta gcctgaacaa cgagtatact 300
atcatcaact gtatggagaa caacagcggt tggaaggtgt ctctgaacta tggtgagatc 360
atttggacct tgcaggacac ccaagagatc aagcagcgcg tcgtgttcaa gtactctcaa 420
atgatcaaca tttccgatta cattaatcgt tggatcttcg tgaccattac gaataaccgt 480
ctgaataaca gcaagattta catcaatggt cgcttgatcg atcagaaacc gattagcaac 540
ctgggtaata tccacgcaag caacaacatt atgttcaaat tggacggttg ccgcgatacc 600
catcgttata tctggatcaa gtatttcaac ctgtttgata aagaactgaa tgagaaggag 660
atcaaagatt tgtatgacaa ccaatctaac agcggcattt tgaaggactt ctggggcgat 720
tatctgcaat acgataagcc gtactatatg ctgaacctgt atgatccgaa caaatatgtg 780
gatgtcaata atgtgggtat tcgtggttac atgtatttga agggtccgcg tggcagcgtt 840
atgacgacca acatttacct gaactctagc ctgtaccgtg gtacgaaatt catcattaag 900
aaatatgcca gcggcaacaa agataacatt gtgcgtaata acgatcgtgt ctacatcaac 960
gtggtcgtga agaataaaga gtaccgtctg gcgaccaacg cttcgcaggc gggtgttgag 1020
aaaattctga gcgcgttgga gatccctgat gtcggtaatc tgagccaagt cgtggttatg 1080
aagagcaaga acgaccaggg tatcactaac aagtgcaaga tgaacctgca agacaacaat 1140
ggtaacgaca tcggctttat tggtttccac cagttcaaca atattgctaa actggtagcg 1200
agcaattggt acaatcgtca gattgagcgc agcagccgta ctttgggctg tagctgggag 1260
tttatcccgg tcgatgatgg ttggggcgaa cgtccgctg 1299
<210> 8
<211> 433
<212> PRT
<213> 人工序列
<220>
<223> rHC/A的多肽序列
<400> 8
Met His His His His His His Lys Asn Ile Ile Asn Thr Ser Ile Leu
1 5 10 15
Asn Leu Arg Tyr Glu Ser Asn His Leu Ile Asp Leu Ser Arg Tyr Ala
20 25 30
Ser Lys Ile Asn Ile Gly Ser Lys Val Asn Phe Asp Pro Ile Asp Lys
35 40 45
Asn Gln Ile Gln Leu Phe Asn Leu Glu Ser Ser Lys Ile Glu Val Ile
50 55 60
Leu Lys Asn Ala Ile Val Tyr Asn Ser Met Tyr Glu Asn Phe Ser Thr
65 70 75 80
Ser Phe Trp Ile Arg Ile Pro Lys Tyr Phe Asn Ser Ile Ser Leu Asn
85 90 95
Asn Glu Tyr Thr Ile Ile Asn Cys Met Glu Asn Asn Ser Gly Trp Lys
100 105 110
Val Ser Leu Asn Tyr Gly Glu Ile Ile Trp Thr Leu Gln Asp Thr Gln
115 120 125
Glu Ile Lys Gln Arg Val Val Phe Lys Tyr Ser Gln Met Ile Asn Ile
130 135 140
Ser Asp Tyr Ile Asn Arg Trp Ile Phe Val Thr Ile Thr Asn Asn Arg
145 150 155 160
Leu Asn Asn Ser Lys Ile Tyr Ile Asn Gly Arg Leu Ile Asp Gln Lys
165 170 175
Pro Ile Ser Asn Leu Gly Asn Ile His Ala Ser Asn Asn Ile Met Phe
180 185 190
Lys Leu Asp Gly Cys Arg Asp Thr His Arg Tyr Ile Trp Ile Lys Tyr
195 200 205
Phe Asn Leu Phe Asp Lys Glu Leu Asn Glu Lys Glu Ile Lys Asp Leu
210 215 220
Tyr Asp Asn Gln Ser Asn Ser Gly Ile Leu Lys Asp Phe Trp Gly Asp
225 230 235 240
Tyr Leu Gln Tyr Asp Lys Pro Tyr Tyr Met Leu Asn Leu Tyr Asp Pro
245 250 255
Asn Lys Tyr Val Asp Val Asn Asn Val Gly Ile Arg Gly Tyr Met Tyr
260 265 270
Leu Lys Gly Pro Arg Gly Ser Val Met Thr Thr Asn Ile Tyr Leu Asn
275 280 285
Ser Ser Leu Tyr Arg Gly Thr Lys Phe Ile Ile Lys Lys Tyr Ala Ser
290 295 300
Gly Asn Lys Asp Asn Ile Val Arg Asn Asn Asp Arg Val Tyr Ile Asn
305 310 315 320
Val Val Val Lys Asn Lys Glu Tyr Arg Leu Ala Thr Asn Ala Ser Gln
325 330 335
Ala Gly Val Glu Lys Ile Leu Ser Ala Leu Glu Ile Pro Asp Val Gly
340 345 350
Asn Leu Ser Gln Val Val Val Met Lys Ser Lys Asn Asp Gln Gly Ile
355 360 365
Thr Asn Lys Cys Lys Met Asn Leu Gln Asp Asn Asn Gly Asn Asp Ile
370 375 380
Gly Phe Ile Gly Phe His Gln Phe Asn Asn Ile Ala Lys Leu Val Ala
385 390 395 400
Ser Asn Trp Tyr Asn Arg Gln Ile Glu Arg Ser Ser Arg Thr Leu Gly
405 410 415
Cys Ser Trp Glu Phe Ile Pro Val Asp Asp Gly Trp Gly Glu Arg Pro
420 425 430
Leu
<210> 9
<211> 3873
<212> DNA
<213> 人工序列
<220>
<223> rBoNT/B(0)的核苷酸序列
<400> 9
atgccggtga cgattaacaa cttcaactac aacgacccga ttgacaacaa caacattatc 60
atgatggaac cgccgtttgc acgcggcacg ggccgttatt acaaagcgtt taaaatcacc 120
gatcgtattt ggattatccc ggaacgctac acgtttggtt ataaaccgga agacttcaac 180
aaaagctctg gcatcttcaa ccgtgatgtt tgcgaatact acgatccgga ctacctgaac 240
accaacgata agaaaaacat ttttctgcaa acgatgatca aactgttcaa tcgcattaaa 300
agcaaaccgc tgggtgaaaa actgctggaa atgattatca atggcattcc gtatctgggt 360
gatcgtcgcg tgccgctgga agaatttaac accaatatcg cgagtgttac ggtcaacaaa 420
ctgatttcca atccgggtga agtcgaacgt aaaaaaggca tcttcgccaa cctgatcatc 480
ttcggcccgg gtccggtgct gaacgaaaat gaaaccattg atatcggtat tcagaaccat 540
tttgcctcac gcgaaggctt cggcggtatt atgcaaatga aattttgccc ggaatatgtg 600
tcggttttca acaatgttca ggaaaacaaa ggtgcaagca tctttaatcg tcgcggctat 660
ttctctgatc cggctctgat cctgatgcac caactgattt atgtgctgca cggcctgtat 720
ggtatcaaag tggatgacct gccgatcgtt ccgaacgaga aaaaattttt catgcagagc 780
accgacgcaa ttcaagctga agaactgtat acgtttggcg gtcaggaccc gtctattatc 840
accccgagca ccgacaaaag catctacgat aaagtgctgc aaaactttcg tggcattgtt 900
gaccgcctga ataaagtcct ggtgtgtatc tctgatccga acatcaacat caacatctac 960
aaaaacaaat tcaaagacaa atacaaattc gttgaagatt ctgaaggcaa atatagtatt 1020
gacgtcgaat cctttgataa actgtacaaa agtctgatgt tcggtttcac cgaaacgaac 1080
atcgcggaaa actacaaaat caaaacccgc gcctcctatt tcagcgactc tctgccgccg 1140
gttaaaatca aaaatctgct ggataacgaa atttatacga tcgaagaagg tttcaacatc 1200
agcgataaag acatggaaaa agaataccgt ggccagaata aagcaatcaa caaacaggcg 1260
tatgaagaaa ttagtaaaga acatctggcg gtctacaaaa ttcagatgtg caaatccgtg 1320
aaagccccgg gtatttgtat cgatgttgac aatgaagacc tgtttttcat cgccgataaa 1380
aacagttttt ccgatgacct gtcaaaaaat gaacgcatcg aatacaacac ccaatcgaac 1440
tacatcgaaa acgatttccc gatcaacgaa ctgattctgg atacggacct gattagtaaa 1500
atcgaactgc cgtcagaaaa caccgaatcg ctgacggact ttaatgttga tgtcccggtg 1560
tatgaaaaac agccggcaat taagaaaatt tttaccgatg aaaacacgat cttccagtac 1620
ctgtacagcc aaacctttcc gctggacatt cgcgatatct ctctgacgag ttcctttgat 1680
gacgcactgc tgttcagcaa caaagtgtac tcctttttct caatggatta catcaaaacc 1740
gctaacaaag tggttgaagc gggcctgttt gccggttggg tgaaacagat cgttaacgat 1800
ttcgtcatcg aagccaacaa aagtaacacg atggataaaa ttgctgatat ctccctgatt 1860
gtcccgtata ttggcctggc actgaatgtg ggtaacgaaa cggcgaaagg caattttgaa 1920
aacgccttcg aaattgcagg cgcttcaatc ctgctggaat ttattccgga actgctgatc 1980
ccggtcgtgg gtgcgttcct gctggaatct tacatcgaca acaaaaacaa aatcatcaaa 2040
accattgata acgcgctgac gaaacgtaac gaaaaatggt cagatatgta cggcctgatt 2100
gttgcccagt ggctgagcac cgtcaacacg caattttaca ccatcaaaga aggtatgtac 2160
aaagcgctga attatcaggc gcaagccctg gaagaaatca tcaaataccg ctacaacatc 2220
tacagcgaaa aagaaaaatc taacatcaac atcgacttta atgatatcaa cagcaaactg 2280
aacgaaggta tcaaccaggc aatcgataac atcaacaact tcatcaacgg ctgctcagtg 2340
tcgtatctga tgaagaaaat gatcccgctg gctgttgaaa aactgctgga ttttgacaac 2400
accctgaaga aaaacctgct gaactacatc gatgaaaaca aactgtacct gatcggctca 2460
gccgaatacg aaaaatcgaa agtgaacaaa tacctgaaaa ccatcatgcc gtttgacctg 2520
agtatttaca ccaacgatac gatcctgatc gaaatgttca acaaatacaa ctccgaaatt 2580
ctgaacaata ttatcctgaa cctgcgttac aaagacaaca atctgatcga tctgagcggc 2640
tatggtgcaa aagttgaagt ctacgacggt gtcgaactga acgataaaaa ccagttcaaa 2700
ctgacctcat cggctaactc aaaaattcgt gtgacgcaga accaaaacat catcttcaac 2760
tcggtctttc tggacttcag cgtgtctttc tggattcgca tcccgaaata taaaaatgat 2820
ggcatccaga actacatcca taacgaatac accatcatca actgtatgaa aaacaacagt 2880
ggttggaaaa tttccatccg tggcaaccgc attatctgga ccctgattga tatcaatggt 2940
aaaacgaaaa gcgtgttttt cgaatacaac atccgtgaag atatctctga atacatcaat 3000
cgctggtttt tcgtgaccat tacgaacaat ctgaacaatg cgaaaatcta tatcaacggc 3060
aaactggaaa gtaataccga catcaaagat attcgtgaag ttatcgccaa cggtgaaatc 3120
atcttcaaac tggatggcga catcgatcgc acccagttca tttggatgaa atacttctcc 3180
atcttcaaca cggaactgag tcagtccaat atcgaagaac gctacaaaat ccaatcatac 3240
tcggaatacc tgaaagattt ctggggtaac ccgctgatgt acaacaaaga atactacatg 3300
ttcaacgcgg gcaacaaaaa ctcatacatc aaactgaaaa aagattcgcc ggtgggtgaa 3360
atcctgaccc gtagcaaata caaccagaac tctaaataca tcaactatcg cgatctgtac 3420
attggcgaaa aatttattat ccgtcgcaaa agcaactctc agagtattaa tgatgacatc 3480
gtgcgtaaag aagactacat ctatctggat ttctttaatc tgaaccaaga atggcgcgtt 3540
tatacctaca aatacttcaa aaaagaagaa gagaaactgt tcctggcccc gattagcgac 3600
agcgatgaat tttacaacac catccagatc aaagaatacg atgaacagcc gacgtatagt 3660
tgccaactgc tgttcaaaaa agacgaagaa tccaccgatg aaattggcct gattggtatc 3720
caccgtttct atgaaagcgg tatcgttttc gaagaataca aagattactt ctgtatctct 3780
aaatggtatc tgaaagaagt caaacgcaaa ccgtacaacc tgaaactggg ctgcaactgg 3840
caatttatcc cgaaagacga aggctggacc gaa 3873
<210> 10
<211> 1291
<212> PRT
<213> 人工序列
<220>
<223> rBoNT/B(0)的多肽序列
<400> 10
Met Pro Val Thr Ile Asn Asn Phe Asn Tyr Asn Asp Pro Ile Asp Asn
1 5 10 15
Asn Asn Ile Ile Met Met Glu Pro Pro Phe Ala Arg Gly Thr Gly Arg
20 25 30
Tyr Tyr Lys Ala Phe Lys Ile Thr Asp Arg Ile Trp Ile Ile Pro Glu
35 40 45
Arg Tyr Thr Phe Gly Tyr Lys Pro Glu Asp Phe Asn Lys Ser Ser Gly
50 55 60
Ile Phe Asn Arg Asp Val Cys Glu Tyr Tyr Asp Pro Asp Tyr Leu Asn
65 70 75 80
Thr Asn Asp Lys Lys Asn Ile Phe Leu Gln Thr Met Ile Lys Leu Phe
85 90 95
Asn Arg Ile Lys Ser Lys Pro Leu Gly Glu Lys Leu Leu Glu Met Ile
100 105 110
Ile Asn Gly Ile Pro Tyr Leu Gly Asp Arg Arg Val Pro Leu Glu Glu
115 120 125
Phe Asn Thr Asn Ile Ala Ser Val Thr Val Asn Lys Leu Ile Ser Asn
130 135 140
Pro Gly Glu Val Glu Arg Lys Lys Gly Ile Phe Ala Asn Leu Ile Ile
145 150 155 160
Phe Gly Pro Gly Pro Val Leu Asn Glu Asn Glu Thr Ile Asp Ile Gly
165 170 175
Ile Gln Asn His Phe Ala Ser Arg Glu Gly Phe Gly Gly Ile Met Gln
180 185 190
Met Lys Phe Cys Pro Glu Tyr Val Ser Val Phe Asn Asn Val Gln Glu
195 200 205
Asn Lys Gly Ala Ser Ile Phe Asn Arg Arg Gly Tyr Phe Ser Asp Pro
210 215 220
Ala Leu Ile Leu Met His Gln Leu Ile Tyr Val Leu His Gly Leu Tyr
225 230 235 240
Gly Ile Lys Val Asp Asp Leu Pro Ile Val Pro Asn Glu Lys Lys Phe
245 250 255
Phe Met Gln Ser Thr Asp Ala Ile Gln Ala Glu Glu Leu Tyr Thr Phe
260 265 270
Gly Gly Gln Asp Pro Ser Ile Ile Thr Pro Ser Thr Asp Lys Ser Ile
275 280 285
Tyr Asp Lys Val Leu Gln Asn Phe Arg Gly Ile Val Asp Arg Leu Asn
290 295 300
Lys Val Leu Val Cys Ile Ser Asp Pro Asn Ile Asn Ile Asn Ile Tyr
305 310 315 320
Lys Asn Lys Phe Lys Asp Lys Tyr Lys Phe Val Glu Asp Ser Glu Gly
325 330 335
Lys Tyr Ser Ile Asp Val Glu Ser Phe Asp Lys Leu Tyr Lys Ser Leu
340 345 350
Met Phe Gly Phe Thr Glu Thr Asn Ile Ala Glu Asn Tyr Lys Ile Lys
355 360 365
Thr Arg Ala Ser Tyr Phe Ser Asp Ser Leu Pro Pro Val Lys Ile Lys
370 375 380
Asn Leu Leu Asp Asn Glu Ile Tyr Thr Ile Glu Glu Gly Phe Asn Ile
385 390 395 400
Ser Asp Lys Asp Met Glu Lys Glu Tyr Arg Gly Gln Asn Lys Ala Ile
405 410 415
Asn Lys Gln Ala Tyr Glu Glu Ile Ser Lys Glu His Leu Ala Val Tyr
420 425 430
Lys Ile Gln Met Cys Lys Ser Val Lys Ala Pro Gly Ile Cys Ile Asp
435 440 445
Val Asp Asn Glu Asp Leu Phe Phe Ile Ala Asp Lys Asn Ser Phe Ser
450 455 460
Asp Asp Leu Ser Lys Asn Glu Arg Ile Glu Tyr Asn Thr Gln Ser Asn
465 470 475 480
Tyr Ile Glu Asn Asp Phe Pro Ile Asn Glu Leu Ile Leu Asp Thr Asp
485 490 495
Leu Ile Ser Lys Ile Glu Leu Pro Ser Glu Asn Thr Glu Ser Leu Thr
500 505 510
Asp Phe Asn Val Asp Val Pro Val Tyr Glu Lys Gln Pro Ala Ile Lys
515 520 525
Lys Ile Phe Thr Asp Glu Asn Thr Ile Phe Gln Tyr Leu Tyr Ser Gln
530 535 540
Thr Phe Pro Leu Asp Ile Arg Asp Ile Ser Leu Thr Ser Ser Phe Asp
545 550 555 560
Asp Ala Leu Leu Phe Ser Asn Lys Val Tyr Ser Phe Phe Ser Met Asp
565 570 575
Tyr Ile Lys Thr Ala Asn Lys Val Val Glu Ala Gly Leu Phe Ala Gly
580 585 590
Trp Val Lys Gln Ile Val Asn Asp Phe Val Ile Glu Ala Asn Lys Ser
595 600 605
Asn Thr Met Asp Lys Ile Ala Asp Ile Ser Leu Ile Val Pro Tyr Ile
610 615 620
Gly Leu Ala Leu Asn Val Gly Asn Glu Thr Ala Lys Gly Asn Phe Glu
625 630 635 640
Asn Ala Phe Glu Ile Ala Gly Ala Ser Ile Leu Leu Glu Phe Ile Pro
645 650 655
Glu Leu Leu Ile Pro Val Val Gly Ala Phe Leu Leu Glu Ser Tyr Ile
660 665 670
Asp Asn Lys Asn Lys Ile Ile Lys Thr Ile Asp Asn Ala Leu Thr Lys
675 680 685
Arg Asn Glu Lys Trp Ser Asp Met Tyr Gly Leu Ile Val Ala Gln Trp
690 695 700
Leu Ser Thr Val Asn Thr Gln Phe Tyr Thr Ile Lys Glu Gly Met Tyr
705 710 715 720
Lys Ala Leu Asn Tyr Gln Ala Gln Ala Leu Glu Glu Ile Ile Lys Tyr
725 730 735
Arg Tyr Asn Ile Tyr Ser Glu Lys Glu Lys Ser Asn Ile Asn Ile Asp
740 745 750
Phe Asn Asp Ile Asn Ser Lys Leu Asn Glu Gly Ile Asn Gln Ala Ile
755 760 765
Asp Asn Ile Asn Asn Phe Ile Asn Gly Cys Ser Val Ser Tyr Leu Met
770 775 780
Lys Lys Met Ile Pro Leu Ala Val Glu Lys Leu Leu Asp Phe Asp Asn
785 790 795 800
Thr Leu Lys Lys Asn Leu Leu Asn Tyr Ile Asp Glu Asn Lys Leu Tyr
805 810 815
Leu Ile Gly Ser Ala Glu Tyr Glu Lys Ser Lys Val Asn Lys Tyr Leu
820 825 830
Lys Thr Ile Met Pro Phe Asp Leu Ser Ile Tyr Thr Asn Asp Thr Ile
835 840 845
Leu Ile Glu Met Phe Asn Lys Tyr Asn Ser Glu Ile Leu Asn Asn Ile
850 855 860
Ile Leu Asn Leu Arg Tyr Lys Asp Asn Asn Leu Ile Asp Leu Ser Gly
865 870 875 880
Tyr Gly Ala Lys Val Glu Val Tyr Asp Gly Val Glu Leu Asn Asp Lys
885 890 895
Asn Gln Phe Lys Leu Thr Ser Ser Ala Asn Ser Lys Ile Arg Val Thr
900 905 910
Gln Asn Gln Asn Ile Ile Phe Asn Ser Val Phe Leu Asp Phe Ser Val
915 920 925
Ser Phe Trp Ile Arg Ile Pro Lys Tyr Lys Asn Asp Gly Ile Gln Asn
930 935 940
Tyr Ile His Asn Glu Tyr Thr Ile Ile Asn Cys Met Lys Asn Asn Ser
945 950 955 960
Gly Trp Lys Ile Ser Ile Arg Gly Asn Arg Ile Ile Trp Thr Leu Ile
965 970 975
Asp Ile Asn Gly Lys Thr Lys Ser Val Phe Phe Glu Tyr Asn Ile Arg
980 985 990
Glu Asp Ile Ser Glu Tyr Ile Asn Arg Trp Phe Phe Val Thr Ile Thr
995 1000 1005
Asn Asn Leu Asn Asn Ala Lys Ile Tyr Ile Asn Gly Lys Leu Glu
1010 1015 1020
Ser Asn Thr Asp Ile Lys Asp Ile Arg Glu Val Ile Ala Asn Gly
1025 1030 1035
Glu Ile Ile Phe Lys Leu Asp Gly Asp Ile Asp Arg Thr Gln Phe
1040 1045 1050
Ile Trp Met Lys Tyr Phe Ser Ile Phe Asn Thr Glu Leu Ser Gln
1055 1060 1065
Ser Asn Ile Glu Glu Arg Tyr Lys Ile Gln Ser Tyr Ser Glu Tyr
1070 1075 1080
Leu Lys Asp Phe Trp Gly Asn Pro Leu Met Tyr Asn Lys Glu Tyr
1085 1090 1095
Tyr Met Phe Asn Ala Gly Asn Lys Asn Ser Tyr Ile Lys Leu Lys
1100 1105 1110
Lys Asp Ser Pro Val Gly Glu Ile Leu Thr Arg Ser Lys Tyr Asn
1115 1120 1125
Gln Asn Ser Lys Tyr Ile Asn Tyr Arg Asp Leu Tyr Ile Gly Glu
1130 1135 1140
Lys Phe Ile Ile Arg Arg Lys Ser Asn Ser Gln Ser Ile Asn Asp
1145 1150 1155
Asp Ile Val Arg Lys Glu Asp Tyr Ile Tyr Leu Asp Phe Phe Asn
1160 1165 1170
Leu Asn Gln Glu Trp Arg Val Tyr Thr Tyr Lys Tyr Phe Lys Lys
1175 1180 1185
Glu Glu Glu Lys Leu Phe Leu Ala Pro Ile Ser Asp Ser Asp Glu
1190 1195 1200
Phe Tyr Asn Thr Ile Gln Ile Lys Glu Tyr Asp Glu Gln Pro Thr
1205 1210 1215
Tyr Ser Cys Gln Leu Leu Phe Lys Lys Asp Glu Glu Ser Thr Asp
1220 1225 1230
Glu Ile Gly Leu Ile Gly Ile His Arg Phe Tyr Glu Ser Gly Ile
1235 1240 1245
Val Phe Glu Glu Tyr Lys Asp Tyr Phe Cys Ile Ser Lys Trp Tyr
1250 1255 1260
Leu Lys Glu Val Lys Arg Lys Pro Tyr Asn Leu Lys Leu Gly Cys
1265 1270 1275
Asn Trp Gln Phe Ile Pro Lys Asp Glu Gly Trp Thr Glu
1280 1285 1290
<210> 11
<211> 3873
<212> DNA
<213> 人工序列
<220>
<223> rBoNT/C(0)的核苷酸序列
<400> 11
atgccgatca cgattaataa tttcaactat agcgatccgg tggacaataa gaatattctg 60
tatctggata ctcatctgaa tacgctggct aacgaaccgg agaaagcgtt ccgcatcaca 120
ggcaacatct gggttattcc cgatcgcttt tcacgcaaca gcaaccctaa tctgaacaaa 180
cctcctcgtg tcaccagtcc taaatccggt tattacgacc caaactatct gagtacggat 240
agcgataaag atccctttct gaaagagatc attaagctgt tcaaacgcat taactctcgc 300
gaaattgggg aagagctgat ctatcggctt tcgacagata tcccgttccc aggtaacaat 360
aataccccga ttaatacttt cgactttgat gttgatttca attctgtgga tgtgaaaacg 420
cgtcaaggca ataattgggt gaaaactggt agcattaacc cgagtgtaat tatcacaggt 480
ccccgtgaga acatcatcga cccggaaacc tctaccttca agctgacgaa caacacgttt 540
gctgcacagg aagggtttgg tgccctgtca atcatttcca tctcaccgcg tttcatgtta 600
acctactcca atgccacaaa tgatgttggc gaaggacgtt ttagcaaatc agaattttgc 660
atggacccaa ttctcattct gatgggcacg ctgaacaatg cgatgcacaa cttgtatggc 720
attgctattc caaacgatca aaccattagc tccgttacca gtaatatctt ctatagccag 780
tataatgtca aattggagta tgccgaaatt tacgcctttg gaggcccgac cattgacctg 840
attccgaaat ctgcacgcaa atacttcgaa gaaaaggcgt tagattacta tcgcagcatc 900
gcgaaacgcc tgaactcgat taccacggcc aatccgtcgt cgttcaacaa atacattggt 960
gaatataaac agaaactgat tcgcaaatat cggtttgtcg tagaaagctc tggtgaagtg 1020
actgtaaacc gcaacaaatt tgtcgaactc tacaacgagt tgacccaaat ctttaccgag 1080
tttaactacg caaagatcta taacgtacag aaccgcaaga tttatcttag caatgtatac 1140
acaccggtta ctgcgaacat cttagacgac aatgtgtatg atattcagaa tggctttaac 1200
atcccgaaat caaatctgaa cgttctgttt atgggccaga acctgagtcg taatccagca 1260
ctgcgtaaag tgaacccgga aaatatgctc tacttgttta ccaaattttg ccacaaagcg 1320
attgatggcc gctctctcta taacaaaacg ctggattgtc gtgagttact tgtgaagaac 1380
actgatttac cgttcattgg ggatatctcc gacgtgaaaa ccgatatctt cctgcgcaaa 1440
gacattaatg aagaaacgga agtcatctat taccccgaca atgtgagcgt tgatcaggtc 1500
attttatcga agaacacctc cgaacatggt cagttggatt tgctgtaccc tagcattgac 1560
tcggagagtg aaatccttcc gggcgaaaat caagtgtttt acgacaaccg tacccaaaat 1620
gttgattatt tgaattctta ttactacctg gaatctcaga aattgagcga caatgtggaa 1680
gatttcacgt tcacacgctc cattgaggaa gcgctggata atagcgcgaa agtgtatacg 1740
tatttcccta ccttggcgaa taaagtaaat gctggtgtcc agggaggctt atttctgatg 1800
tgggcgaatg atgtggtaga agattttacg accaatattt tgcgtaagga caccttagat 1860
aaaattagcg atgttagcgc catcatcccc tatattggcc cagcactgaa tatctcgaac 1920
tctgtgcgtc gcggaaactt caccgaagca tttgcggtga ccggggttac tattctgttg 1980
gaagcctttc cggagtttac tattccggcg ctgggtgcgt ttgtgattta ttcgaaagta 2040
caagaacgca atgaaattat caaaaccatc gataattgcc tggaacaacg cattaaacgc 2100
tggaaggatt cttatgaatg gatgatgggc acctggttat cccgtattat cacacagttt 2160
aacaacatct cgtatcagat gtacgattca ctgaactacc aagcaggggc gatcaaagcc 2220
aagatcgact tagaatacaa gaaatattca ggtagcgata aagagaatat taaaagccag 2280
gttgaaaacc tgaagaactc tctggatgtc aaaatttcag aggctatgaa caacattaac 2340
aaatttatcc gcgaatgtag cgtcacgtat ctgtttaaaa acatgctccc gaaagtgatt 2400
gatgagctca acgagtttga tcgcaacaca aaggccaaac tgattaacct gattgatagt 2460
cacaatatta ttttagtcgg tgaagttgac aagctgaagg ctaaggtcaa taacagcttt 2520
cagaacacta ttccgtttaa tattttctcc tatacgaaca atagtctgct gaaagacatt 2580
atcaacgaat acttcaacaa tattaatgac agcaaaattc tgagcctgca gaatcgtaag 2640
aatacgctgg tagataccag tggatataat gcggaagtct cagaagaggg tgatgtacag 2700
ctgaacccga tctttccgtt cgactttaaa ctggggtcta gtggtgaaga tcgcggtaaa 2760
gtgatcgtta cccaaaacga gaacattgtg tataacagca tgtacgagag tttctcaatt 2820
tctttctgga ttcgcatcaa taaatgggtt tctaatttgc ctggctatac catcattgat 2880
agcgtcaaaa acaactcggg ctggtcgatt ggcattatta gcaactttct ggtgtttacc 2940
ctgaaacaga atgaggattc ggaacagagc attaacttct cctacgacat cagcaacaat 3000
gcaccagggt ataacaaatg gttcttcgta acggtgacga acaatatgat gggcaatatg 3060
aaaatctaca ttaacgggaa acttatcgac accattaaag tgaaagagct tactgggatc 3120
aattttagta aaaccattac ctttgagatc aacaaaattc cggacacggg tctgattacc 3180
tccgattcgg ataatatcaa tatgtggatt cgcgactttt atatcttcgc caaagaactt 3240
gatggcaaag atatcaacat tttgtttaat tccctgcagt ataccaatgt cgttaaggac 3300
tattggggca atgatctccg ctacaataaa gaatactaca tggttaacat cgactatctc 3360
aatcgctaca tgtatgctaa ctcgcgtcaa attgtgttta acacacgtcg taacaacaac 3420
gattttaacg aaggttataa aatcattatc aaacggatcc gcggcaatac gaacgatact 3480
cgtgttcgtg gcggtgacat tctgtatttc gacatgacga ttaataataa agcgtacaat 3540
ctgttcatga agaacgaaac catgtacgcc gataaccatt ccactgaaga tatctacgca 3600
atcggacttc gcgaacagac caaagacatt aacgacaaca tcatctttca gattcaaccg 3660
atgaataata cctactacta tgcctcccag atcttcaaaa gtaatttcaa cggcgaaaac 3720
atttcaggca tttgctcaat cggcacttat cggttccggt taggtggtga ttggtatcgt 3780
cacaactacc ttgttcccac agtgaaacaa ggcaactatg catcgctctt agaaagcaca 3840
tctacgcatt ggggttttgt gccagtcagt gaa 3873
<210> 12
<211> 1291
<212> PRT
<213> 人工序列
<220>
<223> rBoNT/C(0)的多肽序列
<400> 12
Met Pro Ile Thr Ile Asn Asn Phe Asn Tyr Ser Asp Pro Val Asp Asn
1 5 10 15
Lys Asn Ile Leu Tyr Leu Asp Thr His Leu Asn Thr Leu Ala Asn Glu
20 25 30
Pro Glu Lys Ala Phe Arg Ile Thr Gly Asn Ile Trp Val Ile Pro Asp
35 40 45
Arg Phe Ser Arg Asn Ser Asn Pro Asn Leu Asn Lys Pro Pro Arg Val
50 55 60
Thr Ser Pro Lys Ser Gly Tyr Tyr Asp Pro Asn Tyr Leu Ser Thr Asp
65 70 75 80
Ser Asp Lys Asp Pro Phe Leu Lys Glu Ile Ile Lys Leu Phe Lys Arg
85 90 95
Ile Asn Ser Arg Glu Ile Gly Glu Glu Leu Ile Tyr Arg Leu Ser Thr
100 105 110
Asp Ile Pro Phe Pro Gly Asn Asn Asn Thr Pro Ile Asn Thr Phe Asp
115 120 125
Phe Asp Val Asp Phe Asn Ser Val Asp Val Lys Thr Arg Gln Gly Asn
130 135 140
Asn Trp Val Lys Thr Gly Ser Ile Asn Pro Ser Val Ile Ile Thr Gly
145 150 155 160
Pro Arg Glu Asn Ile Ile Asp Pro Glu Thr Ser Thr Phe Lys Leu Thr
165 170 175
Asn Asn Thr Phe Ala Ala Gln Glu Gly Phe Gly Ala Leu Ser Ile Ile
180 185 190
Ser Ile Ser Pro Arg Phe Met Leu Thr Tyr Ser Asn Ala Thr Asn Asp
195 200 205
Val Gly Glu Gly Arg Phe Ser Lys Ser Glu Phe Cys Met Asp Pro Ile
210 215 220
Leu Ile Leu Met Gly Thr Leu Asn Asn Ala Met His Asn Leu Tyr Gly
225 230 235 240
Ile Ala Ile Pro Asn Asp Gln Thr Ile Ser Ser Val Thr Ser Asn Ile
245 250 255
Phe Tyr Ser Gln Tyr Asn Val Lys Leu Glu Tyr Ala Glu Ile Tyr Ala
260 265 270
Phe Gly Gly Pro Thr Ile Asp Leu Ile Pro Lys Ser Ala Arg Lys Tyr
275 280 285
Phe Glu Glu Lys Ala Leu Asp Tyr Tyr Arg Ser Ile Ala Lys Arg Leu
290 295 300
Asn Ser Ile Thr Thr Ala Asn Pro Ser Ser Phe Asn Lys Tyr Ile Gly
305 310 315 320
Glu Tyr Lys Gln Lys Leu Ile Arg Lys Tyr Arg Phe Val Val Glu Ser
325 330 335
Ser Gly Glu Val Thr Val Asn Arg Asn Lys Phe Val Glu Leu Tyr Asn
340 345 350
Glu Leu Thr Gln Ile Phe Thr Glu Phe Asn Tyr Ala Lys Ile Tyr Asn
355 360 365
Val Gln Asn Arg Lys Ile Tyr Leu Ser Asn Val Tyr Thr Pro Val Thr
370 375 380
Ala Asn Ile Leu Asp Asp Asn Val Tyr Asp Ile Gln Asn Gly Phe Asn
385 390 395 400
Ile Pro Lys Ser Asn Leu Asn Val Leu Phe Met Gly Gln Asn Leu Ser
405 410 415
Arg Asn Pro Ala Leu Arg Lys Val Asn Pro Glu Asn Met Leu Tyr Leu
420 425 430
Phe Thr Lys Phe Cys His Lys Ala Ile Asp Gly Arg Ser Leu Tyr Asn
435 440 445
Lys Thr Leu Asp Cys Arg Glu Leu Leu Val Lys Asn Thr Asp Leu Pro
450 455 460
Phe Ile Gly Asp Ile Ser Asp Val Lys Thr Asp Ile Phe Leu Arg Lys
465 470 475 480
Asp Ile Asn Glu Glu Thr Glu Val Ile Tyr Tyr Pro Asp Asn Val Ser
485 490 495
Val Asp Gln Val Ile Leu Ser Lys Asn Thr Ser Glu His Gly Gln Leu
500 505 510
Asp Leu Leu Tyr Pro Ser Ile Asp Ser Glu Ser Glu Ile Leu Pro Gly
515 520 525
Glu Asn Gln Val Phe Tyr Asp Asn Arg Thr Gln Asn Val Asp Tyr Leu
530 535 540
Asn Ser Tyr Tyr Tyr Leu Glu Ser Gln Lys Leu Ser Asp Asn Val Glu
545 550 555 560
Asp Phe Thr Phe Thr Arg Ser Ile Glu Glu Ala Leu Asp Asn Ser Ala
565 570 575
Lys Val Tyr Thr Tyr Phe Pro Thr Leu Ala Asn Lys Val Asn Ala Gly
580 585 590
Val Gln Gly Gly Leu Phe Leu Met Trp Ala Asn Asp Val Val Glu Asp
595 600 605
Phe Thr Thr Asn Ile Leu Arg Lys Asp Thr Leu Asp Lys Ile Ser Asp
610 615 620
Val Ser Ala Ile Ile Pro Tyr Ile Gly Pro Ala Leu Asn Ile Ser Asn
625 630 635 640
Ser Val Arg Arg Gly Asn Phe Thr Glu Ala Phe Ala Val Thr Gly Val
645 650 655
Thr Ile Leu Leu Glu Ala Phe Pro Glu Phe Thr Ile Pro Ala Leu Gly
660 665 670
Ala Phe Val Ile Tyr Ser Lys Val Gln Glu Arg Asn Glu Ile Ile Lys
675 680 685
Thr Ile Asp Asn Cys Leu Glu Gln Arg Ile Lys Arg Trp Lys Asp Ser
690 695 700
Tyr Glu Trp Met Met Gly Thr Trp Leu Ser Arg Ile Ile Thr Gln Phe
705 710 715 720
Asn Asn Ile Ser Tyr Gln Met Tyr Asp Ser Leu Asn Tyr Gln Ala Gly
725 730 735
Ala Ile Lys Ala Lys Ile Asp Leu Glu Tyr Lys Lys Tyr Ser Gly Ser
740 745 750
Asp Lys Glu Asn Ile Lys Ser Gln Val Glu Asn Leu Lys Asn Ser Leu
755 760 765
Asp Val Lys Ile Ser Glu Ala Met Asn Asn Ile Asn Lys Phe Ile Arg
770 775 780
Glu Cys Ser Val Thr Tyr Leu Phe Lys Asn Met Leu Pro Lys Val Ile
785 790 795 800
Asp Glu Leu Asn Glu Phe Asp Arg Asn Thr Lys Ala Lys Leu Ile Asn
805 810 815
Leu Ile Asp Ser His Asn Ile Ile Leu Val Gly Glu Val Asp Lys Leu
820 825 830
Lys Ala Lys Val Asn Asn Ser Phe Gln Asn Thr Ile Pro Phe Asn Ile
835 840 845
Phe Ser Tyr Thr Asn Asn Ser Leu Leu Lys Asp Ile Ile Asn Glu Tyr
850 855 860
Phe Asn Asn Ile Asn Asp Ser Lys Ile Leu Ser Leu Gln Asn Arg Lys
865 870 875 880
Asn Thr Leu Val Asp Thr Ser Gly Tyr Asn Ala Glu Val Ser Glu Glu
885 890 895
Gly Asp Val Gln Leu Asn Pro Ile Phe Pro Phe Asp Phe Lys Leu Gly
900 905 910
Ser Ser Gly Glu Asp Arg Gly Lys Val Ile Val Thr Gln Asn Glu Asn
915 920 925
Ile Val Tyr Asn Ser Met Tyr Glu Ser Phe Ser Ile Ser Phe Trp Ile
930 935 940
Arg Ile Asn Lys Trp Val Ser Asn Leu Pro Gly Tyr Thr Ile Ile Asp
945 950 955 960
Ser Val Lys Asn Asn Ser Gly Trp Ser Ile Gly Ile Ile Ser Asn Phe
965 970 975
Leu Val Phe Thr Leu Lys Gln Asn Glu Asp Ser Glu Gln Ser Ile Asn
980 985 990
Phe Ser Tyr Asp Ile Ser Asn Asn Ala Pro Gly Tyr Asn Lys Trp Phe
995 1000 1005
Phe Val Thr Val Thr Asn Asn Met Met Gly Asn Met Lys Ile Tyr
1010 1015 1020
Ile Asn Gly Lys Leu Ile Asp Thr Ile Lys Val Lys Glu Leu Thr
1025 1030 1035
Gly Ile Asn Phe Ser Lys Thr Ile Thr Phe Glu Ile Asn Lys Ile
1040 1045 1050
Pro Asp Thr Gly Leu Ile Thr Ser Asp Ser Asp Asn Ile Asn Met
1055 1060 1065
Trp Ile Arg Asp Phe Tyr Ile Phe Ala Lys Glu Leu Asp Gly Lys
1070 1075 1080
Asp Ile Asn Ile Leu Phe Asn Ser Leu Gln Tyr Thr Asn Val Val
1085 1090 1095
Lys Asp Tyr Trp Gly Asn Asp Leu Arg Tyr Asn Lys Glu Tyr Tyr
1100 1105 1110
Met Val Asn Ile Asp Tyr Leu Asn Arg Tyr Met Tyr Ala Asn Ser
1115 1120 1125
Arg Gln Ile Val Phe Asn Thr Arg Arg Asn Asn Asn Asp Phe Asn
1130 1135 1140
Glu Gly Tyr Lys Ile Ile Ile Lys Arg Ile Arg Gly Asn Thr Asn
1145 1150 1155
Asp Thr Arg Val Arg Gly Gly Asp Ile Leu Tyr Phe Asp Met Thr
1160 1165 1170
Ile Asn Asn Lys Ala Tyr Asn Leu Phe Met Lys Asn Glu Thr Met
1175 1180 1185
Tyr Ala Asp Asn His Ser Thr Glu Asp Ile Tyr Ala Ile Gly Leu
1190 1195 1200
Arg Glu Gln Thr Lys Asp Ile Asn Asp Asn Ile Ile Phe Gln Ile
1205 1210 1215
Gln Pro Met Asn Asn Thr Tyr Tyr Tyr Ala Ser Gln Ile Phe Lys
1220 1225 1230
Ser Asn Phe Asn Gly Glu Asn Ile Ser Gly Ile Cys Ser Ile Gly
1235 1240 1245
Thr Tyr Arg Phe Arg Leu Gly Gly Asp Trp Tyr Arg His Asn Tyr
1250 1255 1260
Leu Val Pro Thr Val Lys Gln Gly Asn Tyr Ala Ser Leu Leu Glu
1265 1270 1275
Ser Thr Ser Thr His Trp Gly Phe Val Pro Val Ser Glu
1280 1285 1290
<210> 13
<211> 3756
<212> DNA
<213> 人工序列
<220>
<223> rBoNT/E(0)的核苷酸序列
<400> 13
atgccgaaaa tcaactcttt caactacaac gacccggtta acgaccgtac catcctgtat 60
atcaaaccgg gtggttgcca ggagttctac aaatctttca acatcatgaa aaacatctgg 120
atcatcccgg aacgtaacgt tatcggtacc accccgcagg acttccaccc gccgacctct 180
ctgaaaaacg gtgactcttc ttactacgac ccgaactacc tccagtctga cgaagaaaaa 240
gaccgtttcc tgaaaatcgt taccaaaatc ttcaaccgta tcaacaacaa cctgtctggt 300
ggtatcctgc tggaagaact gtctaaagct aacccgtacc tgggtaacga caacaccccg 360
gacaaccagt tccacatcgg tgacgcttct gctgttgaaa tcaaattctc taacggttct 420
caggacatcc tgctgccgaa cgttatcatc atgggtgctg aaccggacct gttcgaaacc 480
aactcttcta acatctctct gcgtaacaac tacatgccgt ctaaccacgg tttcggttct 540
atcgctatcg ttaccttctc tccggaatac tctttccgtt tcaacgacaa cagcatgaac 600
gagttcatcc aggacccggc tctgaccctg atgcaccaac tgatctactc tctgcacggt 660
ctgtacggtg ctaaaggtat caccaccaaa tacaccatca cccagaaaca gaacccgctg 720
atcaccaaca tccgtggtac caacatcgaa gagttcctga ccttcggtgg taccgacctg 780
aacatcatca cctctgctca gtctaacgac atctacacca acctgctggc tgactacaaa 840
aaaatcgctt ctaaactgtc taaagttcag gtttctaacc cgctgctgaa cccgtacaaa 900
gacgttttcg aagctaaata cggtctggac aaagacgctt ctggtatcta ctctgttaac 960
atcaacaaat tcaacgacat cttcaaaaaa ctgtactctt tcaccgagtt cgacctggcg 1020
accaaattcc aggttaaatg ccgtcagacc tacatcggtc agtacaaata cttcaaactg 1080
tctaacctgc tgaacgactc tatctacaac atctctgaag gttacaacat caacaacctg 1140
aaagttaact tccgtggtca gaacgctaac ctgaacccgc gtatcatcac cccgatcacc 1200
ggtcgtggtc tggttaaaaa aatcatccgt ttctgcaaga atattgtaag cgttaaagga 1260
ataagaaaaa gtatctgcat cgaaatcaac aacggtgaac tgttcttcgt tgcttctgaa 1320
aactcttaca acgacgacaa catcaacacc ccgaaagaaa tcgacgacac cgttacctct 1380
aacaacaact acgaaaacga cctggaccag gttatcctga acttcaactc tgaatctgct 1440
ccgggtctgt ctgacgaaaa actgaacctg accatccaga acgacgctta catcccgaaa 1500
tacgactcta acggtacctc tgacatcgaa cagcacgacg ttaacgaact gaacgttttc 1560
ttctacctgg acgctcagaa agttccggaa ggtgaaaaca acgttaacct gacctcttct 1620
atcgacaccg ctctgctgga acagccgaaa atctacacct tcttctcttc tgagttcatc 1680
aacaacgtta acaaaccggt tcaggctgct ctgttcgttt cttggattca gcaggttctg 1740
gttgacttca ccaccgaagc taaccagaaa tctaccgttg acaaaatcgc tgacatctct 1800
atcgttgttc cgtacatcgg tctggctctg aacatcggta acgaagctca gaaaggtaac 1860
ttcaaagacg ctctggaact gctgggtgct ggtatcctgc tggagttcga accggaactg 1920
ctgatcccga ccatcctggt tttcaccatc aaatctttcc tgggttcttc tgacaacaaa 1980
aacaaagtta tcaaagctat caacaacgct ctgaaagaac gtgacgaaaa atggaaagaa 2040
gtttactctt tcatcgtttc taactggatg accaaaatca acacccagtt caacaaacgt 2100
aaagaacaga tgtaccaggc tctccagaac caggttaacg ctatcaaaac catcatcgaa 2160
tctaaataca actcttacac cctggaagaa aaaaacgaac tgaccaacaa atacgacatc 2220
aaacagatcg aaaacgaact gaaccagaaa gtttctatcg ctatgaacaa catcgaccgt 2280
ttcctgaccg aatcttctat ctcttacctg atgaaactca tcaacgaagt taaaatcaac 2340
aaactgcgtg aatacgacga aaacgttaaa acctacctgc tgaactacat catccagcac 2400
ggttctatcc tgggtgaatc tcagcaggaa ctgaactcta tggttaccga caccctgaac 2460
aactctatcc cgttcaaact gtcttcttac accgacgaca aaatcctgat ctcttacttc 2520
aacaaattct ttaaacgcat taagagttca tcggttctga atatgcggta caaaaatgat 2580
aaatatgtcg atacttctgg atatgatagc aatatcaaca ttaacggcga cgtgtataaa 2640
tatccgacaa ataaaaacca gtttgggata tataacgaca agctgtcgga ggtcaatatt 2700
tctcaaaacg actatatcat ttacgataat aaatataaaa actttagcat tagtttttgg 2760
gttcgtatac ctaattatga caataaaatt gtaaatgtga ataacgagta taccattata 2820
aactgtatgc gcgacaataa cagtggttgg aaggtatcgc tgaaccataa tgagattatc 2880
tggaccctgc aggataatgc aggtataaac cagaaactgg cttttaacta tggaaacgca 2940
aatgggatct cagattacat taataaatgg atttttgtta ccattacgaa cgatcgctta 3000
ggcgactcaa aactttatat taatggcaat ctgatagatc agaaatcaat cttaaatttg 3060
ggcaatattc atgtctctga taacatcttg ttcaagatcg ttaattgcag ttacactcgt 3120
tatattggca ttcgttactt taatatcttc gataaagaac tggacgagac ggaaatccag 3180
actctgtatt caaacgagcc caatactaat atattgaaag atttttgggg taactatctt 3240
ttatatgata aagaatacta tctcctgaat gtattgaagc caaacaattt catagataga 3300
cgcaaggata gcacattaag tatcaacaat atcagatcta ctatactgtt agcaaatcgc 3360
ctctactccg gtattaaagt gaagattcag cgggttaata actccagtac caatgataat 3420
ctggtccgta agaacgatca ggtatacatc aatttcgtcg cgagcaaaac tcatctcttc 3480
ccgctttacg ccgatacagc tacgacaaac aaggaaaaaa ccataaaaat ttccagctcc 3540
ggaaacagat tcaatcaagt agttgtaatg aactctgtgg gtaataattg tacgatgaac 3600
tttaagaata acaatgggaa caatattgga cttttgggct tcaaagccga cacagtggtg 3660
gcgtccacct ggtattacac gcacatgcgg gaccatacga attcgaacgg ttgcttctgg 3720
aactttatct cggaagaaca cgggtggcaa gaaaaa 3756
<210> 14
<211> 1252
<212> PRT
<213> 人工序列
<220>
<223> rBoNT/E(0)的多肽序列
<400> 14
Met Pro Lys Ile Asn Ser Phe Asn Tyr Asn Asp Pro Val Asn Asp Arg
1 5 10 15
Thr Ile Leu Tyr Ile Lys Pro Gly Gly Cys Gln Glu Phe Tyr Lys Ser
20 25 30
Phe Asn Ile Met Lys Asn Ile Trp Ile Ile Pro Glu Arg Asn Val Ile
35 40 45
Gly Thr Thr Pro Gln Asp Phe His Pro Pro Thr Ser Leu Lys Asn Gly
50 55 60
Asp Ser Ser Tyr Tyr Asp Pro Asn Tyr Leu Gln Ser Asp Glu Glu Lys
65 70 75 80
Asp Arg Phe Leu Lys Ile Val Thr Lys Ile Phe Asn Arg Ile Asn Asn
85 90 95
Asn Leu Ser Gly Gly Ile Leu Leu Glu Glu Leu Ser Lys Ala Asn Pro
100 105 110
Tyr Leu Gly Asn Asp Asn Thr Pro Asp Asn Gln Phe His Ile Gly Asp
115 120 125
Ala Ser Ala Val Glu Ile Lys Phe Ser Asn Gly Ser Gln Asp Ile Leu
130 135 140
Leu Pro Asn Val Ile Ile Met Gly Ala Glu Pro Asp Leu Phe Glu Thr
145 150 155 160
Asn Ser Ser Asn Ile Ser Leu Arg Asn Asn Tyr Met Pro Ser Asn His
165 170 175
Gly Phe Gly Ser Ile Ala Ile Val Thr Phe Ser Pro Glu Tyr Ser Phe
180 185 190
Arg Phe Asn Asp Asn Ser Met Asn Glu Phe Ile Gln Asp Pro Ala Leu
195 200 205
Thr Leu Met His Gln Leu Ile Tyr Ser Leu His Gly Leu Tyr Gly Ala
210 215 220
Lys Gly Ile Thr Thr Lys Tyr Thr Ile Thr Gln Lys Gln Asn Pro Leu
225 230 235 240
Ile Thr Asn Ile Arg Gly Thr Asn Ile Glu Glu Phe Leu Thr Phe Gly
245 250 255
Gly Thr Asp Leu Asn Ile Ile Thr Ser Ala Gln Ser Asn Asp Ile Tyr
260 265 270
Thr Asn Leu Leu Ala Asp Tyr Lys Lys Ile Ala Ser Lys Leu Ser Lys
275 280 285
Val Gln Val Ser Asn Pro Leu Leu Asn Pro Tyr Lys Asp Val Phe Glu
290 295 300
Ala Lys Tyr Gly Leu Asp Lys Asp Ala Ser Gly Ile Tyr Ser Val Asn
305 310 315 320
Ile Asn Lys Phe Asn Asp Ile Phe Lys Lys Leu Tyr Ser Phe Thr Glu
325 330 335
Phe Asp Leu Ala Thr Lys Phe Gln Val Lys Cys Arg Gln Thr Tyr Ile
340 345 350
Gly Gln Tyr Lys Tyr Phe Lys Leu Ser Asn Leu Leu Asn Asp Ser Ile
355 360 365
Tyr Asn Ile Ser Glu Gly Tyr Asn Ile Asn Asn Leu Lys Val Asn Phe
370 375 380
Arg Gly Gln Asn Ala Asn Leu Asn Pro Arg Ile Ile Thr Pro Ile Thr
385 390 395 400
Gly Arg Gly Leu Val Lys Lys Ile Ile Arg Phe Cys Lys Asn Ile Val
405 410 415
Ser Val Lys Gly Ile Arg Lys Ser Ile Cys Ile Glu Ile Asn Asn Gly
420 425 430
Glu Leu Phe Phe Val Ala Ser Glu Asn Ser Tyr Asn Asp Asp Asn Ile
435 440 445
Asn Thr Pro Lys Glu Ile Asp Asp Thr Val Thr Ser Asn Asn Asn Tyr
450 455 460
Glu Asn Asp Leu Asp Gln Val Ile Leu Asn Phe Asn Ser Glu Ser Ala
465 470 475 480
Pro Gly Leu Ser Asp Glu Lys Leu Asn Leu Thr Ile Gln Asn Asp Ala
485 490 495
Tyr Ile Pro Lys Tyr Asp Ser Asn Gly Thr Ser Asp Ile Glu Gln His
500 505 510
Asp Val Asn Glu Leu Asn Val Phe Phe Tyr Leu Asp Ala Gln Lys Val
515 520 525
Pro Glu Gly Glu Asn Asn Val Asn Leu Thr Ser Ser Ile Asp Thr Ala
530 535 540
Leu Leu Glu Gln Pro Lys Ile Tyr Thr Phe Phe Ser Ser Glu Phe Ile
545 550 555 560
Asn Asn Val Asn Lys Pro Val Gln Ala Ala Leu Phe Val Ser Trp Ile
565 570 575
Gln Gln Val Leu Val Asp Phe Thr Thr Glu Ala Asn Gln Lys Ser Thr
580 585 590
Val Asp Lys Ile Ala Asp Ile Ser Ile Val Val Pro Tyr Ile Gly Leu
595 600 605
Ala Leu Asn Ile Gly Asn Glu Ala Gln Lys Gly Asn Phe Lys Asp Ala
610 615 620
Leu Glu Leu Leu Gly Ala Gly Ile Leu Leu Glu Phe Glu Pro Glu Leu
625 630 635 640
Leu Ile Pro Thr Ile Leu Val Phe Thr Ile Lys Ser Phe Leu Gly Ser
645 650 655
Ser Asp Asn Lys Asn Lys Val Ile Lys Ala Ile Asn Asn Ala Leu Lys
660 665 670
Glu Arg Asp Glu Lys Trp Lys Glu Val Tyr Ser Phe Ile Val Ser Asn
675 680 685
Trp Met Thr Lys Ile Asn Thr Gln Phe Asn Lys Arg Lys Glu Gln Met
690 695 700
Tyr Gln Ala Leu Gln Asn Gln Val Asn Ala Ile Lys Thr Ile Ile Glu
705 710 715 720
Ser Lys Tyr Asn Ser Tyr Thr Leu Glu Glu Lys Asn Glu Leu Thr Asn
725 730 735
Lys Tyr Asp Ile Lys Gln Ile Glu Asn Glu Leu Asn Gln Lys Val Ser
740 745 750
Ile Ala Met Asn Asn Ile Asp Arg Phe Leu Thr Glu Ser Ser Ile Ser
755 760 765
Tyr Leu Met Lys Leu Ile Asn Glu Val Lys Ile Asn Lys Leu Arg Glu
770 775 780
Tyr Asp Glu Asn Val Lys Thr Tyr Leu Leu Asn Tyr Ile Ile Gln His
785 790 795 800
Gly Ser Ile Leu Gly Glu Ser Gln Gln Glu Leu Asn Ser Met Val Thr
805 810 815
Asp Thr Leu Asn Asn Ser Ile Pro Phe Lys Leu Ser Ser Tyr Thr Asp
820 825 830
Asp Lys Ile Leu Ile Ser Tyr Phe Asn Lys Phe Phe Lys Arg Ile Lys
835 840 845
Ser Ser Ser Val Leu Asn Met Arg Tyr Lys Asn Asp Lys Tyr Val Asp
850 855 860
Thr Ser Gly Tyr Asp Ser Asn Ile Asn Ile Asn Gly Asp Val Tyr Lys
865 870 875 880
Tyr Pro Thr Asn Lys Asn Gln Phe Gly Ile Tyr Asn Asp Lys Leu Ser
885 890 895
Glu Val Asn Ile Ser Gln Asn Asp Tyr Ile Ile Tyr Asp Asn Lys Tyr
900 905 910
Lys Asn Phe Ser Ile Ser Phe Trp Val Arg Ile Pro Asn Tyr Asp Asn
915 920 925
Lys Ile Val Asn Val Asn Asn Glu Tyr Thr Ile Ile Asn Cys Met Arg
930 935 940
Asp Asn Asn Ser Gly Trp Lys Val Ser Leu Asn His Asn Glu Ile Ile
945 950 955 960
Trp Thr Leu Gln Asp Asn Ala Gly Ile Asn Gln Lys Leu Ala Phe Asn
965 970 975
Tyr Gly Asn Ala Asn Gly Ile Ser Asp Tyr Ile Asn Lys Trp Ile Phe
980 985 990
Val Thr Ile Thr Asn Asp Arg Leu Gly Asp Ser Lys Leu Tyr Ile Asn
995 1000 1005
Gly Asn Leu Ile Asp Gln Lys Ser Ile Leu Asn Leu Gly Asn Ile
1010 1015 1020
His Val Ser Asp Asn Ile Leu Phe Lys Ile Val Asn Cys Ser Tyr
1025 1030 1035
Thr Arg Tyr Ile Gly Ile Arg Tyr Phe Asn Ile Phe Asp Lys Glu
1040 1045 1050
Leu Asp Glu Thr Glu Ile Gln Thr Leu Tyr Ser Asn Glu Pro Asn
1055 1060 1065
Thr Asn Ile Leu Lys Asp Phe Trp Gly Asn Tyr Leu Leu Tyr Asp
1070 1075 1080
Lys Glu Tyr Tyr Leu Leu Asn Val Leu Lys Pro Asn Asn Phe Ile
1085 1090 1095
Asp Arg Arg Lys Asp Ser Thr Leu Ser Ile Asn Asn Ile Arg Ser
1100 1105 1110
Thr Ile Leu Leu Ala Asn Arg Leu Tyr Ser Gly Ile Lys Val Lys
1115 1120 1125
Ile Gln Arg Val Asn Asn Ser Ser Thr Asn Asp Asn Leu Val Arg
1130 1135 1140
Lys Asn Asp Gln Val Tyr Ile Asn Phe Val Ala Ser Lys Thr His
1145 1150 1155
Leu Phe Pro Leu Tyr Ala Asp Thr Ala Thr Thr Asn Lys Glu Lys
1160 1165 1170
Thr Ile Lys Ile Ser Ser Ser Gly Asn Arg Phe Asn Gln Val Val
1175 1180 1185
Val Met Asn Ser Val Gly Asn Asn Cys Thr Met Asn Phe Lys Asn
1190 1195 1200
Asn Asn Gly Asn Asn Ile Gly Leu Leu Gly Phe Lys Ala Asp Thr
1205 1210 1215
Val Val Ala Ser Thr Trp Tyr Tyr Thr His Met Arg Asp His Thr
1220 1225 1230
Asn Ser Asn Gly Cys Phe Trp Asn Phe Ile Ser Glu Glu His Gly
1235 1240 1245
Trp Gln Glu Lys
1250
<210> 15
<211> 3834
<212> DNA
<213> 人工序列
<220>
<223> rBoNT/F(0)的核苷酸序列
<400> 15
atgccggtgg tcatcaacag cttcaactac aacgacccag taaacgacga cacgatcctg 60
tatatgcaaa tcccgtatga agagaagagc aagaagtact ataaggcctt tgaaatcatg 120
cgcaatgtgt ggattattcc ggagcgtaat acgattggta ctgacccaag cgacttcgat 180
ccacctgcgt ctttggaaaa cggctcgtcc gcatattacg acccgaatta cctgaccacc 240
gatgcggaga aagatcgtta tttgaaaacc accatcaagc tgttcaaacg cattaacagc 300
aatccggcag gtgaggtcct gctgcaagag attagctacg caaagcctta tctgggtaat 360
gagcatacgc ctattaacga gtttcacccg gttacccgca ctaccagcgt taacatcaag 420
tcctcgacca acgtgaagtc tagcattatc ctgaacctgc tggttctggg tgccggtccg 480
gacatcttcg aaaactctag ctacccggtg cgtaaactga tggatagcgg cggtgtttat 540
gacccgagca atgacggttt tggcagcatc aatatcgtga cgtttagccc ggagtacgag 600
tacaccttca atgatatcag cggtggttac aattcttcta ccgagagctt catcgccgac 660
ccggcgatca gcctggcaca ccaactgatc tatgcattgc atggcttgta cggtgcccgt 720
ggtgtgacgt ataaagagac tatcaaggtt aagcaggcac ctctgatgat tgcggaaaag 780
ccgattcgcc tggaagagtt cctgaccttc ggcggtcaag atttgaacat cattacctcg 840
gccatgaaag agaaaatcta taacaatttg ctggccaact atgaaaagat tgcaacgcgc 900
ttgtctcgtg ttaactccgc tccgccggaa tacgacatta atgagtacaa agactacttt 960
caatggaaat atggcctgga caaaaatgcg gatggttctt ataccgtgaa tgaaaacaaa 1020
ttcaatgaaa tctacaagaa actgtacagc ttcaccgaaa tcgatctggc gaacaagttc 1080
aaagtcaaat gtcgtaatac ctacttcatc aaatatggct tcctgaaagt cccgaacctg 1140
ctggacgatg acatctatac cgtcagcgaa ggcttcaaca tcggcaatct ggccgtgaat 1200
aatcgtggtc agaacatcaa actgaatccg aaaatcattg actccatccc agacaagggc 1260
ctggttgaga aaatcgtgaa gttctgcaaa agcgttattc cgcgtaaagg tacgaaagca 1320
ccgcctcgcc tgtgcattcg cgttaacaac cgtgagttgt tctttgtggc atctgaaagc 1380
agctacaacg agaacgacat caacacccct aaagaaattg atgataccac gaacctgaat 1440
aacaattatc gcaacaatct ggacgaggtg atcctggatt acaattcgga aaccattccg 1500
caaattagca atcagacgct gaacaccctg gttcaggacg atagctacgt tccgcgttac 1560
gactccaatg gtactagcga gattgaagaa cacaacgtag tggacttgaa cgttttcttt 1620
tatctgcacg cccagaaggt tccggagggc gaaaccaata ttagcctgac cagctcgatc 1680
gacaccgcgc tgtctgagga gagccaagtc tacacctttt tcagcagcga gtttatcaac 1740
actattaaca agccagttca tgctgcattg tttatctctt ggattaacca ggtgattcgc 1800
gactttacga cggaggcgac ccagaagtct accttcgaca aaattgcaga catctccctg 1860
gtcgtcccat acgtcggcct ggcgttgaat attggcaatg aagttcaaaa agagaacttc 1920
aaagaagcgt tcgagctgct gggtgcaggc atcctgctgg agttcgtgcc ggaactgttg 1980
atcccgacca tcctggtgtt caccattaag agcttcattg gatcctccga gaataagaac 2040
aagatcatca aggcgatcaa taacagcctg atggagcgtg aaacgaagtg gaaagaaatc 2100
tatagctgga ttgttagcaa ttggctgact cgtattaaca cgcaattcaa caagcgtaaa 2160
gagcaaatgt accaagccct gcaaaaccaa gttgacgcca tcaaaacggt aattgaatac 2220
aagtacaaca attacacgag cgatgagcgc aaccgcctgg aaagcgaata caacatcaac 2280
aacattcgcg aagaattgaa caagaaagtg agcctggcga tggagaacat tgagcgtttt 2340
atcaccgaaa gcagcatctt ttacctgatg aaattgatta atgaggcgaa agtctcgaaa 2400
ctgcgtgagt acgacgaagg tgtgaaagag tatctgctgg attacattag cgagcaccgt 2460
agcatcttgg gtaactcggt tcaggagctg aacgatctgg tgacctctac cctgaacaat 2520
agcatcccgt tcgaactgag cagctatacc aatgacaaga ttctgattct gtatttcaat 2580
aaactgtata agaagatcaa ggataacagc attctggata tgcgttacga aaacaataag 2640
tttatcgaca tttctggtta cggcagcaac atttccatca atggcgatgt ctacatctac 2700
agcaccaatc gcaaccagtt cggcatctac tctagcaaac cgagcgaagt taacatcgca 2760
cagaacaatg atattattta taacggtcgt tatcaaaact tctctatcag cttttgggtc 2820
cgtatcccga agtacttcaa taaagtcaat ctgaataatg aatacacgat catcgactgc 2880
attcgcaata acaacagcgg ttggaaaatc agcctgaatt acaacaaaat tatttggacc 2940
ctgcaagata cggcgggtaa caatcagaaa ctggtgttta actacacgca aatgatcagc 3000
atttctgact atatcaacaa gtggatcttt gttaccatca ccaataatcg tctgggcaat 3060
agccgtattt acatcaacgg taacctgatt gatgagaaaa gcatcagcaa cctgggcgat 3120
attcacgtca gcgacaacat tctgttcaaa attgttggtt gtaacgatac ccgttacgtc 3180
ggcatccgtt atttcaaggt tttcgatacg gagctgggta aaacggaaat cgaaacgttg 3240
tactccgatg aaccagatcc gagcattctg aaggactttt ggggtaacta cttgctgtac 3300
aataaacgtt actatctgct gaatctgttg cgcaccgaca agagcattac ccaaaacagc 3360
aatttcctga acattaatca gcaacgcggc gtataccaaa aaccgaacat cttcagcaat 3420
acgcgcctgt atactggtgt tgaagtgatc attcgtaaga acggtagcac cgacattagc 3480
aacacggaca atttcgtccg taagaatgac ctggcgtaca ttaacgtcgt ggaccgtgat 3540
gtcgagtatc gtctgtacgc agacatcagc attgcgaaac cggaaaagat tatcaagctg 3600
atccgtacca gcaacagcaa caacagcctg ggtcagatca ttgtgatgga cagcattggt 3660
aataactgca cgatgaactt ccagaacaac aatggtggta atatcggtct gctgggtttt 3720
cacagcaata atctggttgc ttccagctgg tactacaata acattcgtaa aaacacgtct 3780
agcaatggtt gtttttggag ctttatcagc aaagagcacg gctggcaaga aaat 3834
<210> 16
<211> 1278
<212> PRT
<213> 人工序列
<220>
<223> rBoNT/F(0)的多肽序列
<400> 16
Met Pro Val Val Ile Asn Ser Phe Asn Tyr Asn Asp Pro Val Asn Asp
1 5 10 15
Asp Thr Ile Leu Tyr Met Gln Ile Pro Tyr Glu Glu Lys Ser Lys Lys
20 25 30
Tyr Tyr Lys Ala Phe Glu Ile Met Arg Asn Val Trp Ile Ile Pro Glu
35 40 45
Arg Asn Thr Ile Gly Thr Asp Pro Ser Asp Phe Asp Pro Pro Ala Ser
50 55 60
Leu Glu Asn Gly Ser Ser Ala Tyr Tyr Asp Pro Asn Tyr Leu Thr Thr
65 70 75 80
Asp Ala Glu Lys Asp Arg Tyr Leu Lys Thr Thr Ile Lys Leu Phe Lys
85 90 95
Arg Ile Asn Ser Asn Pro Ala Gly Glu Val Leu Leu Gln Glu Ile Ser
100 105 110
Tyr Ala Lys Pro Tyr Leu Gly Asn Glu His Thr Pro Ile Asn Glu Phe
115 120 125
His Pro Val Thr Arg Thr Thr Ser Val Asn Ile Lys Ser Ser Thr Asn
130 135 140
Val Lys Ser Ser Ile Ile Leu Asn Leu Leu Val Leu Gly Ala Gly Pro
145 150 155 160
Asp Ile Phe Glu Asn Ser Ser Tyr Pro Val Arg Lys Leu Met Asp Ser
165 170 175
Gly Gly Val Tyr Asp Pro Ser Asn Asp Gly Phe Gly Ser Ile Asn Ile
180 185 190
Val Thr Phe Ser Pro Glu Tyr Glu Tyr Thr Phe Asn Asp Ile Ser Gly
195 200 205
Gly Tyr Asn Ser Ser Thr Glu Ser Phe Ile Ala Asp Pro Ala Ile Ser
210 215 220
Leu Ala His Gln Leu Ile Tyr Ala Leu His Gly Leu Tyr Gly Ala Arg
225 230 235 240
Gly Val Thr Tyr Lys Glu Thr Ile Lys Val Lys Gln Ala Pro Leu Met
245 250 255
Ile Ala Glu Lys Pro Ile Arg Leu Glu Glu Phe Leu Thr Phe Gly Gly
260 265 270
Gln Asp Leu Asn Ile Ile Thr Ser Ala Met Lys Glu Lys Ile Tyr Asn
275 280 285
Asn Leu Leu Ala Asn Tyr Glu Lys Ile Ala Thr Arg Leu Ser Arg Val
290 295 300
Asn Ser Ala Pro Pro Glu Tyr Asp Ile Asn Glu Tyr Lys Asp Tyr Phe
305 310 315 320
Gln Trp Lys Tyr Gly Leu Asp Lys Asn Ala Asp Gly Ser Tyr Thr Val
325 330 335
Asn Glu Asn Lys Phe Asn Glu Ile Tyr Lys Lys Leu Tyr Ser Phe Thr
340 345 350
Glu Ile Asp Leu Ala Asn Lys Phe Lys Val Lys Cys Arg Asn Thr Tyr
355 360 365
Phe Ile Lys Tyr Gly Phe Leu Lys Val Pro Asn Leu Leu Asp Asp Asp
370 375 380
Ile Tyr Thr Val Ser Glu Gly Phe Asn Ile Gly Asn Leu Ala Val Asn
385 390 395 400
Asn Arg Gly Gln Asn Ile Lys Leu Asn Pro Lys Ile Ile Asp Ser Ile
405 410 415
Pro Asp Lys Gly Leu Val Glu Lys Ile Val Lys Phe Cys Lys Ser Val
420 425 430
Ile Pro Arg Lys Gly Thr Lys Ala Pro Pro Arg Leu Cys Ile Arg Val
435 440 445
Asn Asn Arg Glu Leu Phe Phe Val Ala Ser Glu Ser Ser Tyr Asn Glu
450 455 460
Asn Asp Ile Asn Thr Pro Lys Glu Ile Asp Asp Thr Thr Asn Leu Asn
465 470 475 480
Asn Asn Tyr Arg Asn Asn Leu Asp Glu Val Ile Leu Asp Tyr Asn Ser
485 490 495
Glu Thr Ile Pro Gln Ile Ser Asn Gln Thr Leu Asn Thr Leu Val Gln
500 505 510
Asp Asp Ser Tyr Val Pro Arg Tyr Asp Ser Asn Gly Thr Ser Glu Ile
515 520 525
Glu Glu His Asn Val Val Asp Leu Asn Val Phe Phe Tyr Leu His Ala
530 535 540
Gln Lys Val Pro Glu Gly Glu Thr Asn Ile Ser Leu Thr Ser Ser Ile
545 550 555 560
Asp Thr Ala Leu Ser Glu Glu Ser Gln Val Tyr Thr Phe Phe Ser Ser
565 570 575
Glu Phe Ile Asn Thr Ile Asn Lys Pro Val His Ala Ala Leu Phe Ile
580 585 590
Ser Trp Ile Asn Gln Val Ile Arg Asp Phe Thr Thr Glu Ala Thr Gln
595 600 605
Lys Ser Thr Phe Asp Lys Ile Ala Asp Ile Ser Leu Val Val Pro Tyr
610 615 620
Val Gly Leu Ala Leu Asn Ile Gly Asn Glu Val Gln Lys Glu Asn Phe
625 630 635 640
Lys Glu Ala Phe Glu Leu Leu Gly Ala Gly Ile Leu Leu Glu Phe Val
645 650 655
Pro Glu Leu Leu Ile Pro Thr Ile Leu Val Phe Thr Ile Lys Ser Phe
660 665 670
Ile Gly Ser Ser Glu Asn Lys Asn Lys Ile Ile Lys Ala Ile Asn Asn
675 680 685
Ser Leu Met Glu Arg Glu Thr Lys Trp Lys Glu Ile Tyr Ser Trp Ile
690 695 700
Val Ser Asn Trp Leu Thr Arg Ile Asn Thr Gln Phe Asn Lys Arg Lys
705 710 715 720
Glu Gln Met Tyr Gln Ala Leu Gln Asn Gln Val Asp Ala Ile Lys Thr
725 730 735
Val Ile Glu Tyr Lys Tyr Asn Asn Tyr Thr Ser Asp Glu Arg Asn Arg
740 745 750
Leu Glu Ser Glu Tyr Asn Ile Asn Asn Ile Arg Glu Glu Leu Asn Lys
755 760 765
Lys Val Ser Leu Ala Met Glu Asn Ile Glu Arg Phe Ile Thr Glu Ser
770 775 780
Ser Ile Phe Tyr Leu Met Lys Leu Ile Asn Glu Ala Lys Val Ser Lys
785 790 795 800
Leu Arg Glu Tyr Asp Glu Gly Val Lys Glu Tyr Leu Leu Asp Tyr Ile
805 810 815
Ser Glu His Arg Ser Ile Leu Gly Asn Ser Val Gln Glu Leu Asn Asp
820 825 830
Leu Val Thr Ser Thr Leu Asn Asn Ser Ile Pro Phe Glu Leu Ser Ser
835 840 845
Tyr Thr Asn Asp Lys Ile Leu Ile Leu Tyr Phe Asn Lys Leu Tyr Lys
850 855 860
Lys Ile Lys Asp Asn Ser Ile Leu Asp Met Arg Tyr Glu Asn Asn Lys
865 870 875 880
Phe Ile Asp Ile Ser Gly Tyr Gly Ser Asn Ile Ser Ile Asn Gly Asp
885 890 895
Val Tyr Ile Tyr Ser Thr Asn Arg Asn Gln Phe Gly Ile Tyr Ser Ser
900 905 910
Lys Pro Ser Glu Val Asn Ile Ala Gln Asn Asn Asp Ile Ile Tyr Asn
915 920 925
Gly Arg Tyr Gln Asn Phe Ser Ile Ser Phe Trp Val Arg Ile Pro Lys
930 935 940
Tyr Phe Asn Lys Val Asn Leu Asn Asn Glu Tyr Thr Ile Ile Asp Cys
945 950 955 960
Ile Arg Asn Asn Asn Ser Gly Trp Lys Ile Ser Leu Asn Tyr Asn Lys
965 970 975
Ile Ile Trp Thr Leu Gln Asp Thr Ala Gly Asn Asn Gln Lys Leu Val
980 985 990
Phe Asn Tyr Thr Gln Met Ile Ser Ile Ser Asp Tyr Ile Asn Lys Trp
995 1000 1005
Ile Phe Val Thr Ile Thr Asn Asn Arg Leu Gly Asn Ser Arg Ile
1010 1015 1020
Tyr Ile Asn Gly Asn Leu Ile Asp Glu Lys Ser Ile Ser Asn Leu
1025 1030 1035
Gly Asp Ile His Val Ser Asp Asn Ile Leu Phe Lys Ile Val Gly
1040 1045 1050
Cys Asn Asp Thr Arg Tyr Val Gly Ile Arg Tyr Phe Lys Val Phe
1055 1060 1065
Asp Thr Glu Leu Gly Lys Thr Glu Ile Glu Thr Leu Tyr Ser Asp
1070 1075 1080
Glu Pro Asp Pro Ser Ile Leu Lys Asp Phe Trp Gly Asn Tyr Leu
1085 1090 1095
Leu Tyr Asn Lys Arg Tyr Tyr Leu Leu Asn Leu Leu Arg Thr Asp
1100 1105 1110
Lys Ser Ile Thr Gln Asn Ser Asn Phe Leu Asn Ile Asn Gln Gln
1115 1120 1125
Arg Gly Val Tyr Gln Lys Pro Asn Ile Phe Ser Asn Thr Arg Leu
1130 1135 1140
Tyr Thr Gly Val Glu Val Ile Ile Arg Lys Asn Gly Ser Thr Asp
1145 1150 1155
Ile Ser Asn Thr Asp Asn Phe Val Arg Lys Asn Asp Leu Ala Tyr
1160 1165 1170
Ile Asn Val Val Asp Arg Asp Val Glu Tyr Arg Leu Tyr Ala Asp
1175 1180 1185
Ile Ser Ile Ala Lys Pro Glu Lys Ile Ile Lys Leu Ile Arg Thr
1190 1195 1200
Ser Asn Ser Asn Asn Ser Leu Gly Gln Ile Ile Val Met Asp Ser
1205 1210 1215
Ile Gly Asn Asn Cys Thr Met Asn Phe Gln Asn Asn Asn Gly Gly
1220 1225 1230
Asn Ile Gly Leu Leu Gly Phe His Ser Asn Asn Leu Val Ala Ser
1235 1240 1245
Ser Trp Tyr Tyr Asn Asn Ile Arg Lys Asn Thr Ser Ser Asn Gly
1250 1255 1260
Cys Phe Trp Ser Phe Ile Ser Lys Glu His Gly Trp Gln Glu Asn
1265 1270 1275
<210> 17
<211> 3942
<212> DNA
<213> 人工序列
<220>
<223> rBoNT/A(0)的核苷酸序列 (His-标签的)
<400> 17
atgccgtttg tgaacaagca gttcaactat aaagatccgg ttaatggtgt ggatatcgcc 60
tatatcaaaa ttccgaatgc aggtcagatg cagccggtta aagcctttaa aatccataac 120
aaaatttggg tgattccgga acgtgatacc tttaccaatc cggaagaagg tgatctgaat 180
ccgcctccgg aagcaaaaca ggttccggtt agctattatg atagcaccta tctgagcacc 240
gataacgaga aagataacta tctgaaaggt gtgaccaaac tgtttgaacg catttatagt 300
accgatctgg gtcgtatgct gctgaccagc attgttcgtg gtattccgtt ttggggtggt 360
agcaccattg ataccgaact gaaagttatt gacaccaact gcattaatgt gattcagccg 420
gatggtagct atcgtagcga agaactgaat ctggttatta ttggtccgag cgcagatatc 480
attcagtttg aatgtaaaag ctttggccac gaagttctga atctgacccg taatggttat 540
ggtagtaccc agtatattcg tttcagtccg gattttacct ttggctttga agaaagcctg 600
gaagttgata caaatccgct gttaggtgca ggtaaatttg caaccgatcc ggcagttacc 660
ctggcacacc agctgattta tgccggtcat cgtctgtatg gtattgccat taatccgaat 720
cgtgtgttca aagtgaatac caacgcctat tatgaaatga gcggtctgga agtgagtttt 780
gaagaactgc gtacctttgg tggtcatgat gccaaattta tcgatagcct gcaagaaaat 840
gaatttcgcc tgtactacta taacaaattc aaggatattg cgagcaccct gaataaagcc 900
aaaagcattg ttggcaccac cgcaagcctg cagtatatga aaaatgtgtt taaagaaaaa 960
tatctgctga gcgaagatac cagcggtaaa tttagcgttg acaaactgaa attcgataaa 1020
ctgtacaaga tgctgaccga gatttatacc gaagataact tcgtgaagtt tttcaaagtg 1080
ctgaaccgca aaacctacct gaactttgat aaagccgtgt tcaaaatcaa catcgtgccg 1140
aaagtgaact ataccatcta tgatggtttt aacctgcgca ataccaatct ggcagcaaac 1200
tttaatggtc agaacaccga aatcaacaac atgaacttta ccaaactgaa gaacttcacc 1260
ggtctgttcg aattttacaa actgctgtgt gttcgtggca ttattaccag caaaaccaaa 1320
agtctggata aaggctacaa taaagccctg aatgatctgt gcattaaggt gaataattgg 1380
gacctgtttt ttagcccgag cgaggataat ttcaccaacg atctgaacaa aggcgaagaa 1440
attaccagcg ataccaatat tgaagcagcc gaagaaaaca ttagcctgga tctgattcag 1500
cagtattatc tgaccttcaa cttcgataat gagccggaaa atatcagcat tgaaaacctg 1560
agcagcgata ttattggcca gctggaactg atgccgaata ttgaacgttt tccgaacggc 1620
aaaaaatacg agctggataa atacaccatg ttccattatc tgcgtgccca agaatttgaa 1680
catggtaaaa gccgtattgc actgaccaat agcgttaatg aagcactgct gaacccgagc 1740
cgtgtttata ccttttttag cagcgattac gtgaaaaagg ttaacaaagc aaccgaagca 1800
gccatgtttt taggttgggt tgaacagctg gtttatgatt tcaccgatga aaccagcgaa 1860
gttagcacca ccgataaaat tgcagatatt accatcatca tcccgtatat cggtccggca 1920
ctgaatattg gcaatatgct gtataaagac gattttgtgg gtgccctgat ttttagcggt 1980
gcagttattc tgctggaatt tattccggaa attgccattc cggttctggg cacctttgca 2040
ctggtgagct atattgcaaa taaagttctg accgtgcaga ccatcgataa tgcactgagc 2100
aaacgtaacg aaaaatggga tgaagtgtac aagtatatcg tgaccaattg gctggcaaaa 2160
gttaacaccc agattgacct gattcgcaag aagatgaaag aagcactgga aaatcaggca 2220
gaagcaacca aagccattat caactatcag tataaccagt acaccgaaga agagaaaaat 2280
aacatcaact tcaacatcga cgatctgtcc agcaaactga acgaaagcat caacaaagcc 2340
atgattaaca ttaacaaatt tctgaaccag tgcagcgtga gctatctgat gaatagcatg 2400
attccgtatg gtgtgaaacg tctggaagat tttgatgcaa gcctgaaaga tgccctgctg 2460
aaatatatct atgataatcg tggcaccctg attggtcagg ttgatcgtct gaaagataaa 2520
gtgaacaaca ccctgagtac cgatattcct tttcagctga gcaaatatgt ggataatcag 2580
cgtctgctgt caacctttac cgaatacatt aagaacatca tcaacaccag cattctgaac 2640
ctgcgttatg aaagcaatca tctgattgat ctgagccgtt atgccagcaa aatcaatata 2700
ggcagcaagg ttaacttcga cccgattgac aaaaatcaga tacagctgtt taatctggaa 2760
agcagcaaaa ttgaggtgat cctgaaaaac gccattgtgt ataatagcat gtacgagaat 2820
ttctcgacca gcttttggat tcgtatcccg aaatacttta atagcatcag cctgaacaac 2880
gagtacacca ttattaactg catggaaaac aatagcggct ggaaagttag cctgaattat 2940
ggcgaaatta tctggaccct gcaggatacc caagaaatca aacagcgtgt ggttttcaaa 3000
tacagccaga tgattaatat cagcgactat atcaaccgct ggatttttgt gaccattacc 3060
aataatcgcc tgaataacag caagatctat attaacggtc gtctgattga ccagaaaccg 3120
attagtaatc tgggtaatat tcatgcgagc aacaacatca tgtttaaact ggatggttgt 3180
cgtgataccc atcgttatat ttggatcaag tacttcaacc tgttcgataa agagttgaac 3240
gaaaaagaaa ttaaagacct gtatgataac cagagcaaca gcggtattct gaaggatttt 3300
tggggagatt atctgcagta tgacaaaccg tattatatgc tgaatctgta cgacccgaat 3360
aaatacgtgg atgtgaataa tgttggcatc cgtggttata tgtacctgaa aggtccgcgt 3420
ggtagcgtta tgaccacaaa catttatctg aatagcagcc tgtatcgcgg aaccaaattc 3480
atcattaaaa agtatgccag cggcaacaag gataatattg tgcgtaataa tgatcgcgtg 3540
tacattaacg ttgtggtgaa gaataaagaa tatcgcctgg caaccaatgc aagccaggca 3600
ggcgttgaaa aaattctgag tgccctggaa attccggatg ttggtaatct gagccaggtt 3660
gttgtgatga aaagcaaaaa tgatcagggc atcaccaaca agtgcaaaat gaatctgcag 3720
gacaataacg gcaacgatat tggttttatt ggcttccacc agttcaacaa tattgcgaaa 3780
ctggttgcaa gcaattggta taatcgtcag attgaacgta gcagtcgtac cctgggttgt 3840
agctgggaat ttatccctgt ggatgatggt tggggtgaac gtccgctgga aaacctgtat 3900
tttcaaggtg caagtcatca tcaccatcac caccatcatt aa 3942
<210> 18
<211> 1313
<212> PRT
<213> 人工序列
<220>
<223> rBoNT/A(0)的多肽序列 (His-标签的)
<400> 18
Met Pro Phe Val Asn Lys Gln Phe Asn Tyr Lys Asp Pro Val Asn Gly
1 5 10 15
Val Asp Ile Ala Tyr Ile Lys Ile Pro Asn Ala Gly Gln Met Gln Pro
20 25 30
Val Lys Ala Phe Lys Ile His Asn Lys Ile Trp Val Ile Pro Glu Arg
35 40 45
Asp Thr Phe Thr Asn Pro Glu Glu Gly Asp Leu Asn Pro Pro Pro Glu
50 55 60
Ala Lys Gln Val Pro Val Ser Tyr Tyr Asp Ser Thr Tyr Leu Ser Thr
65 70 75 80
Asp Asn Glu Lys Asp Asn Tyr Leu Lys Gly Val Thr Lys Leu Phe Glu
85 90 95
Arg Ile Tyr Ser Thr Asp Leu Gly Arg Met Leu Leu Thr Ser Ile Val
100 105 110
Arg Gly Ile Pro Phe Trp Gly Gly Ser Thr Ile Asp Thr Glu Leu Lys
115 120 125
Val Ile Asp Thr Asn Cys Ile Asn Val Ile Gln Pro Asp Gly Ser Tyr
130 135 140
Arg Ser Glu Glu Leu Asn Leu Val Ile Ile Gly Pro Ser Ala Asp Ile
145 150 155 160
Ile Gln Phe Glu Cys Lys Ser Phe Gly His Glu Val Leu Asn Leu Thr
165 170 175
Arg Asn Gly Tyr Gly Ser Thr Gln Tyr Ile Arg Phe Ser Pro Asp Phe
180 185 190
Thr Phe Gly Phe Glu Glu Ser Leu Glu Val Asp Thr Asn Pro Leu Leu
195 200 205
Gly Ala Gly Lys Phe Ala Thr Asp Pro Ala Val Thr Leu Ala His Gln
210 215 220
Leu Ile Tyr Ala Gly His Arg Leu Tyr Gly Ile Ala Ile Asn Pro Asn
225 230 235 240
Arg Val Phe Lys Val Asn Thr Asn Ala Tyr Tyr Glu Met Ser Gly Leu
245 250 255
Glu Val Ser Phe Glu Glu Leu Arg Thr Phe Gly Gly His Asp Ala Lys
260 265 270
Phe Ile Asp Ser Leu Gln Glu Asn Glu Phe Arg Leu Tyr Tyr Tyr Asn
275 280 285
Lys Phe Lys Asp Ile Ala Ser Thr Leu Asn Lys Ala Lys Ser Ile Val
290 295 300
Gly Thr Thr Ala Ser Leu Gln Tyr Met Lys Asn Val Phe Lys Glu Lys
305 310 315 320
Tyr Leu Leu Ser Glu Asp Thr Ser Gly Lys Phe Ser Val Asp Lys Leu
325 330 335
Lys Phe Asp Lys Leu Tyr Lys Met Leu Thr Glu Ile Tyr Thr Glu Asp
340 345 350
Asn Phe Val Lys Phe Phe Lys Val Leu Asn Arg Lys Thr Tyr Leu Asn
355 360 365
Phe Asp Lys Ala Val Phe Lys Ile Asn Ile Val Pro Lys Val Asn Tyr
370 375 380
Thr Ile Tyr Asp Gly Phe Asn Leu Arg Asn Thr Asn Leu Ala Ala Asn
385 390 395 400
Phe Asn Gly Gln Asn Thr Glu Ile Asn Asn Met Asn Phe Thr Lys Leu
405 410 415
Lys Asn Phe Thr Gly Leu Phe Glu Phe Tyr Lys Leu Leu Cys Val Arg
420 425 430
Gly Ile Ile Thr Ser Lys Thr Lys Ser Leu Asp Lys Gly Tyr Asn Lys
435 440 445
Ala Leu Asn Asp Leu Cys Ile Lys Val Asn Asn Trp Asp Leu Phe Phe
450 455 460
Ser Pro Ser Glu Asp Asn Phe Thr Asn Asp Leu Asn Lys Gly Glu Glu
465 470 475 480
Ile Thr Ser Asp Thr Asn Ile Glu Ala Ala Glu Glu Asn Ile Ser Leu
485 490 495
Asp Leu Ile Gln Gln Tyr Tyr Leu Thr Phe Asn Phe Asp Asn Glu Pro
500 505 510
Glu Asn Ile Ser Ile Glu Asn Leu Ser Ser Asp Ile Ile Gly Gln Leu
515 520 525
Glu Leu Met Pro Asn Ile Glu Arg Phe Pro Asn Gly Lys Lys Tyr Glu
530 535 540
Leu Asp Lys Tyr Thr Met Phe His Tyr Leu Arg Ala Gln Glu Phe Glu
545 550 555 560
His Gly Lys Ser Arg Ile Ala Leu Thr Asn Ser Val Asn Glu Ala Leu
565 570 575
Leu Asn Pro Ser Arg Val Tyr Thr Phe Phe Ser Ser Asp Tyr Val Lys
580 585 590
Lys Val Asn Lys Ala Thr Glu Ala Ala Met Phe Leu Gly Trp Val Glu
595 600 605
Gln Leu Val Tyr Asp Phe Thr Asp Glu Thr Ser Glu Val Ser Thr Thr
610 615 620
Asp Lys Ile Ala Asp Ile Thr Ile Ile Ile Pro Tyr Ile Gly Pro Ala
625 630 635 640
Leu Asn Ile Gly Asn Met Leu Tyr Lys Asp Asp Phe Val Gly Ala Leu
645 650 655
Ile Phe Ser Gly Ala Val Ile Leu Leu Glu Phe Ile Pro Glu Ile Ala
660 665 670
Ile Pro Val Leu Gly Thr Phe Ala Leu Val Ser Tyr Ile Ala Asn Lys
675 680 685
Val Leu Thr Val Gln Thr Ile Asp Asn Ala Leu Ser Lys Arg Asn Glu
690 695 700
Lys Trp Asp Glu Val Tyr Lys Tyr Ile Val Thr Asn Trp Leu Ala Lys
705 710 715 720
Val Asn Thr Gln Ile Asp Leu Ile Arg Lys Lys Met Lys Glu Ala Leu
725 730 735
Glu Asn Gln Ala Glu Ala Thr Lys Ala Ile Ile Asn Tyr Gln Tyr Asn
740 745 750
Gln Tyr Thr Glu Glu Glu Lys Asn Asn Ile Asn Phe Asn Ile Asp Asp
755 760 765
Leu Ser Ser Lys Leu Asn Glu Ser Ile Asn Lys Ala Met Ile Asn Ile
770 775 780
Asn Lys Phe Leu Asn Gln Cys Ser Val Ser Tyr Leu Met Asn Ser Met
785 790 795 800
Ile Pro Tyr Gly Val Lys Arg Leu Glu Asp Phe Asp Ala Ser Leu Lys
805 810 815
Asp Ala Leu Leu Lys Tyr Ile Tyr Asp Asn Arg Gly Thr Leu Ile Gly
820 825 830
Gln Val Asp Arg Leu Lys Asp Lys Val Asn Asn Thr Leu Ser Thr Asp
835 840 845
Ile Pro Phe Gln Leu Ser Lys Tyr Val Asp Asn Gln Arg Leu Leu Ser
850 855 860
Thr Phe Thr Glu Tyr Ile Lys Asn Ile Ile Asn Thr Ser Ile Leu Asn
865 870 875 880
Leu Arg Tyr Glu Ser Asn His Leu Ile Asp Leu Ser Arg Tyr Ala Ser
885 890 895
Lys Ile Asn Ile Gly Ser Lys Val Asn Phe Asp Pro Ile Asp Lys Asn
900 905 910
Gln Ile Gln Leu Phe Asn Leu Glu Ser Ser Lys Ile Glu Val Ile Leu
915 920 925
Lys Asn Ala Ile Val Tyr Asn Ser Met Tyr Glu Asn Phe Ser Thr Ser
930 935 940
Phe Trp Ile Arg Ile Pro Lys Tyr Phe Asn Ser Ile Ser Leu Asn Asn
945 950 955 960
Glu Tyr Thr Ile Ile Asn Cys Met Glu Asn Asn Ser Gly Trp Lys Val
965 970 975
Ser Leu Asn Tyr Gly Glu Ile Ile Trp Thr Leu Gln Asp Thr Gln Glu
980 985 990
Ile Lys Gln Arg Val Val Phe Lys Tyr Ser Gln Met Ile Asn Ile Ser
995 1000 1005
Asp Tyr Ile Asn Arg Trp Ile Phe Val Thr Ile Thr Asn Asn Arg
1010 1015 1020
Leu Asn Asn Ser Lys Ile Tyr Ile Asn Gly Arg Leu Ile Asp Gln
1025 1030 1035
Lys Pro Ile Ser Asn Leu Gly Asn Ile His Ala Ser Asn Asn Ile
1040 1045 1050
Met Phe Lys Leu Asp Gly Cys Arg Asp Thr His Arg Tyr Ile Trp
1055 1060 1065
Ile Lys Tyr Phe Asn Leu Phe Asp Lys Glu Leu Asn Glu Lys Glu
1070 1075 1080
Ile Lys Asp Leu Tyr Asp Asn Gln Ser Asn Ser Gly Ile Leu Lys
1085 1090 1095
Asp Phe Trp Gly Asp Tyr Leu Gln Tyr Asp Lys Pro Tyr Tyr Met
1100 1105 1110
Leu Asn Leu Tyr Asp Pro Asn Lys Tyr Val Asp Val Asn Asn Val
1115 1120 1125
Gly Ile Arg Gly Tyr Met Tyr Leu Lys Gly Pro Arg Gly Ser Val
1130 1135 1140
Met Thr Thr Asn Ile Tyr Leu Asn Ser Ser Leu Tyr Arg Gly Thr
1145 1150 1155
Lys Phe Ile Ile Lys Lys Tyr Ala Ser Gly Asn Lys Asp Asn Ile
1160 1165 1170
Val Arg Asn Asn Asp Arg Val Tyr Ile Asn Val Val Val Lys Asn
1175 1180 1185
Lys Glu Tyr Arg Leu Ala Thr Asn Ala Ser Gln Ala Gly Val Glu
1190 1195 1200
Lys Ile Leu Ser Ala Leu Glu Ile Pro Asp Val Gly Asn Leu Ser
1205 1210 1215
Gln Val Val Val Met Lys Ser Lys Asn Asp Gln Gly Ile Thr Asn
1220 1225 1230
Lys Cys Lys Met Asn Leu Gln Asp Asn Asn Gly Asn Asp Ile Gly
1235 1240 1245
Phe Ile Gly Phe His Gln Phe Asn Asn Ile Ala Lys Leu Val Ala
1250 1255 1260
Ser Asn Trp Tyr Asn Arg Gln Ile Glu Arg Ser Ser Arg Thr Leu
1265 1270 1275
Gly Cys Ser Trp Glu Phe Ile Pro Val Asp Asp Gly Trp Gly Glu
1280 1285 1290
Arg Pro Leu Glu Asn Leu Tyr Phe Gln Gly Ala Ser His His His
1295 1300 1305
His His His His His
1310
<210> 19
<211> 2649
<212> DNA
<213> 人工序列
<220>
<223> rLHN/A的核苷酸序列 (His-标签的)
<400> 19
atgccgtttg tgaacaagca gttcaactat aaagatccgg ttaatggtgt ggatatcgcc 60
tatatcaaaa ttccgaatgc aggtcagatg cagccggtta aagcctttaa aatccataac 120
aaaatttggg tgattccgga acgtgatacc tttaccaatc cggaagaagg tgatctgaat 180
ccgcctccgg aagcaaaaca ggttccggtt agctattatg atagcaccta tctgagcacc 240
gataacgaga aagataacta tctgaaaggt gtgaccaaac tgtttgaacg catttatagt 300
accgatctgg gtcgtatgct gctgaccagc attgttcgtg gtattccgtt ttggggtggt 360
agcaccattg ataccgaact gaaagttatt gacaccaact gcattaatgt gattcagccg 420
gatggtagct atcgtagcga agaactgaat ctggttatta ttggtccgag cgcagatatc 480
attcagtttg aatgtaaatc ctttggccac gaagttctga atctgacccg taatggttat 540
ggtagtaccc agtatattcg tttcagtccg gattttacct ttggctttga agaaagcctg 600
gaagttgata caaatccgct gttaggtgca ggtaaatttg caaccgatcc ggcagttacc 660
ctggcacatg aactgattca tgccggtcat cgtctgtatg gtattgcaat taatccgaac 720
cgtgtgttca aagtgaatac caacgcatat tatgaaatga gcggtctgga agtgtcattt 780
gaagaactgc gtacctttgg tggtcatgat gccaaattta tcgatagcct gcaagaaaat 840
gaatttcgcc tgtactacta taacaaattc aaggatattg cgagcaccct gaataaagcc 900
aaaagcattg ttggcaccac cgcaagcctg cagtatatga aaaatgtgtt taaagaaaaa 960
tatctgctga gcgaagatac cagcggtaaa tttagcgttg acaaactgaa attcgataaa 1020
ctgtacaaga tgctgaccga gatttatacc gaagataact tcgtgaagtt tttcaaagtg 1080
ctgaaccgca aaacctacct gaactttgat aaagccgtgt tcaaaatcaa catcgtgccg 1140
aaagtgaact ataccatcta tgatggtttt aacctgcgca ataccaatct ggcagcaaac 1200
tttaatggtc agaacaccga aatcaacaac atgaacttta ccaaactgaa gaacttcacc 1260
ggtctgttcg aattttacaa actgctgtgt gttcgtggca ttattaccag caaaaccaaa 1320
agtctggata aaggctacaa taaagccctg aatgatctgt gcattaaggt gaataattgg 1380
gacctgtttt ttagcccgag cgaggataat ttcaccaacg atctgaacaa aggcgaagaa 1440
attaccagcg ataccaatat tgaagcagcc gaagaaaaca ttagcctgga tctgattcag 1500
cagtattatc tgaccttcaa cttcgataat gagccggaaa atatcagcat tgaaaacctg 1560
agcagcgata ttattggcca gctggaactg atgccgaata ttgaacgttt tccgaacggc 1620
aaaaaatacg agctggataa atacaccatg ttccattatc tgcgtgccca agaatttgaa 1680
catggtaaaa gccgtattgc actgaccaat agcgttaatg aagcactgct gaacccgagc 1740
cgtgtttata ccttttttag cagcgattac gtgaaaaagg ttaacaaagc aaccgaagca 1800
gccatgtttt taggttgggt tgaacagctg gtttatgatt tcaccgatga aaccagcgaa 1860
gttagcacca ccgataaaat tgcagatatt accatcatca tcccgtatat cggtccggca 1920
ctgaatattg gcaatatgct gtataaagac gattttgtgg gtgccctgat ttttagcggt 1980
gcagttattc tgctggaatt tattccggaa attgccattc cggttctggg cacctttgca 2040
ctggtgagct atattgcaaa taaagttctg accgtgcaga ccatcgataa tgcactgagc 2100
aaacgtaacg aaaaatggga tgaagtgtac aagtatatcg tgaccaattg gctggcaaaa 2160
gttaacaccc agattgacct gattcgcaag aagatgaaag aagcactgga aaatcaggca 2220
gaagcaacca aagccattat caactatcag tataaccagt acaccgaaga agagaaaaat 2280
aacatcaact tcaacatcga cgatctgtcc agcaaactga acgaaagcat caacaaagcc 2340
atgattaaca ttaacaaatt tctgaaccag tgcagcgtga gctatctgat gaatagcatg 2400
attccgtatg gtgtgaaacg tctggaagat tttgatgcaa gcctgaaaga tgccctgctg 2460
aaatatatct atgataatcg tggcaccctg attggtcagg ttgatcgtct gaaagataaa 2520
gtgaacaaca ccctgagtac cgatattcct tttcagctga gcaaatatgt ggataatcag 2580
cgtctgctgt caaccgaaaa tctgtatttc cagggtgcaa gtcatcatca ccatcaccac 2640
catcattaa 2649
<210> 20
<211> 882
<212> PRT
<213> 人工序列
<220>
<223> rLHN/A的多肽序列 (His-标签的)
<400> 20
Met Pro Phe Val Asn Lys Gln Phe Asn Tyr Lys Asp Pro Val Asn Gly
1 5 10 15
Val Asp Ile Ala Tyr Ile Lys Ile Pro Asn Ala Gly Gln Met Gln Pro
20 25 30
Val Lys Ala Phe Lys Ile His Asn Lys Ile Trp Val Ile Pro Glu Arg
35 40 45
Asp Thr Phe Thr Asn Pro Glu Glu Gly Asp Leu Asn Pro Pro Pro Glu
50 55 60
Ala Lys Gln Val Pro Val Ser Tyr Tyr Asp Ser Thr Tyr Leu Ser Thr
65 70 75 80
Asp Asn Glu Lys Asp Asn Tyr Leu Lys Gly Val Thr Lys Leu Phe Glu
85 90 95
Arg Ile Tyr Ser Thr Asp Leu Gly Arg Met Leu Leu Thr Ser Ile Val
100 105 110
Arg Gly Ile Pro Phe Trp Gly Gly Ser Thr Ile Asp Thr Glu Leu Lys
115 120 125
Val Ile Asp Thr Asn Cys Ile Asn Val Ile Gln Pro Asp Gly Ser Tyr
130 135 140
Arg Ser Glu Glu Leu Asn Leu Val Ile Ile Gly Pro Ser Ala Asp Ile
145 150 155 160
Ile Gln Phe Glu Cys Lys Ser Phe Gly His Glu Val Leu Asn Leu Thr
165 170 175
Arg Asn Gly Tyr Gly Ser Thr Gln Tyr Ile Arg Phe Ser Pro Asp Phe
180 185 190
Thr Phe Gly Phe Glu Glu Ser Leu Glu Val Asp Thr Asn Pro Leu Leu
195 200 205
Gly Ala Gly Lys Phe Ala Thr Asp Pro Ala Val Thr Leu Ala His Glu
210 215 220
Leu Ile His Ala Gly His Arg Leu Tyr Gly Ile Ala Ile Asn Pro Asn
225 230 235 240
Arg Val Phe Lys Val Asn Thr Asn Ala Tyr Tyr Glu Met Ser Gly Leu
245 250 255
Glu Val Ser Phe Glu Glu Leu Arg Thr Phe Gly Gly His Asp Ala Lys
260 265 270
Phe Ile Asp Ser Leu Gln Glu Asn Glu Phe Arg Leu Tyr Tyr Tyr Asn
275 280 285
Lys Phe Lys Asp Ile Ala Ser Thr Leu Asn Lys Ala Lys Ser Ile Val
290 295 300
Gly Thr Thr Ala Ser Leu Gln Tyr Met Lys Asn Val Phe Lys Glu Lys
305 310 315 320
Tyr Leu Leu Ser Glu Asp Thr Ser Gly Lys Phe Ser Val Asp Lys Leu
325 330 335
Lys Phe Asp Lys Leu Tyr Lys Met Leu Thr Glu Ile Tyr Thr Glu Asp
340 345 350
Asn Phe Val Lys Phe Phe Lys Val Leu Asn Arg Lys Thr Tyr Leu Asn
355 360 365
Phe Asp Lys Ala Val Phe Lys Ile Asn Ile Val Pro Lys Val Asn Tyr
370 375 380
Thr Ile Tyr Asp Gly Phe Asn Leu Arg Asn Thr Asn Leu Ala Ala Asn
385 390 395 400
Phe Asn Gly Gln Asn Thr Glu Ile Asn Asn Met Asn Phe Thr Lys Leu
405 410 415
Lys Asn Phe Thr Gly Leu Phe Glu Phe Tyr Lys Leu Leu Cys Val Arg
420 425 430
Gly Ile Ile Thr Ser Lys Thr Lys Ser Leu Asp Lys Gly Tyr Asn Lys
435 440 445
Ala Leu Asn Asp Leu Cys Ile Lys Val Asn Asn Trp Asp Leu Phe Phe
450 455 460
Ser Pro Ser Glu Asp Asn Phe Thr Asn Asp Leu Asn Lys Gly Glu Glu
465 470 475 480
Ile Thr Ser Asp Thr Asn Ile Glu Ala Ala Glu Glu Asn Ile Ser Leu
485 490 495
Asp Leu Ile Gln Gln Tyr Tyr Leu Thr Phe Asn Phe Asp Asn Glu Pro
500 505 510
Glu Asn Ile Ser Ile Glu Asn Leu Ser Ser Asp Ile Ile Gly Gln Leu
515 520 525
Glu Leu Met Pro Asn Ile Glu Arg Phe Pro Asn Gly Lys Lys Tyr Glu
530 535 540
Leu Asp Lys Tyr Thr Met Phe His Tyr Leu Arg Ala Gln Glu Phe Glu
545 550 555 560
His Gly Lys Ser Arg Ile Ala Leu Thr Asn Ser Val Asn Glu Ala Leu
565 570 575
Leu Asn Pro Ser Arg Val Tyr Thr Phe Phe Ser Ser Asp Tyr Val Lys
580 585 590
Lys Val Asn Lys Ala Thr Glu Ala Ala Met Phe Leu Gly Trp Val Glu
595 600 605
Gln Leu Val Tyr Asp Phe Thr Asp Glu Thr Ser Glu Val Ser Thr Thr
610 615 620
Asp Lys Ile Ala Asp Ile Thr Ile Ile Ile Pro Tyr Ile Gly Pro Ala
625 630 635 640
Leu Asn Ile Gly Asn Met Leu Tyr Lys Asp Asp Phe Val Gly Ala Leu
645 650 655
Ile Phe Ser Gly Ala Val Ile Leu Leu Glu Phe Ile Pro Glu Ile Ala
660 665 670
Ile Pro Val Leu Gly Thr Phe Ala Leu Val Ser Tyr Ile Ala Asn Lys
675 680 685
Val Leu Thr Val Gln Thr Ile Asp Asn Ala Leu Ser Lys Arg Asn Glu
690 695 700
Lys Trp Asp Glu Val Tyr Lys Tyr Ile Val Thr Asn Trp Leu Ala Lys
705 710 715 720
Val Asn Thr Gln Ile Asp Leu Ile Arg Lys Lys Met Lys Glu Ala Leu
725 730 735
Glu Asn Gln Ala Glu Ala Thr Lys Ala Ile Ile Asn Tyr Gln Tyr Asn
740 745 750
Gln Tyr Thr Glu Glu Glu Lys Asn Asn Ile Asn Phe Asn Ile Asp Asp
755 760 765
Leu Ser Ser Lys Leu Asn Glu Ser Ile Asn Lys Ala Met Ile Asn Ile
770 775 780
Asn Lys Phe Leu Asn Gln Cys Ser Val Ser Tyr Leu Met Asn Ser Met
785 790 795 800
Ile Pro Tyr Gly Val Lys Arg Leu Glu Asp Phe Asp Ala Ser Leu Lys
805 810 815
Asp Ala Leu Leu Lys Tyr Ile Tyr Asp Asn Arg Gly Thr Leu Ile Gly
820 825 830
Gln Val Asp Arg Leu Lys Asp Lys Val Asn Asn Thr Leu Ser Thr Asp
835 840 845
Ile Pro Phe Gln Leu Ser Lys Tyr Val Asp Asn Gln Arg Leu Leu Ser
850 855 860
Thr Glu Asn Leu Tyr Phe Gln Gly Ala Ser His His His His His His
865 870 875 880
His His
<210> 21
<211> 1323
<212> DNA
<213> 人工序列
<220>
<223> rHC/A的核苷酸序列 (His-标签的)
<400> 21
atgcatcatc accatcacca cgaaaatcta tacttccaag gaaaaaacat catcaatact 60
agcattctga acctgcgtta cgagagcaat catctgattg atctgagccg ttatgcaagc 120
aagatcaaca tcggtagcaa ggtcaatttt gacccgatcg ataagaacca gatccagctg 180
tttaatctgg aatcgagcaa aattgaggtt atcctgaaaa acgccattgt ctacaactcc 240
atgtacgaga atttctccac cagcttctgg attcgcatcc cgaaatactt caacagcatt 300
agcctgaaca acgagtatac tatcatcaac tgtatggaga acaacagcgg ttggaaggtg 360
tctctgaact atggtgagat catttggacc ttgcaggaca cccaagagat caagcagcgc 420
gtcgtgttca agtactctca aatgatcaac atttccgatt acattaatcg ttggatcttc 480
gtgaccatta cgaataaccg tctgaataac agcaagattt acatcaatgg tcgcttgatc 540
gatcagaaac cgattagcaa cctgggtaat atccacgcaa gcaacaacat tatgttcaaa 600
ttggacggtt gccgcgatac ccatcgttat atctggatca agtatttcaa cctgtttgat 660
aaagaactga atgagaagga gatcaaagat ttgtatgaca accaatctaa cagcggcatt 720
ttgaaggact tctggggcga ttatctgcaa tacgataagc cgtactatat gctgaacctg 780
tatgatccga acaaatatgt ggatgtcaat aatgtgggta ttcgtggtta catgtatttg 840
aagggtccgc gtggcagcgt tatgacgacc aacatttacc tgaactctag cctgtaccgt 900
ggtacgaaat tcatcattaa gaaatatgcc agcggcaaca aagataacat tgtgcgtaat 960
aacgatcgtg tctacatcaa cgtggtcgtg aagaataaag agtaccgtct ggcgaccaac 1020
gcttcgcagg cgggtgttga gaaaattctg agcgcgttgg agatccctga tgtcggtaat 1080
ctgagccaag tcgtggttat gaagagcaag aacgaccagg gtatcactaa caagtgcaag 1140
atgaacctgc aagacaacaa tggtaacgac atcggcttta ttggtttcca ccagttcaac 1200
aatattgcta aactggtagc gagcaattgg tacaatcgtc agattgagcg cagcagccgt 1260
actttgggct gtagctggga gtttatcccg gtcgatgatg gttggggcga acgtccgctg 1320
taa 1323
<210> 22
<211> 440
<212> PRT
<213> 人工序列
<220>
<223> rHC/A的多肽序列 (His-标签的)
<400> 22
Met His His His His His His Glu Asn Leu Tyr Phe Gln Gly Lys Asn
1 5 10 15
Ile Ile Asn Thr Ser Ile Leu Asn Leu Arg Tyr Glu Ser Asn His Leu
20 25 30
Ile Asp Leu Ser Arg Tyr Ala Ser Lys Ile Asn Ile Gly Ser Lys Val
35 40 45
Asn Phe Asp Pro Ile Asp Lys Asn Gln Ile Gln Leu Phe Asn Leu Glu
50 55 60
Ser Ser Lys Ile Glu Val Ile Leu Lys Asn Ala Ile Val Tyr Asn Ser
65 70 75 80
Met Tyr Glu Asn Phe Ser Thr Ser Phe Trp Ile Arg Ile Pro Lys Tyr
85 90 95
Phe Asn Ser Ile Ser Leu Asn Asn Glu Tyr Thr Ile Ile Asn Cys Met
100 105 110
Glu Asn Asn Ser Gly Trp Lys Val Ser Leu Asn Tyr Gly Glu Ile Ile
115 120 125
Trp Thr Leu Gln Asp Thr Gln Glu Ile Lys Gln Arg Val Val Phe Lys
130 135 140
Tyr Ser Gln Met Ile Asn Ile Ser Asp Tyr Ile Asn Arg Trp Ile Phe
145 150 155 160
Val Thr Ile Thr Asn Asn Arg Leu Asn Asn Ser Lys Ile Tyr Ile Asn
165 170 175
Gly Arg Leu Ile Asp Gln Lys Pro Ile Ser Asn Leu Gly Asn Ile His
180 185 190
Ala Ser Asn Asn Ile Met Phe Lys Leu Asp Gly Cys Arg Asp Thr His
195 200 205
Arg Tyr Ile Trp Ile Lys Tyr Phe Asn Leu Phe Asp Lys Glu Leu Asn
210 215 220
Glu Lys Glu Ile Lys Asp Leu Tyr Asp Asn Gln Ser Asn Ser Gly Ile
225 230 235 240
Leu Lys Asp Phe Trp Gly Asp Tyr Leu Gln Tyr Asp Lys Pro Tyr Tyr
245 250 255
Met Leu Asn Leu Tyr Asp Pro Asn Lys Tyr Val Asp Val Asn Asn Val
260 265 270
Gly Ile Arg Gly Tyr Met Tyr Leu Lys Gly Pro Arg Gly Ser Val Met
275 280 285
Thr Thr Asn Ile Tyr Leu Asn Ser Ser Leu Tyr Arg Gly Thr Lys Phe
290 295 300
Ile Ile Lys Lys Tyr Ala Ser Gly Asn Lys Asp Asn Ile Val Arg Asn
305 310 315 320
Asn Asp Arg Val Tyr Ile Asn Val Val Val Lys Asn Lys Glu Tyr Arg
325 330 335
Leu Ala Thr Asn Ala Ser Gln Ala Gly Val Glu Lys Ile Leu Ser Ala
340 345 350
Leu Glu Ile Pro Asp Val Gly Asn Leu Ser Gln Val Val Val Met Lys
355 360 365
Ser Lys Asn Asp Gln Gly Ile Thr Asn Lys Cys Lys Met Asn Leu Gln
370 375 380
Asp Asn Asn Gly Asn Asp Ile Gly Phe Ile Gly Phe His Gln Phe Asn
385 390 395 400
Asn Ile Ala Lys Leu Val Ala Ser Asn Trp Tyr Asn Arg Gln Ile Glu
405 410 415
Arg Ser Ser Arg Thr Leu Gly Cys Ser Trp Glu Phe Ile Pro Val Asp
420 425 430
Asp Gly Trp Gly Glu Arg Pro Leu
435 440
<210> 23
<211> 1326
<212> DNA
<213> 人工序列
<220>
<223> rLC/A的核苷酸序列 (His-标签的)
<400> 23
atgccgtttg tgaacaagca gttcaactat aaagatccgg ttaatggtgt ggatatcgcc 60
tatatcaaaa ttccgaatgc aggtcagatg cagccggtta aagcctttaa aatccataac 120
aaaatttggg tgattccgga acgtgatacc tttaccaatc cggaagaagg tgatctgaat 180
ccgcctccgg aagcaaaaca ggttccggtt agctattatg atagcaccta tctgagcacc 240
gataacgaga aagataacta tctgaaaggt gtgaccaaac tgtttgaacg catttatagt 300
accgatctgg gtcgtatgct gctgaccagc attgttcgtg gtattccgtt ttggggtggt 360
agcaccattg ataccgaact gaaagttatt gacaccaact gcattaatgt gattcagccg 420
gatggtagct atcgtagcga agaactgaat ctggttatta ttggtccgag cgcagatatc 480
attcagtttg aatgtaaatc ctttggccac gaagttctga atctgacccg taatggttat 540
ggtagtaccc agtatattcg tttcagtccg gattttacct ttggctttga agaaagcctg 600
gaagttgata caaatccgct gttaggtgca ggtaaatttg caaccgatcc ggcagttacc 660
ctggcacatg aactgattca tgccggtcat cgtctgtatg gtattgcaat taatccgaac 720
cgtgtgttca aagtgaatac caacgcatat tatgaaatga gcggtctgga agtgtcattt 780
gaagaactgc gtacctttgg tggtcatgat gccaaattta tcgatagcct gcaagaaaat 840
gaatttcgcc tgtactacta taacaaattc aaggatattg cgagcaccct gaataaagcc 900
aaaagcattg ttggcaccac cgcaagcctg cagtatatga aaaatgtgtt taaagaaaaa 960
tatctgctga gcgaagatac cagcggtaaa tttagcgttg acaaactgaa attcgataaa 1020
ctgtacaaga tgctgaccga gatttatacc gaagataact tcgtgaagtt tttcaaagtg 1080
ctgaaccgca aaacctacct gaactttgat aaagccgtgt tcaaaatcaa catcgtgccg 1140
aaagtgaact ataccatcta tgatggtttt aacctgcgca ataccaatct ggcagcaaac 1200
tttaatggtc agaacaccga aatcaacaac atgaacttta ccaaactgaa gaacttcacc 1260
ggtctgtttg aagagaatct gtatttccag ggtgcaagtc atcatcacca tcaccaccat 1320
cattaa 1326
<210> 24
<211> 441
<212> PRT
<213> 人工序列
<220>
<223> rLC/A的多肽序列 (His-标签的)
<400> 24
Met Pro Phe Val Asn Lys Gln Phe Asn Tyr Lys Asp Pro Val Asn Gly
1 5 10 15
Val Asp Ile Ala Tyr Ile Lys Ile Pro Asn Ala Gly Gln Met Gln Pro
20 25 30
Val Lys Ala Phe Lys Ile His Asn Lys Ile Trp Val Ile Pro Glu Arg
35 40 45
Asp Thr Phe Thr Asn Pro Glu Glu Gly Asp Leu Asn Pro Pro Pro Glu
50 55 60
Ala Lys Gln Val Pro Val Ser Tyr Tyr Asp Ser Thr Tyr Leu Ser Thr
65 70 75 80
Asp Asn Glu Lys Asp Asn Tyr Leu Lys Gly Val Thr Lys Leu Phe Glu
85 90 95
Arg Ile Tyr Ser Thr Asp Leu Gly Arg Met Leu Leu Thr Ser Ile Val
100 105 110
Arg Gly Ile Pro Phe Trp Gly Gly Ser Thr Ile Asp Thr Glu Leu Lys
115 120 125
Val Ile Asp Thr Asn Cys Ile Asn Val Ile Gln Pro Asp Gly Ser Tyr
130 135 140
Arg Ser Glu Glu Leu Asn Leu Val Ile Ile Gly Pro Ser Ala Asp Ile
145 150 155 160
Ile Gln Phe Glu Cys Lys Ser Phe Gly His Glu Val Leu Asn Leu Thr
165 170 175
Arg Asn Gly Tyr Gly Ser Thr Gln Tyr Ile Arg Phe Ser Pro Asp Phe
180 185 190
Thr Phe Gly Phe Glu Glu Ser Leu Glu Val Asp Thr Asn Pro Leu Leu
195 200 205
Gly Ala Gly Lys Phe Ala Thr Asp Pro Ala Val Thr Leu Ala His Glu
210 215 220
Leu Ile His Ala Gly His Arg Leu Tyr Gly Ile Ala Ile Asn Pro Asn
225 230 235 240
Arg Val Phe Lys Val Asn Thr Asn Ala Tyr Tyr Glu Met Ser Gly Leu
245 250 255
Glu Val Ser Phe Glu Glu Leu Arg Thr Phe Gly Gly His Asp Ala Lys
260 265 270
Phe Ile Asp Ser Leu Gln Glu Asn Glu Phe Arg Leu Tyr Tyr Tyr Asn
275 280 285
Lys Phe Lys Asp Ile Ala Ser Thr Leu Asn Lys Ala Lys Ser Ile Val
290 295 300
Gly Thr Thr Ala Ser Leu Gln Tyr Met Lys Asn Val Phe Lys Glu Lys
305 310 315 320
Tyr Leu Leu Ser Glu Asp Thr Ser Gly Lys Phe Ser Val Asp Lys Leu
325 330 335
Lys Phe Asp Lys Leu Tyr Lys Met Leu Thr Glu Ile Tyr Thr Glu Asp
340 345 350
Asn Phe Val Lys Phe Phe Lys Val Leu Asn Arg Lys Thr Tyr Leu Asn
355 360 365
Phe Asp Lys Ala Val Phe Lys Ile Asn Ile Val Pro Lys Val Asn Tyr
370 375 380
Thr Ile Tyr Asp Gly Phe Asn Leu Arg Asn Thr Asn Leu Ala Ala Asn
385 390 395 400
Phe Asn Gly Gln Asn Thr Glu Ile Asn Asn Met Asn Phe Thr Lys Leu
405 410 415
Lys Asn Phe Thr Gly Leu Phe Glu Glu Asn Leu Tyr Phe Gln Gly Ala
420 425 430
Ser His His His His His His His His
435 440
<210> 25
<211> 3918
<212> DNA
<213> 人工序列
<220>
<223> rBoNT/FA(0)的核苷酸序列 (His-标签的)
<400> 25
atgccggttg tgattaacag cttcaattat gatgatccgg tgaacgataa caccatcatt 60
tatatccgtc cgccttatta tgaaaccagc aacacctatt tcaaagcctt ccagattatg 120
gataacgtgt ggattattcc ggaacgttat cgtctgggta ttgatccgag cctgtttaat 180
ccgcctgtta gcctgaaagc aggtagtgat ggttattttg atccgaatta tctgagcacc 240
aacaccgaga aaaacaaata cctgcagatt atgatcaagc tgttcaaacg cattaatagc 300
aaaccggcag gtcagattct gctggaagaa atcaaaaatg caattccgta tctgggcaac 360
agctataccc aagaagaaca gtttaccacc aataatcgta ccgtgagctt taatgttaaa 420
ctggccaatg gtaatatcgt tcagcagatg gcaaatctga ttatttgggg tccgggtcct 480
gatctgacca caaataaaac cggtggtatc atctatagcc cgtatcagag catggaagca 540
accccgtata aagatggttt tggtagcatt atgaccgtgg aatttagtcc ggaatatgca 600
accgccttta acgatatttc aattgcaagc catagtccgt cgctgtttat caaagatccg 660
gcactgattc tgatgcacca gctgatttat gttctgcatg gtctgtatgg cacctatatc 720
accgaataca aaattacccc gaatgtggtt cagagctata tgaaagttac caaaccgatt 780
accagcgcag aatttctgac ctttggtggt cgtgatcgca atattgttcc gcagagcatt 840
cagagccagc tgtataacaa agttctgagc gattataaac gtattgccag ccgtctgaat 900
aaagttaata ccgcaaccgc actgatcaac atcgatgaat tcaaaaacct gtacgagtgg 960
aaataccagt ttgccaaaga tagcaatggt gtgtatagcg tggatctgaa caaatttgag 1020
cagctgtaca aaaaaatcta tagcttcacc gaattcaacc tggcctatga gtttaaaatc 1080
aaaacccgtc tgggttatct ggccgaaaat tttggtccgt tttatctgcc gaatctgctg 1140
gatgatagca tttataccga agtggatggt tttaacattg gtgcactgag cattaactat 1200
cagggtcaga atattggcag cgatatcaac agcatcaaaa aactgcaagg tcagggtgtt 1260
gttagccgtg ttgttcgtct gtgtagcaat agcaatacca aaaacagcct gtgcattacc 1320
gttaataatc gcgacctgtt ttttatcgca agccaagaaa gctatggcga gaataccatt 1380
aacacctata aagagattga cgataccacc acactggatc cgagctttga agatattctg 1440
gataaagtga tcctgaactt caacgaacag gttattccgc agatgccgaa tcgtaatgtt 1500
agcaccgata ttcagaaaga caactacatc ccgaaatacg attataaccg caccgacatt 1560
atcgatagct atgaagttgg tcgcaactac aacacctttt tctatctgaa tgcccagaaa 1620
tttagcccga acgaaagcaa tattaccctg accagcagct ttgatacagg tctgttagaa 1680
ggtagcaaag tgtatacctt tttcagcagc gatttcatta acaacatcaa caaaccggtt 1740
caggccctgc tgtttattga atgggttaaa caggtgattc gcgattttac caccgaagca 1800
accaaaacct caaccgttga taaactgaaa gatattagcc tggtggtgcc gtatattggt 1860
ctggcactga atattggtga tgagatctac aaacagcatt ttgcagaagc agttgaactg 1920
gttggtgcag gtctgctgct ggaattttca ccggaatttc ttattccgac gctgctgatt 1980
tttaccatca aaggttatct gaccggtagc attcgcgata aagacaaaat cattaaaacc 2040
ctggataacg ccctgaatgt tcgtgatcag aaatggaaag aactgtatcg ttgggttgtt 2100
agcaaatggc tgaccaccat taatacgcag ttcaacaaac gcaaagaaca aatgtacaaa 2160
gccctgaaaa atcaggccac cgccattaaa aagatcatcg agaacaaata taacaactat 2220
accaccgatg aaaaaagcaa gatcgatagc agctataaca tcaacgaaat tgaacgcacc 2280
ctgaacgaaa aaatcaatct ggccatgaaa aacatcgagc agtttattac cgaaagcagc 2340
attgcctatc tgatcaatat catcaacaac gaaacgatcc agaaactgaa aagctatgat 2400
gacctggttc gtcgttatct gctgggttat attcgtaatc atagcagcat tctgggcaat 2460
agcgttgaag aactgaattc caaagtgaac aaccatctgg ataatggcat tccgtttgaa 2520
ctgagcagtt ataccaatga tagcctgctg atccgctact tcaataaaaa ctatggcgaa 2580
ctgaagtaca actgcattct gaacatcaaa tatgagatgg atcgtgacaa actggttgat 2640
agcagcggtt atcgtagccg tatcaatatt ggtacaggcg tcaaatttag cgagatcgat 2700
aaaaatcaag tgcagctgag caatctggaa tccagcaaaa ttgaagtcat tctgaataac 2760
ggcgtcatct ataacagcat gtatgaaaac ttttcgacca gcttttggat tcgcattccg 2820
aaatactttc gcaacatcaa taacgagtac aagatcatca gctgtatgca gaataatagc 2880
ggttgggaag tgagcctgaa ttttagcaat atgaactcga aaatcatctg gaccctgcag 2940
gataccgaag gtatcaaaaa aaccgttgtg tttcagtaca cccagaacat taacattagc 3000
gactatatca accgctggat ctttgtgacc attacaaata atcgtctgag caacagcaaa 3060
atctacatta atggtcgcct gatcaacgaa gaaagcatta gcgatctggg taatatccat 3120
gccagcaaca acattatgtt taaactggat ggttgccgtg atccgcatcg ttatatctgg 3180
attaaatact ttaacctgtt tgacaaagag ctgaacaaga aagaaattaa agatctgtac 3240
gacaaccaga gcaatagcgg tattctgaaa gatttctggg gtgattatct gcagtatgac 3300
aaaccgtatt atatgctgaa tctgtatgac ccgaataagt atctggatgt gaataatgtt 3360
ggcatccgtg gctatatgta tctgaaaggt ccgcgtggtc gtattgtgac caccaacatt 3420
tatctgaata gcaccctgta tatgggcacc aaattcatca ttaagaaata tgccagcggc 3480
aacaaagata acattgtgcg taataatgat cgcgtgtata ttaacgtggt ggtgaagaat 3540
aaagaatatc gcctggcaac caatgcaagc caggcaggcg ttgaaaaaat tctgagcgca 3600
gttgaaatcc cggatgttgg taatctgagc caggttgttg tgatgaaaag cgaaaatgat 3660
cagggcattc gcaacaagtg taaaatgaat ctgcaagaca ataacggcaa cgatattggc 3720
tttatcggct ttcaccagtt taataacatt gcaaaactgg tggccagcaa ctggtataac 3780
cgtcagattg gtaaagcaag ccgtaccttt ggttgtagct gggaatttat cccggttgat 3840
gatggttggg gtgaaagcag cctggaaaat ctgtatttcc agggtgccag tcatcatcac 3900
caccatcacc atcactga 3918
<210> 26
<211> 1305
<212> PRT
<213> 人工序列
<220>
<223> rBoNT/FA(0)的多肽序列 (His-标签的)
<400> 26
Met Pro Val Val Ile Asn Ser Phe Asn Tyr Asp Asp Pro Val Asn Asp
1 5 10 15
Asn Thr Ile Ile Tyr Ile Arg Pro Pro Tyr Tyr Glu Thr Ser Asn Thr
20 25 30
Tyr Phe Lys Ala Phe Gln Ile Met Asp Asn Val Trp Ile Ile Pro Glu
35 40 45
Arg Tyr Arg Leu Gly Ile Asp Pro Ser Leu Phe Asn Pro Pro Val Ser
50 55 60
Leu Lys Ala Gly Ser Asp Gly Tyr Phe Asp Pro Asn Tyr Leu Ser Thr
65 70 75 80
Asn Thr Glu Lys Asn Lys Tyr Leu Gln Ile Met Ile Lys Leu Phe Lys
85 90 95
Arg Ile Asn Ser Lys Pro Ala Gly Gln Ile Leu Leu Glu Glu Ile Lys
100 105 110
Asn Ala Ile Pro Tyr Leu Gly Asn Ser Tyr Thr Gln Glu Glu Gln Phe
115 120 125
Thr Thr Asn Asn Arg Thr Val Ser Phe Asn Val Lys Leu Ala Asn Gly
130 135 140
Asn Ile Val Gln Gln Met Ala Asn Leu Ile Ile Trp Gly Pro Gly Pro
145 150 155 160
Asp Leu Thr Thr Asn Lys Thr Gly Gly Ile Ile Tyr Ser Pro Tyr Gln
165 170 175
Ser Met Glu Ala Thr Pro Tyr Lys Asp Gly Phe Gly Ser Ile Met Thr
180 185 190
Val Glu Phe Ser Pro Glu Tyr Ala Thr Ala Phe Asn Asp Ile Ser Ile
195 200 205
Ala Ser His Ser Pro Ser Leu Phe Ile Lys Asp Pro Ala Leu Ile Leu
210 215 220
Met His Gln Leu Ile Tyr Val Leu His Gly Leu Tyr Gly Thr Tyr Ile
225 230 235 240
Thr Glu Tyr Lys Ile Thr Pro Asn Val Val Gln Ser Tyr Met Lys Val
245 250 255
Thr Lys Pro Ile Thr Ser Ala Glu Phe Leu Thr Phe Gly Gly Arg Asp
260 265 270
Arg Asn Ile Val Pro Gln Ser Ile Gln Ser Gln Leu Tyr Asn Lys Val
275 280 285
Leu Ser Asp Tyr Lys Arg Ile Ala Ser Arg Leu Asn Lys Val Asn Thr
290 295 300
Ala Thr Ala Leu Ile Asn Ile Asp Glu Phe Lys Asn Leu Tyr Glu Trp
305 310 315 320
Lys Tyr Gln Phe Ala Lys Asp Ser Asn Gly Val Tyr Ser Val Asp Leu
325 330 335
Asn Lys Phe Glu Gln Leu Tyr Lys Lys Ile Tyr Ser Phe Thr Glu Phe
340 345 350
Asn Leu Ala Tyr Glu Phe Lys Ile Lys Thr Arg Leu Gly Tyr Leu Ala
355 360 365
Glu Asn Phe Gly Pro Phe Tyr Leu Pro Asn Leu Leu Asp Asp Ser Ile
370 375 380
Tyr Thr Glu Val Asp Gly Phe Asn Ile Gly Ala Leu Ser Ile Asn Tyr
385 390 395 400
Gln Gly Gln Asn Ile Gly Ser Asp Ile Asn Ser Ile Lys Lys Leu Gln
405 410 415
Gly Gln Gly Val Val Ser Arg Val Val Arg Leu Cys Ser Asn Ser Asn
420 425 430
Thr Lys Asn Ser Leu Cys Ile Thr Val Asn Asn Arg Asp Leu Phe Phe
435 440 445
Ile Ala Ser Gln Glu Ser Tyr Gly Glu Asn Thr Ile Asn Thr Tyr Lys
450 455 460
Glu Ile Asp Asp Thr Thr Thr Leu Asp Pro Ser Phe Glu Asp Ile Leu
465 470 475 480
Asp Lys Val Ile Leu Asn Phe Asn Glu Gln Val Ile Pro Gln Met Pro
485 490 495
Asn Arg Asn Val Ser Thr Asp Ile Gln Lys Asp Asn Tyr Ile Pro Lys
500 505 510
Tyr Asp Tyr Asn Arg Thr Asp Ile Ile Asp Ser Tyr Glu Val Gly Arg
515 520 525
Asn Tyr Asn Thr Phe Phe Tyr Leu Asn Ala Gln Lys Phe Ser Pro Asn
530 535 540
Glu Ser Asn Ile Thr Leu Thr Ser Ser Phe Asp Thr Gly Leu Leu Glu
545 550 555 560
Gly Ser Lys Val Tyr Thr Phe Phe Ser Ser Asp Phe Ile Asn Asn Ile
565 570 575
Asn Lys Pro Val Gln Ala Leu Leu Phe Ile Glu Trp Val Lys Gln Val
580 585 590
Ile Arg Asp Phe Thr Thr Glu Ala Thr Lys Thr Ser Thr Val Asp Lys
595 600 605
Leu Lys Asp Ile Ser Leu Val Val Pro Tyr Ile Gly Leu Ala Leu Asn
610 615 620
Ile Gly Asp Glu Ile Tyr Lys Gln His Phe Ala Glu Ala Val Glu Leu
625 630 635 640
Val Gly Ala Gly Leu Leu Leu Glu Phe Ser Pro Glu Phe Leu Ile Pro
645 650 655
Thr Leu Leu Ile Phe Thr Ile Lys Gly Tyr Leu Thr Gly Ser Ile Arg
660 665 670
Asp Lys Asp Lys Ile Ile Lys Thr Leu Asp Asn Ala Leu Asn Val Arg
675 680 685
Asp Gln Lys Trp Lys Glu Leu Tyr Arg Trp Val Val Ser Lys Trp Leu
690 695 700
Thr Thr Ile Asn Thr Gln Phe Asn Lys Arg Lys Glu Gln Met Tyr Lys
705 710 715 720
Ala Leu Lys Asn Gln Ala Thr Ala Ile Lys Lys Ile Ile Glu Asn Lys
725 730 735
Tyr Asn Asn Tyr Thr Thr Asp Glu Lys Ser Lys Ile Asp Ser Ser Tyr
740 745 750
Asn Ile Asn Glu Ile Glu Arg Thr Leu Asn Glu Lys Ile Asn Leu Ala
755 760 765
Met Lys Asn Ile Glu Gln Phe Ile Thr Glu Ser Ser Ile Ala Tyr Leu
770 775 780
Ile Asn Ile Ile Asn Asn Glu Thr Ile Gln Lys Leu Lys Ser Tyr Asp
785 790 795 800
Asp Leu Val Arg Arg Tyr Leu Leu Gly Tyr Ile Arg Asn His Ser Ser
805 810 815
Ile Leu Gly Asn Ser Val Glu Glu Leu Asn Ser Lys Val Asn Asn His
820 825 830
Leu Asp Asn Gly Ile Pro Phe Glu Leu Ser Ser Tyr Thr Asn Asp Ser
835 840 845
Leu Leu Ile Arg Tyr Phe Asn Lys Asn Tyr Gly Glu Leu Lys Tyr Asn
850 855 860
Cys Ile Leu Asn Ile Lys Tyr Glu Met Asp Arg Asp Lys Leu Val Asp
865 870 875 880
Ser Ser Gly Tyr Arg Ser Arg Ile Asn Ile Gly Thr Gly Val Lys Phe
885 890 895
Ser Glu Ile Asp Lys Asn Gln Val Gln Leu Ser Asn Leu Glu Ser Ser
900 905 910
Lys Ile Glu Val Ile Leu Asn Asn Gly Val Ile Tyr Asn Ser Met Tyr
915 920 925
Glu Asn Phe Ser Thr Ser Phe Trp Ile Arg Ile Pro Lys Tyr Phe Arg
930 935 940
Asn Ile Asn Asn Glu Tyr Lys Ile Ile Ser Cys Met Gln Asn Asn Ser
945 950 955 960
Gly Trp Glu Val Ser Leu Asn Phe Ser Asn Met Asn Ser Lys Ile Ile
965 970 975
Trp Thr Leu Gln Asp Thr Glu Gly Ile Lys Lys Thr Val Val Phe Gln
980 985 990
Tyr Thr Gln Asn Ile Asn Ile Ser Asp Tyr Ile Asn Arg Trp Ile Phe
995 1000 1005
Val Thr Ile Thr Asn Asn Arg Leu Ser Asn Ser Lys Ile Tyr Ile
1010 1015 1020
Asn Gly Arg Leu Ile Asn Glu Glu Ser Ile Ser Asp Leu Gly Asn
1025 1030 1035
Ile His Ala Ser Asn Asn Ile Met Phe Lys Leu Asp Gly Cys Arg
1040 1045 1050
Asp Pro His Arg Tyr Ile Trp Ile Lys Tyr Phe Asn Leu Phe Asp
1055 1060 1065
Lys Glu Leu Asn Lys Lys Glu Ile Lys Asp Leu Tyr Asp Asn Gln
1070 1075 1080
Ser Asn Ser Gly Ile Leu Lys Asp Phe Trp Gly Asp Tyr Leu Gln
1085 1090 1095
Tyr Asp Lys Pro Tyr Tyr Met Leu Asn Leu Tyr Asp Pro Asn Lys
1100 1105 1110
Tyr Leu Asp Val Asn Asn Val Gly Ile Arg Gly Tyr Met Tyr Leu
1115 1120 1125
Lys Gly Pro Arg Gly Arg Ile Val Thr Thr Asn Ile Tyr Leu Asn
1130 1135 1140
Ser Thr Leu Tyr Met Gly Thr Lys Phe Ile Ile Lys Lys Tyr Ala
1145 1150 1155
Ser Gly Asn Lys Asp Asn Ile Val Arg Asn Asn Asp Arg Val Tyr
1160 1165 1170
Ile Asn Val Val Val Lys Asn Lys Glu Tyr Arg Leu Ala Thr Asn
1175 1180 1185
Ala Ser Gln Ala Gly Val Glu Lys Ile Leu Ser Ala Val Glu Ile
1190 1195 1200
Pro Asp Val Gly Asn Leu Ser Gln Val Val Val Met Lys Ser Glu
1205 1210 1215
Asn Asp Gln Gly Ile Arg Asn Lys Cys Lys Met Asn Leu Gln Asp
1220 1225 1230
Asn Asn Gly Asn Asp Ile Gly Phe Ile Gly Phe His Gln Phe Asn
1235 1240 1245
Asn Ile Ala Lys Leu Val Ala Ser Asn Trp Tyr Asn Arg Gln Ile
1250 1255 1260
Gly Lys Ala Ser Arg Thr Phe Gly Cys Ser Trp Glu Phe Ile Pro
1265 1270 1275
Val Asp Asp Gly Trp Gly Glu Ser Ser Leu Glu Asn Leu Tyr Phe
1280 1285 1290
Gln Gly Ala Ser His His His His His His His His
1295 1300 1305
<210> 27
<211> 2634
<212> DNA
<213> 人工序列
<220>
<223> rLHN/FA的核苷酸序列 (His-标签的)
<400> 27
atgccggttg tgattaacag cttcaattat gatgatccgg tgaacgataa caccatcatt 60
tatatccgtc cgccttatta tgaaaccagc aacacctatt tcaaagcctt ccagattatg 120
gataacgtgt ggattattcc ggaacgttat cgtctgggta ttgatccgag cctgtttaat 180
ccgcctgtta gcctgaaagc aggtagtgat ggttattttg atccgaatta tctgagcacc 240
aacaccgaga aaaacaaata cctgcagatt atgatcaagc tgttcaaacg cattaatagc 300
aaaccggcag gtcagattct gctggaagaa atcaaaaatg caattccgta tctgggcaac 360
agctataccc aagaagaaca gtttaccacc aataatcgta ccgtgagctt taatgttaaa 420
ctggccaatg gtaatatcgt tcagcagatg gcaaatctga ttatttgggg tccgggtcct 480
gatctgacca caaataaaac cggtggtatc atctatagcc cgtatcagag catggaagca 540
accccgtata aagatggttt tggtagcatt atgaccgtgg aatttagtcc ggaatatgca 600
accgccttta acgatatttc aattgcaagc catagtccgt cgctgtttat caaagatccg 660
gcactgattc tgatgcatga actgattcat gttctgcatg gtctgtatgg cacctatatt 720
accgaataca aaattacccc gaatgtggtg cagagctata tgaaagttac caaaccgatt 780
accagcgcag aatttctgac ctttggtggt cgtgatcgca atattgttcc gcagagcatt 840
cagagccagc tgtataacaa agttctgagc gattataaac gtattgccag ccgtctgaat 900
aaagttaata ccgcaaccgc actgatcaac atcgatgaat tcaaaaacct gtacgagtgg 960
aaataccagt ttgccaaaga tagcaatggt gtgtatagcg tggatctgaa caaatttgag 1020
cagctgtaca aaaaaatcta tagcttcacc gaattcaacc tggcctatga gtttaaaatc 1080
aaaacccgtc tgggttatct ggccgaaaat tttggtccgt tttatctgcc gaatctgctg 1140
gatgatagca tttataccga agtggatggt tttaacattg gtgcactgag cattaactat 1200
cagggtcaga atattggcag cgatatcaac agcatcaaaa aactgcaagg tcagggtgtt 1260
gttagccgtg ttgttcgtct gtgtagcaat agcaatacca aaaacagcct gtgcattacc 1320
gttaataatc gcgacctgtt ttttatcgca agccaagaaa gctatggcga gaataccatt 1380
aacacctata aagagattga cgataccacc acactggatc cgagctttga agatattctg 1440
gataaagtga tcctgaactt caacgaacag gttattccgc agatgccgaa tcgtaatgtt 1500
agcaccgata ttcagaaaga caactacatc ccgaaatacg attataaccg caccgacatt 1560
atcgatagct atgaagttgg tcgcaactac aacacctttt tctatctgaa tgcccagaaa 1620
tttagcccga acgaaagcaa tattaccctg accagcagct ttgatacagg tctgttagaa 1680
ggtagcaaag tgtatacctt tttcagcagc gatttcatta acaacatcaa caaaccggtt 1740
caggccctgc tgtttattga atgggttaaa caggtgattc gcgattttac caccgaagca 1800
accaaaacct caaccgttga taaactgaaa gatattagcc tggtggtgcc gtatattggt 1860
ctggcactga atattggtga tgagatctac aaacagcatt ttgcagaagc agttgaactg 1920
gttggtgcag gtctgctgct ggaattttca ccggaatttc ttattccgac gctgctgatt 1980
tttaccatca aaggttatct gaccggtagc attcgcgata aagacaaaat cattaaaacc 2040
ctggataacg ccctgaatgt tcgtgatcag aaatggaaag aactgtatcg ttgggttgtt 2100
agcaaatggc tgaccaccat taatacgcag ttcaacaaac gcaaagaaca aatgtacaaa 2160
gccctgaaaa atcaggccac cgccattaaa aagatcatcg agaacaaata taacaactat 2220
accaccgatg aaaaaagcaa gatcgatagc agctataaca tcaacgaaat tgaacgcacc 2280
ctgaacgaaa aaatcaatct ggccatgaaa aacatcgagc agtttattac agaaagcagc 2340
attgcctacc tgatcaatat catcaacaac gaaaccattc agaaactgaa aagctatgat 2400
gacctggttc gtcgttatct gctgggttat attcgtaatc atagcagcat tctgggcaat 2460
agcgttgaag aactgaattc caaagtgaac aaccatctgg ataatggcat tccgtttgaa 2520
ctgagcagtt ataccaatga tagcctgctg atccgctact tcaataaaaa ctatggcgaa 2580
gagaacctgt atttccaggg tgccagtcat catcaccacc atcaccatca ctga 2634
<210> 28
<211> 877
<212> PRT
<213> 人工序列
<220>
<223> rLHN/FA的多肽序列 (His-标签的)
<400> 28
Met Pro Val Val Ile Asn Ser Phe Asn Tyr Asp Asp Pro Val Asn Asp
1 5 10 15
Asn Thr Ile Ile Tyr Ile Arg Pro Pro Tyr Tyr Glu Thr Ser Asn Thr
20 25 30
Tyr Phe Lys Ala Phe Gln Ile Met Asp Asn Val Trp Ile Ile Pro Glu
35 40 45
Arg Tyr Arg Leu Gly Ile Asp Pro Ser Leu Phe Asn Pro Pro Val Ser
50 55 60
Leu Lys Ala Gly Ser Asp Gly Tyr Phe Asp Pro Asn Tyr Leu Ser Thr
65 70 75 80
Asn Thr Glu Lys Asn Lys Tyr Leu Gln Ile Met Ile Lys Leu Phe Lys
85 90 95
Arg Ile Asn Ser Lys Pro Ala Gly Gln Ile Leu Leu Glu Glu Ile Lys
100 105 110
Asn Ala Ile Pro Tyr Leu Gly Asn Ser Tyr Thr Gln Glu Glu Gln Phe
115 120 125
Thr Thr Asn Asn Arg Thr Val Ser Phe Asn Val Lys Leu Ala Asn Gly
130 135 140
Asn Ile Val Gln Gln Met Ala Asn Leu Ile Ile Trp Gly Pro Gly Pro
145 150 155 160
Asp Leu Thr Thr Asn Lys Thr Gly Gly Ile Ile Tyr Ser Pro Tyr Gln
165 170 175
Ser Met Glu Ala Thr Pro Tyr Lys Asp Gly Phe Gly Ser Ile Met Thr
180 185 190
Val Glu Phe Ser Pro Glu Tyr Ala Thr Ala Phe Asn Asp Ile Ser Ile
195 200 205
Ala Ser His Ser Pro Ser Leu Phe Ile Lys Asp Pro Ala Leu Ile Leu
210 215 220
Met His Glu Leu Ile His Val Leu His Gly Leu Tyr Gly Thr Tyr Ile
225 230 235 240
Thr Glu Tyr Lys Ile Thr Pro Asn Val Val Gln Ser Tyr Met Lys Val
245 250 255
Thr Lys Pro Ile Thr Ser Ala Glu Phe Leu Thr Phe Gly Gly Arg Asp
260 265 270
Arg Asn Ile Val Pro Gln Ser Ile Gln Ser Gln Leu Tyr Asn Lys Val
275 280 285
Leu Ser Asp Tyr Lys Arg Ile Ala Ser Arg Leu Asn Lys Val Asn Thr
290 295 300
Ala Thr Ala Leu Ile Asn Ile Asp Glu Phe Lys Asn Leu Tyr Glu Trp
305 310 315 320
Lys Tyr Gln Phe Ala Lys Asp Ser Asn Gly Val Tyr Ser Val Asp Leu
325 330 335
Asn Lys Phe Glu Gln Leu Tyr Lys Lys Ile Tyr Ser Phe Thr Glu Phe
340 345 350
Asn Leu Ala Tyr Glu Phe Lys Ile Lys Thr Arg Leu Gly Tyr Leu Ala
355 360 365
Glu Asn Phe Gly Pro Phe Tyr Leu Pro Asn Leu Leu Asp Asp Ser Ile
370 375 380
Tyr Thr Glu Val Asp Gly Phe Asn Ile Gly Ala Leu Ser Ile Asn Tyr
385 390 395 400
Gln Gly Gln Asn Ile Gly Ser Asp Ile Asn Ser Ile Lys Lys Leu Gln
405 410 415
Gly Gln Gly Val Val Ser Arg Val Val Arg Leu Cys Ser Asn Ser Asn
420 425 430
Thr Lys Asn Ser Leu Cys Ile Thr Val Asn Asn Arg Asp Leu Phe Phe
435 440 445
Ile Ala Ser Gln Glu Ser Tyr Gly Glu Asn Thr Ile Asn Thr Tyr Lys
450 455 460
Glu Ile Asp Asp Thr Thr Thr Leu Asp Pro Ser Phe Glu Asp Ile Leu
465 470 475 480
Asp Lys Val Ile Leu Asn Phe Asn Glu Gln Val Ile Pro Gln Met Pro
485 490 495
Asn Arg Asn Val Ser Thr Asp Ile Gln Lys Asp Asn Tyr Ile Pro Lys
500 505 510
Tyr Asp Tyr Asn Arg Thr Asp Ile Ile Asp Ser Tyr Glu Val Gly Arg
515 520 525
Asn Tyr Asn Thr Phe Phe Tyr Leu Asn Ala Gln Lys Phe Ser Pro Asn
530 535 540
Glu Ser Asn Ile Thr Leu Thr Ser Ser Phe Asp Thr Gly Leu Leu Glu
545 550 555 560
Gly Ser Lys Val Tyr Thr Phe Phe Ser Ser Asp Phe Ile Asn Asn Ile
565 570 575
Asn Lys Pro Val Gln Ala Leu Leu Phe Ile Glu Trp Val Lys Gln Val
580 585 590
Ile Arg Asp Phe Thr Thr Glu Ala Thr Lys Thr Ser Thr Val Asp Lys
595 600 605
Leu Lys Asp Ile Ser Leu Val Val Pro Tyr Ile Gly Leu Ala Leu Asn
610 615 620
Ile Gly Asp Glu Ile Tyr Lys Gln His Phe Ala Glu Ala Val Glu Leu
625 630 635 640
Val Gly Ala Gly Leu Leu Leu Glu Phe Ser Pro Glu Phe Leu Ile Pro
645 650 655
Thr Leu Leu Ile Phe Thr Ile Lys Gly Tyr Leu Thr Gly Ser Ile Arg
660 665 670
Asp Lys Asp Lys Ile Ile Lys Thr Leu Asp Asn Ala Leu Asn Val Arg
675 680 685
Asp Gln Lys Trp Lys Glu Leu Tyr Arg Trp Val Val Ser Lys Trp Leu
690 695 700
Thr Thr Ile Asn Thr Gln Phe Asn Lys Arg Lys Glu Gln Met Tyr Lys
705 710 715 720
Ala Leu Lys Asn Gln Ala Thr Ala Ile Lys Lys Ile Ile Glu Asn Lys
725 730 735
Tyr Asn Asn Tyr Thr Thr Asp Glu Lys Ser Lys Ile Asp Ser Ser Tyr
740 745 750
Asn Ile Asn Glu Ile Glu Arg Thr Leu Asn Glu Lys Ile Asn Leu Ala
755 760 765
Met Lys Asn Ile Glu Gln Phe Ile Thr Glu Ser Ser Ile Ala Tyr Leu
770 775 780
Ile Asn Ile Ile Asn Asn Glu Thr Ile Gln Lys Leu Lys Ser Tyr Asp
785 790 795 800
Asp Leu Val Arg Arg Tyr Leu Leu Gly Tyr Ile Arg Asn His Ser Ser
805 810 815
Ile Leu Gly Asn Ser Val Glu Glu Leu Asn Ser Lys Val Asn Asn His
820 825 830
Leu Asp Asn Gly Ile Pro Phe Glu Leu Ser Ser Tyr Thr Asn Asp Ser
835 840 845
Leu Leu Ile Arg Tyr Phe Asn Lys Asn Tyr Gly Glu Glu Asn Leu Tyr
850 855 860
Phe Gln Gly Ala Ser His His His His His His His His
865 870 875
<210> 29
<211> 1341
<212> DNA
<213> 人工序列
<220>
<223> rHC/FA的核苷酸序列 (His-标签的)
<400> 29
atgctgaagt ataactgcat cctgaacatc aaatatgaga tggatcgtga taaactggtt 60
gatagcagcg gttatcgtag ccgtatcaat attggcaccg gtgtgaaatt tagcgagatc 120
gataaaaatc aggtgcagct gagcaatctg gaaagcagca aaattgaagt gattctgaat 180
aacggcgtga tctacaatag catgtatgaa aacttttcga ccagcttctg gattcgcatt 240
ccgaaatact ttcgcaacat caacaacgag tacaagatta tcagctgtat gcagaataat 300
agcggttggg aagttagcct gaatttcagc aatatgaaca gcaaaatcat ttggaccctg 360
caggataccg aaggtatcaa aaaaaccgtt gtgtttcagt acacccagaa cattaacatc 420
agcgattaca ttaaccgctg gatctttgtg accattacca ataatcgtct gagcaacagc 480
aagatctata ttaacggtcg cctgattaac gaagagagca ttagcgatct gggtaatatt 540
catgccagca acaacatcat gtttaaactg gatggttgtc gtgatccgca tcgttatatt 600
tggatcaaat acttcaacct gtttgataaa gaactgaaca aaaaagaaat caaagacctg 660
tatgataacc agagcaatag cggcattctg aaagattttt ggggtgatta tctgcagtat 720
gacaaaccgt attacatgct gaatctgtac gatccgaaca aatatctgga tgtgaataat 780
gtgggtatcc gtggctatat gtatctgaaa ggtccgcgtg gtcgtattgt taccaccaac 840
atttatctga atagcaccct gtatatgggc accaaattca tcattaaaaa gtatgccagc 900
ggcaacaaag ataacattgt gcgtaataat gatcgcgtgt atatcaatgt ggtggtgaag 960
aataaagaat atcgtctggc caccaatgca agccaggcag gcgttgaaaa aattctgagc 1020
gcagttgaaa ttccggatgt tggtaatctg agccaggttg ttgttatgaa aagcgaaaat 1080
gatcagggca ttcgcaacaa atgcaaaatg aatctgcagg acaataacgg caacgatatt 1140
ggttttattg gcttccacca gttcaacaac attgcaaaac tggtggcgag caattggtat 1200
aatcgtcaga ttggtaaagc aagccgtacc tttggttgta gctgggaatt tattccggtt 1260
gatgatggtt ggggtgaaag cagcctggaa aatctgtatt ttcagggtgc aagtcatcat 1320
caccaccatc accatcatta a 1341
<210> 30
<211> 446
<212> PRT
<213> 人工序列
<220>
<223> rHC/FA的多肽序列 (His-标签的)
<400> 30
Met Leu Lys Tyr Asn Cys Ile Leu Asn Ile Lys Tyr Glu Met Asp Arg
1 5 10 15
Asp Lys Leu Val Asp Ser Ser Gly Tyr Arg Ser Arg Ile Asn Ile Gly
20 25 30
Thr Gly Val Lys Phe Ser Glu Ile Asp Lys Asn Gln Val Gln Leu Ser
35 40 45
Asn Leu Glu Ser Ser Lys Ile Glu Val Ile Leu Asn Asn Gly Val Ile
50 55 60
Tyr Asn Ser Met Tyr Glu Asn Phe Ser Thr Ser Phe Trp Ile Arg Ile
65 70 75 80
Pro Lys Tyr Phe Arg Asn Ile Asn Asn Glu Tyr Lys Ile Ile Ser Cys
85 90 95
Met Gln Asn Asn Ser Gly Trp Glu Val Ser Leu Asn Phe Ser Asn Met
100 105 110
Asn Ser Lys Ile Ile Trp Thr Leu Gln Asp Thr Glu Gly Ile Lys Lys
115 120 125
Thr Val Val Phe Gln Tyr Thr Gln Asn Ile Asn Ile Ser Asp Tyr Ile
130 135 140
Asn Arg Trp Ile Phe Val Thr Ile Thr Asn Asn Arg Leu Ser Asn Ser
145 150 155 160
Lys Ile Tyr Ile Asn Gly Arg Leu Ile Asn Glu Glu Ser Ile Ser Asp
165 170 175
Leu Gly Asn Ile His Ala Ser Asn Asn Ile Met Phe Lys Leu Asp Gly
180 185 190
Cys Arg Asp Pro His Arg Tyr Ile Trp Ile Lys Tyr Phe Asn Leu Phe
195 200 205
Asp Lys Glu Leu Asn Lys Lys Glu Ile Lys Asp Leu Tyr Asp Asn Gln
210 215 220
Ser Asn Ser Gly Ile Leu Lys Asp Phe Trp Gly Asp Tyr Leu Gln Tyr
225 230 235 240
Asp Lys Pro Tyr Tyr Met Leu Asn Leu Tyr Asp Pro Asn Lys Tyr Leu
245 250 255
Asp Val Asn Asn Val Gly Ile Arg Gly Tyr Met Tyr Leu Lys Gly Pro
260 265 270
Arg Gly Arg Ile Val Thr Thr Asn Ile Tyr Leu Asn Ser Thr Leu Tyr
275 280 285
Met Gly Thr Lys Phe Ile Ile Lys Lys Tyr Ala Ser Gly Asn Lys Asp
290 295 300
Asn Ile Val Arg Asn Asn Asp Arg Val Tyr Ile Asn Val Val Val Lys
305 310 315 320
Asn Lys Glu Tyr Arg Leu Ala Thr Asn Ala Ser Gln Ala Gly Val Glu
325 330 335
Lys Ile Leu Ser Ala Val Glu Ile Pro Asp Val Gly Asn Leu Ser Gln
340 345 350
Val Val Val Met Lys Ser Glu Asn Asp Gln Gly Ile Arg Asn Lys Cys
355 360 365
Lys Met Asn Leu Gln Asp Asn Asn Gly Asn Asp Ile Gly Phe Ile Gly
370 375 380
Phe His Gln Phe Asn Asn Ile Ala Lys Leu Val Ala Ser Asn Trp Tyr
385 390 395 400
Asn Arg Gln Ile Gly Lys Ala Ser Arg Thr Phe Gly Cys Ser Trp Glu
405 410 415
Phe Ile Pro Val Asp Asp Gly Trp Gly Glu Ser Ser Leu Glu Asn Leu
420 425 430
Tyr Phe Gln Gly Ala Ser His His His His His His His His
435 440 445
<210> 31
<211> 1347
<212> DNA
<213> 人工序列
<220>
<223> rLC/FA的核苷酸序列 (His-标签的)
<400> 31
atgccggttg tgattaacag cttcaattat gatgatccgg tgaacgataa caccatcatt 60
tatatccgtc cgccttatta tgaaaccagc aacacctatt tcaaagcctt ccagattatg 120
gataacgtgt ggattattcc ggaacgttat cgtctgggta ttgatccgag cctgtttaat 180
ccgcctgtta gcctgaaagc aggtagtgat ggttattttg atccgaatta tctgagcacc 240
aacaccgaga aaaacaaata cctgcagatt atgatcaagc tgttcaaacg cattaatagc 300
aaaccggcag gtcagattct gctggaagaa atcaaaaatg caattccgta tctgggcaac 360
agctataccc aagaagaaca gtttaccacc aataatcgta ccgtgagctt taatgttaaa 420
ctggccaatg gtaatatcgt tcagcagatg gcaaatctga ttatttgggg tccgggtcct 480
gatctgacca caaataaaac cggtggtatc atctatagcc cgtatcagag catggaagca 540
accccgtata aagatggttt tggtagcatt atgaccgtgg aatttagtcc ggaatatgca 600
accgccttta acgatatttc aattgcaagc catagtccgt cgctgtttat caaagatccg 660
gcactgattc tgatgcatga actgattcat gttctgcatg gtctgtatgg cacctatatt 720
accgaataca aaattacccc gaatgtggtg cagagctata tgaaagttac caaaccgatt 780
accagcgcag aatttctgac ctttggtggt cgtgatcgca atattgttcc gcagagcatt 840
cagagccagc tgtataacaa agttctgagc gattataaac gtattgccag ccgtctgaat 900
aaagttaata ccgcaaccgc actgatcaac atcgatgaat tcaaaaacct gtacgagtgg 960
aaataccagt ttgccaaaga tagcaatggt gtgtatagcg tggatctgaa caaatttgag 1020
cagctgtaca aaaaaatcta tagcttcacc gaattcaacc tggcctatga gtttaaaatc 1080
aaaacccgtc tgggttatct ggccgaaaat tttggtccgt tttatctgcc gaatctgctg 1140
gatgatagca tttataccga agtggatggt tttaacattg gtgcactgag cattaactat 1200
cagggtcaga atattggcag cgatatcaac agcatcaaaa aactgcaagg tcagggtgtt 1260
gttagccgtg ttgttcgtct gtgtagcaat agcgaaaatc tgtattttca gggtgccagt 1320
catcatcacc accatcacca tcactga 1347
<210> 32
<211> 448
<212> PRT
<213> 人工序列
<220>
<223> rLC/FA的多肽序列 (His-标签的)
<400> 32
Met Pro Val Val Ile Asn Ser Phe Asn Tyr Asp Asp Pro Val Asn Asp
1 5 10 15
Asn Thr Ile Ile Tyr Ile Arg Pro Pro Tyr Tyr Glu Thr Ser Asn Thr
20 25 30
Tyr Phe Lys Ala Phe Gln Ile Met Asp Asn Val Trp Ile Ile Pro Glu
35 40 45
Arg Tyr Arg Leu Gly Ile Asp Pro Ser Leu Phe Asn Pro Pro Val Ser
50 55 60
Leu Lys Ala Gly Ser Asp Gly Tyr Phe Asp Pro Asn Tyr Leu Ser Thr
65 70 75 80
Asn Thr Glu Lys Asn Lys Tyr Leu Gln Ile Met Ile Lys Leu Phe Lys
85 90 95
Arg Ile Asn Ser Lys Pro Ala Gly Gln Ile Leu Leu Glu Glu Ile Lys
100 105 110
Asn Ala Ile Pro Tyr Leu Gly Asn Ser Tyr Thr Gln Glu Glu Gln Phe
115 120 125
Thr Thr Asn Asn Arg Thr Val Ser Phe Asn Val Lys Leu Ala Asn Gly
130 135 140
Asn Ile Val Gln Gln Met Ala Asn Leu Ile Ile Trp Gly Pro Gly Pro
145 150 155 160
Asp Leu Thr Thr Asn Lys Thr Gly Gly Ile Ile Tyr Ser Pro Tyr Gln
165 170 175
Ser Met Glu Ala Thr Pro Tyr Lys Asp Gly Phe Gly Ser Ile Met Thr
180 185 190
Val Glu Phe Ser Pro Glu Tyr Ala Thr Ala Phe Asn Asp Ile Ser Ile
195 200 205
Ala Ser His Ser Pro Ser Leu Phe Ile Lys Asp Pro Ala Leu Ile Leu
210 215 220
Met His Glu Leu Ile His Val Leu His Gly Leu Tyr Gly Thr Tyr Ile
225 230 235 240
Thr Glu Tyr Lys Ile Thr Pro Asn Val Val Gln Ser Tyr Met Lys Val
245 250 255
Thr Lys Pro Ile Thr Ser Ala Glu Phe Leu Thr Phe Gly Gly Arg Asp
260 265 270
Arg Asn Ile Val Pro Gln Ser Ile Gln Ser Gln Leu Tyr Asn Lys Val
275 280 285
Leu Ser Asp Tyr Lys Arg Ile Ala Ser Arg Leu Asn Lys Val Asn Thr
290 295 300
Ala Thr Ala Leu Ile Asn Ile Asp Glu Phe Lys Asn Leu Tyr Glu Trp
305 310 315 320
Lys Tyr Gln Phe Ala Lys Asp Ser Asn Gly Val Tyr Ser Val Asp Leu
325 330 335
Asn Lys Phe Glu Gln Leu Tyr Lys Lys Ile Tyr Ser Phe Thr Glu Phe
340 345 350
Asn Leu Ala Tyr Glu Phe Lys Ile Lys Thr Arg Leu Gly Tyr Leu Ala
355 360 365
Glu Asn Phe Gly Pro Phe Tyr Leu Pro Asn Leu Leu Asp Asp Ser Ile
370 375 380
Tyr Thr Glu Val Asp Gly Phe Asn Ile Gly Ala Leu Ser Ile Asn Tyr
385 390 395 400
Gln Gly Gln Asn Ile Gly Ser Asp Ile Asn Ser Ile Lys Lys Leu Gln
405 410 415
Gly Gln Gly Val Val Ser Arg Val Val Arg Leu Cys Ser Asn Ser Glu
420 425 430
Asn Leu Tyr Phe Gln Gly Ala Ser His His His His His His His His
435 440 445
<210> 33
<211> 3888
<212> DNA
<213> 人工序列
<220>
<223> rBoNT/F(0)的核苷酸序列 (His-标签的)
<400> 33
atgccggttg tgattaacag cttcaattat aacgatccgg tgaacgatga taccatcctg 60
tatatgcaga ttccgtatga agagaaaagc aaaaagtact acaaagcctt tgagatcatg 120
cgcaacgttt ggattattcc ggaacgtaat accattggca ccgatccgag cgattttgat 180
ccgcctgcaa gcctggaaaa tggtagcagc gcatattatg atccgaatta tctgaccacc 240
gatgccgaaa aagatcgtta tctgaaaacc accatcaaac tgttcaaacg cattaatagc 300
aatccggcag gcgaagttct gctgcaagaa attagctatg caaaaccgta tctgggcaat 360
gaacataccc cgattaatga atttcatccg gttacacgta ccacgagcgt taacattaaa 420
agcagcacca atgtgaagtc cagcattatt ctgaatctgc tggttttagg tgcaggtccg 480
gatatttttg aaaattcaag ctatccggtg cgcaaactga tggatagcgg tggtgtgtat 540
gatccgtcaa atgatggttt tggcagcatt aacattgtga cctttagtcc ggaatatgaa 600
tacaccttca acgatattag cggtggctat aatagcagca ccgaaagttt tattgcagat 660
ccggcaatta gcctggcaca ccagctgatt tatgcactgc atggtctgta tggtgcacgt 720
ggtgttacct ataaagaaac cattaaagtt aaacaggcac cgctgatgat tgcggaaaaa 780
ccgattcgtc tggaagaatt tctgaccttt ggtggtcagg atctgaacat tattaccagc 840
gcaatgaaag agaaaatcta taataacctg ctggccaact atgagaaaat tgcaacccgt 900
ctgagccgtg ttaatagcgc acctcctgaa tatgatatca acgagtataa agactatttt 960
cagtggaaat acggcctgga taaaaatgca gatggtagct ataccgtgaa cgagaacaaa 1020
tttaacgaga tctacaaaaa actgtatagc ttcaccgaaa tcgatctggc caacaaattc 1080
aaagtgaaat gccgcaacac ctacttcatc aaatatggct ttctgaaagt tccgaacctg 1140
cttgatgatg atatctatac cgttagcgaa ggctttaaca ttggtaatct ggccgttaat 1200
aatcgcggtc agaacattaa actgaacccg aaaattatcg atagcatccc ggataaaggc 1260
ctggttgaaa aaattgtgaa attctgcaaa agcgtgattc cgcgtaaagg caccaaagca 1320
ccgcctcgtc tgtgtattcg tgtgaataat cgtgaactgt tttttgttgc aagcgagagc 1380
agctataacg agaatgatat taacaccccg aaagagattg acgataccac caatctgaat 1440
aacaactatc gcaacaatct ggatgaagtg atcctggatt ataacagcga aaccattccg 1500
cagattagca atcagaccct gaataccctg gttcaggatg atagctatgt tccgcgttat 1560
gatagcaatg gcaccagcga aattgaagaa cataatgtgg ttgatctgaa cgtgttcttt 1620
tatctgcatg cacagaaagt gccggaaggt gaaaccaata ttagcctgac cagcagcatt 1680
gataccgcac tgagcgaaga aagccaggtt tatacctttt ttagcagcga attcatcaac 1740
accattaaca aaccggttca tgcagcactg tttattagct ggattaatca ggtgattcgc 1800
gattttacca ccgaagcaac ccagaaaagc acctttgata aaattgccga tattagtctg 1860
gtggtgccgt atgttggtct ggcactgaat attggtaatg aagtgcagaa agagaacttt 1920
aaagaagcct tcgaactgtt aggtgccggt attctgctgg aatttgtgcc ggaactgctg 1980
attccgacca ttctggtttt taccattaag agctttattg gcagcagcga gaacaagaac 2040
aaaatcatta aagccatcaa caacagcctg atggaacgcg aaaccaaatg gaaagaaatt 2100
tacagctgga ttgtgagcaa ttggctgacc cgtatcaata cccagtttaa caaacgcaaa 2160
gaacaaatgt atcaggccct gcagaatcag gttgatgcaa ttaaaaccgt gatcgaatac 2220
aaatacaaca actataccag cgacgaacgt aatcgcctgg aaagcgaata caacattaat 2280
aacattcgcg aagaactgaa caaaaaagtg agcctggcaa tggaaaacat cgaacgtttt 2340
attaccgaaa gcagcatctt ctacctgatg aaactgatta acgaagccaa agttagcaaa 2400
ctgcgcgaat atgatgaagg cgttaaagaa tatctgctgg actatattag cgaacatcgt 2460
agcattctgg gtaatagcgt tcaagagctg aatgatctgg ttaccagcac actgaataat 2520
agcattccgt ttgaactgag cagctacacc aacgataaaa tcctgatcct gtacttcaac 2580
aaactgtaca agaagatcaa ggacaacagc atactggata tgcgctatga aaacaacaag 2640
ttcattgata tcagcggcta tggtagcaac attagcatta atggtgatgt gtatatctac 2700
agcaccaacc gcaatcagtt tggtatttat agcagcaaac cgagcgaagt taatattgcg 2760
cagaataacg atatcatcta caacggtcgc tatcagaact ttagcattag cttttgggtt 2820
cgcattccga aatactttaa caaggtgaac ctgaacaacg agtacaccat tattgattgc 2880
attcgcaata ataacagcgg ctggaaaatc agcctgaact ataacaaaat tatctggacc 2940
ctgcaggata ccgcaggtaa taatcagaaa ctggtgttta actacaccca gatgattagc 3000
atcagcgact atatcaacaa atggatcttt gtgaccatta ccaacaatcg tctgggtaac 3060
agccgcattt atatcaatgg caatctgatc gacgaaaaaa gcatttcaaa tctgggcgat 3120
attcacgtga gcgataacat tctgttcaaa attgttggct gcaacgatac ccgttatgtt 3180
ggtattcgtt acttcaaagt gtttgatacg gaactgggca aaacggaaat tgaaaccctg 3240
tatagtgatg aaccggatcc gagcattctg aaagattttt ggggtaatta tctgctgtac 3300
aacaaacgct actatctgct gaacctgctg cgtaccgata aaagcattac acagaatagc 3360
aactttctga acatcaatca gcagcgtggt gtttatcaga aaccgaacat ttttagcaac 3420
acccgtctgt ataccggtgt ggaagttatt attcgtaaaa acggtagcac cgatatcagc 3480
aacaccgata actttgtgcg taaaaatgac ctggcctata ttaacgttgt tgatcgtgat 3540
gttgagtatc gtctgtatgc ggatattagc attgccaaac cggaaaagat tatcaaactg 3600
atccgtacca gcaacagcaa taattcactg ggtcagatta tcgtgatgga cagcattggt 3660
aacaattgca ccatgaattt ccagaacaat aacggtggta atattggcct gctgggcttt 3720
catagcaata atctggttgc aagcagctgg tattacaaca acatccgtaa aaataccagc 3780
agtaatggtt gcttttggag ctttatcagt aaagaacatg gctggcaaga aaacgagaac 3840
ctgtattttc agggtgcaag tcatcatcac catcaccacc atcattaa 3888
<210> 34
<211> 1295
<212> PRT
<213> 人工序列
<220>
<223> rBoNT/F(0)的多肽序列 (His-标签的)
<400> 34
Met Pro Val Val Ile Asn Ser Phe Asn Tyr Asn Asp Pro Val Asn Asp
1 5 10 15
Asp Thr Ile Leu Tyr Met Gln Ile Pro Tyr Glu Glu Lys Ser Lys Lys
20 25 30
Tyr Tyr Lys Ala Phe Glu Ile Met Arg Asn Val Trp Ile Ile Pro Glu
35 40 45
Arg Asn Thr Ile Gly Thr Asp Pro Ser Asp Phe Asp Pro Pro Ala Ser
50 55 60
Leu Glu Asn Gly Ser Ser Ala Tyr Tyr Asp Pro Asn Tyr Leu Thr Thr
65 70 75 80
Asp Ala Glu Lys Asp Arg Tyr Leu Lys Thr Thr Ile Lys Leu Phe Lys
85 90 95
Arg Ile Asn Ser Asn Pro Ala Gly Glu Val Leu Leu Gln Glu Ile Ser
100 105 110
Tyr Ala Lys Pro Tyr Leu Gly Asn Glu His Thr Pro Ile Asn Glu Phe
115 120 125
His Pro Val Thr Arg Thr Thr Ser Val Asn Ile Lys Ser Ser Thr Asn
130 135 140
Val Lys Ser Ser Ile Ile Leu Asn Leu Leu Val Leu Gly Ala Gly Pro
145 150 155 160
Asp Ile Phe Glu Asn Ser Ser Tyr Pro Val Arg Lys Leu Met Asp Ser
165 170 175
Gly Gly Val Tyr Asp Pro Ser Asn Asp Gly Phe Gly Ser Ile Asn Ile
180 185 190
Val Thr Phe Ser Pro Glu Tyr Glu Tyr Thr Phe Asn Asp Ile Ser Gly
195 200 205
Gly Tyr Asn Ser Ser Thr Glu Ser Phe Ile Ala Asp Pro Ala Ile Ser
210 215 220
Leu Ala His Gln Leu Ile Tyr Ala Leu His Gly Leu Tyr Gly Ala Arg
225 230 235 240
Gly Val Thr Tyr Lys Glu Thr Ile Lys Val Lys Gln Ala Pro Leu Met
245 250 255
Ile Ala Glu Lys Pro Ile Arg Leu Glu Glu Phe Leu Thr Phe Gly Gly
260 265 270
Gln Asp Leu Asn Ile Ile Thr Ser Ala Met Lys Glu Lys Ile Tyr Asn
275 280 285
Asn Leu Leu Ala Asn Tyr Glu Lys Ile Ala Thr Arg Leu Ser Arg Val
290 295 300
Asn Ser Ala Pro Pro Glu Tyr Asp Ile Asn Glu Tyr Lys Asp Tyr Phe
305 310 315 320
Gln Trp Lys Tyr Gly Leu Asp Lys Asn Ala Asp Gly Ser Tyr Thr Val
325 330 335
Asn Glu Asn Lys Phe Asn Glu Ile Tyr Lys Lys Leu Tyr Ser Phe Thr
340 345 350
Glu Ile Asp Leu Ala Asn Lys Phe Lys Val Lys Cys Arg Asn Thr Tyr
355 360 365
Phe Ile Lys Tyr Gly Phe Leu Lys Val Pro Asn Leu Leu Asp Asp Asp
370 375 380
Ile Tyr Thr Val Ser Glu Gly Phe Asn Ile Gly Asn Leu Ala Val Asn
385 390 395 400
Asn Arg Gly Gln Asn Ile Lys Leu Asn Pro Lys Ile Ile Asp Ser Ile
405 410 415
Pro Asp Lys Gly Leu Val Glu Lys Ile Val Lys Phe Cys Lys Ser Val
420 425 430
Ile Pro Arg Lys Gly Thr Lys Ala Pro Pro Arg Leu Cys Ile Arg Val
435 440 445
Asn Asn Arg Glu Leu Phe Phe Val Ala Ser Glu Ser Ser Tyr Asn Glu
450 455 460
Asn Asp Ile Asn Thr Pro Lys Glu Ile Asp Asp Thr Thr Asn Leu Asn
465 470 475 480
Asn Asn Tyr Arg Asn Asn Leu Asp Glu Val Ile Leu Asp Tyr Asn Ser
485 490 495
Glu Thr Ile Pro Gln Ile Ser Asn Gln Thr Leu Asn Thr Leu Val Gln
500 505 510
Asp Asp Ser Tyr Val Pro Arg Tyr Asp Ser Asn Gly Thr Ser Glu Ile
515 520 525
Glu Glu His Asn Val Val Asp Leu Asn Val Phe Phe Tyr Leu His Ala
530 535 540
Gln Lys Val Pro Glu Gly Glu Thr Asn Ile Ser Leu Thr Ser Ser Ile
545 550 555 560
Asp Thr Ala Leu Ser Glu Glu Ser Gln Val Tyr Thr Phe Phe Ser Ser
565 570 575
Glu Phe Ile Asn Thr Ile Asn Lys Pro Val His Ala Ala Leu Phe Ile
580 585 590
Ser Trp Ile Asn Gln Val Ile Arg Asp Phe Thr Thr Glu Ala Thr Gln
595 600 605
Lys Ser Thr Phe Asp Lys Ile Ala Asp Ile Ser Leu Val Val Pro Tyr
610 615 620
Val Gly Leu Ala Leu Asn Ile Gly Asn Glu Val Gln Lys Glu Asn Phe
625 630 635 640
Lys Glu Ala Phe Glu Leu Leu Gly Ala Gly Ile Leu Leu Glu Phe Val
645 650 655
Pro Glu Leu Leu Ile Pro Thr Ile Leu Val Phe Thr Ile Lys Ser Phe
660 665 670
Ile Gly Ser Ser Glu Asn Lys Asn Lys Ile Ile Lys Ala Ile Asn Asn
675 680 685
Ser Leu Met Glu Arg Glu Thr Lys Trp Lys Glu Ile Tyr Ser Trp Ile
690 695 700
Val Ser Asn Trp Leu Thr Arg Ile Asn Thr Gln Phe Asn Lys Arg Lys
705 710 715 720
Glu Gln Met Tyr Gln Ala Leu Gln Asn Gln Val Asp Ala Ile Lys Thr
725 730 735
Val Ile Glu Tyr Lys Tyr Asn Asn Tyr Thr Ser Asp Glu Arg Asn Arg
740 745 750
Leu Glu Ser Glu Tyr Asn Ile Asn Asn Ile Arg Glu Glu Leu Asn Lys
755 760 765
Lys Val Ser Leu Ala Met Glu Asn Ile Glu Arg Phe Ile Thr Glu Ser
770 775 780
Ser Ile Phe Tyr Leu Met Lys Leu Ile Asn Glu Ala Lys Val Ser Lys
785 790 795 800
Leu Arg Glu Tyr Asp Glu Gly Val Lys Glu Tyr Leu Leu Asp Tyr Ile
805 810 815
Ser Glu His Arg Ser Ile Leu Gly Asn Ser Val Gln Glu Leu Asn Asp
820 825 830
Leu Val Thr Ser Thr Leu Asn Asn Ser Ile Pro Phe Glu Leu Ser Ser
835 840 845
Tyr Thr Asn Asp Lys Ile Leu Ile Leu Tyr Phe Asn Lys Leu Tyr Lys
850 855 860
Lys Ile Lys Asp Asn Ser Ile Leu Asp Met Arg Tyr Glu Asn Asn Lys
865 870 875 880
Phe Ile Asp Ile Ser Gly Tyr Gly Ser Asn Ile Ser Ile Asn Gly Asp
885 890 895
Val Tyr Ile Tyr Ser Thr Asn Arg Asn Gln Phe Gly Ile Tyr Ser Ser
900 905 910
Lys Pro Ser Glu Val Asn Ile Ala Gln Asn Asn Asp Ile Ile Tyr Asn
915 920 925
Gly Arg Tyr Gln Asn Phe Ser Ile Ser Phe Trp Val Arg Ile Pro Lys
930 935 940
Tyr Phe Asn Lys Val Asn Leu Asn Asn Glu Tyr Thr Ile Ile Asp Cys
945 950 955 960
Ile Arg Asn Asn Asn Ser Gly Trp Lys Ile Ser Leu Asn Tyr Asn Lys
965 970 975
Ile Ile Trp Thr Leu Gln Asp Thr Ala Gly Asn Asn Gln Lys Leu Val
980 985 990
Phe Asn Tyr Thr Gln Met Ile Ser Ile Ser Asp Tyr Ile Asn Lys Trp
995 1000 1005
Ile Phe Val Thr Ile Thr Asn Asn Arg Leu Gly Asn Ser Arg Ile
1010 1015 1020
Tyr Ile Asn Gly Asn Leu Ile Asp Glu Lys Ser Ile Ser Asn Leu
1025 1030 1035
Gly Asp Ile His Val Ser Asp Asn Ile Leu Phe Lys Ile Val Gly
1040 1045 1050
Cys Asn Asp Thr Arg Tyr Val Gly Ile Arg Tyr Phe Lys Val Phe
1055 1060 1065
Asp Thr Glu Leu Gly Lys Thr Glu Ile Glu Thr Leu Tyr Ser Asp
1070 1075 1080
Glu Pro Asp Pro Ser Ile Leu Lys Asp Phe Trp Gly Asn Tyr Leu
1085 1090 1095
Leu Tyr Asn Lys Arg Tyr Tyr Leu Leu Asn Leu Leu Arg Thr Asp
1100 1105 1110
Lys Ser Ile Thr Gln Asn Ser Asn Phe Leu Asn Ile Asn Gln Gln
1115 1120 1125
Arg Gly Val Tyr Gln Lys Pro Asn Ile Phe Ser Asn Thr Arg Leu
1130 1135 1140
Tyr Thr Gly Val Glu Val Ile Ile Arg Lys Asn Gly Ser Thr Asp
1145 1150 1155
Ile Ser Asn Thr Asp Asn Phe Val Arg Lys Asn Asp Leu Ala Tyr
1160 1165 1170
Ile Asn Val Val Asp Arg Asp Val Glu Tyr Arg Leu Tyr Ala Asp
1175 1180 1185
Ile Ser Ile Ala Lys Pro Glu Lys Ile Ile Lys Leu Ile Arg Thr
1190 1195 1200
Ser Asn Ser Asn Asn Ser Leu Gly Gln Ile Ile Val Met Asp Ser
1205 1210 1215
Ile Gly Asn Asn Cys Thr Met Asn Phe Gln Asn Asn Asn Gly Gly
1220 1225 1230
Asn Ile Gly Leu Leu Gly Phe His Ser Asn Asn Leu Val Ala Ser
1235 1240 1245
Ser Trp Tyr Tyr Asn Asn Ile Arg Lys Asn Thr Ser Ser Asn Gly
1250 1255 1260
Cys Phe Trp Ser Phe Ile Ser Lys Glu His Gly Trp Gln Glu Asn
1265 1270 1275
Glu Asn Leu Tyr Phe Gln Gly Ala Ser His His His His His His
1280 1285 1290
His His
1295
<210> 35
<211> 2649
<212> DNA
<213> 人工序列
<220>
<223> rLHN/F的核苷酸序列 (His-标签的)
<400> 35
atgccggttg tgattaacag cttcaattat aacgatccgg tgaacgatga taccatcctg 60
tatatgcaga ttccgtatga agagaaaagc aaaaagtact acaaagcctt tgagatcatg 120
cgcaacgttt ggattattcc ggaacgtaat accattggca ccgatccgag cgattttgat 180
ccgcctgcaa gcctggaaaa tggtagcagc gcatattatg atccgaatta tctgaccacc 240
gatgccgaaa aagatcgtta tctgaaaacc accatcaaac tgttcaaacg cattaatagc 300
aatccggcag gcgaagttct gctgcaagaa attagctatg caaaaccgta tctgggcaat 360
gaacataccc cgattaatga atttcatccg gttacacgta ccacgagcgt taacattaaa 420
agcagcacca atgtgaagtc cagcattatt ctgaatctgc tggttttagg tgcaggtccg 480
gatatttttg aaaattcaag ctatccggtg cgcaaactga tggatagcgg tggtgtgtat 540
gatccgtcaa atgatggttt tggcagcatt aacattgtga cctttagtcc ggaatatgaa 600
tacaccttca acgatattag cggtggctat aatagcagca ccgaaagttt tattgcagat 660
ccggcaatta gcctggcaca tgaactgatt catgcactgc atggtctgta tggtgcacgt 720
ggtgttacct ataaagaaac cattaaagtt aaacaggcac cgctgatgat tgcggaaaaa 780
ccgattcgtc tggaagaatt tctgaccttt ggtggtcagg atctgaacat tattaccagc 840
gcaatgaaag agaaaatcta taataacctg ctggccaact atgagaaaat tgcaacccgt 900
ctgagccgtg ttaatagcgc acctcctgaa tatgatatca acgagtataa agactatttt 960
cagtggaaat acggcctgga taaaaatgca gatggtagct ataccgtgaa cgagaacaaa 1020
tttaacgaga tctacaaaaa actgtatagc ttcaccgaaa tcgatctggc caacaaattc 1080
aaagtgaaat gccgcaacac ctacttcatc aaatatggct ttctgaaagt tccgaacctg 1140
cttgatgatg atatctatac cgttagcgaa ggctttaaca ttggtaatct ggccgttaat 1200
aatcgcggtc agaacattaa actgaacccg aaaattatcg atagcatccc ggataaaggc 1260
ctggttgaaa aaattgtgaa attctgcaaa agcgtgattc cgcgtaaagg caccaaagca 1320
ccgcctcgtc tgtgtattcg tgtgaataat cgtgaactgt tttttgttgc aagcgagagc 1380
agctataacg agaatgatat taacaccccg aaagagattg acgataccac caatctgaat 1440
aacaactatc gcaacaatct ggatgaagtg atcctggatt ataacagcga aaccattccg 1500
cagattagca atcagaccct gaataccctg gttcaggatg atagctatgt tccgcgttat 1560
gatagcaatg gcaccagcga aattgaagaa cataatgtgg ttgatctgaa cgtgttcttt 1620
tatctgcatg cacagaaagt gccggaaggt gaaaccaata ttagcctgac cagcagcatt 1680
gataccgcac tgagcgaaga aagccaggtt tatacctttt ttagcagcga attcatcaac 1740
accattaaca aaccggttca tgcagcactg tttattagct ggattaatca ggtgattcgc 1800
gattttacca ccgaagcaac ccagaaaagc acctttgata aaattgccga tattagtctg 1860
gtggtgccgt atgttggtct ggcactgaat attggtaatg aagtgcagaa agagaacttt 1920
aaagaagcct tcgaactgtt aggtgccggt attctgctgg aatttgtgcc ggaactgctg 1980
attccgacca ttctggtttt taccattaag agctttattg gcagcagcga gaacaagaac 2040
aaaatcatta aagccatcaa caacagcctg atggaacgcg aaaccaaatg gaaagaaatt 2100
tacagctgga ttgtgagcaa ttggctgacc cgtatcaata cccagtttaa caaacgcaaa 2160
gaacaaatgt atcaggccct gcagaatcag gttgatgcaa ttaaaaccgt gatcgaatac 2220
aaatacaaca actataccag cgacgaacgt aatcgcctgg aaagcgaata caacattaat 2280
aacattcgcg aagaactgaa caaaaaagtg agcctggcaa tggaaaacat cgaacgtttt 2340
attaccgaaa gcagcatctt ctacctgatg aaactgatta acgaagccaa agttagcaaa 2400
ctgcgcgaat atgatgaagg cgttaaagaa tatctgctgg actatattag cgaacatcgt 2460
agcattctgg gtaatagcgt tcaagagctg aatgatctgg ttaccagcac actgaataat 2520
agcattccgt ttgaactgag cagctacacc aacgataaaa tcctgatcct gtacttcaac 2580
aaactgtaca agaaagaaaa cctgtatttt cagggtgcaa gccatcatca ccaccatcac 2640
catcattaa 2649
<210> 36
<211> 882
<212> PRT
<213> 人工序列
<220>
<223> rLHN/F的多肽序列 (His-标签的)
<400> 36
Met Pro Val Val Ile Asn Ser Phe Asn Tyr Asn Asp Pro Val Asn Asp
1 5 10 15
Asp Thr Ile Leu Tyr Met Gln Ile Pro Tyr Glu Glu Lys Ser Lys Lys
20 25 30
Tyr Tyr Lys Ala Phe Glu Ile Met Arg Asn Val Trp Ile Ile Pro Glu
35 40 45
Arg Asn Thr Ile Gly Thr Asp Pro Ser Asp Phe Asp Pro Pro Ala Ser
50 55 60
Leu Glu Asn Gly Ser Ser Ala Tyr Tyr Asp Pro Asn Tyr Leu Thr Thr
65 70 75 80
Asp Ala Glu Lys Asp Arg Tyr Leu Lys Thr Thr Ile Lys Leu Phe Lys
85 90 95
Arg Ile Asn Ser Asn Pro Ala Gly Glu Val Leu Leu Gln Glu Ile Ser
100 105 110
Tyr Ala Lys Pro Tyr Leu Gly Asn Glu His Thr Pro Ile Asn Glu Phe
115 120 125
His Pro Val Thr Arg Thr Thr Ser Val Asn Ile Lys Ser Ser Thr Asn
130 135 140
Val Lys Ser Ser Ile Ile Leu Asn Leu Leu Val Leu Gly Ala Gly Pro
145 150 155 160
Asp Ile Phe Glu Asn Ser Ser Tyr Pro Val Arg Lys Leu Met Asp Ser
165 170 175
Gly Gly Val Tyr Asp Pro Ser Asn Asp Gly Phe Gly Ser Ile Asn Ile
180 185 190
Val Thr Phe Ser Pro Glu Tyr Glu Tyr Thr Phe Asn Asp Ile Ser Gly
195 200 205
Gly Tyr Asn Ser Ser Thr Glu Ser Phe Ile Ala Asp Pro Ala Ile Ser
210 215 220
Leu Ala His Glu Leu Ile His Ala Leu His Gly Leu Tyr Gly Ala Arg
225 230 235 240
Gly Val Thr Tyr Lys Glu Thr Ile Lys Val Lys Gln Ala Pro Leu Met
245 250 255
Ile Ala Glu Lys Pro Ile Arg Leu Glu Glu Phe Leu Thr Phe Gly Gly
260 265 270
Gln Asp Leu Asn Ile Ile Thr Ser Ala Met Lys Glu Lys Ile Tyr Asn
275 280 285
Asn Leu Leu Ala Asn Tyr Glu Lys Ile Ala Thr Arg Leu Ser Arg Val
290 295 300
Asn Ser Ala Pro Pro Glu Tyr Asp Ile Asn Glu Tyr Lys Asp Tyr Phe
305 310 315 320
Gln Trp Lys Tyr Gly Leu Asp Lys Asn Ala Asp Gly Ser Tyr Thr Val
325 330 335
Asn Glu Asn Lys Phe Asn Glu Ile Tyr Lys Lys Leu Tyr Ser Phe Thr
340 345 350
Glu Ile Asp Leu Ala Asn Lys Phe Lys Val Lys Cys Arg Asn Thr Tyr
355 360 365
Phe Ile Lys Tyr Gly Phe Leu Lys Val Pro Asn Leu Leu Asp Asp Asp
370 375 380
Ile Tyr Thr Val Ser Glu Gly Phe Asn Ile Gly Asn Leu Ala Val Asn
385 390 395 400
Asn Arg Gly Gln Asn Ile Lys Leu Asn Pro Lys Ile Ile Asp Ser Ile
405 410 415
Pro Asp Lys Gly Leu Val Glu Lys Ile Val Lys Phe Cys Lys Ser Val
420 425 430
Ile Pro Arg Lys Gly Thr Lys Ala Pro Pro Arg Leu Cys Ile Arg Val
435 440 445
Asn Asn Arg Glu Leu Phe Phe Val Ala Ser Glu Ser Ser Tyr Asn Glu
450 455 460
Asn Asp Ile Asn Thr Pro Lys Glu Ile Asp Asp Thr Thr Asn Leu Asn
465 470 475 480
Asn Asn Tyr Arg Asn Asn Leu Asp Glu Val Ile Leu Asp Tyr Asn Ser
485 490 495
Glu Thr Ile Pro Gln Ile Ser Asn Gln Thr Leu Asn Thr Leu Val Gln
500 505 510
Asp Asp Ser Tyr Val Pro Arg Tyr Asp Ser Asn Gly Thr Ser Glu Ile
515 520 525
Glu Glu His Asn Val Val Asp Leu Asn Val Phe Phe Tyr Leu His Ala
530 535 540
Gln Lys Val Pro Glu Gly Glu Thr Asn Ile Ser Leu Thr Ser Ser Ile
545 550 555 560
Asp Thr Ala Leu Ser Glu Glu Ser Gln Val Tyr Thr Phe Phe Ser Ser
565 570 575
Glu Phe Ile Asn Thr Ile Asn Lys Pro Val His Ala Ala Leu Phe Ile
580 585 590
Ser Trp Ile Asn Gln Val Ile Arg Asp Phe Thr Thr Glu Ala Thr Gln
595 600 605
Lys Ser Thr Phe Asp Lys Ile Ala Asp Ile Ser Leu Val Val Pro Tyr
610 615 620
Val Gly Leu Ala Leu Asn Ile Gly Asn Glu Val Gln Lys Glu Asn Phe
625 630 635 640
Lys Glu Ala Phe Glu Leu Leu Gly Ala Gly Ile Leu Leu Glu Phe Val
645 650 655
Pro Glu Leu Leu Ile Pro Thr Ile Leu Val Phe Thr Ile Lys Ser Phe
660 665 670
Ile Gly Ser Ser Glu Asn Lys Asn Lys Ile Ile Lys Ala Ile Asn Asn
675 680 685
Ser Leu Met Glu Arg Glu Thr Lys Trp Lys Glu Ile Tyr Ser Trp Ile
690 695 700
Val Ser Asn Trp Leu Thr Arg Ile Asn Thr Gln Phe Asn Lys Arg Lys
705 710 715 720
Glu Gln Met Tyr Gln Ala Leu Gln Asn Gln Val Asp Ala Ile Lys Thr
725 730 735
Val Ile Glu Tyr Lys Tyr Asn Asn Tyr Thr Ser Asp Glu Arg Asn Arg
740 745 750
Leu Glu Ser Glu Tyr Asn Ile Asn Asn Ile Arg Glu Glu Leu Asn Lys
755 760 765
Lys Val Ser Leu Ala Met Glu Asn Ile Glu Arg Phe Ile Thr Glu Ser
770 775 780
Ser Ile Phe Tyr Leu Met Lys Leu Ile Asn Glu Ala Lys Val Ser Lys
785 790 795 800
Leu Arg Glu Tyr Asp Glu Gly Val Lys Glu Tyr Leu Leu Asp Tyr Ile
805 810 815
Ser Glu His Arg Ser Ile Leu Gly Asn Ser Val Gln Glu Leu Asn Asp
820 825 830
Leu Val Thr Ser Thr Leu Asn Asn Ser Ile Pro Phe Glu Leu Ser Ser
835 840 845
Tyr Thr Asn Asp Lys Ile Leu Ile Leu Tyr Phe Asn Lys Leu Tyr Lys
850 855 860
Lys Glu Asn Leu Tyr Phe Gln Gly Ala Ser His His His His His His
865 870 875 880
His His
<210> 37
<211> 1296
<212> DNA
<213> 人工序列
<220>
<223> rHC/F的核苷酸序列 (His-标签的)
<400> 37
atgatcaagg ataacagcat tctggatatg cgctatgaga acaacaaatt cattgatatt 60
agcggctatg gcagcaacat tagcattaat ggtgatgtgt atatctacag caccaaccgt 120
aatcagtttg gcatttatag cagcaaaccg agcgaagtta atattgccca gaacaacgat 180
atcatctata acggtcgcta tcagaacttc agcattagct tttgggttcg cattccgaaa 240
tacttcaata aggtgaacct gaacaacgag tataccatca ttgattgcat tcgcaataat 300
aacagcggct ggaaaattag cctgaactac aacaaaatta tctggaccct gcaggatacc 360
gcaggtaata atcagaaact ggtgtttaac tacacccaga tgattagcat cagcgactat 420
atcaacaaat ggatctttgt gaccattacc aataatcgcc tgggtaatag ccgcatttat 480
atcaatggta acctgatcga tgagaaaagc attagcaatc tgggtgatat tcatgtgagc 540
gataacatcc tgtttaaaat cgtgggttgt aacgataccc gttatgttgg tattcgctac 600
ttcaaagtgt ttgataccga actgggtaaa accgaaattg aaaccctgta tagtgatgaa 660
ccggatccga gcattctgaa agatttttgg ggtaattatc tgctgtacaa caaacgctac 720
tatctgctga atctgctgcg taccgataaa tcaattaccc agaatagcaa cttcctgaac 780
attaatcagc agcgtggtgt ttatcagaaa ccgaacattt ttagcaacac ccgtctgtat 840
accggtgtgg aagttattat tcgtaaaaat ggcagcaccg atatcagcaa caccgataac 900
tttgttcgca aaaatgatct ggcgtatatc aacgttgttg atcgtgatgt tgaatatcgt 960
ctgtatgccg atattagcat tgccaaaccg gaaaaaatca tcaaactgat ccgtaccagc 1020
aacagcaata attcactggg tcagattatt gtgatggata gcattggtaa taactgcacc 1080
atgaactttc agaacaataa cggtggtaat attggtctgc tgggctttca tagtaataat 1140
ctggttgcaa gcagctggta ttataacaac atccgtaaaa ataccagcag caatggttgc 1200
ttttggagct ttattagcaa agaacatggc tggcaagaaa acgagaatct gtattttcag 1260
ggtgcaagtc atcatcacca ccatcaccat cattaa 1296
<210> 38
<211> 431
<212> PRT
<213> 人工序列
<220>
<223> rHC/F的多肽序列 (His-标签的)
<400> 38
Met Ile Lys Asp Asn Ser Ile Leu Asp Met Arg Tyr Glu Asn Asn Lys
1 5 10 15
Phe Ile Asp Ile Ser Gly Tyr Gly Ser Asn Ile Ser Ile Asn Gly Asp
20 25 30
Val Tyr Ile Tyr Ser Thr Asn Arg Asn Gln Phe Gly Ile Tyr Ser Ser
35 40 45
Lys Pro Ser Glu Val Asn Ile Ala Gln Asn Asn Asp Ile Ile Tyr Asn
50 55 60
Gly Arg Tyr Gln Asn Phe Ser Ile Ser Phe Trp Val Arg Ile Pro Lys
65 70 75 80
Tyr Phe Asn Lys Val Asn Leu Asn Asn Glu Tyr Thr Ile Ile Asp Cys
85 90 95
Ile Arg Asn Asn Asn Ser Gly Trp Lys Ile Ser Leu Asn Tyr Asn Lys
100 105 110
Ile Ile Trp Thr Leu Gln Asp Thr Ala Gly Asn Asn Gln Lys Leu Val
115 120 125
Phe Asn Tyr Thr Gln Met Ile Ser Ile Ser Asp Tyr Ile Asn Lys Trp
130 135 140
Ile Phe Val Thr Ile Thr Asn Asn Arg Leu Gly Asn Ser Arg Ile Tyr
145 150 155 160
Ile Asn Gly Asn Leu Ile Asp Glu Lys Ser Ile Ser Asn Leu Gly Asp
165 170 175
Ile His Val Ser Asp Asn Ile Leu Phe Lys Ile Val Gly Cys Asn Asp
180 185 190
Thr Arg Tyr Val Gly Ile Arg Tyr Phe Lys Val Phe Asp Thr Glu Leu
195 200 205
Gly Lys Thr Glu Ile Glu Thr Leu Tyr Ser Asp Glu Pro Asp Pro Ser
210 215 220
Ile Leu Lys Asp Phe Trp Gly Asn Tyr Leu Leu Tyr Asn Lys Arg Tyr
225 230 235 240
Tyr Leu Leu Asn Leu Leu Arg Thr Asp Lys Ser Ile Thr Gln Asn Ser
245 250 255
Asn Phe Leu Asn Ile Asn Gln Gln Arg Gly Val Tyr Gln Lys Pro Asn
260 265 270
Ile Phe Ser Asn Thr Arg Leu Tyr Thr Gly Val Glu Val Ile Ile Arg
275 280 285
Lys Asn Gly Ser Thr Asp Ile Ser Asn Thr Asp Asn Phe Val Arg Lys
290 295 300
Asn Asp Leu Ala Tyr Ile Asn Val Val Asp Arg Asp Val Glu Tyr Arg
305 310 315 320
Leu Tyr Ala Asp Ile Ser Ile Ala Lys Pro Glu Lys Ile Ile Lys Leu
325 330 335
Ile Arg Thr Ser Asn Ser Asn Asn Ser Leu Gly Gln Ile Ile Val Met
340 345 350
Asp Ser Ile Gly Asn Asn Cys Thr Met Asn Phe Gln Asn Asn Asn Gly
355 360 365
Gly Asn Ile Gly Leu Leu Gly Phe His Ser Asn Asn Leu Val Ala Ser
370 375 380
Ser Trp Tyr Tyr Asn Asn Ile Arg Lys Asn Thr Ser Ser Asn Gly Cys
385 390 395 400
Phe Trp Ser Phe Ile Ser Lys Glu His Gly Trp Gln Glu Asn Glu Asn
405 410 415
Leu Tyr Phe Gln Gly Ala Ser His His His His His His His His
420 425 430
<210> 39
<211> 1347
<212> DNA
<213> 人工序列
<220>
<223> rLC/F的核苷酸序列 (His-标签的)
<400> 39
atgccggttg tgattaacag cttcaattat aacgatccgg tgaacgatga taccatcctg 60
tatatgcaga ttccgtatga agagaaaagc aaaaagtact acaaagcctt tgagatcatg 120
cgcaacgttt ggattattcc ggaacgtaat accattggca ccgatccgag cgattttgat 180
ccgcctgcaa gcctggaaaa tggtagcagc gcatattatg atccgaatta tctgaccacc 240
gatgccgaaa aagatcgtta tctgaaaacc accatcaaac tgttcaaacg cattaatagc 300
aatccggcag gcgaagttct gctgcaagaa attagctatg caaaaccgta tctgggcaat 360
gaacataccc cgattaatga atttcatccg gttacacgta ccacgagcgt taacattaaa 420
agcagcacca atgtgaagtc cagcattatt ctgaatctgc tggttttagg tgcaggtccg 480
gatatttttg aaaattcaag ctatccggtg cgcaaactga tggatagcgg tggtgtgtat 540
gatccgtcaa atgatggttt tggcagcatt aacattgtga cctttagtcc ggaatatgaa 600
tacaccttca acgatattag cggtggctat aatagcagca ccgaaagttt tattgcagat 660
ccggcaatta gcctggcaca tgaactgatt catgcactgc atggtctgta tggtgcacgt 720
ggtgttacct ataaagaaac cattaaagtt aaacaggcac cgctgatgat tgcggaaaaa 780
ccgattcgtc tggaagaatt tctgaccttt ggtggtcagg atctgaacat tattaccagc 840
gcaatgaaag agaaaatcta taataacctg ctggccaact atgagaaaat tgcaacccgt 900
ctgagccgtg ttaatagcgc acctcctgaa tatgatatca acgagtataa agactatttt 960
cagtggaaat acggcctgga taaaaatgca gatggtagct ataccgtgaa cgagaacaaa 1020
tttaacgaga tctacaaaaa actgtatagc ttcaccgaaa tcgatctggc caacaaattc 1080
aaagtgaaat gccgcaacac ctacttcatc aaatatggct ttctgaaagt tccgaacctg 1140
cttgatgatg atatctatac cgttagcgaa ggctttaaca ttggtaatct ggccgttaat 1200
aatcgcggtc agaacattaa actgaacccg aaaattatcg atagcatccc ggataaaggc 1260
ctggttgaaa aaattgtgaa attctgcaaa agcgagaacc tgtattttca gggtgcaagt 1320
catcatcacc atcaccacca tcattaa 1347
<210> 40
<211> 448
<212> PRT
<213> 人工序列
<220>
<223> rLC/F的多肽序列 (His-标签的)
<400> 40
Met Pro Val Val Ile Asn Ser Phe Asn Tyr Asn Asp Pro Val Asn Asp
1 5 10 15
Asp Thr Ile Leu Tyr Met Gln Ile Pro Tyr Glu Glu Lys Ser Lys Lys
20 25 30
Tyr Tyr Lys Ala Phe Glu Ile Met Arg Asn Val Trp Ile Ile Pro Glu
35 40 45
Arg Asn Thr Ile Gly Thr Asp Pro Ser Asp Phe Asp Pro Pro Ala Ser
50 55 60
Leu Glu Asn Gly Ser Ser Ala Tyr Tyr Asp Pro Asn Tyr Leu Thr Thr
65 70 75 80
Asp Ala Glu Lys Asp Arg Tyr Leu Lys Thr Thr Ile Lys Leu Phe Lys
85 90 95
Arg Ile Asn Ser Asn Pro Ala Gly Glu Val Leu Leu Gln Glu Ile Ser
100 105 110
Tyr Ala Lys Pro Tyr Leu Gly Asn Glu His Thr Pro Ile Asn Glu Phe
115 120 125
His Pro Val Thr Arg Thr Thr Ser Val Asn Ile Lys Ser Ser Thr Asn
130 135 140
Val Lys Ser Ser Ile Ile Leu Asn Leu Leu Val Leu Gly Ala Gly Pro
145 150 155 160
Asp Ile Phe Glu Asn Ser Ser Tyr Pro Val Arg Lys Leu Met Asp Ser
165 170 175
Gly Gly Val Tyr Asp Pro Ser Asn Asp Gly Phe Gly Ser Ile Asn Ile
180 185 190
Val Thr Phe Ser Pro Glu Tyr Glu Tyr Thr Phe Asn Asp Ile Ser Gly
195 200 205
Gly Tyr Asn Ser Ser Thr Glu Ser Phe Ile Ala Asp Pro Ala Ile Ser
210 215 220
Leu Ala His Glu Leu Ile His Ala Leu His Gly Leu Tyr Gly Ala Arg
225 230 235 240
Gly Val Thr Tyr Lys Glu Thr Ile Lys Val Lys Gln Ala Pro Leu Met
245 250 255
Ile Ala Glu Lys Pro Ile Arg Leu Glu Glu Phe Leu Thr Phe Gly Gly
260 265 270
Gln Asp Leu Asn Ile Ile Thr Ser Ala Met Lys Glu Lys Ile Tyr Asn
275 280 285
Asn Leu Leu Ala Asn Tyr Glu Lys Ile Ala Thr Arg Leu Ser Arg Val
290 295 300
Asn Ser Ala Pro Pro Glu Tyr Asp Ile Asn Glu Tyr Lys Asp Tyr Phe
305 310 315 320
Gln Trp Lys Tyr Gly Leu Asp Lys Asn Ala Asp Gly Ser Tyr Thr Val
325 330 335
Asn Glu Asn Lys Phe Asn Glu Ile Tyr Lys Lys Leu Tyr Ser Phe Thr
340 345 350
Glu Ile Asp Leu Ala Asn Lys Phe Lys Val Lys Cys Arg Asn Thr Tyr
355 360 365
Phe Ile Lys Tyr Gly Phe Leu Lys Val Pro Asn Leu Leu Asp Asp Asp
370 375 380
Ile Tyr Thr Val Ser Glu Gly Phe Asn Ile Gly Asn Leu Ala Val Asn
385 390 395 400
Asn Arg Gly Gln Asn Ile Lys Leu Asn Pro Lys Ile Ile Asp Ser Ile
405 410 415
Pro Asp Lys Gly Leu Val Glu Lys Ile Val Lys Phe Cys Lys Ser Glu
420 425 430
Asn Leu Tyr Phe Gln Gly Ala Ser His His His His His His His His
435 440 445
<210> 41
<211> 1317
<212> DNA
<213> 人工序列
<220>
<223> 阳离子rHC/A的核苷酸序列 (His-标签的)
<400> 41
atgatcatca acaccagcat tctgaacctg cgttatgaaa gcaaacatct gattgatctg 60
agccgttatg ccagcaaaat caatataggc agcaaggtta acttcgaccc gattgacaaa 120
aatcagatac agctgtttaa tctggaaagc agcaaaattg aggtgatcct gaaaaaagcg 180
atcgtgtata atagcatgta cgagaatttt tcgaccagct tttggattcg catcccgaaa 240
tactttaaca agattagcct gaacaacgag tataccatca ttaactgcat ggaaaacaat 300
agcggttgga aagtcagcct gaattatggc gaaattatct ggaccctgca ggataccaaa 360
gaaatcaaac agcgtgtggt gttcaaatac agccagatga ttaatatcag cgactatatc 420
aaccgctgga tttttgtgac cattaccaat aatcggctga acaagagcaa gatctatatt 480
aacggtcgtc tgattgacca gaaaccgatt agtaatctgg gtaatattca tgcgagcaac 540
aaaatcatgt ttaaactgga tggttgccgt gatacccatc gttatatttg gatcaaatac 600
ttcaacctgt tcgataaaga gttgaacgaa aaagaaatta aagacctgta cgataaccag 660
agcaatagcg gcatactgaa agatttttgg ggagattatc tgcagtatga caaaccgtat 720
tatatgctga atctgtacga cccgaataaa tacgtggatg ttaataatgt gggcatccgt 780
ggttatatgt acctgaaagg tccgcgtggt agcgttatga ccacaaacat ttatctgaat 840
agcagcctgt atcgcggaac caaattcatc attaaaaagt atgccagcgg caacaaggat 900
aatattgtgc gtaataatga tcgcgtgtac attaacgttg tggtgaagaa taaagaatat 960
cgcctggcaa ccaatgcaag ccaggcaggc gttgaaaaaa ttctgagtgc cctggaaatt 1020
ccggatgttg gtaatctgag ccaggttgtt gtgatgaaaa gcaaaaacga taaaggcatc 1080
accaacaaat gcaagatgaa tctgcaggac aataacggca atgatattgg cttcattggc 1140
tttcaccagt ttaacaacat tgcaaaactg gttgcgagca attggtataa tcgtcagatt 1200
gaacgtagca gtcgtaccct gggttgtagc tgggaattta tccctgtgga tgatggttgg 1260
ggtgaacgtc cgctgaagct tgcggccgca ctcgagcacc accaccacca ccactga 1317
<210> 42
<211> 438
<212> PRT
<213> 人工序列
<220>
<223> 阳离子rHC/A的多肽序列 (His-标签的)
<400> 42
Met Ile Ile Asn Thr Ser Ile Leu Asn Leu Arg Tyr Glu Ser Lys His
1 5 10 15
Leu Ile Asp Leu Ser Arg Tyr Ala Ser Lys Ile Asn Ile Gly Ser Lys
20 25 30
Val Asn Phe Asp Pro Ile Asp Lys Asn Gln Ile Gln Leu Phe Asn Leu
35 40 45
Glu Ser Ser Lys Ile Glu Val Ile Leu Lys Lys Ala Ile Val Tyr Asn
50 55 60
Ser Met Tyr Glu Asn Phe Ser Thr Ser Phe Trp Ile Arg Ile Pro Lys
65 70 75 80
Tyr Phe Asn Lys Ile Ser Leu Asn Asn Glu Tyr Thr Ile Ile Asn Cys
85 90 95
Met Glu Asn Asn Ser Gly Trp Lys Val Ser Leu Asn Tyr Gly Glu Ile
100 105 110
Ile Trp Thr Leu Gln Asp Thr Lys Glu Ile Lys Gln Arg Val Val Phe
115 120 125
Lys Tyr Ser Gln Met Ile Asn Ile Ser Asp Tyr Ile Asn Arg Trp Ile
130 135 140
Phe Val Thr Ile Thr Asn Asn Arg Leu Asn Lys Ser Lys Ile Tyr Ile
145 150 155 160
Asn Gly Arg Leu Ile Asp Gln Lys Pro Ile Ser Asn Leu Gly Asn Ile
165 170 175
His Ala Ser Asn Lys Ile Met Phe Lys Leu Asp Gly Cys Arg Asp Thr
180 185 190
His Arg Tyr Ile Trp Ile Lys Tyr Phe Asn Leu Phe Asp Lys Glu Leu
195 200 205
Asn Glu Lys Glu Ile Lys Asp Leu Tyr Asp Asn Gln Ser Asn Ser Gly
210 215 220
Ile Leu Lys Asp Phe Trp Gly Asp Tyr Leu Gln Tyr Asp Lys Pro Tyr
225 230 235 240
Tyr Met Leu Asn Leu Tyr Asp Pro Asn Lys Tyr Val Asp Val Asn Asn
245 250 255
Val Gly Ile Arg Gly Tyr Met Tyr Leu Lys Gly Pro Arg Gly Ser Val
260 265 270
Met Thr Thr Asn Ile Tyr Leu Asn Ser Ser Leu Tyr Arg Gly Thr Lys
275 280 285
Phe Ile Ile Lys Lys Tyr Ala Ser Gly Asn Lys Asp Asn Ile Val Arg
290 295 300
Asn Asn Asp Arg Val Tyr Ile Asn Val Val Val Lys Asn Lys Glu Tyr
305 310 315 320
Arg Leu Ala Thr Asn Ala Ser Gln Ala Gly Val Glu Lys Ile Leu Ser
325 330 335
Ala Leu Glu Ile Pro Asp Val Gly Asn Leu Ser Gln Val Val Val Met
340 345 350
Lys Ser Lys Asn Asp Lys Gly Ile Thr Asn Lys Cys Lys Met Asn Leu
355 360 365
Gln Asp Asn Asn Gly Asn Asp Ile Gly Phe Ile Gly Phe His Gln Phe
370 375 380
Asn Asn Ile Ala Lys Leu Val Ala Ser Asn Trp Tyr Asn Arg Gln Ile
385 390 395 400
Glu Arg Ser Ser Arg Thr Leu Gly Cys Ser Trp Glu Phe Ile Pro Val
405 410 415
Asp Asp Gly Trp Gly Glu Arg Pro Leu Lys Leu Ala Ala Ala Leu Glu
420 425 430
His His His His His His
435
<210> 43
<211> 1341
<212> DNA
<213> 人工序列
<220>
<223> rHC/AB的核苷酸序列 (His-标签的)
<400> 43
atgattctga acaatattat cctgaacctg cgttacaaag acaacaatct gatcgatctg 60
agcggctatg gtgcaaaagt tgaagtctac gacggtgtcg aactgaacga taaaaaccag 120
ttcaaactga cctcatcggc taactcaaaa attcgtgtga cgcagaacca aaacatcatc 180
ttcaactcgg tctttctgga cttcagcgtg tctttctgga ttcgcatccc gaaatataaa 240
aatgatggca tccagaacta catccataac gaatacacca tcatcaactg tatgaaaaac 300
aacagtggtt ggaaaatttc catccgtggc aaccgcatta tctggaccct gattgatatc 360
aatggtaaaa cgaaaagcgt gtttttcgaa tacaacatcc gtgaagatat ctctgaatac 420
atcaatcgct ggtttttcgt gaccattacg aacaatctga acaatgcgaa aatctatatc 480
aacggcaaac tggaaagtaa taccgacatc aaagatattc gtgaagttat cgccaacggt 540
gaaatcatct tcaaactgga tggcgacatc gatcgcaccc agttcatttg gatgaaatac 600
ttctccatct tcaacacgga actgagtcag tccaatatcg aagaacgcta caaaatccaa 660
tcatactcgg aatacctgaa agatttctgg ggtaacccgc tgatgtacaa caaagaatac 720
tacatgttca acgcgggcaa caaaaactca tacatcaaac tgaaaaaaga ttcgccggtg 780
ggtgaaatcc tgacccgtag caaatacaac cagaactcta aatacatcaa ctatcgcgat 840
ctgtacattg gcgaaaaatt tattatccgt cgcaaaagca actctcagag tattaatgat 900
gacatcgtgc gtaaagaaga ctacatctat ctggatttct ttaatctgaa ccaagaatgg 960
cgcgtttata cctacaaata cttcaaaaaa gaagaaatga aactgttcct ggccccgatt 1020
tacgacagcg atgaatttta caacaccatc cagatcaaag aatacgatga acagccgacg 1080
tatagttgcc aactgctgtt caaaaaagac gaagaatcca ccgatgaaat tggcctgatt 1140
ggtatccacc gtttctatga aagcggtatc gttttcgaag aatacaaaga ttacttctgt 1200
atctctaaat ggtatctgaa agaagtcaaa cgcaaaccgt acaacctgaa actgggctgc 1260
aactggcaat ttatcccgaa agacgaaggc tggaccgaaa agcttgcggc cgcactcgag 1320
caccaccacc accaccactg a 1341
<210> 44
<211> 446
<212> PRT
<213> 人工序列
<220>
<223> rHC/AB的多肽序列 (His-标签的)
<400> 44
Met Ile Leu Asn Asn Ile Ile Leu Asn Leu Arg Tyr Lys Asp Asn Asn
1 5 10 15
Leu Ile Asp Leu Ser Gly Tyr Gly Ala Lys Val Glu Val Tyr Asp Gly
20 25 30
Val Glu Leu Asn Asp Lys Asn Gln Phe Lys Leu Thr Ser Ser Ala Asn
35 40 45
Ser Lys Ile Arg Val Thr Gln Asn Gln Asn Ile Ile Phe Asn Ser Val
50 55 60
Phe Leu Asp Phe Ser Val Ser Phe Trp Ile Arg Ile Pro Lys Tyr Lys
65 70 75 80
Asn Asp Gly Ile Gln Asn Tyr Ile His Asn Glu Tyr Thr Ile Ile Asn
85 90 95
Cys Met Lys Asn Asn Ser Gly Trp Lys Ile Ser Ile Arg Gly Asn Arg
100 105 110
Ile Ile Trp Thr Leu Ile Asp Ile Asn Gly Lys Thr Lys Ser Val Phe
115 120 125
Phe Glu Tyr Asn Ile Arg Glu Asp Ile Ser Glu Tyr Ile Asn Arg Trp
130 135 140
Phe Phe Val Thr Ile Thr Asn Asn Leu Asn Asn Ala Lys Ile Tyr Ile
145 150 155 160
Asn Gly Lys Leu Glu Ser Asn Thr Asp Ile Lys Asp Ile Arg Glu Val
165 170 175
Ile Ala Asn Gly Glu Ile Ile Phe Lys Leu Asp Gly Asp Ile Asp Arg
180 185 190
Thr Gln Phe Ile Trp Met Lys Tyr Phe Ser Ile Phe Asn Thr Glu Leu
195 200 205
Ser Gln Ser Asn Ile Glu Glu Arg Tyr Lys Ile Gln Ser Tyr Ser Glu
210 215 220
Tyr Leu Lys Asp Phe Trp Gly Asn Pro Leu Met Tyr Asn Lys Glu Tyr
225 230 235 240
Tyr Met Phe Asn Ala Gly Asn Lys Asn Ser Tyr Ile Lys Leu Lys Lys
245 250 255
Asp Ser Pro Val Gly Glu Ile Leu Thr Arg Ser Lys Tyr Asn Gln Asn
260 265 270
Ser Lys Tyr Ile Asn Tyr Arg Asp Leu Tyr Ile Gly Glu Lys Phe Ile
275 280 285
Ile Arg Arg Lys Ser Asn Ser Gln Ser Ile Asn Asp Asp Ile Val Arg
290 295 300
Lys Glu Asp Tyr Ile Tyr Leu Asp Phe Phe Asn Leu Asn Gln Glu Trp
305 310 315 320
Arg Val Tyr Thr Tyr Lys Tyr Phe Lys Lys Glu Glu Met Lys Leu Phe
325 330 335
Leu Ala Pro Ile Tyr Asp Ser Asp Glu Phe Tyr Asn Thr Ile Gln Ile
340 345 350
Lys Glu Tyr Asp Glu Gln Pro Thr Tyr Ser Cys Gln Leu Leu Phe Lys
355 360 365
Lys Asp Glu Glu Ser Thr Asp Glu Ile Gly Leu Ile Gly Ile His Arg
370 375 380
Phe Tyr Glu Ser Gly Ile Val Phe Glu Glu Tyr Lys Asp Tyr Phe Cys
385 390 395 400
Ile Ser Lys Trp Tyr Leu Lys Glu Val Lys Arg Lys Pro Tyr Asn Leu
405 410 415
Lys Leu Gly Cys Asn Trp Gln Phe Ile Pro Lys Asp Glu Gly Trp Thr
420 425 430
Glu Lys Leu Ala Ala Ala Leu Glu His His His His His His
435 440 445
<210> 45
<211> 1306
<212> DNA
<213> 人工序列
<220>
<223> rHC/A 变体Y1117V H1253K的核苷酸序列 (His-标签的)
<400> 45
atgatcatca atactagcat tctgaacctg cgttacgaga gcaatcatct gattgatctg 60
agccgttatg caagcaagat caacatcggt agcaaggtca attttgaccc gatcgataag 120
aaccagatcc agctgtttaa tctggaatcg agcaaaattg aggttatcct gaaaaacgcc 180
attgtctaca actccatgta cgagaatttc tccaccagct tctggattcg catcccgaaa 240
tacttcaaca gcattagcct gaacaacgag tatactatca tcaactgtat ggagaacaac 300
agcggttgga aggtgtctct gaactatggt gagatcattt ggaccttgca ggacacccaa 360
gagatcaagc agcgcgtcgt gttcaagtac tctcaaatga tcaacatttc cgattacatt 420
aatcgttgga tcttcgtgac cattacgaat aaccgtctga ataacagcaa gatttacatc 480
aatggtcgct tgatcgatca gaaaccgatt agcaacctgg gtaatatcca cgcaagcaac 540
aacattatgt tcaaattgga cggttgccgc gatacccatc gttatatctg gatcaagtat 600
ttcaacctgt ttgataaaga actgaatgag aaggagatca aagatttgta tgacaaccaa 660
tctaacagcg gcattttgaa ggacttctgg ggcgattatc tgcaatacga taagccgtac 720
tatatgctga acctggttga tccgaacaaa tatgtggatg tcaataatgt gggtattcgt 780
ggttacatgt atttgaaggg tccgcgtggc agcgttatga cgaccaacat ttacctgaac 840
tctagcctgt accgtggtac gaaattcatc attaagaaat atgccagcgg caacaaagat 900
aacattgtgc gtaataacga tcgtgtctac atcaacgtgg tcgtgaagaa taaagagtac 960
cgtctggcga ccaacgcttc gcaggcgggt gttgagaaaa ttctgagcgc gttggagatc 1020
cctgatgtcg gtaatctgag ccaagtcgtg gttatgaaga gcaagaacga ccagggtatc 1080
actaacaagt gcaagatgaa cctgcaagac aacaatggta acgacatcgg ctttattggt 1140
ttcaaacagt tcaacaatat tgctaaactg gtagcgagca attggtacaa tcgtcagatt 1200
gagcgcagca gccgtacttt gggctgtagc tgggagttta tcccggtcga tgatggttgg 1260
ggcgaacgtc cgctgcacca tcaccatcac catcaccatc accatt 1306
<210> 46
<211> 435
<212> PRT
<213> 人工序列
<220>
<223> rHC/A 变体Y1117V H1253K的多肽序列 (His-标签的)
<400> 46
Met Ile Ile Asn Thr Ser Ile Leu Asn Leu Arg Tyr Glu Ser Asn His
1 5 10 15
Leu Ile Asp Leu Ser Arg Tyr Ala Ser Lys Ile Asn Ile Gly Ser Lys
20 25 30
Val Asn Phe Asp Pro Ile Asp Lys Asn Gln Ile Gln Leu Phe Asn Leu
35 40 45
Glu Ser Ser Lys Ile Glu Val Ile Leu Lys Asn Ala Ile Val Tyr Asn
50 55 60
Ser Met Tyr Glu Asn Phe Ser Thr Ser Phe Trp Ile Arg Ile Pro Lys
65 70 75 80
Tyr Phe Asn Ser Ile Ser Leu Asn Asn Glu Tyr Thr Ile Ile Asn Cys
85 90 95
Met Glu Asn Asn Ser Gly Trp Lys Val Ser Leu Asn Tyr Gly Glu Ile
100 105 110
Ile Trp Thr Leu Gln Asp Thr Gln Glu Ile Lys Gln Arg Val Val Phe
115 120 125
Lys Tyr Ser Gln Met Ile Asn Ile Ser Asp Tyr Ile Asn Arg Trp Ile
130 135 140
Phe Val Thr Ile Thr Asn Asn Arg Leu Asn Asn Ser Lys Ile Tyr Ile
145 150 155 160
Asn Gly Arg Leu Ile Asp Gln Lys Pro Ile Ser Asn Leu Gly Asn Ile
165 170 175
His Ala Ser Asn Asn Ile Met Phe Lys Leu Asp Gly Cys Arg Asp Thr
180 185 190
His Arg Tyr Ile Trp Ile Lys Tyr Phe Asn Leu Phe Asp Lys Glu Leu
195 200 205
Asn Glu Lys Glu Ile Lys Asp Leu Tyr Asp Asn Gln Ser Asn Ser Gly
210 215 220
Ile Leu Lys Asp Phe Trp Gly Asp Tyr Leu Gln Tyr Asp Lys Pro Tyr
225 230 235 240
Tyr Met Leu Asn Leu Val Asp Pro Asn Lys Tyr Val Asp Val Asn Asn
245 250 255
Val Gly Ile Arg Gly Tyr Met Tyr Leu Lys Gly Pro Arg Gly Ser Val
260 265 270
Met Thr Thr Asn Ile Tyr Leu Asn Ser Ser Leu Tyr Arg Gly Thr Lys
275 280 285
Phe Ile Ile Lys Lys Tyr Ala Ser Gly Asn Lys Asp Asn Ile Val Arg
290 295 300
Asn Asn Asp Arg Val Tyr Ile Asn Val Val Val Lys Asn Lys Glu Tyr
305 310 315 320
Arg Leu Ala Thr Asn Ala Ser Gln Ala Gly Val Glu Lys Ile Leu Ser
325 330 335
Ala Leu Glu Ile Pro Asp Val Gly Asn Leu Ser Gln Val Val Val Met
340 345 350
Lys Ser Lys Asn Asp Gln Gly Ile Thr Asn Lys Cys Lys Met Asn Leu
355 360 365
Gln Asp Asn Asn Gly Asn Asp Ile Gly Phe Ile Gly Phe Lys Gln Phe
370 375 380
Asn Asn Ile Ala Lys Leu Val Ala Ser Asn Trp Tyr Asn Arg Gln Ile
385 390 395 400
Glu Arg Ser Ser Arg Thr Leu Gly Cys Ser Trp Glu Phe Ile Pro Val
405 410 415
Asp Asp Gly Trp Gly Glu Arg Pro Leu His His His His His His His
420 425 430
His His His
435
<210> 47
<211> 1308
<212> DNA
<213> 人工序列
<220>
<223> rHC/A 变体Y1117V F1252Y H1253K L1278F的核苷酸序列
(His-标签的)
<400> 47
atgatcatca atactagcat tctgaacctg cgttacgaga gcaatcatct gattgatctg 60
agccgttatg caagcaagat caacatcggt agcaaggtca attttgaccc gatcgataag 120
aaccagatcc agctgtttaa tctggaatcg agcaaaattg aggttatcct gaaaaacgcc 180
attgtctaca actccatgta cgagaatttc tccaccagct tctggattcg catcccgaaa 240
tacttcaaca gcattagcct gaacaacgag tatactatca tcaactgtat ggagaacaac 300
agcggttgga aggtgtctct gaactatggt gagatcattt ggaccttgca ggacacccaa 360
gagatcaagc agcgcgtcgt gttcaagtac tctcaaatga tcaacatttc cgattacatt 420
aatcgttgga tcttcgtgac cattacgaat aaccgtctga ataacagcaa gatttacatc 480
aatggtcgct tgatcgatca gaaaccgatt agcaacctgg gtaatatcca cgcaagcaac 540
aacattatgt tcaaattgga cggttgccgc gatacccatc gttatatctg gatcaagtat 600
ttcaacctgt ttgataaaga actgaatgag aaggagatca aagatttgta tgacaaccaa 660
tctaacagcg gcattttgaa ggacttctgg ggcgattatc tgcaatacga taagccgtac 720
tatatgctga acctggttga tccgaacaaa tatgtggatg tcaataatgt gggtattcgt 780
ggttacatgt atttgaaggg tccgcgtggc agcgttatga cgaccaacat ttacctgaac 840
tctagcctgt accgtggtac gaaattcatc attaagaaat atgccagcgg caacaaagat 900
aacattgtgc gtaataacga tcgtgtctac atcaacgtgg tcgtgaagaa taaagagtac 960
cgtctggcga ccaacgcttc gcaggcgggt gttgagaaaa ttctgagcgc gttggagatc 1020
cctgatgtcg gtaatctgag ccaagtcgtg gttatgaaga gcaagaacga ccagggtatc 1080
actaacaagt gcaagatgaa cctgcaagac aacaatggta acgacatcgg ctttattggt 1140
tacaaacagt tcaacaatat tgctaaactg gtagcgagca attggtacaa tcgtcagatt 1200
gagcgcagca gccgtacttt tggctgtagc tgggagttta tcccggtcga tgatggttgg 1260
ggcgaacgtc cgctgcacca tcaccatcac catcaccatc accattaa 1308
<210> 48
<211> 435
<212> PRT
<213> 人工序列
<220>
<223> rHC/A 变体Y1117V F1252Y H1253K L1278F的多肽序列(His-标签的)
<400> 48
Met Ile Ile Asn Thr Ser Ile Leu Asn Leu Arg Tyr Glu Ser Asn His
1 5 10 15
Leu Ile Asp Leu Ser Arg Tyr Ala Ser Lys Ile Asn Ile Gly Ser Lys
20 25 30
Val Asn Phe Asp Pro Ile Asp Lys Asn Gln Ile Gln Leu Phe Asn Leu
35 40 45
Glu Ser Ser Lys Ile Glu Val Ile Leu Lys Asn Ala Ile Val Tyr Asn
50 55 60
Ser Met Tyr Glu Asn Phe Ser Thr Ser Phe Trp Ile Arg Ile Pro Lys
65 70 75 80
Tyr Phe Asn Ser Ile Ser Leu Asn Asn Glu Tyr Thr Ile Ile Asn Cys
85 90 95
Met Glu Asn Asn Ser Gly Trp Lys Val Ser Leu Asn Tyr Gly Glu Ile
100 105 110
Ile Trp Thr Leu Gln Asp Thr Gln Glu Ile Lys Gln Arg Val Val Phe
115 120 125
Lys Tyr Ser Gln Met Ile Asn Ile Ser Asp Tyr Ile Asn Arg Trp Ile
130 135 140
Phe Val Thr Ile Thr Asn Asn Arg Leu Asn Asn Ser Lys Ile Tyr Ile
145 150 155 160
Asn Gly Arg Leu Ile Asp Gln Lys Pro Ile Ser Asn Leu Gly Asn Ile
165 170 175
His Ala Ser Asn Asn Ile Met Phe Lys Leu Asp Gly Cys Arg Asp Thr
180 185 190
His Arg Tyr Ile Trp Ile Lys Tyr Phe Asn Leu Phe Asp Lys Glu Leu
195 200 205
Asn Glu Lys Glu Ile Lys Asp Leu Tyr Asp Asn Gln Ser Asn Ser Gly
210 215 220
Ile Leu Lys Asp Phe Trp Gly Asp Tyr Leu Gln Tyr Asp Lys Pro Tyr
225 230 235 240
Tyr Met Leu Asn Leu Val Asp Pro Asn Lys Tyr Val Asp Val Asn Asn
245 250 255
Val Gly Ile Arg Gly Tyr Met Tyr Leu Lys Gly Pro Arg Gly Ser Val
260 265 270
Met Thr Thr Asn Ile Tyr Leu Asn Ser Ser Leu Tyr Arg Gly Thr Lys
275 280 285
Phe Ile Ile Lys Lys Tyr Ala Ser Gly Asn Lys Asp Asn Ile Val Arg
290 295 300
Asn Asn Asp Arg Val Tyr Ile Asn Val Val Val Lys Asn Lys Glu Tyr
305 310 315 320
Arg Leu Ala Thr Asn Ala Ser Gln Ala Gly Val Glu Lys Ile Leu Ser
325 330 335
Ala Leu Glu Ile Pro Asp Val Gly Asn Leu Ser Gln Val Val Val Met
340 345 350
Lys Ser Lys Asn Asp Gln Gly Ile Thr Asn Lys Cys Lys Met Asn Leu
355 360 365
Gln Asp Asn Asn Gly Asn Asp Ile Gly Phe Ile Gly Tyr Lys Gln Phe
370 375 380
Asn Asn Ile Ala Lys Leu Val Ala Ser Asn Trp Tyr Asn Arg Gln Ile
385 390 395 400
Glu Arg Ser Ser Arg Thr Phe Gly Cys Ser Trp Glu Phe Ile Pro Val
405 410 415
Asp Asp Gly Trp Gly Glu Arg Pro Leu His His His His His His His
420 425 430
His His His
435
<210> 49
<211> 1293
<212> DNA
<213> 人工序列
<220>
<223> rHC/A 变体Y1117V F1252Y H1253K L1278H的核苷酸序列
(His-标签的)
<400> 49
atgatcatca atactagcat tctgaacctg cgttacgaga gcaatcatct gattgatctg 60
agccgttatg caagcaagat caacatcggt agcaaggtca attttgaccc gatcgataag 120
aaccagatcc agctgtttaa tctggaatcg agcaaaattg aggttatcct gaaaaacgcc 180
attgtctaca actccatgta cgagaatttc tccaccagct tctggattcg catcccgaaa 240
tacttcaaca gcattagcct gaacaacgag tatactatca tcaactgtat ggagaacaac 300
agcggttgga aggtgtctct gaactatggt gagatcattt ggaccttgca ggacacccaa 360
gagatcaagc agcgcgtcgt gttcaagtac tctcaaatga tcaacatttc cgattacatt 420
aatcgttgga tcttcgtgac cattacgaat aaccgtctga ataacagcaa gatttacatc 480
aatggtcgct tgatcgatca gaaaccgatt agcaacctgg gtaatatcca cgcaagcaac 540
aacattatgt tcaaattgga cggttgccgc gatacccatc gttatatctg gatcaagtat 600
ttcaacctgt ttgataaaga actgaatgag aaggagatca aagatttgta tgacaaccaa 660
tctaacagcg gcattttgaa ggacttctgg ggcgattatc tgcaatacga taagccgtac 720
tatatgctga acctggttga tccgaacaaa tatgtggatg tcaataatgt gggtattcgt 780
ggttacatgt atttgaaggg tccgcgtggc agcgttatga cgaccaacat ttacctgaac 840
tctagcctgt accgtggtac gaaattcatc attaagaaat atgccagcgg caacaaagat 900
aacattgtgc gtaataacga tcgtgtctac atcaacgtgg tcgtgaagaa taaagagtac 960
cgtctggcga ccaacgcttc gcaggcgggt gttgagaaaa ttctgagcgc gttggagatc 1020
cctgatgtcg gtaatctgag ccaagtcgtg gttatgaaga gcaagaacga ccagggtatc 1080
actaacaagt gcaagatgaa cctgcaagac aacaatggta acgacatcgg ctttattggt 1140
tacaaacagt tcaacaatat tgctaaactg gtagcgagca attggtacaa tcgtcagatt 1200
gagcgcagca gccgtactca tggctgtagc tgggagttta tcccggtcga tgatggttgg 1260
ggcgaacgtc cgctgcacca tcaccatcac cat 1293
<210> 50
<211> 431
<212> PRT
<213> 人工序列
<220>
<223> rHC/A 变体Y1117V F1252Y H1253K L1278H的多肽序列
(His-标签的)
<400> 50
Met Ile Ile Asn Thr Ser Ile Leu Asn Leu Arg Tyr Glu Ser Asn His
1 5 10 15
Leu Ile Asp Leu Ser Arg Tyr Ala Ser Lys Ile Asn Ile Gly Ser Lys
20 25 30
Val Asn Phe Asp Pro Ile Asp Lys Asn Gln Ile Gln Leu Phe Asn Leu
35 40 45
Glu Ser Ser Lys Ile Glu Val Ile Leu Lys Asn Ala Ile Val Tyr Asn
50 55 60
Ser Met Tyr Glu Asn Phe Ser Thr Ser Phe Trp Ile Arg Ile Pro Lys
65 70 75 80
Tyr Phe Asn Ser Ile Ser Leu Asn Asn Glu Tyr Thr Ile Ile Asn Cys
85 90 95
Met Glu Asn Asn Ser Gly Trp Lys Val Ser Leu Asn Tyr Gly Glu Ile
100 105 110
Ile Trp Thr Leu Gln Asp Thr Gln Glu Ile Lys Gln Arg Val Val Phe
115 120 125
Lys Tyr Ser Gln Met Ile Asn Ile Ser Asp Tyr Ile Asn Arg Trp Ile
130 135 140
Phe Val Thr Ile Thr Asn Asn Arg Leu Asn Asn Ser Lys Ile Tyr Ile
145 150 155 160
Asn Gly Arg Leu Ile Asp Gln Lys Pro Ile Ser Asn Leu Gly Asn Ile
165 170 175
His Ala Ser Asn Asn Ile Met Phe Lys Leu Asp Gly Cys Arg Asp Thr
180 185 190
His Arg Tyr Ile Trp Ile Lys Tyr Phe Asn Leu Phe Asp Lys Glu Leu
195 200 205
Asn Glu Lys Glu Ile Lys Asp Leu Tyr Asp Asn Gln Ser Asn Ser Gly
210 215 220
Ile Leu Lys Asp Phe Trp Gly Asp Tyr Leu Gln Tyr Asp Lys Pro Tyr
225 230 235 240
Tyr Met Leu Asn Leu Val Asp Pro Asn Lys Tyr Val Asp Val Asn Asn
245 250 255
Val Gly Ile Arg Gly Tyr Met Tyr Leu Lys Gly Pro Arg Gly Ser Val
260 265 270
Met Thr Thr Asn Ile Tyr Leu Asn Ser Ser Leu Tyr Arg Gly Thr Lys
275 280 285
Phe Ile Ile Lys Lys Tyr Ala Ser Gly Asn Lys Asp Asn Ile Val Arg
290 295 300
Asn Asn Asp Arg Val Tyr Ile Asn Val Val Val Lys Asn Lys Glu Tyr
305 310 315 320
Arg Leu Ala Thr Asn Ala Ser Gln Ala Gly Val Glu Lys Ile Leu Ser
325 330 335
Ala Leu Glu Ile Pro Asp Val Gly Asn Leu Ser Gln Val Val Val Met
340 345 350
Lys Ser Lys Asn Asp Gln Gly Ile Thr Asn Lys Cys Lys Met Asn Leu
355 360 365
Gln Asp Asn Asn Gly Asn Asp Ile Gly Phe Ile Gly Tyr Lys Gln Phe
370 375 380
Asn Asn Ile Ala Lys Leu Val Ala Ser Asn Trp Tyr Asn Arg Gln Ile
385 390 395 400
Glu Arg Ser Ser Arg Thr His Gly Cys Ser Trp Glu Phe Ile Pro Val
405 410 415
Asp Asp Gly Trp Gly Glu Arg Pro Leu His His His His His His
420 425 430
<210> 51
<211> 1296
<212> PRT
<213> 肉毒梭菌
<400> 51
Met Pro Phe Val Asn Lys Gln Phe Asn Tyr Lys Asp Pro Val Asn Gly
1 5 10 15
Val Asp Ile Ala Tyr Ile Lys Ile Pro Asn Val Gly Gln Met Gln Pro
20 25 30
Val Lys Ala Phe Lys Ile His Asn Lys Ile Trp Val Ile Pro Glu Arg
35 40 45
Asp Thr Phe Thr Asn Pro Glu Glu Gly Asp Leu Asn Pro Pro Pro Glu
50 55 60
Ala Lys Gln Val Pro Val Ser Tyr Tyr Asp Ser Thr Tyr Leu Ser Thr
65 70 75 80
Asp Asn Glu Lys Asp Asn Tyr Leu Lys Gly Val Thr Lys Leu Phe Glu
85 90 95
Arg Ile Tyr Ser Thr Asp Leu Gly Arg Met Leu Leu Thr Ser Ile Val
100 105 110
Arg Gly Ile Pro Phe Trp Gly Gly Ser Thr Ile Asp Thr Glu Leu Lys
115 120 125
Val Ile Asp Thr Asn Cys Ile Asn Val Ile Gln Pro Asp Gly Ser Tyr
130 135 140
Arg Ser Glu Glu Leu Asn Leu Val Ile Ile Gly Pro Ser Ala Asp Ile
145 150 155 160
Ile Gln Phe Glu Cys Lys Ser Phe Gly His Glu Val Leu Asn Leu Thr
165 170 175
Arg Asn Gly Tyr Gly Ser Thr Gln Tyr Ile Arg Phe Ser Pro Asp Phe
180 185 190
Thr Phe Gly Phe Glu Glu Ser Leu Glu Val Asp Thr Asn Pro Leu Leu
195 200 205
Gly Ala Gly Lys Phe Ala Thr Asp Pro Ala Val Thr Leu Ala His Glu
210 215 220
Leu Ile His Ala Gly His Arg Leu Tyr Gly Ile Ala Ile Asn Pro Asn
225 230 235 240
Arg Val Phe Lys Val Asn Thr Asn Ala Tyr Tyr Glu Met Ser Gly Leu
245 250 255
Glu Val Ser Phe Glu Glu Leu Arg Thr Phe Gly Gly His Asp Ala Lys
260 265 270
Phe Ile Asp Ser Leu Gln Glu Asn Glu Phe Arg Leu Tyr Tyr Tyr Asn
275 280 285
Lys Phe Lys Asp Ile Ala Ser Thr Leu Asn Lys Ala Lys Ser Ile Val
290 295 300
Gly Thr Thr Ala Ser Leu Gln Tyr Met Lys Asn Val Phe Lys Glu Lys
305 310 315 320
Tyr Leu Leu Ser Glu Asp Thr Ser Gly Lys Phe Ser Val Asp Lys Leu
325 330 335
Lys Phe Asp Lys Leu Tyr Lys Met Leu Thr Glu Ile Tyr Thr Glu Asp
340 345 350
Asn Phe Val Lys Phe Phe Lys Val Leu Asn Arg Lys Thr Tyr Leu Asn
355 360 365
Phe Asp Lys Ala Val Phe Lys Ile Asn Ile Val Pro Lys Val Asn Tyr
370 375 380
Thr Ile Tyr Asp Gly Phe Asn Leu Arg Asn Thr Asn Leu Ala Ala Asn
385 390 395 400
Phe Asn Gly Gln Asn Thr Glu Ile Asn Asn Met Asn Phe Thr Lys Leu
405 410 415
Lys Asn Phe Thr Gly Leu Phe Glu Phe Tyr Lys Leu Leu Cys Val Arg
420 425 430
Gly Ile Ile Thr Ser Lys Thr Lys Ser Leu Asp Lys Gly Tyr Asn Lys
435 440 445
Ala Leu Asn Asp Leu Cys Ile Lys Val Asn Asn Trp Asp Leu Phe Phe
450 455 460
Ser Pro Ser Glu Asp Asn Phe Thr Asn Asp Leu Asn Lys Gly Glu Glu
465 470 475 480
Ile Thr Ser Asp Thr Asn Ile Glu Ala Ala Glu Glu Asn Ile Ser Leu
485 490 495
Asp Leu Ile Gln Gln Tyr Tyr Leu Thr Phe Asn Phe Asp Asn Glu Pro
500 505 510
Glu Asn Ile Ser Ile Glu Asn Leu Ser Ser Asp Ile Ile Gly Gln Leu
515 520 525
Glu Leu Met Pro Asn Ile Glu Arg Phe Pro Asn Gly Lys Lys Tyr Glu
530 535 540
Leu Asp Lys Tyr Thr Met Phe His Tyr Leu Arg Ala Gln Glu Phe Glu
545 550 555 560
His Gly Lys Ser Arg Ile Ala Leu Thr Asn Ser Val Asn Glu Ala Leu
565 570 575
Leu Asn Pro Ser Arg Val Tyr Thr Phe Phe Ser Ser Asp Tyr Val Lys
580 585 590
Lys Val Asn Lys Ala Thr Glu Ala Ala Met Phe Leu Gly Trp Val Glu
595 600 605
Gln Leu Val Tyr Asp Phe Thr Asp Glu Thr Ser Glu Val Ser Thr Thr
610 615 620
Asp Lys Ile Ala Asp Ile Thr Ile Ile Ile Pro Tyr Ile Gly Pro Ala
625 630 635 640
Leu Asn Ile Gly Asn Met Leu Tyr Lys Asp Asp Phe Val Gly Ala Leu
645 650 655
Ile Phe Ser Gly Ala Val Ile Leu Leu Glu Phe Ile Pro Glu Ile Ala
660 665 670
Ile Pro Val Leu Gly Thr Phe Ala Leu Val Ser Tyr Ile Ala Asn Lys
675 680 685
Val Leu Thr Val Gln Thr Ile Asp Asn Ala Leu Ser Lys Arg Asn Glu
690 695 700
Lys Trp Asp Glu Val Tyr Lys Tyr Ile Val Thr Asn Trp Leu Ala Lys
705 710 715 720
Val Asn Thr Gln Ile Asp Leu Ile Arg Lys Lys Met Lys Glu Ala Leu
725 730 735
Glu Asn Gln Ala Glu Ala Thr Lys Ala Ile Ile Asn Tyr Gln Tyr Asn
740 745 750
Gln Tyr Thr Glu Glu Glu Lys Asn Asn Ile Asn Phe Asn Ile Asp Asp
755 760 765
Leu Ser Ser Lys Leu Asn Glu Ser Ile Asn Lys Ala Met Ile Asn Ile
770 775 780
Asn Lys Phe Leu Asn Gln Cys Ser Val Ser Tyr Leu Met Asn Ser Met
785 790 795 800
Ile Pro Tyr Gly Val Lys Arg Leu Glu Asp Phe Asp Ala Ser Leu Lys
805 810 815
Asp Ala Leu Leu Lys Tyr Ile Tyr Asp Asn Arg Gly Thr Leu Ile Gly
820 825 830
Gln Val Asp Arg Leu Lys Asp Lys Val Asn Asn Thr Leu Ser Thr Asp
835 840 845
Ile Pro Phe Gln Leu Ser Lys Tyr Val Asp Asn Gln Arg Leu Leu Ser
850 855 860
Thr Phe Thr Glu Tyr Ile Lys Asn Ile Ile Asn Thr Ser Ile Leu Asn
865 870 875 880
Leu Arg Tyr Glu Ser Asn His Leu Ile Asp Leu Ser Arg Tyr Ala Ser
885 890 895
Lys Ile Asn Ile Gly Ser Lys Val Asn Phe Asp Pro Ile Asp Lys Asn
900 905 910
Gln Ile Gln Leu Phe Asn Leu Glu Ser Ser Lys Ile Glu Val Ile Leu
915 920 925
Lys Asn Ala Ile Val Tyr Asn Ser Met Tyr Glu Asn Phe Ser Thr Ser
930 935 940
Phe Trp Ile Arg Ile Pro Lys Tyr Phe Asn Ser Ile Ser Leu Asn Asn
945 950 955 960
Glu Tyr Thr Ile Ile Asn Cys Met Glu Asn Asn Ser Gly Trp Lys Val
965 970 975
Ser Leu Asn Tyr Gly Glu Ile Ile Trp Thr Leu Gln Asp Thr Gln Glu
980 985 990
Ile Lys Gln Arg Val Val Phe Lys Tyr Ser Gln Met Ile Asn Ile Ser
995 1000 1005
Asp Tyr Ile Asn Arg Trp Ile Phe Val Thr Ile Thr Asn Asn Arg
1010 1015 1020
Leu Asn Asn Ser Lys Ile Tyr Ile Asn Gly Arg Leu Ile Asp Gln
1025 1030 1035
Lys Pro Ile Ser Asn Leu Gly Asn Ile His Ala Ser Asn Asn Ile
1040 1045 1050
Met Phe Lys Leu Asp Gly Cys Arg Asp Thr His Arg Tyr Ile Trp
1055 1060 1065
Ile Lys Tyr Phe Asn Leu Phe Asp Lys Glu Leu Asn Glu Lys Glu
1070 1075 1080
Ile Lys Asp Leu Tyr Asp Asn Gln Ser Asn Ser Gly Ile Leu Lys
1085 1090 1095
Asp Phe Trp Gly Asp Tyr Leu Gln Tyr Asp Lys Pro Tyr Tyr Met
1100 1105 1110
Leu Asn Leu Tyr Asp Pro Asn Lys Tyr Val Asp Val Asn Asn Val
1115 1120 1125
Gly Ile Arg Gly Tyr Met Tyr Leu Lys Gly Pro Arg Gly Ser Val
1130 1135 1140
Met Thr Thr Asn Ile Tyr Leu Asn Ser Ser Leu Tyr Arg Gly Thr
1145 1150 1155
Lys Phe Ile Ile Lys Lys Tyr Ala Ser Gly Asn Lys Asp Asn Ile
1160 1165 1170
Val Arg Asn Asn Asp Arg Val Tyr Ile Asn Val Val Val Lys Asn
1175 1180 1185
Lys Glu Tyr Arg Leu Ala Thr Asn Ala Ser Gln Ala Gly Val Glu
1190 1195 1200
Lys Ile Leu Ser Ala Leu Glu Ile Pro Asp Val Gly Asn Leu Ser
1205 1210 1215
Gln Val Val Val Met Lys Ser Lys Asn Asp Gln Gly Ile Thr Asn
1220 1225 1230
Lys Cys Lys Met Asn Leu Gln Asp Asn Asn Gly Asn Asp Ile Gly
1235 1240 1245
Phe Ile Gly Phe His Gln Phe Asn Asn Ile Ala Lys Leu Val Ala
1250 1255 1260
Ser Asn Trp Tyr Asn Arg Gln Ile Glu Arg Ser Ser Arg Thr Leu
1265 1270 1275
Gly Cys Ser Trp Glu Phe Ile Pro Val Asp Asp Gly Trp Gly Glu
1280 1285 1290
Arg Pro Leu
1295
<210> 52
<211> 1291
<212> PRT
<213> 肉毒梭菌
<400> 52
Met Pro Val Thr Ile Asn Asn Phe Asn Tyr Asn Asp Pro Ile Asp Asn
1 5 10 15
Asn Asn Ile Ile Met Met Glu Pro Pro Phe Ala Arg Gly Thr Gly Arg
20 25 30
Tyr Tyr Lys Ala Phe Lys Ile Thr Asp Arg Ile Trp Ile Ile Pro Glu
35 40 45
Arg Tyr Thr Phe Gly Tyr Lys Pro Glu Asp Phe Asn Lys Ser Ser Gly
50 55 60
Ile Phe Asn Arg Asp Val Cys Glu Tyr Tyr Asp Pro Asp Tyr Leu Asn
65 70 75 80
Thr Asn Asp Lys Lys Asn Ile Phe Leu Gln Thr Met Ile Lys Leu Phe
85 90 95
Asn Arg Ile Lys Ser Lys Pro Leu Gly Glu Lys Leu Leu Glu Met Ile
100 105 110
Ile Asn Gly Ile Pro Tyr Leu Gly Asp Arg Arg Val Pro Leu Glu Glu
115 120 125
Phe Asn Thr Asn Ile Ala Ser Val Thr Val Asn Lys Leu Ile Ser Asn
130 135 140
Pro Gly Glu Val Glu Arg Lys Lys Gly Ile Phe Ala Asn Leu Ile Ile
145 150 155 160
Phe Gly Pro Gly Pro Val Leu Asn Glu Asn Glu Thr Ile Asp Ile Gly
165 170 175
Ile Gln Asn His Phe Ala Ser Arg Glu Gly Phe Gly Gly Ile Met Gln
180 185 190
Met Lys Phe Cys Pro Glu Tyr Val Ser Val Phe Asn Asn Val Gln Glu
195 200 205
Asn Lys Gly Ala Ser Ile Phe Asn Arg Arg Gly Tyr Phe Ser Asp Pro
210 215 220
Ala Leu Ile Leu Met His Glu Leu Ile His Val Leu His Gly Leu Tyr
225 230 235 240
Gly Ile Lys Val Asp Asp Leu Pro Ile Val Pro Asn Glu Lys Lys Phe
245 250 255
Phe Met Gln Ser Thr Asp Ala Ile Gln Ala Glu Glu Leu Tyr Thr Phe
260 265 270
Gly Gly Gln Asp Pro Ser Ile Ile Thr Pro Ser Thr Asp Lys Ser Ile
275 280 285
Tyr Asp Lys Val Leu Gln Asn Phe Arg Gly Ile Val Asp Arg Leu Asn
290 295 300
Lys Val Leu Val Cys Ile Ser Asp Pro Asn Ile Asn Ile Asn Ile Tyr
305 310 315 320
Lys Asn Lys Phe Lys Asp Lys Tyr Lys Phe Val Glu Asp Ser Glu Gly
325 330 335
Lys Tyr Ser Ile Asp Val Glu Ser Phe Asp Lys Leu Tyr Lys Ser Leu
340 345 350
Met Phe Gly Phe Thr Glu Thr Asn Ile Ala Glu Asn Tyr Lys Ile Lys
355 360 365
Thr Arg Ala Ser Tyr Phe Ser Asp Ser Leu Pro Pro Val Lys Ile Lys
370 375 380
Asn Leu Leu Asp Asn Glu Ile Tyr Thr Ile Glu Glu Gly Phe Asn Ile
385 390 395 400
Ser Asp Lys Asp Met Glu Lys Glu Tyr Arg Gly Gln Asn Lys Ala Ile
405 410 415
Asn Lys Gln Ala Tyr Glu Glu Ile Ser Lys Glu His Leu Ala Val Tyr
420 425 430
Lys Ile Gln Met Cys Lys Ser Val Lys Ala Pro Gly Ile Cys Ile Asp
435 440 445
Val Asp Asn Glu Asp Leu Phe Phe Ile Ala Asp Lys Asn Ser Phe Ser
450 455 460
Asp Asp Leu Ser Lys Asn Glu Arg Ile Glu Tyr Asn Thr Gln Ser Asn
465 470 475 480
Tyr Ile Glu Asn Asp Phe Pro Ile Asn Glu Leu Ile Leu Asp Thr Asp
485 490 495
Leu Ile Ser Lys Ile Glu Leu Pro Ser Glu Asn Thr Glu Ser Leu Thr
500 505 510
Asp Phe Asn Val Asp Val Pro Val Tyr Glu Lys Gln Pro Ala Ile Lys
515 520 525
Lys Ile Phe Thr Asp Glu Asn Thr Ile Phe Gln Tyr Leu Tyr Ser Gln
530 535 540
Thr Phe Pro Leu Asp Ile Arg Asp Ile Ser Leu Thr Ser Ser Phe Asp
545 550 555 560
Asp Ala Leu Leu Phe Ser Asn Lys Val Tyr Ser Phe Phe Ser Met Asp
565 570 575
Tyr Ile Lys Thr Ala Asn Lys Val Val Glu Ala Gly Leu Phe Ala Gly
580 585 590
Trp Val Lys Gln Ile Val Asn Asp Phe Val Ile Glu Ala Asn Lys Ser
595 600 605
Asn Thr Met Asp Lys Ile Ala Asp Ile Ser Leu Ile Val Pro Tyr Ile
610 615 620
Gly Leu Ala Leu Asn Val Gly Asn Glu Thr Ala Lys Gly Asn Phe Glu
625 630 635 640
Asn Ala Phe Glu Ile Ala Gly Ala Ser Ile Leu Leu Glu Phe Ile Pro
645 650 655
Glu Leu Leu Ile Pro Val Val Gly Ala Phe Leu Leu Glu Ser Tyr Ile
660 665 670
Asp Asn Lys Asn Lys Ile Ile Lys Thr Ile Asp Asn Ala Leu Thr Lys
675 680 685
Arg Asn Glu Lys Trp Ser Asp Met Tyr Gly Leu Ile Val Ala Gln Trp
690 695 700
Leu Ser Thr Val Asn Thr Gln Phe Tyr Thr Ile Lys Glu Gly Met Tyr
705 710 715 720
Lys Ala Leu Asn Tyr Gln Ala Gln Ala Leu Glu Glu Ile Ile Lys Tyr
725 730 735
Arg Tyr Asn Ile Tyr Ser Glu Lys Glu Lys Ser Asn Ile Asn Ile Asp
740 745 750
Phe Asn Asp Ile Asn Ser Lys Leu Asn Glu Gly Ile Asn Gln Ala Ile
755 760 765
Asp Asn Ile Asn Asn Phe Ile Asn Gly Cys Ser Val Ser Tyr Leu Met
770 775 780
Lys Lys Met Ile Pro Leu Ala Val Glu Lys Leu Leu Asp Phe Asp Asn
785 790 795 800
Thr Leu Lys Lys Asn Leu Leu Asn Tyr Ile Asp Glu Asn Lys Leu Tyr
805 810 815
Leu Ile Gly Ser Ala Glu Tyr Glu Lys Ser Lys Val Asn Lys Tyr Leu
820 825 830
Lys Thr Ile Met Pro Phe Asp Leu Ser Ile Tyr Thr Asn Asp Thr Ile
835 840 845
Leu Ile Glu Met Phe Asn Lys Tyr Asn Ser Glu Ile Leu Asn Asn Ile
850 855 860
Ile Leu Asn Leu Arg Tyr Lys Asp Asn Asn Leu Ile Asp Leu Ser Gly
865 870 875 880
Tyr Gly Ala Lys Val Glu Val Tyr Asp Gly Val Glu Leu Asn Asp Lys
885 890 895
Asn Gln Phe Lys Leu Thr Ser Ser Ala Asn Ser Lys Ile Arg Val Thr
900 905 910
Gln Asn Gln Asn Ile Ile Phe Asn Ser Val Phe Leu Asp Phe Ser Val
915 920 925
Ser Phe Trp Ile Arg Ile Pro Lys Tyr Lys Asn Asp Gly Ile Gln Asn
930 935 940
Tyr Ile His Asn Glu Tyr Thr Ile Ile Asn Cys Met Lys Asn Asn Ser
945 950 955 960
Gly Trp Lys Ile Ser Ile Arg Gly Asn Arg Ile Ile Trp Thr Leu Ile
965 970 975
Asp Ile Asn Gly Lys Thr Lys Ser Val Phe Phe Glu Tyr Asn Ile Arg
980 985 990
Glu Asp Ile Ser Glu Tyr Ile Asn Arg Trp Phe Phe Val Thr Ile Thr
995 1000 1005
Asn Asn Leu Asn Asn Ala Lys Ile Tyr Ile Asn Gly Lys Leu Glu
1010 1015 1020
Ser Asn Thr Asp Ile Lys Asp Ile Arg Glu Val Ile Ala Asn Gly
1025 1030 1035
Glu Ile Ile Phe Lys Leu Asp Gly Asp Ile Asp Arg Thr Gln Phe
1040 1045 1050
Ile Trp Met Lys Tyr Phe Ser Ile Phe Asn Thr Glu Leu Ser Gln
1055 1060 1065
Ser Asn Ile Glu Glu Arg Tyr Lys Ile Gln Ser Tyr Ser Glu Tyr
1070 1075 1080
Leu Lys Asp Phe Trp Gly Asn Pro Leu Met Tyr Asn Lys Glu Tyr
1085 1090 1095
Tyr Met Phe Asn Ala Gly Asn Lys Asn Ser Tyr Ile Lys Leu Lys
1100 1105 1110
Lys Asp Ser Pro Val Gly Glu Ile Leu Thr Arg Ser Lys Tyr Asn
1115 1120 1125
Gln Asn Ser Lys Tyr Ile Asn Tyr Arg Asp Leu Tyr Ile Gly Glu
1130 1135 1140
Lys Phe Ile Ile Arg Arg Lys Ser Asn Ser Gln Ser Ile Asn Asp
1145 1150 1155
Asp Ile Val Arg Lys Glu Asp Tyr Ile Tyr Leu Asp Phe Phe Asn
1160 1165 1170
Leu Asn Gln Glu Trp Arg Val Tyr Thr Tyr Lys Tyr Phe Lys Lys
1175 1180 1185
Glu Glu Glu Lys Leu Phe Leu Ala Pro Ile Ser Asp Ser Asp Glu
1190 1195 1200
Phe Tyr Asn Thr Ile Gln Ile Lys Glu Tyr Asp Glu Gln Pro Thr
1205 1210 1215
Tyr Ser Cys Gln Leu Leu Phe Lys Lys Asp Glu Glu Ser Thr Asp
1220 1225 1230
Glu Ile Gly Leu Ile Gly Ile His Arg Phe Tyr Glu Ser Gly Ile
1235 1240 1245
Val Phe Glu Glu Tyr Lys Asp Tyr Phe Cys Ile Ser Lys Trp Tyr
1250 1255 1260
Leu Lys Glu Val Lys Arg Lys Pro Tyr Asn Leu Lys Leu Gly Cys
1265 1270 1275
Asn Trp Gln Phe Ile Pro Lys Asp Glu Gly Trp Thr Glu
1280 1285 1290
<210> 53
<211> 1291
<212> PRT
<213> 肉毒梭菌
<400> 53
Met Pro Ile Thr Ile Asn Asn Phe Asn Tyr Ser Asp Pro Val Asp Asn
1 5 10 15
Lys Asn Ile Leu Tyr Leu Asp Thr His Leu Asn Thr Leu Ala Asn Glu
20 25 30
Pro Glu Lys Ala Phe Arg Ile Thr Gly Asn Ile Trp Val Ile Pro Asp
35 40 45
Arg Phe Ser Arg Asn Ser Asn Pro Asn Leu Asn Lys Pro Pro Arg Val
50 55 60
Thr Ser Pro Lys Ser Gly Tyr Tyr Asp Pro Asn Tyr Leu Ser Thr Asp
65 70 75 80
Ser Asp Lys Asp Pro Phe Leu Lys Glu Ile Ile Lys Leu Phe Lys Arg
85 90 95
Ile Asn Ser Arg Glu Ile Gly Glu Glu Leu Ile Tyr Arg Leu Ser Thr
100 105 110
Asp Ile Pro Phe Pro Gly Asn Asn Asn Thr Pro Ile Asn Thr Phe Asp
115 120 125
Phe Asp Val Asp Phe Asn Ser Val Asp Val Lys Thr Arg Gln Gly Asn
130 135 140
Asn Trp Val Lys Thr Gly Ser Ile Asn Pro Ser Val Ile Ile Thr Gly
145 150 155 160
Pro Arg Glu Asn Ile Ile Asp Pro Glu Thr Ser Thr Phe Lys Leu Thr
165 170 175
Asn Asn Thr Phe Ala Ala Gln Glu Gly Phe Gly Ala Leu Ser Ile Ile
180 185 190
Ser Ile Ser Pro Arg Phe Met Leu Thr Tyr Ser Asn Ala Thr Asn Asp
195 200 205
Val Gly Glu Gly Arg Phe Ser Lys Ser Glu Phe Cys Met Asp Pro Ile
210 215 220
Leu Ile Leu Met His Glu Leu Asn His Ala Met His Asn Leu Tyr Gly
225 230 235 240
Ile Ala Ile Pro Asn Asp Gln Thr Ile Ser Ser Val Thr Ser Asn Ile
245 250 255
Phe Tyr Ser Gln Tyr Asn Val Lys Leu Glu Tyr Ala Glu Ile Tyr Ala
260 265 270
Phe Gly Gly Pro Thr Ile Asp Leu Ile Pro Lys Ser Ala Arg Lys Tyr
275 280 285
Phe Glu Glu Lys Ala Leu Asp Tyr Tyr Arg Ser Ile Ala Lys Arg Leu
290 295 300
Asn Ser Ile Thr Thr Ala Asn Pro Ser Ser Phe Asn Lys Tyr Ile Gly
305 310 315 320
Glu Tyr Lys Gln Lys Leu Ile Arg Lys Tyr Arg Phe Val Val Glu Ser
325 330 335
Ser Gly Glu Val Thr Val Asn Arg Asn Lys Phe Val Glu Leu Tyr Asn
340 345 350
Glu Leu Thr Gln Ile Phe Thr Glu Phe Asn Tyr Ala Lys Ile Tyr Asn
355 360 365
Val Gln Asn Arg Lys Ile Tyr Leu Ser Asn Val Tyr Thr Pro Val Thr
370 375 380
Ala Asn Ile Leu Asp Asp Asn Val Tyr Asp Ile Gln Asn Gly Phe Asn
385 390 395 400
Ile Pro Lys Ser Asn Leu Asn Val Leu Phe Met Gly Gln Asn Leu Ser
405 410 415
Arg Asn Pro Ala Leu Arg Lys Val Asn Pro Glu Asn Met Leu Tyr Leu
420 425 430
Phe Thr Lys Phe Cys His Lys Ala Ile Asp Gly Arg Ser Leu Tyr Asn
435 440 445
Lys Thr Leu Asp Cys Arg Glu Leu Leu Val Lys Asn Thr Asp Leu Pro
450 455 460
Phe Ile Gly Asp Ile Ser Asp Val Lys Thr Asp Ile Phe Leu Arg Lys
465 470 475 480
Asp Ile Asn Glu Glu Thr Glu Val Ile Tyr Tyr Pro Asp Asn Val Ser
485 490 495
Val Asp Gln Val Ile Leu Ser Lys Asn Thr Ser Glu His Gly Gln Leu
500 505 510
Asp Leu Leu Tyr Pro Ser Ile Asp Ser Glu Ser Glu Ile Leu Pro Gly
515 520 525
Glu Asn Gln Val Phe Tyr Asp Asn Arg Thr Gln Asn Val Asp Tyr Leu
530 535 540
Asn Ser Tyr Tyr Tyr Leu Glu Ser Gln Lys Leu Ser Asp Asn Val Glu
545 550 555 560
Asp Phe Thr Phe Thr Arg Ser Ile Glu Glu Ala Leu Asp Asn Ser Ala
565 570 575
Lys Val Tyr Thr Tyr Phe Pro Thr Leu Ala Asn Lys Val Asn Ala Gly
580 585 590
Val Gln Gly Gly Leu Phe Leu Met Trp Ala Asn Asp Val Val Glu Asp
595 600 605
Phe Thr Thr Asn Ile Leu Arg Lys Asp Thr Leu Asp Lys Ile Ser Asp
610 615 620
Val Ser Ala Ile Ile Pro Tyr Ile Gly Pro Ala Leu Asn Ile Ser Asn
625 630 635 640
Ser Val Arg Arg Gly Asn Phe Thr Glu Ala Phe Ala Val Thr Gly Val
645 650 655
Thr Ile Leu Leu Glu Ala Phe Pro Glu Phe Thr Ile Pro Ala Leu Gly
660 665 670
Ala Phe Val Ile Tyr Ser Lys Val Gln Glu Arg Asn Glu Ile Ile Lys
675 680 685
Thr Ile Asp Asn Cys Leu Glu Gln Arg Ile Lys Arg Trp Lys Asp Ser
690 695 700
Tyr Glu Trp Met Met Gly Thr Trp Leu Ser Arg Ile Ile Thr Gln Phe
705 710 715 720
Asn Asn Ile Ser Tyr Gln Met Tyr Asp Ser Leu Asn Tyr Gln Ala Gly
725 730 735
Ala Ile Lys Ala Lys Ile Asp Leu Glu Tyr Lys Lys Tyr Ser Gly Ser
740 745 750
Asp Lys Glu Asn Ile Lys Ser Gln Val Glu Asn Leu Lys Asn Ser Leu
755 760 765
Asp Val Lys Ile Ser Glu Ala Met Asn Asn Ile Asn Lys Phe Ile Arg
770 775 780
Glu Cys Ser Val Thr Tyr Leu Phe Lys Asn Met Leu Pro Lys Val Ile
785 790 795 800
Asp Glu Leu Asn Glu Phe Asp Arg Asn Thr Lys Ala Lys Leu Ile Asn
805 810 815
Leu Ile Asp Ser His Asn Ile Ile Leu Val Gly Glu Val Asp Lys Leu
820 825 830
Lys Ala Lys Val Asn Asn Ser Phe Gln Asn Thr Ile Pro Phe Asn Ile
835 840 845
Phe Ser Tyr Thr Asn Asn Ser Leu Leu Lys Asp Ile Ile Asn Glu Tyr
850 855 860
Phe Asn Asn Ile Asn Asp Ser Lys Ile Leu Ser Leu Gln Asn Arg Lys
865 870 875 880
Asn Thr Leu Val Asp Thr Ser Gly Tyr Asn Ala Glu Val Ser Glu Glu
885 890 895
Gly Asp Val Gln Leu Asn Pro Ile Phe Pro Phe Asp Phe Lys Leu Gly
900 905 910
Ser Ser Gly Glu Asp Arg Gly Lys Val Ile Val Thr Gln Asn Glu Asn
915 920 925
Ile Val Tyr Asn Ser Met Tyr Glu Ser Phe Ser Ile Ser Phe Trp Ile
930 935 940
Arg Ile Asn Lys Trp Val Ser Asn Leu Pro Gly Tyr Thr Ile Ile Asp
945 950 955 960
Ser Val Lys Asn Asn Ser Gly Trp Ser Ile Gly Ile Ile Ser Asn Phe
965 970 975
Leu Val Phe Thr Leu Lys Gln Asn Glu Asp Ser Glu Gln Ser Ile Asn
980 985 990
Phe Ser Tyr Asp Ile Ser Asn Asn Ala Pro Gly Tyr Asn Lys Trp Phe
995 1000 1005
Phe Val Thr Val Thr Asn Asn Met Met Gly Asn Met Lys Ile Tyr
1010 1015 1020
Ile Asn Gly Lys Leu Ile Asp Thr Ile Lys Val Lys Glu Leu Thr
1025 1030 1035
Gly Ile Asn Phe Ser Lys Thr Ile Thr Phe Glu Ile Asn Lys Ile
1040 1045 1050
Pro Asp Thr Gly Leu Ile Thr Ser Asp Ser Asp Asn Ile Asn Met
1055 1060 1065
Trp Ile Arg Asp Phe Tyr Ile Phe Ala Lys Glu Leu Asp Gly Lys
1070 1075 1080
Asp Ile Asn Ile Leu Phe Asn Ser Leu Gln Tyr Thr Asn Val Val
1085 1090 1095
Lys Asp Tyr Trp Gly Asn Asp Leu Arg Tyr Asn Lys Glu Tyr Tyr
1100 1105 1110
Met Val Asn Ile Asp Tyr Leu Asn Arg Tyr Met Tyr Ala Asn Ser
1115 1120 1125
Arg Gln Ile Val Phe Asn Thr Arg Arg Asn Asn Asn Asp Phe Asn
1130 1135 1140
Glu Gly Tyr Lys Ile Ile Ile Lys Arg Ile Arg Gly Asn Thr Asn
1145 1150 1155
Asp Thr Arg Val Arg Gly Gly Asp Ile Leu Tyr Phe Asp Met Thr
1160 1165 1170
Ile Asn Asn Lys Ala Tyr Asn Leu Phe Met Lys Asn Glu Thr Met
1175 1180 1185
Tyr Ala Asp Asn His Ser Thr Glu Asp Ile Tyr Ala Ile Gly Leu
1190 1195 1200
Arg Glu Gln Thr Lys Asp Ile Asn Asp Asn Ile Ile Phe Gln Ile
1205 1210 1215
Gln Pro Met Asn Asn Thr Tyr Tyr Tyr Ala Ser Gln Ile Phe Lys
1220 1225 1230
Ser Asn Phe Asn Gly Glu Asn Ile Ser Gly Ile Cys Ser Ile Gly
1235 1240 1245
Thr Tyr Arg Phe Arg Leu Gly Gly Asp Trp Tyr Arg His Asn Tyr
1250 1255 1260
Leu Val Pro Thr Val Lys Gln Gly Asn Tyr Ala Ser Leu Leu Glu
1265 1270 1275
Ser Thr Ser Thr His Trp Gly Phe Val Pro Val Ser Glu
1280 1285 1290
<210> 54
<211> 1276
<212> PRT
<213> 肉毒梭菌
<400> 54
Met Thr Trp Pro Val Lys Asp Phe Asn Tyr Ser Asp Pro Val Asn Asp
1 5 10 15
Asn Asp Ile Leu Tyr Leu Arg Ile Pro Gln Asn Lys Leu Ile Thr Thr
20 25 30
Pro Val Lys Ala Phe Met Ile Thr Gln Asn Ile Trp Val Ile Pro Glu
35 40 45
Arg Phe Ser Ser Asp Thr Asn Pro Ser Leu Ser Lys Pro Pro Arg Pro
50 55 60
Thr Ser Lys Tyr Gln Ser Tyr Tyr Asp Pro Ser Tyr Leu Ser Thr Asp
65 70 75 80
Glu Gln Lys Asp Thr Phe Leu Lys Gly Ile Ile Lys Leu Phe Lys Arg
85 90 95
Ile Asn Glu Arg Asp Ile Gly Lys Lys Leu Ile Asn Tyr Leu Val Val
100 105 110
Gly Ser Pro Phe Met Gly Asp Ser Ser Thr Pro Glu Asp Thr Phe Asp
115 120 125
Phe Thr Arg His Thr Thr Asn Ile Ala Val Glu Lys Phe Glu Asn Gly
130 135 140
Ser Trp Lys Val Thr Asn Ile Ile Thr Pro Ser Val Leu Ile Phe Gly
145 150 155 160
Pro Leu Pro Asn Ile Leu Asp Tyr Thr Ala Ser Leu Thr Leu Gln Gly
165 170 175
Gln Gln Ser Asn Pro Ser Phe Glu Gly Phe Gly Thr Leu Ser Ile Leu
180 185 190
Lys Val Ala Pro Glu Phe Leu Leu Thr Phe Ser Asp Val Thr Ser Asn
195 200 205
Gln Ser Ser Ala Val Leu Gly Lys Ser Ile Phe Cys Met Asp Pro Val
210 215 220
Ile Ala Leu Met His Glu Leu Thr His Ser Leu His Gln Leu Tyr Gly
225 230 235 240
Ile Asn Ile Pro Ser Asp Lys Arg Ile Arg Pro Gln Val Ser Glu Gly
245 250 255
Phe Phe Ser Gln Asp Gly Pro Asn Val Gln Phe Glu Glu Leu Tyr Thr
260 265 270
Phe Gly Gly Leu Asp Val Glu Ile Ile Pro Gln Ile Glu Arg Ser Gln
275 280 285
Leu Arg Glu Lys Ala Leu Gly His Tyr Lys Asp Ile Ala Lys Arg Leu
290 295 300
Asn Asn Ile Asn Lys Thr Ile Pro Ser Ser Trp Ile Ser Asn Ile Asp
305 310 315 320
Lys Tyr Lys Lys Ile Phe Ser Glu Lys Tyr Asn Phe Asp Lys Asp Asn
325 330 335
Thr Gly Asn Phe Val Val Asn Ile Asp Lys Phe Asn Ser Leu Tyr Ser
340 345 350
Asp Leu Thr Asn Val Met Ser Glu Val Val Tyr Ser Ser Gln Tyr Asn
355 360 365
Val Lys Asn Arg Thr His Tyr Phe Ser Arg His Tyr Leu Pro Val Phe
370 375 380
Ala Asn Ile Leu Asp Asp Asn Ile Tyr Thr Ile Arg Asp Gly Phe Asn
385 390 395 400
Leu Thr Asn Lys Gly Phe Asn Ile Glu Asn Ser Gly Gln Asn Ile Glu
405 410 415
Arg Asn Pro Ala Leu Gln Lys Leu Ser Ser Glu Ser Val Val Asp Leu
420 425 430
Phe Thr Lys Val Cys Leu Arg Leu Thr Lys Asn Ser Arg Asp Asp Ser
435 440 445
Thr Cys Ile Lys Val Lys Asn Asn Arg Leu Pro Tyr Val Ala Asp Lys
450 455 460
Asp Ser Ile Ser Gln Glu Ile Phe Glu Asn Lys Ile Ile Thr Asp Glu
465 470 475 480
Thr Asn Val Gln Asn Tyr Ser Asp Lys Phe Ser Leu Asp Glu Ser Ile
485 490 495
Leu Asp Gly Gln Val Pro Ile Asn Pro Glu Ile Val Asp Pro Leu Leu
500 505 510
Pro Asn Val Asn Met Glu Pro Leu Asn Leu Pro Gly Glu Glu Ile Val
515 520 525
Phe Tyr Asp Asp Ile Thr Lys Tyr Val Asp Tyr Leu Asn Ser Tyr Tyr
530 535 540
Tyr Leu Glu Ser Gln Lys Leu Ser Asn Asn Val Glu Asn Ile Thr Leu
545 550 555 560
Thr Thr Ser Val Glu Glu Ala Leu Gly Tyr Ser Asn Lys Ile Tyr Thr
565 570 575
Phe Leu Pro Ser Leu Ala Glu Lys Val Asn Lys Gly Val Gln Ala Gly
580 585 590
Leu Phe Leu Asn Trp Ala Asn Glu Val Val Glu Asp Phe Thr Thr Asn
595 600 605
Ile Met Lys Lys Asp Thr Leu Asp Lys Ile Ser Asp Val Ser Val Ile
610 615 620
Ile Pro Tyr Ile Gly Pro Ala Leu Asn Ile Gly Asn Ser Ala Leu Arg
625 630 635 640
Gly Asn Phe Asn Gln Ala Phe Ala Thr Ala Gly Val Ala Phe Leu Leu
645 650 655
Glu Gly Phe Pro Glu Phe Thr Ile Pro Ala Leu Gly Val Phe Thr Phe
660 665 670
Tyr Ser Ser Ile Gln Glu Arg Glu Lys Ile Ile Lys Thr Ile Glu Asn
675 680 685
Cys Leu Glu Gln Arg Val Lys Arg Trp Lys Asp Ser Tyr Gln Trp Met
690 695 700
Val Ser Asn Trp Leu Ser Arg Ile Thr Thr Gln Phe Asn His Ile Asn
705 710 715 720
Tyr Gln Met Tyr Asp Ser Leu Ser Tyr Gln Ala Asp Ala Ile Lys Ala
725 730 735
Lys Ile Asp Leu Glu Tyr Lys Lys Tyr Ser Gly Ser Asp Lys Glu Asn
740 745 750
Ile Lys Ser Gln Val Glu Asn Leu Lys Asn Ser Leu Asp Val Lys Ile
755 760 765
Ser Glu Ala Met Asn Asn Ile Asn Lys Phe Ile Arg Glu Cys Ser Val
770 775 780
Thr Tyr Leu Phe Lys Asn Met Leu Pro Lys Val Ile Asp Glu Leu Asn
785 790 795 800
Lys Phe Asp Leu Arg Thr Lys Thr Glu Leu Ile Asn Leu Ile Asp Ser
805 810 815
His Asn Ile Ile Leu Val Gly Glu Val Asp Arg Leu Lys Ala Lys Val
820 825 830
Asn Glu Ser Phe Glu Asn Thr Met Pro Phe Asn Ile Phe Ser Tyr Thr
835 840 845
Asn Asn Ser Leu Leu Lys Asp Ile Ile Asn Glu Tyr Phe Asn Ser Ile
850 855 860
Asn Asp Ser Lys Ile Leu Ser Leu Gln Asn Lys Lys Asn Ala Leu Val
865 870 875 880
Asp Thr Ser Gly Tyr Asn Ala Glu Val Arg Val Gly Asp Asn Val Gln
885 890 895
Leu Asn Thr Ile Tyr Thr Asn Asp Phe Lys Leu Ser Ser Ser Gly Asp
900 905 910
Lys Ile Ile Val Asn Leu Asn Asn Asn Ile Leu Tyr Ser Ala Ile Tyr
915 920 925
Glu Asn Ser Ser Val Ser Phe Trp Ile Lys Ile Ser Lys Asp Leu Thr
930 935 940
Asn Ser His Asn Glu Tyr Thr Ile Ile Asn Ser Ile Glu Gln Asn Ser
945 950 955 960
Gly Trp Lys Leu Cys Ile Arg Asn Gly Asn Ile Glu Trp Ile Leu Gln
965 970 975
Asp Val Asn Arg Lys Tyr Lys Ser Leu Ile Phe Asp Tyr Ser Glu Ser
980 985 990
Leu Ser His Thr Gly Tyr Thr Asn Lys Trp Phe Phe Val Thr Ile Thr
995 1000 1005
Asn Asn Ile Met Gly Tyr Met Lys Leu Tyr Ile Asn Gly Glu Leu
1010 1015 1020
Lys Gln Ser Gln Lys Ile Glu Asp Leu Asp Glu Val Lys Leu Asp
1025 1030 1035
Lys Thr Ile Val Phe Gly Ile Asp Glu Asn Ile Asp Glu Asn Gln
1040 1045 1050
Met Leu Trp Ile Arg Asp Phe Asn Ile Phe Ser Lys Glu Leu Ser
1055 1060 1065
Asn Glu Asp Ile Asn Ile Val Tyr Glu Gly Gln Ile Leu Arg Asn
1070 1075 1080
Val Ile Lys Asp Tyr Trp Gly Asn Pro Leu Lys Phe Asp Thr Glu
1085 1090 1095
Tyr Tyr Ile Ile Asn Asp Asn Tyr Ile Asp Arg Tyr Ile Ala Pro
1100 1105 1110
Glu Ser Asn Val Leu Val Leu Val Gln Tyr Pro Asp Arg Ser Lys
1115 1120 1125
Leu Tyr Thr Gly Asn Pro Ile Thr Ile Lys Ser Val Ser Asp Lys
1130 1135 1140
Asn Pro Tyr Ser Arg Ile Leu Asn Gly Asp Asn Ile Ile Leu His
1145 1150 1155
Met Leu Tyr Asn Ser Arg Lys Tyr Met Ile Ile Arg Asp Thr Asp
1160 1165 1170
Thr Ile Tyr Ala Thr Gln Gly Gly Glu Cys Ser Gln Asn Cys Val
1175 1180 1185
Tyr Ala Leu Lys Leu Gln Ser Asn Leu Gly Asn Tyr Gly Ile Gly
1190 1195 1200
Ile Phe Ser Ile Lys Asn Ile Val Ser Lys Asn Lys Tyr Cys Ser
1205 1210 1215
Gln Ile Phe Ser Ser Phe Arg Glu Asn Thr Met Leu Leu Ala Asp
1220 1225 1230
Ile Tyr Lys Pro Trp Arg Phe Ser Phe Lys Asn Ala Tyr Thr Pro
1235 1240 1245
Val Ala Val Thr Asn Tyr Glu Thr Lys Leu Leu Ser Thr Ser Ser
1250 1255 1260
Phe Trp Lys Phe Ile Ser Arg Asp Pro Gly Trp Val Glu
1265 1270 1275
<210> 55
<211> 1251
<212> PRT
<213> 肉毒梭菌
<400> 55
Met Pro Lys Ile Asn Ser Phe Asn Tyr Asn Asp Pro Val Asn Asp Arg
1 5 10 15
Thr Ile Leu Tyr Ile Lys Pro Gly Gly Cys Gln Glu Phe Tyr Lys Ser
20 25 30
Phe Asn Ile Met Lys Asn Ile Trp Ile Ile Pro Glu Arg Asn Val Ile
35 40 45
Gly Thr Thr Pro Gln Asp Phe His Pro Pro Thr Ser Leu Lys Asn Gly
50 55 60
Asp Ser Ser Tyr Tyr Asp Pro Asn Tyr Leu Gln Ser Asp Glu Glu Lys
65 70 75 80
Asp Arg Phe Leu Lys Ile Val Thr Lys Ile Phe Asn Arg Ile Asn Asn
85 90 95
Asn Leu Ser Gly Gly Ile Leu Leu Glu Glu Leu Ser Lys Ala Asn Pro
100 105 110
Tyr Leu Gly Asn Asp Asn Thr Pro Asp Asn Gln Phe His Ile Gly Asp
115 120 125
Ala Ser Ala Val Glu Ile Lys Phe Ser Asn Gly Ser Gln Asp Ile Leu
130 135 140
Leu Pro Asn Val Ile Ile Met Gly Ala Glu Pro Asp Leu Phe Glu Thr
145 150 155 160
Asn Ser Ser Asn Ile Ser Leu Arg Asn Asn Tyr Met Pro Ser Asn His
165 170 175
Arg Phe Gly Ser Ile Ala Ile Val Thr Phe Ser Pro Glu Tyr Ser Phe
180 185 190
Arg Phe Asn Asp Asn Cys Met Asn Glu Phe Ile Gln Asp Pro Ala Leu
195 200 205
Thr Leu Met His Glu Leu Ile His Ser Leu His Gly Leu Tyr Gly Ala
210 215 220
Lys Gly Ile Thr Thr Lys Tyr Thr Ile Thr Gln Lys Gln Asn Pro Leu
225 230 235 240
Ile Thr Asn Ile Arg Gly Thr Asn Ile Glu Glu Phe Leu Thr Phe Gly
245 250 255
Gly Thr Asp Leu Asn Ile Ile Thr Ser Ala Gln Ser Asn Asp Ile Tyr
260 265 270
Thr Asn Leu Leu Ala Asp Tyr Lys Lys Ile Ala Ser Lys Leu Ser Lys
275 280 285
Val Gln Val Ser Asn Pro Leu Leu Asn Pro Tyr Lys Asp Val Phe Glu
290 295 300
Ala Lys Tyr Gly Leu Asp Lys Asp Ala Ser Gly Ile Tyr Ser Val Asn
305 310 315 320
Ile Asn Lys Phe Asn Asp Ile Phe Lys Lys Leu Tyr Ser Phe Thr Glu
325 330 335
Phe Asp Leu Arg Thr Lys Phe Gln Val Lys Cys Arg Gln Thr Tyr Ile
340 345 350
Gly Gln Tyr Lys Tyr Phe Lys Leu Ser Asn Leu Leu Asn Asp Ser Ile
355 360 365
Tyr Asn Ile Ser Glu Gly Tyr Asn Ile Asn Asn Leu Lys Val Asn Phe
370 375 380
Arg Gly Gln Asn Ala Asn Leu Asn Pro Arg Ile Ile Thr Pro Ile Thr
385 390 395 400
Gly Arg Gly Leu Val Lys Lys Ile Ile Arg Phe Cys Lys Asn Ile Val
405 410 415
Ser Val Lys Gly Ile Arg Lys Ser Ile Cys Ile Glu Ile Asn Asn Gly
420 425 430
Glu Leu Phe Phe Val Ala Ser Glu Asn Ser Tyr Asn Asp Asp Asn Ile
435 440 445
Asn Thr Pro Lys Glu Ile Asp Asp Thr Val Thr Ser Asn Asn Asn Tyr
450 455 460
Glu Asn Asp Leu Asp Gln Val Ile Leu Asn Phe Asn Ser Glu Ser Ala
465 470 475 480
Pro Gly Leu Ser Asp Glu Lys Leu Asn Leu Thr Ile Gln Asn Asp Ala
485 490 495
Tyr Ile Pro Lys Tyr Asp Ser Asn Gly Thr Ser Asp Ile Glu Gln His
500 505 510
Asp Val Asn Glu Leu Asn Val Phe Phe Tyr Leu Asp Ala Gln Lys Val
515 520 525
Pro Glu Gly Glu Asn Asn Val Asn Leu Thr Ser Ser Ile Asp Thr Ala
530 535 540
Leu Leu Glu Gln Pro Lys Ile Tyr Thr Phe Phe Ser Ser Glu Phe Ile
545 550 555 560
Asn Asn Val Asn Lys Pro Val Gln Ala Ala Leu Phe Val Ser Trp Ile
565 570 575
Gln Gln Val Leu Val Asp Phe Thr Thr Glu Ala Asn Gln Lys Ser Thr
580 585 590
Val Asp Lys Ile Ala Asp Ile Ser Ile Val Val Pro Tyr Ile Gly Leu
595 600 605
Ala Leu Asn Ile Gly Asn Glu Ala Gln Lys Gly Asn Phe Lys Asp Ala
610 615 620
Leu Glu Leu Leu Gly Ala Gly Ile Leu Leu Glu Phe Glu Pro Glu Leu
625 630 635 640
Leu Ile Pro Thr Ile Leu Val Phe Thr Ile Lys Ser Phe Leu Gly Ser
645 650 655
Ser Asp Asn Lys Asn Lys Val Ile Lys Ala Ile Asn Asn Ala Leu Lys
660 665 670
Glu Arg Asp Glu Lys Trp Lys Glu Val Tyr Ser Phe Ile Val Ser Asn
675 680 685
Trp Met Thr Lys Ile Asn Thr Gln Phe Asn Lys Arg Lys Glu Gln Met
690 695 700
Tyr Gln Ala Leu Gln Asn Gln Val Asn Ala Ile Lys Thr Ile Ile Glu
705 710 715 720
Ser Lys Tyr Asn Ser Tyr Thr Leu Glu Glu Lys Asn Glu Leu Thr Asn
725 730 735
Lys Tyr Asp Ile Lys Gln Ile Glu Asn Glu Leu Asn Gln Lys Val Ser
740 745 750
Ile Ala Met Asn Asn Ile Asp Arg Phe Leu Thr Glu Ser Ser Ile Ser
755 760 765
Tyr Leu Met Lys Ile Ile Asn Glu Val Lys Ile Asn Lys Leu Arg Glu
770 775 780
Tyr Asp Glu Asn Val Lys Thr Tyr Leu Leu Asn Tyr Ile Ile Gln His
785 790 795 800
Gly Ser Ile Leu Gly Glu Ser Gln Gln Glu Leu Asn Ser Met Val Thr
805 810 815
Asp Thr Leu Asn Asn Ser Ile Pro Phe Lys Leu Ser Ser Tyr Thr Asp
820 825 830
Asp Lys Ile Leu Ile Ser Tyr Phe Asn Lys Phe Phe Lys Arg Ile Lys
835 840 845
Ser Ser Ser Val Leu Asn Met Arg Tyr Lys Asn Asp Lys Tyr Val Asp
850 855 860
Thr Ser Gly Tyr Asp Ser Asn Ile Asn Ile Asn Gly Asp Val Tyr Lys
865 870 875 880
Tyr Pro Thr Asn Lys Asn Gln Phe Gly Ile Tyr Asn Asp Lys Leu Ser
885 890 895
Glu Val Asn Ile Ser Gln Asn Asp Tyr Ile Ile Tyr Asp Asn Lys Tyr
900 905 910
Lys Asn Phe Ser Ile Ser Phe Trp Val Arg Ile Pro Asn Tyr Asp Asn
915 920 925
Lys Ile Val Asn Val Asn Asn Glu Tyr Thr Ile Ile Asn Cys Met Arg
930 935 940
Asp Asn Asn Ser Gly Trp Lys Val Ser Leu Asn His Asn Glu Ile Ile
945 950 955 960
Trp Thr Phe Glu Asp Asn Arg Gly Ile Asn Gln Lys Leu Ala Phe Asn
965 970 975
Tyr Gly Asn Ala Asn Gly Ile Ser Asp Tyr Ile Asn Lys Trp Ile Phe
980 985 990
Val Thr Ile Thr Asn Asp Arg Leu Gly Asp Ser Lys Leu Tyr Ile Asn
995 1000 1005
Gly Asn Leu Ile Asp Gln Lys Ser Ile Leu Asn Leu Gly Asn Ile
1010 1015 1020
His Val Ser Asp Asn Ile Leu Phe Lys Ile Val Asn Cys Ser Tyr
1025 1030 1035
Thr Arg Tyr Ile Gly Ile Arg Tyr Phe Asn Ile Phe Asp Lys Glu
1040 1045 1050
Leu Asp Glu Thr Glu Ile Gln Thr Leu Tyr Ser Asn Glu Pro Asn
1055 1060 1065
Thr Asn Ile Leu Lys Asp Phe Trp Gly Asn Tyr Leu Leu Tyr Asp
1070 1075 1080
Lys Glu Tyr Tyr Leu Leu Asn Val Leu Lys Pro Asn Asn Phe Ile
1085 1090 1095
Asp Arg Arg Lys Asp Ser Thr Leu Ser Ile Asn Asn Ile Arg Ser
1100 1105 1110
Thr Ile Leu Leu Ala Asn Arg Leu Tyr Ser Gly Ile Lys Val Lys
1115 1120 1125
Ile Gln Arg Val Asn Asn Ser Ser Thr Asn Asp Asn Leu Val Arg
1130 1135 1140
Lys Asn Asp Gln Val Tyr Ile Asn Phe Val Ala Ser Lys Thr His
1145 1150 1155
Leu Phe Pro Leu Tyr Ala Asp Thr Ala Thr Thr Asn Lys Glu Lys
1160 1165 1170
Thr Ile Lys Ile Ser Ser Ser Gly Asn Arg Phe Asn Gln Val Val
1175 1180 1185
Val Met Asn Ser Val Gly Asn Cys Thr Met Asn Phe Lys Asn Asn
1190 1195 1200
Asn Gly Asn Asn Ile Gly Leu Leu Gly Phe Lys Ala Asp Thr Val
1205 1210 1215
Val Ala Ser Thr Trp Tyr Tyr Thr His Met Arg Asp His Thr Asn
1220 1225 1230
Ser Asn Gly Cys Phe Trp Asn Phe Ile Ser Glu Glu His Gly Trp
1235 1240 1245
Gln Glu Lys
1250
<210> 56
<211> 1278
<212> PRT
<213> 肉毒梭菌
<400> 56
Met Pro Val Val Ile Asn Ser Phe Asn Tyr Asn Asp Pro Val Asn Asp
1 5 10 15
Asp Thr Ile Leu Tyr Met Gln Ile Pro Tyr Glu Glu Lys Ser Lys Lys
20 25 30
Tyr Tyr Lys Ala Phe Glu Ile Met Arg Asn Val Trp Ile Ile Pro Glu
35 40 45
Arg Asn Thr Ile Gly Thr Asp Pro Ser Asp Phe Asp Pro Pro Ala Ser
50 55 60
Leu Glu Asn Gly Ser Ser Ala Tyr Tyr Asp Pro Asn Tyr Leu Thr Thr
65 70 75 80
Asp Ala Glu Lys Asp Arg Tyr Leu Lys Thr Thr Ile Lys Leu Phe Lys
85 90 95
Arg Ile Asn Ser Asn Pro Ala Gly Glu Val Leu Leu Gln Glu Ile Ser
100 105 110
Tyr Ala Lys Pro Tyr Leu Gly Asn Glu His Thr Pro Ile Asn Glu Phe
115 120 125
His Pro Val Thr Arg Thr Thr Ser Val Asn Ile Lys Ser Ser Thr Asn
130 135 140
Val Lys Ser Ser Ile Ile Leu Asn Leu Leu Val Leu Gly Ala Gly Pro
145 150 155 160
Asp Ile Phe Glu Asn Ser Ser Tyr Pro Val Arg Lys Leu Met Asp Ser
165 170 175
Gly Gly Val Tyr Asp Pro Ser Asn Asp Gly Phe Gly Ser Ile Asn Ile
180 185 190
Val Thr Phe Ser Pro Glu Tyr Glu Tyr Thr Phe Asn Asp Ile Ser Gly
195 200 205
Gly Tyr Asn Ser Ser Thr Glu Ser Phe Ile Ala Asp Pro Ala Ile Ser
210 215 220
Leu Ala His Glu Leu Ile His Ala Leu His Gly Leu Tyr Gly Ala Arg
225 230 235 240
Gly Val Thr Tyr Lys Glu Thr Ile Lys Val Lys Gln Ala Pro Leu Met
245 250 255
Ile Ala Glu Lys Pro Ile Arg Leu Glu Glu Phe Leu Thr Phe Gly Gly
260 265 270
Gln Asp Leu Asn Ile Ile Thr Ser Ala Met Lys Glu Lys Ile Tyr Asn
275 280 285
Asn Leu Leu Ala Asn Tyr Glu Lys Ile Ala Thr Arg Leu Ser Arg Val
290 295 300
Asn Ser Ala Pro Pro Glu Tyr Asp Ile Asn Glu Tyr Lys Asp Tyr Phe
305 310 315 320
Gln Trp Lys Tyr Gly Leu Asp Lys Asn Ala Asp Gly Ser Tyr Thr Val
325 330 335
Asn Glu Asn Lys Phe Asn Glu Ile Tyr Lys Lys Leu Tyr Ser Phe Thr
340 345 350
Glu Ile Asp Leu Ala Asn Lys Phe Lys Val Lys Cys Arg Asn Thr Tyr
355 360 365
Phe Ile Lys Tyr Gly Phe Leu Lys Val Pro Asn Leu Leu Asp Asp Asp
370 375 380
Ile Tyr Thr Val Ser Glu Gly Phe Asn Ile Gly Asn Leu Ala Val Asn
385 390 395 400
Asn Arg Gly Gln Asn Ile Lys Leu Asn Pro Lys Ile Ile Asp Ser Ile
405 410 415
Pro Asp Lys Gly Leu Val Glu Lys Ile Val Lys Phe Cys Lys Ser Val
420 425 430
Ile Pro Arg Lys Gly Thr Lys Ala Pro Pro Arg Leu Cys Ile Arg Val
435 440 445
Asn Asn Arg Glu Leu Phe Phe Val Ala Ser Glu Ser Ser Tyr Asn Glu
450 455 460
Asn Asp Ile Asn Thr Pro Lys Glu Ile Asp Asp Thr Thr Asn Leu Asn
465 470 475 480
Asn Asn Tyr Arg Asn Asn Leu Asp Glu Val Ile Leu Asp Tyr Asn Ser
485 490 495
Glu Thr Ile Pro Gln Ile Ser Asn Gln Thr Leu Asn Thr Leu Val Gln
500 505 510
Asp Asp Ser Tyr Val Pro Arg Tyr Asp Ser Asn Gly Thr Ser Glu Ile
515 520 525
Glu Glu His Asn Val Val Asp Leu Asn Val Phe Phe Tyr Leu His Ala
530 535 540
Gln Lys Val Pro Glu Gly Glu Thr Asn Ile Ser Leu Thr Ser Ser Ile
545 550 555 560
Asp Thr Ala Leu Ser Glu Glu Ser Gln Val Tyr Thr Phe Phe Ser Ser
565 570 575
Glu Phe Ile Asn Thr Ile Asn Lys Pro Val His Ala Ala Leu Phe Ile
580 585 590
Ser Trp Ile Asn Gln Val Ile Arg Asp Phe Thr Thr Glu Ala Thr Gln
595 600 605
Lys Ser Thr Phe Asp Lys Ile Ala Asp Ile Ser Leu Val Val Pro Tyr
610 615 620
Val Gly Leu Ala Leu Asn Ile Gly Asn Glu Val Gln Lys Glu Asn Phe
625 630 635 640
Lys Glu Ala Phe Glu Leu Leu Gly Ala Gly Ile Leu Leu Glu Phe Val
645 650 655
Pro Glu Leu Leu Ile Pro Thr Ile Leu Val Phe Thr Ile Lys Ser Phe
660 665 670
Ile Gly Ser Ser Glu Asn Lys Asn Lys Ile Ile Lys Ala Ile Asn Asn
675 680 685
Ser Leu Met Glu Arg Glu Thr Lys Trp Lys Glu Ile Tyr Ser Trp Ile
690 695 700
Val Ser Asn Trp Leu Thr Arg Ile Asn Thr Gln Phe Asn Lys Arg Lys
705 710 715 720
Glu Gln Met Tyr Gln Ala Leu Gln Asn Gln Val Asp Ala Ile Lys Thr
725 730 735
Val Ile Glu Tyr Lys Tyr Asn Asn Tyr Thr Ser Asp Glu Arg Asn Arg
740 745 750
Leu Glu Ser Glu Tyr Asn Ile Asn Asn Ile Arg Glu Glu Leu Asn Lys
755 760 765
Lys Val Ser Leu Ala Met Glu Asn Ile Glu Arg Phe Ile Thr Glu Ser
770 775 780
Ser Ile Phe Tyr Leu Met Lys Leu Ile Asn Glu Ala Lys Val Ser Lys
785 790 795 800
Leu Arg Glu Tyr Asp Glu Gly Val Lys Glu Tyr Leu Leu Asp Tyr Ile
805 810 815
Ser Glu His Arg Ser Ile Leu Gly Asn Ser Val Gln Glu Leu Asn Asp
820 825 830
Leu Val Thr Ser Thr Leu Asn Asn Ser Ile Pro Phe Glu Leu Ser Ser
835 840 845
Tyr Thr Asn Asp Lys Ile Leu Ile Leu Tyr Phe Asn Lys Leu Tyr Lys
850 855 860
Lys Ile Lys Asp Asn Ser Ile Leu Asp Met Arg Tyr Glu Asn Asn Lys
865 870 875 880
Phe Ile Asp Ile Ser Gly Tyr Gly Ser Asn Ile Ser Ile Asn Gly Asp
885 890 895
Val Tyr Ile Tyr Ser Thr Asn Arg Asn Gln Phe Gly Ile Tyr Ser Ser
900 905 910
Lys Pro Ser Glu Val Asn Ile Ala Gln Asn Asn Asp Ile Ile Tyr Asn
915 920 925
Gly Arg Tyr Gln Asn Phe Ser Ile Ser Phe Trp Val Arg Ile Pro Lys
930 935 940
Tyr Phe Asn Lys Val Asn Leu Asn Asn Glu Tyr Thr Ile Ile Asp Cys
945 950 955 960
Ile Arg Asn Asn Asn Ser Gly Trp Lys Ile Ser Leu Asn Tyr Asn Lys
965 970 975
Ile Ile Trp Thr Leu Gln Asp Thr Ala Gly Asn Asn Gln Lys Leu Val
980 985 990
Phe Asn Tyr Thr Gln Met Ile Ser Ile Ser Asp Tyr Ile Asn Lys Trp
995 1000 1005
Ile Phe Val Thr Ile Thr Asn Asn Arg Leu Gly Asn Ser Arg Ile
1010 1015 1020
Tyr Ile Asn Gly Asn Leu Ile Asp Glu Lys Ser Ile Ser Asn Leu
1025 1030 1035
Gly Asp Ile His Val Ser Asp Asn Ile Leu Phe Lys Ile Val Gly
1040 1045 1050
Cys Asn Asp Thr Arg Tyr Val Gly Ile Arg Tyr Phe Lys Val Phe
1055 1060 1065
Asp Thr Glu Leu Gly Lys Thr Glu Ile Glu Thr Leu Tyr Ser Asp
1070 1075 1080
Glu Pro Asp Pro Ser Ile Leu Lys Asp Phe Trp Gly Asn Tyr Leu
1085 1090 1095
Leu Tyr Asn Lys Arg Tyr Tyr Leu Leu Asn Leu Leu Arg Thr Asp
1100 1105 1110
Lys Ser Ile Thr Gln Asn Ser Asn Phe Leu Asn Ile Asn Gln Gln
1115 1120 1125
Arg Gly Val Tyr Gln Lys Pro Asn Ile Phe Ser Asn Thr Arg Leu
1130 1135 1140
Tyr Thr Gly Val Glu Val Ile Ile Arg Lys Asn Gly Ser Thr Asp
1145 1150 1155
Ile Ser Asn Thr Asp Asn Phe Val Arg Lys Asn Asp Leu Ala Tyr
1160 1165 1170
Ile Asn Val Val Asp Arg Asp Val Glu Tyr Arg Leu Tyr Ala Asp
1175 1180 1185
Ile Ser Ile Ala Lys Pro Glu Lys Ile Ile Lys Leu Ile Arg Thr
1190 1195 1200
Ser Asn Ser Asn Asn Ser Leu Gly Gln Ile Ile Val Met Asp Ser
1205 1210 1215
Ile Gly Asn Asn Cys Thr Met Asn Phe Gln Asn Asn Asn Gly Gly
1220 1225 1230
Asn Ile Gly Leu Leu Gly Phe His Ser Asn Asn Leu Val Ala Ser
1235 1240 1245
Ser Trp Tyr Tyr Asn Asn Ile Arg Lys Asn Thr Ser Ser Asn Gly
1250 1255 1260
Cys Phe Trp Ser Phe Ile Ser Lys Glu His Gly Trp Gln Glu Asn
1265 1270 1275
<210> 57
<211> 1297
<212> PRT
<213> 肉毒梭菌（Clostridium botulinum）
<220>
<221> misc_feature
<222> (7)..(7)
<223> Xaa可以是任一天然存在的氨基酸
<400> 57
Met Pro Val Asn Ile Lys Xaa Phe Asn Tyr Asn Asp Pro Ile Asn Asn
1 5 10 15
Asp Asp Ile Ile Met Met Glu Pro Phe Asn Asp Pro Gly Pro Gly Thr
20 25 30
Tyr Tyr Lys Ala Phe Arg Ile Ile Asp Arg Ile Trp Ile Val Pro Glu
35 40 45
Arg Phe Thr Tyr Gly Phe Gln Pro Asp Gln Phe Asn Ala Ser Thr Gly
50 55 60
Val Phe Ser Lys Asp Val Tyr Glu Tyr Tyr Asp Pro Thr Tyr Leu Lys
65 70 75 80
Thr Asp Ala Glu Lys Asp Lys Phe Leu Lys Thr Met Ile Lys Leu Phe
85 90 95
Asn Arg Ile Asn Ser Lys Pro Ser Gly Gln Arg Leu Leu Asp Met Ile
100 105 110
Val Asp Ala Ile Pro Tyr Leu Gly Asn Ala Ser Thr Pro Pro Asp Lys
115 120 125
Phe Ala Ala Asn Val Ala Asn Val Ser Ile Asn Lys Lys Ile Ile Gln
130 135 140
Pro Gly Ala Glu Asp Gln Ile Lys Gly Leu Met Thr Asn Leu Ile Ile
145 150 155 160
Phe Gly Pro Gly Pro Val Leu Ser Asp Asn Phe Thr Asp Ser Met Ile
165 170 175
Met Asn Gly His Ser Pro Ile Ser Glu Gly Phe Gly Ala Arg Met Met
180 185 190
Ile Arg Phe Cys Pro Ser Cys Leu Asn Val Phe Asn Asn Val Gln Glu
195 200 205
Asn Lys Asp Thr Ser Ile Phe Ser Arg Arg Ala Tyr Phe Ala Asp Pro
210 215 220
Ala Leu Thr Leu Met His Glu Leu Ile His Val Leu His Gly Leu Tyr
225 230 235 240
Gly Ile Lys Ile Ser Asn Leu Pro Ile Thr Pro Asn Thr Lys Glu Phe
245 250 255
Phe Met Gln His Ser Asp Pro Val Gln Ala Glu Glu Leu Tyr Thr Phe
260 265 270
Gly Gly His Asp Pro Ser Val Ile Ser Pro Ser Thr Asp Met Asn Ile
275 280 285
Tyr Asn Lys Ala Leu Gln Asn Phe Gln Asp Ile Ala Asn Arg Leu Asn
290 295 300
Ile Val Ser Ser Ala Gln Gly Ser Gly Ile Asp Ile Ser Leu Tyr Lys
305 310 315 320
Gln Ile Tyr Lys Asn Lys Tyr Asp Phe Val Glu Asp Pro Asn Gly Lys
325 330 335
Tyr Ser Val Asp Lys Asp Lys Phe Asp Lys Leu Tyr Lys Ala Leu Met
340 345 350
Phe Gly Phe Thr Glu Thr Asn Leu Ala Gly Glu Tyr Gly Ile Lys Thr
355 360 365
Arg Tyr Ser Tyr Phe Ser Glu Tyr Leu Pro Pro Ile Lys Thr Glu Lys
370 375 380
Leu Leu Asp Asn Thr Ile Tyr Thr Gln Asn Glu Gly Phe Asn Ile Ala
385 390 395 400
Ser Lys Asn Leu Lys Thr Glu Phe Asn Gly Gln Asn Lys Ala Val Asn
405 410 415
Lys Glu Ala Tyr Glu Glu Ile Ser Leu Glu His Leu Val Ile Tyr Arg
420 425 430
Ile Ala Met Cys Lys Pro Val Met Tyr Lys Asn Thr Gly Lys Ser Glu
435 440 445
Gln Cys Ile Ile Val Asn Asn Glu Asp Leu Phe Phe Ile Ala Asn Lys
450 455 460
Asp Ser Phe Ser Lys Asp Leu Ala Lys Ala Glu Thr Ile Ala Tyr Asn
465 470 475 480
Thr Gln Asn Asn Thr Ile Glu Asn Asn Phe Ser Ile Asp Gln Leu Ile
485 490 495
Leu Asp Asn Asp Leu Ser Ser Gly Ile Asp Leu Pro Asn Glu Asn Thr
500 505 510
Glu Pro Phe Thr Asn Phe Asp Asp Ile Asp Ile Pro Val Tyr Ile Lys
515 520 525
Gln Ser Ala Leu Lys Lys Ile Phe Val Asp Gly Asp Ser Leu Phe Glu
530 535 540
Tyr Leu His Ala Gln Thr Phe Pro Ser Asn Ile Glu Asn Leu Gln Leu
545 550 555 560
Thr Asn Ser Leu Asn Asp Ala Leu Arg Asn Asn Asn Lys Val Tyr Thr
565 570 575
Phe Phe Ser Thr Asn Leu Val Glu Lys Ala Asn Thr Val Val Gly Ala
580 585 590
Ser Leu Phe Val Asn Trp Val Lys Gly Val Ile Asp Asp Phe Thr Ser
595 600 605
Glu Ser Thr Gln Lys Ser Thr Ile Asp Lys Val Ser Asp Val Ser Ile
610 615 620
Ile Ile Pro Tyr Ile Gly Pro Ala Leu Asn Val Gly Asn Glu Thr Ala
625 630 635 640
Lys Glu Asn Phe Lys Asn Ala Phe Glu Ile Gly Gly Ala Ala Ile Leu
645 650 655
Met Glu Phe Ile Pro Glu Leu Ile Val Pro Ile Val Gly Phe Phe Thr
660 665 670
Leu Glu Ser Tyr Val Gly Asn Lys Gly His Ile Ile Met Thr Ile Ser
675 680 685
Asn Ala Leu Lys Lys Arg Asp Gln Lys Trp Thr Asp Met Tyr Gly Leu
690 695 700
Ile Val Ser Gln Trp Leu Ser Thr Val Asn Thr Gln Phe Tyr Thr Ile
705 710 715 720
Lys Glu Arg Met Tyr Asn Ala Leu Asn Asn Gln Ser Gln Ala Ile Glu
725 730 735
Lys Ile Ile Glu Asp Gln Tyr Asn Arg Tyr Ser Glu Glu Asp Lys Met
740 745 750
Asn Ile Asn Ile Asp Phe Asn Asp Ile Asp Phe Lys Leu Asn Gln Ser
755 760 765
Ile Asn Leu Ala Ile Asn Asn Ile Asp Asp Phe Ile Asn Gln Cys Ser
770 775 780
Ile Ser Tyr Leu Met Asn Arg Met Ile Pro Leu Ala Val Lys Lys Leu
785 790 795 800
Lys Asp Phe Asp Asp Asn Leu Lys Arg Asp Leu Leu Glu Tyr Ile Asp
805 810 815
Thr Asn Glu Leu Tyr Leu Leu Asp Glu Val Asn Ile Leu Lys Ser Lys
820 825 830
Val Asn Arg His Leu Lys Asp Ser Ile Pro Phe Asp Leu Ser Leu Tyr
835 840 845
Thr Lys Asp Thr Ile Leu Ile Gln Val Phe Asn Asn Tyr Ile Ser Asn
850 855 860
Ile Ser Ser Asn Ala Ile Leu Ser Leu Ser Tyr Arg Gly Gly Arg Leu
865 870 875 880
Ile Asp Ser Ser Gly Tyr Gly Ala Thr Met Asn Val Gly Ser Asp Val
885 890 895
Ile Phe Asn Asp Ile Gly Asn Gly Gln Phe Lys Leu Asn Asn Ser Glu
900 905 910
Asn Ser Asn Ile Thr Ala His Gln Ser Lys Phe Val Val Tyr Asp Ser
915 920 925
Met Phe Asp Asn Phe Ser Ile Asn Phe Trp Val Arg Thr Pro Lys Tyr
930 935 940
Asn Asn Asn Asp Ile Gln Thr Tyr Leu Gln Asn Glu Tyr Thr Ile Ile
945 950 955 960
Ser Cys Ile Lys Asn Asp Ser Gly Trp Lys Val Ser Ile Lys Gly Asn
965 970 975
Arg Ile Ile Trp Thr Leu Ile Asp Val Asn Ala Lys Ser Lys Ser Ile
980 985 990
Phe Phe Glu Tyr Ser Ile Lys Asp Asn Ile Ser Asp Tyr Ile Asn Lys
995 1000 1005
Trp Phe Ser Ile Thr Ile Thr Asn Asp Arg Leu Gly Asn Ala Asn
1010 1015 1020
Ile Tyr Ile Asn Gly Ser Leu Lys Lys Ser Glu Lys Ile Leu Asn
1025 1030 1035
Leu Asp Arg Ile Asn Ser Ser Asn Asp Ile Asp Phe Lys Leu Ile
1040 1045 1050
Asn Cys Thr Asp Thr Thr Lys Phe Val Trp Ile Lys Asp Phe Asn
1055 1060 1065
Ile Phe Gly Arg Glu Leu Asn Ala Thr Glu Val Ser Ser Leu Tyr
1070 1075 1080
Trp Ile Gln Ser Ser Thr Asn Thr Leu Lys Asp Phe Trp Gly Asn
1085 1090 1095
Pro Leu Arg Tyr Asp Thr Gln Tyr Tyr Leu Phe Asn Gln Gly Met
1100 1105 1110
Gln Asn Ile Tyr Ile Lys Tyr Phe Ser Lys Ala Ser Met Gly Glu
1115 1120 1125
Thr Ala Pro Arg Thr Asn Phe Asn Asn Ala Ala Ile Asn Tyr Gln
1130 1135 1140
Asn Leu Tyr Leu Gly Leu Arg Phe Ile Ile Lys Lys Ala Ser Asn
1145 1150 1155
Ser Arg Asn Ile Asn Asn Asp Asn Ile Val Arg Glu Gly Asp Tyr
1160 1165 1170
Ile Tyr Leu Asn Ile Asp Asn Ile Ser Asp Glu Ser Tyr Arg Val
1175 1180 1185
Tyr Val Leu Val Asn Ser Lys Glu Ile Gln Thr Gln Leu Phe Leu
1190 1195 1200
Ala Pro Ile Asn Asp Asp Pro Thr Phe Tyr Asp Val Leu Gln Ile
1205 1210 1215
Lys Lys Tyr Tyr Glu Lys Thr Thr Tyr Asn Cys Gln Ile Leu Cys
1220 1225 1230
Glu Lys Asp Thr Lys Thr Phe Gly Leu Phe Gly Ile Gly Lys Phe
1235 1240 1245
Val Lys Asp Tyr Gly Tyr Val Trp Asp Thr Tyr Asp Asn Tyr Phe
1250 1255 1260
Cys Ile Ser Gln Trp Tyr Leu Arg Arg Ile Ser Glu Asn Ile Asn
1265 1270 1275
Lys Leu Arg Leu Gly Cys Asn Trp Gln Phe Ile Pro Val Asp Glu
1280 1285 1290
Gly Trp Thr Glu
1295
<210> 58
<211> 1315
<212> PRT
<213> Clostridium tetani
<400> 58
Met Pro Ile Thr Ile Asn Asn Phe Arg Tyr Ser Asp Pro Val Asn Asn
1 5 10 15
Asp Thr Ile Ile Met Met Glu Pro Pro Tyr Cys Lys Gly Leu Asp Ile
20 25 30
Tyr Tyr Lys Ala Phe Lys Ile Thr Asp Arg Ile Trp Ile Val Pro Glu
35 40 45
Arg Tyr Glu Phe Gly Thr Lys Pro Glu Asp Phe Asn Pro Pro Ser Ser
50 55 60
Leu Ile Glu Gly Ala Ser Glu Tyr Tyr Asp Pro Asn Tyr Leu Arg Thr
65 70 75 80
Asp Ser Asp Lys Asp Arg Phe Leu Gln Thr Met Val Lys Leu Phe Asn
85 90 95
Arg Ile Lys Asn Asn Val Ala Gly Glu Ala Leu Leu Asp Lys Ile Ile
100 105 110
Asn Ala Ile Pro Tyr Leu Gly Asn Ser Tyr Ser Leu Leu Asp Lys Phe
115 120 125
Asp Thr Asn Ser Asn Ser Val Ser Phe Asn Leu Leu Glu Gln Asp Pro
130 135 140
Ser Gly Ala Thr Thr Lys Ser Ala Met Leu Thr Asn Leu Ile Ile Phe
145 150 155 160
Gly Pro Gly Pro Val Leu Asn Lys Asn Glu Val Arg Gly Ile Val Leu
165 170 175
Arg Val Asp Asn Lys Asn Tyr Phe Pro Cys Arg Asp Gly Phe Gly Ser
180 185 190
Ile Met Gln Met Ala Phe Cys Pro Glu Tyr Val Pro Thr Phe Asp Asn
195 200 205
Val Ile Glu Asn Ile Thr Ser Leu Thr Ile Gly Lys Ser Lys Tyr Phe
210 215 220
Gln Asp Pro Ala Leu Leu Leu Met His Glu Leu Ile His Val Leu His
225 230 235 240
Gly Leu Tyr Gly Met Gln Val Ser Ser His Glu Ile Ile Pro Ser Lys
245 250 255
Gln Glu Ile Tyr Met Gln His Thr Tyr Pro Ile Ser Ala Glu Glu Leu
260 265 270
Phe Thr Phe Gly Gly Gln Asp Ala Asn Leu Ile Ser Ile Asp Ile Lys
275 280 285
Asn Asp Leu Tyr Glu Lys Thr Leu Asn Asp Tyr Lys Ala Ile Ala Asn
290 295 300
Lys Leu Ser Gln Val Thr Ser Cys Asn Asp Pro Asn Ile Asp Ile Asp
305 310 315 320
Ser Tyr Lys Gln Ile Tyr Gln Gln Lys Tyr Gln Phe Asp Lys Asp Ser
325 330 335
Asn Gly Gln Tyr Ile Val Asn Glu Asp Lys Phe Gln Ile Leu Tyr Asn
340 345 350
Ser Ile Met Tyr Gly Phe Thr Glu Ile Glu Leu Gly Lys Lys Phe Asn
355 360 365
Ile Lys Thr Arg Leu Ser Tyr Phe Ser Met Asn His Asp Pro Val Lys
370 375 380
Ile Pro Asn Leu Leu Asp Asp Thr Ile Tyr Asn Asp Thr Glu Gly Phe
385 390 395 400
Asn Ile Glu Ser Lys Asp Leu Lys Ser Glu Tyr Lys Gly Gln Asn Met
405 410 415
Arg Val Asn Thr Asn Ala Phe Arg Asn Val Asp Gly Ser Gly Leu Val
420 425 430
Ser Lys Leu Ile Gly Leu Cys Lys Lys Ile Ile Pro Pro Thr Asn Ile
435 440 445
Arg Glu Asn Leu Tyr Asn Arg Thr Ala Ser Leu Thr Asp Leu Gly Gly
450 455 460
Glu Leu Cys Ile Lys Ile Lys Asn Glu Asp Leu Thr Phe Ile Ala Glu
465 470 475 480
Lys Asn Ser Phe Ser Glu Glu Pro Phe Gln Asp Glu Ile Val Ser Tyr
485 490 495
Asn Thr Lys Asn Lys Pro Leu Asn Phe Asn Tyr Ser Leu Asp Lys Ile
500 505 510
Ile Val Asp Tyr Asn Leu Gln Ser Lys Ile Thr Leu Pro Asn Asp Arg
515 520 525
Thr Thr Pro Val Thr Lys Gly Ile Pro Tyr Ala Pro Glu Tyr Lys Ser
530 535 540
Asn Ala Ala Ser Thr Ile Glu Ile His Asn Ile Asp Asp Asn Thr Ile
545 550 555 560
Tyr Gln Tyr Leu Tyr Ala Gln Lys Ser Pro Thr Thr Leu Gln Arg Ile
565 570 575
Thr Met Thr Asn Ser Val Asp Asp Ala Leu Ile Asn Ser Thr Lys Ile
580 585 590
Tyr Ser Tyr Phe Pro Ser Val Ile Ser Lys Val Asn Gln Gly Ala Gln
595 600 605
Gly Ile Leu Phe Leu Gln Trp Val Arg Asp Ile Ile Asp Asp Phe Thr
610 615 620
Asn Glu Ser Ser Gln Lys Thr Thr Ile Asp Lys Ile Ser Asp Val Ser
625 630 635 640
Thr Ile Val Pro Tyr Ile Gly Pro Ala Leu Asn Ile Val Lys Gln Gly
645 650 655
Tyr Glu Gly Asn Phe Ile Gly Ala Leu Glu Thr Thr Gly Val Val Leu
660 665 670
Leu Leu Glu Tyr Ile Pro Glu Ile Thr Leu Pro Val Ile Ala Ala Leu
675 680 685
Ser Ile Ala Glu Ser Ser Thr Gln Lys Glu Lys Ile Ile Lys Thr Ile
690 695 700
Asp Asn Phe Leu Glu Lys Arg Tyr Glu Lys Trp Ile Glu Val Tyr Lys
705 710 715 720
Leu Val Lys Ala Lys Trp Leu Gly Thr Val Asn Thr Gln Phe Gln Lys
725 730 735
Arg Ser Tyr Gln Met Tyr Arg Ser Leu Glu Tyr Gln Val Asp Ala Ile
740 745 750
Lys Lys Ile Ile Asp Tyr Glu Tyr Lys Ile Tyr Ser Gly Pro Asp Lys
755 760 765
Glu Gln Ile Ala Asp Glu Ile Asn Asn Leu Lys Asn Lys Leu Glu Glu
770 775 780
Lys Ala Asn Lys Ala Met Ile Asn Ile Asn Ile Phe Met Arg Glu Ser
785 790 795 800
Ser Arg Ser Phe Leu Val Asn Gln Met Ile Asn Glu Ala Lys Lys Gln
805 810 815
Leu Leu Glu Phe Asp Thr Gln Ser Lys Asn Ile Leu Met Gln Tyr Ile
820 825 830
Lys Ala Asn Ser Lys Phe Ile Gly Ile Thr Glu Leu Lys Lys Leu Glu
835 840 845
Ser Lys Ile Asn Lys Val Phe Ser Thr Pro Ile Pro Phe Ser Tyr Ser
850 855 860
Lys Asn Leu Asp Cys Trp Val Asp Asn Glu Glu Asp Ile Asp Val Ile
865 870 875 880
Leu Lys Lys Ser Thr Ile Leu Asn Leu Asp Ile Asn Asn Asp Ile Ile
885 890 895
Ser Asp Ile Ser Gly Phe Asn Ser Ser Val Ile Thr Tyr Pro Asp Ala
900 905 910
Gln Leu Val Pro Gly Ile Asn Gly Lys Ala Ile His Leu Val Asn Asn
915 920 925
Glu Ser Ser Glu Val Ile Val His Lys Ala Met Asp Ile Glu Tyr Asn
930 935 940
Asp Met Phe Asn Asn Phe Thr Val Ser Phe Trp Leu Arg Val Pro Lys
945 950 955 960
Val Ser Ala Ser His Leu Glu Gln Tyr Gly Thr Asn Glu Tyr Ser Ile
965 970 975
Ile Ser Ser Met Lys Lys His Ser Leu Ser Ile Gly Ser Gly Trp Ser
980 985 990
Val Ser Leu Lys Gly Asn Asn Leu Ile Trp Thr Leu Lys Asp Ser Ala
995 1000 1005
Gly Glu Val Arg Gln Ile Thr Phe Arg Asp Leu Pro Asp Lys Phe
1010 1015 1020
Asn Ala Tyr Leu Ala Asn Lys Trp Val Phe Ile Thr Ile Thr Asn
1025 1030 1035
Asp Arg Leu Ser Ser Ala Asn Leu Tyr Ile Asn Gly Val Leu Met
1040 1045 1050
Gly Ser Ala Glu Ile Thr Gly Leu Gly Ala Ile Arg Glu Asp Asn
1055 1060 1065
Asn Ile Thr Leu Lys Leu Asp Arg Cys Asn Asn Asn Asn Gln Tyr
1070 1075 1080
Val Ser Ile Asp Lys Phe Arg Ile Phe Cys Lys Ala Leu Asn Pro
1085 1090 1095
Lys Glu Ile Glu Lys Leu Tyr Thr Ser Tyr Leu Ser Ile Thr Phe
1100 1105 1110
Leu Arg Asp Phe Trp Gly Asn Pro Leu Arg Tyr Asp Thr Glu Tyr
1115 1120 1125
Tyr Leu Ile Pro Val Ala Ser Ser Ser Lys Asp Val Gln Leu Lys
1130 1135 1140
Asn Ile Thr Asp Tyr Met Tyr Leu Thr Asn Ala Pro Ser Tyr Thr
1145 1150 1155
Asn Gly Lys Leu Asn Ile Tyr Tyr Arg Arg Leu Tyr Asn Gly Leu
1160 1165 1170
Lys Phe Ile Ile Lys Arg Tyr Thr Pro Asn Asn Glu Ile Asp Ser
1175 1180 1185
Phe Val Lys Ser Gly Asp Phe Ile Lys Leu Tyr Val Ser Tyr Asn
1190 1195 1200
Asn Asn Glu His Ile Val Gly Tyr Pro Lys Asp Gly Asn Ala Phe
1205 1210 1215
Asn Asn Leu Asp Arg Ile Leu Arg Val Gly Tyr Asn Ala Pro Gly
1220 1225 1230
Ile Pro Leu Tyr Lys Lys Met Glu Ala Val Lys Leu Arg Asp Leu
1235 1240 1245
Lys Thr Tyr Ser Val Gln Leu Lys Leu Tyr Asp Asp Lys Asn Ala
1250 1255 1260
Ser Leu Gly Leu Val Gly Thr His Asn Gly Gln Ile Gly Asn Asp
1265 1270 1275
Pro Asn Arg Asp Ile Leu Ile Ala Ser Asn Trp Tyr Phe Asn His
1280 1285 1290
Leu Lys Asp Lys Ile Leu Gly Cys Asp Trp Tyr Phe Val Pro Thr
1295 1300 1305
Asp Glu Gly Trp Thr Asn Asp
1310 1315
<210> 59
<211> 1306
<212> PRT
<213> 肉毒梭菌
<400> 59
Met Lys Leu Glu Ile Asn Lys Phe Asn Tyr Asn Asp Pro Ile Asp Gly
1 5 10 15
Ile Asn Val Ile Thr Met Arg Pro Pro Arg His Ser Asp Lys Ile Asn
20 25 30
Lys Gly Lys Gly Pro Phe Lys Ala Phe Gln Val Ile Lys Asn Ile Trp
35 40 45
Ile Val Pro Glu Arg Tyr Asn Phe Thr Asn Asn Thr Asn Asp Leu Asn
50 55 60
Ile Pro Ser Glu Pro Ile Met Glu Ala Asp Ala Ile Tyr Asn Pro Asn
65 70 75 80
Tyr Leu Asn Thr Pro Ser Glu Lys Asp Glu Phe Leu Gln Gly Val Ile
85 90 95
Lys Val Leu Glu Arg Ile Lys Ser Lys Pro Glu Gly Glu Lys Leu Leu
100 105 110
Glu Leu Ile Ser Ser Ser Ile Pro Leu Pro Leu Val Ser Asn Gly Ala
115 120 125
Leu Thr Leu Ser Asp Asn Glu Thr Ile Ala Tyr Gln Glu Asn Asn Asn
130 135 140
Ile Val Ser Asn Leu Gln Ala Asn Leu Val Ile Tyr Gly Pro Gly Pro
145 150 155 160
Asp Ile Ala Asn Asn Ala Thr Tyr Gly Leu Tyr Ser Thr Pro Ile Ser
165 170 175
Asn Gly Glu Gly Thr Leu Ser Glu Val Ser Phe Ser Pro Phe Tyr Leu
180 185 190
Lys Pro Phe Asp Glu Ser Tyr Gly Asn Tyr Arg Ser Leu Val Asn Ile
195 200 205
Val Asn Lys Phe Val Lys Arg Glu Phe Ala Pro Asp Pro Ala Ser Thr
210 215 220
Leu Met His Glu Leu Val His Val Thr His Asn Leu Tyr Gly Ile Ser
225 230 235 240
Asn Arg Asn Phe Tyr Tyr Asn Phe Asp Thr Gly Lys Ile Glu Thr Ser
245 250 255
Arg Gln Gln Asn Ser Leu Ile Phe Glu Glu Leu Leu Thr Phe Gly Gly
260 265 270
Ile Asp Ser Lys Ala Ile Ser Ser Leu Ile Ile Lys Lys Ile Ile Glu
275 280 285
Thr Ala Lys Asn Asn Tyr Thr Thr Leu Ile Ser Glu Arg Leu Asn Thr
290 295 300
Val Thr Val Glu Asn Asp Leu Leu Lys Tyr Ile Lys Asn Lys Ile Pro
305 310 315 320
Val Gln Gly Arg Leu Gly Asn Phe Lys Leu Asp Thr Ala Glu Phe Glu
325 330 335
Lys Lys Leu Asn Thr Ile Leu Phe Val Leu Asn Glu Ser Asn Leu Ala
340 345 350
Gln Arg Phe Ser Ile Leu Val Arg Lys His Tyr Leu Lys Glu Arg Pro
355 360 365
Ile Asp Pro Ile Tyr Val Asn Ile Leu Asp Asp Asn Ser Tyr Ser Thr
370 375 380
Leu Glu Gly Phe Asn Ile Ser Ser Gln Gly Ser Asn Asp Phe Gln Gly
385 390 395 400
Gln Leu Leu Glu Ser Ser Tyr Phe Glu Lys Ile Glu Ser Asn Ala Leu
405 410 415
Arg Ala Phe Ile Lys Ile Cys Pro Arg Asn Gly Leu Leu Tyr Asn Ala
420 425 430
Ile Tyr Arg Asn Ser Lys Asn Tyr Leu Asn Asn Ile Asp Leu Glu Asp
435 440 445
Lys Lys Thr Thr Ser Lys Thr Asn Val Ser Tyr Pro Cys Ser Leu Leu
450 455 460
Asn Gly Cys Ile Glu Val Glu Asn Lys Asp Leu Phe Leu Ile Ser Asn
465 470 475 480
Lys Asp Ser Leu Asn Asp Ile Asn Leu Ser Glu Glu Lys Ile Lys Pro
485 490 495
Glu Thr Thr Val Phe Phe Lys Asp Lys Leu Pro Pro Gln Asp Ile Thr
500 505 510
Leu Ser Asn Tyr Asp Phe Thr Glu Ala Asn Ser Ile Pro Ser Ile Ser
515 520 525
Gln Gln Asn Ile Leu Glu Arg Asn Glu Glu Leu Tyr Glu Pro Ile Arg
530 535 540
Asn Ser Leu Phe Glu Ile Lys Thr Ile Tyr Val Asp Lys Leu Thr Thr
545 550 555 560
Phe His Phe Leu Glu Ala Gln Asn Ile Asp Glu Ser Ile Asp Ser Ser
565 570 575
Lys Ile Arg Val Glu Leu Thr Asp Ser Val Asp Glu Ala Leu Ser Asn
580 585 590
Pro Asn Lys Val Tyr Ser Pro Phe Lys Asn Met Ser Asn Thr Ile Asn
595 600 605
Ser Ile Glu Thr Gly Ile Thr Ser Thr Tyr Ile Phe Tyr Gln Trp Leu
610 615 620
Arg Ser Ile Val Lys Asp Phe Ser Asp Glu Thr Gly Lys Ile Asp Val
625 630 635 640
Ile Asp Lys Ser Ser Asp Thr Leu Ala Ile Val Pro Tyr Ile Gly Pro
645 650 655
Leu Leu Asn Ile Gly Asn Asp Ile Arg His Gly Asp Phe Val Gly Ala
660 665 670
Ile Glu Leu Ala Gly Ile Thr Ala Leu Leu Glu Tyr Val Pro Glu Phe
675 680 685
Thr Ile Pro Ile Leu Val Gly Leu Glu Val Ile Gly Gly Glu Leu Ala
690 695 700
Arg Glu Gln Val Glu Ala Ile Val Asn Asn Ala Leu Asp Lys Arg Asp
705 710 715 720
Gln Lys Trp Ala Glu Val Tyr Asn Ile Thr Lys Ala Gln Trp Trp Gly
725 730 735
Thr Ile His Leu Gln Ile Asn Thr Arg Leu Ala His Thr Tyr Lys Ala
740 745 750
Leu Ser Arg Gln Ala Asn Ala Ile Lys Met Asn Met Glu Phe Gln Leu
755 760 765
Ala Asn Tyr Lys Gly Asn Ile Asp Asp Lys Ala Lys Ile Lys Asn Ala
770 775 780
Ile Ser Glu Thr Glu Ile Leu Leu Asn Lys Ser Val Glu Gln Ala Met
785 790 795 800
Lys Asn Thr Glu Lys Phe Met Ile Lys Leu Ser Asn Ser Tyr Leu Thr
805 810 815
Lys Glu Met Ile Pro Lys Val Gln Asp Asn Leu Lys Asn Phe Asp Leu
820 825 830
Glu Thr Lys Lys Thr Leu Asp Lys Phe Ile Lys Glu Lys Glu Asp Ile
835 840 845
Leu Gly Thr Asn Leu Ser Ser Ser Leu Arg Arg Lys Val Ser Ile Arg
850 855 860
Leu Asn Lys Asn Ile Ala Phe Asp Ile Asn Asp Ile Pro Phe Ser Glu
865 870 875 880
Phe Asp Asp Leu Ile Asn Gln Tyr Lys Asn Glu Ile Glu Asp Tyr Glu
885 890 895
Val Leu Asn Leu Gly Ala Glu Asp Gly Lys Ile Lys Asp Leu Ser Gly
900 905 910
Thr Thr Ser Asp Ile Asn Ile Gly Ser Asp Ile Glu Leu Ala Asp Gly
915 920 925
Arg Glu Asn Lys Ala Ile Lys Ile Lys Gly Ser Glu Asn Ser Thr Ile
930 935 940
Lys Ile Ala Met Asn Lys Tyr Leu Arg Phe Ser Ala Thr Asp Asn Phe
945 950 955 960
Ser Ile Ser Phe Trp Ile Lys His Pro Lys Pro Thr Asn Leu Leu Asn
965 970 975
Asn Gly Ile Glu Tyr Thr Leu Val Glu Asn Phe Asn Gln Arg Gly Trp
980 985 990
Lys Ile Ser Ile Gln Asp Ser Lys Leu Ile Trp Tyr Leu Arg Asp His
995 1000 1005
Asn Asn Ser Ile Lys Ile Val Thr Pro Asp Tyr Ile Ala Phe Asn
1010 1015 1020
Gly Trp Asn Leu Ile Thr Ile Thr Asn Asn Arg Ser Lys Gly Ser
1025 1030 1035
Ile Val Tyr Val Asn Gly Ser Lys Ile Glu Glu Lys Asp Ile Ser
1040 1045 1050
Ser Ile Trp Asn Thr Glu Val Asp Asp Pro Ile Ile Phe Arg Leu
1055 1060 1065
Lys Asn Asn Arg Asp Thr Gln Ala Phe Thr Leu Leu Asp Gln Phe
1070 1075 1080
Ser Ile Tyr Arg Lys Glu Leu Asn Gln Asn Glu Val Val Lys Leu
1085 1090 1095
Tyr Asn Tyr Tyr Phe Asn Ser Asn Tyr Ile Arg Asp Ile Trp Gly
1100 1105 1110
Asn Pro Leu Gln Tyr Asn Lys Lys Tyr Tyr Leu Gln Thr Gln Asp
1115 1120 1125
Lys Pro Gly Lys Gly Leu Ile Arg Glu Tyr Trp Ser Ser Phe Gly
1130 1135 1140
Tyr Asp Tyr Val Ile Leu Ser Asp Ser Lys Thr Ile Thr Phe Pro
1145 1150 1155
Asn Asn Ile Arg Tyr Gly Ala Leu Tyr Asn Gly Ser Lys Val Leu
1160 1165 1170
Ile Lys Asn Ser Lys Lys Leu Asp Gly Leu Val Arg Asn Lys Asp
1175 1180 1185
Phe Ile Gln Leu Glu Ile Asp Gly Tyr Asn Met Gly Ile Ser Ala
1190 1195 1200
Asp Arg Phe Asn Glu Asp Thr Asn Tyr Ile Gly Thr Thr Tyr Gly
1205 1210 1215
Thr Thr His Asp Leu Thr Thr Asp Phe Glu Ile Ile Gln Arg Gln
1220 1225 1230
Glu Lys Tyr Arg Asn Tyr Cys Gln Leu Lys Thr Pro Tyr Asn Ile
1235 1240 1245
Phe His Lys Ser Gly Leu Met Ser Thr Glu Thr Ser Lys Pro Thr
1250 1255 1260
Phe His Asp Tyr Arg Asp Trp Val Tyr Ser Ser Ala Trp Tyr Phe
1265 1270 1275
Gln Asn Tyr Glu Asn Leu Asn Leu Arg Lys His Thr Lys Thr Asn
1280 1285 1290
Trp Tyr Phe Ile Pro Lys Asp Glu Gly Trp Asp Glu Asp
1295 1300 1305
<210> 60
<211> 1296
<212> PRT
<213> 肉毒梭菌
<400> 60
Met Pro Phe Val Asn Lys Gln Phe Asn Tyr Lys Asp Pro Val Asn Gly
1 5 10 15
Val Asp Ile Ala Tyr Ile Lys Ile Pro Asn Ala Gly Gln Met Gln Pro
20 25 30
Val Lys Ala Phe Lys Ile His Asn Lys Ile Trp Val Ile Pro Glu Arg
35 40 45
Asp Thr Phe Thr Asn Pro Glu Glu Gly Asp Leu Asn Pro Pro Pro Glu
50 55 60
Ala Lys Gln Val Pro Val Ser Tyr Tyr Asp Ser Thr Tyr Leu Ser Thr
65 70 75 80
Asp Asn Glu Lys Asp Asn Tyr Leu Lys Gly Val Thr Lys Leu Phe Glu
85 90 95
Arg Ile Tyr Ser Thr Asp Leu Gly Arg Met Leu Leu Thr Ser Ile Val
100 105 110
Arg Gly Ile Pro Phe Trp Gly Gly Ser Thr Ile Asp Thr Glu Leu Lys
115 120 125
Val Ile Asp Thr Asn Cys Ile Asn Val Ile Gln Pro Asp Gly Ser Tyr
130 135 140
Arg Ser Glu Glu Leu Asn Leu Val Ile Ile Gly Pro Ser Ala Asp Ile
145 150 155 160
Ile Gln Phe Glu Cys Lys Ser Phe Gly His Glu Val Leu Asn Leu Thr
165 170 175
Arg Asn Gly Tyr Gly Ser Thr Gln Tyr Ile Arg Phe Ser Pro Asp Phe
180 185 190
Thr Phe Gly Phe Glu Glu Ser Leu Glu Val Asp Thr Asn Pro Leu Leu
195 200 205
Gly Ala Gly Lys Phe Ala Thr Asp Pro Ala Val Thr Leu Ala His Glu
210 215 220
Leu Ile His Ala Gly His Arg Leu Tyr Gly Ile Ala Ile Asn Pro Asn
225 230 235 240
Arg Val Phe Lys Val Asn Thr Asn Ala Tyr Tyr Glu Met Ser Gly Leu
245 250 255
Glu Val Ser Phe Glu Glu Leu Arg Thr Phe Gly Gly His Asp Ala Lys
260 265 270
Phe Ile Asp Ser Leu Gln Glu Asn Glu Phe Arg Leu Tyr Tyr Tyr Asn
275 280 285
Lys Phe Lys Asp Ile Ala Ser Thr Leu Asn Lys Ala Lys Ser Ile Val
290 295 300
Gly Thr Thr Ala Ser Leu Gln Tyr Met Lys Asn Val Phe Lys Glu Lys
305 310 315 320
Tyr Leu Leu Ser Glu Asp Thr Ser Gly Lys Phe Ser Val Asp Lys Leu
325 330 335
Lys Phe Asp Lys Leu Tyr Lys Met Leu Thr Glu Ile Tyr Thr Glu Asp
340 345 350
Asn Phe Val Lys Phe Phe Lys Val Leu Asn Arg Lys Thr Tyr Leu Asn
355 360 365
Phe Asp Lys Ala Val Phe Lys Ile Asn Ile Val Pro Lys Val Asn Tyr
370 375 380
Thr Ile Tyr Asp Gly Phe Asn Leu Arg Asn Thr Asn Leu Ala Ala Asn
385 390 395 400
Phe Asn Gly Gln Asn Thr Glu Ile Asn Asn Met Asn Phe Thr Lys Leu
405 410 415
Lys Asn Phe Thr Gly Leu Phe Glu Phe Tyr Lys Leu Leu Cys Val Arg
420 425 430
Gly Ile Ile Thr Ser Lys Thr Lys Ser Leu Asp Lys Gly Tyr Asn Lys
435 440 445
Ala Leu Asn Asp Leu Cys Ile Lys Val Asn Asn Trp Asp Leu Phe Phe
450 455 460
Ser Pro Ser Glu Asp Asn Phe Thr Asn Asp Leu Asn Lys Gly Glu Glu
465 470 475 480
Ile Thr Ser Asp Thr Asn Ile Glu Ala Ala Glu Glu Asn Ile Ser Leu
485 490 495
Asp Leu Ile Gln Gln Tyr Tyr Leu Thr Phe Asn Phe Asp Asn Glu Pro
500 505 510
Glu Asn Ile Ser Ile Glu Asn Leu Ser Ser Asp Ile Ile Gly Gln Leu
515 520 525
Glu Leu Met Pro Asn Ile Glu Arg Phe Pro Asn Gly Lys Lys Tyr Glu
530 535 540
Leu Asp Lys Tyr Thr Met Phe His Tyr Leu Arg Ala Gln Glu Phe Glu
545 550 555 560
His Gly Lys Ser Arg Ile Ala Leu Thr Asn Ser Val Asn Glu Ala Leu
565 570 575
Leu Asn Pro Ser Arg Val Tyr Thr Phe Phe Ser Ser Asp Tyr Val Lys
580 585 590
Lys Val Asn Lys Ala Thr Glu Ala Ala Met Phe Leu Gly Trp Val Glu
595 600 605
Gln Leu Val Tyr Asp Phe Thr Asp Glu Thr Ser Glu Val Ser Thr Thr
610 615 620
Asp Lys Ile Ala Asp Ile Thr Ile Ile Ile Pro Tyr Ile Gly Pro Ala
625 630 635 640
Leu Asn Ile Gly Asn Met Leu Tyr Lys Asp Asp Phe Val Gly Ala Leu
645 650 655
Ile Phe Ser Gly Ala Val Ile Leu Leu Glu Phe Ile Pro Glu Ile Ala
660 665 670
Ile Pro Val Leu Gly Thr Phe Ala Leu Val Ser Tyr Ile Ala Asn Lys
675 680 685
Val Leu Thr Val Gln Thr Ile Asp Asn Ala Leu Ser Lys Arg Asn Glu
690 695 700
Lys Trp Asp Glu Val Tyr Lys Tyr Ile Val Thr Asn Trp Leu Ala Lys
705 710 715 720
Val Asn Thr Gln Ile Asp Leu Ile Arg Lys Lys Met Lys Glu Ala Leu
725 730 735
Glu Asn Gln Ala Glu Ala Thr Lys Ala Ile Ile Asn Tyr Gln Tyr Asn
740 745 750
Gln Tyr Thr Glu Glu Glu Lys Asn Asn Ile Asn Phe Asn Ile Asp Asp
755 760 765
Leu Ser Ser Lys Leu Asn Glu Ser Ile Asn Lys Ala Met Ile Asn Ile
770 775 780
Asn Lys Phe Leu Asn Gln Cys Ser Val Ser Tyr Leu Met Asn Ser Met
785 790 795 800
Ile Pro Tyr Gly Val Lys Arg Leu Glu Asp Phe Asp Ala Ser Leu Lys
805 810 815
Asp Ala Leu Leu Lys Tyr Ile Tyr Asp Asn Arg Gly Thr Leu Ile Gly
820 825 830
Gln Val Asp Arg Leu Lys Asp Lys Val Asn Asn Thr Leu Ser Thr Asp
835 840 845
Ile Pro Phe Gln Leu Ser Lys Tyr Val Asp Asn Gln Arg Leu Leu Ser
850 855 860
Thr Phe Thr Glu Tyr Ile Lys Asn Ile Ile Asn Thr Ser Ile Leu Asn
865 870 875 880
Leu Arg Tyr Glu Ser Asn His Leu Ile Asp Leu Ser Arg Tyr Ala Ser
885 890 895
Lys Ile Asn Ile Gly Ser Lys Val Asn Phe Asp Pro Ile Asp Lys Asn
900 905 910
Gln Ile Gln Leu Phe Asn Leu Glu Ser Ser Lys Ile Glu Val Ile Leu
915 920 925
Lys Asn Ala Ile Val Tyr Asn Ser Met Tyr Glu Asn Phe Ser Thr Ser
930 935 940
Phe Trp Ile Arg Ile Pro Lys Tyr Phe Asn Ser Ile Ser Leu Asn Asn
945 950 955 960
Glu Tyr Thr Ile Ile Asn Cys Met Glu Asn Asn Ser Gly Trp Lys Val
965 970 975
Ser Leu Asn Tyr Gly Glu Ile Ile Trp Thr Leu Gln Asp Thr Gln Glu
980 985 990
Ile Lys Gln Arg Val Val Phe Lys Tyr Ser Gln Met Ile Asn Ile Ser
995 1000 1005
Asp Tyr Ile Asn Arg Trp Ile Phe Val Thr Ile Thr Asn Asn Arg
1010 1015 1020
Leu Asn Asn Ser Lys Ile Tyr Ile Asn Gly Arg Leu Ile Asp Gln
1025 1030 1035
Lys Pro Ile Ser Asn Leu Gly Asn Ile His Ala Ser Asn Asn Ile
1040 1045 1050
Met Phe Lys Leu Asp Gly Cys Arg Asp Thr His Arg Tyr Ile Trp
1055 1060 1065
Ile Lys Tyr Phe Asn Leu Phe Asp Lys Glu Leu Asn Glu Lys Glu
1070 1075 1080
Ile Lys Asp Leu Tyr Asp Asn Gln Ser Asn Ser Gly Ile Leu Lys
1085 1090 1095
Asp Phe Trp Gly Asp Tyr Leu Gln Tyr Asp Lys Pro Tyr Tyr Met
1100 1105 1110
Leu Asn Leu Tyr Asp Pro Asn Lys Tyr Val Asp Val Asn Asn Val
1115 1120 1125
Gly Ile Arg Gly Tyr Met Tyr Leu Lys Gly Pro Arg Gly Ser Val
1130 1135 1140
Met Thr Thr Asn Ile Tyr Leu Asn Ser Ser Leu Tyr Arg Gly Thr
1145 1150 1155
Lys Phe Ile Ile Lys Lys Tyr Ala Ser Gly Asn Lys Asp Asn Ile
1160 1165 1170
Val Arg Asn Asn Asp Arg Val Tyr Ile Asn Val Val Val Lys Asn
1175 1180 1185
Lys Glu Tyr Arg Leu Ala Thr Asn Ala Ser Gln Ala Gly Val Glu
1190 1195 1200
Lys Ile Leu Ser Ala Leu Glu Ile Pro Asp Val Gly Asn Leu Ser
1205 1210 1215
Gln Val Val Val Met Lys Ser Lys Asn Asp Gln Gly Ile Thr Asn
1220 1225 1230
Lys Cys Lys Met Asn Leu Gln Asp Asn Asn Gly Asn Asp Ile Gly
1235 1240 1245
Phe Ile Gly Phe His Gln Phe Asn Asn Ile Ala Lys Leu Val Ala
1250 1255 1260
Ser Asn Trp Tyr Asn Arg Gln Ile Glu Arg Ser Ser Arg Thr Leu
1265 1270 1275
Gly Cys Ser Trp Glu Phe Ile Pro Val Asp Asp Gly Trp Gly Glu
1280 1285 1290
Arg Pro Leu
1295
<210> 61
<211> 1304
<212> PRT
<213> 人工序列
<220>
<223> mrBoNT/AB(0)的多肽序列
<400> 61
Met Pro Phe Val Asn Lys Gln Phe Asn Tyr Lys Asp Pro Val Asn Gly
1 5 10 15
Val Asp Ile Ala Tyr Ile Lys Ile Pro Asn Ala Gly Gln Met Gln Pro
20 25 30
Val Lys Ala Phe Lys Ile His Asn Lys Ile Trp Val Ile Pro Glu Arg
35 40 45
Asp Thr Phe Thr Asn Pro Glu Glu Gly Asp Leu Asn Pro Pro Pro Glu
50 55 60
Ala Lys Gln Val Pro Val Ser Tyr Tyr Asp Ser Thr Tyr Leu Ser Thr
65 70 75 80
Asp Asn Glu Lys Asp Asn Tyr Leu Lys Gly Val Thr Lys Leu Phe Glu
85 90 95
Arg Ile Tyr Ser Thr Asp Leu Gly Arg Met Leu Leu Thr Ser Ile Val
100 105 110
Arg Gly Ile Pro Phe Trp Gly Gly Ser Thr Ile Asp Thr Glu Leu Lys
115 120 125
Val Ile Asp Thr Asn Cys Ile Asn Val Ile Gln Pro Asp Gly Ser Tyr
130 135 140
Arg Ser Glu Glu Leu Asn Leu Val Ile Ile Gly Pro Ser Ala Asp Ile
145 150 155 160
Ile Gln Phe Glu Cys Lys Ser Phe Gly His Glu Val Leu Asn Leu Thr
165 170 175
Arg Asn Gly Tyr Gly Ser Thr Gln Tyr Ile Arg Phe Ser Pro Asp Phe
180 185 190
Thr Phe Gly Phe Glu Glu Ser Leu Glu Val Asp Thr Asn Pro Leu Leu
195 200 205
Gly Ala Gly Lys Phe Ala Thr Asp Pro Ala Val Thr Leu Ala His Gln
210 215 220
Leu Ile Tyr Ala Gly His Arg Leu Tyr Gly Ile Ala Ile Asn Pro Asn
225 230 235 240
Arg Val Phe Lys Val Asn Thr Asn Ala Tyr Tyr Glu Met Ser Gly Leu
245 250 255
Glu Val Ser Phe Glu Glu Leu Arg Thr Phe Gly Gly His Asp Ala Lys
260 265 270
Phe Ile Asp Ser Leu Gln Glu Asn Glu Phe Arg Leu Tyr Tyr Tyr Asn
275 280 285
Lys Phe Lys Asp Ile Ala Ser Thr Leu Asn Lys Ala Lys Ser Ile Val
290 295 300
Gly Thr Thr Ala Ser Leu Gln Tyr Met Lys Asn Val Phe Lys Glu Lys
305 310 315 320
Tyr Leu Leu Ser Glu Asp Thr Ser Gly Lys Phe Ser Val Asp Lys Leu
325 330 335
Lys Phe Asp Lys Leu Tyr Lys Met Leu Thr Glu Ile Tyr Thr Glu Asp
340 345 350
Asn Phe Val Lys Phe Phe Lys Val Leu Asn Arg Lys Thr Tyr Leu Asn
355 360 365
Phe Asp Lys Ala Val Phe Lys Ile Asn Ile Val Pro Lys Val Asn Tyr
370 375 380
Thr Ile Tyr Asp Gly Phe Asn Leu Arg Asn Thr Asn Leu Ala Ala Asn
385 390 395 400
Phe Asn Gly Gln Asn Thr Glu Ile Asn Asn Met Asn Phe Thr Lys Leu
405 410 415
Lys Asn Phe Thr Gly Leu Phe Glu Phe Tyr Lys Leu Leu Cys Val Arg
420 425 430
Gly Ile Ile Thr Ser Lys Thr Lys Ser Leu Asp Lys Gly Tyr Asn Lys
435 440 445
Ala Leu Asn Asp Leu Cys Ile Lys Val Asn Asn Trp Asp Leu Phe Phe
450 455 460
Ser Pro Ser Glu Asp Asn Phe Thr Asn Asp Leu Asn Lys Gly Glu Glu
465 470 475 480
Ile Thr Ser Asp Thr Asn Ile Glu Ala Ala Glu Glu Asn Ile Ser Leu
485 490 495
Asp Leu Ile Gln Gln Tyr Tyr Leu Thr Phe Asn Phe Asp Asn Glu Pro
500 505 510
Glu Asn Ile Ser Ile Glu Asn Leu Ser Ser Asp Ile Ile Gly Gln Leu
515 520 525
Glu Leu Met Pro Asn Ile Glu Arg Phe Pro Asn Gly Lys Lys Tyr Glu
530 535 540
Leu Asp Lys Tyr Thr Met Phe His Tyr Leu Arg Ala Gln Glu Phe Glu
545 550 555 560
His Gly Lys Ser Arg Ile Ala Leu Thr Asn Ser Val Asn Glu Ala Leu
565 570 575
Leu Asn Pro Ser Arg Val Tyr Thr Phe Phe Ser Ser Asp Tyr Val Lys
580 585 590
Lys Val Asn Lys Ala Thr Glu Ala Ala Met Phe Leu Gly Trp Val Glu
595 600 605
Gln Leu Val Tyr Asp Phe Thr Asp Glu Thr Ser Glu Val Ser Thr Thr
610 615 620
Asp Lys Ile Ala Asp Ile Thr Ile Ile Ile Pro Tyr Ile Gly Pro Ala
625 630 635 640
Leu Asn Ile Gly Asn Met Leu Tyr Lys Asp Asp Phe Val Gly Ala Leu
645 650 655
Ile Phe Ser Gly Ala Val Ile Leu Leu Glu Phe Ile Pro Glu Ile Ala
660 665 670
Ile Pro Val Leu Gly Thr Phe Ala Leu Val Ser Tyr Ile Ala Asn Lys
675 680 685
Val Leu Thr Val Gln Thr Ile Asp Asn Ala Leu Ser Lys Arg Asn Glu
690 695 700
Lys Trp Asp Glu Val Tyr Lys Tyr Ile Val Thr Asn Trp Leu Ala Lys
705 710 715 720
Val Asn Thr Gln Ile Asp Leu Ile Arg Lys Lys Met Lys Glu Ala Leu
725 730 735
Glu Asn Gln Ala Glu Ala Thr Lys Ala Ile Ile Asn Tyr Gln Tyr Asn
740 745 750
Gln Tyr Thr Glu Glu Glu Lys Asn Asn Ile Asn Phe Asn Ile Asp Asp
755 760 765
Leu Ser Ser Lys Leu Asn Glu Ser Ile Asn Lys Ala Met Ile Asn Ile
770 775 780
Asn Lys Phe Leu Asn Gln Cys Ser Val Ser Tyr Leu Met Asn Ser Met
785 790 795 800
Ile Pro Tyr Gly Val Lys Arg Leu Glu Asp Phe Asp Ala Ser Leu Lys
805 810 815
Asp Ala Leu Leu Lys Tyr Ile Tyr Asp Asn Arg Gly Thr Leu Ile Gly
820 825 830
Gln Val Asp Arg Leu Lys Asp Lys Val Asn Asn Thr Leu Ser Thr Asp
835 840 845
Ile Pro Phe Gln Leu Ser Lys Tyr Val Asp Asn Gln Arg Leu Leu Ser
850 855 860
Thr Phe Thr Glu Tyr Ile Lys Asn Ile Leu Asn Asn Ile Ile Leu Asn
865 870 875 880
Leu Arg Tyr Lys Asp Asn Asn Leu Ile Asp Leu Ser Gly Tyr Gly Ala
885 890 895
Lys Val Glu Val Tyr Asp Gly Val Glu Leu Asn Asp Lys Asn Gln Phe
900 905 910
Lys Leu Thr Ser Ser Ala Asn Ser Lys Ile Arg Val Thr Gln Asn Gln
915 920 925
Asn Ile Ile Phe Asn Ser Val Phe Leu Asp Phe Ser Val Ser Phe Trp
930 935 940
Ile Arg Ile Pro Lys Tyr Lys Asn Asp Gly Ile Gln Asn Tyr Ile His
945 950 955 960
Asn Glu Tyr Thr Ile Ile Asn Cys Met Lys Asn Asn Ser Gly Trp Lys
965 970 975
Ile Ser Ile Arg Gly Asn Arg Ile Ile Trp Thr Leu Ile Asp Ile Asn
980 985 990
Gly Lys Thr Lys Ser Val Phe Phe Glu Tyr Asn Ile Arg Glu Asp Ile
995 1000 1005
Ser Glu Tyr Ile Asn Arg Trp Phe Phe Val Thr Ile Thr Asn Asn
1010 1015 1020
Leu Asn Asn Ala Lys Ile Tyr Ile Asn Gly Lys Leu Glu Ser Asn
1025 1030 1035
Thr Asp Ile Lys Asp Ile Arg Glu Val Ile Ala Asn Gly Glu Ile
1040 1045 1050
Ile Phe Lys Leu Asp Gly Asp Ile Asp Arg Thr Gln Phe Ile Trp
1055 1060 1065
Met Lys Tyr Phe Ser Ile Phe Asn Thr Glu Leu Ser Gln Ser Asn
1070 1075 1080
Ile Glu Glu Arg Tyr Lys Ile Gln Ser Tyr Ser Glu Tyr Leu Lys
1085 1090 1095
Asp Phe Trp Gly Asn Pro Leu Met Tyr Asn Lys Glu Tyr Tyr Met
1100 1105 1110
Phe Asn Ala Gly Asn Lys Asn Ser Tyr Ile Lys Leu Lys Lys Asp
1115 1120 1125
Ser Pro Val Gly Glu Ile Leu Thr Arg Ser Lys Tyr Asn Gln Asn
1130 1135 1140
Ser Lys Tyr Ile Asn Tyr Arg Asp Leu Tyr Ile Gly Glu Lys Phe
1145 1150 1155
Ile Ile Arg Arg Lys Ser Asn Ser Gln Ser Ile Asn Asp Asp Ile
1160 1165 1170
Val Arg Lys Glu Asp Tyr Ile Tyr Leu Asp Phe Phe Asn Leu Asn
1175 1180 1185
Gln Glu Trp Arg Val Tyr Thr Tyr Lys Tyr Phe Lys Lys Glu Glu
1190 1195 1200
Met Lys Leu Phe Leu Ala Pro Ile Tyr Asp Ser Asp Glu Phe Tyr
1205 1210 1215
Asn Thr Ile Gln Ile Lys Glu Tyr Asp Glu Gln Pro Thr Tyr Ser
1220 1225 1230
Cys Gln Leu Leu Phe Lys Lys Asp Glu Glu Ser Thr Asp Glu Ile
1235 1240 1245
Gly Leu Ile Gly Ile His Arg Phe Tyr Glu Ser Gly Ile Val Phe
1250 1255 1260
Glu Glu Tyr Lys Asp Tyr Phe Cys Ile Ser Lys Trp Tyr Leu Lys
1265 1270 1275
Glu Val Lys Arg Lys Pro Tyr Asn Leu Lys Leu Gly Cys Asn Trp
1280 1285 1290
Gln Phe Ile Pro Lys Asp Glu Gly Trp Thr Glu
1295 1300
<210> 62
<211> 1296
<212> PRT
<213> 人工序列
<220>
<223> mrBoNT/A(0)的多肽序列
<400> 62
Met Pro Phe Val Asn Lys Gln Phe Asn Tyr Lys Asp Pro Val Asn Gly
1 5 10 15
Val Asp Ile Ala Tyr Ile Lys Ile Pro Asn Ala Gly Gln Met Gln Pro
20 25 30
Val Lys Ala Phe Lys Ile His Asn Lys Ile Trp Val Ile Pro Glu Arg
35 40 45
Asp Thr Phe Thr Asn Pro Glu Glu Gly Asp Leu Asn Pro Pro Pro Glu
50 55 60
Ala Lys Gln Val Pro Val Ser Tyr Tyr Asp Ser Thr Tyr Leu Ser Thr
65 70 75 80
Asp Asn Glu Lys Asp Asn Tyr Leu Lys Gly Val Thr Lys Leu Phe Glu
85 90 95
Arg Ile Tyr Ser Thr Asp Leu Gly Arg Met Leu Leu Thr Ser Ile Val
100 105 110
Arg Gly Ile Pro Phe Trp Gly Gly Ser Thr Ile Asp Thr Glu Leu Lys
115 120 125
Val Ile Asp Thr Asn Cys Ile Asn Val Ile Gln Pro Asp Gly Ser Tyr
130 135 140
Arg Ser Glu Glu Leu Asn Leu Val Ile Ile Gly Pro Ser Ala Asp Ile
145 150 155 160
Ile Gln Phe Glu Cys Lys Ser Phe Gly His Glu Val Leu Asn Leu Thr
165 170 175
Arg Asn Gly Tyr Gly Ser Thr Gln Tyr Ile Arg Phe Ser Pro Asp Phe
180 185 190
Thr Phe Gly Phe Glu Glu Ser Leu Glu Val Asp Thr Asn Pro Leu Leu
195 200 205
Gly Ala Gly Lys Phe Ala Thr Asp Pro Ala Val Thr Leu Ala His Gln
210 215 220
Leu Ile Tyr Ala Gly His Arg Leu Tyr Gly Ile Ala Ile Asn Pro Asn
225 230 235 240
Arg Val Phe Lys Val Asn Thr Asn Ala Tyr Tyr Glu Met Ser Gly Leu
245 250 255
Glu Val Ser Phe Glu Glu Leu Arg Thr Phe Gly Gly His Asp Ala Lys
260 265 270
Phe Ile Asp Ser Leu Gln Glu Asn Glu Phe Arg Leu Tyr Tyr Tyr Asn
275 280 285
Lys Phe Lys Asp Ile Ala Ser Thr Leu Asn Lys Ala Lys Ser Ile Val
290 295 300
Gly Thr Thr Ala Ser Leu Gln Tyr Met Lys Asn Val Phe Lys Glu Lys
305 310 315 320
Tyr Leu Leu Ser Glu Asp Thr Ser Gly Lys Phe Ser Val Asp Lys Leu
325 330 335
Lys Phe Asp Lys Leu Tyr Lys Met Leu Thr Glu Ile Tyr Thr Glu Asp
340 345 350
Asn Phe Val Lys Phe Phe Lys Val Leu Asn Arg Lys Thr Tyr Leu Asn
355 360 365
Phe Asp Lys Ala Val Phe Lys Ile Asn Ile Val Pro Lys Val Asn Tyr
370 375 380
Thr Ile Tyr Asp Gly Phe Asn Leu Arg Asn Thr Asn Leu Ala Ala Asn
385 390 395 400
Phe Asn Gly Gln Asn Thr Glu Ile Asn Asn Met Asn Phe Thr Lys Leu
405 410 415
Lys Asn Phe Thr Gly Leu Phe Glu Phe Tyr Lys Leu Leu Cys Val Arg
420 425 430
Gly Ile Ile Thr Ser Lys Thr Lys Ser Leu Asp Lys Gly Tyr Asn Lys
435 440 445
Ala Leu Asn Asp Leu Cys Ile Lys Val Asn Asn Trp Asp Leu Phe Phe
450 455 460
Ser Pro Ser Glu Asp Asn Phe Thr Asn Asp Leu Asn Lys Gly Glu Glu
465 470 475 480
Ile Thr Ser Asp Thr Asn Ile Glu Ala Ala Glu Glu Asn Ile Ser Leu
485 490 495
Asp Leu Ile Gln Gln Tyr Tyr Leu Thr Phe Asn Phe Asp Asn Glu Pro
500 505 510
Glu Asn Ile Ser Ile Glu Asn Leu Ser Ser Asp Ile Ile Gly Gln Leu
515 520 525
Glu Leu Met Pro Asn Ile Glu Arg Phe Pro Asn Gly Lys Lys Tyr Glu
530 535 540
Leu Asp Lys Tyr Thr Met Phe His Tyr Leu Arg Ala Gln Glu Phe Glu
545 550 555 560
His Gly Lys Ser Arg Ile Ala Leu Thr Asn Ser Val Asn Glu Ala Leu
565 570 575
Leu Asn Pro Ser Arg Val Tyr Thr Phe Phe Ser Ser Asp Tyr Val Lys
580 585 590
Lys Val Asn Lys Ala Thr Glu Ala Ala Met Phe Leu Gly Trp Val Glu
595 600 605
Gln Leu Val Tyr Asp Phe Thr Asp Glu Thr Ser Glu Val Ser Thr Thr
610 615 620
Asp Lys Ile Ala Asp Ile Thr Ile Ile Ile Pro Tyr Ile Gly Pro Ala
625 630 635 640
Leu Asn Ile Gly Asn Met Leu Tyr Lys Asp Asp Phe Val Gly Ala Leu
645 650 655
Ile Phe Ser Gly Ala Val Ile Leu Leu Glu Phe Ile Pro Glu Ile Ala
660 665 670
Ile Pro Val Leu Gly Thr Phe Ala Leu Val Ser Tyr Ile Ala Asn Lys
675 680 685
Val Leu Thr Val Gln Thr Ile Asp Asn Ala Leu Ser Lys Arg Asn Glu
690 695 700
Lys Trp Asp Glu Val Tyr Lys Tyr Ile Val Thr Asn Trp Leu Ala Lys
705 710 715 720
Val Asn Thr Gln Ile Asp Leu Ile Arg Lys Lys Met Lys Glu Ala Leu
725 730 735
Glu Asn Gln Ala Glu Ala Thr Lys Ala Ile Ile Asn Tyr Gln Tyr Asn
740 745 750
Gln Tyr Thr Glu Glu Glu Lys Asn Asn Ile Asn Phe Asn Ile Asp Asp
755 760 765
Leu Ser Ser Lys Leu Asn Glu Ser Ile Asn Lys Ala Met Ile Asn Ile
770 775 780
Asn Lys Phe Leu Asn Gln Cys Ser Val Ser Tyr Leu Met Asn Ser Met
785 790 795 800
Ile Pro Tyr Gly Val Lys Arg Leu Glu Asp Phe Asp Ala Ser Leu Lys
805 810 815
Asp Ala Leu Leu Lys Tyr Ile Tyr Asp Asn Arg Gly Thr Leu Ile Gly
820 825 830
Gln Val Asp Arg Leu Lys Asp Lys Val Asn Asn Thr Leu Ser Thr Asp
835 840 845
Ile Pro Phe Gln Leu Ser Lys Tyr Val Asp Asn Gln Arg Leu Leu Ser
850 855 860
Thr Phe Thr Glu Tyr Ile Lys Asn Ile Ile Asn Thr Ser Ile Leu Asn
865 870 875 880
Leu Arg Tyr Glu Ser Lys His Leu Ile Asp Leu Ser Arg Tyr Ala Ser
885 890 895
Lys Ile Asn Ile Gly Ser Lys Val Asn Phe Asp Pro Ile Asp Lys Asn
900 905 910
Gln Ile Gln Leu Phe Asn Leu Glu Ser Ser Lys Ile Glu Val Ile Leu
915 920 925
Lys Lys Ala Ile Val Tyr Asn Ser Met Tyr Glu Asn Phe Ser Thr Ser
930 935 940
Phe Trp Ile Arg Ile Pro Lys Tyr Phe Asn Lys Ile Ser Leu Asn Asn
945 950 955 960
Glu Tyr Thr Ile Ile Asn Cys Met Glu Asn Asn Ser Gly Trp Lys Val
965 970 975
Ser Leu Asn Tyr Gly Glu Ile Ile Trp Thr Leu Gln Asp Thr Lys Glu
980 985 990
Ile Lys Gln Arg Val Val Phe Lys Tyr Ser Gln Met Ile Asn Ile Ser
995 1000 1005
Asp Tyr Ile Asn Arg Trp Ile Phe Val Thr Ile Thr Asn Asn Arg
1010 1015 1020
Leu Asn Lys Ser Lys Ile Tyr Ile Asn Gly Arg Leu Ile Asp Gln
1025 1030 1035
Lys Pro Ile Ser Asn Leu Gly Asn Ile His Ala Ser Asn Lys Ile
1040 1045 1050
Met Phe Lys Leu Asp Gly Cys Arg Asp Thr His Arg Tyr Ile Trp
1055 1060 1065
Ile Lys Tyr Phe Asn Leu Phe Asp Lys Glu Leu Asn Glu Lys Glu
1070 1075 1080
Ile Lys Asp Leu Tyr Asp Asn Gln Ser Asn Ser Gly Ile Leu Lys
1085 1090 1095
Asp Phe Trp Gly Asp Tyr Leu Gln Tyr Asp Lys Pro Tyr Tyr Met
1100 1105 1110
Leu Asn Leu Tyr Asp Pro Asn Lys Tyr Val Asp Val Asn Asn Val
1115 1120 1125
Gly Ile Arg Gly Tyr Met Tyr Leu Lys Gly Pro Arg Gly Ser Val
1130 1135 1140
Met Thr Thr Asn Ile Tyr Leu Asn Ser Ser Leu Tyr Arg Gly Thr
1145 1150 1155
Lys Phe Ile Ile Lys Lys Tyr Ala Ser Gly Asn Lys Asp Asn Ile
1160 1165 1170
Val Arg Asn Asn Asp Arg Val Tyr Ile Asn Val Val Val Lys Asn
1175 1180 1185
Lys Glu Tyr Arg Leu Ala Thr Asn Ala Ser Gln Ala Gly Val Glu
1190 1195 1200
Lys Ile Leu Ser Ala Leu Glu Ile Pro Asp Val Gly Asn Leu Ser
1205 1210 1215
Gln Val Val Val Met Lys Ser Lys Asn Asp Lys Gly Ile Thr Asn
1220 1225 1230
Lys Cys Lys Met Asn Leu Gln Asp Asn Asn Gly Asn Asp Ile Gly
1235 1240 1245
Phe Ile Gly Phe His Gln Phe Asn Asn Ile Ala Lys Leu Val Ala
1250 1255 1260
Ser Asn Trp Tyr Asn Arg Gln Ile Glu Arg Ser Ser Arg Thr Leu
1265 1270 1275
Gly Cys Ser Trp Glu Phe Ile Pro Val Asp Asp Gly Trp Gly Glu
1280 1285 1290
Arg Pro Leu
1295
<210> 63
<211> 1341
<212> PRT
<213> 人工序列
<220>
<223> BoNT/XB(0)的多肽序列 (His-标签的)
<400> 63
Met Gly Ser Met Lys Leu Glu Ile Asn Lys Phe Asn Tyr Asn Asp Pro
1 5 10 15
Ile Asp Gly Ile Asn Val Ile Thr Met Arg Pro Pro Arg His Ser Asp
20 25 30
Lys Ile Asn Lys Gly Lys Gly Pro Phe Lys Ala Phe Gln Val Ile Lys
35 40 45
Asn Ile Trp Ile Val Pro Glu Arg Tyr Asn Phe Thr Asn Asn Thr Asn
50 55 60
Asp Leu Asn Ile Pro Ser Glu Pro Ile Met Glu Ala Asp Ala Ile Tyr
65 70 75 80
Asn Pro Asn Tyr Leu Asn Thr Pro Ser Glu Lys Asp Glu Phe Leu Gln
85 90 95
Gly Val Ile Lys Val Leu Glu Arg Ile Lys Ser Lys Pro Glu Gly Glu
100 105 110
Lys Leu Leu Glu Leu Ile Ser Ser Ser Ile Pro Leu Pro Leu Val Ser
115 120 125
Asn Gly Ala Leu Thr Leu Ser Asp Asn Glu Thr Ile Ala Tyr Gln Glu
130 135 140
Asn Asn Asn Ile Val Ser Asn Leu Gln Ala Asn Leu Val Ile Tyr Gly
145 150 155 160
Pro Gly Pro Asp Ile Ala Asn Asn Ala Thr Tyr Gly Leu Tyr Ser Thr
165 170 175
Pro Ile Ser Asn Gly Glu Gly Thr Leu Ser Glu Val Ser Phe Ser Pro
180 185 190
Phe Tyr Leu Lys Pro Phe Asp Glu Ser Tyr Gly Asn Tyr Arg Ser Leu
195 200 205
Val Asn Ile Val Asn Lys Phe Val Lys Arg Glu Phe Ala Pro Asp Pro
210 215 220
Ala Ser Thr Leu Met His Gln Leu Val Tyr Val Thr His Asn Leu Tyr
225 230 235 240
Gly Ile Ser Asn Arg Asn Phe Tyr Tyr Asn Phe Asp Thr Gly Lys Ile
245 250 255
Glu Thr Ser Arg Gln Gln Asn Ser Leu Ile Phe Glu Glu Leu Leu Thr
260 265 270
Phe Gly Gly Ile Asp Ser Lys Ala Ile Ser Ser Leu Ile Ile Lys Lys
275 280 285
Ile Ile Glu Thr Ala Lys Asn Asn Tyr Thr Thr Leu Ile Ser Glu Arg
290 295 300
Leu Asn Thr Val Thr Val Glu Asn Asp Leu Leu Lys Tyr Ile Lys Asn
305 310 315 320
Lys Ile Pro Val Gln Gly Arg Leu Gly Asn Phe Lys Leu Asp Thr Ala
325 330 335
Glu Phe Glu Lys Lys Leu Asn Thr Ile Leu Phe Val Leu Asn Glu Ser
340 345 350
Asn Leu Ala Gln Arg Phe Ser Ile Leu Val Arg Lys His Tyr Leu Lys
355 360 365
Glu Arg Pro Ile Asp Pro Ile Tyr Val Asn Ile Leu Asp Asp Asn Ser
370 375 380
Tyr Ser Thr Leu Glu Gly Phe Asn Ile Ser Ser Gln Gly Ser Asn Asp
385 390 395 400
Phe Gln Gly Gln Leu Leu Glu Ser Ser Tyr Phe Glu Lys Ile Glu Ser
405 410 415
Asn Ala Leu Arg Ala Phe Ile Lys Ile Cys His Lys Ala Ile Asp Gly
420 425 430
Arg Ser Leu Tyr Asn Lys Thr Leu Asp Cys Ile Glu Val Glu Asn Lys
435 440 445
Asp Leu Phe Leu Ile Ser Asn Lys Asp Ser Leu Asn Asp Ile Asn Leu
450 455 460
Ser Glu Glu Lys Ile Lys Pro Glu Thr Thr Val Phe Phe Lys Asp Lys
465 470 475 480
Leu Pro Pro Gln Asp Ile Thr Leu Ser Asn Tyr Asp Phe Thr Glu Ala
485 490 495
Asn Ser Ile Pro Ser Ile Ser Gln Gln Asn Ile Leu Glu Arg Asn Glu
500 505 510
Glu Leu Tyr Glu Pro Ile Arg Asn Ser Leu Phe Glu Ile Lys Thr Ile
515 520 525
Tyr Val Asp Lys Leu Thr Thr Phe His Phe Leu Glu Ala Gln Asn Ile
530 535 540
Asp Glu Ser Ile Asp Ser Ser Lys Ile Arg Val Glu Leu Thr Asp Ser
545 550 555 560
Val Asp Glu Ala Leu Ser Asn Pro Asn Lys Val Tyr Ser Pro Phe Lys
565 570 575
Asn Met Ser Asn Thr Ile Asn Ser Ile Glu Thr Gly Ile Thr Ser Thr
580 585 590
Tyr Ile Phe Tyr Gln Trp Leu Arg Ser Ile Val Lys Asp Phe Ser Asp
595 600 605
Glu Thr Gly Lys Ile Asp Val Ile Asp Lys Ser Ser Asp Thr Leu Ala
610 615 620
Ile Val Pro Tyr Ile Gly Pro Leu Leu Asn Ile Gly Asn Asp Ile Arg
625 630 635 640
His Gly Asp Phe Val Gly Ala Ile Glu Leu Ala Gly Ile Thr Ala Leu
645 650 655
Leu Glu Tyr Val Pro Glu Phe Thr Ile Pro Ile Leu Val Gly Leu Glu
660 665 670
Val Ile Gly Gly Glu Leu Ala Arg Glu Gln Val Glu Ala Ile Val Asn
675 680 685
Asn Ala Leu Asp Lys Arg Asp Gln Lys Trp Ala Glu Val Tyr Asn Ile
690 695 700
Thr Lys Ala Gln Trp Trp Gly Thr Ile His Leu Gln Ile Asn Thr Arg
705 710 715 720
Leu Ala His Thr Tyr Lys Ala Leu Ser Arg Gln Ala Asn Ala Ile Lys
725 730 735
Met Asn Met Glu Phe Gln Leu Ala Asn Tyr Lys Gly Asn Ile Asp Asp
740 745 750
Lys Ala Lys Ile Lys Asn Ala Ile Ser Glu Thr Glu Ile Leu Leu Asn
755 760 765
Lys Ser Val Glu Gln Ala Met Lys Asn Thr Glu Lys Phe Met Ile Lys
770 775 780
Leu Ser Asn Ser Tyr Leu Thr Lys Glu Met Ile Pro Lys Val Gln Asp
785 790 795 800
Asn Leu Lys Asn Phe Asp Leu Glu Thr Lys Lys Thr Leu Asp Lys Phe
805 810 815
Ile Lys Glu Lys Glu Asp Ile Leu Gly Thr Asn Leu Ser Ser Ser Leu
820 825 830
Arg Arg Lys Val Ser Ile Arg Leu Asn Lys Asn Ile Ala Phe Asp Ile
835 840 845
Asn Asp Ile Pro Phe Ser Glu Phe Asp Asp Leu Ile Asn Gln Tyr Lys
850 855 860
Asn Glu Ile Glu Asp Tyr Glu Val Leu Asn Leu Gly Ala Glu Asp Gly
865 870 875 880
Lys Ile Lys Asp Leu Ser Gly Thr Thr Ser Asp Ile Asn Ile Gly Ser
885 890 895
Asp Ile Glu Ile Leu Asn Asn Ile Ile Leu Asn Leu Arg Tyr Lys Asp
900 905 910
Asn Asn Leu Ile Asp Leu Ser Gly Tyr Gly Ala Lys Val Glu Val Tyr
915 920 925
Asp Gly Val Glu Leu Asn Asp Lys Asn Gln Phe Lys Leu Thr Ser Ser
930 935 940
Ala Asn Ser Lys Ile Arg Val Thr Gln Asn Gln Asn Ile Ile Phe Asn
945 950 955 960
Ser Val Phe Leu Asp Phe Ser Val Ser Phe Trp Ile Arg Ile Pro Lys
965 970 975
Tyr Lys Asn Asp Gly Ile Gln Asn Tyr Ile His Asn Glu Tyr Thr Ile
980 985 990
Ile Asn Cys Met Lys Asn Asn Ser Gly Trp Lys Ile Ser Ile Arg Gly
995 1000 1005
Asn Arg Ile Ile Trp Thr Leu Ile Asp Ile Asn Gly Lys Thr Lys
1010 1015 1020
Ser Val Phe Phe Glu Tyr Asn Ile Arg Glu Asp Ile Ser Glu Tyr
1025 1030 1035
Ile Asn Arg Trp Phe Phe Val Thr Ile Thr Asn Asn Leu Asn Asn
1040 1045 1050
Ala Lys Ile Tyr Ile Asn Gly Lys Leu Glu Ser Asn Thr Asp Ile
1055 1060 1065
Lys Asp Ile Arg Glu Val Ile Ala Asn Gly Glu Ile Ile Phe Lys
1070 1075 1080
Leu Asp Gly Asp Ile Asp Arg Thr Gln Phe Ile Trp Met Lys Tyr
1085 1090 1095
Phe Ser Ile Phe Asn Thr Glu Leu Ser Gln Ser Asn Ile Glu Glu
1100 1105 1110
Arg Tyr Lys Ile Gln Ser Tyr Ser Glu Tyr Leu Lys Asp Phe Trp
1115 1120 1125
Gly Asn Pro Leu Met Tyr Asn Lys Glu Tyr Tyr Met Phe Asn Ala
1130 1135 1140
Gly Asn Lys Asn Ser Tyr Ile Lys Leu Lys Lys Asp Ser Pro Val
1145 1150 1155
Gly Glu Ile Leu Thr Arg Ser Lys Tyr Asn Gln Asn Ser Lys Tyr
1160 1165 1170
Ile Asn Tyr Arg Asp Leu Tyr Ile Gly Glu Lys Phe Ile Ile Arg
1175 1180 1185
Arg Lys Ser Asn Ser Gln Ser Ile Asn Asp Asp Ile Val Arg Lys
1190 1195 1200
Glu Asp Tyr Ile Tyr Leu Asp Phe Phe Asn Leu Asn Gln Glu Trp
1205 1210 1215
Arg Val Tyr Thr Tyr Lys Tyr Phe Lys Lys Glu Glu Met Lys Leu
1220 1225 1230
Phe Leu Ala Pro Ile Tyr Asp Ser Asp Glu Phe Tyr Asn Thr Ile
1235 1240 1245
Gln Ile Lys Glu Tyr Asp Glu Gln Pro Thr Tyr Ser Cys Gln Leu
1250 1255 1260
Leu Phe Lys Lys Asp Glu Glu Ser Thr Asp Glu Ile Gly Leu Ile
1265 1270 1275
Gly Ile His Arg Phe Tyr Glu Ser Gly Ile Val Phe Glu Glu Tyr
1280 1285 1290
Lys Asp Tyr Phe Cys Ile Ser Lys Trp Tyr Leu Lys Glu Val Lys
1295 1300 1305
Arg Lys Pro Tyr Asn Leu Lys Leu Gly Cys Asn Trp Gln Phe Ile
1310 1315 1320
Pro Lys Asp Glu Gly Trp Thr Glu His His His His His His His
1325 1330 1335
His His His
1340
<210> 64
<211> 1331
<212> PRT
<213> 人工序列
<220>
<223> BoNT/XB(0)的多肽序列
<400> 64
Met Gly Ser Met Lys Leu Glu Ile Asn Lys Phe Asn Tyr Asn Asp Pro
1 5 10 15
Ile Asp Gly Ile Asn Val Ile Thr Met Arg Pro Pro Arg His Ser Asp
20 25 30
Lys Ile Asn Lys Gly Lys Gly Pro Phe Lys Ala Phe Gln Val Ile Lys
35 40 45
Asn Ile Trp Ile Val Pro Glu Arg Tyr Asn Phe Thr Asn Asn Thr Asn
50 55 60
Asp Leu Asn Ile Pro Ser Glu Pro Ile Met Glu Ala Asp Ala Ile Tyr
65 70 75 80
Asn Pro Asn Tyr Leu Asn Thr Pro Ser Glu Lys Asp Glu Phe Leu Gln
85 90 95
Gly Val Ile Lys Val Leu Glu Arg Ile Lys Ser Lys Pro Glu Gly Glu
100 105 110
Lys Leu Leu Glu Leu Ile Ser Ser Ser Ile Pro Leu Pro Leu Val Ser
115 120 125
Asn Gly Ala Leu Thr Leu Ser Asp Asn Glu Thr Ile Ala Tyr Gln Glu
130 135 140
Asn Asn Asn Ile Val Ser Asn Leu Gln Ala Asn Leu Val Ile Tyr Gly
145 150 155 160
Pro Gly Pro Asp Ile Ala Asn Asn Ala Thr Tyr Gly Leu Tyr Ser Thr
165 170 175
Pro Ile Ser Asn Gly Glu Gly Thr Leu Ser Glu Val Ser Phe Ser Pro
180 185 190
Phe Tyr Leu Lys Pro Phe Asp Glu Ser Tyr Gly Asn Tyr Arg Ser Leu
195 200 205
Val Asn Ile Val Asn Lys Phe Val Lys Arg Glu Phe Ala Pro Asp Pro
210 215 220
Ala Ser Thr Leu Met His Gln Leu Val Tyr Val Thr His Asn Leu Tyr
225 230 235 240
Gly Ile Ser Asn Arg Asn Phe Tyr Tyr Asn Phe Asp Thr Gly Lys Ile
245 250 255
Glu Thr Ser Arg Gln Gln Asn Ser Leu Ile Phe Glu Glu Leu Leu Thr
260 265 270
Phe Gly Gly Ile Asp Ser Lys Ala Ile Ser Ser Leu Ile Ile Lys Lys
275 280 285
Ile Ile Glu Thr Ala Lys Asn Asn Tyr Thr Thr Leu Ile Ser Glu Arg
290 295 300
Leu Asn Thr Val Thr Val Glu Asn Asp Leu Leu Lys Tyr Ile Lys Asn
305 310 315 320
Lys Ile Pro Val Gln Gly Arg Leu Gly Asn Phe Lys Leu Asp Thr Ala
325 330 335
Glu Phe Glu Lys Lys Leu Asn Thr Ile Leu Phe Val Leu Asn Glu Ser
340 345 350
Asn Leu Ala Gln Arg Phe Ser Ile Leu Val Arg Lys His Tyr Leu Lys
355 360 365
Glu Arg Pro Ile Asp Pro Ile Tyr Val Asn Ile Leu Asp Asp Asn Ser
370 375 380
Tyr Ser Thr Leu Glu Gly Phe Asn Ile Ser Ser Gln Gly Ser Asn Asp
385 390 395 400
Phe Gln Gly Gln Leu Leu Glu Ser Ser Tyr Phe Glu Lys Ile Glu Ser
405 410 415
Asn Ala Leu Arg Ala Phe Ile Lys Ile Cys His Lys Ala Ile Asp Gly
420 425 430
Arg Ser Leu Tyr Asn Lys Thr Leu Asp Cys Ile Glu Val Glu Asn Lys
435 440 445
Asp Leu Phe Leu Ile Ser Asn Lys Asp Ser Leu Asn Asp Ile Asn Leu
450 455 460
Ser Glu Glu Lys Ile Lys Pro Glu Thr Thr Val Phe Phe Lys Asp Lys
465 470 475 480
Leu Pro Pro Gln Asp Ile Thr Leu Ser Asn Tyr Asp Phe Thr Glu Ala
485 490 495
Asn Ser Ile Pro Ser Ile Ser Gln Gln Asn Ile Leu Glu Arg Asn Glu
500 505 510
Glu Leu Tyr Glu Pro Ile Arg Asn Ser Leu Phe Glu Ile Lys Thr Ile
515 520 525
Tyr Val Asp Lys Leu Thr Thr Phe His Phe Leu Glu Ala Gln Asn Ile
530 535 540
Asp Glu Ser Ile Asp Ser Ser Lys Ile Arg Val Glu Leu Thr Asp Ser
545 550 555 560
Val Asp Glu Ala Leu Ser Asn Pro Asn Lys Val Tyr Ser Pro Phe Lys
565 570 575
Asn Met Ser Asn Thr Ile Asn Ser Ile Glu Thr Gly Ile Thr Ser Thr
580 585 590
Tyr Ile Phe Tyr Gln Trp Leu Arg Ser Ile Val Lys Asp Phe Ser Asp
595 600 605
Glu Thr Gly Lys Ile Asp Val Ile Asp Lys Ser Ser Asp Thr Leu Ala
610 615 620
Ile Val Pro Tyr Ile Gly Pro Leu Leu Asn Ile Gly Asn Asp Ile Arg
625 630 635 640
His Gly Asp Phe Val Gly Ala Ile Glu Leu Ala Gly Ile Thr Ala Leu
645 650 655
Leu Glu Tyr Val Pro Glu Phe Thr Ile Pro Ile Leu Val Gly Leu Glu
660 665 670
Val Ile Gly Gly Glu Leu Ala Arg Glu Gln Val Glu Ala Ile Val Asn
675 680 685
Asn Ala Leu Asp Lys Arg Asp Gln Lys Trp Ala Glu Val Tyr Asn Ile
690 695 700
Thr Lys Ala Gln Trp Trp Gly Thr Ile His Leu Gln Ile Asn Thr Arg
705 710 715 720
Leu Ala His Thr Tyr Lys Ala Leu Ser Arg Gln Ala Asn Ala Ile Lys
725 730 735
Met Asn Met Glu Phe Gln Leu Ala Asn Tyr Lys Gly Asn Ile Asp Asp
740 745 750
Lys Ala Lys Ile Lys Asn Ala Ile Ser Glu Thr Glu Ile Leu Leu Asn
755 760 765
Lys Ser Val Glu Gln Ala Met Lys Asn Thr Glu Lys Phe Met Ile Lys
770 775 780
Leu Ser Asn Ser Tyr Leu Thr Lys Glu Met Ile Pro Lys Val Gln Asp
785 790 795 800
Asn Leu Lys Asn Phe Asp Leu Glu Thr Lys Lys Thr Leu Asp Lys Phe
805 810 815
Ile Lys Glu Lys Glu Asp Ile Leu Gly Thr Asn Leu Ser Ser Ser Leu
820 825 830
Arg Arg Lys Val Ser Ile Arg Leu Asn Lys Asn Ile Ala Phe Asp Ile
835 840 845
Asn Asp Ile Pro Phe Ser Glu Phe Asp Asp Leu Ile Asn Gln Tyr Lys
850 855 860
Asn Glu Ile Glu Asp Tyr Glu Val Leu Asn Leu Gly Ala Glu Asp Gly
865 870 875 880
Lys Ile Lys Asp Leu Ser Gly Thr Thr Ser Asp Ile Asn Ile Gly Ser
885 890 895
Asp Ile Glu Ile Leu Asn Asn Ile Ile Leu Asn Leu Arg Tyr Lys Asp
900 905 910
Asn Asn Leu Ile Asp Leu Ser Gly Tyr Gly Ala Lys Val Glu Val Tyr
915 920 925
Asp Gly Val Glu Leu Asn Asp Lys Asn Gln Phe Lys Leu Thr Ser Ser
930 935 940
Ala Asn Ser Lys Ile Arg Val Thr Gln Asn Gln Asn Ile Ile Phe Asn
945 950 955 960
Ser Val Phe Leu Asp Phe Ser Val Ser Phe Trp Ile Arg Ile Pro Lys
965 970 975
Tyr Lys Asn Asp Gly Ile Gln Asn Tyr Ile His Asn Glu Tyr Thr Ile
980 985 990
Ile Asn Cys Met Lys Asn Asn Ser Gly Trp Lys Ile Ser Ile Arg Gly
995 1000 1005
Asn Arg Ile Ile Trp Thr Leu Ile Asp Ile Asn Gly Lys Thr Lys
1010 1015 1020
Ser Val Phe Phe Glu Tyr Asn Ile Arg Glu Asp Ile Ser Glu Tyr
1025 1030 1035
Ile Asn Arg Trp Phe Phe Val Thr Ile Thr Asn Asn Leu Asn Asn
1040 1045 1050
Ala Lys Ile Tyr Ile Asn Gly Lys Leu Glu Ser Asn Thr Asp Ile
1055 1060 1065
Lys Asp Ile Arg Glu Val Ile Ala Asn Gly Glu Ile Ile Phe Lys
1070 1075 1080
Leu Asp Gly Asp Ile Asp Arg Thr Gln Phe Ile Trp Met Lys Tyr
1085 1090 1095
Phe Ser Ile Phe Asn Thr Glu Leu Ser Gln Ser Asn Ile Glu Glu
1100 1105 1110
Arg Tyr Lys Ile Gln Ser Tyr Ser Glu Tyr Leu Lys Asp Phe Trp
1115 1120 1125
Gly Asn Pro Leu Met Tyr Asn Lys Glu Tyr Tyr Met Phe Asn Ala
1130 1135 1140
Gly Asn Lys Asn Ser Tyr Ile Lys Leu Lys Lys Asp Ser Pro Val
1145 1150 1155
Gly Glu Ile Leu Thr Arg Ser Lys Tyr Asn Gln Asn Ser Lys Tyr
1160 1165 1170
Ile Asn Tyr Arg Asp Leu Tyr Ile Gly Glu Lys Phe Ile Ile Arg
1175 1180 1185
Arg Lys Ser Asn Ser Gln Ser Ile Asn Asp Asp Ile Val Arg Lys
1190 1195 1200
Glu Asp Tyr Ile Tyr Leu Asp Phe Phe Asn Leu Asn Gln Glu Trp
1205 1210 1215
Arg Val Tyr Thr Tyr Lys Tyr Phe Lys Lys Glu Glu Met Lys Leu
1220 1225 1230
Phe Leu Ala Pro Ile Tyr Asp Ser Asp Glu Phe Tyr Asn Thr Ile
1235 1240 1245
Gln Ile Lys Glu Tyr Asp Glu Gln Pro Thr Tyr Ser Cys Gln Leu
1250 1255 1260
Leu Phe Lys Lys Asp Glu Glu Ser Thr Asp Glu Ile Gly Leu Ile
1265 1270 1275
Gly Ile His Arg Phe Tyr Glu Ser Gly Ile Val Phe Glu Glu Tyr
1280 1285 1290
Lys Asp Tyr Phe Cys Ile Ser Lys Trp Tyr Leu Lys Glu Val Lys
1295 1300 1305
Arg Lys Pro Tyr Asn Leu Lys Leu Gly Cys Asn Trp Gln Phe Ile
1310 1315 1320
Pro Lys Asp Glu Gly Trp Thr Glu
1325 1330
<210> 65
<211> 1308
<212> PRT
<213> 人工序列
<220>
<223> BoNT/XB(0)变体的多肽序列(His-标签的)
<400> 65
Met Gly Ser Met Lys Leu Glu Ile Asn Lys Phe Asn Tyr Asn Asp Pro
1 5 10 15
Ile Asp Gly Ile Asn Val Ile Thr Met Arg Pro Pro Arg His Ser Asp
20 25 30
Lys Ile Asn Lys Gly Lys Gly Pro Phe Lys Ala Phe Gln Val Ile Lys
35 40 45
Asn Ile Trp Ile Val Pro Glu Arg Tyr Asn Phe Thr Asn Asn Thr Asn
50 55 60
Asp Leu Asn Ile Pro Ser Glu Pro Ile Met Glu Ala Asp Ala Ile Tyr
65 70 75 80
Asn Pro Asn Tyr Leu Asn Thr Pro Ser Glu Lys Asp Glu Phe Leu Gln
85 90 95
Gly Val Ile Lys Val Leu Glu Arg Ile Lys Ser Lys Pro Glu Gly Glu
100 105 110
Lys Leu Leu Glu Leu Ile Ser Ser Ser Ile Pro Leu Pro Leu Val Ser
115 120 125
Asn Gly Ala Leu Thr Leu Ser Asp Asn Glu Thr Ile Ala Tyr Gln Glu
130 135 140
Asn Asn Asn Ile Val Ser Asn Leu Gln Ala Asn Leu Val Ile Tyr Gly
145 150 155 160
Pro Gly Pro Asp Ile Ala Asn Asn Ala Thr Tyr Gly Leu Tyr Ser Thr
165 170 175
Pro Ile Ser Asn Gly Glu Gly Thr Leu Ser Glu Val Ser Phe Ser Pro
180 185 190
Phe Tyr Leu Lys Pro Phe Asp Glu Ser Tyr Gly Asn Tyr Arg Ser Leu
195 200 205
Val Asn Ile Val Asn Lys Phe Val Lys Arg Glu Phe Ala Pro Asp Pro
210 215 220
Ala Ser Thr Leu Met His Gln Leu Val Tyr Val Thr His Asn Leu Tyr
225 230 235 240
Gly Ile Ser Asn Arg Asn Phe Tyr Tyr Asn Phe Asp Thr Gly Lys Ile
245 250 255
Glu Thr Ser Arg Gln Gln Asn Ser Leu Ile Phe Glu Glu Leu Leu Thr
260 265 270
Phe Gly Gly Ile Asp Ser Lys Ala Ile Ser Ser Leu Ile Ile Lys Lys
275 280 285
Ile Ile Glu Thr Ala Lys Asn Asn Tyr Thr Thr Leu Ile Ser Glu Arg
290 295 300
Leu Asn Thr Val Thr Val Glu Asn Asp Leu Leu Lys Tyr Ile Lys Asn
305 310 315 320
Lys Ile Pro Val Gln Gly Arg Leu Gly Asn Phe Lys Leu Asp Thr Ala
325 330 335
Glu Phe Glu Lys Lys Leu Asn Thr Ile Leu Phe Val Leu Asn Glu Ser
340 345 350
Asn Leu Ala Gln Arg Phe Ser Ile Leu Val Arg Lys His Tyr Leu Lys
355 360 365
Glu Arg Pro Ile Asp Pro Ile Tyr Val Asn Ile Leu Asp Asp Asn Ser
370 375 380
Tyr Ser Thr Leu Glu Gly Phe Asn Ile Ser Ser Gln Gly Ser Asn Asp
385 390 395 400
Phe Gln Gly Gln Leu Leu Glu Ser Ser Tyr Phe Glu Lys Ile Glu Ser
405 410 415
Asn Ala Leu Arg Ala Phe Ile Lys Ile Cys His Lys Ala Ile Asp Gly
420 425 430
Arg Ser Leu Tyr Asn Lys Thr Leu Asp Cys Ile Glu Val Glu Asn Lys
435 440 445
Asp Leu Phe Leu Ile Ser Asn Lys Asp Ser Leu Asn Asp Ile Asn Leu
450 455 460
Ser Glu Glu Lys Ile Lys Pro Glu Thr Thr Val Phe Phe Lys Asp Lys
465 470 475 480
Leu Pro Pro Gln Asp Ile Thr Leu Ser Asn Tyr Asp Phe Thr Glu Ala
485 490 495
Asn Ser Ile Pro Ser Ile Ser Gln Gln Asn Ile Leu Glu Arg Asn Glu
500 505 510
Glu Leu Tyr Glu Pro Ile Arg Asn Ser Leu Phe Glu Ile Lys Thr Ile
515 520 525
Tyr Val Asp Lys Leu Thr Thr Phe His Phe Leu Glu Ala Gln Asn Ile
530 535 540
Asp Glu Ser Ile Asp Ser Ser Lys Ile Arg Val Glu Leu Thr Asp Ser
545 550 555 560
Val Asp Glu Ala Leu Ser Asn Pro Asn Lys Val Tyr Ser Pro Phe Lys
565 570 575
Asn Met Ser Asn Thr Ile Asn Ser Ile Glu Thr Gly Ile Thr Ser Thr
580 585 590
Tyr Ile Phe Tyr Gln Trp Leu Arg Ser Ile Val Lys Asp Phe Ser Asp
595 600 605
Glu Thr Gly Lys Ile Asp Val Ile Asp Lys Ser Ser Asp Thr Leu Ala
610 615 620
Ile Val Pro Tyr Ile Gly Pro Leu Leu Asn Ile Gly Asn Asp Ile Arg
625 630 635 640
His Gly Asp Phe Val Gly Ala Ile Glu Leu Ala Gly Ile Thr Ala Leu
645 650 655
Leu Glu Tyr Val Pro Glu Phe Thr Ile Pro Ile Leu Val Gly Leu Glu
660 665 670
Val Ile Gly Gly Glu Leu Ala Arg Glu Gln Val Glu Ala Ile Val Asn
675 680 685
Asn Ala Leu Asp Lys Arg Asp Gln Lys Trp Ala Glu Val Tyr Asn Ile
690 695 700
Thr Lys Ala Gln Trp Trp Gly Thr Ile His Leu Gln Ile Asn Thr Arg
705 710 715 720
Leu Ala His Thr Tyr Lys Ala Leu Ser Arg Gln Ala Asn Ala Ile Lys
725 730 735
Met Asn Met Glu Phe Gln Leu Ala Asn Tyr Lys Gly Asn Ile Asp Asp
740 745 750
Lys Ala Lys Ile Lys Asn Ala Ile Ser Glu Thr Glu Ile Leu Leu Asn
755 760 765
Lys Ser Val Glu Gln Ala Met Lys Asn Thr Glu Lys Phe Met Ile Lys
770 775 780
Leu Ser Asn Ser Tyr Leu Thr Lys Glu Met Ile Pro Lys Val Gln Asp
785 790 795 800
Asn Leu Lys Asn Phe Asp Leu Glu Thr Lys Lys Thr Leu Asp Lys Phe
805 810 815
Ile Lys Glu Lys Glu Asp Ile Leu Gly Thr Asn Leu Ser Ser Ser Leu
820 825 830
Arg Arg Lys Val Ser Ile Arg Leu Asn Lys Asn Ile Ala Phe Asp Ile
835 840 845
Asn Asp Ile Pro Phe Ser Glu Phe Asp Asp Leu Ile Asn Gln Tyr Lys
850 855 860
Asn Glu Ile Leu Asn Asn Ile Ile Leu Asn Leu Arg Tyr Lys Asp Asn
865 870 875 880
Asn Leu Ile Asp Leu Ser Gly Tyr Gly Ala Lys Val Glu Val Tyr Asp
885 890 895
Gly Val Glu Leu Asn Asp Lys Asn Gln Phe Lys Leu Thr Ser Ser Ala
900 905 910
Asn Ser Lys Ile Arg Val Thr Gln Asn Gln Asn Ile Ile Phe Asn Ser
915 920 925
Val Phe Leu Asp Phe Ser Val Ser Phe Trp Ile Arg Ile Pro Lys Tyr
930 935 940
Lys Asn Asp Gly Ile Gln Asn Tyr Ile His Asn Glu Tyr Thr Ile Ile
945 950 955 960
Asn Cys Met Lys Asn Asn Ser Gly Trp Lys Ile Ser Ile Arg Gly Asn
965 970 975
Arg Ile Ile Trp Thr Leu Ile Asp Ile Asn Gly Lys Thr Lys Ser Val
980 985 990
Phe Phe Glu Tyr Asn Ile Arg Glu Asp Ile Ser Glu Tyr Ile Asn Arg
995 1000 1005
Trp Phe Phe Val Thr Ile Thr Asn Asn Leu Asn Asn Ala Lys Ile
1010 1015 1020
Tyr Ile Asn Gly Lys Leu Glu Ser Asn Thr Asp Ile Lys Asp Ile
1025 1030 1035
Arg Glu Val Ile Ala Asn Gly Glu Ile Ile Phe Lys Leu Asp Gly
1040 1045 1050
Asp Ile Asp Arg Thr Gln Phe Ile Trp Met Lys Tyr Phe Ser Ile
1055 1060 1065
Phe Asn Thr Glu Leu Ser Gln Ser Asn Ile Glu Glu Arg Tyr Lys
1070 1075 1080
Ile Gln Ser Tyr Ser Glu Tyr Leu Lys Asp Phe Trp Gly Asn Pro
1085 1090 1095
Leu Met Tyr Asn Lys Glu Tyr Tyr Met Phe Asn Ala Gly Asn Lys
1100 1105 1110
Asn Ser Tyr Ile Lys Leu Lys Lys Asp Ser Pro Val Gly Glu Ile
1115 1120 1125
Leu Thr Arg Ser Lys Tyr Asn Gln Asn Ser Lys Tyr Ile Asn Tyr
1130 1135 1140
Arg Asp Leu Tyr Ile Gly Glu Lys Phe Ile Ile Arg Arg Lys Ser
1145 1150 1155
Asn Ser Gln Ser Ile Asn Asp Asp Ile Val Arg Lys Glu Asp Tyr
1160 1165 1170
Ile Tyr Leu Asp Phe Phe Asn Leu Asn Gln Glu Trp Arg Val Tyr
1175 1180 1185
Thr Tyr Lys Tyr Phe Lys Lys Glu Glu Met Lys Leu Phe Leu Ala
1190 1195 1200
Pro Ile Tyr Asp Ser Asp Glu Phe Tyr Asn Thr Ile Gln Ile Lys
1205 1210 1215
Glu Tyr Asp Glu Gln Pro Thr Tyr Ser Cys Gln Leu Leu Phe Lys
1220 1225 1230
Lys Asp Glu Glu Ser Thr Asp Glu Ile Gly Leu Ile Gly Ile His
1235 1240 1245
Arg Phe Tyr Glu Ser Gly Ile Val Phe Glu Glu Tyr Lys Asp Tyr
1250 1255 1260
Phe Cys Ile Ser Lys Trp Tyr Leu Lys Glu Val Lys Arg Lys Pro
1265 1270 1275
Tyr Asn Leu Lys Leu Gly Cys Asn Trp Gln Phe Ile Pro Lys Asp
1280 1285 1290
Glu Gly Trp Thr Glu His His His His His His His His His His
1295 1300 1305
<210> 66
<211> 1298
<212> PRT
<213> 人工序列
<220>
<223> BoNT/XB(0)变体的多肽序列
<400> 66
Met Gly Ser Met Lys Leu Glu Ile Asn Lys Phe Asn Tyr Asn Asp Pro
1 5 10 15
Ile Asp Gly Ile Asn Val Ile Thr Met Arg Pro Pro Arg His Ser Asp
20 25 30
Lys Ile Asn Lys Gly Lys Gly Pro Phe Lys Ala Phe Gln Val Ile Lys
35 40 45
Asn Ile Trp Ile Val Pro Glu Arg Tyr Asn Phe Thr Asn Asn Thr Asn
50 55 60
Asp Leu Asn Ile Pro Ser Glu Pro Ile Met Glu Ala Asp Ala Ile Tyr
65 70 75 80
Asn Pro Asn Tyr Leu Asn Thr Pro Ser Glu Lys Asp Glu Phe Leu Gln
85 90 95
Gly Val Ile Lys Val Leu Glu Arg Ile Lys Ser Lys Pro Glu Gly Glu
100 105 110
Lys Leu Leu Glu Leu Ile Ser Ser Ser Ile Pro Leu Pro Leu Val Ser
115 120 125
Asn Gly Ala Leu Thr Leu Ser Asp Asn Glu Thr Ile Ala Tyr Gln Glu
130 135 140
Asn Asn Asn Ile Val Ser Asn Leu Gln Ala Asn Leu Val Ile Tyr Gly
145 150 155 160
Pro Gly Pro Asp Ile Ala Asn Asn Ala Thr Tyr Gly Leu Tyr Ser Thr
165 170 175
Pro Ile Ser Asn Gly Glu Gly Thr Leu Ser Glu Val Ser Phe Ser Pro
180 185 190
Phe Tyr Leu Lys Pro Phe Asp Glu Ser Tyr Gly Asn Tyr Arg Ser Leu
195 200 205
Val Asn Ile Val Asn Lys Phe Val Lys Arg Glu Phe Ala Pro Asp Pro
210 215 220
Ala Ser Thr Leu Met His Gln Leu Val Tyr Val Thr His Asn Leu Tyr
225 230 235 240
Gly Ile Ser Asn Arg Asn Phe Tyr Tyr Asn Phe Asp Thr Gly Lys Ile
245 250 255
Glu Thr Ser Arg Gln Gln Asn Ser Leu Ile Phe Glu Glu Leu Leu Thr
260 265 270
Phe Gly Gly Ile Asp Ser Lys Ala Ile Ser Ser Leu Ile Ile Lys Lys
275 280 285
Ile Ile Glu Thr Ala Lys Asn Asn Tyr Thr Thr Leu Ile Ser Glu Arg
290 295 300
Leu Asn Thr Val Thr Val Glu Asn Asp Leu Leu Lys Tyr Ile Lys Asn
305 310 315 320
Lys Ile Pro Val Gln Gly Arg Leu Gly Asn Phe Lys Leu Asp Thr Ala
325 330 335
Glu Phe Glu Lys Lys Leu Asn Thr Ile Leu Phe Val Leu Asn Glu Ser
340 345 350
Asn Leu Ala Gln Arg Phe Ser Ile Leu Val Arg Lys His Tyr Leu Lys
355 360 365
Glu Arg Pro Ile Asp Pro Ile Tyr Val Asn Ile Leu Asp Asp Asn Ser
370 375 380
Tyr Ser Thr Leu Glu Gly Phe Asn Ile Ser Ser Gln Gly Ser Asn Asp
385 390 395 400
Phe Gln Gly Gln Leu Leu Glu Ser Ser Tyr Phe Glu Lys Ile Glu Ser
405 410 415
Asn Ala Leu Arg Ala Phe Ile Lys Ile Cys His Lys Ala Ile Asp Gly
420 425 430
Arg Ser Leu Tyr Asn Lys Thr Leu Asp Cys Ile Glu Val Glu Asn Lys
435 440 445
Asp Leu Phe Leu Ile Ser Asn Lys Asp Ser Leu Asn Asp Ile Asn Leu
450 455 460
Ser Glu Glu Lys Ile Lys Pro Glu Thr Thr Val Phe Phe Lys Asp Lys
465 470 475 480
Leu Pro Pro Gln Asp Ile Thr Leu Ser Asn Tyr Asp Phe Thr Glu Ala
485 490 495
Asn Ser Ile Pro Ser Ile Ser Gln Gln Asn Ile Leu Glu Arg Asn Glu
500 505 510
Glu Leu Tyr Glu Pro Ile Arg Asn Ser Leu Phe Glu Ile Lys Thr Ile
515 520 525
Tyr Val Asp Lys Leu Thr Thr Phe His Phe Leu Glu Ala Gln Asn Ile
530 535 540
Asp Glu Ser Ile Asp Ser Ser Lys Ile Arg Val Glu Leu Thr Asp Ser
545 550 555 560
Val Asp Glu Ala Leu Ser Asn Pro Asn Lys Val Tyr Ser Pro Phe Lys
565 570 575
Asn Met Ser Asn Thr Ile Asn Ser Ile Glu Thr Gly Ile Thr Ser Thr
580 585 590
Tyr Ile Phe Tyr Gln Trp Leu Arg Ser Ile Val Lys Asp Phe Ser Asp
595 600 605
Glu Thr Gly Lys Ile Asp Val Ile Asp Lys Ser Ser Asp Thr Leu Ala
610 615 620
Ile Val Pro Tyr Ile Gly Pro Leu Leu Asn Ile Gly Asn Asp Ile Arg
625 630 635 640
His Gly Asp Phe Val Gly Ala Ile Glu Leu Ala Gly Ile Thr Ala Leu
645 650 655
Leu Glu Tyr Val Pro Glu Phe Thr Ile Pro Ile Leu Val Gly Leu Glu
660 665 670
Val Ile Gly Gly Glu Leu Ala Arg Glu Gln Val Glu Ala Ile Val Asn
675 680 685
Asn Ala Leu Asp Lys Arg Asp Gln Lys Trp Ala Glu Val Tyr Asn Ile
690 695 700
Thr Lys Ala Gln Trp Trp Gly Thr Ile His Leu Gln Ile Asn Thr Arg
705 710 715 720
Leu Ala His Thr Tyr Lys Ala Leu Ser Arg Gln Ala Asn Ala Ile Lys
725 730 735
Met Asn Met Glu Phe Gln Leu Ala Asn Tyr Lys Gly Asn Ile Asp Asp
740 745 750
Lys Ala Lys Ile Lys Asn Ala Ile Ser Glu Thr Glu Ile Leu Leu Asn
755 760 765
Lys Ser Val Glu Gln Ala Met Lys Asn Thr Glu Lys Phe Met Ile Lys
770 775 780
Leu Ser Asn Ser Tyr Leu Thr Lys Glu Met Ile Pro Lys Val Gln Asp
785 790 795 800
Asn Leu Lys Asn Phe Asp Leu Glu Thr Lys Lys Thr Leu Asp Lys Phe
805 810 815
Ile Lys Glu Lys Glu Asp Ile Leu Gly Thr Asn Leu Ser Ser Ser Leu
820 825 830
Arg Arg Lys Val Ser Ile Arg Leu Asn Lys Asn Ile Ala Phe Asp Ile
835 840 845
Asn Asp Ile Pro Phe Ser Glu Phe Asp Asp Leu Ile Asn Gln Tyr Lys
850 855 860
Asn Glu Ile Leu Asn Asn Ile Ile Leu Asn Leu Arg Tyr Lys Asp Asn
865 870 875 880
Asn Leu Ile Asp Leu Ser Gly Tyr Gly Ala Lys Val Glu Val Tyr Asp
885 890 895
Gly Val Glu Leu Asn Asp Lys Asn Gln Phe Lys Leu Thr Ser Ser Ala
900 905 910
Asn Ser Lys Ile Arg Val Thr Gln Asn Gln Asn Ile Ile Phe Asn Ser
915 920 925
Val Phe Leu Asp Phe Ser Val Ser Phe Trp Ile Arg Ile Pro Lys Tyr
930 935 940
Lys Asn Asp Gly Ile Gln Asn Tyr Ile His Asn Glu Tyr Thr Ile Ile
945 950 955 960
Asn Cys Met Lys Asn Asn Ser Gly Trp Lys Ile Ser Ile Arg Gly Asn
965 970 975
Arg Ile Ile Trp Thr Leu Ile Asp Ile Asn Gly Lys Thr Lys Ser Val
980 985 990
Phe Phe Glu Tyr Asn Ile Arg Glu Asp Ile Ser Glu Tyr Ile Asn Arg
995 1000 1005
Trp Phe Phe Val Thr Ile Thr Asn Asn Leu Asn Asn Ala Lys Ile
1010 1015 1020
Tyr Ile Asn Gly Lys Leu Glu Ser Asn Thr Asp Ile Lys Asp Ile
1025 1030 1035
Arg Glu Val Ile Ala Asn Gly Glu Ile Ile Phe Lys Leu Asp Gly
1040 1045 1050
Asp Ile Asp Arg Thr Gln Phe Ile Trp Met Lys Tyr Phe Ser Ile
1055 1060 1065
Phe Asn Thr Glu Leu Ser Gln Ser Asn Ile Glu Glu Arg Tyr Lys
1070 1075 1080
Ile Gln Ser Tyr Ser Glu Tyr Leu Lys Asp Phe Trp Gly Asn Pro
1085 1090 1095
Leu Met Tyr Asn Lys Glu Tyr Tyr Met Phe Asn Ala Gly Asn Lys
1100 1105 1110
Asn Ser Tyr Ile Lys Leu Lys Lys Asp Ser Pro Val Gly Glu Ile
1115 1120 1125
Leu Thr Arg Ser Lys Tyr Asn Gln Asn Ser Lys Tyr Ile Asn Tyr
1130 1135 1140
Arg Asp Leu Tyr Ile Gly Glu Lys Phe Ile Ile Arg Arg Lys Ser
1145 1150 1155
Asn Ser Gln Ser Ile Asn Asp Asp Ile Val Arg Lys Glu Asp Tyr
1160 1165 1170
Ile Tyr Leu Asp Phe Phe Asn Leu Asn Gln Glu Trp Arg Val Tyr
1175 1180 1185
Thr Tyr Lys Tyr Phe Lys Lys Glu Glu Met Lys Leu Phe Leu Ala
1190 1195 1200
Pro Ile Tyr Asp Ser Asp Glu Phe Tyr Asn Thr Ile Gln Ile Lys
1205 1210 1215
Glu Tyr Asp Glu Gln Pro Thr Tyr Ser Cys Gln Leu Leu Phe Lys
1220 1225 1230
Lys Asp Glu Glu Ser Thr Asp Glu Ile Gly Leu Ile Gly Ile His
1235 1240 1245
Arg Phe Tyr Glu Ser Gly Ile Val Phe Glu Glu Tyr Lys Asp Tyr
1250 1255 1260
Phe Cys Ile Ser Lys Trp Tyr Leu Lys Glu Val Lys Arg Lys Pro
1265 1270 1275
Tyr Asn Leu Lys Leu Gly Cys Asn Trp Gln Phe Ile Pro Lys Asp
1280 1285 1290
Glu Gly Trp Thr Glu
1295
<210> 67
<211> 1331
<212> PRT
<213> 人工序列
<220>
<223> BoNT/XA(0)的多肽序列
<400> 67
Met Gly Ser Met Lys Leu Glu Ile Asn Lys Phe Asn Tyr Asn Asp Pro
1 5 10 15
Ile Asp Gly Ile Asn Val Ile Thr Met Arg Pro Pro Arg His Ser Asp
20 25 30
Lys Ile Asn Lys Gly Lys Gly Pro Phe Lys Ala Phe Gln Val Ile Lys
35 40 45
Asn Ile Trp Ile Val Pro Glu Arg Tyr Asn Phe Thr Asn Asn Thr Asn
50 55 60
Asp Leu Asn Ile Pro Ser Glu Pro Ile Met Glu Ala Asp Ala Ile Tyr
65 70 75 80
Asn Pro Asn Tyr Leu Asn Thr Pro Ser Glu Lys Asp Glu Phe Leu Gln
85 90 95
Gly Val Ile Lys Val Leu Glu Arg Ile Lys Ser Lys Pro Glu Gly Glu
100 105 110
Lys Leu Leu Glu Leu Ile Ser Ser Ser Ile Pro Leu Pro Leu Val Ser
115 120 125
Asn Gly Ala Leu Thr Leu Ser Asp Asn Glu Thr Ile Ala Tyr Gln Glu
130 135 140
Asn Asn Asn Ile Val Ser Asn Leu Gln Ala Asn Leu Val Ile Tyr Gly
145 150 155 160
Pro Gly Pro Asp Ile Ala Asn Asn Ala Thr Tyr Gly Leu Tyr Ser Thr
165 170 175
Pro Ile Ser Asn Gly Glu Gly Thr Leu Ser Glu Val Ser Phe Ser Pro
180 185 190
Phe Tyr Leu Lys Pro Phe Asp Glu Ser Tyr Gly Asn Tyr Arg Ser Leu
195 200 205
Val Asn Ile Val Asn Lys Phe Val Lys Arg Glu Phe Ala Pro Asp Pro
210 215 220
Ala Ser Thr Leu Met His Gln Leu Val Tyr Val Thr His Asn Leu Tyr
225 230 235 240
Gly Ile Ser Asn Arg Asn Phe Tyr Tyr Asn Phe Asp Thr Gly Lys Ile
245 250 255
Glu Thr Ser Arg Gln Gln Asn Ser Leu Ile Phe Glu Glu Leu Leu Thr
260 265 270
Phe Gly Gly Ile Asp Ser Lys Ala Ile Ser Ser Leu Ile Ile Lys Lys
275 280 285
Ile Ile Glu Thr Ala Lys Asn Asn Tyr Thr Thr Leu Ile Ser Glu Arg
290 295 300
Leu Asn Thr Val Thr Val Glu Asn Asp Leu Leu Lys Tyr Ile Lys Asn
305 310 315 320
Lys Ile Pro Val Gln Gly Arg Leu Gly Asn Phe Lys Leu Asp Thr Ala
325 330 335
Glu Phe Glu Lys Lys Leu Asn Thr Ile Leu Phe Val Leu Asn Glu Ser
340 345 350
Asn Leu Ala Gln Arg Phe Ser Ile Leu Val Arg Lys His Tyr Leu Lys
355 360 365
Glu Arg Pro Ile Asp Pro Ile Tyr Val Asn Ile Leu Asp Asp Asn Ser
370 375 380
Tyr Ser Thr Leu Glu Gly Phe Asn Ile Ser Ser Gln Gly Ser Asn Asp
385 390 395 400
Phe Gln Gly Gln Leu Leu Glu Ser Ser Tyr Phe Glu Lys Ile Glu Ser
405 410 415
Asn Ala Leu Arg Ala Phe Ile Lys Ile Cys His Lys Ala Ile Asp Gly
420 425 430
Arg Ser Leu Glu Val Leu Phe Gln Gly Pro Leu Tyr Asn Lys Thr Leu
435 440 445
Asp Cys Ile Glu Val Glu Asn Lys Asp Leu Phe Leu Ile Ser Asn Lys
450 455 460
Asp Ser Leu Asn Asp Ile Asn Leu Ser Glu Glu Lys Ile Lys Pro Glu
465 470 475 480
Thr Thr Val Phe Phe Lys Asp Lys Leu Pro Pro Gln Asp Ile Thr Leu
485 490 495
Ser Asn Tyr Asp Phe Thr Glu Ala Asn Ser Ile Pro Ser Ile Ser Gln
500 505 510
Gln Asn Ile Leu Glu Arg Asn Glu Glu Leu Tyr Glu Pro Ile Arg Asn
515 520 525
Ser Leu Phe Glu Ile Lys Thr Ile Tyr Val Asp Lys Leu Thr Thr Phe
530 535 540
His Phe Leu Glu Ala Gln Asn Ile Asp Glu Ser Ile Asp Ser Ser Lys
545 550 555 560
Ile Arg Val Glu Leu Thr Asp Ser Val Asp Glu Ala Leu Ser Asn Pro
565 570 575
Asn Lys Val Tyr Ser Pro Phe Lys Asn Met Ser Asn Thr Ile Asn Ser
580 585 590
Ile Glu Thr Gly Ile Thr Ser Thr Tyr Ile Phe Tyr Gln Trp Leu Arg
595 600 605
Ser Ile Val Lys Asp Phe Ser Asp Glu Thr Gly Lys Ile Asp Val Ile
610 615 620
Asp Lys Ser Ser Asp Thr Leu Ala Ile Val Pro Tyr Ile Gly Pro Leu
625 630 635 640
Leu Asn Ile Gly Asn Asp Ile Arg His Gly Asp Phe Val Gly Ala Ile
645 650 655
Glu Leu Ala Gly Ile Thr Ala Leu Leu Glu Tyr Val Pro Glu Phe Thr
660 665 670
Ile Pro Ile Leu Val Gly Leu Glu Val Ile Gly Gly Glu Leu Ala Arg
675 680 685
Glu Gln Val Glu Ala Ile Val Asn Asn Ala Leu Asp Lys Arg Asp Gln
690 695 700
Lys Trp Ala Glu Val Tyr Asn Ile Thr Lys Ala Gln Trp Trp Gly Thr
705 710 715 720
Ile His Leu Gln Ile Asn Thr Arg Leu Ala His Thr Tyr Lys Ala Leu
725 730 735
Ser Arg Gln Ala Asn Ala Ile Lys Met Asn Met Glu Phe Gln Leu Ala
740 745 750
Asn Tyr Lys Gly Asn Ile Asp Asp Lys Ala Lys Ile Lys Asn Ala Ile
755 760 765
Ser Glu Thr Glu Ile Leu Leu Asn Lys Ser Val Glu Gln Ala Met Lys
770 775 780
Asn Thr Glu Lys Phe Met Ile Lys Leu Ser Asn Ser Tyr Leu Thr Lys
785 790 795 800
Glu Met Ile Pro Lys Val Gln Asp Asn Leu Lys Asn Phe Asp Leu Glu
805 810 815
Thr Lys Lys Thr Leu Asp Lys Phe Ile Lys Glu Lys Glu Asp Ile Leu
820 825 830
Gly Thr Asn Leu Ser Ser Ser Leu Arg Arg Lys Val Ser Ile Arg Leu
835 840 845
Asn Lys Asn Ile Ala Phe Asp Ile Asn Asp Ile Pro Phe Ser Glu Phe
850 855 860
Asp Asp Leu Ile Asn Gln Tyr Lys Asn Glu Ile Glu Asp Tyr Glu Val
865 870 875 880
Leu Asn Leu Gly Ala Glu Asp Gly Lys Ile Lys Asp Leu Ser Gly Thr
885 890 895
Thr Ser Asp Ile Asn Ile Gly Ser Asp Ile Glu Ile Ile Asn Thr Ser
900 905 910
Ile Leu Asn Leu Arg Tyr Glu Ser Asn His Leu Ile Asp Leu Ser Arg
915 920 925
Tyr Ala Ser Lys Ile Asn Ile Gly Ser Lys Val Asn Phe Asp Pro Ile
930 935 940
Asp Lys Asn Gln Ile Gln Leu Phe Asn Leu Glu Ser Ser Lys Ile Glu
945 950 955 960
Val Ile Leu Lys Asn Ala Ile Val Tyr Asn Ser Met Tyr Glu Asn Phe
965 970 975
Ser Thr Ser Phe Trp Ile Arg Ile Pro Lys Tyr Phe Asn Ser Ile Ser
980 985 990
Leu Asn Asn Glu Tyr Thr Ile Ile Asn Cys Met Glu Asn Asn Ser Gly
995 1000 1005
Trp Lys Val Ser Leu Asn Tyr Gly Glu Ile Ile Trp Thr Leu Gln
1010 1015 1020
Asp Thr Gln Glu Ile Lys Gln Arg Val Val Phe Lys Tyr Ser Gln
1025 1030 1035
Met Ile Asn Ile Ser Asp Tyr Ile Asn Arg Trp Ile Phe Val Thr
1040 1045 1050
Ile Thr Asn Asn Arg Leu Asn Asn Ser Lys Ile Tyr Ile Asn Gly
1055 1060 1065
Arg Leu Ile Asp Gln Lys Pro Ile Ser Asn Leu Gly Asn Ile His
1070 1075 1080
Ala Ser Asn Asn Ile Met Phe Lys Leu Asp Gly Cys Arg Asp Thr
1085 1090 1095
His Arg Tyr Ile Trp Ile Lys Tyr Phe Asn Leu Phe Asp Lys Glu
1100 1105 1110
Leu Asn Glu Lys Glu Ile Lys Asp Leu Tyr Asp Asn Gln Ser Asn
1115 1120 1125
Ser Gly Ile Leu Lys Asp Phe Trp Gly Asp Tyr Leu Gln Tyr Asp
1130 1135 1140
Lys Pro Tyr Tyr Met Leu Asn Leu Tyr Asp Pro Asn Lys Tyr Val
1145 1150 1155
Asp Val Asn Asn Val Gly Ile Arg Gly Tyr Met Tyr Leu Lys Gly
1160 1165 1170
Pro Arg Gly Ser Val Met Thr Thr Asn Ile Tyr Leu Asn Ser Ser
1175 1180 1185
Leu Tyr Arg Gly Thr Lys Phe Ile Ile Lys Lys Tyr Ala Ser Gly
1190 1195 1200
Asn Lys Asp Asn Ile Val Arg Asn Asn Asp Arg Val Tyr Ile Asn
1205 1210 1215
Val Val Val Lys Asn Lys Glu Tyr Arg Leu Ala Thr Asn Ala Ser
1220 1225 1230
Gln Ala Gly Val Glu Lys Ile Leu Ser Ala Leu Glu Ile Pro Asp
1235 1240 1245
Val Gly Asn Leu Ser Gln Val Val Val Met Lys Ser Lys Asn Asp
1250 1255 1260
Gln Gly Ile Thr Asn Lys Cys Lys Met Asn Leu Gln Asp Asn Asn
1265 1270 1275
Gly Asn Asp Ile Gly Phe Ile Gly Phe His Gln Phe Asn Asn Ile
1280 1285 1290
Ala Lys Leu Val Ala Ser Asn Trp Tyr Asn Arg Gln Ile Glu Arg
1295 1300 1305
Ser Ser Arg Thr Leu Gly Cys Ser Trp Glu Phe Ile Pro Val Asp
1310 1315 1320
Asp Gly Trp Gly Glu Arg Pro Leu
1325 1330
<210> 68
<211> 1298
<212> PRT
<213> 人工序列
<220>
<223> BoNT/XA(0)变体的多肽序列
<400> 68
Met Gly Ser Met Lys Leu Glu Ile Asn Lys Phe Asn Tyr Asn Asp Pro
1 5 10 15
Ile Asp Gly Ile Asn Val Ile Thr Met Arg Pro Pro Arg His Ser Asp
20 25 30
Lys Ile Asn Lys Gly Lys Gly Pro Phe Lys Ala Phe Gln Val Ile Lys
35 40 45
Asn Ile Trp Ile Val Pro Glu Arg Tyr Asn Phe Thr Asn Asn Thr Asn
50 55 60
Asp Leu Asn Ile Pro Ser Glu Pro Ile Met Glu Ala Asp Ala Ile Tyr
65 70 75 80
Asn Pro Asn Tyr Leu Asn Thr Pro Ser Glu Lys Asp Glu Phe Leu Gln
85 90 95
Gly Val Ile Lys Val Leu Glu Arg Ile Lys Ser Lys Pro Glu Gly Glu
100 105 110
Lys Leu Leu Glu Leu Ile Ser Ser Ser Ile Pro Leu Pro Leu Val Ser
115 120 125
Asn Gly Ala Leu Thr Leu Ser Asp Asn Glu Thr Ile Ala Tyr Gln Glu
130 135 140
Asn Asn Asn Ile Val Ser Asn Leu Gln Ala Asn Leu Val Ile Tyr Gly
145 150 155 160
Pro Gly Pro Asp Ile Ala Asn Asn Ala Thr Tyr Gly Leu Tyr Ser Thr
165 170 175
Pro Ile Ser Asn Gly Glu Gly Thr Leu Ser Glu Val Ser Phe Ser Pro
180 185 190
Phe Tyr Leu Lys Pro Phe Asp Glu Ser Tyr Gly Asn Tyr Arg Ser Leu
195 200 205
Val Asn Ile Val Asn Lys Phe Val Lys Arg Glu Phe Ala Pro Asp Pro
210 215 220
Ala Ser Thr Leu Met His Gln Leu Val Tyr Val Thr His Asn Leu Tyr
225 230 235 240
Gly Ile Ser Asn Arg Asn Phe Tyr Tyr Asn Phe Asp Thr Gly Lys Ile
245 250 255
Glu Thr Ser Arg Gln Gln Asn Ser Leu Ile Phe Glu Glu Leu Leu Thr
260 265 270
Phe Gly Gly Ile Asp Ser Lys Ala Ile Ser Ser Leu Ile Ile Lys Lys
275 280 285
Ile Ile Glu Thr Ala Lys Asn Asn Tyr Thr Thr Leu Ile Ser Glu Arg
290 295 300
Leu Asn Thr Val Thr Val Glu Asn Asp Leu Leu Lys Tyr Ile Lys Asn
305 310 315 320
Lys Ile Pro Val Gln Gly Arg Leu Gly Asn Phe Lys Leu Asp Thr Ala
325 330 335
Glu Phe Glu Lys Lys Leu Asn Thr Ile Leu Phe Val Leu Asn Glu Ser
340 345 350
Asn Leu Ala Gln Arg Phe Ser Ile Leu Val Arg Lys His Tyr Leu Lys
355 360 365
Glu Arg Pro Ile Asp Pro Ile Tyr Val Asn Ile Leu Asp Asp Asn Ser
370 375 380
Tyr Ser Thr Leu Glu Gly Phe Asn Ile Ser Ser Gln Gly Ser Asn Asp
385 390 395 400
Phe Gln Gly Gln Leu Leu Glu Ser Ser Tyr Phe Glu Lys Ile Glu Ser
405 410 415
Asn Ala Leu Arg Ala Phe Ile Lys Ile Cys His Lys Ala Ile Asp Gly
420 425 430
Arg Ser Leu Glu Val Leu Phe Gln Gly Pro Leu Tyr Asn Lys Thr Leu
435 440 445
Asp Cys Ile Glu Val Glu Asn Lys Asp Leu Phe Leu Ile Ser Asn Lys
450 455 460
Asp Ser Leu Asn Asp Ile Asn Leu Ser Glu Glu Lys Ile Lys Pro Glu
465 470 475 480
Thr Thr Val Phe Phe Lys Asp Lys Leu Pro Pro Gln Asp Ile Thr Leu
485 490 495
Ser Asn Tyr Asp Phe Thr Glu Ala Asn Ser Ile Pro Ser Ile Ser Gln
500 505 510
Gln Asn Ile Leu Glu Arg Asn Glu Glu Leu Tyr Glu Pro Ile Arg Asn
515 520 525
Ser Leu Phe Glu Ile Lys Thr Ile Tyr Val Asp Lys Leu Thr Thr Phe
530 535 540
His Phe Leu Glu Ala Gln Asn Ile Asp Glu Ser Ile Asp Ser Ser Lys
545 550 555 560
Ile Arg Val Glu Leu Thr Asp Ser Val Asp Glu Ala Leu Ser Asn Pro
565 570 575
Asn Lys Val Tyr Ser Pro Phe Lys Asn Met Ser Asn Thr Ile Asn Ser
580 585 590
Ile Glu Thr Gly Ile Thr Ser Thr Tyr Ile Phe Tyr Gln Trp Leu Arg
595 600 605
Ser Ile Val Lys Asp Phe Ser Asp Glu Thr Gly Lys Ile Asp Val Ile
610 615 620
Asp Lys Ser Ser Asp Thr Leu Ala Ile Val Pro Tyr Ile Gly Pro Leu
625 630 635 640
Leu Asn Ile Gly Asn Asp Ile Arg His Gly Asp Phe Val Gly Ala Ile
645 650 655
Glu Leu Ala Gly Ile Thr Ala Leu Leu Glu Tyr Val Pro Glu Phe Thr
660 665 670
Ile Pro Ile Leu Val Gly Leu Glu Val Ile Gly Gly Glu Leu Ala Arg
675 680 685
Glu Gln Val Glu Ala Ile Val Asn Asn Ala Leu Asp Lys Arg Asp Gln
690 695 700
Lys Trp Ala Glu Val Tyr Asn Ile Thr Lys Ala Gln Trp Trp Gly Thr
705 710 715 720
Ile His Leu Gln Ile Asn Thr Arg Leu Ala His Thr Tyr Lys Ala Leu
725 730 735
Ser Arg Gln Ala Asn Ala Ile Lys Met Asn Met Glu Phe Gln Leu Ala
740 745 750
Asn Tyr Lys Gly Asn Ile Asp Asp Lys Ala Lys Ile Lys Asn Ala Ile
755 760 765
Ser Glu Thr Glu Ile Leu Leu Asn Lys Ser Val Glu Gln Ala Met Lys
770 775 780
Asn Thr Glu Lys Phe Met Ile Lys Leu Ser Asn Ser Tyr Leu Thr Lys
785 790 795 800
Glu Met Ile Pro Lys Val Gln Asp Asn Leu Lys Asn Phe Asp Leu Glu
805 810 815
Thr Lys Lys Thr Leu Asp Lys Phe Ile Lys Glu Lys Glu Asp Ile Leu
820 825 830
Gly Thr Asn Leu Ser Ser Ser Leu Arg Arg Lys Val Ser Ile Arg Leu
835 840 845
Asn Lys Asn Ile Ala Phe Asp Ile Asn Asp Ile Pro Phe Ser Glu Phe
850 855 860
Asp Asp Leu Ile Asn Gln Tyr Lys Asn Glu Ile Ile Asn Thr Ser Ile
865 870 875 880
Leu Asn Leu Arg Tyr Glu Ser Asn His Leu Ile Asp Leu Ser Arg Tyr
885 890 895
Ala Ser Lys Ile Asn Ile Gly Ser Lys Val Asn Phe Asp Pro Ile Asp
900 905 910
Lys Asn Gln Ile Gln Leu Phe Asn Leu Glu Ser Ser Lys Ile Glu Val
915 920 925
Ile Leu Lys Asn Ala Ile Val Tyr Asn Ser Met Tyr Glu Asn Phe Ser
930 935 940
Thr Ser Phe Trp Ile Arg Ile Pro Lys Tyr Phe Asn Ser Ile Ser Leu
945 950 955 960
Asn Asn Glu Tyr Thr Ile Ile Asn Cys Met Glu Asn Asn Ser Gly Trp
965 970 975
Lys Val Ser Leu Asn Tyr Gly Glu Ile Ile Trp Thr Leu Gln Asp Thr
980 985 990
Gln Glu Ile Lys Gln Arg Val Val Phe Lys Tyr Ser Gln Met Ile Asn
995 1000 1005
Ile Ser Asp Tyr Ile Asn Arg Trp Ile Phe Val Thr Ile Thr Asn
1010 1015 1020
Asn Arg Leu Asn Asn Ser Lys Ile Tyr Ile Asn Gly Arg Leu Ile
1025 1030 1035
Asp Gln Lys Pro Ile Ser Asn Leu Gly Asn Ile His Ala Ser Asn
1040 1045 1050
Asn Ile Met Phe Lys Leu Asp Gly Cys Arg Asp Thr His Arg Tyr
1055 1060 1065
Ile Trp Ile Lys Tyr Phe Asn Leu Phe Asp Lys Glu Leu Asn Glu
1070 1075 1080
Lys Glu Ile Lys Asp Leu Tyr Asp Asn Gln Ser Asn Ser Gly Ile
1085 1090 1095
Leu Lys Asp Phe Trp Gly Asp Tyr Leu Gln Tyr Asp Lys Pro Tyr
1100 1105 1110
Tyr Met Leu Asn Leu Tyr Asp Pro Asn Lys Tyr Val Asp Val Asn
1115 1120 1125
Asn Val Gly Ile Arg Gly Tyr Met Tyr Leu Lys Gly Pro Arg Gly
1130 1135 1140
Ser Val Met Thr Thr Asn Ile Tyr Leu Asn Ser Ser Leu Tyr Arg
1145 1150 1155
Gly Thr Lys Phe Ile Ile Lys Lys Tyr Ala Ser Gly Asn Lys Asp
1160 1165 1170
Asn Ile Val Arg Asn Asn Asp Arg Val Tyr Ile Asn Val Val Val
1175 1180 1185
Lys Asn Lys Glu Tyr Arg Leu Ala Thr Asn Ala Ser Gln Ala Gly
1190 1195 1200
Val Glu Lys Ile Leu Ser Ala Leu Glu Ile Pro Asp Val Gly Asn
1205 1210 1215
Leu Ser Gln Val Val Val Met Lys Ser Lys Asn Asp Gln Gly Ile
1220 1225 1230
Thr Asn Lys Cys Lys Met Asn Leu Gln Asp Asn Asn Gly Asn Asp
1235 1240 1245
Ile Gly Phe Ile Gly Phe His Gln Phe Asn Asn Ile Ala Lys Leu
1250 1255 1260
Val Ala Ser Asn Trp Tyr Asn Arg Gln Ile Glu Arg Ser Ser Arg
1265 1270 1275
Thr Leu Gly Cys Ser Trp Glu Phe Ile Pro Val Asp Asp Gly Trp
1280 1285 1290
Gly Glu Arg Pro Leu
1295
<210> 69
<211> 1311
<212> PRT
<213> 人工序列
<220>
<223> BoNT/XD(0)的多肽序列
<400> 69
Met Gly Ser Met Lys Leu Glu Ile Asn Lys Phe Asn Tyr Asn Asp Pro
1 5 10 15
Ile Asp Gly Ile Asn Val Ile Thr Met Arg Pro Pro Arg His Ser Asp
20 25 30
Lys Ile Asn Lys Gly Lys Gly Pro Phe Lys Ala Phe Gln Val Ile Lys
35 40 45
Asn Ile Trp Ile Val Pro Glu Arg Tyr Asn Phe Thr Asn Asn Thr Asn
50 55 60
Asp Leu Asn Ile Pro Ser Glu Pro Ile Met Glu Ala Asp Ala Ile Tyr
65 70 75 80
Asn Pro Asn Tyr Leu Asn Thr Pro Ser Glu Lys Asp Glu Phe Leu Gln
85 90 95
Gly Val Ile Lys Val Leu Glu Arg Ile Lys Ser Lys Pro Glu Gly Glu
100 105 110
Lys Leu Leu Glu Leu Ile Ser Ser Ser Ile Pro Leu Pro Leu Val Ser
115 120 125
Asn Gly Ala Leu Thr Leu Ser Asp Asn Glu Thr Ile Ala Tyr Gln Glu
130 135 140
Asn Asn Asn Ile Val Ser Asn Leu Gln Ala Asn Leu Val Ile Tyr Gly
145 150 155 160
Pro Gly Pro Asp Ile Ala Asn Asn Ala Thr Tyr Gly Leu Tyr Ser Thr
165 170 175
Pro Ile Ser Asn Gly Glu Gly Thr Leu Ser Glu Val Ser Phe Ser Pro
180 185 190
Phe Tyr Leu Lys Pro Phe Asp Glu Ser Tyr Gly Asn Tyr Arg Ser Leu
195 200 205
Val Asn Ile Val Asn Lys Phe Val Lys Arg Glu Phe Ala Pro Asp Pro
210 215 220
Ala Ser Thr Leu Met His Gln Leu Val Tyr Val Thr His Asn Leu Tyr
225 230 235 240
Gly Ile Ser Asn Arg Asn Phe Tyr Tyr Asn Phe Asp Thr Gly Lys Ile
245 250 255
Glu Thr Ser Arg Gln Gln Asn Ser Leu Ile Phe Glu Glu Leu Leu Thr
260 265 270
Phe Gly Gly Ile Asp Ser Lys Ala Ile Ser Ser Leu Ile Ile Lys Lys
275 280 285
Ile Ile Glu Thr Ala Lys Asn Asn Tyr Thr Thr Leu Ile Ser Glu Arg
290 295 300
Leu Asn Thr Val Thr Val Glu Asn Asp Leu Leu Lys Tyr Ile Lys Asn
305 310 315 320
Lys Ile Pro Val Gln Gly Arg Leu Gly Asn Phe Lys Leu Asp Thr Ala
325 330 335
Glu Phe Glu Lys Lys Leu Asn Thr Ile Leu Phe Val Leu Asn Glu Ser
340 345 350
Asn Leu Ala Gln Arg Phe Ser Ile Leu Val Arg Lys His Tyr Leu Lys
355 360 365
Glu Arg Pro Ile Asp Pro Ile Tyr Val Asn Ile Leu Asp Asp Asn Ser
370 375 380
Tyr Ser Thr Leu Glu Gly Phe Asn Ile Ser Ser Gln Gly Ser Asn Asp
385 390 395 400
Phe Gln Gly Gln Leu Leu Glu Ser Ser Tyr Phe Glu Lys Ile Glu Ser
405 410 415
Asn Ala Leu Arg Ala Phe Ile Lys Ile Cys His Lys Ala Ile Asp Gly
420 425 430
Arg Ser Leu Tyr Asn Lys Thr Leu Asp Cys Ile Glu Val Glu Asn Lys
435 440 445
Asp Leu Phe Leu Ile Ser Asn Lys Asp Ser Leu Asn Asp Ile Asn Leu
450 455 460
Ser Glu Glu Lys Ile Lys Pro Glu Thr Thr Val Phe Phe Lys Asp Lys
465 470 475 480
Leu Pro Pro Gln Asp Ile Thr Leu Ser Asn Tyr Asp Phe Thr Glu Ala
485 490 495
Asn Ser Ile Pro Ser Ile Ser Gln Gln Asn Ile Leu Glu Arg Asn Glu
500 505 510
Glu Leu Tyr Glu Pro Ile Arg Asn Ser Leu Phe Glu Ile Lys Thr Ile
515 520 525
Tyr Val Asp Lys Leu Thr Thr Phe His Phe Leu Glu Ala Gln Asn Ile
530 535 540
Asp Glu Ser Ile Asp Ser Ser Lys Ile Arg Val Glu Leu Thr Asp Ser
545 550 555 560
Val Asp Glu Ala Leu Ser Asn Pro Asn Lys Val Tyr Ser Pro Phe Lys
565 570 575
Asn Met Ser Asn Thr Ile Asn Ser Ile Glu Thr Gly Ile Thr Ser Thr
580 585 590
Tyr Ile Phe Tyr Gln Trp Leu Arg Ser Ile Val Lys Asp Phe Ser Asp
595 600 605
Glu Thr Gly Lys Ile Asp Val Ile Asp Lys Ser Ser Asp Thr Leu Ala
610 615 620
Ile Val Pro Tyr Ile Gly Pro Leu Leu Asn Ile Gly Asn Asp Ile Arg
625 630 635 640
His Gly Asp Phe Val Gly Ala Ile Glu Leu Ala Gly Ile Thr Ala Leu
645 650 655
Leu Glu Tyr Val Pro Glu Phe Thr Ile Pro Ile Leu Val Gly Leu Glu
660 665 670
Val Ile Gly Gly Glu Leu Ala Arg Glu Gln Val Glu Ala Ile Val Asn
675 680 685
Asn Ala Leu Asp Lys Arg Asp Gln Lys Trp Ala Glu Val Tyr Asn Ile
690 695 700
Thr Lys Ala Gln Trp Trp Gly Thr Ile His Leu Gln Ile Asn Thr Arg
705 710 715 720
Leu Ala His Thr Tyr Lys Ala Leu Ser Arg Gln Ala Asn Ala Ile Lys
725 730 735
Met Asn Met Glu Phe Gln Leu Ala Asn Tyr Lys Gly Asn Ile Asp Asp
740 745 750
Lys Ala Lys Ile Lys Asn Ala Ile Ser Glu Thr Glu Ile Leu Leu Asn
755 760 765
Lys Ser Val Glu Gln Ala Met Lys Asn Thr Glu Lys Phe Met Ile Lys
770 775 780
Leu Ser Asn Ser Tyr Leu Thr Lys Glu Met Ile Pro Lys Val Gln Asp
785 790 795 800
Asn Leu Lys Asn Phe Asp Leu Glu Thr Lys Lys Thr Leu Asp Lys Phe
805 810 815
Ile Lys Glu Lys Glu Asp Ile Leu Gly Thr Asn Leu Ser Ser Ser Leu
820 825 830
Arg Arg Lys Val Ser Ile Arg Leu Asn Lys Asn Ile Ala Phe Asp Ile
835 840 845
Asn Asp Ile Pro Phe Ser Glu Phe Asp Asp Leu Ile Asn Gln Tyr Lys
850 855 860
Asn Glu Ile Glu Asp Tyr Glu Val Leu Asn Leu Gly Ala Glu Asp Gly
865 870 875 880
Lys Ile Lys Asp Leu Ser Gly Thr Thr Ser Asp Ile Asn Ile Gly Ser
885 890 895
Asp Ile Glu Asn Asp Ser Lys Ile Leu Ser Leu Gln Asn Lys Lys Asn
900 905 910
Ala Leu Val Asp Thr Ser Gly Tyr Asn Ala Glu Val Arg Val Gly Asp
915 920 925
Asn Val Gln Leu Asn Thr Ile Tyr Thr Asn Asp Phe Lys Leu Ser Ser
930 935 940
Ser Gly Asp Lys Ile Ile Val Asn Leu Asn Asn Asn Ile Leu Tyr Ser
945 950 955 960
Ala Ile Tyr Glu Asn Ser Ser Val Ser Phe Trp Ile Lys Ile Ser Lys
965 970 975
Asp Leu Thr Asn Ser His Asn Glu Tyr Thr Ile Ile Asn Ser Ile Glu
980 985 990
Gln Asn Ser Gly Trp Lys Leu Cys Ile Arg Asn Gly Asn Ile Glu Trp
995 1000 1005
Ile Leu Gln Asp Val Asn Arg Lys Tyr Lys Ser Leu Ile Phe Asp
1010 1015 1020
Tyr Ser Glu Ser Leu Ser His Thr Gly Tyr Thr Asn Lys Trp Phe
1025 1030 1035
Phe Val Thr Ile Thr Asn Asn Ile Met Gly Tyr Met Lys Leu Tyr
1040 1045 1050
Ile Asn Gly Glu Leu Lys Gln Ser Gln Lys Ile Glu Asp Leu Asp
1055 1060 1065
Glu Val Lys Leu Asp Lys Thr Ile Val Phe Gly Ile Asp Glu Asn
1070 1075 1080
Ile Asp Glu Asn Gln Met Leu Trp Ile Arg Asp Phe Asn Ile Phe
1085 1090 1095
Ser Lys Glu Leu Ser Asn Glu Asp Ile Asn Ile Val Tyr Glu Gly
1100 1105 1110
Gln Ile Leu Arg Asn Val Ile Lys Asp Tyr Trp Gly Asn Pro Leu
1115 1120 1125
Lys Phe Asp Thr Glu Tyr Tyr Ile Ile Asn Asp Asn Tyr Ile Asp
1130 1135 1140
Arg Tyr Ile Ala Pro Glu Ser Asn Val Leu Val Leu Val Gln Tyr
1145 1150 1155
Pro Asp Arg Ser Lys Leu Tyr Thr Gly Asn Pro Ile Thr Ile Lys
1160 1165 1170
Ser Val Ser Asp Lys Asn Pro Tyr Ser Arg Ile Leu Asn Gly Asp
1175 1180 1185
Asn Ile Ile Leu His Met Leu Tyr Asn Ser Arg Lys Tyr Met Ile
1190 1195 1200
Ile Arg Asp Thr Asp Thr Ile Tyr Ala Thr Gln Gly Gly Glu Cys
1205 1210 1215
Ser Gln Asn Cys Val Tyr Ala Leu Lys Leu Gln Ser Asn Leu Gly
1220 1225 1230
Asn Tyr Gly Ile Gly Ile Phe Ser Ile Lys Asn Ile Val Ser Lys
1235 1240 1245
Asn Lys Tyr Cys Ser Gln Ile Phe Ser Ser Phe Arg Glu Asn Thr
1250 1255 1260
Met Leu Leu Ala Asp Ile Tyr Lys Pro Trp Arg Phe Ser Phe Lys
1265 1270 1275
Asn Ala Tyr Thr Pro Val Ala Val Thr Asn Tyr Glu Thr Lys Leu
1280 1285 1290
Leu Ser Thr Ser Ser Phe Trp Lys Phe Ile Ser Arg Asp Pro Gly
1295 1300 1305
Trp Val Glu
1310
<210> 70
<211> 1312
<212> PRT
<213> 人工序列
<220>
<223> BoNT/XF(0)的多肽序列
<400> 70
Met Gly Ser Met Lys Leu Glu Ile Asn Lys Phe Asn Tyr Asn Asp Pro
1 5 10 15
Ile Asp Gly Ile Asn Val Ile Thr Met Arg Pro Pro Arg His Ser Asp
20 25 30
Lys Ile Asn Lys Gly Lys Gly Pro Phe Lys Ala Phe Gln Val Ile Lys
35 40 45
Asn Ile Trp Ile Val Pro Glu Arg Tyr Asn Phe Thr Asn Asn Thr Asn
50 55 60
Asp Leu Asn Ile Pro Ser Glu Pro Ile Met Glu Ala Asp Ala Ile Tyr
65 70 75 80
Asn Pro Asn Tyr Leu Asn Thr Pro Ser Glu Lys Asp Glu Phe Leu Gln
85 90 95
Gly Val Ile Lys Val Leu Glu Arg Ile Lys Ser Lys Pro Glu Gly Glu
100 105 110
Lys Leu Leu Glu Leu Ile Ser Ser Ser Ile Pro Leu Pro Leu Val Ser
115 120 125
Asn Gly Ala Leu Thr Leu Ser Asp Asn Glu Thr Ile Ala Tyr Gln Glu
130 135 140
Asn Asn Asn Ile Val Ser Asn Leu Gln Ala Asn Leu Val Ile Tyr Gly
145 150 155 160
Pro Gly Pro Asp Ile Ala Asn Asn Ala Thr Tyr Gly Leu Tyr Ser Thr
165 170 175
Pro Ile Ser Asn Gly Glu Gly Thr Leu Ser Glu Val Ser Phe Ser Pro
180 185 190
Phe Tyr Leu Lys Pro Phe Asp Glu Ser Tyr Gly Asn Tyr Arg Ser Leu
195 200 205
Val Asn Ile Val Asn Lys Phe Val Lys Arg Glu Phe Ala Pro Asp Pro
210 215 220
Ala Ser Thr Leu Met His Gln Leu Val Tyr Val Thr His Asn Leu Tyr
225 230 235 240
Gly Ile Ser Asn Arg Asn Phe Tyr Tyr Asn Phe Asp Thr Gly Lys Ile
245 250 255
Glu Thr Ser Arg Gln Gln Asn Ser Leu Ile Phe Glu Glu Leu Leu Thr
260 265 270
Phe Gly Gly Ile Asp Ser Lys Ala Ile Ser Ser Leu Ile Ile Lys Lys
275 280 285
Ile Ile Glu Thr Ala Lys Asn Asn Tyr Thr Thr Leu Ile Ser Glu Arg
290 295 300
Leu Asn Thr Val Thr Val Glu Asn Asp Leu Leu Lys Tyr Ile Lys Asn
305 310 315 320
Lys Ile Pro Val Gln Gly Arg Leu Gly Asn Phe Lys Leu Asp Thr Ala
325 330 335
Glu Phe Glu Lys Lys Leu Asn Thr Ile Leu Phe Val Leu Asn Glu Ser
340 345 350
Asn Leu Ala Gln Arg Phe Ser Ile Leu Val Arg Lys His Tyr Leu Lys
355 360 365
Glu Arg Pro Ile Asp Pro Ile Tyr Val Asn Ile Leu Asp Asp Asn Ser
370 375 380
Tyr Ser Thr Leu Glu Gly Phe Asn Ile Ser Ser Gln Gly Ser Asn Asp
385 390 395 400
Phe Gln Gly Gln Leu Leu Glu Ser Ser Tyr Phe Glu Lys Ile Glu Ser
405 410 415
Asn Ala Leu Arg Ala Phe Ile Lys Ile Cys His Lys Ala Ile Asp Gly
420 425 430
Arg Ser Leu Tyr Asn Lys Thr Leu Asp Cys Ile Glu Val Glu Asn Lys
435 440 445
Asp Leu Phe Leu Ile Ser Asn Lys Asp Ser Leu Asn Asp Ile Asn Leu
450 455 460
Ser Glu Glu Lys Ile Lys Pro Glu Thr Thr Val Phe Phe Lys Asp Lys
465 470 475 480
Leu Pro Pro Gln Asp Ile Thr Leu Ser Asn Tyr Asp Phe Thr Glu Ala
485 490 495
Asn Ser Ile Pro Ser Ile Ser Gln Gln Asn Ile Leu Glu Arg Asn Glu
500 505 510
Glu Leu Tyr Glu Pro Ile Arg Asn Ser Leu Phe Glu Ile Lys Thr Ile
515 520 525
Tyr Val Asp Lys Leu Thr Thr Phe His Phe Leu Glu Ala Gln Asn Ile
530 535 540
Asp Glu Ser Ile Asp Ser Ser Lys Ile Arg Val Glu Leu Thr Asp Ser
545 550 555 560
Val Asp Glu Ala Leu Ser Asn Pro Asn Lys Val Tyr Ser Pro Phe Lys
565 570 575
Asn Met Ser Asn Thr Ile Asn Ser Ile Glu Thr Gly Ile Thr Ser Thr
580 585 590
Tyr Ile Phe Tyr Gln Trp Leu Arg Ser Ile Val Lys Asp Phe Ser Asp
595 600 605
Glu Thr Gly Lys Ile Asp Val Ile Asp Lys Ser Ser Asp Thr Leu Ala
610 615 620
Ile Val Pro Tyr Ile Gly Pro Leu Leu Asn Ile Gly Asn Asp Ile Arg
625 630 635 640
His Gly Asp Phe Val Gly Ala Ile Glu Leu Ala Gly Ile Thr Ala Leu
645 650 655
Leu Glu Tyr Val Pro Glu Phe Thr Ile Pro Ile Leu Val Gly Leu Glu
660 665 670
Val Ile Gly Gly Glu Leu Ala Arg Glu Gln Val Glu Ala Ile Val Asn
675 680 685
Asn Ala Leu Asp Lys Arg Asp Gln Lys Trp Ala Glu Val Tyr Asn Ile
690 695 700
Thr Lys Ala Gln Trp Trp Gly Thr Ile His Leu Gln Ile Asn Thr Arg
705 710 715 720
Leu Ala His Thr Tyr Lys Ala Leu Ser Arg Gln Ala Asn Ala Ile Lys
725 730 735
Met Asn Met Glu Phe Gln Leu Ala Asn Tyr Lys Gly Asn Ile Asp Asp
740 745 750
Lys Ala Lys Ile Lys Asn Ala Ile Ser Glu Thr Glu Ile Leu Leu Asn
755 760 765
Lys Ser Val Glu Gln Ala Met Lys Asn Thr Glu Lys Phe Met Ile Lys
770 775 780
Leu Ser Asn Ser Tyr Leu Thr Lys Glu Met Ile Pro Lys Val Gln Asp
785 790 795 800
Asn Leu Lys Asn Phe Asp Leu Glu Thr Lys Lys Thr Leu Asp Lys Phe
805 810 815
Ile Lys Glu Lys Glu Asp Ile Leu Gly Thr Asn Leu Ser Ser Ser Leu
820 825 830
Arg Arg Lys Val Ser Ile Arg Leu Asn Lys Asn Ile Ala Phe Asp Ile
835 840 845
Asn Asp Ile Pro Phe Ser Glu Phe Asp Asp Leu Ile Asn Gln Tyr Lys
850 855 860
Asn Glu Ile Glu Asp Tyr Glu Val Leu Asn Leu Gly Ala Glu Asp Gly
865 870 875 880
Lys Ile Lys Asp Leu Ser Gly Thr Thr Ser Asp Ile Asn Ile Gly Ser
885 890 895
Asp Ile Glu Ile Lys Asp Asn Ser Ile Leu Asp Met Arg Tyr Glu Asn
900 905 910
Asn Lys Phe Ile Asp Ile Ser Gly Tyr Gly Ser Asn Ile Ser Ile Asn
915 920 925
Gly Asp Val Tyr Ile Tyr Ser Thr Asn Arg Asn Gln Phe Gly Ile Tyr
930 935 940
Ser Ser Lys Pro Ser Glu Val Asn Ile Ala Gln Asn Asn Asp Ile Ile
945 950 955 960
Tyr Asn Gly Arg Tyr Gln Asn Phe Ser Ile Ser Phe Trp Val Arg Ile
965 970 975
Pro Lys Tyr Phe Asn Lys Val Asn Leu Asn Asn Glu Tyr Thr Ile Ile
980 985 990
Asp Cys Ile Arg Asn Asn Asn Ser Gly Trp Lys Ile Ser Leu Asn Tyr
995 1000 1005
Asn Lys Ile Ile Trp Thr Leu Gln Asp Thr Ala Gly Asn Asn Gln
1010 1015 1020
Lys Leu Val Phe Asn Tyr Thr Gln Met Ile Ser Ile Ser Asp Tyr
1025 1030 1035
Ile Asn Lys Trp Ile Phe Val Thr Ile Thr Asn Asn Arg Leu Gly
1040 1045 1050
Asn Ser Arg Ile Tyr Ile Asn Gly Asn Leu Ile Asp Glu Lys Ser
1055 1060 1065
Ile Ser Asn Leu Gly Asp Ile His Val Ser Asp Asn Ile Leu Phe
1070 1075 1080
Lys Ile Val Gly Cys Asn Asp Thr Arg Tyr Val Gly Ile Arg Tyr
1085 1090 1095
Phe Lys Val Phe Asp Thr Glu Leu Gly Lys Thr Glu Ile Glu Thr
1100 1105 1110
Leu Tyr Ser Asp Glu Pro Asp Pro Ser Ile Leu Lys Asp Phe Trp
1115 1120 1125
Gly Asn Tyr Leu Leu Tyr Asn Lys Arg Tyr Tyr Leu Leu Asn Leu
1130 1135 1140
Leu Arg Thr Asp Lys Ser Ile Thr Gln Asn Ser Asn Phe Leu Asn
1145 1150 1155
Ile Asn Gln Gln Arg Gly Val Tyr Gln Lys Pro Asn Ile Phe Ser
1160 1165 1170
Asn Thr Arg Leu Tyr Thr Gly Val Glu Val Ile Ile Arg Lys Asn
1175 1180 1185
Gly Ser Thr Asp Ile Ser Asn Thr Asp Asn Phe Val Arg Lys Asn
1190 1195 1200
Asp Leu Ala Tyr Ile Asn Val Val Asp Arg Asp Val Glu Tyr Arg
1205 1210 1215
Leu Tyr Ala Asp Ile Ser Ile Ala Lys Pro Glu Lys Ile Ile Lys
1220 1225 1230
Leu Ile Arg Thr Ser Asn Ser Asn Asn Ser Leu Gly Gln Ile Ile
1235 1240 1245
Val Met Asp Ser Ile Gly Asn Asn Cys Thr Met Asn Phe Gln Asn
1250 1255 1260
Asn Asn Gly Gly Asn Ile Gly Leu Leu Gly Phe Lys Ser Asn Asn
1265 1270 1275
Leu Val Ala Ser Ser Trp Tyr Tyr Asn Asn Ile Arg Lys Asn Thr
1280 1285 1290
Ser Ser Asn Gly Cys Phe Trp Ser Phe Ile Ser Lys Glu His Gly
1295 1300 1305
Trp Gln Glu Asn
1310
<210> 71
<211> 8
<212> PRT
<213> 人工序列
<220>
<223> C1激活环共有序列
<220>
<221> SITE
<222> (2)..(2)
<223> Xaa是1-10个残基的任意氨基酸
<220>
<221> 变体
<222> (4)..(4)
<223> Xaa是Asp或Glu
<220>
<221> SITE
<222> (7)..(7)
<223> Xaa是4-15个残基的任意氨基酸
<400> 71
Cys Xaa Ile Xaa Gly Arg Xaa Cys
<210> 72
<211> 17
<212> PRT
<213> 人工序列
<220>
<223> C1激活环
<400> 72
Cys His Lys Ala Ile Asp Gly Arg Ser Leu Tyr Asn Lys Thr Leu Asp
1 5 10 15
Cys
<210> 73
<211> 17
<212> PRT
<213> 人工序列
<220>
<223> C1激活环变体
<400> 73
Cys His Lys Ala Ile Glu Gly Arg Ser Leu Tyr Asn Lys Thr Leu Asp
1 5 10 15
Cys
<210> 74
<211> 441
<212> PRT
<213> 人工序列
<220>
<223> rLC/A(0)的多肽序列 (His-标签的)
<400> 74
Met Pro Phe Val Asn Lys Gln Phe Asn Tyr Lys Asp Pro Val Asn Gly
1 5 10 15
Val Asp Ile Ala Tyr Ile Lys Ile Pro Asn Ala Gly Gln Met Gln Pro
20 25 30
Val Lys Ala Phe Lys Ile His Asn Lys Ile Trp Val Ile Pro Glu Arg
35 40 45
Asp Thr Phe Thr Asn Pro Glu Glu Gly Asp Leu Asn Pro Pro Pro Glu
50 55 60
Ala Lys Gln Val Pro Val Ser Tyr Tyr Asp Ser Thr Tyr Leu Ser Thr
65 70 75 80
Asp Asn Glu Lys Asp Asn Tyr Leu Lys Gly Val Thr Lys Leu Phe Glu
85 90 95
Arg Ile Tyr Ser Thr Asp Leu Gly Arg Met Leu Leu Thr Ser Ile Val
100 105 110
Arg Gly Ile Pro Phe Trp Gly Gly Ser Thr Ile Asp Thr Glu Leu Lys
115 120 125
Val Ile Asp Thr Asn Cys Ile Asn Val Ile Gln Pro Asp Gly Ser Tyr
130 135 140
Arg Ser Glu Glu Leu Asn Leu Val Ile Ile Gly Pro Ser Ala Asp Ile
145 150 155 160
Ile Gln Phe Glu Cys Lys Ser Phe Gly His Glu Val Leu Asn Leu Thr
165 170 175
Arg Asn Gly Tyr Gly Ser Thr Gln Tyr Ile Arg Phe Ser Pro Asp Phe
180 185 190
Thr Phe Gly Phe Glu Glu Ser Leu Glu Val Asp Thr Asn Pro Leu Leu
195 200 205
Gly Ala Gly Lys Phe Ala Thr Asp Pro Ala Val Thr Leu Ala His Gln
210 215 220
Leu Ile Tyr Ala Gly His Arg Leu Tyr Gly Ile Ala Ile Asn Pro Asn
225 230 235 240
Arg Val Phe Lys Val Asn Thr Asn Ala Tyr Tyr Glu Met Ser Gly Leu
245 250 255
Glu Val Ser Phe Glu Glu Leu Arg Thr Phe Gly Gly His Asp Ala Lys
260 265 270
Phe Ile Asp Ser Leu Gln Glu Asn Glu Phe Arg Leu Tyr Tyr Tyr Asn
275 280 285
Lys Phe Lys Asp Ile Ala Ser Thr Leu Asn Lys Ala Lys Ser Ile Val
290 295 300
Gly Thr Thr Ala Ser Leu Gln Tyr Met Lys Asn Val Phe Lys Glu Lys
305 310 315 320
Tyr Leu Leu Ser Glu Asp Thr Ser Gly Lys Phe Ser Val Asp Lys Leu
325 330 335
Lys Phe Asp Lys Leu Tyr Lys Met Leu Thr Glu Ile Tyr Thr Glu Asp
340 345 350
Asn Phe Val Lys Phe Phe Lys Val Leu Asn Arg Lys Thr Tyr Leu Asn
355 360 365
Phe Asp Lys Ala Val Phe Lys Ile Asn Ile Val Pro Lys Val Asn Tyr
370 375 380
Thr Ile Tyr Asp Gly Phe Asn Leu Arg Asn Thr Asn Leu Ala Ala Asn
385 390 395 400
Phe Asn Gly Gln Asn Thr Glu Ile Asn Asn Met Asn Phe Thr Lys Leu
405 410 415
Lys Asn Phe Thr Gly Leu Phe Glu Glu Asn Leu Tyr Phe Gln Gly Ala
420 425 430
Ser His His His His His His His His
435 440
<210> 75
<211> 882
<212> PRT
<213> 人工序列
<220>
<223> rLHN/A(0)的多肽序列 (His-标签的)
<400> 75
Met Pro Phe Val Asn Lys Gln Phe Asn Tyr Lys Asp Pro Val Asn Gly
1 5 10 15
Val Asp Ile Ala Tyr Ile Lys Ile Pro Asn Ala Gly Gln Met Gln Pro
20 25 30
Val Lys Ala Phe Lys Ile His Asn Lys Ile Trp Val Ile Pro Glu Arg
35 40 45
Asp Thr Phe Thr Asn Pro Glu Glu Gly Asp Leu Asn Pro Pro Pro Glu
50 55 60
Ala Lys Gln Val Pro Val Ser Tyr Tyr Asp Ser Thr Tyr Leu Ser Thr
65 70 75 80
Asp Asn Glu Lys Asp Asn Tyr Leu Lys Gly Val Thr Lys Leu Phe Glu
85 90 95
Arg Ile Tyr Ser Thr Asp Leu Gly Arg Met Leu Leu Thr Ser Ile Val
100 105 110
Arg Gly Ile Pro Phe Trp Gly Gly Ser Thr Ile Asp Thr Glu Leu Lys
115 120 125
Val Ile Asp Thr Asn Cys Ile Asn Val Ile Gln Pro Asp Gly Ser Tyr
130 135 140
Arg Ser Glu Glu Leu Asn Leu Val Ile Ile Gly Pro Ser Ala Asp Ile
145 150 155 160
Ile Gln Phe Glu Cys Lys Ser Phe Gly His Glu Val Leu Asn Leu Thr
165 170 175
Arg Asn Gly Tyr Gly Ser Thr Gln Tyr Ile Arg Phe Ser Pro Asp Phe
180 185 190
Thr Phe Gly Phe Glu Glu Ser Leu Glu Val Asp Thr Asn Pro Leu Leu
195 200 205
Gly Ala Gly Lys Phe Ala Thr Asp Pro Ala Val Thr Leu Ala His Gln
210 215 220
Leu Ile Tyr Ala Gly His Arg Leu Tyr Gly Ile Ala Ile Asn Pro Asn
225 230 235 240
Arg Val Phe Lys Val Asn Thr Asn Ala Tyr Tyr Glu Met Ser Gly Leu
245 250 255
Glu Val Ser Phe Glu Glu Leu Arg Thr Phe Gly Gly His Asp Ala Lys
260 265 270
Phe Ile Asp Ser Leu Gln Glu Asn Glu Phe Arg Leu Tyr Tyr Tyr Asn
275 280 285
Lys Phe Lys Asp Ile Ala Ser Thr Leu Asn Lys Ala Lys Ser Ile Val
290 295 300
Gly Thr Thr Ala Ser Leu Gln Tyr Met Lys Asn Val Phe Lys Glu Lys
305 310 315 320
Tyr Leu Leu Ser Glu Asp Thr Ser Gly Lys Phe Ser Val Asp Lys Leu
325 330 335
Lys Phe Asp Lys Leu Tyr Lys Met Leu Thr Glu Ile Tyr Thr Glu Asp
340 345 350
Asn Phe Val Lys Phe Phe Lys Val Leu Asn Arg Lys Thr Tyr Leu Asn
355 360 365
Phe Asp Lys Ala Val Phe Lys Ile Asn Ile Val Pro Lys Val Asn Tyr
370 375 380
Thr Ile Tyr Asp Gly Phe Asn Leu Arg Asn Thr Asn Leu Ala Ala Asn
385 390 395 400
Phe Asn Gly Gln Asn Thr Glu Ile Asn Asn Met Asn Phe Thr Lys Leu
405 410 415
Lys Asn Phe Thr Gly Leu Phe Glu Phe Tyr Lys Leu Leu Cys Val Arg
420 425 430
Gly Ile Ile Thr Ser Lys Thr Lys Ser Leu Asp Lys Gly Tyr Asn Lys
435 440 445
Ala Leu Asn Asp Leu Cys Ile Lys Val Asn Asn Trp Asp Leu Phe Phe
450 455 460
Ser Pro Ser Glu Asp Asn Phe Thr Asn Asp Leu Asn Lys Gly Glu Glu
465 470 475 480
Ile Thr Ser Asp Thr Asn Ile Glu Ala Ala Glu Glu Asn Ile Ser Leu
485 490 495
Asp Leu Ile Gln Gln Tyr Tyr Leu Thr Phe Asn Phe Asp Asn Glu Pro
500 505 510
Glu Asn Ile Ser Ile Glu Asn Leu Ser Ser Asp Ile Ile Gly Gln Leu
515 520 525
Glu Leu Met Pro Asn Ile Glu Arg Phe Pro Asn Gly Lys Lys Tyr Glu
530 535 540
Leu Asp Lys Tyr Thr Met Phe His Tyr Leu Arg Ala Gln Glu Phe Glu
545 550 555 560
His Gly Lys Ser Arg Ile Ala Leu Thr Asn Ser Val Asn Glu Ala Leu
565 570 575
Leu Asn Pro Ser Arg Val Tyr Thr Phe Phe Ser Ser Asp Tyr Val Lys
580 585 590
Lys Val Asn Lys Ala Thr Glu Ala Ala Met Phe Leu Gly Trp Val Glu
595 600 605
Gln Leu Val Tyr Asp Phe Thr Asp Glu Thr Ser Glu Val Ser Thr Thr
610 615 620
Asp Lys Ile Ala Asp Ile Thr Ile Ile Ile Pro Tyr Ile Gly Pro Ala
625 630 635 640
Leu Asn Ile Gly Asn Met Leu Tyr Lys Asp Asp Phe Val Gly Ala Leu
645 650 655
Ile Phe Ser Gly Ala Val Ile Leu Leu Glu Phe Ile Pro Glu Ile Ala
660 665 670
Ile Pro Val Leu Gly Thr Phe Ala Leu Val Ser Tyr Ile Ala Asn Lys
675 680 685
Val Leu Thr Val Gln Thr Ile Asp Asn Ala Leu Ser Lys Arg Asn Glu
690 695 700
Lys Trp Asp Glu Val Tyr Lys Tyr Ile Val Thr Asn Trp Leu Ala Lys
705 710 715 720
Val Asn Thr Gln Ile Asp Leu Ile Arg Lys Lys Met Lys Glu Ala Leu
725 730 735
Glu Asn Gln Ala Glu Ala Thr Lys Ala Ile Ile Asn Tyr Gln Tyr Asn
740 745 750
Gln Tyr Thr Glu Glu Glu Lys Asn Asn Ile Asn Phe Asn Ile Asp Asp
755 760 765
Leu Ser Ser Lys Leu Asn Glu Ser Ile Asn Lys Ala Met Ile Asn Ile
770 775 780
Asn Lys Phe Leu Asn Gln Cys Ser Val Ser Tyr Leu Met Asn Ser Met
785 790 795 800
Ile Pro Tyr Gly Val Lys Arg Leu Glu Asp Phe Asp Ala Ser Leu Lys
805 810 815
Asp Ala Leu Leu Lys Tyr Ile Tyr Asp Asn Arg Gly Thr Leu Ile Gly
820 825 830
Gln Val Asp Arg Leu Lys Asp Lys Val Asn Asn Thr Leu Ser Thr Asp
835 840 845
Ile Pro Phe Gln Leu Ser Lys Tyr Val Asp Asn Gln Arg Leu Leu Ser
850 855 860
Thr Glu Asn Leu Tyr Phe Gln Gly Ala Ser His His His His His His
865 870 875 880
His His
<210> 76
<211> 438
<212> PRT
<213> 人工序列
<220>
<223> rLC/X(0)的多肽序列
<400> 76
Met Lys Leu Glu Ile Asn Lys Phe Asn Tyr Asn Asp Pro Ile Asp Gly
1 5 10 15
Ile Asn Val Ile Thr Met Arg Pro Pro Arg His Ser Asp Lys Ile Asn
20 25 30
Lys Gly Lys Gly Pro Phe Lys Ala Phe Gln Val Ile Lys Asn Ile Trp
35 40 45
Ile Val Pro Glu Arg Tyr Asn Phe Thr Asn Asn Thr Asn Asp Leu Asn
50 55 60
Ile Pro Ser Glu Pro Ile Met Glu Ala Asp Ala Ile Tyr Asn Pro Asn
65 70 75 80
Tyr Leu Asn Thr Pro Ser Glu Lys Asp Glu Phe Leu Gln Gly Val Ile
85 90 95
Lys Val Leu Glu Arg Ile Lys Ser Lys Pro Glu Gly Glu Lys Leu Leu
100 105 110
Glu Leu Ile Ser Ser Ser Ile Pro Leu Pro Leu Val Ser Asn Gly Ala
115 120 125
Leu Thr Leu Ser Asp Asn Glu Thr Ile Ala Tyr Gln Glu Asn Asn Asn
130 135 140
Ile Val Ser Asn Leu Gln Ala Asn Leu Val Ile Tyr Gly Pro Gly Pro
145 150 155 160
Asp Ile Ala Asn Asn Ala Thr Tyr Gly Leu Tyr Ser Thr Pro Ile Ser
165 170 175
Asn Gly Glu Gly Thr Leu Ser Glu Val Ser Phe Ser Pro Phe Tyr Leu
180 185 190
Lys Pro Phe Asp Glu Ser Tyr Gly Asn Tyr Arg Ser Leu Val Asn Ile
195 200 205
Val Asn Lys Phe Val Lys Arg Glu Phe Ala Pro Asp Pro Ala Ser Thr
210 215 220
Leu Met His Gln Leu Val Tyr Val Thr His Asn Leu Tyr Gly Ile Ser
225 230 235 240
Asn Arg Asn Phe Tyr Tyr Asn Phe Asp Thr Gly Lys Ile Glu Thr Ser
245 250 255
Arg Gln Gln Asn Ser Leu Ile Phe Glu Glu Leu Leu Thr Phe Gly Gly
260 265 270
Ile Asp Ser Lys Ala Ile Ser Ser Leu Ile Ile Lys Lys Ile Ile Glu
275 280 285
Thr Ala Lys Asn Asn Tyr Thr Thr Leu Ile Ser Glu Arg Leu Asn Thr
290 295 300
Val Thr Val Glu Asn Asp Leu Leu Lys Tyr Ile Lys Asn Lys Ile Pro
305 310 315 320
Val Gln Gly Arg Leu Gly Asn Phe Lys Leu Asp Thr Ala Glu Phe Glu
325 330 335
Lys Lys Leu Asn Thr Ile Leu Phe Val Leu Asn Glu Ser Asn Leu Ala
340 345 350
Gln Arg Phe Ser Ile Leu Val Arg Lys His Tyr Leu Lys Glu Arg Pro
355 360 365
Ile Asp Pro Ile Tyr Val Asn Ile Leu Asp Asp Asn Ser Tyr Ser Thr
370 375 380
Leu Glu Gly Phe Asn Ile Ser Ser Gln Gly Ser Asn Asp Phe Gln Gly
385 390 395 400
Gln Leu Leu Glu Ser Ser Tyr Phe Glu Lys Ile Glu Ser Asn Ala Leu
405 410 415
Arg Ala Phe Ile Lys Ile Ala Pro Arg Asn Gly Leu Leu Tyr Asn Ala
420 425 430
Ile Tyr Arg Asn Ser Lys
435
<210> 77
<211> 8
<212> PRT
<213> 人工序列
<220>
<223> 前切割蛋白酶位点
<400> 77
Leu Glu Val Leu Phe Gln Gly Pro
1 5
<210> 78
<211> 25
<212> PRT
<213> 人工序列
<220>
<223> C1激活环变体2
<400> 78
Cys His Lys Ala Ile Asp Gly Arg Ser Leu Glu Val Leu Phe Gln Gly
1 5 10 15
Pro Leu Tyr Asn Lys Thr Leu Asp Cys
20 25

Claims (69)

1.用于治疗疼痛的多肽，其中该多肽包含梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)，其中当该多肽包含梭菌神经毒素L链时，该L链无催化活性。
2.用于治疗疼痛的方法，该方法包括对受试者施用多肽，其中该多肽包含梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)，其中当该多肽包含梭菌神经毒素L链时，该L链无催化活性。
3.多肽在制备用于治疗疼痛的药物中的用途，其中该多肽包含梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)，其中当该多肽包含梭菌神经毒素L链时，该L链无催化活性。
4.根据权利要求1所述的用途的多肽、根据权利要求2所述的方法或根据权利要求3所述的用途，其中该多肽不通过以下任一种治疗疼痛：不通过促进神经元生长、不通过促进神经元修复或不通过促进神经元生长和修复。
5.用于治疗炎性疾病的多肽，其中该多肽包含梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)，其中当该多肽包含梭菌神经毒素L链时，该L链无催化活性。
6.用于治疗炎性疾病的方法，该方法包括对受试者施用多肽，其中该多肽包含梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)，其中当该多肽包含梭菌神经毒素L链时，该L链无催化活性。
7.多肽在制备用于治疗炎性疾病的药物中的用途，其中该多肽包含梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)，其中当该多肽包含梭菌神经毒素L链时，该L链无催化活性。
8.根据权利要求5所述的用途的多肽、根据权利要求6所述的方法或根据权利要求7所述的用途，其中该多肽不通过以下任一种治疗炎性疾病：不通过促进神经元生长、不通过促进神经元修复或不通过促进神经元生长和修复。
9.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该多肽不包含进一步的催化活性结构域。
10.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该多肽不包含除梭菌神经毒素L链、HN结构域和/或HC结构域之外的治疗或诊断剂(例如共价或非共价结合的治疗或诊断剂)。
11.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该多肽不与另外的治疗剂或诊断剂一起施用(例如顺序地或随后地)。
12.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该多肽包含无催化活性梭菌神经毒素L链。
13.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该多肽包含梭菌神经毒素L链、HN结构域和HC结构域，其中该L链无催化活性。
14.根据权利要求1-12中任一项所述的用途的多肽、方法或用途，其中该多肽基本上由梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)组成，其中在该多肽包含梭菌神经毒素L链或基本上由其组成时，该L链无催化活性。
15.根据权利要求1-12或14中任一项所述的用途的多肽、方法或用途，其中该多肽基本上由梭菌神经毒素轻链(L链)和梭菌神经毒素转运结构域(HN结构域)组成，其中该L链无催化活性。
16.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该多肽基本上由梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和梭菌神经毒素受体结合结构域(HC结构域)组成，其中该L链无催化活性。
17.根据权利要求1-12或14中任一项所述的用途的多肽、方法或用途，其中该多肽由梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和/或梭菌神经毒素受体结合结构域(HC结构域)组成，其中在该多肽包含梭菌神经毒素L链或由其组成时，该L链无催化活性。
18.根据权利要求1-12、15或17中任一项所述的用途的多肽、方法或用途，其中该多肽由梭菌神经毒素轻链(L链)和梭菌神经毒素转运结构域(HN结构域)组成，其中该L链无催化活性。
19.根据权利要求1-13、14、16或17中任一项所述的用途的多肽、方法或用途，其中该多肽由梭菌神经毒素轻链(L链)、梭菌神经毒素转运结构域(HN结构域)和梭菌神经毒素受体结合结构域(HC结构域)组成，其中该L链无催化活性。
20.根据权利要求1-12、14、15、17或18中任一项所述的用途的多肽、方法或用途，其中该多肽不包含梭菌神经毒素HN结构域和HC结构域。
21.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该多肽不进一步包含非梭菌催化结构域。
22.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该疼痛是慢性疼痛。
23.根据权利要求1-21中任一项所述的用途的多肽、方法或用途，其中该疼痛是急性疼痛。
24.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该疼痛是炎性疼痛。
25.根据权利要求24所述的用途的多肽、方法或用途，其中该炎性疼痛由晒伤、UV诱导的损伤、关节炎引起的疾病、自身免疫性疾病、结缔组织疾病、损伤、感染、神经炎、关节炎症或头痛(优选肌肉/肌源性头痛、血管性头痛、高血压性头痛、激素性头痛、反弹性头痛、慢性鼻窦炎性头痛、器质性头痛或发作性头痛)引起或与之相关。
26.根据权利要求1-23中任一项所述的用途的多肽、方法或用途，其中该疼痛是神经病性疼痛。
27.根据权利要求26所述的用途的多肽、方法或用途，其中该神经病性疼痛是神经痛、传入神经阻滞、复杂性区域疼痛综合征(CRPS)或神经病(例如中枢或外周神经病)(或由其引起或与之相关)。
28.根据权利要求1-23中任一项所述的用途的多肽、方法或用途，其中该疼痛是混合性疼痛。
29.根据权利要求1-23中任一项所述的用途的多肽、方法或用途，其中该疼痛是异常性疼痛。
30.根据权利要求1-23中任一项所述的用途的多肽、方法或用途，其中该疼痛是内脏疼痛。
31.根据权利要求30所述的用途的多肽、方法或用途，其中该内脏疼痛是功能性内脏疼痛、慢性胃肠道炎症、自身免疫性疼痛、器质性内脏疼痛或治疗诱导的内脏疼痛(或由其引起或与之相关)。
32.根据权利要求1-23中任一项所述的用途的多肽、方法或用途，其中该疼痛是头痛(例如偏头痛)。
33.根据权利要求32所述的用途的多肽、方法或用途，其中该疼痛是偏头痛。
34.根据权利要求32所述的用途的多肽、方法或用途，其中该头痛由肌肉/肌源性头痛、血管性头痛、高血压性头痛、激素性头痛、反弹性头痛、慢性鼻窦炎性头痛、器质性头痛或发作性头痛引起或与之相关。
35.根据权利要求1-23中任一项所述的用途的多肽、方法或用途，其中该疼痛是术后疼痛。
36.根据权利要求1-23中任一项所述的用途的多肽、方法或用途，其中该疼痛是牵涉性疼痛。
37.根据权利要求1-23中任一项所述的用途的多肽、方法或用途，其中该疼痛是躯体疼痛。
38.根据权利要求37所述的用途的多肽、方法或用途，其中该疼痛是由过度肌肉张力、重复运动障碍、肌肉障碍、肌痛、感染或药物引起或与之相关的躯体疼痛。
39.根据权利要求1-23中任一项所述的用途的多肽、方法或用途，其中该疼痛是膀胱疼痛综合征，优选其中该膀胱疼痛由间质性膀胱炎引起或与之相关。
40.根据权利要求1-23中任一项所述的用途的多肽、方法或用途，其中该疼痛是幻肢疼痛。
41.根据权利要求5-21中任一项所述的用途的多肽、方法或用途，其中该炎性疾病是选自膀胱炎、子宫内膜异位症、类风湿性关节炎、复杂性区域疼痛综合征和神经炎的一种或多种。
42.根据权利要求41所述的用途的多肽、方法或用途，其中该膀胱炎是间质性膀胱炎。
43.根据权利要求41所述的用途的多肽、方法或用途，其中该神经炎是外周神经炎。
44.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中所施用的该多肽的单个剂量大于250μg。
45.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中所施用的该多肽的单个剂量为251μg至10g。
46.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中所施用的该多肽的单个剂量为251μg至1g。
47.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中所施用的该多肽的单个剂量为251-1000μg。
48.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该多肽反复施用(例如作为疼痛治疗方案的一部分)。
49.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该多肽皮内施用。
50.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该多肽包含与SEQ ID NO:2、4、6、8、10、12、14、16、18、20、22、24、26、28、30、32、34、36、38、40、42、44、46、48、50、51、52、53、54、55、56、57、58、59、60、61、62、63、64、65、66、67、68、69、70、74、75或76中的任一个具有至少70％序列同一性的多肽序列，条件是在该多肽包含梭菌神经毒素L链时，该L链无催化活性。
51.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该多肽包含与SEQ ID NO:2、4、6、8、10、12、14、16、18、20、22、24、26、28、30、32、34、36、38、40、42、44、46、48、50、51、52、53、54、55、56、57、58、59、60、61、62、63、64、65、66、67、68、69、70、74、75或76中的任一个具有至少80％序列同一性的多肽序列，条件是在该多肽包含梭菌神经毒素L链时，该L链无催化活性。
52.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该多肽包含与SEQ ID NO:2、4、6、8、10、12、14、16、18、20、22、24、26、28、30、32、34、36、38、40、42、44、46、48、50、51、52、53、54、55、56、57、58、59、60、61、62、63、64、65、66、67、68、69、70、74、75或76中的任一个具有至少90％序列同一性的多肽序列，条件是在该多肽包含梭菌神经毒素L链时，该L链无催化活性。
53.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该多肽包含SEQID NO:2、4、6、8、10、12、14、16、18、20、22、24、26、28、30、32、34、36、38、40、42、44、46、48、50、51、52、53、54、55、56、57、58、59、60、61、62、63、64、65、66、67、68、69、70、74、75或76中任一个的多肽序列。
54.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该多肽是无催化活性BoNT/A。
55.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该多肽是经修饰的梭菌神经毒素，如嵌合梭菌神经毒素或杂合梭菌神经毒素，优选其中该多肽不包含天然梭菌神经毒素H链。
56.根据权利要求1-53或55中任一项所述的用途的多肽、方法或用途，其中该多肽缺乏梭菌神经毒素的功能性HCC结构域或HC结构域。
57.根据权利要求1-53或55-56中任一项所述的用途的多肽、方法或用途，其中该多肽是包含非梭菌靶向部分(TM)的重靶向梭菌神经毒素。
58.根据权利要求1-53或55-56中任一项所述的用途的多肽、方法或用途，其中该多肽缺乏梭菌神经毒素的功能性HC结构域，并且还缺乏任何功能等同的外源性配体靶向部分(TM)。
59.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该多肽不在该个体的细胞中表达，例如其中该用途或方法不包括在该受试者的细胞中表达编码该多肽的核酸。
60.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该多肽进一步包含一个或多个非梭菌神经毒素序列。
61.根据权利要求60所述的用途的多肽、方法或用途，其中该一个或多个非梭菌神经毒素序列不与细胞受体结合。
62.根据权利要求60或61所述的用途的多肽、方法或用途，其中该一个或多个非梭菌神经毒素序列不包含细胞受体的配体。
63.根据权利要求1-53、55或59-62中任一项所述的用途的多肽、方法或用途，其中该多肽是嵌合肉毒神经毒素(BoNT)，其包含无催化活性BoNT/A轻链以及转运结构域和BoNT/B受体结合结构域(HC结构域)。
64.根据权利要求1-55或59-62中任一项所述的用途的多肽、方法或用途，其中该多肽包含经修饰的BoNT/A HC结构域，该结构域在选自以下的一个或多个氨基酸残基处包含修饰：ASN 886、ASN 905、GLN 915、ASN 918、GLU 920、ASN 930、ASN 954、SER 955、GLN 991、GLU 992、GLN 995、ASN 1006、ASN 1025、ASN 1026、ASN 1032、ASN 1043、ASN 1046、ASN1052、ASP 1058、HIS1064、ASN 1080、GLU 1081、GLU 1083、ASP 1086、ASN 1188、ASP 1213、GLY 1215、ASN 1216、GLN 1229、ASN 1242、ASN 1243、SER 1274和THR 1277，其中该修饰选自：
i.用碱性氨基酸残基取代表面暴露的酸性氨基酸残基；
ii.用不带电荷的氨基酸残基取代表面暴露的酸性氨基酸残基；
iii.用碱性氨基酸残基取代表面暴露的不带电荷的氨基酸残基；
iv.插入碱性氨基酸残基；和
v.缺失表面暴露的酸性氨基酸残基。
65.根据权利要求1-53或55-62中任一项所述的用途的多肽、方法或用途，其中该多肽包含无催化活性肉毒神经毒素血清型X(BoNT/X)L链、BoNT/X HN结构域和/或BoNT/X HC结构域。
66.根据权利要求1-53、59-62或65中任一项所述的用途的多肽、方法或用途，其中该多肽是嵌合肉毒神经毒素(BoNT)，其包含无催化活性BoNT/X轻链和转运结构域，以及来自不同(即，非BoNT/X)梭菌神经毒素的受体结合结构域(HC结构域)。
67.根据权利要求1-53、59-62或65-66中任一项所述的用途的多肽、方法或用途，其中该多肽是嵌合肉毒神经毒素(BoNT)，其包含无催化活性BoNT/X轻链和转运结构域以及BoNT/B受体结合结构域(HC结构域)。
68.根据权利要求65-67中任一项所述的用途的多肽、方法或用途，其中该疼痛是炎性疼痛。
69.根据前述权利要求中任一项所述的用途的多肽、方法或用途，其中该多肽包含Cys-(Xaa)a-Ile-Asp/Glu-Gly-Arg-(Yaa)b-Cys(SEQ ID NO:71)，其中a＝1-10，且b＝4-15。
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