CN115720581A - 冠状病毒疫苗配制品 - Google Patents
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Abstract
本文公开了适用于在疫苗中使用的冠状病毒纤突(S)蛋白和包含所述蛋白的纳米颗粒。所述纳米颗粒呈递来自包围洗涤剂核心并与之缔合的病原体的抗原，从而产生增强的稳定性和良好的免疫原性。还公开了所述疫苗和纳米颗粒的剂量、配制品和用于制备所述疫苗和纳米颗粒的方法。
Description
相关申请的交叉引用
本申请要求以下申请的优先权，将其中的每个申请出于所有目的通过引用以其整体而并入：2020年1月27日提交的美国临时申请号62/966,271；2020年2月14日提交的美国临时申请号62/976,858；2020年2月28日提交的美国临时申请号62/983,180；2020年7月7日提交的美国临时申请号63/048,945；2020年7月14日提交的美国临时申请号63/051,706；2020年7月20日提交的美国临时申请号63/054,182；2020年12月22日提交的美国临时申请号63/129,392；以及2020年8月19日提交的美国非临时申请号16/997,001。
电子提交文本文件的说明
将以电子方式随同提交的文本文件的内容通过引用以其整体并入本文：计算机可读格式拷贝的序列表(文件名：NOVV_088_08WO_SeqList_ST25.txt，记录日期：2021年1月26日，文件大小：577千字节)。
技术领域
本公开文本总体上涉及非天然存在的冠状病毒(CoV)纤突(S)多肽以及包含所述多肽的纳米颗粒和疫苗，其可用于刺激免疫反应。所述纳米颗粒提供了任选地与洗涤剂核心缔合的抗原(例如，糖蛋白抗原)，并且通常使用重组方法产生。所述纳米颗粒具有改善的稳定性和增强的表位呈递。本公开文本还提供了含有所述纳米颗粒的组合物、用于产生所述组合物的方法以及刺激免疫反应的方法。
背景技术
感染性疾病仍然是遍及全世界的问题。尽管在开发针对一些病原体的疫苗方面已取得进展，但许多病原体仍对人类健康构成威胁。最近，SARS-CoV-2冠状病毒已蔓延到美国、泰国、韩国、中国台湾和日本。SARS-CoV-2冠状病毒与在过去17年中已导致数百人死亡的严重急性呼吸系统综合征冠状病毒(SARS-CoV)和中东呼吸系统综合征冠状病毒(MERS-CoV)属于同一病毒家族。SARS-CoV-2导致疾病COVID-19。
期望开发预防威胁生命的感染性疾病(像SARS-CoV-2冠状病毒)或减轻其严重性的疫苗。然而，由于病原体的逃避机制非常复杂并且难以稳定疫苗，因此人类疫苗的开发仍然具有挑战性。最佳地，疫苗必须同时诱导阻断或中和感染原的抗体，并且在各种环境(包括不能制冷的环境)中保持稳定。
发明内容
本公开文本提供了适用于诱导针对SARS-CoV-2(也称为CoV和2019-nCoV)的免疫反应的非天然存在的CoV S多肽。本公开文本还提供了含有所述糖蛋白的纳米颗粒以及刺激免疫反应的方法。
本公开文本还提供了适用于诱导针对多种冠状病毒的免疫反应的CoV S多肽，所述冠状病毒包括SARS-CoV-2、中东呼吸系统综合征(MERS)和严重急性呼吸系统综合征(SARS)。
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所述专利或申请文件含有至少一张彩色附图。在请求并支付必要的费用后，官方将会提供带有彩色附图的本专利或专利申请公开案的副本。
图1示出了SARS-CoV-2纤突(S)蛋白的野生型氨基酸序列(SEQ ID NO:1)的示意图。弗林蛋白酶切割位点RRAR(SEQ ID NO:6)以粗体突出显示，并且信号肽加下划线。
图2示出了SARS-CoV-2S多肽的一级结构，其具有无活性弗林蛋白酶切割位点、融合肽缺失以及K986P和V987P突变。结构域位置相对于来自含有信号肽的SARS-CoV-2的野生型CoV S多肽的氨基酸序列(SEQ ID NO:1)来编号。
图3示出了BV2378 CoV S多肽的一级结构，其具有无活性弗林蛋白酶切割位点、氨基酸819-828的融合肽缺失以及K986P和V987P突变。结构域位置相对于来自含有信号肽的SARS-CoV-2的野生型CoV S多肽的氨基酸序列(SEQ ID NO:1)来编号。
图4示出了CoV S多肽BV2364、BV2365、BV2366、BV2367、BV2368、BV2369、BV2373、BV2374和BV2375的纯化。数据揭示具有含有QQAQ的氨基酸序列(SEQ ID NO:7)的无活性弗林蛋白酶切割位点的BV2365(SEQ ID NO:4)和BV2373(SEQ ID NO:87)被表达为单链(S0)。相比之下，含有完整弗林蛋白酶切割位点的CoV S多肽(例如BV2364、BV2366和BV2374)被切割，如通过切割产物S2的存在所证实的。
图5通过生物膜层干涉测量术显示，CoV S多肽BV2361、BV2365、BV2369、BV2365、BV2373和BV2374与人血管紧张素转化酶2前体(hACE2)结合。
图6通过生物膜层干涉测量术显示，来自SARS-CoV-2的BV2361不结合MERS-CoV受体二肽基肽酶IV(DPP4)，并且MERS S蛋白不与人血管紧张素转化酶2前体(hACE2)结合。
图7通过酶联免疫吸附测定(ELISA)显示，BV2361与hACE2结合。
图8示出了BV2373 CoV S多肽的一级结构和对弗林蛋白酶切割位点的修饰K986P和V987P。
图9示出了野生型CoV S多肽以及CoV S多肽BV2365和BV2373的纯化。
图10示出了覆盖在SARS-CoV-2纤突蛋白(EMB ID：21374)的冷冻电子显微术(cryoEM)结构上的BV2373 CoV S多肽的cryoEM结构。
图11A-图11F显示CoV S纤突多肽BV2365和BV2373与hACE2结合。生物膜层干涉测量术揭示，BV2365(图11B)和BV2373(图11C)以与野生型CoV S多肽(图11A)类似的解离动力学与hACE2结合。ELISA表明，野生型CoV S多肽(图11D)和BV2365(图11E)以类似的亲和力与hACE2结合，而BV2373以更高的亲和力与hACE2结合(图11F)。
图12A-图12B示出了胁迫条件(诸如温度、两个冷冻/解冻循环、氧化、搅拌和pH极端值)对CoV S多肽BV2373(图12A)和BV2365(图12B)与hACE2的结合的影响。
图13A-图13B示出了在用两剂(图13A)和一剂(图13B)的0.1μg至10μg的BV2373、加或不加级分A和级分C iscom基质(例如，MATRIX-MTM)免疫小鼠后13天、21天和28天的抗CoVS多肽IgG滴度。
图14示出了在用一剂或两剂的0.1μg至10μg的BV2373、加或不加MATRIX-MTM免疫的小鼠中阻断hACE2相互作用的抗体的诱导。
图15示出了在用一剂或两剂的0.1μg至10μg的BV2373、加或不加MATRIX-MTM免疫的小鼠中检测到的病毒中和抗体。
图16示出了用单剂的BV2373或两剂的BV2373间隔14天、加或不加MATRIX-MTM免疫的Ad/CMV/hACE2小鼠的肺中的病毒载量(SARS-CoV-2)。
图17A-图17C示出了用BV2373免疫后小鼠表现出的体重减轻。图17A示出了用单一0.01μg、0.1μg、1μg或10μg的BV2373+MATRIX-MTM免疫对体重减轻的影响。图17B示出了用两剂的BV2373(0.01μg、0.1μg、1μg)+MATRIX-MTM免疫对体重减轻的影响。图17C示出了在存在或不存在MATRIX-MTM的情况下用两剂的BV2373(10μg)免疫对体重减轻的影响。
图18A-图18B示出了BV2373在感染SARS-CoV-2后四天(图18A)或七天(图18B)对小鼠肺组织病理学的影响。
图19示出了与在存在MATRIX-MTM的情况下用BV2373免疫的小鼠相比，在不存在佐剂的情况下用BV2373免疫的小鼠的脾脏中离体刺激后IFN-γ分泌性细胞的数量。
图20A-图20E示出了在存在或不存在MATRIX-MTM的情况下用BV2373免疫的小鼠的脾脏中细胞因子分泌性CD4+T细胞的频率。图20A示出了IFN-γ分泌性CD4+T细胞的频率。图20B示出了TNF-α分泌性CD4+T细胞的频率。图20C示出了IL-2分泌性CD4+T细胞的频率。图20D示出了分泌选自IFN-γ、TNF-α和IL-2的两种细胞因子的CD4+T细胞的频率。图20E示出了表达IFN-γ、TNF-α和IL-2的CD4+T细胞的频率。
图21A-图21E示出了在存在或不存在MATRIX-MTM的情况下用BV2373免疫的小鼠的脾脏中细胞因子分泌性CD8+T细胞的频率。图21A示出了IFN-γ分泌性CD8+T细胞的频率。图21B示出了TNF-α分泌性CD8+T细胞的频率。图21C示出了IL-2分泌性CD8+T细胞的频率。图20D示出了分泌选自IFN-γ、TNF-α和IL-2的两种细胞因子的CD8+T细胞的频率。图21E示出了表达IFN-γ、TNF-α和IL-2的CD8+T细胞的频率。
图22展示了在存在或不存在MATRIX-MTM的情况下用BV2373免疫的小鼠的脾脏中表达选自IFN-γ、TNF-α和IL-2的一种(单一)、两种(双重)或三种(三重)细胞因子的CD4+或CD8+细胞的频率。
图23A-图23C展示了在存在或不存在MATRIX-MTM的情况下用BV2373免疫对CD4+T细胞分泌2型细胞因子的影响。图23A示出了IL-4分泌性细胞的频率。图23B示出了IL-5分泌性CD4+细胞的频率。图23C示出了IFN-γ分泌性与IL-4分泌性CD4+T细胞的比率。
图24A-图24B通过评估CD4+滤泡辅助性T细胞(TFH)的存在展示了在存在或不存在MATRIX-MTM的情况下用BV2373免疫小鼠对生发中心形成的影响。图24A示出了脾脏中CD4+滤泡辅助性T细胞的频率，并且图24B示出了CD4+滤泡辅助性T细胞的表型(例如CD4+CXCR5+PD-1+)。
图25A-图25B通过评估生发中心(GC)B细胞的存在展示了在存在或不存在MATRIX-MTM的情况下用BV2373免疫小鼠对生发中心形成的影响。图25A示出了脾脏中GC B细胞的频率，并且图25B揭示了CD4+滤泡辅助性T细胞的表型(例如CD19+GL7+CD-95+)。
图26A-图26C示出了在存在或不存在MATRIX-MTM的情况下用BV2373免疫对橄榄狒狒的抗体反应的影响。图26A示出了用BV2373免疫后狒狒中的抗SARS-CoV-2S多肽IgG滴度。图26B示出了在存在MATRIX-MTM的情况下用5μg或25μg的BV2373单次免疫后在狒狒中hACE2受体阻断抗体的存在。图26C示出了用BV2373和MATRIX-MTM单次免疫后病毒中和抗体的滴度。
图27示出了用BV2373免疫后橄榄狒狒中抗SARS-CoV-2S多肽IgG与中和抗体滴度之间的显著相关性。
图28示出了在存在或不存在MATRIX-MTM的情况下用BV2373免疫的橄榄狒狒的外周血单核细胞(PBMC)中IFN-γ分泌性细胞的频率。
图29A-图29E示出了在存在或不存在MATRIX-MTM的情况下用BV2373免疫的橄榄狒狒的PBMC中细胞因子分泌性CD4+T细胞的频率。图29A示出了IFN-γ分泌性CD4+T细胞的频率。图29B示出了IL-2分泌性CD4+T细胞的频率。图29C示出了TNF-α分泌性CD4+T细胞的频率。图29D示出了分泌选自IFN-γ、TNF-α和IL-2的两种细胞因子的CD4+T细胞的频率。图29E示出了表达IFN-γ、TNF-α和IL-2的CD4+T细胞的频率。
图30示出了冠状病毒纤突(S)蛋白(SEQ ID NO:109)(BV2384)的示意图。弗林蛋白酶切割位点GSAS(SEQ ID NO:97)加单下划线，并且K986P和V987P突变加双下划线。
图31示出了冠状病毒纤突(S)蛋白(SEQ ID NO:86)(BV2373)的示意图。弗林蛋白酶切割位点QQAQ(SEQ ID NO:7)加单下划线，并且K986P和V987P突变加双下划线。
图32示出了CoV S多肽BV2373(SEQ ID NO:87)和BV2384(SEQ ID NO:109)的纯化。
图33示出了纯化后BV2384(SEQ ID NO:109)纯度的扫描光密度测定图。
图34示出了纯化后BV2373(SEQ ID NO:87)纯度的扫描光密度测定图。
图35A-图35B展示了响应于BV2373和MATRIX-MTM的施用抗S抗体(图35A)和中和抗体(图35B)的诱导。向食蟹猴施用一剂或两剂(第0天和第21天)的2.5μg、5μg或25μg的BV2373加25μg或50μg MATRIX-MTM佐剂。对照既未接受BV2373又未接受MATRIX-MTM。在第21天和第33天测量抗体。
图36A-图36B展示了本文所公开的疫苗配制品对SARS-CoV-2病毒复制的减少，如在食蟹猴的支气管肺泡灌洗液(BAL)中所评估的。如图所示，向食蟹猴施用BV2373和MATRIX-MTM。受试者在第0天进行免疫，并且在用两剂的组中在第0天和第21天进行免疫。第37天用1x104 pfu SARS-CoV-2病毒攻击受试动物。在用感染性病毒攻击后2天和4天(d2pi和d4pi)，在支气管灌洗液(BAL)中评估病毒RNA(图36A，对应于存在的总RNA)和病毒亚基因组RNA(图36B，对应于复制病毒)水平。大多数受试者未显示出病毒RNA。在第2天，在一些受试者中测量到少量RNA。到第4天，除了接受2.5μg最低剂量的两名受试者以外，都未测量到RNA。除了同样接受最低剂量的1名受试者以外，在2天或4天均未检测到亚基因组RNA。
图37A-图37B展示了本文所公开的疫苗配制品对SARS-CoV-2病毒复制的减少，如在食蟹猴的鼻拭子中所评估的。如图所示，向食蟹猴施用BV2373加MATRIX-MTM。受试者在第0天进行免疫，并且在用两剂的组中在第0天和第21天进行免疫。第37天用1x104SARS-CoV-2病毒攻击受试动物。在感染后2天和4天(d2pi和d4pi)通过鼻拭子评估病毒RNA(图37A)和病毒亚基因组(sg)RNA(图37B)。大多数受试者未显示出病毒RNA。在第2天和第4天，在一些受试者中测量到少量RNA。在2天或4天均未检测到亚基因组RNA。受试者在第0天进行免疫，并且在用两剂的组中在第0天和第21天进行免疫。这些数据表明，疫苗将鼻总病毒RNA降低100-1000倍，并将sgRNA降低至不可检测的水平，并且确认对疫苗的免疫反应将阻断病毒复制并防止病毒传播。
图38A-图38B示出了用一剂(图38A)或两剂(图38B)的BV2373和25μg或50μg的MATRIX-MTM免疫食蟹猴后21天和35天的抗CoV S多肽IgG滴度。
图38C-图38D示出了用一剂(图38C)或两剂(图38D)的BV2373(5μg)和MATRIX-MTM(25μg或50μg)免疫食蟹猴后21天和35天食蟹猴的hACE2抑制滴度。
图38E示出了在施用BV2373和MATRIX-MTM后在食蟹猴中抗CoV S多肽IgG滴度与hACE2抑制滴度之间的显著相关性。示出了表4的第2-6组的数据。
图39示出了在用两剂的BV2373和MATRIX-MTM免疫后或在用表4的恢复期人血清(第2、4和6组)免疫后35天食蟹猴的抗CoV S多肽滴度和hACE2抑制滴度。这些数据表明，用BV2373和MATRIX-MTM免疫的食蟹猴的抗CoV S多肽和hACE2抑制滴度优于用恢复期血清免疫的食蟹猴。
图40A-图40B示出了用BV2373和MATRIX-MTM免疫的食蟹猴的SARS-CoV-2中和滴度，如通过致细胞病变效应(CPE)(图40A)和噬斑减少中和测试(PRNT)(图40B)所确定的。
图41示出了评价包含BV2373和任选的MATRIX-MTM的疫苗的安全性和功效的临床试验的施用时间。AESI表示特别关注的不良事件。MAEE表示医疗护理不良事件，并且SAE表示严重不良事件。
图42A-图42B示出了在评价包含BV2373和MATRIX-MTM的疫苗的临床试验中患者经历的局部(图42A)和全身(图42B)不良事件。在表5中标识了组A-E。数据表明，疫苗具有良好的耐受性和安全性。
图43A-图43B示出了在评价包含BV2373和MATRIX-MTM的疫苗的临床试验中免疫参与者后21天和35天的抗CoV S多肽IgG(图43A)和中和(图43B)滴度。水平条分别表示四分位距(IRQ)和中值曲线下面积。须端点等于低于或高于中值±1.5倍IQR的最大值和最小值。恢复期血清图包括来自美国贝勒医学院(Baylor College of Medicine)的PCR确认的COVID-19参与者的样本(29个样本用于ELISA且32个样本用于微量中和(MN IC>99))。COVID-19的严重性表示为住院治疗患者的红色标记(包括重症监护环境)、门诊治疗患者的蓝色标记(在急诊科收集的样品)和无症状(暴露)患者的绿色标记(从接触/暴露评估中收集的样品)。
图44A-图44C示出了在施用恢复期血清(图44A)、两剂25μg的BV2373(图44B)和两剂(5μg和25μg)的BV2373加MATRIX-MTM(图44C)的患者中抗CoV S多肽IgG与中和抗体滴度之间的相关性。在用恢复期血清或加佐剂的BV2373治疗的患者中在中和抗体滴度与抗CoV-SIgG滴度之间观察到很强的相关性，而在不存在佐剂的情况下用BV2373治疗的患者中并未观察到。
图45A-图45D示出了在A组(安慰剂，图45A)、B组(25μg BV2373，图45B)、C组(5μgBV2373和50μg MATRIX-MTM，图45C)和D组(25μg BV2373和50μg MATRIX-MTM，图45D)中在用BV2373刺激后参与者的产生T辅助1(Th1)细胞因子干扰素-γ(IFN-γ)、肿瘤坏死因子-α(TNF-α)以及白介素(IL)-2和T辅助2(Th2)细胞因子IL-5和IL-13指示的细胞因子的抗原特异性CD4+T细胞的频率。Th1细胞因子图中的“任何2”意指可以同时产生两种类型的Th1细胞因子的CD4+T细胞。“全部3”指示同时产生IFN-γ、TNF-α和IL-2的CD4+T细胞。Th2图中的“两者”意指可以同时产生Th2细胞因子IL-5和IL-13的CD4+T细胞。
图46A示出了含有信号肽的野生型SARS-CoV-2S多肽的一级结构，其相对于SEQ IDNO:1来编号。图46B示出了没有信号肽的野生型SARS-CoV-2S多肽的一级结构，其相对于SEQID NO:2来编号。
具体实施方式
定义
如本文和所附权利要求所用，单数形式“一个/一种(a、an)”和“所述(the)”包括复数指代物，除非上下文另外明确地规定。因此，例如，提及“一种蛋白质”可以指一种蛋白质或这种蛋白质的混合物，并且提及“所述方法”包括提及本领域技术人员已知的等同步骤和/或方法，等等。
如本文所用，术语“佐剂”是指当与免疫原组合使用时增加或以其他方式改变或修饰针对所述免疫原诱导的免疫反应的化合物。免疫反应的修饰可以包括强化或扩大抗体免疫反应和细胞免疫反应中任一者或两者的特异性。
如本文所用，当在数值之前时，术语“约”或“大约”指示所述值加或减10％的范围。例如，“约100”包括90和110。
如本文所用，术语“免疫原”、“抗原”和“表位”是指能够引发免疫反应的物质，诸如蛋白质(包括糖蛋白)和肽。
如本文所用，“免疫原性组合物”是包含抗原的组合物，其中向受试者施用所述组合物导致在所述受试者中产生针对所述抗原的体液免疫反应和/或细胞免疫反应。
如本文所用，“亚单位”组合物(例如疫苗)包括一种或多种选定抗原，但不是来自病原体的所有抗原。这样的组合物基本上不含完整的病毒或此类细胞或颗粒的裂解物，并且通常由至少部分纯化的(通常基本上纯化的)来自所述病原体的免疫原性多肽制备。通常使用杆状病毒系统，通常以重组方式制备本文所公开的亚单位组合物中的抗原。
如本文所用，“基本上”是指这样分离物质(例如化合物、多核苷酸或多肽)，使得所述物质形成包含其的样品的大多数百分比。例如，在样品中，基本上纯化的组分构成所述样品的85％，优选85％-90％，更优选至少95％-99.5％，最优选至少99％。如果组分基本上被替代，则在样品中剩余的量小于或等于约0.5％至约10％，优选小于约0.5％至约1.0％。
如本文所用，术语“治疗”(“treat”、“treatment”.
和“treating”)是指用于获得有益或希望的结果(例如，临床结果)的方法。出于本公开文本的目的，有益或希望的结果可以包括抑制或压制感染或疾病的开始或进展；改善感染或疾病的症状或减轻其发展；或其组合。
如本文所用，“预防”(“prevention”)与“防预”(“prophylaxis”)可互换使用，并且可以意指完全预防感染或疾病，或预防这种感染或疾病的症状的发展；延迟感染或疾病或其症状的发作；或降低随后发展的感染或疾病或其症状的严重性。
如本文所用，“有效剂量”或“有效量”是指免疫原的这样的量，所述量足以诱导减轻病原体感染的至少一种症状的免疫反应。可以例如通过测量中和分泌抗体和/或血清抗体的量，例如通过噬斑中和、补体结合、酶联免疫吸附(ELISA)或微量中和测定来确定有效剂量或有效量。
如本文所用，术语“疫苗”是指用于诱导针对病原体的免疫反应的免疫原性组合物，诸如源自所述病原体的免疫原，所述免疫反应提供保护性免疫力(例如，保护受试者免于所述病原体感染和/或降低由所述病原体感染引起的疾病或病症的严重性的免疫力)。保护性免疫反应可以包括抗体和/或细胞介导的反应的形成。取决于上下文，术语“疫苗”也可以指施用至受试者以产生保护性免疫力的免疫原悬浮液或溶液。
如本文所用，术语“受试者”包括人和其他动物。通常，受试者是人。例如，受试者可以是成人、青少年、儿童(2岁至14岁)、婴儿(出生至2岁)或新生儿(多至2个月)。在特定方面，受试者多至4个月大或多至6个月大。在一些方面，成人是约65岁或更大或约60岁或更大的年长者。在一些方面，受试者是孕妇或打算怀孕的妇女。在其他方面，受试者不是人；例如非人灵长类动物；例如，狒狒、黑猩猩、大猩猩或猕猴。在某些方面，受试者可以是宠物，诸如狗或猫。
如本文所用，术语“药学上可接受的”意指由美国联邦政府或州政府的监管机构批准或在美国药典、欧洲药典或其他公认药典中列出用于哺乳动物，更特别地用于人。这些组合物可以用作用于在脊椎动物中诱导保护性免疫反应的疫苗和/或抗原组合物。
如本文所用，术语“约”意指加或减所指示数值的10％。
如本文所用，术语“NVX-CoV2373”是指包含BV2373纤突糖蛋白(SEQ ID NO:87)以及级分A和级分C iscom基质(例如，MATRIX-MTM)的疫苗组合物。
如本文所用，术语“修饰”当提及CoV S多肽时是指CoV S多肽的一个或多个氨基酸的突变、缺失或添加。CoV S多肽内修饰的位置可以基于将多肽的序列与SEQ ID NO:1(含有信号肽的CoV S多肽)或SEQ ID NO:2(缺少信号肽的成熟CoV S多肽)比对来确定。
含有冠状病毒(CoV)纤突(S)蛋白的疫苗组合物
本公开文本提供了非天然存在的冠状病毒(CoV)纤突(S)多肽、含有CoV S多肽的纳米颗粒以及含有非天然存在的CoV S多肽或含有CoV S多肽的纳米颗粒的免疫原性组合物和疫苗组合物。在实施方案中，本文提供了使用CoV S多肽、纳米颗粒、免疫原性组合物和疫苗组合物来刺激免疫反应的方法。
本文还提供了制造所述纳米颗粒和疫苗组合物的方法。有利地，所述方法提供了基本上没有被其他蛋白质污染的纳米颗粒，所述其他蛋白质是诸如与蛋白质在昆虫细胞中的重组表达相关的蛋白质。在实施方案中，表达发生在杆状病毒/Sf9系统中。
CoV S多肽抗原
本公开文本的疫苗组合物含有非天然存在的CoV S多肽。CoV S多肽可以源自冠状病毒，包括但不限于SARS-CoV-2，例如源自SARS-CoV-2、源自MERS CoV和源自SARS CoV。在实施方案中，所述CoV S多肽源自SARS-CoV-2的变体。在实施方案中，SARS-CoV-2的变体是SARS-CoV-2VUI 202012/01、B.1.1.7、501Y.V2、Cal.20C或P.1。与SARS-CoV S相比，SARS-CoV-2S蛋白在S1/S2切割位点中具有四个氨基酸的插入，从而产生多碱性RRAR弗林蛋白酶样切割基序。SARS-CoV-2S蛋白被合成为无活性前体(S0)，其在弗林蛋白酶切割位点处被蛋白水解切割成S1和S2亚基，这些亚基保持非共价连接以形成融合前三聚体。SARS-CoV-2S蛋白的S2结构域包含融合肽(FP)、两个七肽重复区(HR1和HR2)、跨膜(TM)结构域和胞质尾(CT)。SARS-CoV-2S蛋白的S1结构域折叠成四个不同的结构域：N末端结构域(NTD)和含有受体结合结构域(RBD)的C末端结构域、以及两个亚结构域SD1和SD2。融合前SARS-CoV-2S蛋白三聚体在S蛋白受体结合和切割后经历从融合前到融合后构象的结构重排。
在实施方案中，由于翻译后糖基化，所述CoV S多肽是糖蛋白。所述糖蛋白包含信号肽、S1亚基、S2亚基、NTD、RBD、两个亚结构域(SD1和SD2，在图46A-图46B中标记为SD1/2并且在本文中称为“SD1/2”)、完整或修饰的融合肽，HR1结构域、HR2结构域、TM和CD中的一种或多种。在实施方案中，每个结构域的氨基酸在图2和图46A(根据SEQ ID NO:1示出)、图46B(根据SEQ ID NO:2示出)和图3(对应于SEQ ID NO:1示出)中给出。在实施方案中，每个结构域可以与如在SEQ ID NO:1或SEQ ID NO:2中的每个结构域的序列具有至少95％、至少96％、至少97％、至少98％、至少99％或至少99.5％同一性。每个结构域可以具有与SEQ IDNO:1或SEQ ID NO:2中所示的那些相比多至约1、多至约2、多至约3、多至约4、多至约5、多至约10、多至约20或多至约30个氨基酸的缺失、插入或突变。每个结构域可以具有与SEQ IDNO:1或SEQ ID NO:2中所示的那些相比在约1与约5个之间的氨基酸、在约3与约10个之间的氨基酸、在约5与10个之间的氨基酸、在约8与12个之间的氨基酸、在约10与15个之间的氨基酸、在约12与17个之间的氨基酸、在约15与20个之间的氨基酸、在约18与23个之间的氨基酸、在约20与25个之间的氨基酸、在约22与约27个之间的氨基酸或在约25与30个之间的氨基酸的缺失、插入或突变。注意，图2和图3展示了成熟肽中不存在的13个氨基酸的N末端信号肽。所述CoV S多肽可以用于刺激针对天然CoV纤突(S)多肽的免疫反应。
在实施方案中，修饰天然CoV纤突(S)多肽(SEQ ID NO:2)，从而产生非天然存在的CoV纤突(S)多肽(图1)。在实施方案中，所述CoV纤突(S)糖蛋白包含S1亚基和S2亚基，其中所述S1亚基包含NTD、RBD、SD1/2和无活性弗林蛋白酶切割位点(氨基酸669-672)，并且其中所述S2亚基包含氨基酸973和974的突变；
其中所述NTD(氨基酸1-318)任选地包含选自以下的一个或多个修饰：
(a)一个或多个氨基酸从所述NTD中的缺失，任选地其中所述一个或多个修饰选自氨基酸56、57、131、132、229、230、231及其组合；以及
(b)选自氨基酸67、229、202、139、5、233、7、13、125、177或其组合的一个或多个氨基酸的突变；
其中所述RBD任选地包含选自氨基酸488、404、471、439、426、440及其组合的一个或多个氨基酸的突变；
其中所述SD1/2任选地包含选自601、557、668、642及其组合的一个或多个氨基酸的突变；并且
其中所述S2亚基任选地包含选自以下的一个或多个修饰：
(a)来自676-702、702-711、775-793的一个或多个氨基酸的缺失；
(b)一个或多个氨基酸从融合肽(氨基酸806-815)中的缺失；
(c)选自973、974、703、1105、688、969和1014的一个或多个氨基酸的突变；以及
(d)一个或多个氨基酸从跨膜和胞质结构域(TMCT)(氨基酸1201-1260)中的缺失，
其中所述CoV S糖蛋白的氨基酸相对于SEQ ID NO:2来编号。
图3示出了称为BV2378的CoV S多肽，其具有无活性弗林蛋白酶切割位点、缺失的融合肽(例如，氨基酸819-828的缺失)、K986P和V987突变，其中氨基酸相对于SEQ ID NO:1来编号。成熟BV2378多肽缺少信号肽的一个或多个氨基酸，所述信号肽为SEQ ID NO:1的氨基酸1-13。
在实施方案中，本文所述的CoV S多肽以融合前构象存在。在实施方案中，本文所述的CoV S多肽包含柔性HR2结构域。除非另外提及，否则结构域的柔性通过透射电子显微术(TEM)和2D类别取平均值来确定。电子密度的降低对应于柔性结构域。CoV S多肽抗原-对S1亚基的修饰
在实施方案中，所述CoV S多肽含有对具有SEQ ID NO:121的氨基酸序列的S1亚基的一个或多个修饰。
所述S1亚基的氨基酸序列(SEQ ID NO:121)示于下文。QCVNLTTRTQLPPAYTNSFTRGVYYPDKVFRSSVLHSTQDLFLPFFSNVTWFHAIHVSGTNGTKRFDNPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIVNNATNVVIKVCEFQFCNDPFLGVYYHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMDLEGKQGNFKNLREFVFKNIDGYFKIYSKHTPINLVRDLPQGFSALEPLVDLPIGINITRFQTLLALHRSYLTPGDSSSGWTAGAAAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFNFNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLYQDVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEHVNNSYECDIPIGAGICASYQTQTNSPRRAR
SEQ ID NO:121的加下划线的区域表示S1亚基内的可以修饰的氨基酸。
在实施方案中，本文所述的CoV S多肽包含与SEQ ID NO:1或SEQ ID NO:2的S1亚基具有至少95％、至少96％、至少97％、至少98％、至少99％或至少99.5％同一性的S1亚基。所述S1亚基可以具有与SEQ ID NO:1或SEQ ID NO:2的S1亚基的氨基酸序列相比多至约1、多至约2、多至约3、多至约4、多至约5、多至约10、多至约15、多至约20、多至约25或多至约30个氨基酸的缺失、插入或突变。所述S1亚基可以具有与SEQ ID NO:1或SEQ ID NO:2的S1亚基相比在约1与约5个之间的氨基酸、在约3与约10个之间的氨基酸、在约5与10个之间的氨基酸、在约8与12个之间的氨基酸、在约10与15个之间的氨基酸、在约12与17个之间的氨基酸、在约15与20个之间的氨基酸、在约18与23个之间的氨基酸、在约20与25个之间的氨基酸、在约22与约27个之间的氨基酸或在约25与30个之间的氨基酸的缺失、插入或突变。
在实施方案中，所述S1亚基可以含有表1A中所示的修饰的任何组合。
表1A
CoV S多肽抗原-对S1亚基-NTD的修饰
在实施方案中，所述CoV S多肽含有对NTD的一个或多个修饰。在实施方案中，所述NTD具有SEQ ID NO:118的氨基酸序列，其对应于SEQ ID NO:1的氨基酸14-305或SEQ IDNO:2的氨基酸1-292。
NTD的氨基酸序列(SEQ ID NO:118)示于下文。
QCVNLTTRTQLPPAYTNSFTRGVYYPDKVFRSSVLHSTQDLFLPFFSNVTWFHAIHVSGTNGTKRFDNPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIVNNATNVVIKVCEFQFCNDPFLGVYYHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMDLEGKQGNFKNLREFVFKNIDGYFKIYSKHTPINLVRDLPQGFSALEPLVDLPIGINITRFQTLLALHRSYLTPGDSSSGWTAGAAAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKS
SEQ ID NO:118的加下划线的区域表示NTD内的可以修饰的氨基酸。
在实施方案中，所述NTD具有SEQ ID NO:45的氨基酸序列，其对应于SEQ ID NO:1的氨基酸14至331或SEQ ID NO:2的氨基酸1-318。NTD的氨基酸序列(SEQ ID NO:45)示于下文。
QCVNLTTRTQLPPAYTNSFTRGVYYPDKVFRSSVLHSTQDLFLPFFSNVTWFHAIHVSGTNGTKRFDNPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIVNNATNVVIKVCEFQFCNDPFLGVYYHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMDLEGKQGNFKNLREFVFKNIDGYFKIYSKHTPINLVRDLPQGFSALEPLVDLPIGINITRFQTLLALHRSYLTPGDSSSGWTAGAAAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPN
在实施方案中，所述NTD和RBD重叠多至约1个氨基酸、多至约5个氨基酸、多至约10个氨基酸或多至约20个氨基酸。
在实施方案中，如本文所提供的NTD可以在C末端延伸多至5、多至10、多至15、多至20、多至25或多至30个氨基酸。
在实施方案中，本文所述的CoV S多肽包含与SEQ ID NO:1或SEQ ID NO:2的NTD具有至少95％、至少96％、至少97％、至少98％、至少99％或至少99.5％同一性的NTD。所述NTD可以具有与SEQ ID NO:1或SEQ ID NO:2的NTD的氨基酸序列相比多至约1、多至约2、多至约3、多至约4、多至约5、多至约10、多至约15、多至约20、多至约25或多至约30个氨基酸的缺失、插入或突变。所述NTD可以具有与SEQ ID NO:1或SEQ ID NO:2的NTD相比在约1与约5个之间的氨基酸、在约3与约10个之间的氨基酸、在约5与10个之间的氨基酸、在约8与12个之间的氨基酸、在约10与15个之间的氨基酸、在约12与17个之间的氨基酸、在约15与20个之间的氨基酸、在约18与23个之间的氨基酸、在约20与25个之间的氨基酸、在约22与约27个之间的氨基酸或在约25与30个之间的氨基酸的缺失、插入或突变。
在实施方案中，所述CoV S多肽含有一个或多个氨基酸从N末端结构域(NTD)(对应于SEQ ID NO:2的氨基酸1-292)中的缺失。在一些实施方案中，所述CoV S多肽含有NTD的多至约10、20、30、40、50、60、70、80、90、100、110、120、130、140、150、160、170、180、190、200、210、220、230、240、250、260、270、280、290或292个氨基酸的缺失。
在一些实施方案中，所述CoV S多肽含有一个或多个氨基酸从NTD(对应于SEQ IDNO:2的氨基酸1-318)中的缺失。在实施方案中，所述CoV S多肽含有SEQ ID NO:2的NTD的氨基酸1-318的缺失。在实施方案中，NTD的缺失增强了CoV纤突(S)多肽的蛋白质表达。在实施方案中，具有NTD缺失的CoV S多肽具有由SEQ ID NO:46、48、49、51、52和54表示的氨基酸序列。在实施方案中，具有NTD缺失的CoV S多肽由选自SEQ ID NO:47、SEQ ID NO:50和SEQ IDNO:53的分离的核酸序列编码。
在实施方案中，所述NTD可以含有表1B中所示的修饰的任何组合。修饰相对于供参考的成熟S多肽序列SEQ ID NO:2示出。
表1B
CoV S多肽抗原-对S1亚基-RBD的修饰
在实施方案中，所述CoV S多肽含有对RBD的一个或多个修饰。
在实施方案中，所述RBD具有SEQ ID NO:126的氨基酸序列，其对应于SEQ ID NO:1的氨基酸331-527或SEQ ID NO:2的氨基酸318-514。
所述RBD的氨基酸序列(SEQ ID NO:126)示于下文：
NITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGP
SEQ ID NO:126的加下划线的区域表示RBD亚基内的可以修饰的氨基酸。
在实施方案中，所述RBD具有SEQ ID NO:116的氨基酸序列，其对应于SEQ ID NO:1的氨基酸335-530或SEQ ID NO:2的氨基酸322-517。
所述RBD的氨基酸序列(SEQ ID NO:116)示于下文。
LCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKS
SEQ ID NO:116的加下划线的区域表示RBD亚基内的可以修饰的氨基酸。
在实施方案中，如本文所提供的RBD可以在N末端或C末端延伸多至1个氨基酸、多至5个氨基酸、多至10个氨基酸、多至15个氨基酸、多至20个氨基酸、多至25个氨基酸或多至30个氨基酸。
在实施方案中，本文所述的CoV S多肽包含与SEQ ID NO:1或SEQ ID NO:2的RBD具有至少95％、至少96％、至少97％、至少98％、至少99％或至少99.5％同一性的RBD。所述RBD可以具有与SEQ ID NO:1或SEQ ID NO:2的RBD的氨基酸序列相比多至约1、多至约2、多至约3、多至约4、多至约5、多至约10、多至约15、多至约20、多至约25或多至约30个氨基酸的缺失、插入或突变。所述RBD可以具有与SEQ ID NO:1或SEQ ID NO:2的RBD相比在约1与约5个之间的氨基酸、在约3与约10个之间的氨基酸、在约5与10个之间的氨基酸、在约8与12个之间的氨基酸、在约10与15个之间的氨基酸、在约12与17个之间的氨基酸、在约15与20个之间的氨基酸、在约18与23个之间的氨基酸、在约20与25个之间的氨基酸、在约22与约27个之间的氨基酸或在约25与30个之间的氨基酸的缺失、插入或突变。
在实施方案中，所述CoV S多肽在RBD中具有至少一、至少二、至少三、至少四、至少5、至少6、至少7、至少8、至少9、至少10、至少11、至少12、至少13、至少14、至少15、至少16、至少17、至少18、至少19或至少20个突变。在实施方案中，所述RBD可以含有如表1C中所示的修饰的任何组合。
表1C
CoV S多肽抗原-对SD1/2的修饰
在实施方案中，所述CoV S多肽含有对具有SEQ ID NO:122的氨基酸序列的SD1/2的一个或多个修饰，所述氨基酸序列对应于SEQ ID NO:1的氨基酸542-681或SEQ ID NO:2的氨基酸529-668。
所述SD1/2的氨基酸序列(SEQ ID NO:122)示于下文。
NFNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLYQDVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEHVNNSYECDIPIGAGICASYQTQTNSP
SEQ ID NO:122的加下划线的区域表示SD1/2内的可以修饰的氨基酸。
在实施方案中，本文所述的CoV S多肽包含与SEQ ID NO:1或SEQ ID NO:2的SD1/2具有至少95％、至少96％、至少97％、至少98％、至少99％或至少99.5％同一性的SD1/2。所述SD1/2可以具有与SEQ ID NO:1或SEQ ID NO:2的SD1/2的氨基酸序列相比多至约1、多至约2、多至约3、多至约4、多至约5、多至约10、多至约15、多至约20、多至约25或多至约30个氨基酸的缺失、插入或突变。所述SD1/2可以具有与SEQ ID NO:1或SEQ ID NO:2的SD1/2相比在约1与约5个之间的氨基酸、在约3与约10个之间的氨基酸、在约5与10个之间的氨基酸、在约8与12个之间的氨基酸、在约10与15个之间的氨基酸、在约12与17个之间的氨基酸、在约15与20个之间的氨基酸、在约18与23个之间的氨基酸、在约20与25个之间的氨基酸、在约22与约27个之间的氨基酸或在约25与30个之间的氨基酸的缺失、插入或突变。
在实施方案中，所述CoV S多肽在SD1/2中具有至少一、至少二、至少三、至少四、至少5、至少6、至少7、至少8、至少9、至少10、至少11、至少12、至少13、至少14、至少15、至少16、至少17、至少18、至少19或至少20个突变。在实施方案中，所述SD1/2可以含有如表1D中所示的修饰的任何组合。
表1D
CoV S多肽抗原-对弗林蛋白酶切割位点的修饰
在实施方案中，所述CoV S多肽含有被一个或多个突变灭活的弗林蛋白酶位点(RRAR)，其对应于SEQ ID NO:1的氨基酸682-685或SEQ ID NO:2的氨基酸669-672。弗林蛋白酶切割位点的灭活防止弗林蛋白酶切割CoV S多肽。在实施方案中，本文所述的含有灭活的弗林蛋白酶切割位点的CoV S多肽被表达为单链。
在实施方案中，构成天然弗林蛋白酶切割位点的一个或多个氨基酸被突变为任何天然氨基酸。在实施方案中，所述氨基酸是L-氨基酸。氨基酸的非限制性例子包括丙氨酸、精氨酸、甘氨酸、天冬酰胺、天冬氨酸、半胱氨酸、谷氨酰胺、谷氨酸、丝氨酸、苏氨酸、组氨酸、赖氨酸、甲硫氨酸、脯氨酸、缬氨酸、异亮氨酸、酪氨酸、色氨酸和苯丙氨酸。
在实施方案中，构成天然弗林蛋白酶切割位点的一个或多个氨基酸被突变为谷氨酰胺。在实施方案中，1、2、3或4个氨基酸可以被突变为谷氨酰胺。在实施方案中，构成天然弗林蛋白酶切割位点的一个精氨酸被突变为谷氨酰胺。在实施方案中，构成天然弗林蛋白酶切割位点的两个精氨酸被突变为谷氨酰胺。在实施方案中，构成天然弗林蛋白酶切割位点的三个精氨酸被突变为谷氨酰胺。
在实施方案中，构成天然弗林蛋白酶切割位点的一个或多个氨基酸被突变为丙氨酸。在实施方案中，1、2、3或4个氨基酸可以被突变为丙氨酸。在实施方案中，构成天然弗林蛋白酶切割位点的一个精氨酸被突变为丙氨酸。在实施方案中，构成天然弗林蛋白酶切割位点的两个精氨酸被突变为丙氨酸。在实施方案中，构成天然弗林蛋白酶切割位点的三个精氨酸被突变为丙氨酸。
在实施方案中，构成天然弗林蛋白酶切割位点的一个或多个氨基酸被突变为甘氨酸。在实施方案中，1、2、3或4个氨基酸可以被突变为甘氨酸。在实施方案中，天然弗林蛋白酶切割位点的一个精氨酸被突变为甘氨酸。在实施方案中，构成天然弗林蛋白酶切割位点的两个精氨酸被突变为甘氨酸。在实施方案中，构成天然弗林蛋白酶切割位点的三个精氨酸被突变为甘氨酸。
在实施方案中，构成天然弗林蛋白酶切割位点的一个或多个氨基酸被突变为天冬酰胺。例如，1、2、3或4个氨基酸可以被突变为天冬酰胺。在实施方案中，构成天然弗林蛋白酶切割位点的一个精氨酸被突变为天冬酰胺。在实施方案中，构成天然弗林蛋白酶切割位点的两个精氨酸被突变为天冬酰胺。在实施方案中，构成天然弗林蛋白酶切割位点的三个精氨酸被突变为天冬酰胺。
包含在所述CoV S多肽内的灭活的弗林蛋白酶位点的氨基酸序列的非限制性例子可见于表1E。
表1E：灭活的弗林蛋白酶切割位点
| 弗林蛋白酶切割位点的氨基酸序列 | 有活性或无活性的弗林蛋白酶切割位点 |
| RRAR(SEQ ID NO:6) | 有活性 |
| QQAQ(SEQ ID NO:7) | 无活性 |
| QRAR(SEQ ID NO:8) | 无活性 |
| RQAR(SEQ ID NO:9) | 无活性 |
| RRAQ(SEQ ID NO:10) | 无活性 |
| QQAR(SEQ ID NO:11) | 无活性 |
| RQAQ(SEQ ID NO:12) | 无活性 |
| QRAQ(SEQ ID NO:13) | 无活性 |
| NNAN(SEQ ID NO:14) | 无活性 |
| NRAR(SEQ ID NO:15) | 无活性 |
| RNAR(SEQ ID NO:16) | 无活性 |
| RRAN(SEQ ID NO:17) | 无活性 |
| NNAR(SEQ ID NO:18) | 无活性 |
| RNAN(SEQ ID NO:19) | 无活性 |
| NRAN(SEQ ID NO:20) | 无活性 |
| AAAA(SEQ ID NO:21) | 无活性 |
| ARAR(SEQ ID NO:22) | 无活性 |
| RAAR(SEQ ID NO:23) | 无活性 |
| RRAA(SEQ ID NO:24) | 无活性 |
| AAAR(SEQ ID NO:25) | 无活性 |
| RAAA(SEQ ID NO:26) | 无活性 |
| ARAA(SEQ ID NO:27) | 无活性 |
| GGAG(SEQ ID NO:28) | 无活性 |
| GRAR(SEQ ID NO:29) | 无活性 |
| RGAR(SEQ ID NO:30) | 无活性 |
| RRAG(SEQ ID NO:31) | 无活性 |
| GGAR(SEQ ID NO:32) | 无活性 |
| RGAG(SEQ ID NO:33) | 无活性 |
| GRAG(SEQ ID NO:34) | 无活性 |
| GSAS(SEQ ID NO:97) | 无活性 |
| GSGA(SEQ ID NO:111) | 无活性 |
在实施方案中，代替有活性的弗林蛋白酶切割位点(SEQ ID NO:6)，本文所述的CoV S多肽含有灭活的弗林蛋白酶切割位点。在实施方案中，灭活的弗林蛋白酶切割位点的氨基酸序列由SEQ ID NO:7-34或SEQ ID NO:97中的任一个表示。在实施方案中，灭活的弗林蛋白酶切割位点的氨基酸序列为QQAQ(SEQ ID NO:7)。在实施方案中，灭活的弗林蛋白酶切割位点的氨基酸序列为GSAS(SEQ ID NO:97)。在实施方案中，灭活的弗林蛋白酶切割位点的氨基酸序列为GSGA(SEQ ID NO:111)。
CoV S多肽抗原-对S2亚基的修饰
在实施方案中，所述CoV S多肽含有对具有SEQ ID NO:120的氨基酸序列的S2亚基的一个或多个修饰，所述氨基酸序列对应于SEQ ID NO:1的氨基酸686-1273或SEQ ID NO:2的氨基酸673-1260。
所述S2亚基的氨基酸序列(SEQ ID NO:120)示于下文。SVASQSIIAYTMSLGAENSVAYSNNSIAIPTNFTISVTTEILPVSMTKTSVDCTMYICGDSTECSNLLLQYGSFCTQLNRALTGIAVEQDKNTQEVFAQVKQIYKTPPIKDFGGFNFSQILPDPSKPSKRSFIEDLLFNKVTLADAGFIKQYGDCLGDIAARDLICAQKFNGLTVLPPLLTDEMIAQYTSALLAGTITSGWTFGAGAALQIPFAMQMAYRFNGIGVTQNVLYENQKLIANQFNSAIGKIQDSLSSTASALGKLQDVVNQNAQALNTLVKQLSSNFGAISSVLNDILSRLDKVEAEVQIDRLITGRLQSLQTYVTQQLIRAAEIRASANLAATKMSECVLGQSKRVDFCGKGYHLMSFPQSAPHGVVFLHVTYVPAQEKNFTTAPAICHDGKAHFPREGVFVSNGTHWFVTQRNFYEPQIITTDNTFVSGNCDVVIGIVNNTVYDPLQPELDSFKEELDKYFKNHTSPDVDLGDISGINASVVNIQKEIDRLNEVAKNLNESLIDLQELGKYEQYIKWPWYIWLGFIAGLIAIVMVTIMLCCMTSCCSCLKGCCSCGSCCKFDEDDSEPVLKGVKLHYT
SEQ ID NO:120的加下划线的区域表示S2亚基内的可以修饰的氨基酸。
在实施方案中，本文所述的CoV S多肽包含与SEQ ID NO:1或SEQ ID NO:2的S2亚基具有至少95％、至少96％、至少97％、至少98％、至少99％或至少99.5％同一性的S2亚基。所述S2亚基可以具有与SEQ ID NO:1或SEQ ID NO:2的S2亚基的氨基酸序列相比多至约1、多至约2、多至约3、多至约4、多至约5、多至约10、多至约15、多至约20、多至约25或多至约30个氨基酸的缺失、插入或突变。所述S2亚基可以具有与SEQ ID NO:1或SEQ ID NO:2的S2亚基相比在约1与约5个之间的氨基酸、在约3与约10个之间的氨基酸、在约5与10个之间的氨基酸、在约8与12个之间的氨基酸、在约10与15个之间的氨基酸、在约12与17个之间的氨基酸、在约15与20个之间的氨基酸、在约18与23个之间的氨基酸、在约20与25个之间的氨基酸、在约22与约27个之间的氨基酸或在约25与30个之间的氨基酸的缺失、插入或突变。
在实施方案中，所述S2亚基可以含有如表1F中所示的修饰的任何组合。
表1F
在实施方案中，所述CoV S多肽含有缺失，其对应于天然CoV纤突(S)多肽(SEQ IDNO:2)的氨基酸676-685内的一个或多个缺失。在实施方案中，天然CoV纤突(S)多肽(SEQ IDNO:2)的氨基酸676-685的1、2、3、4、5、6、7、8、9或10个氨基酸被缺失。在实施方案中，氨基酸676-685内的氨基酸的缺失是连续的，例如氨基酸676和677被缺失或氨基酸680和681被缺失。在实施方案中，氨基酸676-685内的氨基酸的缺失是非连续的，例如氨基酸676和680被缺失或氨基酸677和682被缺失。在实施方案中，含有对应于氨基酸676-685内的一个或多个缺失的缺失的CoV S多肽具有选自SEQ ID NO:62和SEQ ID NO:63的氨基酸序列。
在实施方案中，所述CoV S多肽含有缺失，其对应于天然CoV纤突(S)多肽(SEQ IDNO:2)的氨基酸702-711内的一个或多个缺失。在实施方案中，天然SARS-CoV-2纤突(S)多肽(SEQ ID NO:2)的氨基酸702-711的1、2、3、4、5、6、7、8、9或10个氨基酸被缺失。在实施方案中，氨基酸702-711内的氨基酸的一个或多个缺失是连续的，例如氨基酸702和703被缺失或氨基酸708和709被缺失。在实施方案中，氨基酸702-711内的氨基酸的缺失是非连续的，例如氨基酸702和704被缺失或氨基酸707和710被缺失。在实施方案中，含有对应于氨基酸702-711内的一个或多个缺失的缺失的CoV S多肽具有选自SEQ ID NO:64和SEQ ID NO:65的氨基酸序列。
在实施方案中，所述CoV S多肽含有缺失，其对应于天然CoV S多肽(SEQ ID NO:2)的氨基酸775-793内的一个或多个缺失。在实施方案中，天然SARS-CoV-2纤突(S)多肽(SEQID NO:2)的氨基酸775-793的多至约1、2、3、4、5、6、7、8、9、10、11、12、13、14、15、16、17、18或19个氨基酸被缺失。在实施方案中，氨基酸775-793内的氨基酸的一个或多个缺失是连续的，例如氨基酸776和777被缺失或氨基酸780和781被缺失。在实施方案中，氨基酸775-793内的氨基酸的缺失是非连续的，例如氨基酸775和790被缺失或氨基酸777和781被缺失。
在实施方案中，所述CoV S多肽含有融合肽(SEQ ID NO:104)的缺失，所述融合肽对应于SEQ ID NO:2的氨基酸806-815。在实施方案中，CoV纤突(S)多肽(SEQ ID NO:2)的融合肽的1、2、3、4、5、6、7、8、9或10个氨基酸被缺失。在实施方案中，融合肽内的氨基酸的缺失是连续的，例如氨基酸806和807被缺失或氨基酸809和810被缺失。在实施方案中，融合肽内的氨基酸的缺失是非连续的，例如氨基酸806和808被缺失或氨基酸810和813被缺失。在实施方案中，含有对应于融合肽的一个或多个氨基酸的缺失的CoV S多肽具有选自SEQ IDNO:66、77和105-108的氨基酸序列。
在实施方案中，所述CoV S多肽在天然CoV纤突(S)多肽(SEQ ID NO:2)的Lys-973处含有突变。在实施方案中，Lys-973被突变为任何天然氨基酸。在实施方案中，Lys-973被突变为脯氨酸。在实施方案中，Lys-973被突变为甘氨酸。在实施方案中，在氨基酸973处含有突变的CoV S多肽选自SEQ ID NO:84-89、105-106和109-110。
在实施方案中，所述CoV S多肽在天然CoV纤突(S)多肽(SEQ ID NO:2)的Val-974处含有突变。在实施方案中，Val-974被突变为任何天然氨基酸。在实施方案中，Val-974被突变为脯氨酸。在实施方案中，Val-974被突变为甘氨酸。在实施方案中，在氨基酸974处含有突变的CoV S多肽选自SEQ ID NO:84-89、105-106和109-110。
在实施方案中，所述CoV S多肽在天然CoV纤突(S)多肽(SEQ ID NO:2)的Lys-973和Val-974处含有突变。在实施方案中，Lys-973和Val-974被突变为任何天然氨基酸。在实施方案中，Lys-973和Val-974被突变为脯氨酸。在实施方案中，在氨基酸973和974处含有突变的CoV S多肽选自SEQ ID NO:84-89、105-106和109-110。
CoV S多肽抗原-对S2亚基-HR1结构域的修饰
在实施方案中，所述CoV S多肽含有对具有SEQ ID NO:119的氨基酸序列的HR1结构域的一个或多个修饰，所述氨基酸序列对应于SEQ ID NO:1的氨基酸912-984或SEQ IDNO:2的氨基酸889-971。
所述HR1结构域的氨基酸序列(SEQ ID NO:119)示于下文。MAYRFNGIGVTQNVLYENQKLIANQFNSAIGKIQDSLSSTASALGKLQDVVNQNAQALNTLVKQLSSNFGAISSVLNDILSRL
SEQ ID NO:119的加下划线的区域表示HR1结构域内的可以修饰的氨基酸。
在实施方案中，本文所述的CoV S多肽包含与SEQ ID NO:1或SEQ ID NO:2的HR1结构域具有至少95％、至少96％、至少97％、至少98％、至少99％或至少99.5％同一性的HR1结构域。所述HR1结构域可以具有与SEQ ID NO:1或SEQ ID NO:2的HR1结构域的氨基酸序列相比多至约1、多至约2、多至约3、多至约4、多至约5、多至约10、多至约15、多至约20、多至约25或多至约30个氨基酸的缺失、插入或突变。所述HR1结构域可以具有与SEQ ID NO:1或SEQID NO:2的HR1结构域相比在约1与约5个之间的氨基酸、在约3与约10个之间的氨基酸、在约5与10个之间的氨基酸、在约8与12个之间的氨基酸、在约10与15个之间的氨基酸、在约12与17个之间的氨基酸、在约15与20个之间的氨基酸、在约18与23个之间的氨基酸、在约20与25个之间的氨基酸、在约22与约27个之间的氨基酸或在约25与30个之间的氨基酸的缺失、插入或突变。
在实施方案中，所述HR1结构域可以含有如表1G中所示的修饰的任何组合。
表1G
CoV S多肽抗原-对S2亚基-HR2结构域的修饰
在实施方案中，所述CoV S多肽含有对具有SEQ ID NO:125的氨基酸序列的HR2结构域的一个或多个修饰，所述氨基酸序列对应于SEQ ID NO:1的氨基酸1163-1213或SEQ IDNO:2的氨基酸1150-1200。
所述HR2结构域的氨基酸序列(SEQ ID NO:125)示于下文。DVDLGDISGINASVVNIQKEIDRLNEVAKNLNESLIDLQELGKYEQYIKWP
在实施方案中，本文所述的CoV S多肽包含与SEQ ID NO:1或SEQ ID NO:2的HR2结构域具有至少95％、至少96％、至少97％、至少98％、至少99％或至少99.5％同一性的HR2结构域。所述HR2结构域可以具有与SEQ ID NO:1或SEQ ID NO:2的HR2结构域的氨基酸序列相比多至约1、多至约2、多至约3、多至约4、多至约5、多至约10、多至约15、多至约20、多至约25或多至约30个氨基酸的缺失、插入或突变。所述HR2结构域可以具有与SEQ ID NO:1或SEQID NO:2的HR2结构域相比在约1与约5个之间的氨基酸、在约3与约10个之间的氨基酸、在约5与10个之间的氨基酸、在约8与12个之间的氨基酸、在约10与15个之间的氨基酸、在约12与17个之间的氨基酸、在约15与20个之间的氨基酸、在约18与23个之间的氨基酸、在约20与25个之间的氨基酸、在约22与约27个之间的氨基酸或在约25与30个之间的氨基酸的缺失、插入或突变。
CoV S多肽抗原-对TM结构域的修饰
在实施方案中，所述CoV S多肽含有对具有SEQ ID NO:123的氨基酸序列的TM结构域的一个或多个修饰，所述氨基酸序列对应于SEQ ID NO:1的氨基酸1214-1237或SEQ IDNO:2的氨基酸1201-1224。
所述TM结构域的氨基酸序列(SEQ ID NO:123)示于下文。WYIWLGFIAGLIAIVMVTIMLCCM
在实施方案中，本文所述的CoV S多肽包含与SEQ ID NO:1或SEQ ID NO:2的TM结构域具有至少95％、至少96％、至少97％、至少98％、至少99％或至少99.5％同一性的TM结构域。所述TM结构域可以具有与SEQ ID NO:1或SEQ ID NO:2的TM结构域的氨基酸序列相比多至约1、多至约2、多至约3、多至约4、多至约5、多至约10、多至约15、多至约20、多至约25或多至约30个氨基酸的缺失、插入或突变。所述TM结构域可以具有与SEQ ID NO:1或SEQ IDNO:2的TM结构域相比在约1与约5个之间的氨基酸、在约3与约10个之间的氨基酸、在约5与10个之间的氨基酸、在约8与12个之间的氨基酸、在约10与15个之间的氨基酸、在约12与17个之间的氨基酸、在约15与20个之间的氨基酸、在约18与23个之间的氨基酸、在约20与25个之间的氨基酸、在约22与约27个之间的氨基酸或在约25与30个之间的氨基酸的缺失、插入或突变。
在实施方案中，本文所述的CoV S多肽缺少整个TM结构域。在实施方案中，所述CoVS多肽包含TM结构域。
CoV S多肽抗原-对CT的修饰
在实施方案中，所述CoV S多肽含有对具有SEQ ID NO:124的氨基酸序列的CT的一个或多个修饰，所述氨基酸序列对应于SEQ ID NO:1的氨基酸1238-1273或SEQ ID NO:2的氨基酸1225-1260。
所述CT的氨基酸序列(SEQ ID NO:124)示于下文：
TSCCSCLKGCCSCGSCCKFDEDDSEPVLKGVKLHYT
在实施方案中，本文所述的CoV S多肽包含与SEQ ID NO:1或SEQ ID NO:2的CT具有至少95％、至少96％、至少97％、至少98％、至少99％或至少99.5％同一性的CT。所述CT可以具有与SEQ ID NO:1或SEQ ID NO:2的CT的氨基酸序列相比多至约1、多至约2、多至约3、多至约4、多至约5、多至约10、多至约15、多至约20、多至约25或多至约30个氨基酸的缺失、插入或突变。所述CT可以具有与SEQ ID NO:1或SEQ ID NO:2的CT相比在约1与约5个之间的氨基酸、在约3与约10个之间的氨基酸、在约5与10个之间的氨基酸、在约8与12个之间的氨基酸、在约10与15个之间的氨基酸、在约12与17个之间的氨基酸、在约15与20个之间的氨基酸、在约18与23个之间的氨基酸、在约20与25个之间的氨基酸、在约22与约27个之间的氨基酸或在约25与30个之间的氨基酸的缺失、插入或突变。
在实施方案中，本文所述的CoV S多肽缺少CT。在实施方案中，所述CoV S多肽包含CT。
在实施方案中，所述CoV S多肽包含TM和CT。在实施方案中，所述CoV纤突(S)多肽含有一个或多个氨基酸从跨膜和胞质尾(TMCT)(对应于氨基酸1201-1260)中的缺失。所述TMCT的氨基酸序列由SEQ ID NO:39表示。在实施方案中，具有TMCT的一个或多个残基的缺失的CoV S多肽具有增强的蛋白质表达。在实施方案中，具有来自TMCT的一个或多个缺失的CoV纤突(S)多肽具有选自SEQ ID NO:40、41、42、52、54、59、61、88和89的氨基酸序列。在实施方案中，具有来自TM-CD的一个或多个缺失的CoV S多肽由选自SEQ ID NO:39、43、53和60的分离的核酸序列编码。
CoV S多肽抗原-突变的非限制性组合
在实施方案中，所述CoV S多肽含有天然CoV纤突(S)多肽(SEQ ID NO:2)的氨基酸56和57的缺失。
在实施方案中，所述CoV S多肽含有天然CoV纤突(S)多肽(SEQ ID NO:2)的氨基酸131和132的缺失。
在实施方案中，所述CoV S多肽含有天然CoV纤突(S)多肽(SEQ ID NO:2)的氨基酸56和131的缺失。在实施方案中，所述CoV S多肽含有天然CoV纤突(S)多肽(SEQ ID NO:2)的氨基酸57和131的缺失。
在实施方案中，所述CoV S多肽含有天然CoV纤突(S)多肽(SEQ ID NO:2)的氨基酸56、57和131的缺失。
在实施方案中，所述CoV S多肽含有天然CoV纤突(S)多肽(SEQ ID NO:2)的氨基酸56和132的缺失。
在实施方案中，所述CoV S多肽含有天然CoV纤突(S)多肽(SEQ ID NO:2)的氨基酸57和132的缺失。
在实施方案中，所述CoV S多肽含有天然CoV纤突(S)多肽(SEQ ID NO:2)的氨基酸56、57和132的缺失。
在实施方案中，所述CoV S多肽含有天然CoV纤突(S)多肽(SEQ ID NO:2)的氨基酸56、57、131和132的缺失。
在实施方案中，所述CoV S多肽含有使CoV S多肽的融合前构象稳定的突变。在实施方案中，所述CoV S多肽含有使融合前构象稳定的脯氨酸或甘氨酸取代。此策略已被用于开发如以下文献中所述的融合前稳定的MERS-CoV S蛋白，将这些文献各自通过引用以其整体并入本文：Proc Natl Acad Sci USA.2017年8月29日；114(35):E7348-E7357；SciRep.2018年10月24日；8(1):15701；美国公开号2020/0061185；以及PCT申请号PCT/US2017/058370。
在实施方案中，所述CoV S多肽含有在Lys-973和Val-974处的突变以及灭活的弗林蛋白酶切割位点。在实施方案中，所述CoV S多肽含有Lys-973和Val-974至脯氨酸的突变以及具有QQAQ(SEQ ID NO:7)或GSAS(SEQ ID NO:96)的氨基酸序列的灭活的弗林蛋白酶切割位点。在图8中描绘了含有在Lys-973和Val-974处的突变以及灭活的弗林蛋白酶切割位点的示例性CoV S多肽。在实施方案中，含有Lys-973和Val-974至脯氨酸的突变以及灭活的弗林蛋白酶切割位点的CoV S多肽具有SEQ ID NO:86或87的氨基酸序列和SEQ ID NO:96的核酸序列。
在实施方案中，所述CoV S多肽含有在Lys-973和Val-974处的突变、灭活的弗林蛋白酶切割位点以及融合肽的一个或多个氨基酸的缺失。在实施方案中，所述CoV S多肽含有Lys-973和Val-974至脯氨酸的突变、具有QQAQ(SEQ ID NO:7)或GSAS(SEQ ID NO:96)的氨基酸序列的灭活的弗林蛋白酶切割位点以及融合肽的一个或多个氨基酸的缺失。在实施方案中，含有Lys-973和Val-974至脯氨酸的突变、灭活的弗林蛋白酶切割位点以及融合肽的一个或多个氨基酸的缺失的CoV S多肽具有SEQ ID NO:105或106的氨基酸序列。在实施方案中，相对于天然CoV纤突(S)多肽(SEQ ID NO:2)，所述CoV S多肽含有Leu-5至苯丙氨酸的突变、Thr-7至天冬酰胺的突变、Pro-13至丝氨酸的突变、Asp-125至酪氨酸的突变、Arg-177至丝氨酸的突变、Lys-404至苏氨酸的突变、Glu-471至赖氨酸的突变、Asn-488至酪氨酸的突变、His-642至酪氨酸的突变、Thr-1014至异亮氨酸的突变、Lys-973和Val-974至脯氨酸的突变以及具有QQAQ(SEQ ID NO:7)或GSAS(SEQ ID NO:96)的氨基酸序列的灭活的弗林蛋白酶切割位点。
在实施方案中，相对于天然CoV纤突(S)多肽(SEQ ID NO:2)，所述CoV S多肽含有Trp-139至半胱氨酸的突变、Leu-439至精氨酸的突变、Lys-973和Val-974至脯氨酸的突变以及具有QQAQ(SEQ ID NO:7)或GSAS(SEQ ID NO:96)的氨基酸序列的灭活的弗林蛋白酶切割位点。在实施方案中，相对于天然CoV纤突(S)多肽(SEQ ID NO:1)，所述CoV S多肽含有Trp-152至半胱氨酸的突变、Leu-452至精氨酸的突变、Ser-13至异亮氨酸的突变、Lys-986和Val-987至脯氨酸的突变以及具有QQAQ(SEQ ID NO:7)或GSAS(SEQ ID NO:96)的氨基酸序列的灭活的弗林蛋白酶切割位点。
在实施方案中，相对于天然CoV纤突(S)多肽(SEQ ID NO:2)，所述CoV S多肽含有Lys-404至苏氨酸或天冬酰胺的突变、Glu-471至赖氨酸的突变、Asn-488至酪氨酸的突变、Leu-5至苯丙氨酸的突变、Asp-67至丙氨酸的突变、Asp-202至甘氨酸的突变、氨基酸229-231中的一个或多个的缺失、Arg-233至异亮氨酸的突变、Lys-973和Val-974至脯氨酸的突变以及具有QQAQ(SEQ ID NO:7)或GSAS(SEQ ID NO:96)的氨基酸序列的灭活的弗林蛋白酶切割位点。
在实施方案中，相对于天然CoV纤突(S)多肽(SEQ ID NO:2)，所述CoV S多肽含有Asn-488至酪氨酸的突变、Lys-973和Val-974至脯氨酸的突变以及具有QQAQ(SEQ ID NO:7)或GSAS(SEQ ID NO:96)的氨基酸序列的灭活的弗林蛋白酶切割位点。在实施方案中，具有Asn-488至酪氨酸的突变、Lys-973和Val-974至脯氨酸的突变以及具有QQAQ(SEQ ID NO:7)或GSAS(SEQ ID NO:96)的氨基酸序列的灭活的弗林蛋白酶切割位点的CoV S多肽包含SEQID NO:112的氨基酸序列。
在实施方案中，相对于天然CoV纤突(S)多肽(SEQ ID NO:2)，所述CoV S多肽含有Asp-601至甘氨酸的突变、Asn-488至酪氨酸的突变、Lys-973和Val-974至脯氨酸的突变以及具有QQAQ(SEQ ID NO:7)或GSAS(SEQ ID NO:96)的氨基酸序列的灭活的弗林蛋白酶切割位点。在实施方案中，具有Asn-488至酪氨酸的突变、Lys-973和Val-974至脯氨酸的突变以及具有QQAQ(SEQ ID NO:7)或GSAS(SEQ ID NO:96)的氨基酸序列的灭活的弗林蛋白酶切割位点的CoV S多肽包含SEQ ID NO:113的氨基酸序列。
在实施方案中，相对于天然CoV纤突(S)多肽(SEQ ID NO:2)，所述CoV S多肽含有氨基酸56、57和131的缺失、Asn-488至酪氨酸的突变、Ala-557至天冬氨酸的突变、Asp-601至甘氨酸的突变、Pro-668至组氨酸的突变、Thr-703至异亮氨酸的突变、Ser-969至丙氨酸的突变、Asp-1105至组氨酸的突变、Lys-973和Val-974至脯氨酸的突变以及具有QQAQ(SEQID NO:7)或GSAS(SEQ ID NO:96)的氨基酸序列的灭活的弗林蛋白酶切割位点。在实施方案中，具有氨基酸56、57和131的缺失、Asn-488至酪氨酸的突变、Ala-557至天冬氨酸的突变、Asp-601至甘氨酸的突变、Pro-668至组氨酸的突变、Thr-703至异亮氨酸的突变、Ser-969至丙氨酸的突变、Asp-1105至组氨酸的突变、Lys-973和Val-974至脯氨酸的突变以及具有QQAQ(SEQ ID NO:7)或GSAS(SEQ ID NO:96)的氨基酸序列的灭活的弗林蛋白酶切割位点的CoV S多肽包含SEQ ID NO:114的氨基酸序列。
在实施方案中，相对于天然CoV纤突(S)多肽(SEQ ID NO:2)，所述CoV S多肽含有氨基酸56、57和132的缺失、Asn-488至酪氨酸的突变、Ala-557至天冬氨酸的突变、Asp-601至甘氨酸的突变、Pro-668至组氨酸的突变、Thr-703至异亮氨酸的突变、Ser-969至丙氨酸的突变、Asp-1105至组氨酸的突变、Lys-973和Val-974至脯氨酸的突变以及具有QQAQ(SEQID NO:7)或GSAS(SEQ ID NO:96)的氨基酸序列的灭活的弗林蛋白酶切割位点。在实施方案中，具有氨基酸56、57和132的缺失、Asn-488至酪氨酸的突变、Ala-557至天冬氨酸的突变、Asp-601至甘氨酸的突变、Pro-668至组氨酸的突变、Thr-703至异亮氨酸的突变、Ser-969至丙氨酸的突变、Asp-1105至组氨酸的突变、Lys-973和Val-974至脯氨酸的突变以及具有QQAQ(SEQ ID NO:7)或GSAS(SEQ ID NO:96)的氨基酸序列的灭活的弗林蛋白酶切割位点的CoV S多肽包含SEQ ID NO:114的氨基酸序列。
在实施方案中，相对于天然CoV纤突(S)多肽(SEQ ID NO:2)，所述CoV S多肽含有Asn-488至酪氨酸的突变、Asp-67至丙氨酸的突变、Leu-229至组氨酸的突变、Asp-202至甘氨酸的突变、Lys-404至天冬酰胺的突变、Glu-471至赖氨酸的突变、Ala-688至缬氨酸的突变、Asp-601至甘氨酸的突变、Lys-973和Val-974至脯氨酸的突变以及具有QQAQ(SEQ IDNO:7)或GSAS(SEQ ID NO:96)的氨基酸序列的灭活的弗林蛋白酶切割位点。在实施方案中，具有Asn-488至酪氨酸的突变、Asp-67至丙氨酸的突变、Leu-229至组氨酸的突变、Asp-202至甘氨酸的突变、Lys-404至天冬酰胺的突变、Glu-471至赖氨酸的突变、Ala-688至缬氨酸的突变、Asp-601至甘氨酸的突变、Lys-973和Val-974至脯氨酸的突变以及具有QQAQ(SEQID NO:7)或GSAS(SEQ ID NO:96)的氨基酸序列的灭活的弗林蛋白酶切割位点的CoV S多肽包含SEQ ID NO:115的氨基酸序列。
在实施方案中，所述CoV纤突(S)多肽包含多肽接头。在实施方案中，所述多肽接头含有甘氨酸和丝氨酸。在实施方案中，所述接头具有约50％、约55％、约60％、约65％、约70％、约75％、约80％、约85％、约90％、约95％或约100％甘氨酸。
在实施方案中，所述多肽接头具有重复序列(SGGG)n(SEQ ID NO:91)，其中n是1至50的整数(例如1、2、3、4、5、6、7、8、9、10、11、12、13、14、15、16、17、18、19、20、21、22、23、24、25、26、27、28、29、30、31、32、33、34、35、36、37、38、39、40、41、42、43、44、45、46、47、48、49或50)。在实施方案中，所述多肽接头具有对应于SEQ ID NO:90的氨基酸序列。
在实施方案中，所述多肽接头具有重复序列(GGGGS)n(SEQ ID NO:93)，其中n是1至50的整数(例如1、2、3、4、5、6、7、8、9、10、11、12、13、14、15、16、17、18、19、20、21、22、23、24、25、26、27、28、29、30、31、32、33、34、35、36、37、38、39、40、41、42、43、44、45、46、47、48、49或50)。
在实施方案中，所述多肽接头具有重复序列(GGGS)n(SEQ ID NO:92)，其中n是1至50的整数(例如1、2、3、4、5、6、7、8、9、10、11、12、13、14、15、16、17、18、19、20、21、22、23、24、25、26、27、28、29、30、31、32、33、34、35、36、37、38、39、40、41、42、43、44、45、46、47、48、49或50)。
在一些方面，所述多肽接头是聚(Gly)n接头，其中n是1、2、3、4、5、6、7、8、9、10、15、16、17、18、19或20。在其他实施方案中，所述接头选自：二肽、三肽和四肽。在实施方案中，所述接头是选自丙氨酸-丝氨酸(AS)、亮氨酸-谷氨酸(LE)和丝氨酸-精氨酸(SR)的二肽。
在实施方案中，所述多肽接头包含天然存在的CoV S多肽或本文所公开的CoV S多肽的1至100个之间的连续氨基酸。在实施方案中，所述多肽接头具有对应于SEQ ID NO:94的氨基酸序列。
在实施方案中，所述CoV纤突(S)多肽包含折叠子。在实施方案中，所述TMCT被折叠子替代。在实施方案中，折叠子引起CoV纤突(S)多肽三聚化。在实施方案中，所述折叠子是本领域已知的氨基酸序列。在实施方案中，所述折叠子具有SEQ ID NO:68的氨基酸序列。在实施方案中，所述折叠子是T4次要纤维蛋白(fibritin)三聚化基序。在实施方案中，T4次要纤维蛋白三聚化结构域具有SEQ ID NO:103的氨基酸序列。在实施方案中，所述折叠子的氨基酸序列由多肽接头与CoV纤突(S)多肽分开。多肽接头的非限制性例子可见于整个本公开文本。
在实施方案中，本公开文本提供了包含冠状病毒S蛋白的片段的CoV S多肽以及包含所述多肽的纳米颗粒和疫苗。在实施方案中，冠状病毒S蛋白的片段的长度在10与1500个氨基酸之间(例如长度为约10、约20、约30、约40、约50、约60、约70、约80、约90、约100、约150、约200、约250、约300、约350、约400、约450、约500、约550、约600、约650、约700、约750、约800、约850、约900、约950、约1000、约1050、约1100、约1150、约1200、约1250、约1300、约1350、约1400、约1450或约1500个氨基酸)。在实施方案中，冠状病毒S蛋白的片段选自受体结合结构域(RBD)、亚结构域1、亚结构域2、上螺旋、融合肽、连接区、七肽重复区1、中央螺旋、七肽重复区2、NTD和TMCT。
在实施方案中，所述CoV S多肽包含RBD和亚结构域1。在实施方案中，包含RBD和亚结构域1的CoV S多肽是SEQ ID NO:1的氨基酸319至591。
在实施方案中，所述CoV S多肽含有冠状病毒S蛋白的片段，其中所述冠状病毒S蛋白的片段是RBD。RBD的非限制性例子包括SARS-CoV-2(氨基酸序列＝SEQ ID NO:69)的RBD、SARS(氨基酸序列＝SEQ ID NO:70)的RBD和MERS(氨基酸序列＝SEQ ID NO:71)的RBD。
在实施方案中，所述CoV S多肽含有通过多肽接头连接的两个或更多个RBD。在实施方案中，所述多肽接头具有SEQ ID NO:90或SEQ ID NO:94的氨基酸序列。
在实施方案中，所述CoV S多肽含有1、2、3、4、5、6、7、8、9、10、11、12、13、14、15、16、17、18、19或20个RBD。
在一些实施方案中，所述CoV S多肽含有通过多肽接头连接的两个或更多个SARS-CoV-2RBD。在实施方案中，含有两个或更多个SARS-CoV-2RBD的抗原具有对应于SEQ ID NO:72-75之一的氨基酸序列。
在实施方案中，所述CoV S多肽含有SARS-CoV-2RBD和SARS RBD。在实施方案中，所述CoV S多肽包含SARS-CoV-2RBD和SARS RBD，其中每个RBD由多肽接头分开。在实施方案中，包含SARS-CoV-2RBD和SARS RBD的CoV S多肽具有选自SEQ ID NO:76-79的氨基酸序列。
在实施方案中，所述CoV S多肽含有SARS-CoV-2RBD和MERS RBD。在实施方案中，所述CoV S多肽包含SARS-CoV-2RBD和MERS RBD，其中每个RBD由多肽接头分开。
在实施方案中，所述CoV S多肽包含SARS RBD和MERS RBD。在实施方案中，所述CoVS多肽包含SARS RBD和MERS RBD，其中每个RBD由多肽接头分开。
在实施方案中，所述CoV S多肽包含SARS-CoV-2RBD、SARS RBD和MERS RBD。在实施方案中，所述CoV S多肽含有SARS-CoV-2RBD、SARS RBD和MERS RBD，其中每个RBD由多肽接头分开。在实施方案中，包含SARS-CoV-2RBD、SARS RBD和MERS RBD的CoV S多肽具有选自SEQ ID NO:80-83的氨基酸序列。
在实施方案中，本文所述的CoV S多肽与N末端信号肽一起表达。在实施方案中，所述N末端信号肽具有SEQ ID NO:5(MFVFLVLLPLVSS)的氨基酸序列。在实施方案中，所述N末端信号肽具有SEQ ID NO:117(MFVFLVLLPLVSI)的氨基酸序列。在实施方案中，所述信号肽可以被使得能够表达CoV S蛋白的任何信号肽替代。在实施方案中，CoV S蛋白信号肽氨基酸中的一个或多个可以被缺失或突变。维持起始的甲硫氨酸残基以起始表达。在实施方案中，所述CoV S多肽由选自下组的核酸序列编码：SEQ ID NO:35、SEQ ID NO:37、SEQ ID NO:95、SEQ ID NO:43、SEQ ID NO:47、SEQ ID NO:50、SEQ ID NO:53、SEQ ID NO:55、SEQ IDNO:57、SEQ ID NO:96和SEQ ID NO:60。在实施方案中，相对于天然CoV纤突(S)信号多肽(SEQ ID NO:5)，所述CoV S多肽的N末端信号肽在Ser-13处包含突变。在实施方案中，Ser-13被突变为任何天然氨基酸。在实施方案中，Ser-13被突变为丙氨酸、甲硫氨酸、异亮氨酸、亮氨酸、苏氨酸或缬氨酸。在实施方案中，Ser-13被突变为异亮氨酸。
所述CoV S蛋白在宿主细胞中表达后，N末端信号肽被切割以提供成熟CoV蛋白序列(SEQ ID NO:2、4、38、41、44、48、51、54、58、61、63、65、67、73、75、78、79、82、83、85、87、89、106和110)。在实施方案中，所述信号肽被宿主细胞蛋白酶切割。在方面中，可以从宿主细胞中分离全长蛋白质，随后切割信号肽。
在表达和纯化期间从具有对应于SEQ ID NO:1、3、36、40、42、46、49、52、56、59、62、64、66、72、74、76、77、80、81、84、86、87、105、107、88和109的氨基酸序列的CoV纤突(S)多肽中切割信号肽后，获得具有选自SEQ ID NO:2、4、38、41、44、48、51、54、58、61、63、65、67、73、75、78、79、82、83、85、106、108、89和110或112-115的氨基酸序列的成熟多肽，并且将其用于产生CoV S纳米颗粒疫苗或CoV S纳米颗粒。
有利地，相对于天然CoV纤突(S)蛋白，所公开的CoV S多肽可以具有增强的蛋白质表达和稳定性。
在实施方案中，本文所述的CoV S多肽含有相对于天然冠状病毒S蛋白(SEQ IDNO:2)的进一步修饰。在实施方案中，本文所述的冠状病毒S蛋白与天然冠状病毒S蛋白表现出至少80％、或至少90％、或至少95％、或至少97％、或至少99％同一性。本领域技术人员将使用已知技术来计算重组冠状病毒S蛋白与天然蛋白或本文所述的任何CoV S多肽的同一性百分比。例如，可以使用可在线获得的工具CLUSTALW2或基本局部比对搜索工具(BasicLocal Alignment Search Tool，BLAST)来计算同一性百分比。可以将以下默认参数用于CLUSTALW2配对比对：蛋白质权重矩阵＝Gonnet；空位开放＝10；空位延伸＝0.1。
在实施方案中，本文所述的CoV S多肽与具有SEQ ID NO:87的氨基酸序列的CoV S多肽具有至少95％、至少96％、至少97％、至少98％、至少99％或至少99.5％同一性。与具有SEQ ID NO:87的氨基酸序列的CoV S多肽的氨基酸序列相比，CoV S多肽可以具有多至约1、多至约2、多至约3、多至约4、多至约5、多至约10、多至约15、多至约20、多至约25、多至约30、多至约35、多至约40、多至约45或多至约50个氨基酸的缺失、插入或突变。与具有SEQ IDNO:87的氨基酸序列的CoV S多肽相比，CoV S多肽可以具有在约1与约5个之间的氨基酸、在约3与约10个之间的氨基酸、在约5与10个之间的氨基酸、在约8与12个之间的氨基酸、在约10与15个之间的氨基酸、在约12与17个之间的氨基酸、在约15与20个之间的氨基酸、在约18与23个之间的氨基酸、在约20与25个之间的氨基酸、在约22与约27个之间的氨基酸、在约25与30个之间的氨基酸、在约30与35个之间的氨基酸、在约35与40个之间的氨基酸、在约40与45个之间的氨基酸或在约45与50个之间的氨基酸的缺失、插入或突变。在实施方案中，与冠状病毒S蛋白(SEQ ID NO:87)相比，本文所述的CoV S多肽包含约1、约2、约3、约4、约5、约6、约7、约8、约9、约10、约11、约12、约13、约14、约15、约16、约17、约18、约19、约20、约21、约22、约23、约24或约25个取代。
在实施方案中，所述冠状病毒S多肽在N末端、C末端或N末端和C末端两者处延伸。在一些方面，所述延伸物是对诸如纯化或检测等功能有用的标签。在一些方面，所述标签含有表位。例如，所述标签可以是聚谷氨酸标签、FLAG标签、HA标签、聚His标签(具有约5-10个组氨酸)(SEQ ID NO:101)、六组氨酸标签(SEQ ID NO:100)、8X-His标签(具有八个组氨酸)(SEQ ID NO:102)、Myc标签、谷胱甘肽-S-转移酶标签、绿色荧光蛋白标签、麦芽糖结合蛋白标签、硫氧还蛋白标签或Fc标签。在其他方面，所述延伸物可以是与蛋白质融合以增强表达的N末端信号肽。尽管此类信号肽通常在细胞中表达期间被切割，但一些纳米颗粒可以含有具有完整信号肽的抗原。因此，当纳米颗粒包含抗原时，所述抗原可以含有延伸物，因此当掺入纳米颗粒中时可以是融合蛋白。出于计算与序列的同一性的目的，不包括延伸物。在实施方案中，所述标签是蛋白酶切割位点。蛋白酶切割位点的非限制性例子包括HRV3C蛋白酶切割位点、胰凝乳蛋白酶、胰蛋白酶、弹性蛋白酶、内肽酶、半胱天冬酶1、半胱天冬酶2、半胱天冬酶3、半胱天冬酶4、半胱天冬酶5、半胱天冬酶6、半胱天冬酶7、半胱天冬酶8、半胱天冬酶9、半胱天冬酶10、肠激酶、因子Xa、颗粒酶B、TEV蛋白酶和凝血酶。在实施方案中，所述蛋白酶切割位点是HRV3C蛋白酶切割位点。在实施方案中，所述蛋白酶切割位点包含SEQ IDNO:98的氨基酸序列。
在实施方案中，所述CoV S糖蛋白包含融合蛋白。在实施方案中，所述CoV S糖蛋白包含N末端融合蛋白。在实施方案中，所述CoV S糖蛋白包含C末端融合蛋白。在实施方案中，所述融合蛋白包括可用于蛋白质表达、纯化或检测的标签。在实施方案中，所述标签是聚His标签(具有约5-10个组氨酸)、Myc标签、谷胱甘肽-S-转移酶标签、绿色荧光蛋白标签、麦芽糖结合蛋白标签、硫氧还蛋白标签、Strep标签、Twin-Strep标签或Fc标签。在实施方案中，所述标签是Fc标签。在实施方案中，所述Fc标签是单体、二聚体或三聚体的。在实施方案中，所述标签是六组氨酸标签，例如含有六个组氨酸的聚His标签(SEQ ID NO:100)。在实施方案中，所述标签是具有SEQ ID NO:99的氨基酸序列的Twin-Strep标签。
在实施方案中，所述CoV S多肽是包含另一种冠状病毒蛋白的融合蛋白。在实施方案中，另一种冠状病毒蛋白来自相同的冠状病毒。在实施方案中，另一种冠状病毒蛋白来自不同的冠状病毒。
在一些方面，所述CoV S蛋白可以被截短。例如，N末端可以被截短约10个氨基酸、约30个氨基酸、约50个氨基酸、约75个氨基酸、约100个氨基酸或约200个氨基酸。代替N末端或除了N末端之外，C末端可以被截短。例如，C末端可以被截短约10个氨基酸、约30个氨基酸、约50个氨基酸、约75个氨基酸、约100个氨基酸或约200个氨基酸。出于计算与具有截短的蛋白质的同一性的目的，在蛋白质的剩余部分上测量同一性。
含有CoV纤突(S)多肽的纳米颗粒
在实施方案中，使用成熟CoV S多肽抗原来产生包含冠状病毒S纳米颗粒的疫苗。在实施方案中，本公开文本的纳米颗粒包含本文所述的CoV S多肽。在实施方案中，本公开文本的纳米颗粒包含与洗涤剂核心缔合的CoV S多肽。洗涤剂的存在通过形成组织并呈递抗原的核心来促进纳米颗粒的形成。在实施方案中，所述纳米颗粒以含有组装成多寡聚糖蛋白-洗涤剂(例如PS80)纳米颗粒的CoV S多肽，其中头部区域向外突出并且疏水区域和PS80洗涤剂形成被糖蛋白包围的中央核心。在实施方案中，所述CoV S多肽固有地含有或适于含有跨膜结构域以促进将所述蛋白质缔合到洗涤剂核心中。在实施方案中，所述CoV S多肽含有头部结构域。图10示出了本公开文本的CoV S多肽的示例性结构。主要地，CoV S多肽三聚体的跨膜结构域与洗涤剂缔合；然而，多肽的其他部分也可以相互作用。有利地，所述纳米颗粒具有改善的对环境胁迫的抗性，使得由于在洗涤剂周围组织蛋白质的多个拷贝，它们提供增强的稳定性和/或改善的对免疫系统的呈递。
在实施方案中，所述洗涤剂核心是非离子型洗涤剂核心。在实施方案中，所述CoVS多肽与非离子型洗涤剂核心缔合。在实施方案中，所述洗涤剂选自聚山梨醇酯20(PS20)、聚山梨醇酯40(PS40)、聚山梨醇酯60(PS60)、聚山梨醇酯65(PS65)和聚山梨醇酯80(PS80)。
在实施方案中，所述洗涤剂是PS80。
在实施方案中，所述CoV S多肽形成三聚体。在实施方案中，所述CoV S多肽纳米颗粒由围绕非离子型洗涤剂核心的多个多肽三聚体构成。在实施方案中，所述纳米颗粒含有至少约1个或多个三聚体。在实施方案中，所述纳米颗粒含有纤突蛋白的至少约5个三聚体至约30个三聚体。在实施方案中，每个纳米颗粒可以含有1、2、3、4、5、6、7、8、9、10、11、12、或15、20、25或30个三聚体，包括在中间的所有值和范围。本文所公开的组合物可以含有具有不同数量的三聚体的纳米颗粒。例如，组合物可以含有三聚体的数量范围从2-9的纳米颗粒；在实施方案中，组合物中的纳米颗粒可以含有2-6个三聚体。在实施方案中，所述组合物含有纳米颗粒的异质群体，每个纳米颗粒具有2至6个三聚体，或者每个纳米颗粒具有2至9个三聚体。在实施方案中，所述组合物可以含有纳米颗粒的基本上均质的群体。例如，所述群体可以含有约95％的具有5个三聚体的纳米颗粒。
本文所公开的纳米颗粒的粒度具有一定范围。在实施方案中，本文所公开的纳米颗粒的粒度的Z-ave尺寸范围从约20nm至约60nm、约20nm至约50nm、约20nm至约45nm、约20nm至约35nm、约20nm至约30nm、约25nm至约35nm或约25nm至约45nm。除非另外说明，否则使用Zetasizer NanoZS(Malvern，英国)通过动态光散射(DLS)来测量粒度(Z-ave)。
在实施方案中，与包含野生型CoV S多肽的纳米颗粒相比，包含本文所公开的CoVS多肽的纳米颗粒具有减小的粒度。在实施方案中，所述CoV S多肽的粒度小至少约40％，例如粒度小至少约40％、至少约45％、至少约50％、至少约55％、至少约60％、至少约65％、至少约70％、至少约75％、至少约80％或至少约85％。
包含本文所公开的CoV S多肽的纳米颗粒在尺寸、形状和质量方面比包含野生型CoV S多肽的纳米颗粒更均匀。多分散性指数(PDI)是异质性的度量，除非另外说明，否则使用Malvern Setasizer通过动态光散射来测量。在实施方案中，本文测量的颗粒的PDI从约0.2至约0.45，例如约0.2、约0.25、约0.29、约0.3、约0.35、约0.40或约0.45。在实施方案中，本文测量的纳米颗粒的PDI比包含野生型CoV S多肽的纳米颗粒的PDI小至少约25％，例如小至少约25％、至少约30％、至少约35％、至少约40％、至少约45％、至少约50％、至少约55％或至少约60％。
与野生型CoV S多肽或其纳米颗粒相比，所述CoV S多肽和包含所述多肽的纳米颗粒具有改善的热稳定性。除非另外说明，否则使用差示扫描量热法(DSC)来测量CoV S多肽的热稳定性。转变焓(ΔHcal)是使CoV S多肽展开所需的能量。在实施方案中，与野生型CoVS多肽相比，所述CoV S多肽具有增加的ΔHcal。在实施方案中，CoV S多肽的ΔHcal是野生型CoV S多肽的ΔHcal的约2倍、约3倍、约4倍、约5倍、约6倍、约7倍、约8倍、约9倍或约10倍。
在本文所公开的疫苗组合物中可以包括若干种纳米颗粒类型。在一些方面，纳米颗粒类型呈各向异性棒的形式，其可以是二聚体或单体。在其他方面，纳米颗粒类型是球形寡聚体。在又其他方面，所述纳米颗粒可以被描述为中间体纳米颗粒，其具有介于前两种类型之间的沉降特性。可以通过控制产生过程期间的洗涤剂和蛋白质浓度来调节纳米颗粒类型的形成。可以通过测量沉降系数来确定纳米颗粒类型。
含有CoV S多肽抗原的纳米颗粒的产生
本公开文本的纳米颗粒是非天然存在的产物，其组分在自然界中不会一起出现。通常，本文所公开的方法使用洗涤剂交换方法，其中使用第一洗涤剂来分离蛋白质，然后将该第一洗涤剂交换成第二洗涤剂以形成纳米颗粒。
所述纳米颗粒中包含的抗原通常是通过在宿主细胞中重组表达而产生的。可以使用标准重组技术。在实施方案中，使用杆状病毒系统在昆虫宿主细胞中表达所述CoV S多肽。在实施方案中，所述杆状病毒是组织蛋白酶-L敲除的杆状病毒、几丁质酶敲除的杆状病毒。任选地，所述杆状病毒是组织蛋白酶-L和几丁质酶两者的双重敲除。在昆虫细胞表达系统中可以获得高水平的表达。昆虫细胞的非限制性例子是草地贪夜蛾(Spodopterafrugiperda)(Sf)细胞(例如Sf9、Sf21)、粉纹夜蛾(Trichoplusiani)细胞(例如High Five细胞)和果蝇(Drosophila)S2细胞。在实施方案中，在任何合适的宿主细胞中产生本文所述的CoV S多肽。在实施方案中，所述宿主细胞是昆虫细胞。在实施方案中，所述昆虫细胞是Sf9细胞。
典型的转染和细胞生长方法可以用于培养所述细胞。可以根据本领域熟知的方法将载体(例如，包含编码融合蛋白的多核苷酸的载体)转染到宿主细胞中。例如，可以通过磷酸钙共沉淀、电穿孔、显微注射、脂质转染和采用多胺转染试剂的转染来实现将核酸引入真核细胞中。在一个实施方案中，所述载体是重组杆状病毒。
使宿主细胞生长的方法包括但不限于分批、补料分批、连续和灌注细胞培养技术。细胞培养意指细胞在生物反应器(发酵室)中的生长和繁殖，在所述生物反应器中细胞繁殖并表达蛋白质(例如重组蛋白)以用于纯化和分离。通常，细胞培养是在生物反应器中在无菌、受控的温度和大气条件下进行。生物反应器是可以监测环境条件诸如温度、大气、搅拌和/或pH的用于培养细胞的室。在一个实施方案中，所述生物反应器是不锈钢室。在另一个实施方案中，所述生物反应器是预灭菌的塑料袋(例如
Wave Biotech，新泽西州布里奇沃特)。在其他实施方案中，所述预灭菌的塑料袋是约50L至3500L的袋。
含有CoV纤突(S)蛋白抗原的纳米颗粒的提取和纯化
在宿主细胞生长后，可以使用洗涤剂和纯化方案从宿主细胞中收获所述蛋白质。一旦宿主细胞已生长48至96小时，便从培养基中分离细胞，并且添加含洗涤剂的溶液以溶解细胞膜，从而使蛋白质释放到洗涤剂提取物中。Triton X-100和
壬基酚乙氧基化物(也称为NP-9)各自是用于提取的优选洗涤剂。可以将洗涤剂添加至约0.1％至约1.0％的最终浓度。例如，浓度可以为约0.1％、约0.2％、约0.3％、约0.5％、约0.7％、约0.8％或约1.0％。范围可以是约0.1％至约0.3％。在方面中，浓度为约0.5％。
在其他方面，不同的第一洗涤剂可以用于从宿主细胞中分离所述蛋白质。例如，所述第一洗涤剂可以是双(聚乙二醇双[咪唑基羰基])、壬苯醇醚-9、双(聚乙二醇双[咪唑基羰基])、
聚乙二醇十二烷基醚35、
聚乙二醇(3)十六烷基醚56、
醇乙氧基化物72、
聚乙二醇2硬脂基醚76、
聚乙二醇单油烯基醚92V、
聚氧乙烯(10)油烯基醚97、
聚乙二醇十六烷基醚58P、
EL聚乙二醇甘油蓖麻酸酯、十乙二醇单十二烷基醚、N-癸酰基-N-甲基葡糖胺、正癸基α-D吡喃葡萄糖苷、癸基β-D-麦芽吡喃糖苷、正十二烷酰基-N-甲基葡糖酰胺、正十二烷基α-D-麦芽糖苷、正十二烷基β-D-麦芽糖苷、正十二烷基β-D-麦芽糖苷、七乙二醇单癸基醚、七乙二醇单十二烷基醚、七乙二醇单十四烷基醚、正十六烷基β-D-麦芽糖苷、六乙二醇单十二烷基醚、六乙二醇单十六烷基醚、六乙二醇单十八烷基醚、六乙二醇单十四烷基醚、Igepal CA-630、IgepalCA-630、甲基-6-0-(N-庚基氨基甲酰基)-α-D-吡喃葡萄糖苷、九乙二醇单十二烷基醚、N-壬酰基-N-甲基葡糖胺、N-壬酰基N-甲基葡糖胺、八乙二醇单癸基醚、八乙二醇单十二烷基醚、八乙二醇单十六烷基醚、八乙二醇单十八烷基醚、八乙二醇单十四烷基醚、辛基-β-D吡喃葡萄糖苷、五乙二醇单癸基醚、五乙二醇单十二烷基醚、五乙二醇单十六烷基醚、五乙二醇单己基醚、五乙二醇单十八烷基醚、五乙二醇单辛基醚、聚乙二醇二缩水甘油醚、聚乙二醇醚W-1、聚氧乙烯10十三烷基醚、聚氧乙烯100硬脂酸酯、聚氧乙烯20异十六烷基醚、聚氧乙烯20油烯基醚、聚氧乙烯40硬脂酸酯、聚氧乙烯50硬脂酸酯、聚氧乙烯8硬脂酸酯、聚氧乙烯双(咪唑基羰基)、聚氧乙烯25丙二醇硬脂酸酯、皂草苷(Saponin from Quillaja bark)、
20山梨聚糖月桂酸酯、
40山梨聚糖单棕榈酸酯、
60山梨聚糖硬脂酸酯、
65山梨聚糖三硬脂酸酯、
80山梨聚糖单油酸酯、
85山梨聚糖三油酸酯、
仲醇乙氧基化物15-S-12型、
仲醇乙氧基化物15-S-30型、
仲醇乙氧基化物15-S-5型、
仲醇乙氧基化物15-S-7型、
仲醇乙氧基化物15-S-9型、
壬基酚乙氧基化物NP-10型、
壬基酚乙氧基化物NP-4型、
壬基酚乙氧基化物NP-40型、
壬基酚乙氧基化物NP-7型、
壬基酚乙氧基化物NP-9型、
支链仲醇乙氧基化物TMN-10型、
支链仲醇乙氧基化物TMN-6型、TRITONTM X-100聚乙二醇叔辛基苯基醚或其组合。
然后可以使用离心从细胞碎片中分离所述纳米颗粒。在实施方案中，可以使用诸如采用氯化铯、蔗糖和碘克沙醇的梯度离心。其他技术可以用作替代或补充，诸如标准纯化技术，包括例如离子交换色谱、亲和色谱和凝胶过滤色谱。
例如，第一柱可以是离子交换色谱树脂，诸如
EMD基于甲基丙烯酸酯的聚合物珠TMAE(EMD Millipore)；第二柱可以是小扁豆(Lens culinaris)凝集素亲和树脂；并且第三柱可以是阳离子交换柱，诸如
EMD基于甲基丙烯酸酯的聚合物珠SO3(EMD Millipore)树脂。在其他方面，阳离子交换柱可以是MMC柱或Nuvia CPrime柱(Bio-Rad Laboratories,Inc)。优选地，本文所公开的方法不使用洗涤剂提取柱；例如疏水相互作用柱。这样的柱通常在纯化期间用于去除洗涤剂，但可能对此处公开的方法产生负面影响。
含有CoV S多肽抗原的纳米颗粒的洗涤剂交换
为了形成纳米颗粒，用于从宿主细胞中提取蛋白质的第一洗涤剂基本上被第二洗涤剂替代以得到纳米颗粒结构。NP-9是优选的提取洗涤剂。通常，当通过HPLC测量时，所述纳米颗粒不含有可检测的NP-9。所述第二洗涤剂通常选自PS20、PS40、PS60、PS65和PS80。优选地，所述第二洗涤剂是PS80。
在特定方面，使用亲和色谱以结合糖蛋白(经由其碳水化合物部分)来进行洗涤剂交换。例如，亲和色谱可以使用豆类凝集素柱。豆类凝集素是最初在植物中鉴定出的蛋白质，并且发现其与碳水化合物残基特异性地且可逆地相互作用。参见例如，Sharon和Lis,“Legume lectins--a large family of homologous proteins,”FASEB J.1990年11月；4(14):3198-208；Liener,“The Lectins:Properties,Functions,and Applications inBiology and Medicine,”Elsevier,2012。合适的凝集素包括伴刀豆球蛋白A(con A)、豌豆凝集素、红豆草凝集素和小扁豆凝集素。小扁豆凝集素由于其结合特性是用于洗涤剂交换的优选柱。凝集素柱可商购获得；例如，Capto小扁豆凝集素可从GE Healthcare获得。在某些方面，小扁豆凝集素柱可以使用重组凝集素。在分子水平上，认为碳水化合物部分与小扁豆凝集素结合，释放蛋白质的氨基酸以在洗涤剂周围聚结，导致形成洗涤剂核心，从而提供具有多个抗原拷贝(例如，糖蛋白寡聚体)的纳米颗粒，所述糖蛋白寡聚体可以是锚定在洗涤剂中的二聚体、三聚体或四聚体。在实施方案中，所述CoV S多肽形成三聚体。在实施方案中，所述CoV S多肽三聚体锚定在洗涤剂中。在实施方案中，每个CoV S多肽纳米颗粒含有至少一个与非离子型核心缔合的三聚体。
当在洗涤剂交换期间与蛋白质一起孵育形成纳米颗粒时，洗涤剂在早期纯化步骤期间可以按多至约0.1％(w/v)存在，并且使此量降低以获得具有最佳稳定性的最终纳米颗粒。例如，所述非离子型洗涤剂(例如，PS80)可以是约0.005％(v/v)至约0.1％(v/v)，例如约0.005％(v/v)、约0.006％(v/v)、约0.007％(v/v)、约0.008％(v/v)、约0.009％(v/v)、约0.01％(v/v)、约0.015％(v/v)、约0.02％(v/v)、约0.025％(v/v)、约0.03％(v/v)、约0.035％(v/v)、约0.04％(v/v)、约0.045％(v/v)、约0.05％(v/v)、约0.055％(v/v)、约0.06％(v/v)、约0.065％(v/v)、约0.07％(v/v)、约0.075％(v/v)、约0.08％(v/v)、约0.085％(v/v)、约0.09％(v/v)、约0.095％(v/v)或约0.1％(v/v)PS80。在实施方案中，所述纳米颗粒含有约0.03％至约0.05％ PS80。在实施方案中，所述纳米颗粒含有约0.01％(v/v)PS80。
在实施方案中，对经纯化的CoV S多肽进行透析。在实施方案中，在纯化后进行透析。在实施方案中，在包含磷酸钠、NaCl和PS80的溶液中对所述CoV S多肽进行透析。在实施方案中，包含磷酸钠的透析溶液含有在约5mM与约100mM之间的磷酸钠，例如约5mM、约10mM、约15mM、约20mM、约25mM、约30mM、约35mM、约40mM、约45mM、约50mM、约55mM、约60mM、约65mM、约70mM、约75mM、约80mM、约85mM、约90mM、约95mM或约100mM磷酸钠。在实施方案中，包含磷酸钠的溶液的pH为约6.5、约6.6、约6.7、约6.8、约6.9、约7.0、约7.1、约7.2、约7.3、约7.4或约7.5。在实施方案中，包含氯化钠的透析溶液包含约50mM NaCl至约500mM NaCl，例如约50mM、约60mM、约70mM、约80mM、约90mM、约100mM、约110mM、约120mM、约130mM、约140mM、约150mM、约160mM、约170mM、约180mM、约190mM、约200mM、约210mM、约220mM、约230mM、约240mM、约250mM、约260mM、约270mM、约280mM、约290mM、约300mM、约310mM、约320mM、约330mM、约340mM、约350mM、约360mM、约370mM、约380mM、约390mM、约400mM、约410mM、约420mM、约430mM、约440mM、约450mM、约460mM、约470mM、约480mM、约490mM或约500mM NaCl。在实施方案中，包含PS80的透析溶液包含约0.005％(v/v)、约0.006％(v/v)、约0.007％(v/v)、约0.008％(v/v)、约0.009％(v/v)、约0.01％(v/v)、约0.015％(v/v)、约0.02％(v/v)、约0.025％(v/v)、约0.03％(v/v)、约0.035％(v/v)、约0.04％(v/v)、约0.045％(v/v)、约0.05％(v/v)、约0.055％(v/v)、约0.06％(v/v)、约0.065％(v/v)、约0.07％(v/v)、约0.075％(v/v)、约0.08％(v/v)、约0.085％(v/v)、约0.09％(v/v)、约0.095％(v/v)或约0.1％(v/v)PS80。在实施方案中，所述透析溶液包含约25mM磷酸钠(pH 7.2)、约300mM NaCl和约0.01％(v/v)PS80。
可以使用如下蛋白质进行洗涤剂交换，将所述蛋白质如上文所讨论地纯化，并且纯化、冷冻储存，然后解冻以进行洗涤剂交换。
本文所公开的组合物的稳定性可以按多种方式测量。在一种方法中，可以制备肽图谱，以在设计成通过模拟苛刻的储存条件来胁迫纳米颗粒的各种处理后确定抗原蛋白的完整性。因此，稳定性的度量是与对照样品相比胁迫样品中抗原肽的相对丰度。例如，可以通过将含有CoV S多肽的纳米颗粒暴露于各种pH、蛋白酶、盐、氧化剂(包括但不限于过氧化氢)、各种温度、冷冻/解冻循环和搅拌来评价所述纳米颗粒的稳定性。图12A-图12B显示BV2373(SEQ ID NO:87)和BV2365(SEQ ID NO:4)在多种胁迫条件下保持与hACE2结合。认为糖蛋白锚定在洗涤剂核心中的位置通过减少不希望的相互作用而提供增强的稳定性。例如，可以通过屏蔽作用来实现针对基于蛋白酶的降解的改善的保护，由此以本文所公开的摩尔比将糖蛋白锚定在核心中导致了空间位阻，从而阻断蛋白酶进入。也可以通过监测完整蛋白质来测量稳定性。图33和图34比较了含有分别具有SEQ ID NO:109和87的氨基酸序列的CoV多肽的纳米颗粒。图34表明，具有SEQ ID NO:87的氨基酸序列的CoV S多肽在纯化期间显示出特别好的稳定性。图34的多肽包含具有QQAQ(SEQ ID NO:7)的氨基酸序列的弗林蛋白酶切割位点。
含有CoV S多肽抗原的疫苗组合物
本公开文本提供了例如在纳米颗粒中包含CoV S多肽的疫苗组合物。在一些方面，所述疫苗组合物可以含有具有抗原的纳米颗粒，所述抗原来自同一病毒物种的多于一种病毒株。在另一个实施方案中，本公开文本提供了包含一个或多个容器的药物包装或试剂盒，所述一个或多个容器装有所述疫苗组合物的一种或多种组分。
本文所公开的组合物可以预防性地或治疗性地使用，但是通常将是预防性的。因此，本公开文本包括用于治疗或预防感染的方法。所述方法涉及向受试者施用治疗量或预防量的本公开文本的免疫原性组合物。优选地，所述药物组合物是提供保护作用的疫苗组合物。在其他方面，所述保护作用可以包括在一定百分比的暴露人群中改善与感染相关的症状。例如，与未经治疗的受试者相比，所述组合物可以预防或减轻选自以下的一种或多种病毒疾病症状：发热、疲劳、肌肉疼痛、头痛、喉咙痛、呕吐、腹泻、皮疹、肾和肝功能受损的症状、内出血和外出血。
可以在存在各种赋形剂、缓冲剂等的情况下将所述纳米颗粒配制用于作为疫苗施用。例如，所述疫苗组合物可以含有磷酸钠、氯化钠和/或组氨酸。磷酸钠可以按约10mM至约50mM、约15mM至约25mM或约25mM存在；在特定情况下，存在约22mM磷酸钠。组氨酸可以按约0.1％(w/v)、约0.5％(w/v)、约0.7％(w/v)、约1％(w/v)、约1.5％(w/v)、约2％(w/v)或约2.5％(w/v)存在。氯化钠(当存在时)可以为约150mM。在某些组合物中，氯化钠可以按较高浓度(例如从约200mM至约500mM)存在。在实施方案中，氯化钠以高浓度(包括但不限于约200mM、约250mM、约300mM、约350mM、约400mM、约450mM或约500mM)存在。
在实施方案中，本文所述的纳米颗粒在某些pH水平下具有改善的稳定性。在实施方案中，所述纳米颗粒在微酸性pH水平下是稳定的。例如，所述纳米颗粒在微酸性pH(例如从pH 5.8至pH 7.0)下是稳定的。在实施方案中，所述纳米颗粒和含有纳米颗粒的组合物在范围从约pH 5.8至约pH 7.0的pH(包括约pH 5.9至约pH 6.8、约pH 6.0至约pH 6.5、约pH6.1至约pH 6.4、约pH 6.1至约pH 6.3或约pH 6.2)下可以是稳定的。在实施方案中，本文所述的纳米颗粒和组合物在中性pH(包括从约pH 7.0至约pH 7.4)下是稳定的。在实施方案中，本文所述的纳米颗粒和组合物在微碱性pH(例如从约pH 7.0至约pH 8.5、从约pH 7.0至约pH 8.0或从约pH 7.0至约pH 7.5，包括在中间的所有值和范围)下是稳定的。
佐剂
在某些实施方案中，本文所公开的组合物可以与一种或多种佐剂组合以增强免疫反应。在其他实施方案中，所述组合物是在没有佐剂的情况下制备的，因此可用来作为无佐剂组合物施用。有利地，当作为单剂施用时，本文所公开的无佐剂组合物可以提供保护性免疫反应。诱导稳健的免疫反应的无明矾组合物尤其可用于约60岁及以上的成人。
基于铝的助剂
在实施方案中，所述佐剂可以是明矾(例如AlPO4或Al(OH)3)。通常，所述纳米颗粒基本上结合至明矾。例如，所述纳米颗粒可以是至少80％结合、至少85％结合、至少90％结合或至少95％结合至明矾。通常，在组合物中，所述纳米颗粒是92％至97％结合至明矾。每剂存在的明矾的量通常在约400μg至约1250μg之间的范围内。例如，明矾可以按每剂约300μg至约900μg、约400μg至约800μg、约500μg至约700μg、约400μg至约600μg或约400μg至约500μg的量存在。通常，对于120μg蛋白纳米颗粒的剂量，明矾以约400μg存在。
皂苷佐剂
含有皂苷的佐剂也可以与本文所公开的免疫原组合。皂苷是源自皂树(Quillajasaponaria Molina)树皮的糖苷。通常，使用多步纯化过程制备皂苷，从而产生多种级分。如本文所用，术语“来自皂树的皂苷级分”一般用于描述皂树的半纯化或确定的皂苷级分或其基本上纯的级分。
皂苷级分
用于产生皂苷级分的若干种方法是合适的。级分A、B和C描述于美国专利号6,352,697中，并且可以如下制备。通过色谱法分离来自Quil A(一种粗水性皂树提取物)的亲脂级分，并且将其用水中的70％乙腈洗脱以回收亲脂级分。然后通过半制备型HPLC分离这种亲脂级分，使用在酸性水中的从25％至60％乙腈的梯度进行洗脱。在本文中称为“级分A”或“QH-A”的级分是或对应于在大约39％乙腈下洗脱的级分。在本文中称为“级分B”或“QH-B”的级分是或对应于在大约47％乙腈下洗脱的级分。在本文中称为“级分C”或“QH-C”的级分是或对应于在大约49％乙腈下洗脱的级分。关于级分纯化的另外信息可见于美国专利号5,057,540。当如本文所述制备时，皂树的级分A、B和C各自表示具有可定义特性的化学上紧密相关分子的组或家族。获得所述级分的色谱条件使得就洗脱曲线和生物活性而言批次间重现性是高度一致的。
已描述了其他皂苷级分。级分B3、B4和B4b描述于EP 0436620中。级分QA1-QA22是描述的EP03632279 B2，Q-VAC(Nor-Feed,AS丹麦)，皂树(Quillaja saponaria Molina)Spikoside(lsconova AB,Ultunaallén 2B,75651乌普萨拉,瑞典)。可以使用EP 0 3632279B2中的级分QA-1、QA-2、QA-3、QA-4、QA-5、QA-6、QA-7、QA-8、QA-9、QA-10、QA-11、QA-12、QA-13、QA-14、QA-15、QA-16、QA-17、QA-18、QA-19、QA-20、QA-21和QA-22，尤其是QA-7、QA-17、QA-18和QA-21。如EP 0 3632 279B2中(尤其是在第6页以及在第8和9页中的实施例1中)所述获得所述级分。
本文所述并用于形成佐剂的皂苷级分通常是基本上纯的级分；也就是说，所述级分基本上不存在来自其他材料的污染。在特定方面，基本上纯的皂苷级分可以含有按重量计至多40％、按重量计至多30％、按重量计至多25％、按重量计至多20％、按重量计至多15％、按重量计至多10％、按重量计至多7％、按重量计至多5％、按重量计至多2％、按重量计至多1％、按重量计至多0.5％或按重量计至多0.1％的其他化合物，诸如其他皂苷或其他佐剂材料。
ISCOM结构
皂苷级分能以称为ISCOM(免疫刺激复合物)的笼状颗粒的形式施用。ISCOM可以如EP0109942B1、EP0242380B1和EP0180546 B1中所述制备。在特定实施方案中，可以使用转运和/或乘客抗原，如EP 9600647-3(PCT/SE97/00289)中所述。
基质佐剂
在实施方案中，所述ISCOM是ISCOM基质复合物。ISCOM基质复合物包含至少一种皂苷级分和脂质。所述脂质至少是固醇，诸如胆固醇。在特定方面，所述ISCOM基质复合物还含有磷脂。所述ISCOM基质复合物还可以含有一种或多种其他免疫调节(佐剂活性)物质，不一定是糖苷，并且可以如EP0436620B1中所述产生，将所述专利通过引用以其整体并入本文。
在其他方面，所述ISCOM是ISCOM复合物。ISCOM复合物含有至少一种皂苷、至少一种脂质和至少一种抗原或表位。所述ISCOM复合物含有通过洗涤剂处理缔合的抗原，使得一部分抗原整合到颗粒中。相比之下，ISCOM基质被配制为与抗原的混合物，并且ISCOM基质颗粒与抗原之间的缔合通过静电和/或疏水相互作用来介导。
根据一个实施方案，整合到ISCOM基质复合物或ISCOM复合物中的皂苷级分或也整合到ISCOM或ISCOM基质复合物中或与其混合的至少一种另外的佐剂选自皂树的级分A、级分B或级分C、皂树的半纯化制剂、皂树的纯化制剂或任何经纯化的亚级分(例如，QA 1-21)。
在特定方面，每个ISCOM颗粒可以含有至少两种皂苷级分。可以使用不同皂苷级分的重量％的任何组合。可以使用任何两种级分的重量％的任何组合。例如，所述颗粒可以分别含有任何重量％的级分A和任何重量％的另一种皂苷级分，诸如粗皂苷级分或级分C。因此，在特定方面，每个ISCOM基质颗粒或每个ISCOM复合物颗粒可以含有按重量计0.1％至99.9％、按重量计5％至95％、按重量计10％至90％、按重量计15％至85％、按重量计20％至80％、按重量计25％至75％、按重量计30％至70％、按重量计35％至65％、按重量计40％至60％、按重量计45％至55％、按重量计40％至60％或按重量计50％的一种皂苷级分(例如级分A)以及在每种情况下其余至多100％的另一种皂苷(例如任何粗级分或任何其他级分，例如级分C)。将重量计算为皂苷级分的总重量。ISCOM基质复合物和ISCOM复合物佐剂的例子公开于美国公开的申请号2013/0129770中，将所述申请通过引用以其整体并入本文。
在特定实施方案中，所述ISCOM基质或ISCOM复合物包含按重量计5％-99％的一种级分(例如级分A)和其余按重量计至多100％的另一种级分(例如粗皂苷级分或级分C)。将重量计算为皂苷级分的总重量。
在另一个实施方案中，所述ISCOM基质或ISCOM复合物包含按重量计40％至99％的一种级分(例如级分A)和按重量计1％至60％的另一种级分(例如粗皂苷级分或级分C)。将重量计算为皂苷级分的总重量。
在又另一个实施方案中，所述ISCOM基质或ISCOM复合物包含按重量计70％至95％的一种级分(例如，级分A)和按重量计30％至5％的另一种级分(例如，粗皂苷级分或级分C)。将重量计算为皂苷级分的总重量。在其他实施方案中，来自皂树的皂苷级分选自QA 1-21中的任一种。
除了含有皂苷级分的混合物的颗粒之外，ISCOM基质颗粒和ISCOM复合物颗粒各自都可以仅使用一种皂苷级分形成。本文所公开的组合物可以含有多个颗粒，其中每个颗粒仅含有一种皂苷级分。也就是说，某些组合物可以含有一种或多种不同类型的ISCOM-基质复合物颗粒和/或一种或多种不同类型的ISCOM复合物颗粒，其中每个单独的颗粒含有来自皂树的一种皂苷级分，其中一种复合物中的皂苷级分与另一种复合物颗粒中的皂苷级分不同。
在特定方面，一种类型的皂苷级分或粗皂苷级分可以整合到一种ISCOM基质复合物或颗粒中，并且另一种类型的基本上纯的皂苷级分或粗皂苷级分可以整合到另一种ISCOM基质复合物或颗粒中。组合物或疫苗可以包含至少两种类型的复合物或颗粒，每种类型具有一种类型的整合到物理上不同的颗粒中的皂苷。
在所述组合物中，可以使用ISCOM基质复合物颗粒和/或ISCOM复合物颗粒的混合物，其中将一种皂苷级分皂树和另一种皂苷级分皂树分开掺入不同的ISCOM基质复合物颗粒和/或ISCOM复合物颗粒中。
各自具有一种皂苷级分的ISCOM基质或ISCOM复合物颗粒可以按重量％的任何组合存在于组合物中。在特定方面，组合物可以含有按重量计0.1％至99.9％、按重量计5％至95％、按重量计10％至90％、按重量计15％至85％、按重量计20％至80％、按重量计25％至75％、按重量计30％至70％、按重量计35％至65％、按重量计40％至60％、按重量计45％至55％、按重量计40至60％或按重量计50％的含有第一皂苷级分的ISCOM基质或复合物，其余部分由含有不同皂苷级分的ISCOM基质或复合物组成。在一些方面，所述其余部分是一种或多种ISCOM基质或复合物，其中每个基质或复合物颗粒仅含有一种皂苷级分。在其他方面，所述ISCOM基质或复合物颗粒可以含有多于一种皂苷级分。
在特定组合物中，第一ISCOM基质或ISCOM复合物颗粒中唯一的皂苷级分是级分A，并且第二ISCOM基质或ISCOM复合物颗粒中唯一的皂苷级分是级分C。
优选的组合物包含含有级分A的第一ISCOM基质和含有级分C的第二ISCOM基质，其中级分A ISCOM基质占总皂苷佐剂重量的约70％，并且级分C ISCOM基质占总皂苷佐剂重量的约30％。在另一种优选的组合物中，级分A ISCOM基质占总皂苷佐剂重量的约85％，并且级分C ISCOM基质占总皂苷佐剂重量的约15％。因此，在某些组合物中，级分A ISCOM基质存在的范围为所述组合物中皂苷佐剂总重量的约70％至约85％，并且级分C ISCOM基质存在的范围为约15％至约30％。在实施方案中，分别地，级分A ISCOM基质占佐剂中级分A ISCOM基质和级分C ISCOM的重量之和的按重量计50％-96％，并且级分C ISCOM基质占剩余部分。在特别优选的组合物(在本文中称为MATRIX-MTM)中，级分AISCOM基质以所述组合物中皂苷佐剂总重量的约85％存在，并且级分C ISCOM基质以约15％存在。MATRIX-MTM可以可互换地称为Matrix-M1。
示例性QS-7和QS-21级分、其产生及其用途描述于美国专利号5,057,540；6,231,859；6,352,697；6,524,584；6,846,489；7,776,343；和8,173,141中，将所述专利通过引用并入本文。
在一些组合物中，可以另外或替代地使用其他佐剂。包括在以下文献中所述的任何佐剂设想在本公开文本的范围内：Vogel等人,“A Compendiumof Vaccine Adjuvantsand Excipients(第2版),”(出于所有目的通过引用以其整体并入本文)。其他佐剂包括完全弗氏佐剂(免疫反应的非特异性刺激剂，含有杀死的结核分枝杆菌(Mycobacteriumtuberculosis))、不完全弗氏佐剂和氢氧化铝佐剂。其他佐剂包括GMCSP、BCG、MDP化合物(诸如thur-MDP和nor-MDP)、CGP(MTP-PE)、脂质A和单磷酰脂质A(MPL)、MF-59、RIBI(其含有从细菌中提取的三种组分)、MPL、海藻糖二霉菌酸酯(TDM)和在2％角鲨烯/
聚山梨醇酯80乳剂中的细胞壁骨架(CWS)。在实施方案中，所述佐剂可以是少层状(paucilamellar)脂质囊泡；例如
是范围从约100nm至约500nm的少层状非磷脂囊泡。它们包含
醇乙氧基化物72、胆固醇、油酸和角鲨烯。已显示
是有效的佐剂(参见美国专利号5,629,021、6,387,373和4,911,928)。
施用和剂量
在实施方案中，本公开文本提供了用于引发针对一种或多种冠状病毒的免疫反应的方法。在实施方案中，所述反应是针对SARS-CoV-2病毒、MERS和SARS中的一种或多种。所述方法涉及向受试者施用免疫有效量的含有纳米颗粒或含有重组CoV纤突(S)多肽的组合物。有利地，本文所公开的蛋白质诱导一种或多种特别有用的抗冠状病毒反应。
在实施方案中，将所述纳米颗粒或CoV S多肽与佐剂一起施用。在其他方面，在没有佐剂的情况下施用所述纳米颗粒或CoV S多肽。在一些方面，所述佐剂可以诸如通过非共价相互作用结合至所述纳米颗粒。在其他方面，将所述佐剂与所述纳米颗粒共同施用，但是所述佐剂和纳米颗粒基本上不相互作用。
在实施方案中，所述纳米颗粒可以用于预防和/或治疗SARS-CoV-2感染、SARS感染或MERS感染中的一种或多种。因此，本公开文本提供了用于引发针对SARS-CoV-2病毒、MERS和SARS中的一种或多种的免疫反应的方法。所述方法涉及向受试者施用免疫有效量的含有纳米颗粒或CoV S多肽的组合物。有利地，本文所公开的蛋白质诱导特别有用的抗冠状病毒反应。
本文所公开的组合物可以经由全身途径或粘膜途径或透皮途径施用或直接施用到特定组织中。如本文所用，术语“全身施用”包括肠胃外施用途径。特别地，肠胃外施用包括皮下、腹膜内、静脉内、动脉内、肌内或胸骨内注射、静脉内或肾脏透析输注技术。通常，全身肠胃外施用是肌内注射。如本文所用，术语“粘膜施用”包括口服、鼻内、阴道内、直肠内、气管内、肠和眼部施用。优选地，施用是肌内的。
可以按单剂方案或多剂方案施用组合物。可以在初次免疫方案或在加强免疫方案中使用多剂。在多剂方案中，各个剂量可以通过相同或不同的途径给予，所述途径是例如为肠胃外引发和粘膜加强、粘膜引发和肠胃外加强等。在一些方面，在先前剂量后约2周、约3周、约4周、约5周或约6周施用后续加强剂量。在实施方案中，在施用先前剂量后3周施用后续加强剂量。在实施方案中，第一剂量在第0天施用，并且加强剂量在第21天施用。在实施方案中，第一剂量在第0天施用，并且加强剂量在第28天施用。
在实施方案中，如以μg测量的剂量可以是包括溶质的剂量的总重量、或CoV S多肽纳米颗粒的重量、或CoV S多肽的重量。使用蛋白质浓度测定(A280或ELISA)来测量剂量。
包括用于小儿施用的抗原剂量可以在约5μg至约25μg、约1μg至约300μg、约90μg至约270μg、约100μg至约160μg、约110μg至约150μg、约120μg至约140μg或约140μg至约160μg的范围内。在实施方案中，所述剂量为约120μg，与明矾一起施用。在一些方面，小儿剂量可以在约1μg至约90μg的范围内。在实施方案中，CoV纤突(S)多肽的剂量为约1μg、约2μg、约3μg、约4μg、约5μg、约6μg、约7μg、约8μg、约9μg、约10μg、约11μg、约12μg、约13μg、约14μg、约15μg、约16μg、约17μg、约18μg、约19μg、约20μg、约21μg、约22μg、约23μg、约24μg、约25μg、约26μg、约27μg、约28μg、约29μg、约30μg、约40μg、约50μg、约60μg、约70μg、约80μg、约90μg、约100μg、约110μg、约120μg、约130μg、约140μg、约150μg、约160μg、约170μg、约180μg、约190μg、约200μg、约210μg、约220μg、约230μg、约240μg、约250μg、约260μg、约270μg、约280μg、约290μg或约300μg，包括在中间的所有值和范围。在实施方案中，CoV S多肽的剂量为5μg。在实施方案中，CoV S多肽的剂量为25μg。
可以在有或没有佐剂的情况下向某些人群施用。在某些方面，组合物可以不含添加的佐剂。在此类情况下，可以将剂量增加约10％。
在实施方案中，与非天然存在的CoV S多肽一起施用的佐剂的剂量为约1μg至约100μg，例如约1μg、约2μg、约3μg、约4μg、约5μg、约6μg、约7μg、约8μg、约9μg、约10μg、约11μg、约12μg、约13μg、约14μg、约15μg、约16μg、约17μg、约18μg、约19μg、约20μg、约21μg、约22μg、约23μg、约24μg、约25μg、约26μg、约27μg、约28μg、约29μg、约30μg、约31μg、约32μg、约33μg、约34μg、约35μg、约36μg、约37μg、约38μg、约39μg、约40μg、约41μg、约42μg、约43μg、约44μg、约45μg、约46μg、约47μg、约48μg、约49μg、约50μg、约51μg、约52μg、约53μg、约54μg、约55μg、约56μg、约57μg、约58μg、约59μg、约60μg、约61μg、约62μg、约63μg、约64μg、约65μg、约66μg、约67μg、约68μg、约69μg、约70μg、约71μg、约72μg、约73μg、约74μg、约75μg、约76μg、约77μg、约78μg、约79μg、约80μg、约81μg、约82μg、约83μg、约84μg、约85μg、约86μg、约87μg、约88μg、约89μg、约90μg、约91μg、约92μg、约93μg、约94μg、约95μg、约96μg、约97μg、约98μg、约99μg或约100μg的佐剂。在实施方案中，佐剂的剂量为约50μg。在实施方案中，所述佐剂是皂苷佐剂，例如MATRIX-MTM。
在实施方案中，施用的剂量的体积为约0.1mL至约1.5mL，例如约0.1mL、约0.2mL、约0.25mL、约0.3mL、约0.4mL、约0.5mL、约0.6mL、约0.7mL、约0.8mL、约0.9mL、约1.0mL、约1.1mL、约1.2mL、约1.3mL、约1.4mL或约1.5mL。在实施方案中，施用的剂量的体积为0.25mL。在实施方案中，施用的剂量的体积为0.5mL。在实施方案中，施用的剂量的体积为0.6mL。
在针对MERS、SARS或SARS-CoV-2冠状病毒的疫苗的特定实施方案中，所述剂量可以包含以下浓度的CoV S多肽：约1μg/mL至约50μg/mL、10μg/mL至约100μg/mL、约10μg/mL至约50μg/mL、约175μg/mL至约325μg/mL、约200μg/mL至约300μg/mL、约220μg/mL至约280μg/mL或约240μg/mL至约260μg/mL。
在另一个实施方案中，本公开文本提供了配制疫苗组合物的方法，所述疫苗组合物诱导针对哺乳动物的感染或其至少一种疾病症状的免疫力，所述方法包括向所述组合物中添加有效剂量的纳米颗粒或CoV S多肽。所公开的CoV S多肽和纳米颗粒可用于制备刺激免疫反应的组合物，所述免疫反应赋予针对感染原的免疫力或实质性免疫力。因此，在一个实施方案中，本公开文本提供了在受试者中诱导针对感染或其至少一种疾病症状的免疫力的方法，其方法包括施用至少一个有效剂量的纳米颗粒和/或CoV S多肽。
在实施方案中，将所述CoV S多肽或包含所述多肽的纳米颗粒与另外的免疫原性组合物组合施用。在实施方案中，所述另外的免疫原性组合物诱导针对SARS-CoV-2的免疫反应。在实施方案中，将所述另外的免疫原性组合物在所公开的CoV S多肽或包含所述多肽的纳米颗粒的约1分钟、约5分钟、约10分钟、约20分钟、约30分钟、约40分钟、约50分钟、约1小时、约2小时、约3小时、约4小时、约5小时、约6小时、约7小时、约8小时、约9小时、约10小时、约11小时、约12小时、约13小时、约14小时、约15小时、约16小时、约17小时、约18小时、约19小时、约20小时、约21小时、约22小时、约23小时、约1天、约2天、约3天、约4天、约5天、约6天、约7天、约8天、约9天、约10天、约11天、约12天、约13天、约14天、约15天、约16天、约17天、约18天、约19天、约20天、约21天、约22天、约23天、约24天、约25天、约26天、约27天、约28天、约29天、约30天或约31天内施用。在实施方案中，将所述另外的组合物与第一剂量的包含CoV S多肽或含有所述多肽的纳米颗粒的组合物一起施用。在实施方案中，将所述另外的组合物与加强剂量的包含CoV S多肽或含有所述多肽的纳米颗粒的组合物一起施用。
在实施方案中，所述另外的免疫原性组合物包含编码SARS-CoV-2纤突糖蛋白的mRNA、编码SARS-CoV-2纤突糖蛋白的质粒DNA、编码SARS-CoV-2纤突糖蛋白的病毒载体或灭活的SARS-CoV-2病毒。
在实施方案中，所述另外的免疫原性组合物包含编码CoV S多肽的mRNA。在实施方案中，所述mRNA编码在SEQ ID NO:1的位置986和987处包含脯氨酸取代的CoV S多肽。在实施方案中，所述mRNA编码包含完整弗林蛋白酶切割位点的CoV S多肽。在实施方案中，所述mRNA编码包含在SEQ ID NO:1的位置986和987处的脯氨酸取代和完整弗林蛋白酶切割位点的CoV S多肽。在实施方案中，所述mRNA编码包含在SEQ ID NO:1的位置986和987处的脯氨酸取代和无活性的弗林蛋白酶切割位点的CoV S多肽。在实施方案中，所述mRNA编码具有SEQ ID NO:87的氨基酸序列的CoV S多肽。在实施方案中，编码CoV S多肽的mRNA被封装在脂质纳米颗粒中。包含编码CoV S多肽的mRNA的示例性免疫原性组合物描述于Jackson等人N.Eng.J.Med.2020.An mRNAVaccine against SARS-CoV-2-preliminary report中，将所述文献通过引用以其整体并入本文。在实施方案中，将包含编码CoV S多肽的mRNA的组合物以25μg、100μg或250μg的剂量施用。
在实施方案中，所述另外的免疫原性组合物包含编码CoV S多肽的腺病毒载体。在实施方案中，所述AAV载体编码野生型CoV S多肽。在实施方案中，所述AAV载体编码包含在SEQ ID NO:1的位置986和987处的脯氨酸取代和完整弗林蛋白酶切割位点的CoV S多肽。在实施方案中，所述AAV载体编码包含在SEQ ID NO:1的位置986和987处的脯氨酸取代和无活性的弗林蛋白酶切割位点的CoV S多肽。在实施方案中，所述AAV载体编码具有SEQ ID NO:87的氨基酸序列的CoV S多肽。以下出版物描述了包含编码CoV S多肽的腺病毒载体的免疫原性组合物，将其中的每个出版物通过引用以其整体并入本文：van Doremalen N.等人Asingle dose of ChAdOx1 MERS provides protective immunity in rhesusmacaques.Science Advances,2020；van Doremalen N.等人ChAdOx1 nCoV-19vaccinationprevents SARS-CoV-2pneumonia in rhesus macaques.bioRxiv,(2020)。
在实施方案中，所述另外的免疫原性组合物包含脱氧核糖核酸(DNA)。在实施方案中，所述另外的免疫原性组合物包含质粒DNA。在实施方案中，所述质粒DNA编码CoV S多肽。在实施方案中，所述DNA编码包含在SEQ ID NO:1的位置986和987处的脯氨酸取代和完整弗林蛋白酶切割位点的CoV S多肽。在实施方案中，所述DNA编码包含在SEQ ID NO:1的位置986和987处的脯氨酸取代和无活性的弗林蛋白酶切割位点的CoV S多肽。在实施方案中，所述DNA编码具有SEQ ID NO:87的氨基酸序列的CoV S多肽。
在实施方案中，所述另外的免疫原性组合物包含灭活的病毒疫苗。
在实施方案中，所述CoV S蛋白或包含CoV S蛋白的纳米颗粒可用于制备免疫原性组合物以刺激赋予针对MERS、SARS和SARS-CoV-2中的一种或多种的免疫力或实质性免疫力的免疫反应。粘膜免疫力和细胞免疫力都可以有助于针对感染和疾病的免疫力。上呼吸道局部分泌的抗体是抵抗天然感染的主要因子。分泌型免疫球蛋白A(sIgA)参与保护上呼吸道，并且血清IgG参与保护下呼吸道。感染诱导的免疫反应预防相同病毒或抗原性相似的病毒株再感染。用本文所公开的纳米颗粒免疫后在宿主中产生的抗体也可以施用于其他宿主，从而在受试者中提供被动施用。
在实施方案中，所述CoV S蛋白或包含CoV S蛋白的纳米颗粒诱导针对含有具有选自以下的一个或多个修饰的S蛋白的SARS-CoV-2病毒的交叉中和抗体：
(a)NTD的一个或多个氨基酸的缺失，其中所述一个或多个氨基酸选自氨基酸56、57、131、132、229、230、231或其组合；以及
(b)NTD的一个或多个氨基酸的突变，其中所述一个或多个突变选自氨基酸67、229、202、139、5、233、7、13、125、177或其组合；
(c)RBD的一个或多个氨基酸的突变，其中所述一个或多个突变选自氨基酸488、404、471、439、426、440及其组合；
(d)SD1/2的一个或多个氨基酸的突变，其中所述一个或多个氨基酸选自601、557、668、642及其组合；
(e)无活性的弗林蛋白酶切割位点(对应于氨基酸669-672中的一个或多个突变)；
(f)S2亚基的一个或多个氨基酸的缺失，其中所述氨基酸选自676-702、702-711、775-793、806-815及其组合；
(g)S2亚基的一个或多个氨基酸的突变，其中所述氨基酸选自973、974、703、1105、688、969和1014及其组合；
(h)一个或多个氨基酸从TMCT(氨基酸1201-1260)中的缺失，其中所述CoV S糖蛋白的氨基酸相对于SEQ ID NO:2来编号。
在实施方案中，所述CoV S蛋白或包含CoV S蛋白的纳米颗粒诱导针对含有具有选自以下的一个或多个修饰的S蛋白的SARS-CoV-2病毒的交叉中和抗体：氨基酸56的缺失、氨基酸57的缺失、氨基酸131的缺失、N488Y、A557D、D601G、P668H、T703I、S969A、D1105H、N426K和Y440F，其中所述氨基酸相对于具有SEQ ID NO:2的氨基酸序列的CoV S多肽来编号。
在实施方案中，本公开文本提供了产生高亲和力抗MERS-CoV、抗SARS-CoV和抗SARS-CoV-2病毒抗体中的一种或多种的方法。通过用本文所公开的纳米颗粒免疫而产生的高亲和力抗体通过以下方式来产生：向动物施用包含S CoV多肽或含有S CoV多肽的纳米颗粒的免疫原性组合物，从所述动物中收集血清和/或血浆，并且从血清/和或血浆中纯化抗体。在一个实施方案中，所述动物是人。在实施方案中，所述动物是鸡、小鼠、豚鼠、大鼠、兔、山羊、人、马、羊或牛。在一个实施方案中，所述动物是牛或马。在另一个实施方案中，所述牛或马动物是转基因的。在又另一个实施方案中，所述转基因牛或马动物产生人抗体。在实施方案中，所述动物产生单克隆抗体。在实施方案中，所述动物产生多克隆抗体。在一个实施方案中，所述方法还包括施用佐剂或免疫刺激化合物。在另一个实施方案中，向人类受试者施用经纯化的高亲和力抗体。在一个实施方案中，所述人类受试者有感染MERS、SARS和SARS-CoV-2中的一种或多种的风险。
在实施方案中，将所述CoV S蛋白或纳米颗粒与流感糖蛋白或包含流感糖蛋白的纳米颗粒共同施用。合适的糖蛋白和纳米颗粒描述于美国公开号2018/0133308和美国公开号2019/0314487中，将其中的每个公开案通过引用以其整体并入本文。在实施方案中，将所述CoV S蛋白或纳米颗粒与以下各项共同施用：(a)洗涤剂-核心纳米颗粒，其中所述洗涤剂-核心纳米颗粒包含来自B型流感毒株的重组流感血凝素(HA)糖蛋白；和(b)血凝素皂苷基质纳米颗粒(HaSMaN)，其中所述HaSMaN包含来自A型流感毒株的重组流感HA糖蛋白和ISCOM基质佐剂。在实施方案中，将所述CoV S蛋白或纳米颗粒与包含非离子型洗涤剂核心和流感HA糖蛋白的纳米颗粒共同施用，其中所述流感HA糖蛋白含有从非离子型洗涤剂核心向外突出的头部区域和与非离子型洗涤剂核心缔合的跨膜结构域，其中所述流感HA糖蛋白是HA0糖蛋白，其中所述流感HA糖蛋白的氨基酸序列与天然流感HA蛋白的氨基酸序列具有100％同一性。在实施方案中，将所述流感糖蛋白或纳米颗粒与所述CoV S蛋白或纳米颗粒共同配制。
将本公开文本中引用的所有专利、专利申请、参考文献和期刊文章出于所有目的通过引用以其整体明确地并入本文。
实施例
实施例1
冠状病毒纤突(S)多肽纳米颗粒的表达和纯化
已在杆状病毒表达系统中表达了天然冠状病毒纤突(S)多肽(SEQ ID NO:1和SEQID NO:2)和CoV纤突多肽(其具有对应于SEQ ID NO:3、4、38、41、44、48、51、54、58、61、63、65、67、73、75、78、79、82、83、85、87、106、108和89的氨基酸序列)，并且挑选并确认了表达冠状病毒纤突(S)多肽的重组噬斑。在每种情况下，信号肽是SEQ ID NO:5。图4和图9示出了CoV纤突多肽BV2364、BV2365、BV2366、BV2367、BV2368、BV2369、BV2373、BV2374和BV2375的成功纯化。表2示出了上述CoV纤突多肽的序列特征。
表2：选定的CoV纤突多肽
野生型BV2361蛋白(SEQ ID NO:2)与人血管紧张素转化酶2前体(hACE2)结合。进行了生物膜层干涉测量术和ELISA，以评估CoV S多肽的结合。
生物膜层干涉测量术(BLI)：
使用Octet QK384系统(Pall Forté Bio，加利福尼亚州弗里蒙特)进行了BLI实验。将His标记的人ACE2(2μg mL-1)固定在带镍的Ni-NTA生物传感器尖端上。在基线后，将含SARS-CoV-2S蛋白的样品连续稀释2倍，并且允许其缔合600秒，然后再使其解离900秒。使用Octet软件HT 101:1全局曲线拟合来分析数据。
CoV S多肽BV2361、BV2365、BV2369、BV2365、BV2373、BV2374保留与hACE2结合的能力(图5、图11A-图11C)。解离动力学表明，在不存在流体相S蛋白的情况下，如通过观察900秒的最小解离或无解离所证实的，S蛋白保持紧密结合(图11A-图11C)。
此外，结合是特异性的。野生型CoV S蛋白BV2361以及CoV S多肽BV2365和BV2373不结合MERS-CoV受体二肽基肽酶IV(DPP4)。此外，MERS S蛋白不与人血管紧张素转化酶2前体(hACE2)结合(图6和图11D-图11F)。
ELISA
通过ELISA确认了CoV S多肽对hACE2的特异性。将96孔板在4℃下用100μLSARS-CoV-2纤突蛋白(2μg/mL)包被过夜。将板用含0.05％Tween的磷酸盐缓冲盐水(PBS-T)缓冲液洗涤，并且用TBS Startblock封闭缓冲液(ThermoFisher,Scientific)封闭。将His标记的hACE2和hDPP4受体连续稀释3倍(5-0.0001μg mL-1)，并且在室温下将其添加到包被的孔中持续2小时。将板用PBS-T洗涤。添加最适稀释的辣根过氧化物酶(HRP)缀合的抗组氨酸，并且通过添加3,3’,5,5’-四甲基联苯胺过氧化物酶底物(TMB，T0440-IL，Sigma，美国密苏里州圣路易斯)来显色。用SpectraMax Plus板读取器(Molecular Devices，美国加利福尼亚州桑尼维尔)在450nm的OD下读取板，并且用SoftMax软件分析数据。使用GraphPad Prism7.05软件通过4参数拟合来计算EC50值。
ELISA结果表明，野生型CoV S多肽(BV2361)、BV2365和BV2373蛋白特异性地结合hACE2，但未能结合由MERS-CoV使用的hDPP-4受体(IC50>5000ng mL-1)。野生型CoV S多肽和BV2365以相似的亲和力结合至hACE2(IC50＝36-38ng/mL)，而BV2373在低2倍的浓度下达到50％的hACE2结合的饱和(IC50＝18ng/mL)(图7、图11D-图11F)。
蛋白质和纳米颗粒产生
通过感染Sf9昆虫细胞来扩增重组病毒。在约3MOI(感染复数＝病毒ffu或pfu/细胞)下用杆状病毒感染昆虫细胞培养物。感染后48-72小时收获培养物和上清液。通过以大约800x g离心15分钟来澄清大约30mL的粗细胞收获物。如下文所述，将含有冠状病毒纤突(S)蛋白的所得粗细胞收获物纯化为纳米颗粒。
为了产生纳米颗粒，将非离子型表面活性剂
壬基酚乙氧基化物NP-9用于膜蛋白提取方案中。经由通过阴离子交换色谱、小扁豆凝集素亲和/HIC和阳离子交换色谱进一步纯化粗提取物。通过洗涤剂处理使洗涤过的细胞裂解，然后对其进行低pH处理，这导致BV和Sf9宿主细胞DNA和蛋白质沉淀。使经中和的低pH处理裂解液澄清，并且在进行第二次低pH处理之前在阴离子交换和亲和色谱上进一步纯化。
使用亲和色谱来去除Sf9/BV蛋白、DNA和NP-9，并且浓缩冠状病毒纤突(S)蛋白。简言之，小扁豆凝集素是一种含有钙和锰的金属蛋白，其可逆地结合含有葡萄糖或甘露糖的多糖和糖基化蛋白。将含冠状病毒纤突(S)蛋白的阴离子交换流过级分加载到小扁豆凝集素亲和色谱树脂(Capto小扁豆凝集素，GE Healthcare)上。糖基化冠状病毒纤突(S)蛋白选择性地结合至树脂，而非糖基化蛋白和DNA在柱流过物中被去除。通过含有高盐和低摩尔浓度的甲基α-D-吡喃甘露糖苷(MMP)的缓冲液去除弱结合的糖蛋白。
还使用柱洗涤物来将NP-9洗涤剂用表面活性剂聚山梨醇酯80(PS80)进行洗涤剂交换。用高浓度的MMP将冠状病毒纤突(S)多肽以纳米颗粒结构从小扁豆凝集素柱中洗脱。在洗脱后，将冠状病毒纤突(S)蛋白三聚体组装成由包含在洗涤剂核心中的冠状病毒纤突(S)蛋白三聚体和PS80构成的纳米颗粒。
实施例2
冠状病毒纤突(S)多肽纳米颗粒疫苗在小鼠中的免疫原性
使用雌性BALB/c小鼠(7-9周龄；Harlan Laboratories Inc.，马里兰州弗雷德里克)评价了如实施例1所述的包含SEQ ID NO:87的CoV S多肽(也称为“BV2373”)的冠状病毒纤突(S)蛋白组合物在鼠类模型中的免疫原性和毒性。在存在和不存在皂苷佐剂(例如，MATRIX-MTM)的情况下评价了组合物。含有MATRIX-MTM的组合物含有5μg的MATRIX-MTM。将含有不同剂量(包括0.01μg、0.1μg、1μg和10μg)的冠状病毒纤突(S)多肽的疫苗作为单剂(也称为单次引发剂量)(研究第14天)或作为间隔14天(研究第0天和第14天)的两剂(也称为引发/加强方案)肌内施用。安慰剂组用作非免疫对照。在研究第-1天、第13天、第21天和第28天收集血清进行分析。在一次(单剂)或两次(两剂)免疫后42天，用SARS-CoV-2鼻内攻击接种疫苗的动物和对照动物。
疫苗免疫原性
用0.1-10μg BV2373和MATRIX-MTM的单次引发剂量免疫的动物具有升高的抗S IgG滴度，其在单次免疫后21-28天检测到(图13B)。用10μg剂量的BV2373和MATRIX-MTM免疫的小鼠产生了阻断hACE2受体与CoV S蛋白结合的抗体和在单次引发剂量后21-28天检测到的病毒中和抗体(图14和图15)。用引发/加强方案(两剂)免疫的动物具有显著升高的抗S IgG滴度，其在所有剂量水平下在加强免疫后7-16天均检测到(图13A)。用BV2373(1μg和10μg)和MATRIX-MTM免疫的动物在免疫后具有相似的高抗S IgG滴度(分别地，GMT＝139,000和84,000)。与在没有佐剂的情况下用10μg BV2373免疫的小鼠相比，用BV2373(0.1μg、1μg或10μg)和MATRIX-MTM免疫的小鼠具有显著(p≤0.05和p≤0.0001)更高的抗S IgG滴度(图13A)。这些结果表明了由MATRIX-MTM佐剂提供10倍至100倍剂量节省的可能性。此外，用两剂的BV2373和MATRIX-MTM免疫产生了高滴度抗体，其在所有剂量水平下均阻断hACE2受体与S蛋白结合(IC50＝218-1642)并中和SARS-CoV-2对Vero E6细胞的致细胞病变效应(CPE)(CPE的100％阻断＝7680-20,000)(图14和图15)。
SARS CoV-2攻击
为了评价保护性免疫力的诱导，用SARS-CoV-2攻击了经免疫的小鼠。由于小鼠不支持野生型SARS-CoV-2病毒的复制，因此在初始疫苗接种后第52天，用表达hACE2的腺病毒(Ad/hACE2)鼻内感染小鼠以允许其复制。向小鼠鼻内接种1.5x 105pfu的SARS-CoV-2，在各鼻孔之间分50μL。在感染当天称量受攻击的小鼠，并且在感染后每天称重持续7天。在感染后4天和7天，处死来自疫苗接种组和对照组各自的5只小鼠，并且收获肺并准备进行肺组织学检查。
通过噬斑测定对病毒滴度进行了定量。简言之，使用1.0mm玻璃珠(SigmaAldrich)和Beadruptor(Omini International Inc.)在PBS中使所收获的肺均质化。将匀浆添加到Vero E6近汇合培养物中，并且使用6点稀释曲线通过计数噬斑形成单位(pfu)来确定SARS-CoV-2病毒滴度。
在感染后4天，经安慰剂治疗的小鼠具有104SARS-CoV-2pfu/肺，而在没有MATRIX-MTM的情况下用BV2363免疫的小鼠具有103pfu/肺(图16)。BV2373加MATRIX-MTM仅引发组的小鼠表现出病毒滴度的剂量依赖性降低，其中在感染后第4天，10μg BV2373剂量的接受者没有可检测的病毒。与接种安慰剂的小鼠相比，接受1μg、0.1μg和0.01μg BV2373剂量的小鼠均显示出滴度显著降低。在引发/加强组中，用10μg、1μg和0.1μg剂量免疫的小鼠具有几乎不可检测的肺病毒载量，而0.01μg组相对于安慰剂动物显示降低1个对数的降低。
体重减轻与病毒载量的发现结果平行。与未接种疫苗的安慰剂动物相比，接受单剂的BV2373(0.1μg、1μg和10μg)和MATRIX-MTM的动物显示出显著预防体重减轻(图17A)。在所有剂量水平下，接受引发和加强剂量加佐剂的小鼠也均展现出显著预防体重减轻(图17B-图17C)。评价了佐剂的存在对预防体重减轻的作用。相对于安慰剂，接受引发/加强(两剂)+佐剂的小鼠显著被保护免于体重减轻，而在没有佐剂的情况下免疫的组则没有(图17C)。这些结果表明，BV2373赋予预防SARS-CoV-2，并且与较低的血清学反应相关的低剂量疫苗不会加剧体重减轻或展现出疾病加重。
在感染后第4天和第7天评价了肺组织病理学(图18A和图18B)。在感染后第4天，经安慰剂免疫的小鼠显示出在大气道中的上皮细胞剥蚀以及由混合炎性细胞群包围的肺泡隔增厚。在整个肺中观察到外周小动脉成套有炎症细胞，其主要由嗜中性粒细胞和巨噬细胞组成。到感染后第7天，经安慰剂治疗的小鼠显示细支气管周炎症和外周小动脉成套增加。肺泡隔增厚在整个肺泡隔中仍伴有弥漫性间质性炎症增加(图18B)。
经BV2373免疫的小鼠在感染后第4天和第7天都显示出肺部病理以剂量依赖性方式显著降低。与安慰剂小鼠相比，仅引发组在10μg和1μg剂量下显示炎症减少，其中支气管和小动脉周围炎症减少。在较低剂量的仅引发组中，肺部炎症类似于安慰剂组，与体重减轻和肺部病毒滴度相关。对于所有测试剂量，引发/加强免疫组都显示肺部炎症显著降低，这再次与肺部病毒滴度和体重减轻数据相关。在第4天和第7天，基本上保留大支气管和小支气管中的上皮细胞，其中细支气管脱落和病毒感染迹象最小。用10μg、1μg和0.1μg剂量免疫的动物的小动脉具有最少的炎症，其中在0.01μg剂量的情况下仅观察到适度的成套，类似于安慰剂。在接受较高剂量的动物中肺泡炎症减少，其中仅较低的0.01μg剂量与炎症相关(图18A-图18B)。这些数据证明，BV2373减少攻击后的肺部炎症，并且甚至引起最小或没有引起可检测的中和活性的BV2373的剂量和方案也与对病毒的炎症反应加剧不相关。此外，疫苗不会在受攻击的小鼠中引起疫苗相关的增强型呼吸系统疾病(VAERD)。
T细胞反应
评价了包含SEQ ID NO:87的CoV S多肽的疫苗组合物对T细胞反应的影响。以间隔21天的2剂用10μg BV2373加或不加5μg MATRIX-MTM肌内免疫BALB/c小鼠(每组N＝6)。在第二次免疫后7天(研究第28天)收集脾脏。未接种疫苗的组(N＝3)用作对照。
通过ELISPOTTM酶联免疫吸附测定和细胞内细胞因子染色(ICCS)测量来自第二次免疫后7天(研究第28天)收集的脾脏的抗原特异性T细胞反应。与单独的BV2373相比，在用BV2373和MATRIX-MTM免疫的小鼠的脾脏中，离体刺激后IFN-γ分泌性细胞的数量增加了20倍(p＝0.002)，如通过ELISPOTTM测定所测量的(图19)。为了分开检查CD4+和CD8+T细胞反应，与表面标记物染色组合进行了ICCS测定。所示出的数据在CD44hi CD62L-效应记忆T细胞群上进行门控。与在没有佐剂的情况下免疫的小鼠相比，在来自用BV2373免疫的小鼠的脾脏中IFN-γ+、TNF-α+和IL-2+细胞因子分泌性CD4+和CD8+T细胞的频率显著更高(p<0.0001)(图20A-图20C和图21A-图21C)。此外，与在不存在佐剂的情况下免疫的小鼠相比，在来自经BV2373/MATRIX-MTM免疫的小鼠的脾脏中同时产生至少两种或三种细胞因子的多功能性CD4+和CD8+T细胞的频率也显著增加(p<0.0001)(图20D-图20E和图21D-图21E)。用BV2373/MATRIX-MTM免疫在CD4+和CD8+T细胞群内都产生了更高比例的多功能表型(例如，分泌IFN-γ、TNF-α和IL-2中的多于一种的T细胞)。在记忆CD4+T细胞中检测到的多功能表型的比例高于在CD8+T细胞中的比例(图22)。
还分别通过ICCS和ELISPOTTM测定了CD4+T细胞的2型细胞因子IL-4和IL-5分泌。与单独用BV2373免疫相比，用BV2373/MATRIX-MTM免疫还增加了2型细胞因子IL-4和IL-5分泌(2倍)，但程度小于1型细胞因子产生的增强(例如IFN-γ增加20倍)(图23A-图23C)。这些结果表明，MATRIX-MTM佐剂的施用使CD4+T细胞偏向产生Th1反应。
通过测量脾脏中CD4+滤泡辅助性T(TFH)细胞和生发中心(GC)B细胞的频率来评估免疫对生发中心形成的影响。MATRIX-MTM施用显著增加了脾脏中TFH细胞(CD4+CXCR5+PD-1+)的频率(p＝0.01)以及GC B细胞(CD19+GL7+CD95+)的频率(p＝0.0002)(图24A-图24B和图25A-图25B)。
实施例3
冠状病毒纤突(S)多肽纳米颗粒疫苗在橄榄狒狒中的免疫原性
评估了包含BV2373的疫苗组合物在狒狒中的免疫原性。通过肌内(IM)注射以间隔21天的两剂施用剂量范围(1μg、5μg和25μg)的BV2373和50μg MATRIX-MTM佐剂来免疫成年橄榄狒狒。为了评估MATRIX-MTM在非人灵长类动物中的辅助活性，在没有MATRIX-MTM的情况下用25μg的BV2373免疫另一组动物。在所有剂量水平下，在用BV2373/MATRIX-MTM免疫的动物中，在单次引发免疫的21天内均检测到抗S蛋白IgG滴度(GMT＝1249-19,000)。在所有剂量水平下，加强免疫后1至2周(第28天和第35天)内，抗S蛋白IgG滴度均增加超过一个对数(GMT＝33,000-174,000)。(图26A)。
用BV2373(5μg或25μg)和MATRIX-MTM单次免疫后，在动物中检测到低水平的hACE2受体阻断抗体(GMT＝22-37)。在用BV2373/MATRIX-MTM免疫的所有组中，在加强免疫的一至二周内，受体阻断抗体滴度均显著增加(GMT＝150-600)(图26B)。在用BV2373/MATRIX-MTM单次免疫后，在所有剂量组中，病毒中和抗体均升高(GMT＝190-446)。单独用25μg BV2373免疫的动物没有可检测的阻断S蛋白与hACE2结合的抗体(图26C)。加强免疫后一周，中和滴度增加6倍至8倍(GMT＝1160-3846)。第二次免疫后，中和滴度再增加25倍至38倍(GMT＝6400-17,000)(图26C)。抗S IgG水平与中和抗体滴度之间存在显著相关性(p<0.0001)(图27)。加佐剂的疫苗在非人灵长类动物中的免疫原性与实施例2的结果一致，并且进一步支持MATRIX-MTM在促进中和抗体的产生和剂量节省中的作用。
在第二次免疫后7天(第28天)收集PBMC，并且通过ELISPOT测定测量了T细胞反应。来自用BV2373(5μg或25μg)和MATRIX-MTM免疫的动物的PBMC具有最高数量的IFN-γ分泌性细胞，其是单独用25μg BV2373或BV2373(1μg)和MATRIX-MTM免疫的动物的所述数量的5倍(图28)。通过ICCS分析，用BV2373(5μg)和MATRIX-MTM免疫显示出IFN-γ+、IL-2+和TNF-α+CD4+T细胞的最高频率(图29A-图29C)。这种趋势对于同时产生至少两种或三种1型细胞因子的多功能性CD4+T细胞也是如此(图29D-图29E)。
实施例4.
冠状病毒纤突(S)多肽纳米颗粒疫苗的结构表征
使用透射电子显微术(TEM)和二维(2D)类别取平均值来确定BV2373的超微结构。负染色的BV2373的高放大率(67,000倍和100,000倍)TEM图像显示出对应于S蛋白同三聚体的颗粒。
使用自动挑选方案来构建2D类别平均图像(Lander G.C.等人J StructBiol.166,95-102(2009)；Sorzano C.O.等人,J Struct Biol.148,194-204(2004).)。对同三聚体结构进行的两轮2D类别取平均值揭示出了具有15nm长度和13nm宽度的三角形颗粒外观(图10，左上方)。将最近解析的SARS-CoV-2纤突蛋白(EMD ID：21374)的cryoEM结构覆盖在2D BV2373图像上显示出与冠形S1(NTD和RBD)和S2茎的良好符合(图10，左下方)。在2D图像中同样明显的是从与NTD/RBD冠相对的三聚体结构的尖端突出的微弱投影(图10，右上方)。使用较大盒尺寸进行的2D类别取平均值表明，这些微弱的投影形成了S三聚体与无定形结构之间的连接。(图10，右下方)。
动态光散射(DLS)表明，与BV2365(33.4nm)和BV2373(27.2nm)的小2倍的粒度相比，野生型CoV S蛋白具有69.53nm的Z-avg粒径。多分散性指数(PDI)表明，与野生型纤突蛋白(PDI＝0.46)相比，BV2365和BV2373颗粒在尺寸、形状和质量方面总体上是均匀的(PDI＝0.25-0.29)(表3)。
表3：SARS-CoV-2三聚体纤突蛋白的粒度和热稳定性
通过差示扫描量热法(DSC)确定了S三聚体的热稳定性。野生型CoV S蛋白的热转变温度(Tmax＝58.6℃)与分别为Tmax＝61.3℃和60.4℃的BV2365和BV2373相似(表3)。更重要的是，与展开WT纤突蛋白所需的较低焓(ΔHcal＝153kJ/mol)相比，展开BV2365和BV2373变体所需的转变焓增加3-5倍(分别为ΔHcal＝466和732kJ/mol)。这些结果与和WT纤突蛋白的热稳定性相比BV2365和BV2373的热稳定性得到改善一致(表3)。
通过动态光散射评价了CoV纤突(S)多肽纳米颗粒疫苗的稳定性。使用各种pH、温度、盐浓度和蛋白酶来比较CoV纤突(S)多肽纳米颗粒疫苗与含有天然CoV纤突(S)多肽的纳米颗粒疫苗的稳定性。
实施例5.
冠状病毒纤突(S)多肽纳米颗粒疫苗的稳定性
通过动态光散射评价了CoV纤突(S)多肽纳米颗粒疫苗的稳定性。使用各种pH、温度、盐浓度和蛋白酶来比较CoV纤突(S)多肽纳米颗粒疫苗与含有天然CoV纤突(S)多肽的纳米颗粒疫苗的稳定性。在不同的环境胁迫条件下使用hACE2捕获ELISA评估了没有2-脯氨酸取代的BV2365和具有两个脯氨酸取代的BV2373的稳定性。将BV2373在极端pH下(在pH 4和pH 9下48小时)、伴随长时间搅拌(48小时)和通过冷冻/解冻(2个循环)以及在升高的温度下(在25℃和37℃下48小时)孵育对hACE2受体结合没有影响(IC50＝14.0-18.3ng mL-1)。
使用过氧化氢的氧化条件将hACE2与BV2373的结合降低了8倍(IC50＝120ng mL-1)(图12A)。没有2-脯氨酸取代的BV2365不太稳定，如通过在多种条件下hACE2结合的显著损失所确定的(图12B)。
比较了BV2384(SEQ ID NO:110)和BV2373(SEQ ID NO:87)的稳定性。BV2384具有GSAS(SEQ ID NO:97)的弗林蛋白酶切割位点序列，而BV2373具有QQAQ(SEQ ID NO:7)的弗林蛋白酶切割位点。如通过SDS-PAGE和蛋白质印迹所展现的，与BV2373相比，BV2384显示出广泛的降解(图32)。此外，与BV2373(图34)相比，扫描光密度测定法和回收率数据展现了全长CoV S蛋白BV2384的意外损失、较低的纯度和回收率(图33)。
实施例6
在食蟹猴中的免疫反应
评估了BV2373在食蟹猴SARS-CoV-2感染模型中诱导的免疫反应。第1-6组按表4所示进行治疗。
表4：食蟹猴研究的第1-6组
施用包含BV2373的疫苗导致诱导抗CoV-S抗体(图35A)，包括中和抗体(图35B)。在施用一剂(图38A)或两剂(图38B)的BV2373后诱导了抗CoV-S抗体。施用包含BV2373的疫苗还导致产生阻断CoV S蛋白与hACE2结合的抗体(图38C和图38D)。在施用BV2373后，食蟹猴中抗CoV S多肽IgG滴度与hACE2抑制滴度之间存在显著相关性(图38E)。通过致细胞病变效应(CPE)(图40A)和噬斑减少中和测试(PRNT)(图40B)评价了BV2373诱导中和抗体产生的能力。数据揭示，与对照相比，表4的疫苗配制品产生了SARS-CoV-2中和滴度。
将包含能够在食蟹猴中诱导抗CoV-S抗体和阻断hACE2与CoV S蛋白结合的抗体的BV2373的疫苗与人恢复期血清进行了比较。数据揭示，与人恢复期血清相比，BV2373疫苗配制品诱导了优异的抗CoV S多肽和hACE2抑制滴度(图39)。
BV2373疫苗配制品还引起了SARS-CoV-2病毒复制的减少(图36A-图36B)。在用感染性病毒攻击后2天和4天(d2pi和d4pi)，在支气管灌洗液(BAL)中评估病毒RNA(图36A，对应于存在的总RNA)和病毒亚基因组RNA(sgRNA)(图36B，对应于复制病毒)水平。大多数受试者未显示出病毒RNA。在第2天，在一些受试者中测量到少量RNA。到第4天，除了接受2.5μg最低剂量的两名受试者以外，都未测量到RNA。除了同样接受最低剂量的1名受试者以外，在2天或4天均未检测到亚基因组RNA。在感染后2天和4天(d2pi和d4pi)通过鼻拭子评估病毒RNA(图37A)和病毒亚基因组(sg)RNA(图37B)。大多数受试者未显示出病毒RNA。在第2天和第4天，在一些受试者中测量到少量RNA。在2天或4天均未检测到亚基因组RNA。受试者在第0天进行免疫，并且在用两剂的组中在第0天和第21天进行免疫。这些数据表明，疫苗将鼻总病毒RNA降低100-1000倍，并将sgRNA降低至不可检测的水平，并且确认对疫苗的免疫反应将阻断病毒复制并防止病毒传播。
实施例7
CoV S多肽纳米颗粒疫苗在人中的评价
在131名18-59岁健康参与者的随机、观察者盲性、安慰剂对照的1期临床试验中评估了包含BV2373的疫苗的安全性和功效。参与者进行了两次肌内注射免疫，间隔21天。参与者接受加或不加MATRIX-MTM的BV2373(n＝106)或安慰剂(n＝25)。组A-E按表5所示进行治疗。图41示出了临床终点的评价的时间线。
表5：1期人类研究的组A-E
总体反应原性是轻度的，并且疫苗接种耐受性良好。在用BV2373和MATRIX-MTM治疗的患者中，局部反应原性更为常见(图42A-图42B)。
评价了加或不加MATRIX-MTM的BV2373的免疫原性。疫苗接种后21天，所有疫苗方案均检测到抗CoV-S抗体(图43A)。包含MATRIX-MTM的疫苗方案的几何平均倍数上升(GMFR)超过了未加佐剂的BV2373诱导的几何平均倍数上升。第二次疫苗接种后7天(第28天)，抗CoV-S滴度相对于第一次疫苗接种时观察到的反应再增加了八倍，并且在14天(第35天)内反应又再次增加超过一倍，达到GMFR相对于用单独BV2373观察到的GMFR是大约100倍。用BV2373/MATRIX-MTM单次疫苗接种达到的抗CoV-S滴度水平与无症状(暴露)COVID-19患者中的水平相似。第二次疫苗接种达到的GMEU水平超过了门诊治疗的COVID-19患者的恢复期血清六倍，达到了与COVID-19住院治疗患者的恢复期血清相似的水平，并且超过总恢复期血清抗CoV-S抗体近六倍。两剂5μg和25μg BV2373/MATRIX-MTM方案的反应是相似的。这突出了佐剂(MATRIX-MTM)使得能够节省剂量的能力。
在用BV2373治疗的所有组中均诱导了中和抗体(图43B)。用BV2373和MATRIX-MTM方案治疗的组表现出的GMFR为单独用BV2373治疗的组的大约五倍(图43B)。与在没有佐剂的情况下进行单次疫苗接种相比，在有佐剂的情况下进行第二次疫苗接种对中和抗体滴度具有深远的影响，诱导>100倍的升高。当与恢复期血清相比时，用BV2373/MATRIX-MTM进行第二次疫苗接种达到的GMT水平是门诊治疗的COVID-19患者的四倍，其水平跨越COVID-19住院治疗患者的水平，并且超过了总体恢复期血清GMT四倍。
从具有需要医疗护理的临床症状的COVID-19患者获得的恢复期血清展现出随着疾病严重性增加而增加的成比例的抗CoV-S IgG和中和滴度(图43A-图43B)。
在用BV2373和MATRIX-MTM治疗的患者中观察到中和抗体滴度与抗CoV-S IgG之间的强相关性(r＝0.9466，图44C)，类似于在用恢复期血清治疗的患者中观察到的情况(r＝0.958)(图44A)。在施用未加佐剂的BV2373的受试者中未观察到这种相关性(r＝0.7616)(图44B)。5μg和25μg BV2373/MATRIX-MTM(表5的组C-E)均展现出相似的两剂反应幅度，并且当利用两剂方案时，每名参与者都使用任一测定测量进行了血清转化。
16名参与者(组A至D各四名参与者)的T细胞反应表明，在用BV2373刺激后，在IFN-γ、IL-2和TNF-α产生方面，BV2373/MATRIX-MTM方案诱导了抗原特异性多功能性CD4+T细胞反应。存在对Th1细胞因子产生的强烈偏向(图45A-图45D)。
实施例8
下一代CoV S多肽纳米颗粒的表达、纯化和评价
在杆状病毒表达系统中表达具有SEQ ID NO:112、SEQ ID NO:113、SEQ ID NO:114或SEQ ID NO:115的氨基酸序列的CoV S多肽，并且挑选并确认表达冠状病毒纤突(S)多肽的重组噬斑。使用具有SEQ ID NO:5的氨基酸序列的N末端信号肽来表达具有SEQ ID NO:112、SEQ ID NO:113、SEQ ID NO:114和SEQ ID NO:115的序列的CoV S多肽。
具有SEQ ID NO:112的序列的CoV S多肽包含Asn-488至酪氨酸的突变、Lys-973和Val-974至脯氨酸的突变以及具有QQAQ(SEQ ID NO:7)的氨基酸序列的灭活的弗林蛋白酶切割位点。
具有SEQ ID NO:113的序列的CoV S多肽包含Asp-601至甘氨酸的突变、Asn-488至酪氨酸的突变、Lys-973和Val-974至脯氨酸的突变以及具有QQAQ(SEQ ID NO:7)的氨基酸序列的灭活的弗林蛋白酶切割位点。
具有SEQ ID NO:114的序列的CoV S多肽包含氨基酸56、57和131的缺失、Asn-488至酪氨酸的突变、Ala-557至天冬氨酸的突变、Asp-601至甘氨酸的突变、Pro-668至组氨酸的突变、Thr-703至异亮氨酸的突变、Ser-969至丙氨酸的突变、Asp-1105至组氨酸的突变、Lys-973和Val-974至脯氨酸的突变以及具有QQAQ(SEQ ID NO:7)的氨基酸序列的灭活的弗林蛋白酶切割位点。
具有SEQ ID NO:115的序列的CoV S多肽包含Asn-488至酪氨酸的突变、Asp-67至丙氨酸的突变、Leu-229至组氨酸的突变、Asp-202至甘氨酸的突变、Lys-404至天冬酰胺的突变、Glu-471至赖氨酸的突变、Ala-688至缬氨酸的突变、Asp-601至甘氨酸的突变、Lys-973和Val-974至脯氨酸的突变以及具有QQAQ的氨基酸序列的灭活的弗林蛋白酶切割位点。
如实施例1中那样产生CoV S多肽纳米颗粒。如实施例2-7中那样评价具有SEQ IDNO:112、SEQ ID NO:112、SEQ ID NO:113和SEQ ID NO:115的氨基酸序列的CoV S多肽的稳定性和免疫原性。
编号实施方案
1.一种免疫原性组合物，其包含：
(i)包含具有SEQ ID NO:87的氨基酸序列的冠状病毒S(CoV S)糖蛋白和非离子型洗涤剂核心的纳米颗粒；
(ii)药学上可接受的缓冲液；以及
(iii)皂苷佐剂。
2.根据实施方案1所述的免疫原性组合物，其包含在约5μg与约25μg之间的CoV S糖蛋白。
3.根据实施方案2所述的免疫原性组合物，其包含约5μg的CoV S糖蛋白。
4.根据实施方案1所述的免疫原性组合物，其中所述皂苷佐剂包含至少两种iscom颗粒，其中：
第一iscom颗粒包含皂树(Quillaja Saponaria Molina)的级分A，而不包含皂树的级分C；并且
第二iscom颗粒包含皂树的级分C，而不包含皂树的级分A。
5.根据实施方案4所述的免疫原性组合物，其中分别地，皂树的级分A占在所述佐剂中皂树的级分A和皂树的级分C的重量之和的按重量计50％-96％，并且皂树的级分C占剩余部分。
6.根据实施方案4所述的免疫原性组合物，其中皂树的级分A和皂树的级分C分别占在所述佐剂中皂树的级分A和皂树的级分C的重量之和的按重量计约85％和按重量计约15％。
7.根据实施方案1所述的免疫原性组合物，其包含约50μg的皂苷佐剂。
8.根据实施方案1所述的免疫原性组合物，其中所述非离子型洗涤剂核心选自聚山梨醇酯20(PS20)、聚山梨醇酯40(PS40)、聚山梨醇酯60(PS60)、聚山梨醇酯65(PS65)和聚山梨醇酯80(PS80)。
9.一种在受试者中刺激针对SARS-CoV-2的免疫反应的方法，其包括施用根据实施方案1所述的免疫原性组合物。
10.根据实施方案9所述的方法，其包含在约5μg与约25μg之间的CoV S糖蛋白。
11.根据实施方案10所述的方法，其包含5μg的CoV S糖蛋白。
12.根据实施方案9所述的方法，其中所述皂苷佐剂包含至少两种iscom颗粒，其中：
第一iscom颗粒包含皂树的级分A，而不包含皂树的级分C；并且
第二iscom颗粒包含皂树的级分C，而不包含皂树的级分A。
13.根据实施方案12所述的方法，其中分别地，皂树的级分A占在所述佐剂中皂树的级分A和皂树的级分C的重量之和的按重量计50％-96％，并且皂树的级分C占剩余部分。
14.根据实施方案12所述的方法，其中皂树的级分A和皂树的级分C分别占在所述佐剂中皂树的级分A和皂树的级分C的重量之和的按重量计约85％和按重量计约15％。
15.根据实施方案9所述的方法，其包含约50μg的皂苷佐剂。
16.根据实施方案9所述的方法，其中所述非离子型洗涤剂核心选自聚山梨醇酯20(PS20)、聚山梨醇酯40(PS40)、聚山梨醇酯60(PS60)、聚山梨醇酯65(PS65)和聚山梨醇酯80(PS80)。
17.根据实施方案9所述的方法，其中向所述受试者在第0天施用第一剂量，并且在第21天施用加强剂量。
18.根据实施方案9所述的方法，其中施用单剂的所述免疫原性组合物。
19.根据实施方案9所述的方法，其包括施用不同于所述第一免疫原性组合物的第二免疫原性组合物。
20.根据实施方案19所述的方法，其中所述第二免疫原性组合物包含编码SARS-CoV-2纤突糖蛋白的mRNA、编码SARS-CoV-2纤突糖蛋白的质粒DNA、编码SARS-CoV-2纤突糖蛋白的病毒载体或灭活的SARS-CoV-2病毒。
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序列表
<110> 诺瓦瓦克斯股份有限公司
<120> 冠状病毒疫苗配制品
<130> NOVV-088/08CN 305047-3008
<150> US 63/129,392
<151> 2020-12-22
<150> US 16/997,001
<151> 2020-08-19
<150> US 63/054,182
<151> 2020-07-20
<150> US 63/051,706
<151> 2020-07-14
<150> US 63/048,945
<151> 2020-07-07
<150> US 62/983,180
<151> 2020-02-28
<150> US 62/976,858
<151> 2020-02-14
<150> US 62/966,271
<151> 2020-01-27
<160> 126
<170> PatentIn version 3.5
<210> 1
<211> 1273
<212> PRT
<213> β冠状病毒严重急性呼吸系统综合征冠状病毒2
<400> 1
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Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe
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Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu
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His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp
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Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp
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Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu
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Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser
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Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile
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Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr
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Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
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Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
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Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
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Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
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Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
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Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
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Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
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Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
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Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
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Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
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Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
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Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
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Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
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Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
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Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
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Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
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Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
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Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
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Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
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Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
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Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
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Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
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Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
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Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
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Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
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Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
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Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val
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Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
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Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
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Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile
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His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
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Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
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Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
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Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala Arg Ser Val Ala
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Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser
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Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile
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Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val
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Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu
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Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr
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Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln
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Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe
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Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser
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Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly
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Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp
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Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu
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Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly
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Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln Ile
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Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr
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Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn
915 920 925
Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala
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Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn
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Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val
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Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala Glu Val Gln
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Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val
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Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn
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Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys
1025 1030 1035
Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro
1040 1045 1050
Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val
1055 1060 1065
Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His
1070 1075 1080
Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn
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Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln
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Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val
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Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro
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Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn
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His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn
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Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu
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Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu
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Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile Trp Leu
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Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr Ile Met
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Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly Cys Cys
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Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser Glu Pro
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Val Leu Lys Gly Val Lys Leu His Tyr Thr
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<213> β冠状病毒严重急性呼吸系统综合征冠状病毒2
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Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
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Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
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Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
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Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
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Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn
290 295 300
Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr
305 310 315 320
Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser
325 330 335
Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr
340 345 350
Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly
355 360 365
Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala
370 375 380
Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly
385 390 395 400
Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe
405 410 415
Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val
420 425 430
Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu
435 440 445
Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser
450 455 460
Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln
465 470 475 480
Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg
485 490 495
Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys
500 505 510
Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe
515 520 525
Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys
530 535 540
Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr
545 550 555 560
Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro
565 570 575
Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser
580 585 590
Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro
595 600 605
Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser
610 615 620
Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala
625 630 635 640
Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly
645 650 655
Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala Arg
660 665 670
Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala
675 680 685
Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn
690 695 700
Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys
705 710 715 720
Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys
725 730 735
Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg
740 745 750
Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val
755 760 765
Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe
770 775 780
Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser
785 790 795 800
Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala
805 810 815
Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala
820 825 830
Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu
835 840 845
Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu
850 855 860
Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala
865 870 875 880
Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile
885 890 895
Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn
900 905 910
Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr
915 920 925
Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln
930 935 940
Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile
945 950 955 960
Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala
965 970 975
Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln
980 985 990
Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser
995 1000 1005
Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln
1010 1015 1020
Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser
1025 1030 1035
Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr
1040 1045 1050
Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile
1055 1060 1065
Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val
1070 1075 1080
Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu
1085 1090 1095
Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys
1100 1105 1110
Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu
1115 1120 1125
Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe
1130 1135 1140
Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly
1145 1150 1155
Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu
1160 1165 1170
Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln
1175 1180 1185
Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile
1190 1195 1200
Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr
1205 1210 1215
Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly
1220 1225 1230
Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser
1235 1240 1245
Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr
1250 1255 1260
<210> 3
<211> 1273
<212> PRT
<213> 人工序列
<220>
<223> 重组S蛋白
<400> 3
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val
1 5 10 15
Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe
20 25 30
Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu
35 40 45
His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp
50 55 60
Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp
65 70 75 80
Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu
85 90 95
Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser
100 105 110
Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile
115 120 125
Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr
130 135 140
Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
145 150 155 160
Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
165 170 175
Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
180 185 190
Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
195 200 205
Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
210 215 220
Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
225 230 235 240
Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
245 250 255
Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
260 265 270
Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
275 280 285
Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
290 295 300
Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
305 310 315 320
Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
325 330 335
Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
340 345 350
Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
355 360 365
Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
370 375 380
Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
385 390 395 400
Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
405 410 415
Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
420 425 430
Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
435 440 445
Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
450 455 460
Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
465 470 475 480
Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
485 490 495
Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
500 505 510
Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
515 520 525
Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
530 535 540
Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
545 550 555 560
Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val
565 570 575
Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
580 585 590
Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
595 600 605
Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile
610 615 620
His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
625 630 635 640
Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
645 650 655
Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
660 665 670
Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln Ser Val Ala
675 680 685
Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser
690 695 700
Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile
705 710 715 720
Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val
725 730 735
Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu
740 745 750
Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr
755 760 765
Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln
770 775 780
Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe
785 790 795 800
Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser
805 810 815
Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly
820 825 830
Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp
835 840 845
Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu
850 855 860
Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly
865 870 875 880
Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln Ile
885 890 895
Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr
900 905 910
Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn
915 920 925
Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala
930 935 940
Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn
945 950 955 960
Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val
965 970 975
Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala Glu Val Gln
980 985 990
Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val
995 1000 1005
Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn
1010 1015 1020
Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys
1025 1030 1035
Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro
1040 1045 1050
Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val
1055 1060 1065
Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His
1070 1075 1080
Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn
1085 1090 1095
Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln
1100 1105 1110
Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val
1115 1120 1125
Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro
1130 1135 1140
Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn
1145 1150 1155
His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn
1160 1165 1170
Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu
1175 1180 1185
Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu
1190 1195 1200
Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile Trp Leu
1205 1210 1215
Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr Ile Met
1220 1225 1230
Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly Cys Cys
1235 1240 1245
Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser Glu Pro
1250 1255 1260
Val Leu Lys Gly Val Lys Leu His Tyr Thr
1265 1270
<210> 4
<211> 1260
<212> PRT
<213> 人工序列
<220>
<223> 重组S蛋白
<400> 4
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn
290 295 300
Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr
305 310 315 320
Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser
325 330 335
Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr
340 345 350
Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly
355 360 365
Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala
370 375 380
Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly
385 390 395 400
Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe
405 410 415
Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val
420 425 430
Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu
435 440 445
Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser
450 455 460
Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln
465 470 475 480
Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg
485 490 495
Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys
500 505 510
Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe
515 520 525
Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys
530 535 540
Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr
545 550 555 560
Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro
565 570 575
Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser
580 585 590
Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro
595 600 605
Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser
610 615 620
Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala
625 630 635 640
Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly
645 650 655
Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln
660 665 670
Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala
675 680 685
Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn
690 695 700
Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys
705 710 715 720
Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys
725 730 735
Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg
740 745 750
Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val
755 760 765
Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe
770 775 780
Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser
785 790 795 800
Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala
805 810 815
Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala
820 825 830
Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu
835 840 845
Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu
850 855 860
Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala
865 870 875 880
Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile
885 890 895
Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn
900 905 910
Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr
915 920 925
Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln
930 935 940
Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile
945 950 955 960
Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala
965 970 975
Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln
980 985 990
Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser
995 1000 1005
Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln
1010 1015 1020
Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser
1025 1030 1035
Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr
1040 1045 1050
Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile
1055 1060 1065
Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val
1070 1075 1080
Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu
1085 1090 1095
Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys
1100 1105 1110
Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu
1115 1120 1125
Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe
1130 1135 1140
Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly
1145 1150 1155
Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu
1160 1165 1170
Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln
1175 1180 1185
Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile
1190 1195 1200
Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr
1205 1210 1215
Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly
1220 1225 1230
Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser
1235 1240 1245
Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr
1250 1255 1260
<210> 5
<211> 13
<212> PRT
<213> 人工序列
<220>
<223> 信号肽
<400> 5
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser
1 5 10
<210> 6
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 6
Arg Arg Ala Arg
1
<210> 7
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 7
Gln Gln Ala Gln
1
<210> 8
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 8
Gln Arg Ala Arg
1
<210> 9
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 9
Arg Gln Ala Arg
1
<210> 10
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 10
Arg Arg Ala Gln
1
<210> 11
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 11
Gln Gln Ala Arg
1
<210> 12
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 12
Arg Gln Ala Gln
1
<210> 13
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 13
Gln Arg Ala Gln
1
<210> 14
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 14
Asn Asn Ala Asn
1
<210> 15
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 15
Asn Arg Ala Arg
1
<210> 16
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 16
Arg Asn Ala Arg
1
<210> 17
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 17
Arg Arg Ala Asn
1
<210> 18
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 18
Asn Asn Ala Arg
1
<210> 19
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 19
Arg Asn Ala Asn
1
<210> 20
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 20
Asn Arg Ala Asn
1
<210> 21
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 21
Ala Ala Ala Ala
1
<210> 22
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 22
Ala Arg Ala Arg
1
<210> 23
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 23
Arg Ala Ala Arg
1
<210> 24
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 24
Arg Arg Ala Ala
1
<210> 25
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 25
Ala Ala Ala Arg
1
<210> 26
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 26
Arg Ala Ala Ala
1
<210> 27
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 27
Ala Arg Ala Ala
1
<210> 28
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 28
Gly Gly Ala Gly
1
<210> 29
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 29
Gly Arg Ala Arg
1
<210> 30
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 30
Arg Gly Ala Arg
1
<210> 31
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 31
Arg Arg Ala Gly
1
<210> 32
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 32
Gly Gly Ala Arg
1
<210> 33
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 33
Arg Gly Ala Gly
1
<210> 34
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 34
Gly Arg Ala Gly
1
<210> 35
<211> 3819
<212> DNA
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 35
atgttcgtct tcctggtgct gctgcccctg gtgtccagcc agtgcgtgaa cctgaccact 60
aggactcagc tgcctcccgc ttacaccaac tcattcactc gcggtgtgta ctaccctgac 120
aaggtcttcc gttcttcagt gctgcactca actcaggacc tgttcctgcc cttcttctcc 180
aacgtcacct ggttccacgc catccacgtg tccggcacca acggcactaa gcgcttcgac 240
aacccagtgc tgcctttcaa cgacggtgtc tacttcgctt caaccgagaa gtccaacatc 300
atccgtggat ggatcttcgg caccactctg gacagcaaga ctcagtctct gctgatcgtc 360
aacaacgcca ccaacgtggt catcaaggtc tgcgaattcc agttctgcaa cgacccattc 420
ctgggcgtct actaccacaa gaacaacaag tcatggatgg agtccgaatt ccgcgtctac 480
tccagcgcta acaactgcac tttcgagtac gtgtcccagc ctttcctgat ggacctggaa 540
ggaaagcagg gtaacttcaa gaacctgagg gagttcgtgt tcaagaacat cgacggatac 600
ttcaagattt acagcaagca caccccaatc aacctggtgc gcgacctgcc tcagggtttc 660
tctgctctgg agccactggt ggacctgcct atcggcatca acatcacccg cttccagact 720
ctgctggctc tgcaccgttc ctacctgact ccaggcgact catcttctgg atggactgct 780
ggagctgctg cttactacgt gggctacctg cagcctcgca ccttcctgct gaagtacaac 840
gaaaacggaa ccatcactga cgccgtcgac tgcgctctgg accctctgtc agaaaccaag 900
tgcactctga agtccttcac cgtggagaag ggcatctacc agacttcaaa cttcagggtg 960
cagcccaccg aatccatcgt cagattccct aacatcacta acctgtgccc cttcggagag 1020
gtcttcaacg ccacccgctt cgcttccgtg tacgcctgga acaggaagag aatctcaaac 1080
tgcgtcgctg actactccgt gctgtacaac tcagcctcct tcagcacctt caagtgctac 1140
ggcgtgtcac caactaagct gaacgacctg tgcttcacca acgtctacgc cgactccttc 1200
gtgatcaggg gagacgaggt cagacagatc gctcctggcc agactggaaa gatcgccgac 1260
tacaactaca agctgcccga cgacttcacc ggttgcgtca tcgcttggaa cagcaacaac 1320
ctggactcta aagtgggtgg caactacaac tacctgtacc gcctgttccg taagtcaaac 1380
ctgaagccat tcgagaggga catcagcact gaaatctacc aggccggatc taccccttgc 1440
aacggtgtcg agggcttcaa ctgctacttc cccctgcagt cctacggttt ccagccaact 1500
aacggtgtgg gctaccagcc ttacagagtg gtcgtgctga gcttcgaact gctccacgct 1560
cctgctactg tgtgcggtcc aaagaagtct accaacctgg tcaagaacaa gtgcgtgaac 1620
ttcaacttca acggcctgac cggaactggt gtcctgaccg agagcaacaa gaagttcctg 1680
cccttccagc agttcggaag ggacatcgct gacaccactg acgctgtgcg cgaccctcag 1740
accctggaaa tcctggacat cactccatgc tcattcggag gtgtctccgt gatcacccct 1800
ggcaccaaca cttctaacca ggtcgctgtg ctgtaccagg acgtcaactg caccgaggtc 1860
cctgtggcca tccacgctga ccagctgacc cccacttggc gcgtgtactc caccggcagc 1920
aacgtgttcc agactcgtgc tggttgcctg atcggcgccg agcacgtgaa caacagctac 1980
gaatgcgaca tccccatcgg cgctggaatc tgcgcctctt accagaccca gactaacagc 2040
ccacgcaggg ctcgctctgt ggcctctcag tcaatcatcg cttacaccat gtcactgggc 2100
gctgaaaact ccgtggccta ctctaacaac tcaatcgcca tccccaccaa cttcactatc 2160
agcgtgacca ctgagatcct gccagtcagc atgaccaaga cttctgtgga ctgcactatg 2220
tacatctgcg gagacagcac cgaatgctct aacctgctgc tgcagtacgg ctctttctgc 2280
acccagctga accgtgctct gactggaatc gccgtggagc aggacaagaa cactcaggaa 2340
gtcttcgctc aggtgaagca aatctacaag accccaccta tcaaggactt cggcggattc 2400
aacttctccc agatcctgcc tgacccctcc aagccaagca agcgctcttt catcgaggac 2460
ctgctgttca acaaggtcac tctggccgac gctggattca tcaagcagta cggagactgc 2520
ctgggtgaca tcgccgctcg tgacctgatc tgcgctcaga agttcaacgg tctgaccgtg 2580
ctgcccccac tgctgactga cgaaatgatc gcccagtaca ctagcgccct gctggctgga 2640
accatcactt ctggttggac cttcggtgct ggcgccgctc tgcagatccc tttcgctatg 2700
cagatggcct accgtttcaa cggaatcggt gtcacccaga acgtgctgta cgagaaccag 2760
aagctgatcg ctaaccagtt caactcagcc atcggaaaga tccaggacag cctgagctct 2820
actgcctctg ctctgggcaa gctgcaggac gtcgtgaacc agaacgccca ggctctgaac 2880
accctggtca agcagctgtc atccaacttc ggtgctatca gctctgtgct gaacgacatc 2940
ctgtcccgcc tggacaaggt cgaggccgaa gtgcagatcg accgcctgat cactggccgt 3000
ctgcagtcac tgcagaccta cgtgactcag cagctgatca gggccgctga aatcagagcc 3060
tccgctaacc tggccgctac caagatgagc gagtgcgtcc tgggtcaatc taagcgtgtg 3120
gacttctgcg gcaagggata ccacctgatg tcattccctc agtctgctcc ccacggtgtg 3180
gtgttcctgc acgtcaccta cgtgccagcc caggagaaga acttcaccac tgcccctgct 3240
atctgccacg acggcaaggc tcacttcccc agggaaggtg tcttcgtgag caacggcacc 3300
cactggttcg tcactcagag aaacttctac gagccacaga tcatcaccac tgacaacact 3360
ttcgtgtctg gaaactgcga cgtggtcatc ggtatcgtca acaacaccgt gtacgacccc 3420
ctgcagccag agctggactc attcaaggag gaactggaca agtacttcaa gaaccacacc 3480
tcccctgacg tcgacctggg cgacatctca ggaatcaacg cttccgtcgt gaacatccag 3540
aaggagatcg accgcctgaa cgaagtggcc aagaacctga acgaaagcct gatcgacctg 3600
caggagctgg gcaagtacga acagtacatc aagtggcctt ggtacatctg gctgggtttc 3660
atcgctggcc tcatcgctat cgtgatggtg accatcatgc tgtgctgcat gacttcatgc 3720
tgctcctgcc tgaagggctg ctgcagctgc ggatcttgct gcaagttcga cgaggacgac 3780
tctgaacccg tcctgaaggg cgtgaagctg cactacacc 3819
<210> 36
<211> 1279
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 36
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val
1 5 10 15
Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe
20 25 30
Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu
35 40 45
His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp
50 55 60
Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp
65 70 75 80
Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu
85 90 95
Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser
100 105 110
Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile
115 120 125
Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr
130 135 140
Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
145 150 155 160
Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
165 170 175
Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
180 185 190
Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
195 200 205
Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
210 215 220
Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
225 230 235 240
Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
245 250 255
Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
260 265 270
Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
275 280 285
Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
290 295 300
Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
305 310 315 320
Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
325 330 335
Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
340 345 350
Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
355 360 365
Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
370 375 380
Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
385 390 395 400
Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
405 410 415
Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
420 425 430
Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
435 440 445
Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
450 455 460
Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
465 470 475 480
Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
485 490 495
Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
500 505 510
Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
515 520 525
Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
530 535 540
Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
545 550 555 560
Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val
565 570 575
Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
580 585 590
Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
595 600 605
Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile
610 615 620
His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
625 630 635 640
Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
645 650 655
Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
660 665 670
Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala Arg Ser Val Ala
675 680 685
Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser
690 695 700
Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile
705 710 715 720
Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val
725 730 735
Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu
740 745 750
Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr
755 760 765
Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln
770 775 780
Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe
785 790 795 800
Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser
805 810 815
Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly
820 825 830
Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp
835 840 845
Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu
850 855 860
Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly
865 870 875 880
Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln Ile
885 890 895
Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr
900 905 910
Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn
915 920 925
Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala
930 935 940
Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn
945 950 955 960
Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val
965 970 975
Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala Glu Val Gln
980 985 990
Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val
995 1000 1005
Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn
1010 1015 1020
Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys
1025 1030 1035
Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro
1040 1045 1050
Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val
1055 1060 1065
Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His
1070 1075 1080
Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn
1085 1090 1095
Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln
1100 1105 1110
Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val
1115 1120 1125
Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro
1130 1135 1140
Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn
1145 1150 1155
His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn
1160 1165 1170
Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu
1175 1180 1185
Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu
1190 1195 1200
Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile Trp Leu
1205 1210 1215
Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr Ile Met
1220 1225 1230
Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly Cys Cys
1235 1240 1245
Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser Glu Pro
1250 1255 1260
Val Leu Lys Gly Val Lys Leu His Tyr Thr His His His His His
1265 1270 1275
His
<210> 37
<211> 3837
<212> DNA
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 37
atgttcgtct tcctggtgct gctgcccctg gtgtccagcc agtgcgtgaa cctgaccact 60
aggactcagc tgcctcccgc ttacaccaac tcattcactc gcggtgtgta ctaccctgac 120
aaggtcttcc gttcttcagt gctgcactca actcaggacc tgttcctgcc cttcttctcc 180
aacgtcacct ggttccacgc catccacgtg tccggcacca acggcactaa gcgcttcgac 240
aacccagtgc tgcctttcaa cgacggtgtc tacttcgctt caaccgagaa gtccaacatc 300
atccgtggat ggatcttcgg caccactctg gacagcaaga ctcagtctct gctgatcgtc 360
aacaacgcca ccaacgtggt catcaaggtc tgcgaattcc agttctgcaa cgacccattc 420
ctgggcgtct actaccacaa gaacaacaag tcatggatgg agtccgaatt ccgcgtctac 480
tccagcgcta acaactgcac tttcgagtac gtgtcccagc ctttcctgat ggacctggaa 540
ggaaagcagg gtaacttcaa gaacctgagg gagttcgtgt tcaagaacat cgacggatac 600
ttcaagattt acagcaagca caccccaatc aacctggtgc gcgacctgcc tcagggtttc 660
tctgctctgg agccactggt ggacctgcct atcggcatca acatcacccg cttccagact 720
ctgctggctc tgcaccgttc ctacctgact ccaggcgact catcttctgg atggactgct 780
ggagctgctg cttactacgt gggctacctg cagcctcgca ccttcctgct gaagtacaac 840
gaaaacggaa ccatcactga cgccgtcgac tgcgctctgg accctctgtc agaaaccaag 900
tgcactctga agtccttcac cgtggagaag ggcatctacc agacttcaaa cttcagggtg 960
cagcccaccg aatccatcgt cagattccct aacatcacta acctgtgccc cttcggagag 1020
gtcttcaacg ccacccgctt cgcttccgtg tacgcctgga acaggaagag aatctcaaac 1080
tgcgtcgctg actactccgt gctgtacaac tcagcctcct tcagcacctt caagtgctac 1140
ggcgtgtcac caactaagct gaacgacctg tgcttcacca acgtctacgc cgactccttc 1200
gtgatcaggg gagacgaggt cagacagatc gctcctggcc agactggaaa gatcgccgac 1260
tacaactaca agctgcccga cgacttcacc ggttgcgtca tcgcttggaa cagcaacaac 1320
ctggactcta aagtgggtgg caactacaac tacctgtacc gcctgttccg taagtcaaac 1380
ctgaagccat tcgagaggga catcagcact gaaatctacc aggccggatc taccccttgc 1440
aacggtgtcg agggcttcaa ctgctacttc cccctgcagt cctacggttt ccagccaact 1500
aacggtgtgg gctaccagcc ttacagagtg gtcgtgctga gcttcgaact gctccacgct 1560
cctgctactg tgtgcggtcc aaagaagtct accaacctgg tcaagaacaa gtgcgtgaac 1620
ttcaacttca acggcctgac cggaactggt gtcctgaccg agagcaacaa gaagttcctg 1680
cccttccagc agttcggaag ggacatcgct gacaccactg acgctgtgcg cgaccctcag 1740
accctggaaa tcctggacat cactccatgc tcattcggag gtgtctccgt gatcacccct 1800
ggcaccaaca cttctaacca ggtcgctgtg ctgtaccagg acgtcaactg caccgaggtc 1860
cctgtggcca tccacgctga ccagctgacc cccacttggc gcgtgtactc caccggcagc 1920
aacgtgttcc agactcgtgc tggttgcctg atcggcgccg agcacgtgaa caacagctac 1980
gaatgcgaca tccccatcgg cgctggaatc tgcgcctctt accagaccca gactaacagc 2040
ccacgcaggg ctcgctctgt ggcctctcag tcaatcatcg cttacaccat gtcactgggc 2100
gctgaaaact ccgtggccta ctctaacaac tcaatcgcca tccccaccaa cttcactatc 2160
agcgtgacca ctgagatcct gccagtcagc atgaccaaga cttctgtgga ctgcactatg 2220
tacatctgcg gagacagcac cgaatgctct aacctgctgc tgcagtacgg ctctttctgc 2280
acccagctga accgtgctct gactggaatc gccgtggagc aggacaagaa cactcaggaa 2340
gtcttcgctc aggtgaagca aatctacaag accccaccta tcaaggactt cggcggattc 2400
aacttctccc agatcctgcc tgacccctcc aagccaagca agcgctcttt catcgaggac 2460
ctgctgttca acaaggtcac tctggccgac gctggattca tcaagcagta cggagactgc 2520
ctgggtgaca tcgccgctcg tgacctgatc tgcgctcaga agttcaacgg tctgaccgtg 2580
ctgcccccac tgctgactga cgaaatgatc gcccagtaca ctagcgccct gctggctgga 2640
accatcactt ctggttggac cttcggtgct ggcgccgctc tgcagatccc tttcgctatg 2700
cagatggcct accgtttcaa cggaatcggt gtcacccaga acgtgctgta cgagaaccag 2760
aagctgatcg ctaaccagtt caactcagcc atcggaaaga tccaggacag cctgagctct 2820
actgcctctg ctctgggcaa gctgcaggac gtcgtgaacc agaacgccca ggctctgaac 2880
accctggtca agcagctgtc atccaacttc ggtgctatca gctctgtgct gaacgacatc 2940
ctgtcccgcc tggacaaggt cgaggccgaa gtgcagatcg accgcctgat cactggccgt 3000
ctgcagtcac tgcagaccta cgtgactcag cagctgatca gggccgctga aatcagagcc 3060
tccgctaacc tggccgctac caagatgagc gagtgcgtcc tgggtcaatc taagcgtgtg 3120
gacttctgcg gcaagggata ccacctgatg tcattccctc agtctgctcc ccacggtgtg 3180
gtgttcctgc acgtcaccta cgtgccagcc caggagaaga acttcaccac tgcccctgct 3240
atctgccacg acggcaaggc tcacttcccc agggaaggtg tcttcgtgag caacggcacc 3300
cactggttcg tcactcagag aaacttctac gagccacaga tcatcaccac tgacaacact 3360
ttcgtgtctg gaaactgcga cgtggtcatc ggtatcgtca acaacaccgt gtacgacccc 3420
ctgcagccag agctggactc attcaaggag gaactggaca agtacttcaa gaaccacacc 3480
tcccctgacg tcgacctggg cgacatctca ggaatcaacg cttccgtcgt gaacatccag 3540
aaggagatcg accgcctgaa cgaagtggcc aagaacctga acgaaagcct gatcgacctg 3600
caggagctgg gcaagtacga acagtacatc aagtggcctt ggtacatctg gctgggtttc 3660
atcgctggcc tcatcgctat cgtgatggtg accatcatgc tgtgctgcat gacttcatgc 3720
tgctcctgcc tgaagggctg ctgcagctgc ggatcttgct gcaagttcga cgaggacgac 3780
tctgaacccg tcctgaaggg cgtgaagctg cactacaccc accaccacca ccaccac 3837
<210> 38
<211> 1266
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 38
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn
290 295 300
Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr
305 310 315 320
Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser
325 330 335
Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr
340 345 350
Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly
355 360 365
Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala
370 375 380
Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly
385 390 395 400
Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe
405 410 415
Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val
420 425 430
Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu
435 440 445
Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser
450 455 460
Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln
465 470 475 480
Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg
485 490 495
Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys
500 505 510
Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe
515 520 525
Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys
530 535 540
Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr
545 550 555 560
Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro
565 570 575
Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser
580 585 590
Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro
595 600 605
Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser
610 615 620
Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala
625 630 635 640
Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly
645 650 655
Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala Arg
660 665 670
Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala
675 680 685
Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn
690 695 700
Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys
705 710 715 720
Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys
725 730 735
Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg
740 745 750
Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val
755 760 765
Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe
770 775 780
Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser
785 790 795 800
Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala
805 810 815
Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala
820 825 830
Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu
835 840 845
Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu
850 855 860
Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala
865 870 875 880
Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile
885 890 895
Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn
900 905 910
Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr
915 920 925
Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln
930 935 940
Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile
945 950 955 960
Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala
965 970 975
Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln
980 985 990
Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser
995 1000 1005
Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln
1010 1015 1020
Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser
1025 1030 1035
Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr
1040 1045 1050
Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile
1055 1060 1065
Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val
1070 1075 1080
Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu
1085 1090 1095
Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys
1100 1105 1110
Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu
1115 1120 1125
Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe
1130 1135 1140
Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly
1145 1150 1155
Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu
1160 1165 1170
Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln
1175 1180 1185
Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile
1190 1195 1200
Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr
1205 1210 1215
Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly
1220 1225 1230
Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser
1235 1240 1245
Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr His His His
1250 1255 1260
His His His
1265
<210> 39
<211> 60
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 39
Trp Tyr Ile Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met
1 5 10 15
Val Thr Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys
20 25 30
Gly Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser
35 40 45
Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr
50 55 60
<210> 40
<211> 1213
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 40
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val
1 5 10 15
Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe
20 25 30
Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu
35 40 45
His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp
50 55 60
Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp
65 70 75 80
Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu
85 90 95
Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser
100 105 110
Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile
115 120 125
Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr
130 135 140
Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
145 150 155 160
Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
165 170 175
Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
180 185 190
Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
195 200 205
Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
210 215 220
Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
225 230 235 240
Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
245 250 255
Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
260 265 270
Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
275 280 285
Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
290 295 300
Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
305 310 315 320
Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
325 330 335
Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
340 345 350
Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
355 360 365
Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
370 375 380
Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
385 390 395 400
Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
405 410 415
Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
420 425 430
Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
435 440 445
Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
450 455 460
Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
465 470 475 480
Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
485 490 495
Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
500 505 510
Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
515 520 525
Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
530 535 540
Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
545 550 555 560
Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val
565 570 575
Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
580 585 590
Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
595 600 605
Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile
610 615 620
His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
625 630 635 640
Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
645 650 655
Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
660 665 670
Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala Arg Ser Val Ala
675 680 685
Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser
690 695 700
Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile
705 710 715 720
Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val
725 730 735
Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu
740 745 750
Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr
755 760 765
Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln
770 775 780
Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe
785 790 795 800
Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser
805 810 815
Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly
820 825 830
Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp
835 840 845
Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu
850 855 860
Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly
865 870 875 880
Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln Ile
885 890 895
Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr
900 905 910
Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn
915 920 925
Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala
930 935 940
Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn
945 950 955 960
Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val
965 970 975
Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala Glu Val Gln
980 985 990
Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val
995 1000 1005
Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn
1010 1015 1020
Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys
1025 1030 1035
Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro
1040 1045 1050
Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val
1055 1060 1065
Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His
1070 1075 1080
Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn
1085 1090 1095
Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln
1100 1105 1110
Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val
1115 1120 1125
Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro
1130 1135 1140
Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn
1145 1150 1155
His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn
1160 1165 1170
Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu
1175 1180 1185
Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu
1190 1195 1200
Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro
1205 1210
<210> 41
<211> 1200
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 41
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn
290 295 300
Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr
305 310 315 320
Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser
325 330 335
Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr
340 345 350
Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly
355 360 365
Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala
370 375 380
Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly
385 390 395 400
Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe
405 410 415
Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val
420 425 430
Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu
435 440 445
Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser
450 455 460
Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln
465 470 475 480
Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg
485 490 495
Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys
500 505 510
Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe
515 520 525
Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys
530 535 540
Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr
545 550 555 560
Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro
565 570 575
Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser
580 585 590
Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro
595 600 605
Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser
610 615 620
Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala
625 630 635 640
Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly
645 650 655
Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala Arg
660 665 670
Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala
675 680 685
Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn
690 695 700
Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys
705 710 715 720
Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys
725 730 735
Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg
740 745 750
Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val
755 760 765
Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe
770 775 780
Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser
785 790 795 800
Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala
805 810 815
Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala
820 825 830
Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu
835 840 845
Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu
850 855 860
Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala
865 870 875 880
Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile
885 890 895
Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn
900 905 910
Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr
915 920 925
Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln
930 935 940
Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile
945 950 955 960
Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala
965 970 975
Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln
980 985 990
Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser
995 1000 1005
Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln
1010 1015 1020
Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser
1025 1030 1035
Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr
1040 1045 1050
Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile
1055 1060 1065
Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val
1070 1075 1080
Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu
1085 1090 1095
Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys
1100 1105 1110
Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu
1115 1120 1125
Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe
1130 1135 1140
Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly
1145 1150 1155
Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu
1160 1165 1170
Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln
1175 1180 1185
Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro
1190 1195 1200
<210> 42
<211> 1219
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 42
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val
1 5 10 15
Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe
20 25 30
Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu
35 40 45
His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp
50 55 60
Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp
65 70 75 80
Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu
85 90 95
Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser
100 105 110
Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile
115 120 125
Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr
130 135 140
Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
145 150 155 160
Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
165 170 175
Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
180 185 190
Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
195 200 205
Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
210 215 220
Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
225 230 235 240
Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
245 250 255
Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
260 265 270
Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
275 280 285
Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
290 295 300
Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
305 310 315 320
Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
325 330 335
Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
340 345 350
Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
355 360 365
Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
370 375 380
Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
385 390 395 400
Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
405 410 415
Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
420 425 430
Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
435 440 445
Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
450 455 460
Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
465 470 475 480
Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
485 490 495
Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
500 505 510
Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
515 520 525
Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
530 535 540
Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
545 550 555 560
Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val
565 570 575
Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
580 585 590
Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
595 600 605
Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile
610 615 620
His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
625 630 635 640
Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
645 650 655
Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
660 665 670
Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala Arg Ser Val Ala
675 680 685
Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser
690 695 700
Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile
705 710 715 720
Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val
725 730 735
Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu
740 745 750
Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr
755 760 765
Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln
770 775 780
Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe
785 790 795 800
Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser
805 810 815
Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly
820 825 830
Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp
835 840 845
Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu
850 855 860
Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly
865 870 875 880
Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln Ile
885 890 895
Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr
900 905 910
Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn
915 920 925
Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala
930 935 940
Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn
945 950 955 960
Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val
965 970 975
Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala Glu Val Gln
980 985 990
Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val
995 1000 1005
Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn
1010 1015 1020
Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys
1025 1030 1035
Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro
1040 1045 1050
Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val
1055 1060 1065
Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His
1070 1075 1080
Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn
1085 1090 1095
Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln
1100 1105 1110
Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val
1115 1120 1125
Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro
1130 1135 1140
Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn
1145 1150 1155
His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn
1160 1165 1170
Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu
1175 1180 1185
Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu
1190 1195 1200
Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro His His His His His
1205 1210 1215
His
<210> 43
<211> 3657
<212> DNA
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 43
atgttcgtct tcctggtgct gctgcccctg gtgtccagcc agtgcgtgaa cctgaccact 60
aggactcagc tgcctcccgc ttacaccaac tcattcactc gcggtgtgta ctaccctgac 120
aaggtcttcc gttcttcagt gctgcactca actcaggacc tgttcctgcc cttcttctcc 180
aacgtcacct ggttccacgc catccacgtg tccggcacca acggcactaa gcgcttcgac 240
aacccagtgc tgcctttcaa cgacggtgtc tacttcgctt caaccgagaa gtccaacatc 300
atccgtggat ggatcttcgg caccactctg gacagcaaga ctcagtctct gctgatcgtc 360
aacaacgcca ccaacgtggt catcaaggtc tgcgaattcc agttctgcaa cgacccattc 420
ctgggcgtct actaccacaa gaacaacaag tcatggatgg agtccgaatt ccgcgtctac 480
tccagcgcta acaactgcac tttcgagtac gtgtcccagc ctttcctgat ggacctggaa 540
ggaaagcagg gtaacttcaa gaacctgagg gagttcgtgt tcaagaacat cgacggatac 600
ttcaagattt acagcaagca caccccaatc aacctggtgc gcgacctgcc tcagggtttc 660
tctgctctgg agccactggt ggacctgcct atcggcatca acatcacccg cttccagact 720
ctgctggctc tgcaccgttc ctacctgact ccaggcgact catcttctgg atggactgct 780
ggagctgctg cttactacgt gggctacctg cagcctcgca ccttcctgct gaagtacaac 840
gaaaacggaa ccatcactga cgccgtcgac tgcgctctgg accctctgtc agaaaccaag 900
tgcactctga agtccttcac cgtggagaag ggcatctacc agacttcaaa cttcagggtg 960
cagcccaccg aatccatcgt cagattccct aacatcacta acctgtgccc cttcggagag 1020
gtcttcaacg ccacccgctt cgcttccgtg tacgcctgga acaggaagag aatctcaaac 1080
tgcgtcgctg actactccgt gctgtacaac tcagcctcct tcagcacctt caagtgctac 1140
ggcgtgtcac caactaagct gaacgacctg tgcttcacca acgtctacgc cgactccttc 1200
gtgatcaggg gagacgaggt cagacagatc gctcctggcc agactggaaa gatcgccgac 1260
tacaactaca agctgcccga cgacttcacc ggttgcgtca tcgcttggaa cagcaacaac 1320
ctggactcta aagtgggtgg caactacaac tacctgtacc gcctgttccg taagtcaaac 1380
ctgaagccat tcgagaggga catcagcact gaaatctacc aggccggatc taccccttgc 1440
aacggtgtcg agggcttcaa ctgctacttc cccctgcagt cctacggttt ccagccaact 1500
aacggtgtgg gctaccagcc ttacagagtg gtcgtgctga gcttcgaact gctccacgct 1560
cctgctactg tgtgcggtcc aaagaagtct accaacctgg tcaagaacaa gtgcgtgaac 1620
ttcaacttca acggcctgac cggaactggt gtcctgaccg agagcaacaa gaagttcctg 1680
cccttccagc agttcggaag ggacatcgct gacaccactg acgctgtgcg cgaccctcag 1740
accctggaaa tcctggacat cactccatgc tcattcggag gtgtctccgt gatcacccct 1800
ggcaccaaca cttctaacca ggtcgctgtg ctgtaccagg acgtcaactg caccgaggtc 1860
cctgtggcca tccacgctga ccagctgacc cccacttggc gcgtgtactc caccggcagc 1920
aacgtgttcc agactcgtgc tggttgcctg atcggcgccg agcacgtgaa caacagctac 1980
gaatgcgaca tccccatcgg cgctggaatc tgcgcctctt accagaccca gactaacagc 2040
ccacgcaggg ctcgctctgt ggcctctcag tcaatcatcg cttacaccat gtcactgggc 2100
gctgaaaact ccgtggccta ctctaacaac tcaatcgcca tccccaccaa cttcactatc 2160
agcgtgacca ctgagatcct gccagtcagc atgaccaaga cttctgtgga ctgcactatg 2220
tacatctgcg gagacagcac cgaatgctct aacctgctgc tgcagtacgg ctctttctgc 2280
acccagctga accgtgctct gactggaatc gccgtggagc aggacaagaa cactcaggaa 2340
gtcttcgctc aggtgaagca aatctacaag accccaccta tcaaggactt cggcggattc 2400
aacttctccc agatcctgcc tgacccctcc aagccaagca agcgctcttt catcgaggac 2460
ctgctgttca acaaggtcac tctggccgac gctggattca tcaagcagta cggagactgc 2520
ctgggtgaca tcgccgctcg tgacctgatc tgcgctcaga agttcaacgg tctgaccgtg 2580
ctgcccccac tgctgactga cgaaatgatc gcccagtaca ctagcgccct gctggctgga 2640
accatcactt ctggttggac cttcggtgct ggcgccgctc tgcagatccc tttcgctatg 2700
cagatggcct accgtttcaa cggaatcggt gtcacccaga acgtgctgta cgagaaccag 2760
aagctgatcg ctaaccagtt caactcagcc atcggaaaga tccaggacag cctgagctct 2820
actgcctctg ctctgggcaa gctgcaggac gtcgtgaacc agaacgccca ggctctgaac 2880
accctggtca agcagctgtc atccaacttc ggtgctatca gctctgtgct gaacgacatc 2940
ctgtcccgcc tggacaaggt cgaggccgaa gtgcagatcg accgcctgat cactggccgt 3000
ctgcagtcac tgcagaccta cgtgactcag cagctgatca gggccgctga aatcagagcc 3060
tccgctaacc tggccgctac caagatgagc gagtgcgtcc tgggtcaatc taagcgtgtg 3120
gacttctgcg gcaagggata ccacctgatg tcattccctc agtctgctcc ccacggtgtg 3180
gtgttcctgc acgtcaccta cgtgccagcc caggagaaga acttcaccac tgcccctgct 3240
atctgccacg acggcaaggc tcacttcccc agggaaggtg tcttcgtgag caacggcacc 3300
cactggttcg tcactcagag aaacttctac gagccacaga tcatcaccac tgacaacact 3360
ttcgtgtctg gaaactgcga cgtggtcatc ggtatcgtca acaacaccgt gtacgacccc 3420
ctgcagccag agctggactc attcaaggag gaactggaca agtacttcaa gaaccacacc 3480
tcccctgacg tcgacctggg cgacatctca ggaatcaacg cttccgtcgt gaacatccag 3540
aaggagatcg accgcctgaa cgaagtggcc aagaacctga acgaaagcct gatcgacctg 3600
caggagctgg gcaagtacga acagtacatc aagtggcctc accaccacca ccaccac 3657
<210> 44
<211> 1219
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 44
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val
1 5 10 15
Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe
20 25 30
Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu
35 40 45
His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp
50 55 60
Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp
65 70 75 80
Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu
85 90 95
Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser
100 105 110
Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile
115 120 125
Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr
130 135 140
Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
145 150 155 160
Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
165 170 175
Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
180 185 190
Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
195 200 205
Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
210 215 220
Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
225 230 235 240
Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
245 250 255
Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
260 265 270
Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
275 280 285
Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
290 295 300
Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
305 310 315 320
Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
325 330 335
Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
340 345 350
Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
355 360 365
Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
370 375 380
Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
385 390 395 400
Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
405 410 415
Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
420 425 430
Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
435 440 445
Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
450 455 460
Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
465 470 475 480
Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
485 490 495
Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
500 505 510
Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
515 520 525
Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
530 535 540
Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
545 550 555 560
Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val
565 570 575
Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
580 585 590
Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
595 600 605
Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile
610 615 620
His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
625 630 635 640
Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
645 650 655
Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
660 665 670
Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala Arg Ser Val Ala
675 680 685
Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser
690 695 700
Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile
705 710 715 720
Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val
725 730 735
Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu
740 745 750
Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr
755 760 765
Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln
770 775 780
Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe
785 790 795 800
Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser
805 810 815
Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly
820 825 830
Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp
835 840 845
Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu
850 855 860
Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly
865 870 875 880
Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln Ile
885 890 895
Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr
900 905 910
Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn
915 920 925
Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala
930 935 940
Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn
945 950 955 960
Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val
965 970 975
Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala Glu Val Gln
980 985 990
Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val
995 1000 1005
Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn
1010 1015 1020
Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys
1025 1030 1035
Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro
1040 1045 1050
Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val
1055 1060 1065
Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His
1070 1075 1080
Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn
1085 1090 1095
Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln
1100 1105 1110
Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val
1115 1120 1125
Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro
1130 1135 1140
Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn
1145 1150 1155
His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn
1160 1165 1170
Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu
1175 1180 1185
Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu
1190 1195 1200
Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro His His His His His
1205 1210 1215
His
<210> 45
<211> 318
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 45
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn
290 295 300
Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn
305 310 315
<210> 46
<211> 955
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 46
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Ile Thr Asn
1 5 10 15
Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val
20 25 30
Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser
35 40 45
Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val
50 55 60
Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp
65 70 75 80
Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln
85 90 95
Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr
100 105 110
Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly
115 120 125
Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys
130 135 140
Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr
145 150 155 160
Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser
165 170 175
Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val
180 185 190
Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly
195 200 205
Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn
210 215 220
Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys
225 230 235 240
Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp
245 250 255
Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys
260 265 270
Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn
275 280 285
Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val
290 295 300
Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr
305 310 315 320
Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu
325 330 335
His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile
340 345 350
Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala Arg Ser
355 360 365
Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu
370 375 380
Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe
385 390 395 400
Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr
405 410 415
Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser
420 425 430
Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala
435 440 445
Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe
450 455 460
Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly
465 470 475 480
Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys
485 490 495
Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp
500 505 510
Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala
515 520 525
Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro
530 535 540
Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu
545 550 555 560
Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu
565 570 575
Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly
580 585 590
Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln
595 600 605
Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala
610 615 620
Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala
625 630 635 640
Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser
645 650 655
Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala Glu
660 665 670
Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr
675 680 685
Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala
690 695 700
Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys
705 710 715 720
Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro Gln
725 730 735
Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val Pro Ala
740 745 750
Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His Asp Gly Lys
755 760 765
Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn Gly Thr His Trp
770 775 780
Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln Ile Ile Thr Thr Asp
785 790 795 800
Asn Thr Phe Val Ser Gly Asn Cys Asp Val Val Ile Gly Ile Val Asn
805 810 815
Asn Thr Val Tyr Asp Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys Glu
820 825 830
Glu Leu Asp Lys Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp Leu
835 840 845
Gly Asp Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu
850 855 860
Ile Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile
865 870 875 880
Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp
885 890 895
Tyr Ile Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val
900 905 910
Thr Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly
915 920 925
Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser Glu
930 935 940
Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr
945 950 955
<210> 47
<211> 2868
<212> DNA
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 47
atgttcgtct tcctggtgct gctgcccctg gtgtccagca tcactaacct gtgccccttc 60
ggagaggtct tcaacgccac ccgcttcgct tccgtgtacg cctggaacag gaagagaatc 120
tcaaactgcg tcgctgacta ctccgtgctg tacaactcag cctccttcag caccttcaag 180
tgctacggcg tgtcaccaac taagctgaac gacctgtgct tcaccaacgt ctacgccgac 240
tccttcgtga tcaggggaga cgaggtcaga cagatcgctc ctggccagac tggaaagatc 300
gccgactaca actacaagct gcccgacgac ttcaccggtt gcgtcatcgc ttggaacagc 360
aacaacctgg actctaaagt gggtggcaac tacaactacc tgtaccgcct gttccgtaag 420
tcaaacctga agccattcga gagggacatc agcactgaaa tctaccaggc cggatctacc 480
ccttgcaacg gtgtcgaggg cttcaactgc tacttccccc tgcagtccta cggtttccag 540
ccaactaacg gtgtgggcta ccagccttac agagtggtcg tgctgagctt cgaactgctc 600
cacgctcctg ctactgtgtg cggtccaaag aagtctacca acctggtcaa gaacaagtgc 660
gtgaacttca acttcaacgg cctgaccgga actggtgtcc tgaccgagag caacaagaag 720
ttcctgccct tccagcagtt cggaagggac atcgctgaca ccactgacgc tgtgcgcgac 780
cctcagaccc tggaaatcct ggacatcact ccatgctcat tcggaggtgt ctccgtgatc 840
acccctggca ccaacacttc taaccaggtc gctgtgctgt accaggacgt caactgcacc 900
gaggtccctg tggccatcca cgctgaccag ctgaccccca cttggcgcgt gtactccacc 960
ggcagcaacg tgttccagac tcgtgctggt tgcctgatcg gcgccgagca cgtgaacaac 1020
agctacgaat gcgacatccc catcggcgct ggaatctgcg cctcttacca gacccagact 1080
aacagcccac gcagggctcg ctctgtggcc tctcagtcaa tcatcgctta caccatgtca 1140
ctgggcgctg aaaactccgt ggcctactct aacaactcaa tcgccatccc caccaacttc 1200
actatcagcg tgaccactga gatcctgcca gtcagcatga ccaagacttc tgtggactgc 1260
actatgtaca tctgcggaga cagcaccgaa tgctctaacc tgctgctgca gtacggctct 1320
ttctgcaccc agctgaaccg tgctctgact ggaatcgccg tggagcagga caagaacact 1380
caggaagtct tcgctcaggt gaagcaaatc tacaagaccc cacctatcaa ggacttcggc 1440
ggattcaact tctcccagat cctgcctgac ccctccaagc caagcaagcg ctctttcatc 1500
gaggacctgc tgttcaacaa ggtcactctg gccgacgctg gattcatcaa gcagtacgga 1560
gactgcctgg gtgacatcgc cgctcgtgac ctgatctgcg ctcagaagtt caacggtctg 1620
accgtgctgc ccccactgct gactgacgaa atgatcgccc agtacactag cgccctgctg 1680
gctggaacca tcacttctgg ttggaccttc ggtgctggcg ccgctctgca gatccctttc 1740
gctatgcaga tggcctaccg tttcaacgga atcggtgtca cccagaacgt gctgtacgag 1800
aaccagaagc tgatcgctaa ccagttcaac tcagccatcg gaaagatcca ggacagcctg 1860
agctctactg cctctgctct gggcaagctg caggacgtcg tgaaccagaa cgcccaggct 1920
ctgaacaccc tggtcaagca gctgtcatcc aacttcggtg ctatcagctc tgtgctgaac 1980
gacatcctgt cccgcctgga caaggtcgag gccgaagtgc agatcgaccg cctgatcact 2040
ggccgtctgc agtcactgca gacctacgtg actcagcagc tgatcagggc cgctgaaatc 2100
agagcctccg ctaacctggc cgctaccaag atgagcgagt gcgtcctggg tcaatctaag 2160
cgtgtggact tctgcggcaa gggataccac ctgatgtcat tccctcagtc tgctccccac 2220
ggtgtggtgt tcctgcacgt cacctacgtg ccagcccagg agaagaactt caccactgcc 2280
cctgctatct gccacgacgg caaggctcac ttccccaggg aaggtgtctt cgtgagcaac 2340
ggcacccact ggttcgtcac tcagagaaac ttctacgagc cacagatcat caccactgac 2400
aacactttcg tgtctggaaa ctgcgacgtg gtcatcggta tcgtcaacaa caccgtgtac 2460
gaccccctgc agccagagct ggactcattc aaggaggaac tggacaagta cttcaagaac 2520
cacacctccc ctgacgtcga cctgggcgac atctcaggaa tcaacgcttc cgtcgtgaac 2580
atccagaagg agatcgaccg cctgaacgaa gtggccaaga acctgaacga aagcctgatc 2640
gacctgcagg agctgggcaa gtacgaacag tacatcaagt ggccttggta catctggctg 2700
ggtttcatcg ctggcctcat cgctatcgtg atggtgacca tcatgctgtg ctgcatgact 2760
tcatgctgct cctgcctgaa gggctgctgc agctgcggat cttgctgcaa gttcgacgag 2820
gacgactctg aacccgtcct gaagggcgtg aagctgcact acacctaa 2868
<210> 48
<211> 942
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 48
Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe
1 5 10 15
Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala
20 25 30
Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys
35 40 45
Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val
50 55 60
Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala
65 70 75 80
Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp
85 90 95
Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser
100 105 110
Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser
115 120 125
Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala
130 135 140
Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro
145 150 155 160
Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro
165 170 175
Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr
180 185 190
Val Cys Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val
195 200 205
Asn Phe Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser
210 215 220
Asn Lys Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp
225 230 235 240
Thr Thr Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile
245 250 255
Thr Pro Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn
260 265 270
Thr Ser Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu
275 280 285
Val Pro Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val
290 295 300
Tyr Ser Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile
305 310 315 320
Gly Ala Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly
325 330 335
Ala Gly Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg
340 345 350
Ala Arg Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu
355 360 365
Gly Ala Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro
370 375 380
Thr Asn Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met
385 390 395 400
Thr Lys Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr
405 410 415
Glu Cys Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu
420 425 430
Asn Arg Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln
435 440 445
Glu Val Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys
450 455 460
Asp Phe Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys
465 470 475 480
Pro Ser Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr
485 490 495
Leu Ala Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp
500 505 510
Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr
515 520 525
Val Leu Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser
530 535 540
Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly
545 550 555 560
Ala Ala Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn
565 570 575
Gly Ile Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile
580 585 590
Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser
595 600 605
Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn
610 615 620
Ala Gln Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly
625 630 635 640
Ala Ile Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val
645 650 655
Glu Ala Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser
660 665 670
Leu Gln Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg
675 680 685
Ala Ser Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly
690 695 700
Gln Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser
705 710 715 720
Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr
725 730 735
Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His
740 745 750
Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn Gly
755 760 765
Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln Ile Ile
770 775 780
Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val Val Ile Gly
785 790 795 800
Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro Glu Leu Asp Ser
805 810 815
Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn His Thr Ser Pro Asp
820 825 830
Val Asp Leu Gly Asp Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile
835 840 845
Gln Lys Glu Ile Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu
850 855 860
Ser Leu Ile Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys
865 870 875 880
Trp Pro Trp Tyr Ile Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile
885 890 895
Val Met Val Thr Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys
900 905 910
Leu Lys Gly Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp
915 920 925
Asp Ser Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr
930 935 940
<210> 49
<211> 960
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 49
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Ile Thr Asn
1 5 10 15
Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val
20 25 30
Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser
35 40 45
Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val
50 55 60
Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp
65 70 75 80
Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln
85 90 95
Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr
100 105 110
Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly
115 120 125
Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys
130 135 140
Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr
145 150 155 160
Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser
165 170 175
Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val
180 185 190
Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly
195 200 205
Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn
210 215 220
Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys
225 230 235 240
Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp
245 250 255
Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys
260 265 270
Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn
275 280 285
Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val
290 295 300
Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr
305 310 315 320
Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu
325 330 335
His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile
340 345 350
Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala Arg Ser
355 360 365
Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu
370 375 380
Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe
385 390 395 400
Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr
405 410 415
Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser
420 425 430
Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala
435 440 445
Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe
450 455 460
Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly
465 470 475 480
Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys
485 490 495
Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp
500 505 510
Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala
515 520 525
Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro
530 535 540
Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu
545 550 555 560
Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu
565 570 575
Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly
580 585 590
Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln
595 600 605
Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala
610 615 620
Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala
625 630 635 640
Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser
645 650 655
Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala Glu
660 665 670
Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr
675 680 685
Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala
690 695 700
Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys
705 710 715 720
Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro Gln
725 730 735
Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val Pro Ala
740 745 750
Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His Asp Gly Lys
755 760 765
Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn Gly Thr His Trp
770 775 780
Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln Ile Ile Thr Thr Asp
785 790 795 800
Asn Thr Phe Val Ser Gly Asn Cys Asp Val Val Ile Gly Ile Val Asn
805 810 815
Asn Thr Val Tyr Asp Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys Glu
820 825 830
Glu Leu Asp Lys Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp Leu
835 840 845
Gly Asp Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu
850 855 860
Ile Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile
865 870 875 880
Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp
885 890 895
Tyr Ile Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val
900 905 910
Thr Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly
915 920 925
Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser Glu
930 935 940
Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr His His His His His
945 950 955 960
<210> 50
<211> 2883
<212> DNA
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 50
atgttcgtct tcctggtgct gctgcccctg gtgtccagca tcactaacct gtgccccttc 60
ggagaggtct tcaacgccac ccgcttcgct tccgtgtacg cctggaacag gaagagaatc 120
tcaaactgcg tcgctgacta ctccgtgctg tacaactcag cctccttcag caccttcaag 180
tgctacggcg tgtcaccaac taagctgaac gacctgtgct tcaccaacgt ctacgccgac 240
tccttcgtga tcaggggaga cgaggtcaga cagatcgctc ctggccagac tggaaagatc 300
gccgactaca actacaagct gcccgacgac ttcaccggtt gcgtcatcgc ttggaacagc 360
aacaacctgg actctaaagt gggtggcaac tacaactacc tgtaccgcct gttccgtaag 420
tcaaacctga agccattcga gagggacatc agcactgaaa tctaccaggc cggatctacc 480
ccttgcaacg gtgtcgaggg cttcaactgc tacttccccc tgcagtccta cggtttccag 540
ccaactaacg gtgtgggcta ccagccttac agagtggtcg tgctgagctt cgaactgctc 600
cacgctcctg ctactgtgtg cggtccaaag aagtctacca acctggtcaa gaacaagtgc 660
gtgaacttca acttcaacgg cctgaccgga actggtgtcc tgaccgagag caacaagaag 720
ttcctgccct tccagcagtt cggaagggac atcgctgaca ccactgacgc tgtgcgcgac 780
cctcagaccc tggaaatcct ggacatcact ccatgctcat tcggaggtgt ctccgtgatc 840
acccctggca ccaacacttc taaccaggtc gctgtgctgt accaggacgt caactgcacc 900
gaggtccctg tggccatcca cgctgaccag ctgaccccca cttggcgcgt gtactccacc 960
ggcagcaacg tgttccagac tcgtgctggt tgcctgatcg gcgccgagca cgtgaacaac 1020
agctacgaat gcgacatccc catcggcgct ggaatctgcg cctcttacca gacccagact 1080
aacagcccac gcagggctcg ctctgtggcc tctcagtcaa tcatcgctta caccatgtca 1140
ctgggcgctg aaaactccgt ggcctactct aacaactcaa tcgccatccc caccaacttc 1200
actatcagcg tgaccactga gatcctgcca gtcagcatga ccaagacttc tgtggactgc 1260
actatgtaca tctgcggaga cagcaccgaa tgctctaacc tgctgctgca gtacggctct 1320
ttctgcaccc agctgaaccg tgctctgact ggaatcgccg tggagcagga caagaacact 1380
caggaagtct tcgctcaggt gaagcaaatc tacaagaccc cacctatcaa ggacttcggc 1440
ggattcaact tctcccagat cctgcctgac ccctccaagc caagcaagcg ctctttcatc 1500
gaggacctgc tgttcaacaa ggtcactctg gccgacgctg gattcatcaa gcagtacgga 1560
gactgcctgg gtgacatcgc cgctcgtgac ctgatctgcg ctcagaagtt caacggtctg 1620
accgtgctgc ccccactgct gactgacgaa atgatcgccc agtacactag cgccctgctg 1680
gctggaacca tcacttctgg ttggaccttc ggtgctggcg ccgctctgca gatccctttc 1740
gctatgcaga tggcctaccg tttcaacgga atcggtgtca cccagaacgt gctgtacgag 1800
aaccagaagc tgatcgctaa ccagttcaac tcagccatcg gaaagatcca ggacagcctg 1860
agctctactg cctctgctct gggcaagctg caggacgtcg tgaaccagaa cgcccaggct 1920
ctgaacaccc tggtcaagca gctgtcatcc aacttcggtg ctatcagctc tgtgctgaac 1980
gacatcctgt cccgcctgga caaggtcgag gccgaagtgc agatcgaccg cctgatcact 2040
ggccgtctgc agtcactgca gacctacgtg actcagcagc tgatcagggc cgctgaaatc 2100
agagcctccg ctaacctggc cgctaccaag atgagcgagt gcgtcctggg tcaatctaag 2160
cgtgtggact tctgcggcaa gggataccac ctgatgtcat tccctcagtc tgctccccac 2220
ggtgtggtgt tcctgcacgt cacctacgtg ccagcccagg agaagaactt caccactgcc 2280
cctgctatct gccacgacgg caaggctcac ttccccaggg aaggtgtctt cgtgagcaac 2340
ggcacccact ggttcgtcac tcagagaaac ttctacgagc cacagatcat caccactgac 2400
aacactttcg tgtctggaaa ctgcgacgtg gtcatcggta tcgtcaacaa caccgtgtac 2460
gaccccctgc agccagagct ggactcattc aaggaggaac tggacaagta cttcaagaac 2520
cacacctccc ctgacgtcga cctgggcgac atctcaggaa tcaacgcttc cgtcgtgaac 2580
atccagaagg agatcgaccg cctgaacgaa gtggccaaga acctgaacga aagcctgatc 2640
gacctgcagg agctgggcaa gtacgaacag tacatcaagt ggccttggta catctggctg 2700
ggtttcatcg ctggcctcat cgctatcgtg atggtgacca tcatgctgtg ctgcatgact 2760
tcatgctgct cctgcctgaa gggctgctgc agctgcggat cttgctgcaa gttcgacgag 2820
gacgactctg aacccgtcct gaagggcgtg aagctgcact acacccacca ccaccaccac 2880
cac 2883
<210> 51
<211> 947
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 51
Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe
1 5 10 15
Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala
20 25 30
Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys
35 40 45
Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val
50 55 60
Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala
65 70 75 80
Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp
85 90 95
Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser
100 105 110
Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser
115 120 125
Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala
130 135 140
Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro
145 150 155 160
Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro
165 170 175
Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr
180 185 190
Val Cys Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val
195 200 205
Asn Phe Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser
210 215 220
Asn Lys Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp
225 230 235 240
Thr Thr Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile
245 250 255
Thr Pro Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn
260 265 270
Thr Ser Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu
275 280 285
Val Pro Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val
290 295 300
Tyr Ser Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile
305 310 315 320
Gly Ala Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly
325 330 335
Ala Gly Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg
340 345 350
Ala Arg Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu
355 360 365
Gly Ala Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro
370 375 380
Thr Asn Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met
385 390 395 400
Thr Lys Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr
405 410 415
Glu Cys Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu
420 425 430
Asn Arg Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln
435 440 445
Glu Val Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys
450 455 460
Asp Phe Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys
465 470 475 480
Pro Ser Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr
485 490 495
Leu Ala Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp
500 505 510
Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr
515 520 525
Val Leu Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser
530 535 540
Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly
545 550 555 560
Ala Ala Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn
565 570 575
Gly Ile Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile
580 585 590
Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser
595 600 605
Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn
610 615 620
Ala Gln Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly
625 630 635 640
Ala Ile Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val
645 650 655
Glu Ala Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser
660 665 670
Leu Gln Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg
675 680 685
Ala Ser Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly
690 695 700
Gln Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser
705 710 715 720
Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr
725 730 735
Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His
740 745 750
Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn Gly
755 760 765
Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln Ile Ile
770 775 780
Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val Val Ile Gly
785 790 795 800
Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro Glu Leu Asp Ser
805 810 815
Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn His Thr Ser Pro Asp
820 825 830
Val Asp Leu Gly Asp Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile
835 840 845
Gln Lys Glu Ile Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu
850 855 860
Ser Leu Ile Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys
865 870 875 880
Trp Pro Trp Tyr Ile Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile
885 890 895
Val Met Val Thr Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys
900 905 910
Leu Lys Gly Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp
915 920 925
Asp Ser Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr His His
930 935 940
His His His
945
<210> 52
<211> 901
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 52
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Ile Thr Asn
1 5 10 15
Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val
20 25 30
Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser
35 40 45
Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val
50 55 60
Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp
65 70 75 80
Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln
85 90 95
Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr
100 105 110
Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly
115 120 125
Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys
130 135 140
Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr
145 150 155 160
Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser
165 170 175
Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val
180 185 190
Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly
195 200 205
Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn
210 215 220
Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys
225 230 235 240
Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp
245 250 255
Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys
260 265 270
Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn
275 280 285
Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val
290 295 300
Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr
305 310 315 320
Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu
325 330 335
His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile
340 345 350
Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala Arg Ser
355 360 365
Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu
370 375 380
Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe
385 390 395 400
Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr
405 410 415
Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser
420 425 430
Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala
435 440 445
Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe
450 455 460
Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly
465 470 475 480
Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys
485 490 495
Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp
500 505 510
Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala
515 520 525
Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro
530 535 540
Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu
545 550 555 560
Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu
565 570 575
Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly
580 585 590
Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln
595 600 605
Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala
610 615 620
Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala
625 630 635 640
Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser
645 650 655
Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala Glu
660 665 670
Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr
675 680 685
Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala
690 695 700
Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys
705 710 715 720
Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro Gln
725 730 735
Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val Pro Ala
740 745 750
Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His Asp Gly Lys
755 760 765
Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn Gly Thr His Trp
770 775 780
Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln Ile Ile Thr Thr Asp
785 790 795 800
Asn Thr Phe Val Ser Gly Asn Cys Asp Val Val Ile Gly Ile Val Asn
805 810 815
Asn Thr Val Tyr Asp Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys Glu
820 825 830
Glu Leu Asp Lys Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp Leu
835 840 845
Gly Asp Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu
850 855 860
Ile Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile
865 870 875 880
Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro His
885 890 895
His His His His His
900
<210> 53
<211> 2706
<212> DNA
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 53
atgttcgtct tcctggtgct gctgcccctg gtgtccagca tcactaacct gtgccccttc 60
ggagaggtct tcaacgccac ccgcttcgct tccgtgtacg cctggaacag gaagagaatc 120
tcaaactgcg tcgctgacta ctccgtgctg tacaactcag cctccttcag caccttcaag 180
tgctacggcg tgtcaccaac taagctgaac gacctgtgct tcaccaacgt ctacgccgac 240
tccttcgtga tcaggggaga cgaggtcaga cagatcgctc ctggccagac tggaaagatc 300
gccgactaca actacaagct gcccgacgac ttcaccggtt gcgtcatcgc ttggaacagc 360
aacaacctgg actctaaagt gggtggcaac tacaactacc tgtaccgcct gttccgtaag 420
tcaaacctga agccattcga gagggacatc agcactgaaa tctaccaggc cggatctacc 480
ccttgcaacg gtgtcgaggg cttcaactgc tacttccccc tgcagtccta cggtttccag 540
ccaactaacg gtgtgggcta ccagccttac agagtggtcg tgctgagctt cgaactgctc 600
cacgctcctg ctactgtgtg cggtccaaag aagtctacca acctggtcaa gaacaagtgc 660
gtgaacttca acttcaacgg cctgaccgga actggtgtcc tgaccgagag caacaagaag 720
ttcctgccct tccagcagtt cggaagggac atcgctgaca ccactgacgc tgtgcgcgac 780
cctcagaccc tggaaatcct ggacatcact ccatgctcat tcggaggtgt ctccgtgatc 840
acccctggca ccaacacttc taaccaggtc gctgtgctgt accaggacgt caactgcacc 900
gaggtccctg tggccatcca cgctgaccag ctgaccccca cttggcgcgt gtactccacc 960
ggcagcaacg tgttccagac tcgtgctggt tgcctgatcg gcgccgagca cgtgaacaac 1020
agctacgaat gcgacatccc catcggcgct ggaatctgcg cctcttacca gacccagact 1080
aacagcccac gcagggctcg ctctgtggcc tctcagtcaa tcatcgctta caccatgtca 1140
ctgggcgctg aaaactccgt ggcctactct aacaactcaa tcgccatccc caccaacttc 1200
actatcagcg tgaccactga gatcctgcca gtcagcatga ccaagacttc tgtggactgc 1260
actatgtaca tctgcggaga cagcaccgaa tgctctaacc tgctgctgca gtacggctct 1320
ttctgcaccc agctgaaccg tgctctgact ggaatcgccg tggagcagga caagaacact 1380
caggaagtct tcgctcaggt gaagcaaatc tacaagaccc cacctatcaa ggacttcggc 1440
ggattcaact tctcccagat cctgcctgac ccctccaagc caagcaagcg ctctttcatc 1500
gaggacctgc tgttcaacaa ggtcactctg gccgacgctg gattcatcaa gcagtacgga 1560
gactgcctgg gtgacatcgc cgctcgtgac ctgatctgcg ctcagaagtt caacggtctg 1620
accgtgctgc ccccactgct gactgacgaa atgatcgccc agtacactag cgccctgctg 1680
gctggaacca tcacttctgg ttggaccttc ggtgctggcg ccgctctgca gatccctttc 1740
gctatgcaga tggcctaccg tttcaacgga atcggtgtca cccagaacgt gctgtacgag 1800
aaccagaagc tgatcgctaa ccagttcaac tcagccatcg gaaagatcca ggacagcctg 1860
agctctactg cctctgctct gggcaagctg caggacgtcg tgaaccagaa cgcccaggct 1920
ctgaacaccc tggtcaagca gctgtcatcc aacttcggtg ctatcagctc tgtgctgaac 1980
gacatcctgt cccgcctgga caaggtcgag gccgaagtgc agatcgaccg cctgatcact 2040
ggccgtctgc agtcactgca gacctacgtg actcagcagc tgatcagggc cgctgaaatc 2100
agagcctccg ctaacctggc cgctaccaag atgagcgagt gcgtcctggg tcaatctaag 2160
cgtgtggact tctgcggcaa gggataccac ctgatgtcat tccctcagtc tgctccccac 2220
ggtgtggtgt tcctgcacgt cacctacgtg ccagcccagg agaagaactt caccactgcc 2280
cctgctatct gccacgacgg caaggctcac ttccccaggg aaggtgtctt cgtgagcaac 2340
ggcacccact ggttcgtcac tcagagaaac ttctacgagc cacagatcat caccactgac 2400
aacactttcg tgtctggaaa ctgcgacgtg gtcatcggta tcgtcaacaa caccgtgtac 2460
gaccccctgc agccagagct ggactcattc aaggaggaac tggacaagta cttcaagaac 2520
cacacctccc ctgacgtcga cctgggcgac atctcaggaa tcaacgcttc cgtcgtgaac 2580
atccagaagg agatcgaccg cctgaacgaa gtggccaaga acctgaacga aagcctgatc 2640
gacctgcagg agctgggcaa gtacgaacag tacatcaagt ggcctcacca ccaccaccac 2700
caccac 2706
<210> 54
<211> 888
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 54
Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe
1 5 10 15
Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala
20 25 30
Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys
35 40 45
Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val
50 55 60
Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala
65 70 75 80
Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp
85 90 95
Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser
100 105 110
Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser
115 120 125
Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala
130 135 140
Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro
145 150 155 160
Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro
165 170 175
Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr
180 185 190
Val Cys Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val
195 200 205
Asn Phe Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser
210 215 220
Asn Lys Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp
225 230 235 240
Thr Thr Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile
245 250 255
Thr Pro Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn
260 265 270
Thr Ser Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu
275 280 285
Val Pro Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val
290 295 300
Tyr Ser Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile
305 310 315 320
Gly Ala Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly
325 330 335
Ala Gly Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg
340 345 350
Ala Arg Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu
355 360 365
Gly Ala Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro
370 375 380
Thr Asn Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met
385 390 395 400
Thr Lys Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr
405 410 415
Glu Cys Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu
420 425 430
Asn Arg Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln
435 440 445
Glu Val Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys
450 455 460
Asp Phe Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys
465 470 475 480
Pro Ser Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr
485 490 495
Leu Ala Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp
500 505 510
Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr
515 520 525
Val Leu Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser
530 535 540
Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly
545 550 555 560
Ala Ala Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn
565 570 575
Gly Ile Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile
580 585 590
Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser
595 600 605
Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn
610 615 620
Ala Gln Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly
625 630 635 640
Ala Ile Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val
645 650 655
Glu Ala Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser
660 665 670
Leu Gln Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg
675 680 685
Ala Ser Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly
690 695 700
Gln Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser
705 710 715 720
Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr
725 730 735
Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His
740 745 750
Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn Gly
755 760 765
Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln Ile Ile
770 775 780
Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val Val Ile Gly
785 790 795 800
Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro Glu Leu Asp Ser
805 810 815
Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn His Thr Ser Pro Asp
820 825 830
Val Asp Leu Gly Asp Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile
835 840 845
Gln Lys Glu Ile Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu
850 855 860
Ser Leu Ile Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys
865 870 875 880
Trp Pro His His His His His His
885
<210> 55
<211> 3819
<212> DNA
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 55
atgttcgtct tcctggtgct gctgcccctg gtgtccagcc agtgcgtgaa cctgaccact 60
aggactcagc tgcctcccgc ttacaccaac tcattcactc gcggtgtgta ctaccctgac 120
aaggtcttcc gttcttcagt gctgcactca actcaggacc tgttcctgcc cttcttctcc 180
aacgtcacct ggttccacgc catccacgtg tccggcacca acggcactaa gcgcttcgac 240
aacccagtgc tgcctttcaa cgacggtgtc tacttcgctt caaccgagaa gtccaacatc 300
atccgtggat ggatcttcgg caccactctg gacagcaaga ctcagtctct gctgatcgtc 360
aacaacgcca ccaacgtggt catcaaggtc tgcgaattcc agttctgcaa cgacccattc 420
ctgggcgtct actaccacaa gaacaacaag tcatggatgg agtccgaatt ccgcgtctac 480
tccagcgcta acaactgcac tttcgagtac gtgtcccagc ctttcctgat ggacctggaa 540
ggaaagcagg gtaacttcaa gaacctgagg gagttcgtgt tcaagaacat cgacggatac 600
ttcaagattt acagcaagca caccccaatc aacctggtgc gcgacctgcc tcagggtttc 660
tctgctctgg agccactggt ggacctgcct atcggcatca acatcacccg cttccagact 720
ctgctggctc tgcaccgttc ctacctgact ccaggcgact catcttctgg atggactgct 780
ggagctgctg cttactacgt gggctacctg cagcctcgca ccttcctgct gaagtacaac 840
gaaaacggaa ccatcactga cgccgtcgac tgcgctctgg accctctgtc agaaaccaag 900
tgcactctga agtccttcac cgtggagaag ggcatctacc agacttcaaa cttcagggtg 960
cagcccaccg aatccatcgt cagattccct aacatcacta acctgtgccc cttcggagag 1020
gtcttcaacg ccacccgctt cgcttccgtg tacgcctgga acaggaagag aatctcaaac 1080
tgcgtcgctg actactccgt gctgtacaac tcagcctcct tcagcacctt caagtgctac 1140
ggcgtgtcac caactaagct gaacgacctg tgcttcacca acgtctacgc cgactccttc 1200
gtgatcaggg gagacgaggt cagacagatc gctcctggcc agactggaaa gatcgccgac 1260
tacaactaca agctgcccga cgacttcacc ggttgcgtca tcgcttggaa cagcaacaac 1320
ctggactcta aagtgggtgg caactacaac tacctgtacc gcctgttccg taagtcaaac 1380
ctgaagccat tcgagaggga catcagcact gaaatctacc aggccggatc taccccttgc 1440
aacggtgtcg agggcttcaa ctgctacttc cccctgcagt cctacggttt ccagccaact 1500
aacggtgtgg gctaccagcc ttacagagtg gtcgtgctga gcttcgaact gctccacgct 1560
cctgctactg tgtgcggtcc aaagaagtct accaacctgg tcaagaacaa gtgcgtgaac 1620
ttcaacttca acggcctgac cggaactggt gtcctgaccg agagcaacaa gaagttcctg 1680
cccttccagc agttcggaag ggacatcgct gacaccactg acgctgtgcg cgaccctcag 1740
accctggaaa tcctggacat cactccatgc tcattcggag gtgtctccgt gatcacccct 1800
ggcaccaaca cttctaacca ggtcgctgtg ctgtaccagg acgtcaactg caccgaggtc 1860
cctgtggcca tccacgctga ccagctgacc cccacttggc gcgtgtactc caccggcagc 1920
aacgtgttcc agactcgtgc tggttgcctg atcggcgccg agcacgtgaa caacagctac 1980
gaatgcgaca tccccatcgg cgctggaatc tgcgcctctt accagaccca gactaacagc 2040
ccacagcagg ctcagtctgt ggcctctcag tcaatcatcg cttacaccat gtcactgggc 2100
gctgaaaact ccgtggccta ctctaacaac tcaatcgcca tccccaccaa cttcactatc 2160
agcgtgacca ctgagatcct gccagtcagc atgaccaaga cttctgtgga ctgcactatg 2220
tacatctgcg gagacagcac cgaatgctct aacctgctgc tgcagtacgg ctctttctgc 2280
acccagctga accgtgctct gactggaatc gccgtggagc aggacaagaa cactcaggaa 2340
gtcttcgctc aggtgaagca aatctacaag accccaccta tcaaggactt cggcggattc 2400
aacttctccc agatcctgcc tgacccctcc aagccaagca agcgctcttt catcgaggac 2460
ctgctgttca acaaggtcac tctggccgac gctggattca tcaagcagta cggagactgc 2520
ctgggtgaca tcgccgctcg tgacctgatc tgcgctcaga agttcaacgg tctgaccgtg 2580
ctgcccccac tgctgactga cgaaatgatc gcccagtaca ctagcgccct gctggctgga 2640
accatcactt ctggttggac cttcggtgct ggcgccgctc tgcagatccc tttcgctatg 2700
cagatggcct accgtttcaa cggaatcggt gtcacccaga acgtgctgta cgagaaccag 2760
aagctgatcg ctaaccagtt caactcagcc atcggaaaga tccaggacag cctgagctct 2820
actgcctctg ctctgggcaa gctgcaggac gtcgtgaacc agaacgccca ggctctgaac 2880
accctggtca agcagctgtc atccaacttc ggtgctatca gctctgtgct gaacgacatc 2940
ctgtcccgcc tggacaaggt cgaggccgaa gtgcagatcg accgcctgat cactggccgt 3000
ctgcagtcac tgcagaccta cgtgactcag cagctgatca gggccgctga aatcagagcc 3060
tccgctaacc tggccgctac caagatgagc gagtgcgtcc tgggtcaatc taagcgtgtg 3120
gacttctgcg gcaagggata ccacctgatg tcattccctc agtctgctcc ccacggtgtg 3180
gtgttcctgc acgtcaccta cgtgccagcc caggagaaga acttcaccac tgcccctgct 3240
atctgccacg acggcaaggc tcacttcccc agggaaggtg tcttcgtgag caacggcacc 3300
cactggttcg tcactcagag aaacttctac gagccacaga tcatcaccac tgacaacact 3360
ttcgtgtctg gaaactgcga cgtggtcatc ggtatcgtca acaacaccgt gtacgacccc 3420
ctgcagccag agctggactc attcaaggag gaactggaca agtacttcaa gaaccacacc 3480
tcccctgacg tcgacctggg cgacatctca ggaatcaacg cttccgtcgt gaacatccag 3540
aaggagatcg accgcctgaa cgaagtggcc aagaacctga acgaaagcct gatcgacctg 3600
caggagctgg gcaagtacga acagtacatc aagtggcctt ggtacatctg gctgggtttc 3660
atcgctggcc tcatcgctat cgtgatggtg accatcatgc tgtgctgcat gacttcatgc 3720
tgctcctgcc tgaagggctg ctgcagctgc ggatcttgct gcaagttcga cgaggacgac 3780
tctgaacccg tcctgaaggg cgtgaagctg cactacacc 3819
<210> 56
<211> 1279
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 56
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val
1 5 10 15
Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe
20 25 30
Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu
35 40 45
His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp
50 55 60
Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp
65 70 75 80
Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu
85 90 95
Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser
100 105 110
Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile
115 120 125
Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr
130 135 140
Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
145 150 155 160
Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
165 170 175
Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
180 185 190
Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
195 200 205
Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
210 215 220
Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
225 230 235 240
Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
245 250 255
Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
260 265 270
Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
275 280 285
Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
290 295 300
Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
305 310 315 320
Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
325 330 335
Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
340 345 350
Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
355 360 365
Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
370 375 380
Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
385 390 395 400
Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
405 410 415
Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
420 425 430
Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
435 440 445
Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
450 455 460
Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
465 470 475 480
Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
485 490 495
Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
500 505 510
Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
515 520 525
Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
530 535 540
Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
545 550 555 560
Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val
565 570 575
Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
580 585 590
Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
595 600 605
Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile
610 615 620
His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
625 630 635 640
Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
645 650 655
Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
660 665 670
Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln Ser Val Ala
675 680 685
Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser
690 695 700
Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile
705 710 715 720
Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val
725 730 735
Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu
740 745 750
Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr
755 760 765
Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln
770 775 780
Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe
785 790 795 800
Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser
805 810 815
Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly
820 825 830
Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp
835 840 845
Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu
850 855 860
Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly
865 870 875 880
Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln Ile
885 890 895
Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr
900 905 910
Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn
915 920 925
Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala
930 935 940
Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn
945 950 955 960
Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val
965 970 975
Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala Glu Val Gln
980 985 990
Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val
995 1000 1005
Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn
1010 1015 1020
Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys
1025 1030 1035
Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro
1040 1045 1050
Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val
1055 1060 1065
Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His
1070 1075 1080
Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn
1085 1090 1095
Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln
1100 1105 1110
Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val
1115 1120 1125
Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro
1130 1135 1140
Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn
1145 1150 1155
His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn
1160 1165 1170
Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu
1175 1180 1185
Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu
1190 1195 1200
Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile Trp Leu
1205 1210 1215
Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr Ile Met
1220 1225 1230
Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly Cys Cys
1235 1240 1245
Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser Glu Pro
1250 1255 1260
Val Leu Lys Gly Val Lys Leu His Tyr Thr His His His His His
1265 1270 1275
His
<210> 57
<211> 3837
<212> DNA
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 57
atgttcgtct tcctggtgct gctgcccctg gtgtccagcc agtgcgtgaa cctgaccact 60
aggactcagc tgcctcccgc ttacaccaac tcattcactc gcggtgtgta ctaccctgac 120
aaggtcttcc gttcttcagt gctgcactca actcaggacc tgttcctgcc cttcttctcc 180
aacgtcacct ggttccacgc catccacgtg tccggcacca acggcactaa gcgcttcgac 240
aacccagtgc tgcctttcaa cgacggtgtc tacttcgctt caaccgagaa gtccaacatc 300
atccgtggat ggatcttcgg caccactctg gacagcaaga ctcagtctct gctgatcgtc 360
aacaacgcca ccaacgtggt catcaaggtc tgcgaattcc agttctgcaa cgacccattc 420
ctgggcgtct actaccacaa gaacaacaag tcatggatgg agtccgaatt ccgcgtctac 480
tccagcgcta acaactgcac tttcgagtac gtgtcccagc ctttcctgat ggacctggaa 540
ggaaagcagg gtaacttcaa gaacctgagg gagttcgtgt tcaagaacat cgacggatac 600
ttcaagattt acagcaagca caccccaatc aacctggtgc gcgacctgcc tcagggtttc 660
tctgctctgg agccactggt ggacctgcct atcggcatca acatcacccg cttccagact 720
ctgctggctc tgcaccgttc ctacctgact ccaggcgact catcttctgg atggactgct 780
ggagctgctg cttactacgt gggctacctg cagcctcgca ccttcctgct gaagtacaac 840
gaaaacggaa ccatcactga cgccgtcgac tgcgctctgg accctctgtc agaaaccaag 900
tgcactctga agtccttcac cgtggagaag ggcatctacc agacttcaaa cttcagggtg 960
cagcccaccg aatccatcgt cagattccct aacatcacta acctgtgccc cttcggagag 1020
gtcttcaacg ccacccgctt cgcttccgtg tacgcctgga acaggaagag aatctcaaac 1080
tgcgtcgctg actactccgt gctgtacaac tcagcctcct tcagcacctt caagtgctac 1140
ggcgtgtcac caactaagct gaacgacctg tgcttcacca acgtctacgc cgactccttc 1200
gtgatcaggg gagacgaggt cagacagatc gctcctggcc agactggaaa gatcgccgac 1260
tacaactaca agctgcccga cgacttcacc ggttgcgtca tcgcttggaa cagcaacaac 1320
ctggactcta aagtgggtgg caactacaac tacctgtacc gcctgttccg taagtcaaac 1380
ctgaagccat tcgagaggga catcagcact gaaatctacc aggccggatc taccccttgc 1440
aacggtgtcg agggcttcaa ctgctacttc cccctgcagt cctacggttt ccagccaact 1500
aacggtgtgg gctaccagcc ttacagagtg gtcgtgctga gcttcgaact gctccacgct 1560
cctgctactg tgtgcggtcc aaagaagtct accaacctgg tcaagaacaa gtgcgtgaac 1620
ttcaacttca acggcctgac cggaactggt gtcctgaccg agagcaacaa gaagttcctg 1680
cccttccagc agttcggaag ggacatcgct gacaccactg acgctgtgcg cgaccctcag 1740
accctggaaa tcctggacat cactccatgc tcattcggag gtgtctccgt gatcacccct 1800
ggcaccaaca cttctaacca ggtcgctgtg ctgtaccagg acgtcaactg caccgaggtc 1860
cctgtggcca tccacgctga ccagctgacc cccacttggc gcgtgtactc caccggcagc 1920
aacgtgttcc agactcgtgc tggttgcctg atcggcgccg agcacgtgaa caacagctac 1980
gaatgcgaca tccccatcgg cgctggaatc tgcgcctctt accagaccca gactaacagc 2040
ccacagcagg ctcagtctgt ggcctctcag tcaatcatcg cttacaccat gtcactgggc 2100
gctgaaaact ccgtggccta ctctaacaac tcaatcgcca tccccaccaa cttcactatc 2160
agcgtgacca ctgagatcct gccagtcagc atgaccaaga cttctgtgga ctgcactatg 2220
tacatctgcg gagacagcac cgaatgctct aacctgctgc tgcagtacgg ctctttctgc 2280
acccagctga accgtgctct gactggaatc gccgtggagc aggacaagaa cactcaggaa 2340
gtcttcgctc aggtgaagca aatctacaag accccaccta tcaaggactt cggcggattc 2400
aacttctccc agatcctgcc tgacccctcc aagccaagca agcgctcttt catcgaggac 2460
ctgctgttca acaaggtcac tctggccgac gctggattca tcaagcagta cggagactgc 2520
ctgggtgaca tcgccgctcg tgacctgatc tgcgctcaga agttcaacgg tctgaccgtg 2580
ctgcccccac tgctgactga cgaaatgatc gcccagtaca ctagcgccct gctggctgga 2640
accatcactt ctggttggac cttcggtgct ggcgccgctc tgcagatccc tttcgctatg 2700
cagatggcct accgtttcaa cggaatcggt gtcacccaga acgtgctgta cgagaaccag 2760
aagctgatcg ctaaccagtt caactcagcc atcggaaaga tccaggacag cctgagctct 2820
actgcctctg ctctgggcaa gctgcaggac gtcgtgaacc agaacgccca ggctctgaac 2880
accctggtca agcagctgtc atccaacttc ggtgctatca gctctgtgct gaacgacatc 2940
ctgtcccgcc tggacaaggt cgaggccgaa gtgcagatcg accgcctgat cactggccgt 3000
ctgcagtcac tgcagaccta cgtgactcag cagctgatca gggccgctga aatcagagcc 3060
tccgctaacc tggccgctac caagatgagc gagtgcgtcc tgggtcaatc taagcgtgtg 3120
gacttctgcg gcaagggata ccacctgatg tcattccctc agtctgctcc ccacggtgtg 3180
gtgttcctgc acgtcaccta cgtgccagcc caggagaaga acttcaccac tgcccctgct 3240
atctgccacg acggcaaggc tcacttcccc agggaaggtg tcttcgtgag caacggcacc 3300
cactggttcg tcactcagag aaacttctac gagccacaga tcatcaccac tgacaacact 3360
ttcgtgtctg gaaactgcga cgtggtcatc ggtatcgtca acaacaccgt gtacgacccc 3420
ctgcagccag agctggactc attcaaggag gaactggaca agtacttcaa gaaccacacc 3480
tcccctgacg tcgacctggg cgacatctca ggaatcaacg cttccgtcgt gaacatccag 3540
aaggagatcg accgcctgaa cgaagtggcc aagaacctga acgaaagcct gatcgacctg 3600
caggagctgg gcaagtacga acagtacatc aagtggcctt ggtacatctg gctgggtttc 3660
atcgctggcc tcatcgctat cgtgatggtg accatcatgc tgtgctgcat gacttcatgc 3720
tgctcctgcc tgaagggctg ctgcagctgc ggatcttgct gcaagttcga cgaggacgac 3780
tctgaacccg tcctgaaggg cgtgaagctg cactacaccc accaccacca ccaccac 3837
<210> 58
<211> 1266
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 58
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn
290 295 300
Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr
305 310 315 320
Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser
325 330 335
Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr
340 345 350
Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly
355 360 365
Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala
370 375 380
Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly
385 390 395 400
Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe
405 410 415
Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val
420 425 430
Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu
435 440 445
Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser
450 455 460
Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln
465 470 475 480
Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg
485 490 495
Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys
500 505 510
Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe
515 520 525
Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys
530 535 540
Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr
545 550 555 560
Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro
565 570 575
Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser
580 585 590
Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro
595 600 605
Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser
610 615 620
Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala
625 630 635 640
Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly
645 650 655
Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln
660 665 670
Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala
675 680 685
Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn
690 695 700
Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys
705 710 715 720
Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys
725 730 735
Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg
740 745 750
Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val
755 760 765
Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe
770 775 780
Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser
785 790 795 800
Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala
805 810 815
Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala
820 825 830
Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu
835 840 845
Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu
850 855 860
Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala
865 870 875 880
Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile
885 890 895
Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn
900 905 910
Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr
915 920 925
Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln
930 935 940
Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile
945 950 955 960
Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala
965 970 975
Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln
980 985 990
Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser
995 1000 1005
Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln
1010 1015 1020
Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser
1025 1030 1035
Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr
1040 1045 1050
Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile
1055 1060 1065
Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val
1070 1075 1080
Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu
1085 1090 1095
Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys
1100 1105 1110
Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu
1115 1120 1125
Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe
1130 1135 1140
Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly
1145 1150 1155
Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu
1160 1165 1170
Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln
1175 1180 1185
Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile
1190 1195 1200
Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr
1205 1210 1215
Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly
1220 1225 1230
Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser
1235 1240 1245
Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr His His His
1250 1255 1260
His His His
1265
<210> 59
<211> 1219
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 59
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val
1 5 10 15
Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe
20 25 30
Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu
35 40 45
His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp
50 55 60
Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp
65 70 75 80
Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu
85 90 95
Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser
100 105 110
Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile
115 120 125
Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr
130 135 140
Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
145 150 155 160
Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
165 170 175
Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
180 185 190
Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
195 200 205
Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
210 215 220
Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
225 230 235 240
Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
245 250 255
Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
260 265 270
Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
275 280 285
Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
290 295 300
Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
305 310 315 320
Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
325 330 335
Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
340 345 350
Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
355 360 365
Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
370 375 380
Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
385 390 395 400
Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
405 410 415
Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
420 425 430
Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
435 440 445
Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
450 455 460
Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
465 470 475 480
Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
485 490 495
Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
500 505 510
Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
515 520 525
Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
530 535 540
Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
545 550 555 560
Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val
565 570 575
Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
580 585 590
Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
595 600 605
Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile
610 615 620
His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
625 630 635 640
Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
645 650 655
Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
660 665 670
Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln Ser Val Ala
675 680 685
Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser
690 695 700
Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile
705 710 715 720
Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val
725 730 735
Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu
740 745 750
Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr
755 760 765
Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln
770 775 780
Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe
785 790 795 800
Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser
805 810 815
Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly
820 825 830
Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp
835 840 845
Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu
850 855 860
Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly
865 870 875 880
Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln Ile
885 890 895
Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr
900 905 910
Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn
915 920 925
Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala
930 935 940
Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn
945 950 955 960
Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val
965 970 975
Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala Glu Val Gln
980 985 990
Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val
995 1000 1005
Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn
1010 1015 1020
Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys
1025 1030 1035
Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro
1040 1045 1050
Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val
1055 1060 1065
Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His
1070 1075 1080
Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn
1085 1090 1095
Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln
1100 1105 1110
Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val
1115 1120 1125
Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro
1130 1135 1140
Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn
1145 1150 1155
His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn
1160 1165 1170
Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu
1175 1180 1185
Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu
1190 1195 1200
Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro His His His His His
1205 1210 1215
His
<210> 60
<211> 3657
<212> DNA
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 60
atgttcgtct tcctggtgct gctgcccctg gtgtccagcc agtgcgtgaa cctgaccact 60
aggactcagc tgcctcccgc ttacaccaac tcattcactc gcggtgtgta ctaccctgac 120
aaggtcttcc gttcttcagt gctgcactca actcaggacc tgttcctgcc cttcttctcc 180
aacgtcacct ggttccacgc catccacgtg tccggcacca acggcactaa gcgcttcgac 240
aacccagtgc tgcctttcaa cgacggtgtc tacttcgctt caaccgagaa gtccaacatc 300
atccgtggat ggatcttcgg caccactctg gacagcaaga ctcagtctct gctgatcgtc 360
aacaacgcca ccaacgtggt catcaaggtc tgcgaattcc agttctgcaa cgacccattc 420
ctgggcgtct actaccacaa gaacaacaag tcatggatgg agtccgaatt ccgcgtctac 480
tccagcgcta acaactgcac tttcgagtac gtgtcccagc ctttcctgat ggacctggaa 540
ggaaagcagg gtaacttcaa gaacctgagg gagttcgtgt tcaagaacat cgacggatac 600
ttcaagattt acagcaagca caccccaatc aacctggtgc gcgacctgcc tcagggtttc 660
tctgctctgg agccactggt ggacctgcct atcggcatca acatcacccg cttccagact 720
ctgctggctc tgcaccgttc ctacctgact ccaggcgact catcttctgg atggactgct 780
ggagctgctg cttactacgt gggctacctg cagcctcgca ccttcctgct gaagtacaac 840
gaaaacggaa ccatcactga cgccgtcgac tgcgctctgg accctctgtc agaaaccaag 900
tgcactctga agtccttcac cgtggagaag ggcatctacc agacttcaaa cttcagggtg 960
cagcccaccg aatccatcgt cagattccct aacatcacta acctgtgccc cttcggagag 1020
gtcttcaacg ccacccgctt cgcttccgtg tacgcctgga acaggaagag aatctcaaac 1080
tgcgtcgctg actactccgt gctgtacaac tcagcctcct tcagcacctt caagtgctac 1140
ggcgtgtcac caactaagct gaacgacctg tgcttcacca acgtctacgc cgactccttc 1200
gtgatcaggg gagacgaggt cagacagatc gctcctggcc agactggaaa gatcgccgac 1260
tacaactaca agctgcccga cgacttcacc ggttgcgtca tcgcttggaa cagcaacaac 1320
ctggactcta aagtgggtgg caactacaac tacctgtacc gcctgttccg taagtcaaac 1380
ctgaagccat tcgagaggga catcagcact gaaatctacc aggccggatc taccccttgc 1440
aacggtgtcg agggcttcaa ctgctacttc cccctgcagt cctacggttt ccagccaact 1500
aacggtgtgg gctaccagcc ttacagagtg gtcgtgctga gcttcgaact gctccacgct 1560
cctgctactg tgtgcggtcc aaagaagtct accaacctgg tcaagaacaa gtgcgtgaac 1620
ttcaacttca acggcctgac cggaactggt gtcctgaccg agagcaacaa gaagttcctg 1680
cccttccagc agttcggaag ggacatcgct gacaccactg acgctgtgcg cgaccctcag 1740
accctggaaa tcctggacat cactccatgc tcattcggag gtgtctccgt gatcacccct 1800
ggcaccaaca cttctaacca ggtcgctgtg ctgtaccagg acgtcaactg caccgaggtc 1860
cctgtggcca tccacgctga ccagctgacc cccacttggc gcgtgtactc caccggcagc 1920
aacgtgttcc agactcgtgc tggttgcctg atcggcgccg agcacgtgaa caacagctac 1980
gaatgcgaca tccccatcgg cgctggaatc tgcgcctctt accagaccca gactaacagc 2040
ccacagcagg ctcagtctgt ggcctctcag tcaatcatcg cttacaccat gtcactgggc 2100
gctgaaaact ccgtggccta ctctaacaac tcaatcgcca tccccaccaa cttcactatc 2160
agcgtgacca ctgagatcct gccagtcagc atgaccaaga cttctgtgga ctgcactatg 2220
tacatctgcg gagacagcac cgaatgctct aacctgctgc tgcagtacgg ctctttctgc 2280
acccagctga accgtgctct gactggaatc gccgtggagc aggacaagaa cactcaggaa 2340
gtcttcgctc aggtgaagca aatctacaag accccaccta tcaaggactt cggcggattc 2400
aacttctccc agatcctgcc tgacccctcc aagccaagca agcgctcttt catcgaggac 2460
ctgctgttca acaaggtcac tctggccgac gctggattca tcaagcagta cggagactgc 2520
ctgggtgaca tcgccgctcg tgacctgatc tgcgctcaga agttcaacgg tctgaccgtg 2580
ctgcccccac tgctgactga cgaaatgatc gcccagtaca ctagcgccct gctggctgga 2640
accatcactt ctggttggac cttcggtgct ggcgccgctc tgcagatccc tttcgctatg 2700
cagatggcct accgtttcaa cggaatcggt gtcacccaga acgtgctgta cgagaaccag 2760
aagctgatcg ctaaccagtt caactcagcc atcggaaaga tccaggacag cctgagctct 2820
actgcctctg ctctgggcaa gctgcaggac gtcgtgaacc agaacgccca ggctctgaac 2880
accctggtca agcagctgtc atccaacttc ggtgctatca gctctgtgct gaacgacatc 2940
ctgtcccgcc tggacaaggt cgaggccgaa gtgcagatcg accgcctgat cactggccgt 3000
ctgcagtcac tgcagaccta cgtgactcag cagctgatca gggccgctga aatcagagcc 3060
tccgctaacc tggccgctac caagatgagc gagtgcgtcc tgggtcaatc taagcgtgtg 3120
gacttctgcg gcaagggata ccacctgatg tcattccctc agtctgctcc ccacggtgtg 3180
gtgttcctgc acgtcaccta cgtgccagcc caggagaaga acttcaccac tgcccctgct 3240
atctgccacg acggcaaggc tcacttcccc agggaaggtg tcttcgtgag caacggcacc 3300
cactggttcg tcactcagag aaacttctac gagccacaga tcatcaccac tgacaacact 3360
ttcgtgtctg gaaactgcga cgtggtcatc ggtatcgtca acaacaccgt gtacgacccc 3420
ctgcagccag agctggactc attcaaggag gaactggaca agtacttcaa gaaccacacc 3480
tcccctgacg tcgacctggg cgacatctca ggaatcaacg cttccgtcgt gaacatccag 3540
aaggagatcg accgcctgaa cgaagtggcc aagaacctga acgaaagcct gatcgacctg 3600
caggagctgg gcaagtacga acagtacatc aagtggcctc accaccacca ccaccac 3657
<210> 61
<211> 1205
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 61
Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn
1 5 10 15
Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser
20 25 30
Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val
35 40 45
Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg
50 55 60
Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser
65 70 75 80
Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu
85 90 95
Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val
100 105 110
Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly
115 120 125
Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg
130 135 140
Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro
145 150 155 160
Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg
165 170 175
Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys
180 185 190
His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala
195 200 205
Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe
210 215 220
Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser
225 230 235 240
Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu
245 250 255
Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr
260 265 270
Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr
275 280 285
Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe
290 295 300
Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn
305 310 315 320
Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val
325 330 335
Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser
340 345 350
Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val
355 360 365
Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp
370 375 380
Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln
385 390 395 400
Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr
405 410 415
Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly
420 425 430
Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys
435 440 445
Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr
450 455 460
Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser
465 470 475 480
Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val
485 490 495
Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly
500 505 510
Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn
515 520 525
Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys
530 535 540
Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp
545 550 555 560
Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys
565 570 575
Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn
580 585 590
Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val
595 600 605
Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr
610 615 620
Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu
625 630 635 640
His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile
645 650 655
Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln Ser
660 665 670
Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu
675 680 685
Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe
690 695 700
Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr
705 710 715 720
Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser
725 730 735
Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala
740 745 750
Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe
755 760 765
Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly
770 775 780
Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys
785 790 795 800
Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp
805 810 815
Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala
820 825 830
Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro
835 840 845
Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu
850 855 860
Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu
865 870 875 880
Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly
885 890 895
Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln
900 905 910
Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala
915 920 925
Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala
930 935 940
Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser
945 950 955 960
Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala Glu
965 970 975
Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr
980 985 990
Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala
995 1000 1005
Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser
1010 1015 1020
Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe
1025 1030 1035
Pro Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr
1040 1045 1050
Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys
1055 1060 1065
His Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser
1070 1075 1080
Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro
1085 1090 1095
Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp
1100 1105 1110
Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln
1115 1120 1125
Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys
1130 1135 1140
Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile
1145 1150 1155
Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn
1160 1165 1170
Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu
1175 1180 1185
Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro His His His His
1190 1195 1200
His His
1205
<210> 62
<211> 1263
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 62
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val
1 5 10 15
Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe
20 25 30
Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu
35 40 45
His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp
50 55 60
Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp
65 70 75 80
Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu
85 90 95
Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser
100 105 110
Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile
115 120 125
Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr
130 135 140
Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
145 150 155 160
Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
165 170 175
Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
180 185 190
Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
195 200 205
Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
210 215 220
Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
225 230 235 240
Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
245 250 255
Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
260 265 270
Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
275 280 285
Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
290 295 300
Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
305 310 315 320
Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
325 330 335
Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
340 345 350
Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
355 360 365
Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
370 375 380
Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
385 390 395 400
Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
405 410 415
Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
420 425 430
Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
435 440 445
Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
450 455 460
Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
465 470 475 480
Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
485 490 495
Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
500 505 510
Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
515 520 525
Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
530 535 540
Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
545 550 555 560
Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val
565 570 575
Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
580 585 590
Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
595 600 605
Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile
610 615 620
His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
625 630 635 640
Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
645 650 655
Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
660 665 670
Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln Ser Val Ala
675 680 685
Leu Gly Ala Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile
690 695 700
Pro Thr Asn Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser
705 710 715 720
Met Thr Lys Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser
725 730 735
Thr Glu Cys Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln
740 745 750
Leu Asn Arg Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr
755 760 765
Gln Glu Val Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile
770 775 780
Lys Asp Phe Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser
785 790 795 800
Lys Pro Ser Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val
805 810 815
Thr Leu Ala Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly
820 825 830
Asp Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu
835 840 845
Thr Val Leu Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr
850 855 860
Ser Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala
865 870 875 880
Gly Ala Ala Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe
885 890 895
Asn Gly Ile Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu
900 905 910
Ile Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu
915 920 925
Ser Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln
930 935 940
Asn Ala Gln Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe
945 950 955 960
Gly Ala Ile Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys
965 970 975
Val Glu Ala Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln
980 985 990
Ser Leu Gln Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile
995 1000 1005
Arg Ala Ser Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val
1010 1015 1020
Leu Gly Gln Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His
1025 1030 1035
Leu Met Ser Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu
1040 1045 1050
His Val Thr Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala
1055 1060 1065
Pro Ala Ile Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly
1070 1075 1080
Val Phe Val Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn
1085 1090 1095
Phe Tyr Glu Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser
1100 1105 1110
Gly Asn Cys Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr
1115 1120 1125
Asp Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp
1130 1135 1140
Lys Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp
1145 1150 1155
Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile
1160 1165 1170
Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile
1175 1180 1185
Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro
1190 1195 1200
Trp Tyr Ile Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val
1205 1210 1215
Met Val Thr Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys
1220 1225 1230
Leu Lys Gly Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu
1235 1240 1245
Asp Asp Ser Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr
1250 1255 1260
<210> 63
<211> 1250
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 63
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn
290 295 300
Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr
305 310 315 320
Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser
325 330 335
Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr
340 345 350
Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly
355 360 365
Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala
370 375 380
Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly
385 390 395 400
Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe
405 410 415
Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val
420 425 430
Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu
435 440 445
Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser
450 455 460
Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln
465 470 475 480
Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg
485 490 495
Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys
500 505 510
Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe
515 520 525
Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys
530 535 540
Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr
545 550 555 560
Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro
565 570 575
Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser
580 585 590
Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro
595 600 605
Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser
610 615 620
Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala
625 630 635 640
Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly
645 650 655
Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln
660 665 670
Ser Val Ala Leu Gly Ala Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser
675 680 685
Ile Ala Ile Pro Thr Asn Phe Thr Ile Ser Val Thr Thr Glu Ile Leu
690 695 700
Pro Val Ser Met Thr Lys Thr Ser Val Asp Cys Thr Met Tyr Ile Cys
705 710 715 720
Gly Asp Ser Thr Glu Cys Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe
725 730 735
Cys Thr Gln Leu Asn Arg Ala Leu Thr Gly Ile Ala Val Glu Gln Asp
740 745 750
Lys Asn Thr Gln Glu Val Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr
755 760 765
Pro Pro Ile Lys Asp Phe Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro
770 775 780
Asp Pro Ser Lys Pro Ser Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe
785 790 795 800
Asn Lys Val Thr Leu Ala Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp
805 810 815
Cys Leu Gly Asp Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe
820 825 830
Asn Gly Leu Thr Val Leu Pro Pro Leu Leu Thr Asp Glu Met Ile Ala
835 840 845
Gln Tyr Thr Ser Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr
850 855 860
Phe Gly Ala Gly Ala Ala Leu Gln Ile Pro Phe Ala Met Gln Met Ala
865 870 875 880
Tyr Arg Phe Asn Gly Ile Gly Val Thr Gln Asn Val Leu Tyr Glu Asn
885 890 895
Gln Lys Leu Ile Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln
900 905 910
Asp Ser Leu Ser Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln Asp Val
915 920 925
Val Asn Gln Asn Ala Gln Ala Leu Asn Thr Leu Val Lys Gln Leu Ser
930 935 940
Ser Asn Phe Gly Ala Ile Ser Ser Val Leu Asn Asp Ile Leu Ser Arg
945 950 955 960
Leu Asp Lys Val Glu Ala Glu Val Gln Ile Asp Arg Leu Ile Thr Gly
965 970 975
Arg Leu Gln Ser Leu Gln Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala
980 985 990
Ala Glu Ile Arg Ala Ser Ala Asn Leu Ala Ala Thr Lys Met Ser Glu
995 1000 1005
Cys Val Leu Gly Gln Ser Lys Arg Val Asp Phe Cys Gly Lys Gly
1010 1015 1020
Tyr His Leu Met Ser Phe Pro Gln Ser Ala Pro His Gly Val Val
1025 1030 1035
Phe Leu His Val Thr Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr
1040 1045 1050
Thr Ala Pro Ala Ile Cys His Asp Gly Lys Ala His Phe Pro Arg
1055 1060 1065
Glu Gly Val Phe Val Ser Asn Gly Thr His Trp Phe Val Thr Gln
1070 1075 1080
Arg Asn Phe Tyr Glu Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe
1085 1090 1095
Val Ser Gly Asn Cys Asp Val Val Ile Gly Ile Val Asn Asn Thr
1100 1105 1110
Val Tyr Asp Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu
1115 1120 1125
Leu Asp Lys Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp Leu
1130 1135 1140
Gly Asp Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln Lys
1145 1150 1155
Glu Ile Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu Ser
1160 1165 1170
Leu Ile Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys
1175 1180 1185
Trp Pro Trp Tyr Ile Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala
1190 1195 1200
Ile Val Met Val Thr Ile Met Leu Cys Cys Met Thr Ser Cys Cys
1205 1210 1215
Ser Cys Leu Lys Gly Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe
1220 1225 1230
Asp Glu Asp Asp Ser Glu Pro Val Leu Lys Gly Val Lys Leu His
1235 1240 1245
Tyr Thr
1250
<210> 64
<211> 1263
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 64
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val
1 5 10 15
Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe
20 25 30
Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu
35 40 45
His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp
50 55 60
Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp
65 70 75 80
Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu
85 90 95
Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser
100 105 110
Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile
115 120 125
Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr
130 135 140
Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
145 150 155 160
Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
165 170 175
Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
180 185 190
Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
195 200 205
Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
210 215 220
Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
225 230 235 240
Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
245 250 255
Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
260 265 270
Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
275 280 285
Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
290 295 300
Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
305 310 315 320
Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
325 330 335
Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
340 345 350
Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
355 360 365
Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
370 375 380
Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
385 390 395 400
Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
405 410 415
Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
420 425 430
Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
435 440 445
Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
450 455 460
Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
465 470 475 480
Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
485 490 495
Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
500 505 510
Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
515 520 525
Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
530 535 540
Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
545 550 555 560
Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val
565 570 575
Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
580 585 590
Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
595 600 605
Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile
610 615 620
His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
625 630 635 640
Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
645 650 655
Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
660 665 670
Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln Ser Val Ala
675 680 685
Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser
690 695 700
Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Glu Ile Leu Pro Val Ser
705 710 715 720
Met Thr Lys Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser
725 730 735
Thr Glu Cys Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln
740 745 750
Leu Asn Arg Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr
755 760 765
Gln Glu Val Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile
770 775 780
Lys Asp Phe Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser
785 790 795 800
Lys Pro Ser Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val
805 810 815
Thr Leu Ala Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly
820 825 830
Asp Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu
835 840 845
Thr Val Leu Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr
850 855 860
Ser Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala
865 870 875 880
Gly Ala Ala Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe
885 890 895
Asn Gly Ile Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu
900 905 910
Ile Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu
915 920 925
Ser Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln
930 935 940
Asn Ala Gln Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe
945 950 955 960
Gly Ala Ile Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys
965 970 975
Val Glu Ala Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln
980 985 990
Ser Leu Gln Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile
995 1000 1005
Arg Ala Ser Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val
1010 1015 1020
Leu Gly Gln Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His
1025 1030 1035
Leu Met Ser Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu
1040 1045 1050
His Val Thr Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala
1055 1060 1065
Pro Ala Ile Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly
1070 1075 1080
Val Phe Val Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn
1085 1090 1095
Phe Tyr Glu Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser
1100 1105 1110
Gly Asn Cys Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr
1115 1120 1125
Asp Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp
1130 1135 1140
Lys Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp
1145 1150 1155
Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile
1160 1165 1170
Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile
1175 1180 1185
Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro
1190 1195 1200
Trp Tyr Ile Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val
1205 1210 1215
Met Val Thr Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys
1220 1225 1230
Leu Lys Gly Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu
1235 1240 1245
Asp Asp Ser Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr
1250 1255 1260
<210> 65
<211> 1250
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 65
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn
290 295 300
Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr
305 310 315 320
Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser
325 330 335
Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr
340 345 350
Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly
355 360 365
Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala
370 375 380
Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly
385 390 395 400
Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe
405 410 415
Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val
420 425 430
Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu
435 440 445
Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser
450 455 460
Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln
465 470 475 480
Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg
485 490 495
Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys
500 505 510
Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe
515 520 525
Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys
530 535 540
Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr
545 550 555 560
Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro
565 570 575
Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser
580 585 590
Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro
595 600 605
Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser
610 615 620
Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala
625 630 635 640
Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly
645 650 655
Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln
660 665 670
Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala
675 680 685
Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Glu Ile Leu
690 695 700
Pro Val Ser Met Thr Lys Thr Ser Val Asp Cys Thr Met Tyr Ile Cys
705 710 715 720
Gly Asp Ser Thr Glu Cys Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe
725 730 735
Cys Thr Gln Leu Asn Arg Ala Leu Thr Gly Ile Ala Val Glu Gln Asp
740 745 750
Lys Asn Thr Gln Glu Val Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr
755 760 765
Pro Pro Ile Lys Asp Phe Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro
770 775 780
Asp Pro Ser Lys Pro Ser Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe
785 790 795 800
Asn Lys Val Thr Leu Ala Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp
805 810 815
Cys Leu Gly Asp Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe
820 825 830
Asn Gly Leu Thr Val Leu Pro Pro Leu Leu Thr Asp Glu Met Ile Ala
835 840 845
Gln Tyr Thr Ser Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr
850 855 860
Phe Gly Ala Gly Ala Ala Leu Gln Ile Pro Phe Ala Met Gln Met Ala
865 870 875 880
Tyr Arg Phe Asn Gly Ile Gly Val Thr Gln Asn Val Leu Tyr Glu Asn
885 890 895
Gln Lys Leu Ile Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln
900 905 910
Asp Ser Leu Ser Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln Asp Val
915 920 925
Val Asn Gln Asn Ala Gln Ala Leu Asn Thr Leu Val Lys Gln Leu Ser
930 935 940
Ser Asn Phe Gly Ala Ile Ser Ser Val Leu Asn Asp Ile Leu Ser Arg
945 950 955 960
Leu Asp Lys Val Glu Ala Glu Val Gln Ile Asp Arg Leu Ile Thr Gly
965 970 975
Arg Leu Gln Ser Leu Gln Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala
980 985 990
Ala Glu Ile Arg Ala Ser Ala Asn Leu Ala Ala Thr Lys Met Ser Glu
995 1000 1005
Cys Val Leu Gly Gln Ser Lys Arg Val Asp Phe Cys Gly Lys Gly
1010 1015 1020
Tyr His Leu Met Ser Phe Pro Gln Ser Ala Pro His Gly Val Val
1025 1030 1035
Phe Leu His Val Thr Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr
1040 1045 1050
Thr Ala Pro Ala Ile Cys His Asp Gly Lys Ala His Phe Pro Arg
1055 1060 1065
Glu Gly Val Phe Val Ser Asn Gly Thr His Trp Phe Val Thr Gln
1070 1075 1080
Arg Asn Phe Tyr Glu Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe
1085 1090 1095
Val Ser Gly Asn Cys Asp Val Val Ile Gly Ile Val Asn Asn Thr
1100 1105 1110
Val Tyr Asp Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu
1115 1120 1125
Leu Asp Lys Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp Leu
1130 1135 1140
Gly Asp Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln Lys
1145 1150 1155
Glu Ile Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu Ser
1160 1165 1170
Leu Ile Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys
1175 1180 1185
Trp Pro Trp Tyr Ile Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala
1190 1195 1200
Ile Val Met Val Thr Ile Met Leu Cys Cys Met Thr Ser Cys Cys
1205 1210 1215
Ser Cys Leu Lys Gly Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe
1220 1225 1230
Asp Glu Asp Asp Ser Glu Pro Val Leu Lys Gly Val Lys Leu His
1235 1240 1245
Tyr Thr
1250
<210> 66
<211> 1263
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 66
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val
1 5 10 15
Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe
20 25 30
Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu
35 40 45
His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp
50 55 60
Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp
65 70 75 80
Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu
85 90 95
Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser
100 105 110
Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile
115 120 125
Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr
130 135 140
Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
145 150 155 160
Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
165 170 175
Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
180 185 190
Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
195 200 205
Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
210 215 220
Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
225 230 235 240
Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
245 250 255
Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
260 265 270
Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
275 280 285
Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
290 295 300
Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
305 310 315 320
Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
325 330 335
Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
340 345 350
Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
355 360 365
Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
370 375 380
Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
385 390 395 400
Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
405 410 415
Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
420 425 430
Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
435 440 445
Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
450 455 460
Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
465 470 475 480
Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
485 490 495
Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
500 505 510
Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
515 520 525
Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
530 535 540
Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
545 550 555 560
Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val
565 570 575
Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
580 585 590
Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
595 600 605
Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile
610 615 620
His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
625 630 635 640
Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
645 650 655
Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
660 665 670
Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln Ser Val Ala
675 680 685
Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser
690 695 700
Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile
705 710 715 720
Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val
725 730 735
Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu
740 745 750
Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr
755 760 765
Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln
770 775 780
Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe
785 790 795 800
Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser
805 810 815
Phe Ile Ala Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly
820 825 830
Asp Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu
835 840 845
Thr Val Leu Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr
850 855 860
Ser Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala
865 870 875 880
Gly Ala Ala Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe
885 890 895
Asn Gly Ile Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu
900 905 910
Ile Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu
915 920 925
Ser Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln
930 935 940
Asn Ala Gln Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe
945 950 955 960
Gly Ala Ile Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys
965 970 975
Val Glu Ala Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln
980 985 990
Ser Leu Gln Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile
995 1000 1005
Arg Ala Ser Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val
1010 1015 1020
Leu Gly Gln Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His
1025 1030 1035
Leu Met Ser Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu
1040 1045 1050
His Val Thr Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala
1055 1060 1065
Pro Ala Ile Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly
1070 1075 1080
Val Phe Val Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn
1085 1090 1095
Phe Tyr Glu Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser
1100 1105 1110
Gly Asn Cys Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr
1115 1120 1125
Asp Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp
1130 1135 1140
Lys Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp
1145 1150 1155
Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile
1160 1165 1170
Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile
1175 1180 1185
Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro
1190 1195 1200
Trp Tyr Ile Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val
1205 1210 1215
Met Val Thr Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys
1220 1225 1230
Leu Lys Gly Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu
1235 1240 1245
Asp Asp Ser Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr
1250 1255 1260
<210> 67
<211> 1250
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 67
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn
290 295 300
Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr
305 310 315 320
Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser
325 330 335
Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr
340 345 350
Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly
355 360 365
Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala
370 375 380
Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly
385 390 395 400
Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe
405 410 415
Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val
420 425 430
Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu
435 440 445
Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser
450 455 460
Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln
465 470 475 480
Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg
485 490 495
Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys
500 505 510
Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe
515 520 525
Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys
530 535 540
Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr
545 550 555 560
Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro
565 570 575
Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser
580 585 590
Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro
595 600 605
Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser
610 615 620
Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala
625 630 635 640
Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly
645 650 655
Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln
660 665 670
Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala
675 680 685
Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn
690 695 700
Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys
705 710 715 720
Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys
725 730 735
Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg
740 745 750
Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val
755 760 765
Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe
770 775 780
Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser
785 790 795 800
Lys Arg Ser Phe Ile Ala Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp
805 810 815
Cys Leu Gly Asp Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe
820 825 830
Asn Gly Leu Thr Val Leu Pro Pro Leu Leu Thr Asp Glu Met Ile Ala
835 840 845
Gln Tyr Thr Ser Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr
850 855 860
Phe Gly Ala Gly Ala Ala Leu Gln Ile Pro Phe Ala Met Gln Met Ala
865 870 875 880
Tyr Arg Phe Asn Gly Ile Gly Val Thr Gln Asn Val Leu Tyr Glu Asn
885 890 895
Gln Lys Leu Ile Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln
900 905 910
Asp Ser Leu Ser Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln Asp Val
915 920 925
Val Asn Gln Asn Ala Gln Ala Leu Asn Thr Leu Val Lys Gln Leu Ser
930 935 940
Ser Asn Phe Gly Ala Ile Ser Ser Val Leu Asn Asp Ile Leu Ser Arg
945 950 955 960
Leu Asp Lys Val Glu Ala Glu Val Gln Ile Asp Arg Leu Ile Thr Gly
965 970 975
Arg Leu Gln Ser Leu Gln Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala
980 985 990
Ala Glu Ile Arg Ala Ser Ala Asn Leu Ala Ala Thr Lys Met Ser Glu
995 1000 1005
Cys Val Leu Gly Gln Ser Lys Arg Val Asp Phe Cys Gly Lys Gly
1010 1015 1020
Tyr His Leu Met Ser Phe Pro Gln Ser Ala Pro His Gly Val Val
1025 1030 1035
Phe Leu His Val Thr Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr
1040 1045 1050
Thr Ala Pro Ala Ile Cys His Asp Gly Lys Ala His Phe Pro Arg
1055 1060 1065
Glu Gly Val Phe Val Ser Asn Gly Thr His Trp Phe Val Thr Gln
1070 1075 1080
Arg Asn Phe Tyr Glu Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe
1085 1090 1095
Val Ser Gly Asn Cys Asp Val Val Ile Gly Ile Val Asn Asn Thr
1100 1105 1110
Val Tyr Asp Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu
1115 1120 1125
Leu Asp Lys Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp Leu
1130 1135 1140
Gly Asp Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln Lys
1145 1150 1155
Glu Ile Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu Ser
1160 1165 1170
Leu Ile Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys
1175 1180 1185
Trp Pro Trp Tyr Ile Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala
1190 1195 1200
Ile Val Met Val Thr Ile Met Leu Cys Cys Met Thr Ser Cys Cys
1205 1210 1215
Ser Cys Leu Lys Gly Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe
1220 1225 1230
Asp Glu Asp Asp Ser Glu Pro Val Leu Lys Gly Val Lys Leu His
1235 1240 1245
Tyr Thr
1250
<210> 68
<211> 31
<212> PRT
<213> 人工序列
<220>
<223> 折叠子
<400> 68
Ser Ala Ile Gly Gly Tyr Ile Pro Glu Ala Pro Arg Asp Gly Gln Ala
1 5 10 15
Tyr Val Arg Lys Asp Gly Glu Trp Val Leu Leu Ser Thr Phe Leu
20 25 30
<210> 69
<211> 202
<212> PRT
<213> β冠状病毒严重急性呼吸系统综合征冠状病毒2
<400> 69
Arg Phe Pro Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn
1 5 10 15
Ala Thr Arg Phe Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser
20 25 30
Asn Cys Val Ala Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser
35 40 45
Thr Phe Lys Cys Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys
50 55 60
Phe Thr Asn Val Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val
65 70 75 80
Arg Gln Ile Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr
85 90 95
Lys Leu Pro Asp Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn
100 105 110
Asn Leu Asp Ser Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu
115 120 125
Phe Arg Lys Ser Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu
130 135 140
Ile Tyr Gln Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn
145 150 155 160
Cys Tyr Phe Pro Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val
165 170 175
Gly Tyr Gln Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His
180 185 190
Ala Pro Ala Thr Val Cys Gly Pro Lys Lys
195 200
<210> 70
<211> 201
<212> PRT
<213> β冠状病毒严重急性呼吸系统综合征相关冠状病毒
<400> 70
Arg Phe Pro Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn
1 5 10 15
Ala Thr Lys Phe Pro Ser Val Tyr Ala Trp Glu Arg Lys Lys Ile Ser
20 25 30
Asn Cys Val Ala Asp Tyr Ser Val Leu Tyr Asn Ser Thr Phe Phe Ser
35 40 45
Thr Phe Lys Cys Tyr Gly Val Ser Ala Thr Lys Leu Asn Asp Leu Cys
50 55 60
Phe Ser Asn Val Tyr Ala Asp Ser Phe Val Val Lys Gly Asp Asp Val
65 70 75 80
Arg Gln Ile Ala Pro Gly Gln Thr Gly Val Ile Ala Asp Tyr Asn Tyr
85 90 95
Lys Leu Pro Asp Asp Phe Met Gly Cys Val Leu Ala Trp Asn Thr Arg
100 105 110
Asn Ile Asp Ala Thr Ser Thr Gly Asn Tyr Asn Tyr Lys Tyr Arg Tyr
115 120 125
Leu Arg His Gly Lys Leu Arg Pro Phe Glu Arg Asp Ile Ser Asn Val
130 135 140
Pro Phe Ser Pro Asp Gly Lys Pro Cys Thr Pro Pro Ala Leu Asn Cys
145 150 155 160
Tyr Trp Pro Leu Asn Asp Tyr Gly Phe Tyr Thr Thr Thr Gly Ile Gly
165 170 175
Tyr Gln Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu Asn Ala
180 185 190
Pro Ala Thr Val Cys Gly Pro Lys Leu
195 200
<210> 71
<211> 239
<212> PRT
<213> β冠状病毒中东呼吸系统综合征相关冠状病毒
<400> 71
Phe Glu Ala Lys Pro Ser Gly Ser Val Val Ala Glu Gly Val Glu Cys
1 5 10 15
Asp Phe Ser Pro Leu Leu Ser Gly Thr Pro Pro Gln Val Tyr Asn Phe
20 25 30
Lys Arg Leu Val Phe Thr Asn Cys Asn Tyr Asn Leu Thr Lys Leu Leu
35 40 45
Ser Leu Phe Ser Val Asn Asp Phe Thr Cys Ser Gln Ile Ser Pro Ala
50 55 60
Ala Ile Ala Ser Asn Cys Tyr Ser Ser Leu Ile Leu Asp Tyr Phe Ser
65 70 75 80
Tyr Pro Leu Ser Met Lys Ser Asp Leu Ser Val Ser Ser Ala Gly Pro
85 90 95
Ile Ser Gln Phe Asn Tyr Lys Gln Ser Phe Ser Asn Pro Thr Cys Leu
100 105 110
Ile Leu Ala Thr Val Pro His Asn Leu Thr Thr Ile Thr Lys Pro Leu
115 120 125
Lys Tyr Ser Tyr Ile Asn Lys Cys Ser Arg Leu Leu Ser Asp Asp Arg
130 135 140
Thr Glu Val Pro Gln Leu Val Asn Ala Asn Gln Tyr Ser Pro Cys Val
145 150 155 160
Ser Ile Val Pro Ser Thr Val Trp Glu Asp Gly Asp Tyr Tyr Arg Lys
165 170 175
Gln Leu Ser Pro Leu Glu Gly Gly Gly Trp Leu Val Ala Ser Gly Ser
180 185 190
Thr Val Ala Met Thr Glu Gln Leu Gln Met Gly Phe Gly Ile Thr Val
195 200 205
Gln Tyr Gly Thr Asp Thr Asn Ser Val Cys Pro Lys Leu Glu Phe Ala
210 215 220
Asn Asp Thr Lys Ile Ala Ser Gln Leu Gly Asn Cys Val Glu Tyr
225 230 235
<210> 72
<211> 544
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 72
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Arg Phe Pro
1 5 10 15
Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg
20 25 30
Phe Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val
35 40 45
Ala Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys
50 55 60
Cys Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn
65 70 75 80
Val Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile
85 90 95
Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro
100 105 110
Asp Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp
115 120 125
Ser Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys
130 135 140
Ser Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln
145 150 155 160
Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe
165 170 175
Pro Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln
180 185 190
Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala
195 200 205
Thr Val Cys Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys
210 215 220
Val Asn Phe Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu
225 230 235 240
Ser Asn Lys Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala
245 250 255
Asp Thr Thr Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp
260 265 270
Ile Thr Pro Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr
275 280 285
Asn Thr Ser Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr
290 295 300
Glu Arg Phe Pro Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe
305 310 315 320
Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile
325 330 335
Ser Asn Cys Val Ala Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe
340 345 350
Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu
355 360 365
Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu
370 375 380
Val Arg Gln Ile Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn
385 390 395 400
Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser
405 410 415
Asn Asn Leu Asp Ser Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg
420 425 430
Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr
435 440 445
Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe
450 455 460
Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly
465 470 475 480
Val Gly Tyr Gln Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu
485 490 495
His Ala Pro Ala Thr Val Cys Gly Pro Lys Lys Ser Ala Ile Gly Gly
500 505 510
Tyr Ile Pro Glu Ala Pro Arg Asp Gly Gln Ala Tyr Val Arg Lys Asp
515 520 525
Gly Glu Trp Val Leu Leu Ser Thr Phe Leu His His His His His His
530 535 540
<210> 73
<211> 531
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 73
Arg Phe Pro Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn
1 5 10 15
Ala Thr Arg Phe Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser
20 25 30
Asn Cys Val Ala Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser
35 40 45
Thr Phe Lys Cys Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys
50 55 60
Phe Thr Asn Val Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val
65 70 75 80
Arg Gln Ile Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr
85 90 95
Lys Leu Pro Asp Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn
100 105 110
Asn Leu Asp Ser Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu
115 120 125
Phe Arg Lys Ser Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu
130 135 140
Ile Tyr Gln Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn
145 150 155 160
Cys Tyr Phe Pro Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val
165 170 175
Gly Tyr Gln Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His
180 185 190
Ala Pro Ala Thr Val Cys Gly Pro Lys Lys Ser Thr Asn Leu Val Lys
195 200 205
Asn Lys Cys Val Asn Phe Asn Phe Asn Gly Leu Thr Gly Thr Gly Val
210 215 220
Leu Thr Glu Ser Asn Lys Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg
225 230 235 240
Asp Ile Ala Asp Thr Thr Asp Ala Val Arg Asp Pro Gln Thr Leu Glu
245 250 255
Ile Leu Asp Ile Thr Pro Cys Ser Phe Gly Gly Val Ser Val Ile Thr
260 265 270
Pro Gly Thr Asn Thr Ser Asn Gln Val Ala Val Leu Tyr Gln Asp Val
275 280 285
Asn Cys Thr Glu Arg Phe Pro Asn Ile Thr Asn Leu Cys Pro Phe Gly
290 295 300
Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala Trp Asn Arg
305 310 315 320
Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu Tyr Asn Ser
325 330 335
Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro Thr Lys Leu
340 345 350
Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe Val Ile Arg
355 360 365
Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly Lys Ile Ala
370 375 380
Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys Val Ile Ala
385 390 395 400
Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn Tyr Asn Tyr
405 410 415
Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe Glu Arg Asp
420 425 430
Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys Asn Gly Val
435 440 445
Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly Phe Gln Pro
450 455 460
Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val Leu Ser Phe
465 470 475 480
Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys Lys Ser Ala
485 490 495
Ile Gly Gly Tyr Ile Pro Glu Ala Pro Arg Asp Gly Gln Ala Tyr Val
500 505 510
Arg Lys Asp Gly Glu Trp Val Leu Leu Ser Thr Phe Leu His His His
515 520 525
His His His
530
<210> 74
<211> 544
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 74
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Arg Phe Pro
1 5 10 15
Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg
20 25 30
Phe Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val
35 40 45
Ala Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys
50 55 60
Cys Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn
65 70 75 80
Val Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile
85 90 95
Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro
100 105 110
Asp Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp
115 120 125
Ser Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys
130 135 140
Ser Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln
145 150 155 160
Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe
165 170 175
Pro Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln
180 185 190
Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala
195 200 205
Thr Val Cys Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys
210 215 220
Val Asn Phe Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu
225 230 235 240
Ser Asn Lys Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala
245 250 255
Asp Thr Thr Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp
260 265 270
Ile Thr Pro Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr
275 280 285
Asn Thr Ser Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr
290 295 300
Glu Arg Phe Pro Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe
305 310 315 320
Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile
325 330 335
Ser Asn Cys Val Ala Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe
340 345 350
Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu
355 360 365
Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu
370 375 380
Val Arg Gln Ile Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn
385 390 395 400
Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser
405 410 415
Asn Asn Leu Asp Ser Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg
420 425 430
Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr
435 440 445
Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe
450 455 460
Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly
465 470 475 480
Val Gly Tyr Gln Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu
485 490 495
His Ala Pro Ala Thr Val Cys Gly Pro Lys Lys Ser Ala Ile Gly Gly
500 505 510
Tyr Ile Pro Glu Ala Pro Arg Asp Gly Gln Ala Tyr Val Arg Lys Asp
515 520 525
Gly Glu Trp Val Leu Leu Ser Thr Phe Leu His His His His His His
530 535 540
<210> 75
<211> 531
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 75
Arg Phe Pro Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn
1 5 10 15
Ala Thr Arg Phe Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser
20 25 30
Asn Cys Val Ala Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser
35 40 45
Thr Phe Lys Cys Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys
50 55 60
Phe Thr Asn Val Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val
65 70 75 80
Arg Gln Ile Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr
85 90 95
Lys Leu Pro Asp Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn
100 105 110
Asn Leu Asp Ser Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu
115 120 125
Phe Arg Lys Ser Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu
130 135 140
Ile Tyr Gln Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn
145 150 155 160
Cys Tyr Phe Pro Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val
165 170 175
Gly Tyr Gln Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His
180 185 190
Ala Pro Ala Thr Val Cys Gly Pro Lys Lys Ser Thr Asn Leu Val Lys
195 200 205
Asn Lys Cys Val Asn Phe Asn Phe Asn Gly Leu Thr Gly Thr Gly Val
210 215 220
Leu Thr Glu Ser Asn Lys Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg
225 230 235 240
Asp Ile Ala Asp Thr Thr Asp Ala Val Arg Asp Pro Gln Thr Leu Glu
245 250 255
Ile Leu Asp Ile Thr Pro Cys Ser Phe Gly Gly Val Ser Val Ile Thr
260 265 270
Pro Gly Thr Asn Thr Ser Asn Gln Val Ala Val Leu Tyr Gln Asp Val
275 280 285
Asn Cys Thr Glu Arg Phe Pro Asn Ile Thr Asn Leu Cys Pro Phe Gly
290 295 300
Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala Trp Asn Arg
305 310 315 320
Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu Tyr Asn Ser
325 330 335
Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro Thr Lys Leu
340 345 350
Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe Val Ile Arg
355 360 365
Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly Lys Ile Ala
370 375 380
Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys Val Ile Ala
385 390 395 400
Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn Tyr Asn Tyr
405 410 415
Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe Glu Arg Asp
420 425 430
Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys Asn Gly Val
435 440 445
Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly Phe Gln Pro
450 455 460
Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val Leu Ser Phe
465 470 475 480
Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys Lys Ser Ala
485 490 495
Ile Gly Gly Tyr Ile Pro Glu Ala Pro Arg Asp Gly Gln Ala Tyr Val
500 505 510
Arg Lys Asp Gly Glu Trp Val Leu Leu Ser Thr Phe Leu His His His
515 520 525
His His His
530
<210> 76
<211> 470
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 76
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Arg Phe Pro
1 5 10 15
Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg
20 25 30
Phe Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val
35 40 45
Ala Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys
50 55 60
Cys Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn
65 70 75 80
Val Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile
85 90 95
Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro
100 105 110
Asp Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp
115 120 125
Ser Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys
130 135 140
Ser Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln
145 150 155 160
Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe
165 170 175
Pro Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln
180 185 190
Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala
195 200 205
Thr Val Cys Gly Pro Lys Lys Ser Gly Gly Gly Ser Gly Gly Gly Ser
210 215 220
Gly Gly Gly Ser Gly Gly Gly Ser Arg Phe Pro Asn Ile Thr Asn Leu
225 230 235 240
Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Lys Phe Pro Ser Val Tyr
245 250 255
Ala Trp Glu Arg Lys Lys Ile Ser Asn Cys Val Ala Asp Tyr Ser Val
260 265 270
Leu Tyr Asn Ser Thr Phe Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser
275 280 285
Ala Thr Lys Leu Asn Asp Leu Cys Phe Ser Asn Val Tyr Ala Asp Ser
290 295 300
Phe Val Val Lys Gly Asp Asp Val Arg Gln Ile Ala Pro Gly Gln Thr
305 310 315 320
Gly Val Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Met Gly
325 330 335
Cys Val Leu Ala Trp Asn Thr Arg Asn Ile Asp Ala Thr Ser Thr Gly
340 345 350
Asn Tyr Asn Tyr Lys Tyr Arg Tyr Leu Arg His Gly Lys Leu Arg Pro
355 360 365
Phe Glu Arg Asp Ile Ser Asn Val Pro Phe Ser Pro Asp Gly Lys Pro
370 375 380
Cys Thr Pro Pro Ala Leu Asn Cys Tyr Trp Pro Leu Asn Asp Tyr Gly
385 390 395 400
Phe Tyr Thr Thr Thr Gly Ile Gly Tyr Gln Pro Tyr Arg Val Val Val
405 410 415
Leu Ser Phe Glu Leu Leu Asn Ala Pro Ala Thr Val Cys Gly Pro Lys
420 425 430
Leu Ser Ala Ile Gly Gly Tyr Ile Pro Glu Ala Pro Arg Asp Gly Gln
435 440 445
Ala Tyr Val Arg Lys Asp Gly Glu Trp Val Leu Leu Ser Thr Phe Leu
450 455 460
His His His His His His
465 470
<210> 77
<211> 464
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 77
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Arg Phe Pro
1 5 10 15
Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg
20 25 30
Phe Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val
35 40 45
Ala Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys
50 55 60
Cys Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn
65 70 75 80
Val Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile
85 90 95
Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro
100 105 110
Asp Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp
115 120 125
Ser Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys
130 135 140
Ser Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln
145 150 155 160
Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe
165 170 175
Pro Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln
180 185 190
Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala
195 200 205
Thr Val Cys Gly Pro Lys Lys Ser Gly Gly Gly Ser Gly Gly Gly Ser
210 215 220
Gly Gly Gly Ser Gly Gly Gly Ser Arg Phe Pro Asn Ile Thr Asn Leu
225 230 235 240
Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Lys Phe Pro Ser Val Tyr
245 250 255
Ala Trp Glu Arg Lys Lys Ile Ser Asn Cys Val Ala Asp Tyr Ser Val
260 265 270
Leu Tyr Asn Ser Thr Phe Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser
275 280 285
Ala Thr Lys Leu Asn Asp Leu Cys Phe Ser Asn Val Tyr Ala Asp Ser
290 295 300
Phe Val Val Lys Gly Asp Asp Val Arg Gln Ile Ala Pro Gly Gln Thr
305 310 315 320
Gly Val Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Met Gly
325 330 335
Cys Val Leu Ala Trp Asn Thr Arg Asn Ile Asp Ala Thr Ser Thr Gly
340 345 350
Asn Tyr Asn Tyr Lys Tyr Arg Tyr Leu Arg His Gly Lys Leu Arg Pro
355 360 365
Phe Glu Arg Asp Ile Ser Asn Val Pro Phe Ser Pro Asp Gly Lys Pro
370 375 380
Cys Thr Pro Pro Ala Leu Asn Cys Tyr Trp Pro Leu Asn Asp Tyr Gly
385 390 395 400
Phe Tyr Thr Thr Thr Gly Ile Gly Tyr Gln Pro Tyr Arg Val Val Val
405 410 415
Leu Ser Phe Glu Leu Leu Asn Ala Pro Ala Thr Val Cys Gly Pro Lys
420 425 430
Leu Ser Ala Ile Gly Gly Tyr Ile Pro Glu Ala Pro Arg Asp Gly Gln
435 440 445
Ala Tyr Val Arg Lys Asp Gly Glu Trp Val Leu Leu Ser Thr Phe Leu
450 455 460
<210> 78
<211> 457
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 78
Arg Phe Pro Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn
1 5 10 15
Ala Thr Arg Phe Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser
20 25 30
Asn Cys Val Ala Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser
35 40 45
Thr Phe Lys Cys Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys
50 55 60
Phe Thr Asn Val Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val
65 70 75 80
Arg Gln Ile Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr
85 90 95
Lys Leu Pro Asp Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn
100 105 110
Asn Leu Asp Ser Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu
115 120 125
Phe Arg Lys Ser Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu
130 135 140
Ile Tyr Gln Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn
145 150 155 160
Cys Tyr Phe Pro Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val
165 170 175
Gly Tyr Gln Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His
180 185 190
Ala Pro Ala Thr Val Cys Gly Pro Lys Lys Ser Gly Gly Gly Ser Gly
195 200 205
Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Arg Phe Pro Asn Ile
210 215 220
Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Lys Phe Pro
225 230 235 240
Ser Val Tyr Ala Trp Glu Arg Lys Lys Ile Ser Asn Cys Val Ala Asp
245 250 255
Tyr Ser Val Leu Tyr Asn Ser Thr Phe Phe Ser Thr Phe Lys Cys Tyr
260 265 270
Gly Val Ser Ala Thr Lys Leu Asn Asp Leu Cys Phe Ser Asn Val Tyr
275 280 285
Ala Asp Ser Phe Val Val Lys Gly Asp Asp Val Arg Gln Ile Ala Pro
290 295 300
Gly Gln Thr Gly Val Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp
305 310 315 320
Phe Met Gly Cys Val Leu Ala Trp Asn Thr Arg Asn Ile Asp Ala Thr
325 330 335
Ser Thr Gly Asn Tyr Asn Tyr Lys Tyr Arg Tyr Leu Arg His Gly Lys
340 345 350
Leu Arg Pro Phe Glu Arg Asp Ile Ser Asn Val Pro Phe Ser Pro Asp
355 360 365
Gly Lys Pro Cys Thr Pro Pro Ala Leu Asn Cys Tyr Trp Pro Leu Asn
370 375 380
Asp Tyr Gly Phe Tyr Thr Thr Thr Gly Ile Gly Tyr Gln Pro Tyr Arg
385 390 395 400
Val Val Val Leu Ser Phe Glu Leu Leu Asn Ala Pro Ala Thr Val Cys
405 410 415
Gly Pro Lys Leu Ser Ala Ile Gly Gly Tyr Ile Pro Glu Ala Pro Arg
420 425 430
Asp Gly Gln Ala Tyr Val Arg Lys Asp Gly Glu Trp Val Leu Leu Ser
435 440 445
Thr Phe Leu His His His His His His
450 455
<210> 79
<211> 451
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 79
Arg Phe Pro Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn
1 5 10 15
Ala Thr Arg Phe Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser
20 25 30
Asn Cys Val Ala Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser
35 40 45
Thr Phe Lys Cys Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys
50 55 60
Phe Thr Asn Val Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val
65 70 75 80
Arg Gln Ile Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr
85 90 95
Lys Leu Pro Asp Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn
100 105 110
Asn Leu Asp Ser Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu
115 120 125
Phe Arg Lys Ser Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu
130 135 140
Ile Tyr Gln Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn
145 150 155 160
Cys Tyr Phe Pro Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val
165 170 175
Gly Tyr Gln Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His
180 185 190
Ala Pro Ala Thr Val Cys Gly Pro Lys Lys Ser Gly Gly Gly Ser Gly
195 200 205
Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Arg Phe Pro Asn Ile
210 215 220
Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Lys Phe Pro
225 230 235 240
Ser Val Tyr Ala Trp Glu Arg Lys Lys Ile Ser Asn Cys Val Ala Asp
245 250 255
Tyr Ser Val Leu Tyr Asn Ser Thr Phe Phe Ser Thr Phe Lys Cys Tyr
260 265 270
Gly Val Ser Ala Thr Lys Leu Asn Asp Leu Cys Phe Ser Asn Val Tyr
275 280 285
Ala Asp Ser Phe Val Val Lys Gly Asp Asp Val Arg Gln Ile Ala Pro
290 295 300
Gly Gln Thr Gly Val Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp
305 310 315 320
Phe Met Gly Cys Val Leu Ala Trp Asn Thr Arg Asn Ile Asp Ala Thr
325 330 335
Ser Thr Gly Asn Tyr Asn Tyr Lys Tyr Arg Tyr Leu Arg His Gly Lys
340 345 350
Leu Arg Pro Phe Glu Arg Asp Ile Ser Asn Val Pro Phe Ser Pro Asp
355 360 365
Gly Lys Pro Cys Thr Pro Pro Ala Leu Asn Cys Tyr Trp Pro Leu Asn
370 375 380
Asp Tyr Gly Phe Tyr Thr Thr Thr Gly Ile Gly Tyr Gln Pro Tyr Arg
385 390 395 400
Val Val Val Leu Ser Phe Glu Leu Leu Asn Ala Pro Ala Thr Val Cys
405 410 415
Gly Pro Lys Leu Ser Ala Ile Gly Gly Tyr Ile Pro Glu Ala Pro Arg
420 425 430
Asp Gly Gln Ala Tyr Val Arg Lys Asp Gly Glu Trp Val Leu Leu Ser
435 440 445
Thr Phe Leu
450
<210> 80
<211> 726
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 80
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Arg Phe Pro
1 5 10 15
Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg
20 25 30
Phe Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val
35 40 45
Ala Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys
50 55 60
Cys Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn
65 70 75 80
Val Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile
85 90 95
Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro
100 105 110
Asp Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp
115 120 125
Ser Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys
130 135 140
Ser Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln
145 150 155 160
Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe
165 170 175
Pro Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln
180 185 190
Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala
195 200 205
Thr Val Cys Gly Pro Lys Lys Ser Gly Gly Gly Ser Gly Gly Gly Ser
210 215 220
Gly Gly Gly Ser Gly Gly Gly Ser Phe Glu Ala Lys Pro Ser Gly Ser
225 230 235 240
Val Val Ala Glu Gly Val Glu Cys Asp Phe Ser Pro Leu Leu Ser Gly
245 250 255
Thr Pro Pro Gln Val Tyr Asn Phe Lys Arg Leu Val Phe Thr Asn Cys
260 265 270
Asn Tyr Asn Leu Thr Lys Leu Leu Ser Leu Phe Ser Val Asn Asp Phe
275 280 285
Thr Cys Ser Gln Ile Ser Pro Ala Ala Ile Ala Ser Asn Cys Tyr Ser
290 295 300
Ser Leu Ile Leu Asp Tyr Phe Ser Tyr Pro Leu Ser Met Lys Ser Asp
305 310 315 320
Leu Ser Val Ser Ser Ala Gly Pro Ile Ser Gln Phe Asn Tyr Lys Gln
325 330 335
Ser Phe Ser Asn Pro Thr Cys Leu Ile Leu Ala Thr Val Pro His Asn
340 345 350
Leu Thr Thr Ile Thr Lys Pro Leu Lys Tyr Ser Tyr Ile Asn Lys Cys
355 360 365
Ser Arg Leu Leu Ser Asp Asp Arg Thr Glu Val Pro Gln Leu Val Asn
370 375 380
Ala Asn Gln Tyr Ser Pro Cys Val Ser Ile Val Pro Ser Thr Val Trp
385 390 395 400
Glu Asp Gly Asp Tyr Tyr Arg Lys Gln Leu Ser Pro Leu Glu Gly Gly
405 410 415
Gly Trp Leu Val Ala Ser Gly Ser Thr Val Ala Met Thr Glu Gln Leu
420 425 430
Gln Met Gly Phe Gly Ile Thr Val Gln Tyr Gly Thr Asp Thr Asn Ser
435 440 445
Val Cys Pro Lys Leu Glu Phe Ala Asn Asp Thr Lys Ile Ala Ser Gln
450 455 460
Leu Gly Asn Cys Val Glu Tyr Ser Gly Gly Gly Ser Gly Gly Gly Ser
465 470 475 480
Gly Gly Gly Ser Gly Gly Gly Ser Arg Phe Pro Asn Ile Thr Asn Leu
485 490 495
Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Lys Phe Pro Ser Val Tyr
500 505 510
Ala Trp Glu Arg Lys Lys Ile Ser Asn Cys Val Ala Asp Tyr Ser Val
515 520 525
Leu Tyr Asn Ser Thr Phe Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser
530 535 540
Ala Thr Lys Leu Asn Asp Leu Cys Phe Ser Asn Val Tyr Ala Asp Ser
545 550 555 560
Phe Val Val Lys Gly Asp Asp Val Arg Gln Ile Ala Pro Gly Gln Thr
565 570 575
Gly Val Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Met Gly
580 585 590
Cys Val Leu Ala Trp Asn Thr Arg Asn Ile Asp Ala Thr Ser Thr Gly
595 600 605
Asn Tyr Asn Tyr Lys Tyr Arg Tyr Leu Arg His Gly Lys Leu Arg Pro
610 615 620
Phe Glu Arg Asp Ile Ser Asn Val Pro Phe Ser Pro Asp Gly Lys Pro
625 630 635 640
Cys Thr Pro Pro Ala Leu Asn Cys Tyr Trp Pro Leu Asn Asp Tyr Gly
645 650 655
Phe Tyr Thr Thr Thr Gly Ile Gly Tyr Gln Pro Tyr Arg Val Val Val
660 665 670
Leu Ser Phe Glu Leu Leu Asn Ala Pro Ala Thr Val Cys Gly Pro Lys
675 680 685
Leu Ser Ala Ile Gly Gly Tyr Ile Pro Glu Ala Pro Arg Asp Gly Gln
690 695 700
Ala Tyr Val Arg Lys Asp Gly Glu Trp Val Leu Leu Ser Thr Phe Leu
705 710 715 720
His His His His His His
725
<210> 81
<211> 720
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 81
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Arg Phe Pro
1 5 10 15
Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg
20 25 30
Phe Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val
35 40 45
Ala Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys
50 55 60
Cys Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn
65 70 75 80
Val Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile
85 90 95
Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro
100 105 110
Asp Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp
115 120 125
Ser Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys
130 135 140
Ser Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln
145 150 155 160
Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe
165 170 175
Pro Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln
180 185 190
Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala
195 200 205
Thr Val Cys Gly Pro Lys Lys Ser Gly Gly Gly Ser Gly Gly Gly Ser
210 215 220
Gly Gly Gly Ser Gly Gly Gly Ser Phe Glu Ala Lys Pro Ser Gly Ser
225 230 235 240
Val Val Ala Glu Gly Val Glu Cys Asp Phe Ser Pro Leu Leu Ser Gly
245 250 255
Thr Pro Pro Gln Val Tyr Asn Phe Lys Arg Leu Val Phe Thr Asn Cys
260 265 270
Asn Tyr Asn Leu Thr Lys Leu Leu Ser Leu Phe Ser Val Asn Asp Phe
275 280 285
Thr Cys Ser Gln Ile Ser Pro Ala Ala Ile Ala Ser Asn Cys Tyr Ser
290 295 300
Ser Leu Ile Leu Asp Tyr Phe Ser Tyr Pro Leu Ser Met Lys Ser Asp
305 310 315 320
Leu Ser Val Ser Ser Ala Gly Pro Ile Ser Gln Phe Asn Tyr Lys Gln
325 330 335
Ser Phe Ser Asn Pro Thr Cys Leu Ile Leu Ala Thr Val Pro His Asn
340 345 350
Leu Thr Thr Ile Thr Lys Pro Leu Lys Tyr Ser Tyr Ile Asn Lys Cys
355 360 365
Ser Arg Leu Leu Ser Asp Asp Arg Thr Glu Val Pro Gln Leu Val Asn
370 375 380
Ala Asn Gln Tyr Ser Pro Cys Val Ser Ile Val Pro Ser Thr Val Trp
385 390 395 400
Glu Asp Gly Asp Tyr Tyr Arg Lys Gln Leu Ser Pro Leu Glu Gly Gly
405 410 415
Gly Trp Leu Val Ala Ser Gly Ser Thr Val Ala Met Thr Glu Gln Leu
420 425 430
Gln Met Gly Phe Gly Ile Thr Val Gln Tyr Gly Thr Asp Thr Asn Ser
435 440 445
Val Cys Pro Lys Leu Glu Phe Ala Asn Asp Thr Lys Ile Ala Ser Gln
450 455 460
Leu Gly Asn Cys Val Glu Tyr Ser Gly Gly Gly Ser Gly Gly Gly Ser
465 470 475 480
Gly Gly Gly Ser Gly Gly Gly Ser Arg Phe Pro Asn Ile Thr Asn Leu
485 490 495
Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Lys Phe Pro Ser Val Tyr
500 505 510
Ala Trp Glu Arg Lys Lys Ile Ser Asn Cys Val Ala Asp Tyr Ser Val
515 520 525
Leu Tyr Asn Ser Thr Phe Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser
530 535 540
Ala Thr Lys Leu Asn Asp Leu Cys Phe Ser Asn Val Tyr Ala Asp Ser
545 550 555 560
Phe Val Val Lys Gly Asp Asp Val Arg Gln Ile Ala Pro Gly Gln Thr
565 570 575
Gly Val Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Met Gly
580 585 590
Cys Val Leu Ala Trp Asn Thr Arg Asn Ile Asp Ala Thr Ser Thr Gly
595 600 605
Asn Tyr Asn Tyr Lys Tyr Arg Tyr Leu Arg His Gly Lys Leu Arg Pro
610 615 620
Phe Glu Arg Asp Ile Ser Asn Val Pro Phe Ser Pro Asp Gly Lys Pro
625 630 635 640
Cys Thr Pro Pro Ala Leu Asn Cys Tyr Trp Pro Leu Asn Asp Tyr Gly
645 650 655
Phe Tyr Thr Thr Thr Gly Ile Gly Tyr Gln Pro Tyr Arg Val Val Val
660 665 670
Leu Ser Phe Glu Leu Leu Asn Ala Pro Ala Thr Val Cys Gly Pro Lys
675 680 685
Leu Ser Ala Ile Gly Gly Tyr Ile Pro Glu Ala Pro Arg Asp Gly Gln
690 695 700
Ala Tyr Val Arg Lys Asp Gly Glu Trp Val Leu Leu Ser Thr Phe Leu
705 710 715 720
<210> 82
<211> 713
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 82
Arg Phe Pro Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn
1 5 10 15
Ala Thr Arg Phe Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser
20 25 30
Asn Cys Val Ala Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser
35 40 45
Thr Phe Lys Cys Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys
50 55 60
Phe Thr Asn Val Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val
65 70 75 80
Arg Gln Ile Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr
85 90 95
Lys Leu Pro Asp Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn
100 105 110
Asn Leu Asp Ser Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu
115 120 125
Phe Arg Lys Ser Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu
130 135 140
Ile Tyr Gln Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn
145 150 155 160
Cys Tyr Phe Pro Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val
165 170 175
Gly Tyr Gln Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His
180 185 190
Ala Pro Ala Thr Val Cys Gly Pro Lys Lys Ser Gly Gly Gly Ser Gly
195 200 205
Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Phe Glu Ala Lys Pro
210 215 220
Ser Gly Ser Val Val Ala Glu Gly Val Glu Cys Asp Phe Ser Pro Leu
225 230 235 240
Leu Ser Gly Thr Pro Pro Gln Val Tyr Asn Phe Lys Arg Leu Val Phe
245 250 255
Thr Asn Cys Asn Tyr Asn Leu Thr Lys Leu Leu Ser Leu Phe Ser Val
260 265 270
Asn Asp Phe Thr Cys Ser Gln Ile Ser Pro Ala Ala Ile Ala Ser Asn
275 280 285
Cys Tyr Ser Ser Leu Ile Leu Asp Tyr Phe Ser Tyr Pro Leu Ser Met
290 295 300
Lys Ser Asp Leu Ser Val Ser Ser Ala Gly Pro Ile Ser Gln Phe Asn
305 310 315 320
Tyr Lys Gln Ser Phe Ser Asn Pro Thr Cys Leu Ile Leu Ala Thr Val
325 330 335
Pro His Asn Leu Thr Thr Ile Thr Lys Pro Leu Lys Tyr Ser Tyr Ile
340 345 350
Asn Lys Cys Ser Arg Leu Leu Ser Asp Asp Arg Thr Glu Val Pro Gln
355 360 365
Leu Val Asn Ala Asn Gln Tyr Ser Pro Cys Val Ser Ile Val Pro Ser
370 375 380
Thr Val Trp Glu Asp Gly Asp Tyr Tyr Arg Lys Gln Leu Ser Pro Leu
385 390 395 400
Glu Gly Gly Gly Trp Leu Val Ala Ser Gly Ser Thr Val Ala Met Thr
405 410 415
Glu Gln Leu Gln Met Gly Phe Gly Ile Thr Val Gln Tyr Gly Thr Asp
420 425 430
Thr Asn Ser Val Cys Pro Lys Leu Glu Phe Ala Asn Asp Thr Lys Ile
435 440 445
Ala Ser Gln Leu Gly Asn Cys Val Glu Tyr Ser Gly Gly Gly Ser Gly
450 455 460
Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Arg Phe Pro Asn Ile
465 470 475 480
Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Lys Phe Pro
485 490 495
Ser Val Tyr Ala Trp Glu Arg Lys Lys Ile Ser Asn Cys Val Ala Asp
500 505 510
Tyr Ser Val Leu Tyr Asn Ser Thr Phe Phe Ser Thr Phe Lys Cys Tyr
515 520 525
Gly Val Ser Ala Thr Lys Leu Asn Asp Leu Cys Phe Ser Asn Val Tyr
530 535 540
Ala Asp Ser Phe Val Val Lys Gly Asp Asp Val Arg Gln Ile Ala Pro
545 550 555 560
Gly Gln Thr Gly Val Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp
565 570 575
Phe Met Gly Cys Val Leu Ala Trp Asn Thr Arg Asn Ile Asp Ala Thr
580 585 590
Ser Thr Gly Asn Tyr Asn Tyr Lys Tyr Arg Tyr Leu Arg His Gly Lys
595 600 605
Leu Arg Pro Phe Glu Arg Asp Ile Ser Asn Val Pro Phe Ser Pro Asp
610 615 620
Gly Lys Pro Cys Thr Pro Pro Ala Leu Asn Cys Tyr Trp Pro Leu Asn
625 630 635 640
Asp Tyr Gly Phe Tyr Thr Thr Thr Gly Ile Gly Tyr Gln Pro Tyr Arg
645 650 655
Val Val Val Leu Ser Phe Glu Leu Leu Asn Ala Pro Ala Thr Val Cys
660 665 670
Gly Pro Lys Leu Ser Ala Ile Gly Gly Tyr Ile Pro Glu Ala Pro Arg
675 680 685
Asp Gly Gln Ala Tyr Val Arg Lys Asp Gly Glu Trp Val Leu Leu Ser
690 695 700
Thr Phe Leu His His His His His His
705 710
<210> 83
<211> 707
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 83
Arg Phe Pro Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn
1 5 10 15
Ala Thr Arg Phe Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser
20 25 30
Asn Cys Val Ala Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser
35 40 45
Thr Phe Lys Cys Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys
50 55 60
Phe Thr Asn Val Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val
65 70 75 80
Arg Gln Ile Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr
85 90 95
Lys Leu Pro Asp Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn
100 105 110
Asn Leu Asp Ser Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu
115 120 125
Phe Arg Lys Ser Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu
130 135 140
Ile Tyr Gln Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn
145 150 155 160
Cys Tyr Phe Pro Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val
165 170 175
Gly Tyr Gln Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His
180 185 190
Ala Pro Ala Thr Val Cys Gly Pro Lys Lys Ser Gly Gly Gly Ser Gly
195 200 205
Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Phe Glu Ala Lys Pro
210 215 220
Ser Gly Ser Val Val Ala Glu Gly Val Glu Cys Asp Phe Ser Pro Leu
225 230 235 240
Leu Ser Gly Thr Pro Pro Gln Val Tyr Asn Phe Lys Arg Leu Val Phe
245 250 255
Thr Asn Cys Asn Tyr Asn Leu Thr Lys Leu Leu Ser Leu Phe Ser Val
260 265 270
Asn Asp Phe Thr Cys Ser Gln Ile Ser Pro Ala Ala Ile Ala Ser Asn
275 280 285
Cys Tyr Ser Ser Leu Ile Leu Asp Tyr Phe Ser Tyr Pro Leu Ser Met
290 295 300
Lys Ser Asp Leu Ser Val Ser Ser Ala Gly Pro Ile Ser Gln Phe Asn
305 310 315 320
Tyr Lys Gln Ser Phe Ser Asn Pro Thr Cys Leu Ile Leu Ala Thr Val
325 330 335
Pro His Asn Leu Thr Thr Ile Thr Lys Pro Leu Lys Tyr Ser Tyr Ile
340 345 350
Asn Lys Cys Ser Arg Leu Leu Ser Asp Asp Arg Thr Glu Val Pro Gln
355 360 365
Leu Val Asn Ala Asn Gln Tyr Ser Pro Cys Val Ser Ile Val Pro Ser
370 375 380
Thr Val Trp Glu Asp Gly Asp Tyr Tyr Arg Lys Gln Leu Ser Pro Leu
385 390 395 400
Glu Gly Gly Gly Trp Leu Val Ala Ser Gly Ser Thr Val Ala Met Thr
405 410 415
Glu Gln Leu Gln Met Gly Phe Gly Ile Thr Val Gln Tyr Gly Thr Asp
420 425 430
Thr Asn Ser Val Cys Pro Lys Leu Glu Phe Ala Asn Asp Thr Lys Ile
435 440 445
Ala Ser Gln Leu Gly Asn Cys Val Glu Tyr Ser Gly Gly Gly Ser Gly
450 455 460
Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Arg Phe Pro Asn Ile
465 470 475 480
Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Lys Phe Pro
485 490 495
Ser Val Tyr Ala Trp Glu Arg Lys Lys Ile Ser Asn Cys Val Ala Asp
500 505 510
Tyr Ser Val Leu Tyr Asn Ser Thr Phe Phe Ser Thr Phe Lys Cys Tyr
515 520 525
Gly Val Ser Ala Thr Lys Leu Asn Asp Leu Cys Phe Ser Asn Val Tyr
530 535 540
Ala Asp Ser Phe Val Val Lys Gly Asp Asp Val Arg Gln Ile Ala Pro
545 550 555 560
Gly Gln Thr Gly Val Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp
565 570 575
Phe Met Gly Cys Val Leu Ala Trp Asn Thr Arg Asn Ile Asp Ala Thr
580 585 590
Ser Thr Gly Asn Tyr Asn Tyr Lys Tyr Arg Tyr Leu Arg His Gly Lys
595 600 605
Leu Arg Pro Phe Glu Arg Asp Ile Ser Asn Val Pro Phe Ser Pro Asp
610 615 620
Gly Lys Pro Cys Thr Pro Pro Ala Leu Asn Cys Tyr Trp Pro Leu Asn
625 630 635 640
Asp Tyr Gly Phe Tyr Thr Thr Thr Gly Ile Gly Tyr Gln Pro Tyr Arg
645 650 655
Val Val Val Leu Ser Phe Glu Leu Leu Asn Ala Pro Ala Thr Val Cys
660 665 670
Gly Pro Lys Leu Ser Ala Ile Gly Gly Tyr Ile Pro Glu Ala Pro Arg
675 680 685
Asp Gly Gln Ala Tyr Val Arg Lys Asp Gly Glu Trp Val Leu Leu Ser
690 695 700
Thr Phe Leu
705
<210> 84
<211> 1273
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 84
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val
1 5 10 15
Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe
20 25 30
Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu
35 40 45
His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp
50 55 60
Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp
65 70 75 80
Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu
85 90 95
Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser
100 105 110
Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile
115 120 125
Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr
130 135 140
Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
145 150 155 160
Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
165 170 175
Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
180 185 190
Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
195 200 205
Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
210 215 220
Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
225 230 235 240
Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
245 250 255
Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
260 265 270
Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
275 280 285
Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
290 295 300
Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
305 310 315 320
Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
325 330 335
Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
340 345 350
Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
355 360 365
Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
370 375 380
Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
385 390 395 400
Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
405 410 415
Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
420 425 430
Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
435 440 445
Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
450 455 460
Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
465 470 475 480
Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
485 490 495
Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
500 505 510
Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
515 520 525
Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
530 535 540
Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
545 550 555 560
Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val
565 570 575
Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
580 585 590
Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
595 600 605
Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile
610 615 620
His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
625 630 635 640
Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
645 650 655
Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
660 665 670
Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala Arg Ser Val Ala
675 680 685
Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser
690 695 700
Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile
705 710 715 720
Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val
725 730 735
Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu
740 745 750
Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr
755 760 765
Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln
770 775 780
Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe
785 790 795 800
Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser
805 810 815
Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly
820 825 830
Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp
835 840 845
Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu
850 855 860
Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly
865 870 875 880
Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln Ile
885 890 895
Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr
900 905 910
Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn
915 920 925
Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala
930 935 940
Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn
945 950 955 960
Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val
965 970 975
Leu Asn Asp Ile Leu Ser Arg Leu Asp Pro Pro Glu Ala Glu Val Gln
980 985 990
Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val
995 1000 1005
Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn
1010 1015 1020
Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys
1025 1030 1035
Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro
1040 1045 1050
Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val
1055 1060 1065
Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His
1070 1075 1080
Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn
1085 1090 1095
Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln
1100 1105 1110
Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val
1115 1120 1125
Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro
1130 1135 1140
Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn
1145 1150 1155
His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn
1160 1165 1170
Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu
1175 1180 1185
Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu
1190 1195 1200
Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile Trp Leu
1205 1210 1215
Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr Ile Met
1220 1225 1230
Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly Cys Cys
1235 1240 1245
Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser Glu Pro
1250 1255 1260
Val Leu Lys Gly Val Lys Leu His Tyr Thr
1265 1270
<210> 85
<211> 1260
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 85
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn
290 295 300
Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr
305 310 315 320
Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser
325 330 335
Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr
340 345 350
Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly
355 360 365
Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala
370 375 380
Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly
385 390 395 400
Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe
405 410 415
Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val
420 425 430
Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu
435 440 445
Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser
450 455 460
Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln
465 470 475 480
Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg
485 490 495
Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys
500 505 510
Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe
515 520 525
Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys
530 535 540
Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr
545 550 555 560
Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro
565 570 575
Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser
580 585 590
Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro
595 600 605
Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser
610 615 620
Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala
625 630 635 640
Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly
645 650 655
Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala Arg
660 665 670
Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala
675 680 685
Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn
690 695 700
Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys
705 710 715 720
Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys
725 730 735
Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg
740 745 750
Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val
755 760 765
Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe
770 775 780
Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser
785 790 795 800
Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala
805 810 815
Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala
820 825 830
Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu
835 840 845
Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu
850 855 860
Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala
865 870 875 880
Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile
885 890 895
Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn
900 905 910
Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr
915 920 925
Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln
930 935 940
Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile
945 950 955 960
Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Pro Pro Glu Ala
965 970 975
Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln
980 985 990
Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser
995 1000 1005
Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln
1010 1015 1020
Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser
1025 1030 1035
Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr
1040 1045 1050
Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile
1055 1060 1065
Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val
1070 1075 1080
Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu
1085 1090 1095
Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys
1100 1105 1110
Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu
1115 1120 1125
Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe
1130 1135 1140
Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly
1145 1150 1155
Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu
1160 1165 1170
Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln
1175 1180 1185
Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile
1190 1195 1200
Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr
1205 1210 1215
Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly
1220 1225 1230
Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser
1235 1240 1245
Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr
1250 1255 1260
<210> 86
<211> 1273
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 86
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val
1 5 10 15
Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe
20 25 30
Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu
35 40 45
His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp
50 55 60
Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp
65 70 75 80
Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu
85 90 95
Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser
100 105 110
Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile
115 120 125
Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr
130 135 140
Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
145 150 155 160
Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
165 170 175
Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
180 185 190
Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
195 200 205
Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
210 215 220
Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
225 230 235 240
Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
245 250 255
Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
260 265 270
Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
275 280 285
Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
290 295 300
Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
305 310 315 320
Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
325 330 335
Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
340 345 350
Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
355 360 365
Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
370 375 380
Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
385 390 395 400
Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
405 410 415
Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
420 425 430
Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
435 440 445
Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
450 455 460
Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
465 470 475 480
Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
485 490 495
Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
500 505 510
Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
515 520 525
Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
530 535 540
Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
545 550 555 560
Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val
565 570 575
Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
580 585 590
Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
595 600 605
Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile
610 615 620
His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
625 630 635 640
Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
645 650 655
Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
660 665 670
Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln Ser Val Ala
675 680 685
Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser
690 695 700
Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile
705 710 715 720
Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val
725 730 735
Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu
740 745 750
Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr
755 760 765
Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln
770 775 780
Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe
785 790 795 800
Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser
805 810 815
Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly
820 825 830
Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp
835 840 845
Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu
850 855 860
Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly
865 870 875 880
Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln Ile
885 890 895
Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr
900 905 910
Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn
915 920 925
Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala
930 935 940
Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn
945 950 955 960
Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val
965 970 975
Leu Asn Asp Ile Leu Ser Arg Leu Asp Pro Pro Glu Ala Glu Val Gln
980 985 990
Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val
995 1000 1005
Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn
1010 1015 1020
Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys
1025 1030 1035
Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro
1040 1045 1050
Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val
1055 1060 1065
Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His
1070 1075 1080
Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn
1085 1090 1095
Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln
1100 1105 1110
Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val
1115 1120 1125
Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro
1130 1135 1140
Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn
1145 1150 1155
His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn
1160 1165 1170
Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu
1175 1180 1185
Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu
1190 1195 1200
Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile Trp Leu
1205 1210 1215
Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr Ile Met
1220 1225 1230
Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly Cys Cys
1235 1240 1245
Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser Glu Pro
1250 1255 1260
Val Leu Lys Gly Val Lys Leu His Tyr Thr
1265 1270
<210> 87
<211> 1260
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 87
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn
290 295 300
Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr
305 310 315 320
Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser
325 330 335
Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr
340 345 350
Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly
355 360 365
Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala
370 375 380
Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly
385 390 395 400
Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe
405 410 415
Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val
420 425 430
Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu
435 440 445
Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser
450 455 460
Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln
465 470 475 480
Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg
485 490 495
Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys
500 505 510
Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe
515 520 525
Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys
530 535 540
Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr
545 550 555 560
Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro
565 570 575
Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser
580 585 590
Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro
595 600 605
Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser
610 615 620
Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala
625 630 635 640
Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly
645 650 655
Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln
660 665 670
Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala
675 680 685
Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn
690 695 700
Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys
705 710 715 720
Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys
725 730 735
Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg
740 745 750
Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val
755 760 765
Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe
770 775 780
Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser
785 790 795 800
Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala
805 810 815
Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala
820 825 830
Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu
835 840 845
Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu
850 855 860
Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala
865 870 875 880
Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile
885 890 895
Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn
900 905 910
Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr
915 920 925
Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln
930 935 940
Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile
945 950 955 960
Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Pro Pro Glu Ala
965 970 975
Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln
980 985 990
Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser
995 1000 1005
Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln
1010 1015 1020
Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser
1025 1030 1035
Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr
1040 1045 1050
Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile
1055 1060 1065
Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val
1070 1075 1080
Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu
1085 1090 1095
Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys
1100 1105 1110
Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu
1115 1120 1125
Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe
1130 1135 1140
Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly
1145 1150 1155
Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu
1160 1165 1170
Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln
1175 1180 1185
Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile
1190 1195 1200
Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr
1205 1210 1215
Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly
1220 1225 1230
Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser
1235 1240 1245
Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr
1250 1255 1260
<210> 88
<211> 1219
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 88
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val
1 5 10 15
Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe
20 25 30
Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu
35 40 45
His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp
50 55 60
Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp
65 70 75 80
Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu
85 90 95
Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser
100 105 110
Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile
115 120 125
Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr
130 135 140
Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
145 150 155 160
Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
165 170 175
Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
180 185 190
Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
195 200 205
Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
210 215 220
Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
225 230 235 240
Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
245 250 255
Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
260 265 270
Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
275 280 285
Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
290 295 300
Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
305 310 315 320
Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
325 330 335
Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
340 345 350
Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
355 360 365
Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
370 375 380
Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
385 390 395 400
Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
405 410 415
Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
420 425 430
Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
435 440 445
Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
450 455 460
Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
465 470 475 480
Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
485 490 495
Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
500 505 510
Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
515 520 525
Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
530 535 540
Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
545 550 555 560
Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val
565 570 575
Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
580 585 590
Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
595 600 605
Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile
610 615 620
His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
625 630 635 640
Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
645 650 655
Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
660 665 670
Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln Ser Val Ala
675 680 685
Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser
690 695 700
Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile
705 710 715 720
Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val
725 730 735
Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu
740 745 750
Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr
755 760 765
Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln
770 775 780
Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe
785 790 795 800
Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser
805 810 815
Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly
820 825 830
Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp
835 840 845
Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu
850 855 860
Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly
865 870 875 880
Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln Ile
885 890 895
Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr
900 905 910
Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn
915 920 925
Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala
930 935 940
Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn
945 950 955 960
Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val
965 970 975
Leu Asn Asp Ile Leu Ser Arg Leu Asp Pro Pro Glu Ala Glu Val Gln
980 985 990
Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val
995 1000 1005
Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn
1010 1015 1020
Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys
1025 1030 1035
Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro
1040 1045 1050
Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val
1055 1060 1065
Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His
1070 1075 1080
Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn
1085 1090 1095
Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln
1100 1105 1110
Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val
1115 1120 1125
Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro
1130 1135 1140
Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn
1145 1150 1155
His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn
1160 1165 1170
Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu
1175 1180 1185
Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu
1190 1195 1200
Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro His His His His His
1205 1210 1215
His
<210> 89
<211> 1206
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 89
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn
290 295 300
Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr
305 310 315 320
Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser
325 330 335
Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr
340 345 350
Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly
355 360 365
Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala
370 375 380
Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly
385 390 395 400
Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe
405 410 415
Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val
420 425 430
Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu
435 440 445
Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser
450 455 460
Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln
465 470 475 480
Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg
485 490 495
Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys
500 505 510
Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe
515 520 525
Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys
530 535 540
Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr
545 550 555 560
Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro
565 570 575
Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser
580 585 590
Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro
595 600 605
Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser
610 615 620
Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala
625 630 635 640
Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly
645 650 655
Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln
660 665 670
Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala
675 680 685
Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn
690 695 700
Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys
705 710 715 720
Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys
725 730 735
Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg
740 745 750
Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val
755 760 765
Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe
770 775 780
Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser
785 790 795 800
Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala
805 810 815
Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala
820 825 830
Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu
835 840 845
Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu
850 855 860
Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala
865 870 875 880
Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile
885 890 895
Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn
900 905 910
Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr
915 920 925
Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln
930 935 940
Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile
945 950 955 960
Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Pro Pro Glu Ala
965 970 975
Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln
980 985 990
Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser
995 1000 1005
Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln
1010 1015 1020
Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser
1025 1030 1035
Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr
1040 1045 1050
Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile
1055 1060 1065
Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val
1070 1075 1080
Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu
1085 1090 1095
Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys
1100 1105 1110
Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu
1115 1120 1125
Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe
1130 1135 1140
Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly
1145 1150 1155
Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu
1160 1165 1170
Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln
1175 1180 1185
Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro His His His
1190 1195 1200
His His His
1205
<210> 90
<211> 17
<212> PRT
<213> 人工序列
<220>
<223> 接头
<400> 90
Ser Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly
1 5 10 15
Ser
<210> 91
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 接头
<400> 91
Ser Gly Gly Gly
1
<210> 92
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 接头
<400> 92
Gly Gly Gly Ser
1
<210> 93
<211> 5
<212> PRT
<213> 人工序列
<220>
<223> 接头
<400> 93
Gly Gly Gly Gly Ser
1 5
<210> 94
<211> 90
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 94
Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn Gly
1 5 10 15
Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu Pro
20 25 30
Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val Arg
35 40 45
Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe Gly
50 55 60
Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val Ala
65 70 75 80
Val Leu Tyr Gln Asp Val Asn Cys Thr Glu
85 90
<210> 95
<211> 3639
<212> DNA
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 95
atgttcgtct tcctggtgct gctgcccctg gtgtccagcc agtgcgtgaa cctgaccact 60
aggactcagc tgcctcccgc ttacaccaac tcattcactc gcggtgtgta ctaccctgac 120
aaggtcttcc gttcttcagt gctgcactca actcaggacc tgttcctgcc cttcttctcc 180
aacgtcacct ggttccacgc catccacgtg tccggcacca acggcactaa gcgcttcgac 240
aacccagtgc tgcctttcaa cgacggtgtc tacttcgctt caaccgagaa gtccaacatc 300
atccgtggat ggatcttcgg caccactctg gacagcaaga ctcagtctct gctgatcgtc 360
aacaacgcca ccaacgtggt catcaaggtc tgcgaattcc agttctgcaa cgacccattc 420
ctgggcgtct actaccacaa gaacaacaag tcatggatgg agtccgaatt ccgcgtctac 480
tccagcgcta acaactgcac tttcgagtac gtgtcccagc ctttcctgat ggacctggaa 540
ggaaagcagg gtaacttcaa gaacctgagg gagttcgtgt tcaagaacat cgacggatac 600
ttcaagattt acagcaagca caccccaatc aacctggtgc gcgacctgcc tcagggtttc 660
tctgctctgg agccactggt ggacctgcct atcggcatca acatcacccg cttccagact 720
ctgctggctc tgcaccgttc ctacctgact ccaggcgact catcttctgg atggactgct 780
ggagctgctg cttactacgt gggctacctg cagcctcgca ccttcctgct gaagtacaac 840
gaaaacggaa ccatcactga cgccgtcgac tgcgctctgg accctctgtc agaaaccaag 900
tgcactctga agtccttcac cgtggagaag ggcatctacc agacttcaaa cttcagggtg 960
cagcccaccg aatccatcgt cagattccct aacatcacta acctgtgccc cttcggagag 1020
gtcttcaacg ccacccgctt cgcttccgtg tacgcctgga acaggaagag aatctcaaac 1080
tgcgtcgctg actactccgt gctgtacaac tcagcctcct tcagcacctt caagtgctac 1140
ggcgtgtcac caactaagct gaacgacctg tgcttcacca acgtctacgc cgactccttc 1200
gtgatcaggg gagacgaggt cagacagatc gctcctggcc agactggaaa gatcgccgac 1260
tacaactaca agctgcccga cgacttcacc ggttgcgtca tcgcttggaa cagcaacaac 1320
ctggactcta aagtgggtgg caactacaac tacctgtacc gcctgttccg taagtcaaac 1380
ctgaagccat tcgagaggga catcagcact gaaatctacc aggccggatc taccccttgc 1440
aacggtgtcg agggcttcaa ctgctacttc cccctgcagt cctacggttt ccagccaact 1500
aacggtgtgg gctaccagcc ttacagagtg gtcgtgctga gcttcgaact gctccacgct 1560
cctgctactg tgtgcggtcc aaagaagtct accaacctgg tcaagaacaa gtgcgtgaac 1620
ttcaacttca acggcctgac cggaactggt gtcctgaccg agagcaacaa gaagttcctg 1680
cccttccagc agttcggaag ggacatcgct gacaccactg acgctgtgcg cgaccctcag 1740
accctggaaa tcctggacat cactccatgc tcattcggag gtgtctccgt gatcacccct 1800
ggcaccaaca cttctaacca ggtcgctgtg ctgtaccagg acgtcaactg caccgaggtc 1860
cctgtggcca tccacgctga ccagctgacc cccacttggc gcgtgtactc caccggcagc 1920
aacgtgttcc agactcgtgc tggttgcctg atcggcgccg agcacgtgaa caacagctac 1980
gaatgcgaca tccccatcgg cgctggaatc tgcgcctctt accagaccca gactaacagc 2040
ccacgcaggg ctcgctctgt ggcctctcag tcaatcatcg cttacaccat gtcactgggc 2100
gctgaaaact ccgtggccta ctctaacaac tcaatcgcca tccccaccaa cttcactatc 2160
agcgtgacca ctgagatcct gccagtcagc atgaccaaga cttctgtgga ctgcactatg 2220
tacatctgcg gagacagcac cgaatgctct aacctgctgc tgcagtacgg ctctttctgc 2280
acccagctga accgtgctct gactggaatc gccgtggagc aggacaagaa cactcaggaa 2340
gtcttcgctc aggtgaagca aatctacaag accccaccta tcaaggactt cggcggattc 2400
aacttctccc agatcctgcc tgacccctcc aagccaagca agcgctcttt catcgaggac 2460
ctgctgttca acaaggtcac tctggccgac gctggattca tcaagcagta cggagactgc 2520
ctgggtgaca tcgccgctcg tgacctgatc tgcgctcaga agttcaacgg tctgaccgtg 2580
ctgcccccac tgctgactga cgaaatgatc gcccagtaca ctagcgccct gctggctgga 2640
accatcactt ctggttggac cttcggtgct ggcgccgctc tgcagatccc tttcgctatg 2700
cagatggcct accgtttcaa cggaatcggt gtcacccaga acgtgctgta cgagaaccag 2760
aagctgatcg ctaaccagtt caactcagcc atcggaaaga tccaggacag cctgagctct 2820
actgcctctg ctctgggcaa gctgcaggac gtcgtgaacc agaacgccca ggctctgaac 2880
accctggtca agcagctgtc atccaacttc ggtgctatca gctctgtgct gaacgacatc 2940
ctgtcccgcc tggacaaggt cgaggccgaa gtgcagatcg accgcctgat cactggccgt 3000
ctgcagtcac tgcagaccta cgtgactcag cagctgatca gggccgctga aatcagagcc 3060
tccgctaacc tggccgctac caagatgagc gagtgcgtcc tgggtcaatc taagcgtgtg 3120
gacttctgcg gcaagggata ccacctgatg tcattccctc agtctgctcc ccacggtgtg 3180
gtgttcctgc acgtcaccta cgtgccagcc caggagaaga acttcaccac tgcccctgct 3240
atctgccacg acggcaaggc tcacttcccc agggaaggtg tcttcgtgag caacggcacc 3300
cactggttcg tcactcagag aaacttctac gagccacaga tcatcaccac tgacaacact 3360
ttcgtgtctg gaaactgcga cgtggtcatc ggtatcgtca acaacaccgt gtacgacccc 3420
ctgcagccag agctggactc attcaaggag gaactggaca agtacttcaa gaaccacacc 3480
tcccctgacg tcgacctggg cgacatctca ggaatcaacg cttccgtcgt gaacatccag 3540
aaggagatcg accgcctgaa cgaagtggcc aagaacctga acgaaagcct gatcgacctg 3600
caggagctgg gcaagtacga acagtacatc aagtggcct 3639
<210> 96
<211> 3822
<212> DNA
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 96
atgttcgtct tcctggtgct gctgcccctg gtgtccagcc agtgcgtgaa cctgaccact 60
aggactcagc tgcctcccgc ttacaccaac tcattcactc gcggtgtgta ctaccctgac 120
aaggtcttcc gttcttcagt gctgcactca actcaggacc tgttcctgcc cttcttctcc 180
aacgtcacct ggttccacgc catccacgtg tccggcacca acggcactaa gcgcttcgac 240
aacccagtgc tgcctttcaa cgacggtgtc tacttcgctt caaccgagaa gtccaacatc 300
atccgtggat ggatcttcgg caccactctg gacagcaaga ctcagtctct gctgatcgtc 360
aacaacgcca ccaacgtggt catcaaggtc tgcgaattcc agttctgcaa cgacccattc 420
ctgggcgtct actaccacaa gaacaacaag tcatggatgg agtccgaatt ccgcgtctac 480
tccagcgcta acaactgcac tttcgagtac gtgtcccagc ctttcctgat ggacctggaa 540
ggaaagcagg gtaacttcaa gaacctgagg gagttcgtgt tcaagaacat cgacggatac 600
ttcaagattt acagcaagca caccccaatc aacctggtgc gcgacctgcc tcagggtttc 660
tctgctctgg agccactggt ggacctgcct atcggcatca acatcacccg cttccagact 720
ctgctggctc tgcaccgttc ctacctgact ccaggcgact catcttctgg atggactgct 780
ggagctgctg cttactacgt gggctacctg cagcctcgca ccttcctgct gaagtacaac 840
gaaaacggaa ccatcactga cgccgtcgac tgcgctctgg accctctgtc agaaaccaag 900
tgcactctga agtccttcac cgtggagaag ggcatctacc agacttcaaa cttcagggtg 960
cagcccaccg aatccatcgt cagattccct aacatcacta acctgtgccc cttcggagag 1020
gtcttcaacg ccacccgctt cgcttccgtg tacgcctgga acaggaagag aatctcaaac 1080
tgcgtcgctg actactccgt gctgtacaac tcagcctcct tcagcacctt caagtgctac 1140
ggcgtgtcac caactaagct gaacgacctg tgcttcacca acgtctacgc cgactccttc 1200
gtgatcaggg gagacgaggt cagacagatc gctcctggcc agactggaaa gatcgccgac 1260
tacaactaca agctgcccga cgacttcacc ggttgcgtca tcgcttggaa cagcaacaac 1320
ctggactcta aagtgggtgg caactacaac tacctgtacc gcctgttccg taagtcaaac 1380
ctgaagccat tcgagaggga catcagcact gaaatctacc aggccggatc taccccttgc 1440
aacggtgtcg agggcttcaa ctgctacttc cccctgcagt cctacggttt ccagccaact 1500
aacggtgtgg gctaccagcc ttacagagtg gtcgtgctga gcttcgaact gctccacgct 1560
cctgctactg tgtgcggtcc aaagaagtct accaacctgg tcaagaacaa gtgcgtgaac 1620
ttcaacttca acggcctgac cggaactggt gtcctgaccg agagcaacaa gaagttcctg 1680
cccttccagc agttcggaag ggacatcgct gacaccactg acgctgtgcg cgaccctcag 1740
accctggaaa tcctggacat cactccatgc tcattcggag gtgtctccgt gatcacccct 1800
ggcaccaaca cttctaacca ggtcgctgtg ctgtaccagg acgtcaactg caccgaggtc 1860
cctgtggcca tccacgctga ccagctgacc cccacttggc gcgtgtactc caccggcagc 1920
aacgtgttcc agactcgtgc tggttgcctg atcggcgccg agcacgtgaa caacagctac 1980
gaatgcgaca tccccatcgg cgctggaatc tgcgcctctt accagaccca gactaacagc 2040
ccacagcagg ctcagtctgt ggcctctcag tcaatcatcg cttacaccat gtcactgggc 2100
gctgaaaact ccgtggccta ctctaacaac tcaatcgcca tccccaccaa cttcactatc 2160
agcgtgacca ctgagatcct gccagtcagc atgaccaaga cttctgtgga ctgcactatg 2220
tacatctgcg gagacagcac cgaatgctct aacctgctgc tgcagtacgg ctctttctgc 2280
acccagctga accgtgctct gactggaatc gccgtggagc aggacaagaa cactcaggaa 2340
gtcttcgctc aggtgaagca aatctacaag accccaccta tcaaggactt cggcggattc 2400
aacttctccc agatcctgcc tgacccctcc aagccaagca agcgctcttt catcgaggac 2460
ctgctgttca acaaggtcac tctggccgac gctggattca tcaagcagta cggagactgc 2520
ctgggtgaca tcgccgctcg tgacctgatc tgcgctcaga agttcaacgg tctgaccgtg 2580
ctgcccccac tgctgactga cgaaatgatc gcccagtaca ctagcgccct gctggctgga 2640
accatcactt ctggttggac cttcggtgct ggcgccgctc tgcagatccc tttcgctatg 2700
cagatggcct accgtttcaa cggaatcggt gtcacccaga acgtgctgta cgagaaccag 2760
aagctgatcg ctaaccagtt caactcagcc atcggaaaga tccaggacag cctgagctct 2820
actgcctctg ctctgggcaa gctgcaggac gtcgtgaacc agaacgccca ggctctgaac 2880
accctggtca agcagctgtc atccaacttc ggtgctatca gctctgtgct gaacgacatc 2940
ctgtcccgcc tggaccctcc cgaggccgaa gtgcagatcg accgcctgat cactggccgt 3000
ctgcagtcac tgcagaccta cgtgactcag cagctgatca gggccgctga aatcagagcc 3060
tccgctaacc tggccgctac caagatgagc gagtgcgtcc tgggtcaatc taagcgtgtg 3120
gacttctgcg gcaagggata ccacctgatg tcattccctc agtctgctcc ccacggtgtg 3180
gtgttcctgc acgtcaccta cgtgccagcc caggagaaga acttcaccac tgcccctgct 3240
atctgccacg acggcaaggc tcacttcccc agggaaggtg tcttcgtgag caacggcacc 3300
cactggttcg tcactcagag aaacttctac gagccacaga tcatcaccac tgacaacact 3360
ttcgtgtctg gaaactgcga cgtggtcatc ggtatcgtca acaacaccgt gtacgacccc 3420
ctgcagccag agctggactc attcaaggag gaactggaca agtacttcaa gaaccacacc 3480
tcccctgacg tcgacctggg cgacatctca ggaatcaacg cttccgtcgt gaacatccag 3540
aaggagatcg accgcctgaa cgaagtggcc aagaacctga acgaaagcct gatcgacctg 3600
caggagctgg gcaagtacga acagtacatc aagtggcctt ggtacatctg gctgggtttc 3660
atcgctggcc tcatcgctat cgtgatggtg accatcatgc tgtgctgcat gacttcatgc 3720
tgctcctgcc tgaagggctg ctgcagctgc ggatcttgct gcaagttcga cgaggacgac 3780
tctgaacccg tcctgaaggg cgtgaagctg cactacacct aa 3822
<210> 97
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 97
Gly Ser Ala Ser
1
<210> 98
<211> 8
<212> PRT
<213> 人工序列
<220>
<223> HRV3C切割位点
<400> 98
Leu Glu Val Leu Phe Gln Gly Pro
1 5
<210> 99
<211> 8
<212> PRT
<213> 人工序列
<220>
<223> Strep标签
<400> 99
Trp Ser His Pro Gln Phe Glu Lys
1 5
<210> 100
<211> 6
<212> PRT
<213> 人工序列
<220>
<223> 6X-His标签
<400> 100
His His His His His His
1 5
<210> 101
<211> 10
<212> PRT
<213> 人工序列
<220>
<223> His标签
<220>
<221> 尚未归类的特征
<222> (6)..(6)
<223> 可以存在或不存在
<220>
<221> 尚未归类的特征
<222> (7)..(7)
<223> 可以存在或不存在
<220>
<221> 尚未归类的特征
<222> (8)..(8)
<223> 可以存在或不存在
<220>
<221> 尚未归类的特征
<222> (9)..(9)
<223> 可以存在或不存在
<220>
<221> 尚未归类的特征
<222> (10)..(10)
<223> 可以存在或不存在
<400> 101
His His His His His His His His His His
1 5 10
<210> 102
<211> 8
<212> PRT
<213> 人工序列
<220>
<223> 8X-His标签
<400> 102
His His His His His His His His
1 5
<210> 103
<211> 30
<212> PRT
<213> 人工序列
<220>
<223> 折叠子
<400> 103
Gly Tyr Ile Pro Glu Ala Pro Arg Asp Gly Gln Ala Tyr Val Arg Lys
1 5 10 15
Asp Gly Glu Trp Val Leu Leu Ser Thr Phe Leu Ser Pro Ala
20 25 30
<210> 104
<211> 10
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 104
Glu Asp Leu Leu Phe Asn Lys Val Thr Leu
1 5 10
<210> 105
<211> 1263
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 105
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val
1 5 10 15
Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe
20 25 30
Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu
35 40 45
His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp
50 55 60
Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp
65 70 75 80
Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu
85 90 95
Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser
100 105 110
Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile
115 120 125
Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr
130 135 140
Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
145 150 155 160
Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
165 170 175
Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
180 185 190
Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
195 200 205
Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
210 215 220
Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
225 230 235 240
Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
245 250 255
Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
260 265 270
Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
275 280 285
Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
290 295 300
Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
305 310 315 320
Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
325 330 335
Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
340 345 350
Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
355 360 365
Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
370 375 380
Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
385 390 395 400
Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
405 410 415
Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
420 425 430
Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
435 440 445
Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
450 455 460
Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
465 470 475 480
Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
485 490 495
Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
500 505 510
Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
515 520 525
Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
530 535 540
Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
545 550 555 560
Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val
565 570 575
Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
580 585 590
Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
595 600 605
Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile
610 615 620
His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
625 630 635 640
Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
645 650 655
Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
660 665 670
Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln Ser Val Ala
675 680 685
Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser
690 695 700
Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile
705 710 715 720
Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val
725 730 735
Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu
740 745 750
Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr
755 760 765
Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln
770 775 780
Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe
785 790 795 800
Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser
805 810 815
Phe Ile Ala Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly
820 825 830
Asp Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu
835 840 845
Thr Val Leu Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr
850 855 860
Ser Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala
865 870 875 880
Gly Ala Ala Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe
885 890 895
Asn Gly Ile Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu
900 905 910
Ile Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu
915 920 925
Ser Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln
930 935 940
Asn Ala Gln Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe
945 950 955 960
Gly Ala Ile Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Pro
965 970 975
Pro Glu Ala Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln
980 985 990
Ser Leu Gln Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile
995 1000 1005
Arg Ala Ser Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val
1010 1015 1020
Leu Gly Gln Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His
1025 1030 1035
Leu Met Ser Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu
1040 1045 1050
His Val Thr Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala
1055 1060 1065
Pro Ala Ile Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly
1070 1075 1080
Val Phe Val Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn
1085 1090 1095
Phe Tyr Glu Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser
1100 1105 1110
Gly Asn Cys Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr
1115 1120 1125
Asp Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp
1130 1135 1140
Lys Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp
1145 1150 1155
Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile
1160 1165 1170
Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile
1175 1180 1185
Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro
1190 1195 1200
Trp Tyr Ile Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val
1205 1210 1215
Met Val Thr Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys
1220 1225 1230
Leu Lys Gly Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu
1235 1240 1245
Asp Asp Ser Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr
1250 1255 1260
<210> 106
<211> 1250
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 106
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn
290 295 300
Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr
305 310 315 320
Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser
325 330 335
Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr
340 345 350
Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly
355 360 365
Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala
370 375 380
Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly
385 390 395 400
Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe
405 410 415
Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val
420 425 430
Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu
435 440 445
Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser
450 455 460
Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln
465 470 475 480
Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg
485 490 495
Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys
500 505 510
Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe
515 520 525
Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys
530 535 540
Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr
545 550 555 560
Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro
565 570 575
Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser
580 585 590
Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro
595 600 605
Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser
610 615 620
Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala
625 630 635 640
Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly
645 650 655
Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln
660 665 670
Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala
675 680 685
Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn
690 695 700
Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys
705 710 715 720
Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys
725 730 735
Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg
740 745 750
Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val
755 760 765
Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe
770 775 780
Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser
785 790 795 800
Lys Arg Ser Phe Ile Ala Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp
805 810 815
Cys Leu Gly Asp Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe
820 825 830
Asn Gly Leu Thr Val Leu Pro Pro Leu Leu Thr Asp Glu Met Ile Ala
835 840 845
Gln Tyr Thr Ser Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr
850 855 860
Phe Gly Ala Gly Ala Ala Leu Gln Ile Pro Phe Ala Met Gln Met Ala
865 870 875 880
Tyr Arg Phe Asn Gly Ile Gly Val Thr Gln Asn Val Leu Tyr Glu Asn
885 890 895
Gln Lys Leu Ile Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln
900 905 910
Asp Ser Leu Ser Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln Asp Val
915 920 925
Val Asn Gln Asn Ala Gln Ala Leu Asn Thr Leu Val Lys Gln Leu Ser
930 935 940
Ser Asn Phe Gly Ala Ile Ser Ser Val Leu Asn Asp Ile Leu Ser Arg
945 950 955 960
Leu Asp Pro Pro Glu Ala Glu Val Gln Ile Asp Arg Leu Ile Thr Gly
965 970 975
Arg Leu Gln Ser Leu Gln Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala
980 985 990
Ala Glu Ile Arg Ala Ser Ala Asn Leu Ala Ala Thr Lys Met Ser Glu
995 1000 1005
Cys Val Leu Gly Gln Ser Lys Arg Val Asp Phe Cys Gly Lys Gly
1010 1015 1020
Tyr His Leu Met Ser Phe Pro Gln Ser Ala Pro His Gly Val Val
1025 1030 1035
Phe Leu His Val Thr Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr
1040 1045 1050
Thr Ala Pro Ala Ile Cys His Asp Gly Lys Ala His Phe Pro Arg
1055 1060 1065
Glu Gly Val Phe Val Ser Asn Gly Thr His Trp Phe Val Thr Gln
1070 1075 1080
Arg Asn Phe Tyr Glu Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe
1085 1090 1095
Val Ser Gly Asn Cys Asp Val Val Ile Gly Ile Val Asn Asn Thr
1100 1105 1110
Val Tyr Asp Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu
1115 1120 1125
Leu Asp Lys Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp Leu
1130 1135 1140
Gly Asp Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln Lys
1145 1150 1155
Glu Ile Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu Ser
1160 1165 1170
Leu Ile Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys
1175 1180 1185
Trp Pro Trp Tyr Ile Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala
1190 1195 1200
Ile Val Met Val Thr Ile Met Leu Cys Cys Met Thr Ser Cys Cys
1205 1210 1215
Ser Cys Leu Lys Gly Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe
1220 1225 1230
Asp Glu Asp Asp Ser Glu Pro Val Leu Lys Gly Val Lys Leu His
1235 1240 1245
Tyr Thr
1250
<210> 107
<211> 1263
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 107
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val
1 5 10 15
Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe
20 25 30
Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu
35 40 45
His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp
50 55 60
Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp
65 70 75 80
Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu
85 90 95
Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser
100 105 110
Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile
115 120 125
Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr
130 135 140
Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
145 150 155 160
Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
165 170 175
Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
180 185 190
Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
195 200 205
Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
210 215 220
Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
225 230 235 240
Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
245 250 255
Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
260 265 270
Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
275 280 285
Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
290 295 300
Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
305 310 315 320
Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
325 330 335
Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
340 345 350
Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
355 360 365
Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
370 375 380
Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
385 390 395 400
Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
405 410 415
Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
420 425 430
Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
435 440 445
Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
450 455 460
Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
465 470 475 480
Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
485 490 495
Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
500 505 510
Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
515 520 525
Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
530 535 540
Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
545 550 555 560
Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val
565 570 575
Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
580 585 590
Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
595 600 605
Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile
610 615 620
His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
625 630 635 640
Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
645 650 655
Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
660 665 670
Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala Arg Ser Val Ala
675 680 685
Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser
690 695 700
Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile
705 710 715 720
Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val
725 730 735
Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu
740 745 750
Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr
755 760 765
Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln
770 775 780
Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe
785 790 795 800
Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser
805 810 815
Phe Ile Ala Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly
820 825 830
Asp Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu
835 840 845
Thr Val Leu Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr
850 855 860
Ser Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala
865 870 875 880
Gly Ala Ala Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe
885 890 895
Asn Gly Ile Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu
900 905 910
Ile Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu
915 920 925
Ser Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln
930 935 940
Asn Ala Gln Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe
945 950 955 960
Gly Ala Ile Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys
965 970 975
Val Glu Ala Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln
980 985 990
Ser Leu Gln Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile
995 1000 1005
Arg Ala Ser Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val
1010 1015 1020
Leu Gly Gln Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His
1025 1030 1035
Leu Met Ser Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu
1040 1045 1050
His Val Thr Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala
1055 1060 1065
Pro Ala Ile Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly
1070 1075 1080
Val Phe Val Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn
1085 1090 1095
Phe Tyr Glu Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser
1100 1105 1110
Gly Asn Cys Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr
1115 1120 1125
Asp Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp
1130 1135 1140
Lys Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp
1145 1150 1155
Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile
1160 1165 1170
Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile
1175 1180 1185
Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro
1190 1195 1200
Trp Tyr Ile Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val
1205 1210 1215
Met Val Thr Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys
1220 1225 1230
Leu Lys Gly Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu
1235 1240 1245
Asp Asp Ser Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr
1250 1255 1260
<210> 108
<211> 1250
<212> PRT
<213> 人工序列
<220>
<223> 重组纤突蛋白
<400> 108
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn
290 295 300
Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr
305 310 315 320
Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser
325 330 335
Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr
340 345 350
Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly
355 360 365
Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala
370 375 380
Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly
385 390 395 400
Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe
405 410 415
Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val
420 425 430
Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu
435 440 445
Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser
450 455 460
Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln
465 470 475 480
Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg
485 490 495
Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys
500 505 510
Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe
515 520 525
Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys
530 535 540
Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr
545 550 555 560
Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro
565 570 575
Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser
580 585 590
Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro
595 600 605
Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser
610 615 620
Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala
625 630 635 640
Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly
645 650 655
Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala Arg
660 665 670
Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala
675 680 685
Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn
690 695 700
Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys
705 710 715 720
Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys
725 730 735
Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg
740 745 750
Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val
755 760 765
Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe
770 775 780
Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser
785 790 795 800
Lys Arg Ser Phe Ile Ala Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp
805 810 815
Cys Leu Gly Asp Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe
820 825 830
Asn Gly Leu Thr Val Leu Pro Pro Leu Leu Thr Asp Glu Met Ile Ala
835 840 845
Gln Tyr Thr Ser Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr
850 855 860
Phe Gly Ala Gly Ala Ala Leu Gln Ile Pro Phe Ala Met Gln Met Ala
865 870 875 880
Tyr Arg Phe Asn Gly Ile Gly Val Thr Gln Asn Val Leu Tyr Glu Asn
885 890 895
Gln Lys Leu Ile Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln
900 905 910
Asp Ser Leu Ser Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln Asp Val
915 920 925
Val Asn Gln Asn Ala Gln Ala Leu Asn Thr Leu Val Lys Gln Leu Ser
930 935 940
Ser Asn Phe Gly Ala Ile Ser Ser Val Leu Asn Asp Ile Leu Ser Arg
945 950 955 960
Leu Asp Lys Val Glu Ala Glu Val Gln Ile Asp Arg Leu Ile Thr Gly
965 970 975
Arg Leu Gln Ser Leu Gln Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala
980 985 990
Ala Glu Ile Arg Ala Ser Ala Asn Leu Ala Ala Thr Lys Met Ser Glu
995 1000 1005
Cys Val Leu Gly Gln Ser Lys Arg Val Asp Phe Cys Gly Lys Gly
1010 1015 1020
Tyr His Leu Met Ser Phe Pro Gln Ser Ala Pro His Gly Val Val
1025 1030 1035
Phe Leu His Val Thr Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr
1040 1045 1050
Thr Ala Pro Ala Ile Cys His Asp Gly Lys Ala His Phe Pro Arg
1055 1060 1065
Glu Gly Val Phe Val Ser Asn Gly Thr His Trp Phe Val Thr Gln
1070 1075 1080
Arg Asn Phe Tyr Glu Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe
1085 1090 1095
Val Ser Gly Asn Cys Asp Val Val Ile Gly Ile Val Asn Asn Thr
1100 1105 1110
Val Tyr Asp Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu
1115 1120 1125
Leu Asp Lys Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp Leu
1130 1135 1140
Gly Asp Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln Lys
1145 1150 1155
Glu Ile Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu Ser
1160 1165 1170
Leu Ile Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys
1175 1180 1185
Trp Pro Trp Tyr Ile Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala
1190 1195 1200
Ile Val Met Val Thr Ile Met Leu Cys Cys Met Thr Ser Cys Cys
1205 1210 1215
Ser Cys Leu Lys Gly Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe
1220 1225 1230
Asp Glu Asp Asp Ser Glu Pro Val Leu Lys Gly Val Lys Leu His
1235 1240 1245
Tyr Thr
1250
<210> 109
<211> 1273
<212> PRT
<213> 人工序列
<220>
<223> 重组CoV S多肽
<400> 109
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val
1 5 10 15
Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe
20 25 30
Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu
35 40 45
His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp
50 55 60
Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp
65 70 75 80
Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu
85 90 95
Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser
100 105 110
Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile
115 120 125
Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr
130 135 140
Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
145 150 155 160
Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
165 170 175
Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
180 185 190
Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
195 200 205
Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
210 215 220
Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
225 230 235 240
Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
245 250 255
Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
260 265 270
Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
275 280 285
Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
290 295 300
Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
305 310 315 320
Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
325 330 335
Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
340 345 350
Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
355 360 365
Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
370 375 380
Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
385 390 395 400
Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
405 410 415
Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
420 425 430
Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
435 440 445
Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
450 455 460
Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
465 470 475 480
Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
485 490 495
Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
500 505 510
Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
515 520 525
Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
530 535 540
Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
545 550 555 560
Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val
565 570 575
Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
580 585 590
Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
595 600 605
Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile
610 615 620
His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
625 630 635 640
Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
645 650 655
Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
660 665 670
Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gly Ser Ala Ser Ser Val Ala
675 680 685
Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser
690 695 700
Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile
705 710 715 720
Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val
725 730 735
Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu
740 745 750
Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr
755 760 765
Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln
770 775 780
Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe
785 790 795 800
Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser
805 810 815
Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly
820 825 830
Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp
835 840 845
Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu
850 855 860
Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly
865 870 875 880
Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln Ile
885 890 895
Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr
900 905 910
Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn
915 920 925
Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala
930 935 940
Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn
945 950 955 960
Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val
965 970 975
Leu Asn Asp Ile Leu Ser Arg Leu Asp Pro Pro Glu Ala Glu Val Gln
980 985 990
Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val
995 1000 1005
Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn
1010 1015 1020
Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys
1025 1030 1035
Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro
1040 1045 1050
Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val
1055 1060 1065
Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His
1070 1075 1080
Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn
1085 1090 1095
Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln
1100 1105 1110
Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val
1115 1120 1125
Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro
1130 1135 1140
Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn
1145 1150 1155
His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn
1160 1165 1170
Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu
1175 1180 1185
Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu
1190 1195 1200
Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile Trp Leu
1205 1210 1215
Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr Ile Met
1220 1225 1230
Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly Cys Cys
1235 1240 1245
Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser Glu Pro
1250 1255 1260
Val Leu Lys Gly Val Lys Leu His Tyr Thr
1265 1270
<210> 110
<211> 1260
<212> PRT
<213> 人工序列
<220>
<223> 重组CoV S多肽
<400> 110
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn
290 295 300
Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr
305 310 315 320
Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser
325 330 335
Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr
340 345 350
Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly
355 360 365
Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala
370 375 380
Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly
385 390 395 400
Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe
405 410 415
Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val
420 425 430
Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu
435 440 445
Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser
450 455 460
Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln
465 470 475 480
Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg
485 490 495
Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys
500 505 510
Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe
515 520 525
Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys
530 535 540
Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr
545 550 555 560
Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro
565 570 575
Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser
580 585 590
Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro
595 600 605
Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser
610 615 620
Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala
625 630 635 640
Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly
645 650 655
Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gly Ser Ala Ser
660 665 670
Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala
675 680 685
Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn
690 695 700
Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys
705 710 715 720
Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys
725 730 735
Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg
740 745 750
Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val
755 760 765
Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe
770 775 780
Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser
785 790 795 800
Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala
805 810 815
Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala
820 825 830
Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu
835 840 845
Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu
850 855 860
Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala
865 870 875 880
Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile
885 890 895
Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn
900 905 910
Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr
915 920 925
Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln
930 935 940
Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile
945 950 955 960
Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Pro Pro Glu Ala
965 970 975
Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln
980 985 990
Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser
995 1000 1005
Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln
1010 1015 1020
Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser
1025 1030 1035
Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr
1040 1045 1050
Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile
1055 1060 1065
Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val
1070 1075 1080
Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu
1085 1090 1095
Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys
1100 1105 1110
Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu
1115 1120 1125
Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe
1130 1135 1140
Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly
1145 1150 1155
Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu
1160 1165 1170
Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln
1175 1180 1185
Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile
1190 1195 1200
Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr
1205 1210 1215
Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly
1220 1225 1230
Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser
1235 1240 1245
Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr
1250 1255 1260
<210> 111
<211> 4
<212> PRT
<213> 人工序列
<220>
<223> 重组弗林蛋白酶切割位点
<400> 111
Gly Ser Gly Ala
1
<210> 112
<211> 1260
<212> PRT
<213> 人工序列
<220>
<223> 重组SARS-CoV-2纤突蛋白
<400> 112
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn
290 295 300
Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr
305 310 315 320
Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser
325 330 335
Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr
340 345 350
Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly
355 360 365
Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala
370 375 380
Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly
385 390 395 400
Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe
405 410 415
Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val
420 425 430
Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu
435 440 445
Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser
450 455 460
Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln
465 470 475 480
Ser Tyr Gly Phe Gln Pro Thr Tyr Gly Val Gly Tyr Gln Pro Tyr Arg
485 490 495
Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys
500 505 510
Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe
515 520 525
Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys
530 535 540
Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr
545 550 555 560
Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro
565 570 575
Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser
580 585 590
Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro
595 600 605
Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser
610 615 620
Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala
625 630 635 640
Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly
645 650 655
Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln
660 665 670
Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala
675 680 685
Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn
690 695 700
Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys
705 710 715 720
Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys
725 730 735
Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg
740 745 750
Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val
755 760 765
Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe
770 775 780
Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser
785 790 795 800
Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala
805 810 815
Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala
820 825 830
Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu
835 840 845
Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu
850 855 860
Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala
865 870 875 880
Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile
885 890 895
Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn
900 905 910
Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr
915 920 925
Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln
930 935 940
Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile
945 950 955 960
Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Pro Pro Glu Ala
965 970 975
Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln
980 985 990
Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser
995 1000 1005
Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln
1010 1015 1020
Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser
1025 1030 1035
Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr
1040 1045 1050
Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile
1055 1060 1065
Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val
1070 1075 1080
Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu
1085 1090 1095
Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys
1100 1105 1110
Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu
1115 1120 1125
Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe
1130 1135 1140
Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly
1145 1150 1155
Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu
1160 1165 1170
Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln
1175 1180 1185
Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile
1190 1195 1200
Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr
1205 1210 1215
Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly
1220 1225 1230
Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser
1235 1240 1245
Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr
1250 1255 1260
<210> 113
<211> 1260
<212> PRT
<213> 人工序列
<220>
<223> 重组SARS-CoV-2纤突蛋白
<400> 113
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn
290 295 300
Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr
305 310 315 320
Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser
325 330 335
Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr
340 345 350
Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly
355 360 365
Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala
370 375 380
Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly
385 390 395 400
Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe
405 410 415
Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val
420 425 430
Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu
435 440 445
Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser
450 455 460
Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln
465 470 475 480
Ser Tyr Gly Phe Gln Pro Thr Tyr Gly Val Gly Tyr Gln Pro Tyr Arg
485 490 495
Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys
500 505 510
Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe
515 520 525
Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys
530 535 540
Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr
545 550 555 560
Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro
565 570 575
Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser
580 585 590
Asn Gln Val Ala Val Leu Tyr Gln Gly Val Asn Cys Thr Glu Val Pro
595 600 605
Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser
610 615 620
Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala
625 630 635 640
Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly
645 650 655
Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln
660 665 670
Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala
675 680 685
Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn
690 695 700
Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys
705 710 715 720
Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys
725 730 735
Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg
740 745 750
Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val
755 760 765
Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe
770 775 780
Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser
785 790 795 800
Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala
805 810 815
Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala
820 825 830
Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu
835 840 845
Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu
850 855 860
Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala
865 870 875 880
Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile
885 890 895
Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn
900 905 910
Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr
915 920 925
Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln
930 935 940
Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile
945 950 955 960
Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Pro Pro Glu Ala
965 970 975
Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln
980 985 990
Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser
995 1000 1005
Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln
1010 1015 1020
Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser
1025 1030 1035
Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr
1040 1045 1050
Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile
1055 1060 1065
Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val
1070 1075 1080
Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu
1085 1090 1095
Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys
1100 1105 1110
Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu
1115 1120 1125
Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe
1130 1135 1140
Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly
1145 1150 1155
Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu
1160 1165 1170
Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln
1175 1180 1185
Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile
1190 1195 1200
Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr
1205 1210 1215
Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly
1220 1225 1230
Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser
1235 1240 1245
Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr
1250 1255 1260
<210> 114
<211> 1257
<212> PRT
<213> 人工序列
<220>
<223> 重组SARS-CoV-2纤突蛋白
<400> 114
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile Ser Gly Thr Asn Gly Thr Lys Arg Phe
50 55 60
Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr
65 70 75 80
Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp
85 90 95
Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val
100 105 110
Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val
115 120 125
Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
130 135 140
Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
145 150 155 160
Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
165 170 175
Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
180 185 190
Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
195 200 205
Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
210 215 220
Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
225 230 235 240
Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
245 250 255
Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
260 265 270
Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
275 280 285
Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
290 295 300
Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
305 310 315 320
Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
325 330 335
Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
340 345 350
Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
355 360 365
Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
370 375 380
Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
385 390 395 400
Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
405 410 415
Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
420 425 430
Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
435 440 445
Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
450 455 460
Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
465 470 475 480
Phe Gln Pro Thr Tyr Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
485 490 495
Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
500 505 510
Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
515 520 525
Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
530 535 540
Pro Phe Gln Gln Phe Gly Arg Asp Ile Asp Asp Thr Thr Asp Ala Val
545 550 555 560
Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
565 570 575
Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
580 585 590
Ala Val Leu Tyr Gln Gly Val Asn Cys Thr Glu Val Pro Val Ala Ile
595 600 605
His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
610 615 620
Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
625 630 635 640
Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
645 650 655
Ser Tyr Gln Thr Gln Thr Asn Ser His Gln Gln Ala Gln Ser Val Ala
660 665 670
Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser
675 680 685
Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Ile Asn Phe Thr Ile
690 695 700
Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val
705 710 715 720
Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu
725 730 735
Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr
740 745 750
Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln
755 760 765
Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe
770 775 780
Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser
785 790 795 800
Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly
805 810 815
Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp
820 825 830
Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu
835 840 845
Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly
850 855 860
Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln Ile
865 870 875 880
Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr
885 890 895
Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn
900 905 910
Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala
915 920 925
Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn
930 935 940
Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val
945 950 955 960
Leu Asn Asp Ile Leu Ala Arg Leu Asp Pro Pro Glu Ala Glu Val Gln
965 970 975
Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val
980 985 990
Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn Leu
995 1000 1005
Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys Arg
1010 1015 1020
Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro Gln
1025 1030 1035
Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val Pro
1040 1045 1050
Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His Asp
1055 1060 1065
Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn Gly
1070 1075 1080
Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln Ile
1085 1090 1095
Ile Thr Thr His Asn Thr Phe Val Ser Gly Asn Cys Asp Val Val
1100 1105 1110
Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro Glu
1115 1120 1125
Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn His
1130 1135 1140
Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn Ala
1145 1150 1155
Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu Val
1160 1165 1170
Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu Gly
1175 1180 1185
Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile Trp Leu Gly
1190 1195 1200
Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr Ile Met Leu
1205 1210 1215
Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly Cys Cys Ser
1220 1225 1230
Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser Glu Pro Val
1235 1240 1245
Leu Lys Gly Val Lys Leu His Tyr Thr
1250 1255
<210> 115
<211> 1260
<212> PRT
<213> 人工序列
<220>
<223> 重组SARS-CoV-2纤突蛋白
<400> 115
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Ala Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Gly Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu His Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn
290 295 300
Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr
305 310 315 320
Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser
325 330 335
Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr
340 345 350
Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly
355 360 365
Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala
370 375 380
Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly
385 390 395 400
Gln Thr Gly Asn Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe
405 410 415
Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val
420 425 430
Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu
435 440 445
Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser
450 455 460
Thr Pro Cys Asn Gly Val Lys Gly Phe Asn Cys Tyr Phe Pro Leu Gln
465 470 475 480
Ser Tyr Gly Phe Gln Pro Thr Tyr Gly Val Gly Tyr Gln Pro Tyr Arg
485 490 495
Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys
500 505 510
Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe
515 520 525
Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys
530 535 540
Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr
545 550 555 560
Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro
565 570 575
Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser
580 585 590
Asn Gln Val Ala Val Leu Tyr Gln Gly Val Asn Cys Thr Glu Val Pro
595 600 605
Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser
610 615 620
Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala
625 630 635 640
Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly
645 650 655
Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Gln Gln Ala Gln
660 665 670
Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Val
675 680 685
Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn
690 695 700
Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys
705 710 715 720
Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys
725 730 735
Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg
740 745 750
Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val
755 760 765
Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe
770 775 780
Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser
785 790 795 800
Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala
805 810 815
Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala
820 825 830
Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu
835 840 845
Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu
850 855 860
Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala
865 870 875 880
Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile
885 890 895
Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn
900 905 910
Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr
915 920 925
Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln
930 935 940
Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile
945 950 955 960
Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Pro Pro Glu Ala
965 970 975
Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln
980 985 990
Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser
995 1000 1005
Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln
1010 1015 1020
Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser
1025 1030 1035
Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr
1040 1045 1050
Tyr Val Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile
1055 1060 1065
Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val
1070 1075 1080
Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu
1085 1090 1095
Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys
1100 1105 1110
Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu
1115 1120 1125
Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe
1130 1135 1140
Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly
1145 1150 1155
Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu
1160 1165 1170
Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln
1175 1180 1185
Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile
1190 1195 1200
Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr
1205 1210 1215
Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly
1220 1225 1230
Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser
1235 1240 1245
Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr
1250 1255 1260
<210> 116
<211> 196
<212> PRT
<213> β冠状病毒严重急性呼吸系统综合征冠状病毒2
<400> 116
Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val
1 5 10 15
Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser
20 25 30
Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val
35 40 45
Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp
50 55 60
Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln
65 70 75 80
Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr
85 90 95
Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly
100 105 110
Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys
115 120 125
Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr
130 135 140
Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser
145 150 155 160
Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val
165 170 175
Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly
180 185 190
Pro Lys Lys Ser
195
<210> 117
<211> 13
<212> PRT
<213> 人工序列
<220>
<223> 信号肽
<400> 117
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ile
1 5 10
<210> 118
<211> 292
<212> PRT
<213> β冠状病毒严重急性呼吸系统综合征冠状病毒2
<400> 118
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
Thr Leu Lys Ser
290
<210> 119
<211> 83
<212> PRT
<213> β冠状病毒严重急性呼吸系统综合征冠状病毒2
<400> 119
Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr Gln Asn Val Leu Tyr
1 5 10 15
Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys
20 25 30
Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln
35 40 45
Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn Thr Leu Val Lys Gln
50 55 60
Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val Leu Asn Asp Ile Leu
65 70 75 80
Ser Arg Leu
<210> 120
<211> 588
<212> PRT
<213> β冠状病毒严重急性呼吸系统综合征冠状病毒2
<400> 120
Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala
1 5 10 15
Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn
20 25 30
Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys
35 40 45
Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys
50 55 60
Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg
65 70 75 80
Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val
85 90 95
Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe
100 105 110
Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser
115 120 125
Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala
130 135 140
Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala
145 150 155 160
Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu
165 170 175
Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu
180 185 190
Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala
195 200 205
Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile
210 215 220
Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn
225 230 235 240
Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr
245 250 255
Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln
260 265 270
Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile
275 280 285
Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala
290 295 300
Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln
305 310 315 320
Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser
325 330 335
Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser
340 345 350
Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro
355 360 365
Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val Pro
370 375 380
Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His Asp Gly
385 390 395 400
Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn Gly Thr His
405 410 415
Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln Ile Ile Thr Thr
420 425 430
Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val Val Ile Gly Ile Val
435 440 445
Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys
450 455 460
Glu Glu Leu Asp Lys Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp
465 470 475 480
Leu Gly Asp Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln Lys
485 490 495
Glu Ile Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu
500 505 510
Ile Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro
515 520 525
Trp Tyr Ile Trp Leu Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met
530 535 540
Val Thr Ile Met Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys
545 550 555 560
Gly Cys Cys Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser
565 570 575
Glu Pro Val Leu Lys Gly Val Lys Leu His Tyr Thr
580 585
<210> 121
<211> 672
<212> PRT
<213> β冠状病毒严重急性呼吸系统综合征冠状病毒2
<400> 121
Gln Cys Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr
1 5 10 15
Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser
20 25 30
Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn
35 40 45
Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys
50 55 60
Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala
65 70 75 80
Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr
85 90 95
Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn
100 105 110
Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu
115 120 125
Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe
130 135 140
Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln
145 150 155 160
Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu
165 170 175
Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser
180 185 190
Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser
195 200 205
Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg
210 215 220
Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp
225 230 235 240
Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr
245 250 255
Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile
260 265 270
Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys
275 280 285
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Claims (57)
1.一种冠状病毒(CoV)纤突(S)糖蛋白，其包含
(i)具有灭活的弗林蛋白酶切割位点的S1亚基，其中所述S1亚基包含N末端结构域(NTD)、受体结合结构域(RBD)、亚结构域1和2(SD1/2)，其中所述灭活的弗林蛋白酶切割位点具有QQAQ的氨基酸序列；
其中所述NTD任选地包含选自以下的一个或多个修饰：
(a)选自氨基酸56、57、131、132、229、230、231及其组合的一个或多个氨基酸的缺失；和
(b)选自氨基酸67、229、202、139、5、233、7、13、125、177及其组合的一个或多个氨基酸的突变；
其中所述RBD任选地包含选自氨基酸488、404、471、439、426、440及其组合的一个或多个氨基酸的突变；
其中所述SD1/2结构域任选地包含选自601、557、668、642及其组合的一个或多个氨基酸的突变；以及
(ii)S2亚基，其中氨基酸973和974是脯氨酸，
其中所述S2亚基任选地包含选自以下的一个或多个修饰：
(a)来自676-702、702-711、775-793、806-815及其组合的一个或多个氨基酸的缺失；
(b)选自703、1105、688、969和1014及其组合的一个或多个氨基酸的突变；和
(c)一个或多个氨基酸从TMCT中的缺失。
2.根据权利要求1所述的冠状病毒S糖蛋白，其包含氨基酸676-685的缺失。
3.根据权利要求1所述的冠状病毒S糖蛋白，其包含氨基酸702-711的缺失。
4.根据权利要求1所述的冠状病毒S糖蛋白，其包含氨基酸806-815的缺失。
5.根据权利要求1所述的冠状病毒S糖蛋白，其包含氨基酸775-793的缺失。
6.根据权利要求1所述的冠状病毒S糖蛋白，其包含所述NTD的氨基酸1-292的缺失。
7.根据权利要求1所述的冠状病毒S糖蛋白，其包含跨膜和胞质尾(TMCT)的氨基酸1201-1260的缺失。
8.根据权利要求1所述的冠状病毒S糖蛋白，其包含选自SEQ ID NO:85-89、105、106和112-115的氨基酸序列，或由所述氨基酸序列组成。
9.根据权利要求1至8中任一项所述的冠状病毒S糖蛋白，其包含信号肽，任选地其中所述信号肽包含SEQ ID NO:5或SEQ ID NO:117的氨基酸序列。
10.根据权利要求1至9中任一项所述的冠状病毒S糖蛋白，其包含C末端融合蛋白。
11.根据权利要求10所述的冠状病毒S糖蛋白，其中所述C末端融合蛋白是六组氨酸标签。
12.根据权利要求10所述的冠状病毒S糖蛋白，其中所述C末端融合蛋白是折叠子。
13.根据权利要求12所述的冠状病毒S糖蛋白，其中所述折叠子具有对应于SEQ ID NO:68的氨基酸序列。
14.根据权利要求1-13中任一项所述的冠状病毒S糖蛋白，其中ΔHcal是野生型CoV S糖蛋白(SEQ ID NO:2)的ΔHcal的至少2倍。
15.根据权利要求1-14中任一项所述的冠状病毒S糖蛋白，其中所述S2亚基、所述NTD、所述RBD和所述SD1/2各自与具有SEQ ID NO:2的氨基酸序列的CoV S糖蛋白的对应亚基或结构域具有95％同一性。
16.根据权利要求1-14中任一项所述的冠状病毒S糖蛋白，其中所述S2亚基、所述NTD、所述RBD和所述SD1/2各自与具有SEQ ID NO:2的氨基酸序列的CoV S糖蛋白的对应亚基或结构域具有97％同一性。
17.根据权利要求1-14中任一项所述的冠状病毒S糖蛋白，其中所述S2亚基、所述NTD、所述RBD和所述SD1/2各自与具有SEQ ID NO:2的氨基酸序列的CoV S糖蛋白的对应亚基或结构域具有99％同一性。
18.根据权利要求1-14中任一项所述的冠状病毒S糖蛋白，其中所述S2亚基、所述NTD、所述RBD和所述SD1/2各自与具有SEQ ID NO:2的氨基酸序列的CoV S糖蛋白的对应亚基或结构域具有99.5％同一性。
19.一种分离的核酸，其编码根据权利要求1-18中任一项所述的S糖蛋白。
20.一种载体，其包含根据权利要求19所述的核酸。
21.一种纳米颗粒，其包含根据权利要求1-18中任一项所述的冠状病毒S糖蛋白。
22.根据权利要求21所述的纳米颗粒，其中所述纳米颗粒的Zavg直径在约20nm与约35nm之间。
23.根据权利要求21所述的纳米颗粒，其中所述纳米颗粒的多分散性指数为约0.2至约0.45。
24.一种细胞，其表达根据权利要求1-18中任一项所述的冠状病毒S糖蛋白。
25.一种疫苗组合物，其包含根据权利要求1-18中任一项所述的冠状病毒S糖蛋白和药学上可接受的缓冲液。
26.根据权利要求25所述的疫苗组合物，其包含佐剂。
27.根据权利要求26所述的疫苗组合物，其中所述佐剂包含至少两种iscom颗粒，其中：
第一iscom颗粒包含皂树(Quillaja Saponaria Molina)的级分A，而不包含皂树的级分C；并且
第二iscom颗粒包含皂树的级分C，而不包含皂树的级分A。
28.根据权利要求27所述的疫苗组合物，其中皂树的级分A和皂树的级分C分别占在所述佐剂中皂树的级分A和皂树的级分C的重量之和的按重量计约92％和按重量计约8％。
29.根据权利要求26所述的疫苗组合物，其中将所述佐剂以约50μg的剂量施用。
30.一种在受试者中刺激针对SARS-CoV-2的免疫反应的方法，其包括施用根据权利要求25-29中任一项所述的疫苗组合物。
31.根据权利要求30所述的方法，其中向所述受试者在第0天施用第一剂量，并且在第21天施用加强剂量。
32.根据权利要求30所述的方法，其中向所述受试者施用在约5μg与约25μg之间的冠状病毒S糖蛋白。
33.根据权利要求30所述的方法，其中向所述受试者施用约5μg的冠状病毒S糖蛋白。
34.根据权利要求30-33中任一项所述的方法，其中将所述疫苗组合物肌内施用。
35.根据权利要求30、32、33和34中任一项所述的方法，其中施用单剂的所述疫苗组合物。
36.根据权利要求30-34中任一项所述的方法，其中施用多剂的所述疫苗组合物。
37.根据权利要求30-36中任一项所述的方法，其中将所述疫苗组合物与流感糖蛋白共同施用。
38.一种免疫原性组合物，其包含：
(i)包含具有SEQ ID NO:87的氨基酸序列的冠状病毒S(CoV S)糖蛋白和非离子型洗涤剂核心的纳米颗粒；
(ii)药学上可接受的缓冲液；以及
(iii)皂苷佐剂。
39.根据权利要求38所述的免疫原性组合物，其包含在约5μg与约25μg之间的CoV S糖蛋白。
40.根据权利要求39所述的免疫原性组合物，其包含约5μg的CoV S糖蛋白。
41.根据权利要求38所述的免疫原性组合物，其中所述皂苷佐剂包含至少两种iscom颗粒，其中：
第一iscom颗粒包含皂树的级分A，而不包含皂树的级分C；并且
第二iscom颗粒包含皂树的级分C，而不包含皂树的级分A。
42.根据权利要求41所述的免疫原性组合物，其中分别地，皂树的级分A占在所述佐剂中皂树的级分A和皂树的级分C的重量之和的按重量计50％-96％，并且皂树的级分C占剩余部分。
43.根据权利要求41所述的免疫原性组合物，其中皂树的级分A和皂树的级分C分别占在所述佐剂中皂树的级分A和皂树的级分C的重量之和的按重量计约85％和按重量计约15％。
44.根据权利要求38所述的免疫原性组合物，其包含约50μg的皂苷佐剂。
45.根据权利要求38所述的免疫原性组合物，其中所述非离子型洗涤剂核心选自聚山梨醇酯20(PS20)、聚山梨醇酯40(PS40)、聚山梨醇酯60(PS60)、聚山梨醇酯65(PS65)和聚山梨醇酯80(PS80)。
46.一种在受试者中刺激针对SARS-CoV-2的免疫反应的方法，其包括施用根据权利要求38所述的免疫原性组合物。
47.根据权利要求46所述的方法，其包含在约5μg与约25μg之间的CoV S糖蛋白。
48.根据权利要求47所述的方法，其包含5μg的CoV S糖蛋白。
49.根据权利要求46所述的方法，其中所述皂苷佐剂包含至少两种iscom颗粒，其中：
第一iscom颗粒包含皂树的级分A，而不包含皂树的级分C；并且
第二iscom颗粒包含皂树的级分C，而不包含皂树的级分A。
50.根据权利要求49所述的方法，其中分别地，皂树的级分A占在所述佐剂中皂树的级分A和皂树的级分C的重量之和的按重量计50％-96％，并且皂树的级分C占剩余部分。
51.根据权利要求49所述的方法，其中皂树的级分A和皂树的级分C分别占在所述佐剂中皂树的级分A和皂树的级分C的重量之和的按重量计约85％和按重量计约15％。
52.根据权利要求46所述的方法，其包含约50μg的皂苷佐剂。
53.根据权利要求46所述的方法，其中所述非离子型洗涤剂核心选自聚山梨醇酯20(PS20)、聚山梨醇酯40(PS40)、聚山梨醇酯60(PS60)、聚山梨醇酯65(PS65)和聚山梨醇酯80(PS80)。
54.根据权利要求46所述的方法，其中向所述受试者在第0天施用第一剂量，并且在第21天施用加强剂量。
55.根据权利要求46所述的方法，其中施用单剂的所述免疫原性组合物。
56.根据权利要求46所述的方法，其包括施用不同于所述第一免疫原性组合物的第二免疫原性组合物。
57.根据权利要求56所述的方法，其中所述第二免疫原性组合物包含编码SARS-CoV-2纤突糖蛋白的mRNA、编码SARS-CoV-2纤突糖蛋白的质粒DNA、编码SARS-CoV-2纤突糖蛋白的病毒载体或灭活的SARS-CoV-2病毒。
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| EP3046579A1 (en) * | 2013-09-19 | 2016-07-27 | Novavax, Inc. | Immunogenic middle east respiratory syndrome coronavirus (mers-cov) compositions and methods |
| US10426829B2 (en) * | 2015-09-03 | 2019-10-01 | Novavax, Inc. | Vaccine compositions having improved stability and immunogenicity |
| US20170354729A1 (en) * | 2016-03-16 | 2017-12-14 | Novavax, Inc. | Vaccine compositions containing modified zika virus antigens |
| EP3532095A1 (en) * | 2016-10-25 | 2019-09-04 | The U.S.A. as represented by the Secretary, Department of Health and Human Services | Prefusion coronavirus spike proteins and their use |
| MA47790A (fr) * | 2017-03-17 | 2021-05-05 | Modernatx Inc | Vaccins à base d'arn contre des maladies zoonotiques |
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2021
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- 2021-01-27 JP JP2022545141A patent/JP2023512648A/ja active Pending
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| WO2021154812A8 (en) | 2022-02-24 |
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| US20230070886A1 (en) | 2023-03-09 |
| KR20220141302A (ko) | 2022-10-19 |
| JP2023512648A (ja) | 2023-03-28 |
| GB202212336D0 (en) | 2022-10-12 |
| IL295142A (en) | 2022-09-01 |
| BR112022014830A2 (pt) | 2022-09-27 |
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