CN115484976A - 用于移植的调理方案 - Google Patents

用于移植的调理方案 Download PDF

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CN115484976A
CN115484976A CN202180032965.7A CN202180032965A CN115484976A CN 115484976 A CN115484976 A CN 115484976A CN 202180032965 A CN202180032965 A CN 202180032965A CN 115484976 A CN115484976 A CN 115484976A
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罗伯特·博克曼
安娜·卡琳·路易丝·罗伯逊
J·林
科林·查尔斯·安德森
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Lufthansa Biopharmaceutical Co ltd
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Abstract

本发明涉及用于移植细胞、组织或器官、任选地造血干细胞/祖细胞至对象的调理方案。本发明还涉及诱导对象中的造血嵌合体的方法。本发明还涉及预防或治疗对象中的疾病或病症的方法，其中诱导造血嵌合体以便提高后续治疗对对象的益处。后续治疗可以是细胞、组织或器官的移植，或者可以是使用基因改造的造血干细胞/祖细胞进行的基因治疗。

Description

用于移植的调理方案
技术领域
本发明涉及用于移植细胞、组织或器官、任选地造血干细胞/祖细胞(HSPC)至对象的调理方案。本发明还涉及诱导对象中的造血嵌合体的方法。本发明还涉及预防或治疗对象中的疾病或病症的方法，其中诱导造血嵌合体以便提高后续治疗对对象的益处。后续治疗可以是细胞、组织或器官移植，或者可以是使用基因改造的HSPC进行的基因治疗。
背景技术
组织/器官移植体可能因急性和/或慢性排斥反应而受损，这可能导致移植失败(graft failure)。急性和慢性排斥反应二者通常通过免疫抑制剂进行治疗，这可能增加感染风险，增加癌症风险，还可能造成器官衰竭(包括移植体器官衰竭)。可以减少免疫抑制需求的技术(通过建立对移植体的免疫耐受)是在移植细胞、组织或器官之前，通过从同一供者移植造血干细胞和祖细胞(通常是在骨髓移植(BMT)中)来诱导造血嵌合体。诱导的嵌合体实质上产生嵌合免疫系统，该嵌合免疫系统不会攻击具有与供者造血系统相同免疫特征的移植体，同时另外地保留受者的正常免疫能力以对不相关抗原作出反应。
不幸的是，移植HSPC涉及的复杂免疫学可能存在问题，特别是如果在移植前对供者抗原敏化(sensitized)。供者和受者的免疫系统的存在可导致由体液成分和细胞成分介导的急性和慢性排斥反应。剧烈的宿主抗移植体反应(HVG)和移植体抗宿主病(GVHD)都可以观察到。通常，移植的细胞不能成功地在受者体内植入。目前的方法试图通过移植前和移植后的免疫抑制来解决这些问题。移植前进行的步骤可称为调理方案(conditioningregimen，)，并可以包括不仅仅是免疫抑制的治疗。例如，放射可用于耗竭受者中的一些或全部的现有骨髓细胞，为移植细胞的植入创造空间。然而，植入常常不成功。需要用于移植HSPC的改进的调理方案。
发明内容
用于移植造血干细胞和祖细胞(HSPC)的调理方案通常包括T淋巴细胞耗竭和/或处理，以直接地(例如，通过血浆置换或施用吸附供者特异性抗体(DSA)的不匹配血小板输注)或通过抑制抗体产生而间接地(例如，使用利妥昔单抗(rituximab)或硼替佐米(bortezomib))减少DSA。然而，现有的调理方案经常是低效的，而且植入经常不成功。这可能是因为骨髓来源的细胞上MHC的高表达可能会增加对任何剩余的功能性DSA的敏感性。
本发明人出乎意料地表明，包括对对象中的血清IgG酶促失活的调理方案可显著提高植入率(相比于先前使用的抗体耗竭技术)，因此更有可能在对象中产生造血嵌合体。
本发明提供了一种用于移植HSPC至对象的调理方案，包括向对象施用使对象的血清IgG分子失活的酶。所述酶的施用量优选地足以使对象的血清中存在的所有或基本所有的IgG分子失活。
调理方案还可以包括以下中的一个或多个：
(a)向对象施用非致命剂量的放射和/或任何其他耗竭对象的HSPC的药剂；
(b)施用用于减少对象中淋巴细胞的数量和/或下调对象中淋巴细胞的活性的药剂，其中所述淋巴细胞包括：
i.T细胞；和/或
ii.B细胞(任选地包括产生抗体的细胞)；
(c)施用任何其他降低免疫系统活性的药剂或方案，例如补体抑制剂、细胞因子抑制剂、先天免疫细胞抑制剂、耐受性诱导剂。
调理方案优选地包括至少(a)，但最优选地包括至少(a)和(b)。
本发明还提供了一种诱导对象中的造血嵌合体的方法，该方法包括实施本发明的调理方案，并且随后以足以在对象中诱导造血嵌合体的量和在适于在对象中诱导造血嵌合体的条件下向对象施用HSPC。HSPC可以是自体的(使用对象自己的细胞)或同种异体的(细胞来自不同的供者)。HSPC可以是基因改造的，在这种情况下，它们优选地是自体的。基因改造可以用于表达任何基因，但通常是对受者有治疗益处的基因，在这种情况下，HSPC可以被称为表达基因疗法。HSPC优选地是同种异体细胞或经基因改造的自体细胞。HSPC最优选地是同种异体的。
本发明还提供了一种预防或治疗对象的疾病或病症的方法，其中根据本发明的方法诱导对象中的造血嵌合体，以提高该疾病或病症的疗法对对象的益处。该疗法可以是细胞、组织或器官移植体，通常与HSPC来自同一供者。移植的细胞、组织或器官可以属于任何类型，包括肾脏、肝脏、心脏、胰腺、肺、小肠、皮肤、血管/血管组织、面部、手臂、气管、眼睛的部分、胰岛、黑质、骨髓或干细胞。移植的细胞可以属于任何类型，包括该方法本身使用的相同的HSPC，如此不再需要额外的治疗。
换言之，本发明还提供了一种预防或治疗细胞、组织或器官移植体的免疫排斥的方法，该方法包括根据本发明的方法诱导对象中的造血嵌合体，并且向对象施用细胞、组织或器官移植体，任选地其中所述细胞、组织或器官与HSPC来自同一供者。该细胞、组织或器官通常是在诱导对象中的造血嵌合体后施用的。细胞、组织或器官移植体可以属于任何类型，包括肾脏、肝脏、心脏、胰腺、肺、小肠、皮肤、血管/血管组织、面部、手臂、气管、眼睛的部分、胰岛、黑质、骨髓或干细胞。移植的细胞可以属于任何类型，包括与用于诱导造血嵌合体的相同的HSPC，如此不再需要额外的移植体。
以另一种方式表述，本发明提供一种预防或治疗细胞、组织或器官移植体的免疫排斥的方法，包括
(i)实施本发明的调理方案。
(ii)以足以在对象中诱导造血嵌合体的量和在适于在对象中诱导造血嵌合体的条件下将HSPC施用至对象；和
(iii)将与HSPC来自同一供者的细胞、组织或器官移植体施用至对象，任选地其中所述移植体是步骤(ii)中施用的HSPC。
在步骤(iii)中的细胞、组织或器官是步骤(ii)的HSPC时，本发明的方法可以是对通过HSPC移植治疗的疾病或病症进行治疗的方法。在将HSPC基因改造以施用基因治疗时，本发明的方法可以用于预防或治疗该基因治疗所针对的疾病或病症。
附图说明
图1.EndoS抑制单克隆DSA介导的对供者骨髓细胞的杀伤。
对幼稚NOD(小图A和B)或B6.H-2g7(小图C和D)静脉内给予30μg EndoS和/或抗H-2Kb mAb(10μg或100μg)，四小时后输注CFSE标记的NOD/CTV标记的B6骨髓细胞的混合物(BMC；小图A和B)或CFSE标记的B6.H-2g7/CTV标记的NOD.H-2b BMC的混合物(小图C和D)。所示为NOD(小图A)和B6.H-2g7(小图C)的实验方案。所示为骨髓移植(BMT)后1至3小时收集的血液(左图)、BMT后4小时收集的宿主脾脏(中图)和骨髓(BM，右图)中的染料标记的B6与NOD细胞的比率(小图B)或NOD.H-2b细胞与B6.H-2g7细胞的比率(小图D)。以平均值±SEM示出。数据来自五个(小图B)和四个(小图D)独立实验的集合。使用曼-惠特尼U检验(中图和右图)以用于显示比较；*p<0.05和**p<0.01。
图2.EndoS-imlifidase减少敏化受者中DSA介导的对供者BMC的杀伤。
(A-C)用FVB脾细胞免疫幼稚NOD小鼠，四周后施用EndoS-imlifidase。获取免疫前、酶处理前和酶处理后四小时的血清。左边的代表性直方图是用于DSA-IgG Fc(小图A)、DSA-IgG1 Fc(小图B)、DSA-IgG3 Fc(小图C)和DSA-IgG3重链(小图D)，血清稀释度为1:25。右边示出滴定血清中DSA的平均荧光强度(MFI)。以平均值±SEM示出。用比率配对t检验来比较每个血清稀释度下酶处理之前和之后的DSA的MFI，*p<0.05，**p<0.01。(D)E-F中示出实验示意图。用B6.CD45.1脾细胞免疫幼稚NOD小鼠，在四周后注射T细胞耗竭性单抗。在T细胞耗竭后的两天施用EndoS-imlifidase。酶处理后四小时，NOD小鼠被静脉内注射8000万个B6.CD45.2骨髓细胞。分析脾细胞和骨髓细胞的MHC-I H-2Kb和CD45.2表达。(E-F)示出四个不同处理组的代表性点阵图(左)和供者细胞的百分比(右，平均值±SEM)。带霍尔姆-西达克多重比较检验的单因素方差分析用于比较三个敏化组之间的值，*p<0.05。
图3.进行BMT前的硼替佐米/环磷酰胺可减少敏化受者中的骨髓B细胞。
(A)B-E示出实验的示意图。用FVB脾细胞免疫后4周，用环磷酰胺和硼替佐米(CyBor)静脉内处理NOD小鼠。在CyBor处理后四天，用2000万个FVB BMC进行骨髓移植。在BMT后五天收集脾细胞和骨髓细胞进行分析。收集CyBor处理前以及BMT后五天的血清。示出的是给予CyBor或溶媒的小鼠中骨髓(小图B)和脾脏(小图C)中的B细胞和血浆细胞的细胞计数。(D)收集免疫前以及BMT后5天(即CyBor处理后9天)的血清。示出的是来自个体对照的小鼠(左)或CyBor处理小鼠(右)的滴定血清中DSA-IgG Fc的MFI。(E)示出的是1:25稀释度的DSA的MFI在第9天与处理前相比的百分数变化。填充符号和空心符号代表在两个独立实验中收集的数据。
图4.EndoS-imlifidase允许预敏化受者具有造血嵌合体
(A)嵌合体诱导方案的示意图；用FVB脾细胞免疫幼稚B6.H-2g7或NOD小鼠，四周至六周后进行嵌合体诱导。对于嵌合体诱导，在基于BMT当日起往前算的第4天给予CyBor。在BMT前的第2天、BMT后第2天、第6天、第11天和第16天腹膜内注射T细胞耗竭性(TCD)抗体。对已经用FVB脾细胞敏化的一些受者在BMT前的第6天静脉内注射EndoS-imlifidase来进行处理，并在第0天进行BMT前的4小时重复给药。在第0天进行BMT前的4小时给予6Gy全身放射。在第0天给予FVB BMC(80×106)。(B)示出的是外周血中淋巴细胞门中供者细胞随时间的比率。(C)示出的是来自幼稚NOD嵌合体(n＝4，左，平均值±SEM)和致敏的(primed)NOD嵌合体(n＝2，右)的外周血中淋巴细胞门中不同谱系的供者细胞的百分比。数据来自六个独立实验的集合。
图5.imlifidase平均浓度与给药后的标称时间的关系(N＝15)。数据BLQ包括在平均值计算中，作为BLQ/2。用棒表示SD。
图6.imlifidase对rATG随时间的体外切割。柱表示在imlifidase后蛋白质印迹上具有可见完整rATG的对象数量(N＝11)。
序列的简要描述
SEQ ID NO:1是IdeS的完整序列，包括N端甲硫氨酸和信号序列。它也可作为NCBI参考序列号WP_010922160.1获得。
SEQ ID NO:2是IdeS的成熟序列，缺少N端甲硫氨酸和信号序列。它也可作为Genbank登录号ADF13949.1获得。
SEQ ID NO:3是IdeZ的完整序列，包括N端甲硫氨酸和信号序列。它也可作为NCBI参考序列号WP_014622780.1获得。
SEQ ID NO:4是IdeZ的成熟序列，缺少N端甲硫氨酸和信号序列。
SEQ ID NO:5是杂合IdeS/Z的序列。N端基于缺少N端甲硫氨酸和信号序列的IdeZ。
SEQ ID NO:6至25是用于在本发明方法中使用的示例性蛋白酶的序列。
SEQ ID NO:26是IdeS多肽的序列。包含SEQ ID NO:2的序列，带有附加的N端甲硫氨酸和组氨酸标签(内部参考pCART124)。
SEQ ID NO:27是IdeZ多肽的序列。包含SEQ ID NO:4的序列，带有附加的N端甲硫氨酸和组氨酸标签(内部参考pCART144)。
SEQ ID NO:28是IdeS/Z多肽的序列。包含SEQ ID NO:5的序列，带有附加的N端甲硫氨酸和组氨酸标签(内部参考pCART145)。
SEQ ID NO:29是连续序列PLTPEQFRYNN，其对应于SEQ ID NO:3的位置63-73。
SEQ ID NO:30是连续序列PPANFTQG，其对应于SEQ ID NO:1的位置58-65。
SEQ ID NO:31是连续序列DDYQRNATEAYAKEVPHQIT，其对应于SEQ ID NO:3的位置35-54。
SEQ ID NO:32是连续序列DSFSANQEIRYSEVTPYHVT，其对应于SEQ ID NO:1的位置30-49。
SEQ ID NO:33至55是编码上述蛋白酶的核苷酸序列。
SEQ ID NO:56至69是用于在本发明方法中使用的示例性蛋白酶的序列。
SEQ ID NO:70是连续序列NQTN，其对应于SEQ ID NO:1的位置336-339。
SEQ ID NO:71是连续序列DSFSANQEIR YSEVTPYHVT，其对应于SEQ ID NO:1的位置30-49。
SEQ ID NO:72至86是编码本文公开的多肽的核苷酸序列。
SEQ ID NO:87是序列SFSANQEIRY SEVTPYHVT，其对应于SEQ ID NO:1的位置31-49。
SEQ ID NO:88是序列DYQRNATEAY AKEVPHQIT，其对应于IdeZ多肽(NCBI参考序列号WP_014622780.1)的位置36-54。
SEQ ID NO:89是序列DDYQRNATEA YAKEVPHQIT，其可以存在于本发明多肽的N端。
SEQ ID NO:90是EndoS(酿脓链球菌的内切糖苷酶)的成熟序列。
具体实施方式
一般知识
应当理解的是，所公开的产品和方法的不同应用可以根据本领域的具体需要进行调整。还应理解的是，本文使用的术语仅用于描述本发明的特定实施方式的目的，而不是为了限制。
此外，如在本说明书和所附权利要求书中使用的，除非内容另有明确规定，单数形式“一个/一种(a/an)”和“该(the)”包括复数指称。因此，例如，对“多肽(polypeptide)”的提及包括“多个多肽(polypeptides)”等。
“多肽”在本文中以其最广泛的含义使用，是指两个或更多个亚基氨基酸的化合物、氨基酸类似物或其他肽模拟物。因此，术语“多肽”包括短肽序列以及更长的多肽和蛋白质。如本文所用，术语“氨基酸”是指天然和/或非天然或合成的氨基酸，包括D或L旋光异构体二者和氨基酸类似物和肽模拟物。
术语“患者”和“对象”可互换使用，并且通常指的是人。除非另有说明，否则对IgG的提及通常是指人IgG。
如上所述的氨基酸同一性可以使用任何合适的算法来计算。例如，PILEUP和BLAST算法可用于计算同一性或排列序列(例如识别等效序列或相应序列(通常以它们的默认设置)，例如Altschul SF(1993)J Mol Evol 36:290-300；Altschul，S，F等人(1990)J MolBiol 215:403-10中所述。执行BLAST分析的软件可以通过国家生物技术信息中心公开获得。该算法包括首先通过识别查询序列中长度W的短字段(word)来识别高评分序列对(HSP)，这些短字段在与数据库序列中相同长度的字段对齐时匹配或满足一些正值阈值得分T。T被称为邻域字段得分阈值(Altschul等人，同上)。这些初始邻域字段命中(word hit)充当启动搜索以查找包含它们的HSP的种子。字段命中沿着每个序列在两个方向上扩展，只要可以增加累积对齐分数。在以下情况下，字段命中在每个方向上的扩展都会停止：在累积对齐得分从其最大实现值下降数量X；由于一个或多个负得分残基对齐的累积，累积得分变为零或更低；或到达任一序列的末端。BLAST算法参数W、T和X确定对齐的灵敏度和速度。BLAST程序使用以下作为默认值：字长(W)为11，BLOSUM62得分矩阵(参见Henikoff和Henikoff(1992)Proc.Natl.Acad.Sci.USA 89:10915-10919)对齐(B)为50，期望(E)为10，M＝5，N＝4，以及两条链的比较。
BLAST算法对两个序列之间的相似性进行统计分析；参见例如，Karlin和Altschul(1993)Proc.Natl.Acad.Sci.USA 90:5873-5787。BLAST算法提供的相似性的一种度量是最小和概率(smallest sum probability)(P(N))，它提供对两个多核苷酸或氨基酸序列之间的匹配是偶然发生的概率的指示。例如，如果一个序列与另一个序列相比的最小和概率小于约1，优选小于约0.1，更优选小于约0.01，最优选小于约0.001，则第一序列被认为与第二序列相似。或者，UWGCG包提供可以用于计算同一性的BESTFIT程序(例如在其默认设置上使用)(Devereux等人(1984)Nucleic Acids Research 12，387-395)。
本文引用的所有出版物、专利和专利申请，无论在上文还是下文中，通过引用其整体并入本文。
调理方案
本发明提供了一种用于移植细胞、组织或器官至对象的调理方案，包括向对象施用使对象中的血清IgG分子失活的酶。所述酶的施用量优选地足以使对象的血清中存在的所有或基本所有的IgG分子失活。如有需要，可以组合施用一种以上的IgG失活酶，包括同时地或依次地施用，以任意顺序进行。
术语“血清IgG分子”或“血清中存在的IgG分子”是指在实施本发明方法之前存在于人体组织或循环中的任何γ免疫球蛋白(IgG1、IgG2、IgG3和IgG4)分子。这种IgG分子可以是由个体的B细胞内源性产生的，也可以是在实施本发明方法之前已经施用至对象的外源性γ免疫球蛋白，包括任何来源的任何治疗性IgG分子。血清IgG的失活通常意味着IgG分子的Fc受体相互作用的降低。术语“Fc受体”是指存在于细胞上的Fcγ免疫球蛋白受体(FcγR)。在人中，FcγR是指包括FcγRI(CD64)、FcγRIIA(CD32A)、FcγRIIB(CD32B)、FcγRIIC(CD32C)、FcγRIIIA(CD16a)和FcγRIIIB(CD16b)在内的受体家族中的一种、一些或全部。如本文所用，术语FcγR包括FcγRI(CD64)、FcγRIIA(CD32A)、FcγRIIB(CD32B)、FcγRIIC(CD32C)、FcγRIIIA(CD16a)和FcγRIIIB(CD16b)的天然存在的多态性。
本发明方法中使用的酶可以是使血清IgG失活的任何酶，但通常是IgG半胱氨酸蛋白酶，它能切割IgG使抗原结合结构域和Fc相互作用结构域彼此分离。在这种情况下，血清IgG分子的Fc受体相互作用降低，因为血清中完整IgG分子的量降低。作为另一个实例，酶可以是IgG内切糖苷酶，它能切割IgG的Fc相互作用结构域上的聚糖结构，特别是位于Asn-297位置(Kabat编号)的N-链接的双天线聚糖。这种聚糖结构在Fc受体结合和补体激活中具有关键作用。因此，当它被蛋白质完全或部分去除时，这将导致通过另外的完整IgG分子产生的Fc受体结合或补体激活降低。将在下面的章节中更详细地讨论适合用于调理方案的酶。
酶优选地通过静脉内输注施用，但也可以通过任何合适的途径施用，包括例如皮内、皮下、经皮、肌内、动脉内、腹膜内、关节内、骨内或其他适当的施用途径。酶的施用量可以在0.01mg/kg BW和2mg/kg BW之间，0.05mg/kg BW和1.5mg/kg BW之间，0.1mg/kg BW和1mg/kg BW之间，优选地0.15mg/kg BW和0.7mg/kg BW之间，最优选地0.2mg/kg BW和0.3mg/kg BW之间，尤其是0.25mg/kg BW。可以对同一对象多次施用该酶，前提是对象的血清中能够与该酶结合的抗药物抗体(ADA)的量不超过临床医生确定的阈值。对象的血清中能够与该蛋白酶结合的ADA数量可以通过任何合适的方法确定，如药剂特异性CAP FEIA(ImmunoCAP)试验或滴度测定。如果对象中的ADA超过该阈值，则治疗方案可以包括施用替代性酶。
调理方案还可以包括以下中的一个或多个：
(a)向对象施用非致命剂量的放射和/或任何其他耗竭对象的HSPC的药剂；
(b)施用用于减少对象中淋巴细胞的数量和/或下调对象中淋巴细胞的活性的药剂，其中所述淋巴细胞包括：
i.T细胞；和/或
ii.B细胞(任选地包括产生抗体的细胞)；
(c)施用任何其他调节(例如降低)免疫系统活性的药剂或方案，例如补体抑制剂、细胞因子抑制剂、先天免疫细胞抑制剂、耐受性诱导剂。
步骤(a)通常涉及施用足以部分或完全消除(或消融)对象骨髓的放射剂量。部分消除是优选的，因为副作用通常不那么严重，而且也期望保留一些受者骨髓。受者骨髓的消融在骨髓中为供者HSPC的植入创造空间，但也会耗竭对象的淋巴细胞，因此也会以与步骤(b)相同的方式降低免疫系统的活性。因此，调理方案优选地包括至少(a)，但最优选地包括至少(a)和(b)。或者，在步骤(a)中，可以优选地使用无放射的方法来耗竭对象的HSPC，如施用抗CD117和/或抗CD47。这将为供者HSPC的植入创造空间，而没有一些不期望的放射副作用。除了HSPC耗竭外，对象还可以任选地接受供者CD8-α细胞的输注，这可以增加敏化受者中稳定嵌合体的频率。供者T细胞输注可以通过减少宿主T细胞的存活来促进供者HSPC的植入。
步骤(b)可以通过任何合适的方法且使用任何合适的药剂进行。相同药剂或药剂组合可以有效地减少一种以上的淋巴细胞的数量和/或下调一种以上的淋巴细胞的活性。例如，在非人灵长类动物模型中对预敏化受者进行移植的临床前研究表明，将贝拉西普(belatacept)的共刺激阻断与硼替佐米的血浆细胞耗竭疗法相结合可以持久地抑制DSA并且降低抗体介导的排斥反应的风险。
适合用于耗竭T细胞的示例性药剂在本领域是已知的，包括抗胸腺细胞球蛋白(ATG，如兔或马的ATG)；或包括抗CD4、抗CD8和抗CD90的一组抗体；抗CD52抗体(如阿仑单抗(alemtuzumab))；抗CD117抗体；抗CD45抗体；白消安(busulfan)；环磷酰胺；氟达拉滨(fludarabine)；苏消安(treosulfan)；环孢素；他克莫司(tacrolimus)；或针对T细胞的免疫毒素。
适合用于耗竭B细胞(任选地包括血浆细胞)的示例性药剂是本领域已知的，包括抗CD20抗体(如利妥昔单抗)；抗CD19抗体；硼替佐米；氟达拉滨；环磷酰胺；或针对B细胞的免疫毒素，如抗CD20免疫毒素(例如MT-3724)。
实施例中示出了包括步骤(a)和(b)的示例性方案。该方案包括施用非致命剂量的放射，以及施用包括抗CD4、抗CD8和抗CD90的一组抗体以耗竭T细胞，以及施用硼替佐米和环磷酰胺以耗竭B细胞(包括产生抗体的细胞)。
步骤(a)和(b)通常是彼此分开的，必要时也与使对象中血清IgG分子失活的酶的施用分开，以适合施用以达到期望效果的任何时间作为间隔。例如，当步骤(a)和/或(b)包括抗体类药剂时，期望的是在施用酶之后有足够的时间间隔来发生这些步骤，这样酶就不会使步骤(a)或(b)的抗体类药剂失活。实施例2中示出了示例性的时间间隔。最早可在施用imlifidase后四天开始施用rATG(或其他抗体类治疗药物)。或者，可以在抗体类药剂之后以适当间隔添加该酶，这样抗体类药剂已经产生其作用。
施用使血清IgG分子失活的酶和步骤(a)和(b)可以在相对于移植细胞、组织或器官到对象的不同的时间进行。例如，施用酶和步骤(a)和(b)可以在移植细胞、组织或器官之前进行。或者，施用酶和步骤(a)和(b)可以都在移植细胞、组织或器官之后进行。或者，施用酶可以在移植细胞、组织或器官之前进行，而步骤(a)和(b)在移植细胞、组织或器官之后进行。或者，施用酶和步骤(a)(如果存在的话)可以在移植细胞、组织或器官之前进行，而步骤(b)在移植细胞、组织或器官之后进行。典型的方法可以包括施用酶，随后施用细胞、组织或器官移植体(如肾脏移植体)，随后在施用酶之后以适当时间间隔施用ATG。对于HPSC的移植(如骨髓移植)，步骤的顺序通常可以是步骤(b)的抗体类药剂，随后是步骤(a)的受者HPSC耗竭，随后是使对象中血清IgG分子失活的酶，随后是移植体。
诱导造血干细胞嵌合体的方法
本发明提供了一种诱导对象中的造血嵌合体的方法，该方法包括实施本发明的调理方案，随后以足以在对象中诱导造血嵌合体的量和在适于在对象中诱导造血嵌合体的条件下将HSPC施用至对象。该方法也可以被描述为稳定移植HSPC的方法。HSPC可以是自体的(使用患者自己的细胞)或同基因的(细胞来自基因相同的双胞胎)，也可以是同种异体的(细胞来自单独的非同一供者)。
减少HSPC在受者中成功植入的可能性的免疫并发症对同种异体细胞最重要，因此本发明的方法对这种细胞最有利。然而，如果受者先前没有接触过的产物存在表达，即使是自体细胞也会发生免疫并发症。如果自体细胞经过基因改造以表达基因治疗，该细胞可能被充分改变而引起免疫反应。例如，可能存在针对表达的基因治疗产物的免疫反应。如果HSPC经基因改造而表达与受者HLA不匹配的不同HLA类型，也会出现类似的情况。因此，HSPC优选地是同种异体的，或者是基因改造的自体细胞或同基因细胞。HSPC最优选地是同种异体的。在特别优选的实施方式中，HSPC的供者也是待被移植到受者的另一器官或组织的供者。也就是说，同一供者提供HSPC和其他细胞、器官或组织二者。
HSPC存在于成年人的骨髓中，特别是在骨盆、股骨和胸骨。它们也存在于脐带血中，少量存在于外周血中。可以使用本领域建立的任何合适的技术从这些位置获取HSPC。
例如，HSPC可以从人的骨髓获取，通过用大针直接从供者的骨头中心抽出。髂骨后段是常见的获取部位。该技术被称为骨髓获取，可在局部或全身麻醉下进行。当施用的HSPC来自于供者的骨髓时，HSPC的施用可描述为骨髓移植(BMT)。
HSPC可以在婴儿出生后不久从脐带血获取。从脐部将脐带两端夹住，并在夹子之间进行处理(transact)。随后在无菌条件下穿刺脐带静脉，血液在重力作用下自由流进抗凝血的无菌封闭获取系统，由此可以将HSPC分离出来。
HSPC可以从外周血获取，通常通过抽血。然而，由于外周血中的HSPC数量通常很低，首先需要从骨髓调动HSPC。在健康的供者中，这可以通过施用粒细胞集落刺激因子(G-CSF)来实现。如果供者不健康，可能需要替代性策略。如果打算进行的HSPC移植是自体的，就可能经常出现这种情况。
HSPC优选地在获取后尽快使用(即新鲜的)，但可以进行低温保存，之后解冻以用于本发明的方法。低温保存通常包括通过去除红细胞和血浆来减少体积。可以量化获取的干细胞的量，例如通过对样本进行流式细胞术分析，以确定对CD34(干细胞的标志物)阳性的细胞比率。
可以通过任何合适的方法向对象施用HSPC。优选的方法是输注，通常通过中心管进行。在输注之前、期间和之后，患者可以保持在高度清洁或无菌的环境下，例如在正压下的具有高效粒子空气过滤器(HEPA)的房间里，以减少感染的风险。
可以监测方法以确定HSPC移植已成功产生造血嵌合体。这通过确定在施用HSPC后的特定的时间间隔(通常为28天)后从对象提取的血样中存在的供者来源的造血细胞的比率来实现。例如，如果发现样本中的至少5％的淋巴细胞和/或髓样细胞是供者来源的，优选地是发现样本中的至少5％的淋巴细胞是供者来源的，则可定义为实现造血嵌合体。如果发现样品中不超过90％的淋巴细胞和/或髓样细胞是供者来源的(即至少10％仍来自受者)，优选地如果发现样品中不超过90％的淋巴细胞是供者来源的(即至少10％的淋巴细胞仍来自受者)，则描述为混合型嵌合体。如果发现样品中的98％或更多的淋巴细胞和/或髓样细胞是供者来源的，那么描述为完全型嵌合体。对于本发明的方法，混合型嵌合体通常是优选的，因为受者将有更高的免疫力水平。然而，全嵌合体在某些情况下可以是有益的，例如在治疗癌症(例如白血病)时，其目标是消除有可能导致癌症复发的宿主细胞，用移植的HSPC取代它们。
样品中的供者来源的细胞和受者来源的细胞的比率可以通过本领域任何合适的方法确定，如实施例中描述的流式细胞术分析。也可以使用实时PCR。在Agrawal等人的BoneMarrow Transplantation 2004(34)p-12中讨论了其他方法。
治疗或预防疾病或病症的方法
本发明提供了一种预防或治疗对象中的疾病或病症的方法。该方法包括根据上述方法诱导对象中的造血嵌合体，以提高该疾病或病症的治疗对对象的益处，从而治疗或预防该疾病或病症。该疗法可以是细胞、组织或器官移植体，通常与HSPC来自同一供者。移植的细胞、组织或器官可以属于任何类型，包括肾脏、肝脏、心脏、胰腺、肺、小肠、皮肤、血管/血管组织、面部、手臂、气管、眼睛的部分、胰岛、黑质、骨髓。移植的细胞可以属于任何类型，包括用于本方法本身的相同的HSPC，如此不再需要额外的治疗。该疗法可以是使用基因改造的HPSC进行的基因疗法。
以另一种方式表述，本发明还提供了一种预防或治疗细胞、组织或器官移植体的免疫排斥的方法，该方法包括根据本发明的方法诱导对象中的造血嵌合体，并且向对象施用细胞、组织或器官移植体，任选地其中该细胞、组织或器官与HSPC来自同一供者。通常在诱导对象中的造血嵌合体之后施用该细胞、组织或器官，但也可以在诱导对象中的造血嵌合体之前施用该细胞、组织或器官。例如，如果器官取自已故供者，优选地首先进行器官移植，随后使用取自同一已故供者或密切匹配的供者的HSPC诱导造血嵌合体。细胞、组织或器官移植体可以属于任何类型，包括肾脏、肝脏、心脏、胰腺、肺、小肠、皮肤、血管/血管组织、面部、手臂、气管、眼睛的部分、胰岛、黑质、骨髓。移植的细胞可以具有任何类型，包括与用于诱导造血嵌合体的相同的HSPC，如此不再需要额外的移植体。
待被移植的细胞、组织或器官可以来自与受者不同的物种，即可以是异种移植。用于异种移植到人类受者的合适物种可以包括猪或非人灵长类动物。在这种情况下，可以对HSPC进行基因改造，以帮助提高对移植的耐受性。作为异种移植的细胞、组织或器官也可以进行基因改造。
待被治疗的对象可以优选地是敏化的或高度敏化的。"敏化"是指对象已经产生针对人类主要组织相容性(MHC)抗原(也被称为人白细胞抗原(HLA))的抗体。抗HLA抗体起源于同种异体的敏化B细胞，并且通常存在于先前因输血、既往移植或怀孕而敏化的患者。通过在T细胞和B细胞中产生特异性耐受，在敏化患者中实现造血嵌合体可以逆转异体敏化，从而减少供者特异性免疫反应，如DSA。
潜在的移植受者是否敏化可以通过任何合适的方法确定。例如，群体反应性抗体(PRA)测试可用于确定受者是否敏化。PRA得分>30％通常意味着患者是“高免疫风险”或“敏化”的。或者，可以进行交叉匹配测试，即将潜在移植供者的血液样本与预期受者的血液样本混合。交叉匹配阳性意味着受者具有与供者样本发生反应的抗体，表明受者是敏化的，不应进行移植。交叉匹配测试通常是将要移植前作为最后检查进行。
一种预防或治疗细胞、组织或器官移植体的免疫排斥反应的方法，包括：
(i)实施本发明的调理方案；
(ii)以足以在对象中诱导造血嵌合体的量和在适于在对象中诱导造血嵌合体的条件下将HSPC施用至对象。
该方法还可以任选地包括(iii)将细胞、组织或器官移植体施用至对象，该移植体通常与HSPC来自同一供者。在步骤(ii)中施用的HSPC本身可以是移植体，在这种情况下不再需要额外的步骤(iii)。该方法可被认为是对通过细胞、组织或器官移植体治疗的疾病或病症进行治疗的方法。例如，在HSPC本身就是移植体的情况下，该方法可用于预防或治疗通过HSPC移植体治疗的任何疾病或病症。
通常由HSPC移植体治疗的疾病或病症可以是获得性或先天性的。可以由HSPC移植体治疗的获得性疾病或病症包括：
-血液学恶性肿瘤，如白血病(例如，急性淋巴母细胞性白血病(ALL)、急性骨髓性白血病(AML)、慢性淋巴细胞性白血病(CLL)、慢性骨髓性白血病(CML))；淋巴瘤(例如，霍奇金病、非霍奇金淋巴瘤)和骨髓瘤(例如，多发性骨髓瘤(卡勒氏病))。
-实体瘤癌症，如神经母细胞瘤、促结缔组织增生性小圆细胞瘤、尤文氏肉瘤、绒毛膜癌。
-血液病，如吞噬细胞疾病(例如，骨髓发育不良)；贫血(例如，阵发性夜间血红蛋白尿症(PNH；严重再生障碍)、再生障碍性贫血、获得性纯红细胞再生障碍)；骨髓增生性疾病(例如，真性红细胞增多症、原发性血小板增多症、骨髓纤维化)。
-代谢性疾病，如淀粉样变性(例如，淀粉样轻链(AL)淀粉样变性)。
-环境诱导的疾病，如放射中毒。
-病毒性疾病，如人类嗜T淋巴细胞病毒(HTLV)或人类免疫缺陷病毒(HIV)。
-自身免疫性疾病，如多发性硬化症。
可以由HSPC移植体进行治疗的先天性疾病或病症包括：
-溶酶体贮积症，如脂质沉积-脂质贮积症-(例如，神经元蜡样质脂褐质沉积症，婴儿型神经元蜡样质脂褐质沉积症(INCL，桑塔沃里病(Santavuori disease))，詹-比二氏病(Jansky-Bielschowsky disease)(晚期婴儿型神经元蜡样质脂褐质沉积症))；神经鞘脂贮积症(例如，尼曼-匹克氏病(Niemann–Pick disease)、戈谢病(Gaucher disease))；脑白质营养不良(例如，肾上腺脑白质营养不良、肾上腺脑白质营养不良、克拉伯病(Krabbedisease)(球形细胞脑白质营养不良))；黏多糖贮积症(例如，赫尔勒综合征(Hurlersyndrome)(MPS I H，α-L-艾杜糖醛酸酶缺乏症)，施艾氏症(Scheie syndrome)(MPS I S)，Hurler-Scheie综合征(MPS I H-S)，Hunter综合征(MPS II，艾杜糖醛酸硫酸酶缺乏症)，沙费利波综合征(Sanfilippo syndrome)(MPS III)，莫基奥综合征(Morquio syndrome)(MPSIV)，马-拉综合征(Maroteaux–Lamy syndrome)(MPS VI)，Sly综合征(MPS VII)；糖蛋白病(例如，黏脂贮积病Ⅱ型(I型细胞病)、岩藻糖代谢病、天冬氨酰葡萄糖胺尿症、α-甘露糖苷病；或其他病(例如，沃尔曼病(酸性脂酶缺乏症))。
-免疫缺陷症，如T细胞缺陷症(例如，共济失调毛细血管扩张症，迪格奥尔格综合征(DiGeorge syndrome))；T细胞和B细胞联合缺陷症(例如，严重联合免疫缺陷病(SCID)，所有类型)；Wiskott-Aldrich综合征；吞噬细胞病(例如，Kostmann综合征、Shwachman-Diamond综合征)；免疫失调病(例如，格里切利综合征(Griscelli syndrome)，II型)；先天免疫缺陷症(例如，NF-κB基本调节蛋白(NEMO)缺乏症)。
-血液病，如血红蛋白病(例如，镰状细胞病、β地中海贫血症(库利氏贫血症))；贫血(例如，再生障碍性贫血症、戴-布二氏贫血症(Diamond–Blackfan anemia)、范可尼贫血症(Fanconi anemia))；血球减少症(例如，低巨核细胞性血小板减少症(Amegakaryocyticthrombocytopenia))；以及嗜血细胞性综合征(例如，嗜血细胞性淋巴组织细胞增生症(HLH))。
在将HSPC基因改造以进行基因治疗的情况下，本发明的方法可用于预防或治疗该基因治疗针对的疾病或病症。
本发明还提供了用于预防或治疗疾病或病症的方法中的酶，该酶使对象中的血清IgG分子失活，其中该方法如上所述。
本发明还提供了使对象中的血清IgG分子失活的酶在制备药物中的用途，其中该药物用于预防或治疗如上所述的方法中的疾病或病症。
酶
IgG半胱氨酸蛋白酶
用于本发明的IgG半胱氨酸蛋白酶对IgG具有特异性。在优选的实施方式中，用于本发明的方法的蛋白酶是IdeS(酿脓链球菌(S.pyogenes)的免疫球蛋白G-降解酶)，又称imlifidase。IdeS是由人病原体酿脓链球菌产生的细胞外半胱氨酸蛋白酶。IdeS最初是从血清型M1的A组链球菌菌株中分离出来的，但是现在已经在所有测试的A组链球菌菌株中鉴定出ides基因。IdeS具有极其高程度的底物特异性，其唯一识别的底物是IgG。IdeS催化人IgG所有亚类的重链下部铰链区的单一蛋白水解切割。IdeS还催化各种动物中一些IgG亚类的重链的等效切割。IdeS通过两阶段机制有效地将IgG切割为Fc和F(ab')2片段。在第一阶段，IgG的一条(第一)重链被切割以产生具有非共价结合的Fc分子的单切割的IgG(scIgG)分子。scIgG分子实际上是一种中间产物，它保留原始IgG分子的剩余(第二)重链。在该机制的第二阶段，该第二重链被IdeS切割以释放F(ab')2片段和同源二聚体Fc片段。这些是通常在生理条件下观察到的产物。在还原条件下，F(ab')2片段可以解离成两个Fab片段，并且同源二聚体Fc可以解离成其组成单体。IdeS已被证明在切割人的IgG上特别有效。在给药IdeS的几分钟内，整个血浆IgG库被切割，并且血液中IgG水平保持低水平超过一个星期，直到新合成的IgG出现在血浆中。这表明整个细胞外IgG库而不仅仅是血浆库(即，血清IgG分子)被IdeS切割(Winstedt等人；PloS One 2015；10(7)：e0132011)。
SEQ ID NO:1是包含N端甲硫氨酸和信号序列的IdeS的完整序列。它也可作为NCBI参考序列号WP_010922160.1获得。SEQ ID NO:2是缺少N端甲硫氨酸和信号序列的IdeS的成熟序列。它也可作为Genbank登录号ADF13949.1获得。
在替代的实施方式中，用于在本发明方法中使用的蛋白酶是IdeZ，它是由马链球菌兽疫亚种(Streptococcus equi ssp.Zooepidemicu，一种主要发现于马中的细菌)产生的IgG半胱氨酸蛋白酶。SEQ ID NO:3是包含N端甲硫氨酸和信号序列的IdeZ的完整序列。它也可作为NCBI参考序列号WP_014622780.1获得。SEQ ID NO:4是缺少N端甲硫氨酸和信号序列的IdeZ的成熟序列。
在替代的实施方式中，用于在本发明方法中使用的蛋白酶是杂合IdeS/Z，例如SEQID NO:5。N端基于缺少N端甲硫氨酸和信号序列的IdeZ。
在优选的实施方式中，用于在本发明中使用的蛋白酶可以包含SEQ ID NO:2、4或5或由SEQ ID NO:2、4或5组成。用于在本发明中使用的蛋白酶可以包含N端的附加甲硫氨酸(M)残基和/或C端的标签，以协助在标准细菌表达系统中表达和从标准细菌表达系统中分离。合适的标签包括组氨酸标签，其可以直接地连接到多肽的C端或通过任何合适的接头序列(例如3、4或5个甘氨酸残基)间接地连接。组氨酸标签通常由六个组氨酸残基组成，尽管它可以比这更长，通常多达7、8、9、10或20个氨基酸，或者更短，例如5、4、3、2或1个氨基酸。
在进一步优选的实施方式中，用于在本发明中使用的蛋白酶可以包含以下、基本上由以下组成或由以下组成：SEQ ID NO:6至25中任一个的序列。这些序列代表具有增加的蛋白酶活性和/或降低的免疫原性的IdeS和IdeZ多肽。SEQ ID NO:6至25中的每一个可以任选地包含N端的附加甲硫氨酸和/或C端的组氨酸标签。组氨酸标签优选地由六个组氨酸残基组成。组氨酸标签优选地通过3×甘氨酸或5×甘氨酸残基的接头连接到C端。
在进一步优选的实施方式中，用于在本发明中使用的蛋白酶可以包含以下、基本上由以下组成或由以下组成：SEQ ID NO:56至69中任一个的序列。这些序列代表具有增加的蛋白酶活性和/或降低的免疫原性的IdeS多肽。SEQ ID NO:56至69中的每一个可以任选地包含N端的附加甲硫氨酸和/或C端的组氨酸标签。组氨酸标签优选地由六个组氨酸残基组成。组氨酸标签优选地通过3×甘氨酸或5×甘氨酸残基的接头连接到C端。
在进一步优选的实施方式中，用于在本发明中使用的蛋白酶可以包含以下、基本上由以下组成或由以下组成：SEQ ID NO:6至25中任一个的序列，其任选地具有多达3个(例如1、2或3个)氨基酸取代。SEQ ID NO:6至25中的每一个及其变体可以任选地包含N端的附加甲硫氨酸和/或C端的组氨酸标签。
在进一步优选的实施方式中，用于在本发明中使用的蛋白酶可以包含以下、基本上由以下组成或由以下组成：SEQ ID NO:56至69中任一个的序列，其任选地具有多达3个(例如1、2或3个)氨基酸取代。SEQ ID NO:56至69中的每一个及其变体可以任选地包含N端的附加甲硫氨酸和/或C端的组氨酸标签。
本发明多肽的长度通常为至少100、150、200、250、260、270、280、290、300或310个氨基酸。本发明多肽的长度通常为不大于400、350、340、330、320或315个氨基酸。应当理解，上面列出的下限中的任一个都可以与上面列出的上限中的任一个组合以提供本发明多肽的长度范围。例如，多肽的长度可以为100至400个氨基酸、或250至350个氨基酸。该多肽的长度优选地为290至320个氨基酸，最优选地为300至315个氨基酸。
本发明蛋白酶的一级结构(氨基酸序列)是基于IdeS、IdeZ或IdeS/Z的一级结构，特别是分别为SEQ ID NO:2、4或5的氨基酸序列。本发明蛋白酶的序列可以包含SEQ ID NO:2、4或5的氨基酸序列的变体，该变体与SEQ ID NO:2、4或5的氨基酸序列至少80％相同。变体序列可以与SEQ ID NO:2、4或5的序列至少80％、至少85％，优选至少90％、至少95％、至少98％或至少99％相同。变体可以与SEQ ID NO:2、4或5的序列相同，除了包含在WO2016/128558或WO2016/128559中鉴定的一种或多种特定修饰之外。可以在SEQ ID NO:2、4或5所示的序列的至少50、至少100、至少200、至少300个或更多个连续氨基酸的区域上或更优选地在SEQ ID NO:4或5的全长上测量相对于SEQ ID NO:2、4或5序列的同一性。
用于本发明的蛋白酶可以是包含SEQ ID NO:2、4或5的氨基酸序列的变体的IdeS、IdeZ或IdeS/Z多肽，该变体中的修饰(例如，氨基酸添加、缺失或取代)是相对于SEQ ID NO:2、4或5的序列进行的。此类修饰优选地是保守氨基酸取代。保守取代利用化学结构相似、化学性质相似或侧链体积相似的其他氨基酸替换氨基酸。引入的氨基酸可以具有与其所替换的氨基酸相似的极性、亲水性、疏水性、碱性、酸性、中性或电荷。或者，保守取代可以引入芳香族或脂肪族的另一种氨基酸来代替先前存在的芳香族或脂肪族的氨基酸。保守氨基酸变化是本领域熟知的。
IgG半胱氨酸蛋白酶活性可以通过任何合适的方法来评估，例如通过将多肽与含有IgG的样品孵育并且确定IgG切割产物的存在。WO2016/128559中描述了合适的方法。合适的测定包括基于ELISA的测定，例如在WO2016/128559中描述的那些。在这样的测定中，测定板的孔通常用抗体靶标(例如，牛血清白蛋白(BSA))包被。然后将待测试的多肽样品添加到孔中，随后添加靶标特异性抗体的样品，在该实例中该抗体是对BSA特异的。允许多肽和抗体在适合IgG半胱氨酸蛋白酶活性的条件下相互作用。在合适的时间间隔后，洗涤测定板，并且在适合与靶标特异性抗体结合的条件下添加与靶标特异性抗体特异性结合的检测抗体。检测抗体将与每个孔中已与靶标结合的任何完整靶标特异性抗体结合。洗涤后，孔中存在的检测抗体的量将与结合到该孔的靶标特异性抗体的量成比例。检测抗体可以直接地或间接地与标记或另一种报告系统(例如酶)缀合，从而可以确定留在每个孔中的检测抗体的量。孔中的测试多肽的效力越高，完整靶标特异性抗体将保留得越少，并且因此将存在越少的检测抗体。通常，给定的测定板上的至少一个孔将包含IdeS，而不是待测试的多肽，因此可以将测试多肽的效力直接地与IdeS的效力进行比较。也可以包括IdeZ和IdeS/Z用于比较。
其他测定可以通过直接地可视化和/或量化由测试多肽切割IgG产生的IgG片段来确定测试多肽的效力。这种类型的测定也在WO2016/128559中描述。这种测定通常将IgG样品与滴定系列下不同浓度的测试多肽(或与作为对照的IdeS、IdeZ和IdeS/Z中的一种或多种)孵育。然后使用凝胶电泳，例如通过SDS-PAGE分离在每个浓度下孵育所产生的产物。然后完整IgG和由切割IgG产生的片段可以通过大小进行鉴定并且通过使用合适染料的染色强度进行量化。在给定浓度下，切割片段的量越多，测试多肽的效力越大。本发明的多肽通常在比IdeZ和/或IdeS更低的浓度(滴定系列中的较低点)下会产生可检测量的切割片段。这种类型的测定还可以鉴定测试多肽在切割IgG分子的第一重链还是第二重链上更有效，因为也可以确定由每个切割事件产生的不同片段的量。本发明的多肽可以在切割IgG分子的第一链上比切割第二链上更有效，特别是在IgG是IgG2同种型时。本发明的多肽可以在切割IgG1上比切割IgG2上更有效。
IgG内切糖苷酶
该酶可以具有IgG内切糖苷酶的活性，优选地切割IgG的Fc区域上Asn-297(Kabat编号)的聚糖部分。这种蛋白质的实例是EndoS(酿脓链球菌的内切糖苷酶)。EndoS水解正常糖基化IgG的天冬酰胺链接的聚糖的β-1,4-二-N-乙酰壳二糖核心。EndoS的成熟序列被提供为SEQ ID NO:90。该药剂可以是包含SEQ ID NO:90的氨基酸序列或由SEQ ID NO:90的氨基酸序列组成的蛋白质，也可以是其同源物，该同源物来自替代性细菌(例如，马链球菌(Streptococcus equi)或兽疫链球菌(Streptococcus zooepidemicus)，或假结核棒杆菌(Corynebacterium pseudotuberculosis)，粪肠球菌(Enterococcus faecalis)，或脑膜脓毒性伊丽莎白金菌Elizabethkingia meningoseptica)。该药剂可以是CP40、EndoE或EndoF2。
或者，该药剂可以是EndoS蛋白的变体，该变体包含与SEQ ID NO：90至少80％、85％、90％或95％相同的任何氨基酸序列或由与SEQ ID NO：90至少80％、85％、90％或95％相同的任何氨基酸序列组成并且具有IgG内切糖苷酶活性。EndoS蛋白的变体可以包含以下或由以下组成：相对于SEQ ID NO:90的氨基酸序列已经具有多达1、2、3、4、5、10、20、30、40、50、60、70、80、90个或更多个氨基酸取代、插入或缺失的氨基酸序列，前提是该变体具有IgG内切糖苷酶活性。该氨基酸取代优选地是保守的。保守取代如前文所定义。
或者，该药剂可以是包含SEQ ID NO：90的片段或由SEQ ID NO：90的片段组成并且具有IgG内切糖苷酶活性的蛋白质，优选地其中该片段的长度为400至950个、500至950个、600至950个、700至950个或800至950个氨基酸。优选的片段由SEQ ID NO：90的第1至409个氨基酸组成，该片段对应于由链球菌半胱氨酸蛋白酶SpeB切割产生的具有酶活性的EndoSα结构域。该片段可以通过使SEQ ID NO:90的氨基酸序列的一个或多个氨基酸残基缺失而产生。可以使多达1、2、3、4、5、10、20、30、40、50、100、200、300、400、500或550个或更多个残基缺失。缺失的残基可以与其他的相连。
SEQ ID NO:90的任何片段或变体优选地含有SEQ ID NO:90的残基191至199，即SEQ ID NO:90的Leu-191、Asp-192、Gly-193、Leu-194、Asp-195、Val-196、Asp-197、Val-198和Glu-199。这些氨基酸构成了完整的几丁质酶家族18的活性位点，以谷氨酸结束。几丁质酶活性位点中的谷氨酸对酶的活性至关重要。因此，最优选的是，SEQ ID NO:90的变体含有SEQ ID NO:90的Glu-199。SEQ ID NO:90的变体可以含有具有一个或多个保守取代的SEQID NO:90的残基191至199，前提是变体含有SEQ ID NO:90的Glu-199。
多肽的产生
本发明方法中使用的酶是多肽并且可以通过任何合适的方式产生。例如，可以使用本领域已知的标准技术直接地合成多肽，例如Fmoc固相化学、Boc固相化学或通过液相肽合成。或者，可以通过用编码该多肽的核酸分子或载体(vector)转化细胞、通常是细菌细胞来产生多肽。WO2016/128558和WO2016/128559中描述并例证了通过在细菌宿主细胞中表达来产生酶多肽。
包含多肽的组合物和制剂
本发明还提供了用于本发明的方法的包含酶的组合物。例如，本发明提供了一种组合物，该组合物包含一种或多种多肽和至少一种药学上可接受的运载体(carrier)或稀释剂。运载体需要在与组合物的其他成分相容且对被施用组合物的对象无害的意义上是“可接受的”。通常，运载体和最终组合物是无菌且无热原的。
合适组合物的配制可以使用标准的药物制剂化学品和方法进行，所有这些对于相当熟练的技术人员来说都是易于获得的。例如，酶可以与一种或多种药学上可接受的赋形剂或溶媒组合。辅助物质(例如，润湿剂或乳化剂)、pH缓冲物质、还原剂等可以存在于赋形剂或溶媒中。合适的还原剂包括半胱氨酸、硫代甘油、硫氧还原蛋白、谷胱甘肽等。赋形剂、溶媒和辅助物质通常是不会在接受组合物的个体中诱导免疫反应并且可以施用而没有过度毒性的药学药剂。药学上可接受的赋形剂包括但不限于液体如水、盐水、聚乙二醇、透明质酸、甘油、硫代甘油和乙醇。其中还可包含药学上可接受的盐，例如无机酸盐，例如盐酸盐、氢溴酸盐、磷酸盐、硫酸盐等；以及有机酸盐，例如乙酸盐、丙酸盐、丙二酸盐、苯甲酸盐等。药学上可接受的赋形剂、溶媒和辅助物质的深入讨论可在《雷氏药学大全》(Remington's Pharmaceutical Sciences，Mack Pub.Co.,NJ 1991)中获得。
此类组合物可以以适合于单次施用(bolus administration)或持续施用的形式制备、包装或出售。可注射的组合物可以以单位剂型(例如，于含防腐剂的安瓿或于多剂量容器中)制备、包装或出售。组合物包含但不限于混悬剂、溶液剂、油性或水性溶媒中的乳剂、糊剂和可植入的缓释或可生物降解的制剂。此类组合物还可包含一种或多种附加成分，包括但不限于混悬剂、稳定剂或分散剂。在用于肠胃外施用的组合物的一个实施方式中，活性成分以干燥(例如，粉末或颗粒)形式提供，用于用合适的溶媒(例如，无菌无热原水)重构，然后在肠胃外施用重构的组合物。组合物可以以无菌可注射的水性或油性混悬剂或溶液剂的形式制备、包装或出售。该混悬剂或溶液剂可以根据已知技术进行配制，并且可以包含除活性成分外的附加成分，例如本文所述的分散剂、润湿剂或助悬剂。此类无菌可注射制剂可以使用无毒的肠胃外可接受的稀释剂或溶剂(例如，水或1,3-丁二醇)来制备。其它可接受的稀释剂和溶剂包括但不限于林格氏溶液、等渗氯化钠溶液和不挥发油(例如，合成的甘油单酯或甘油二酯)。
其他可用的肠胃外施用的组合物包括这样的组合物，这些组合物包含微晶形式的、在脂质体制剂中的或作为生物可降解聚合物系统的组分的活性成分。用于持续释放或植入的组合物可以包含药学上可接受的聚合物或疏水材料，例如乳液、离子交换树脂、微溶聚合物或微溶盐。组合物可以适合于通过任何合适途径施用，包括例如皮内、皮下、经皮、肌内、动脉内、腹膜内、关节内、骨内或其他适当的施用途径。优选的组合物适合于通过静脉内输注施用。
试剂盒
本发明还提供了一种用于实施本文所述方法的试剂盒。本发明的试剂盒可以包括如上所述的酶或包含酶的组合物。该试剂盒可以包括用于向对象施用该酶或组合物的装置。该试剂盒可以包括本文所述的任何方法中各种组分的使用说明书。
实施例
除非另有说明，否则所使用的方法是标准的生物化学和分子生物学技术。合适的方法教科书的实例包括Sambrook等人，Molecular Cloning，A Laboratory Manual(1989)和Ausubel等人，Current Protocols in Molecular Biology(1995),John Wiley和Sons,Inc。
实施例1
介绍
Imlifidase可切割所有的人类IgG亚型，但只能切割小鼠IgG2c和IgG3，而不能切割小鼠IgG1和IgG2b。有趣的是，EndoS已被证明可以减少小鼠IgG1和IgG2b的补体介导功能和FcγR介导功能。然而，EndoS处理的小鼠IgG2a和IgG2c已被证明可以通过FcγR保持细胞溶解活性，但IgG2c也被证明会取决于条件而失去一些结合亲和力。因此，就以下实验中使用的动物模型而言，使用了imlifidase和EndoS的组合以确保对小鼠对象的血清IgG的影响最大。预计单独的imlifidase或EndoS(或特异性/活性相当的另一种蛋白酶或内切糖苷酶)将足以在人类对象中达到相当的效果。
下面的实验使用敏化NOD受者的严格模型，该模型对放射和耐受性诱导有抵抗力。实验证明，包括imlifidase和EndoS的联合方法允许在这些小鼠中产生混合型造血嵌合体。
材料和方法
动物
成年NOD/ShiLtJ(H-2g7；称为NOD)、FVB/NJ(H-2q；称为FVB)、C57BL/6J(H-2b；称为B6.CD45.2)、B6.SJL-Ptprc a Pepcb./Boy(H-2b，称为B6.CD45.1)、B6.NOD-(D17Mit21-D17Mit10)(H-2g7；称为B6.H-2g7)、NOD.B10Sn-H2b/J(H-2b；称为NOD.H-2b)小鼠购自杰克森实验室(Jackson Laboratory，美国缅因州巴尔港)，在阿尔伯塔大学的无特定病原体室中进行繁殖和饲养。对动物的所有护理和处理都是根据加拿大动物护理委员会的指导方针进行的。用于嵌合体诱导的所有NOD小鼠都是8至10周龄的雌性。
体内实验的试剂
Imlifidase和EndoS由汉莎生物制药股份有限公司(瑞典隆德)提供并经许可使用。抗CD4单抗(克隆Gk1.5，大鼠IgG2b)、抗CD90单抗(克隆YTS154，大鼠IgG2b)、抗CD8α单抗(克隆YTS169.4，大鼠IgG2b)和抗MHC-I H-2Kb mAb(克隆B8.24.3，小鼠IgG2b)是公司内部生产的。生产YTS 169.4抗小鼠CD8α单抗的细胞由H Waldmann教授和SP Cobbold博士(剑桥大学，病理学系)开发，并通过剑桥企业有限公司(Cambridge Enterprise Limited，HauserForum,3Charles Babbage Road,Cambridge CB3 0GT)获得。环磷酰胺(29875)和硼替佐米(A2614)分别购自Sigma(美国密苏里州)和ApexBio(美国得克萨斯州)。
体内EndoS介导的单克隆DSA抑制测定
用溶媒、单独的抗MHC-I H-2Kb(10μg)、或EndoS(30μg)与抗MHC-I H-2Kb(10μg或100μg)的混合物静脉内注射NOD或B6.H-2g7小鼠，作为预处理。EndoS和抗H-2Kb在即将注射前混合。预处理4小时后，将500万个细胞的羧基荧光素琥珀酰亚胺酯(CFSE)标记的NOD和CellTrace Violet染料(CTV)标记的B6骨髓细胞(BMC)的1:1混合物静脉内注射到预处理的NOD小鼠中。同样，将CFSE标记的B6.H-2g7和CTV标记的NOD.H-2b BMC的1:1混合物注射到预处理的B6.H-2g7小鼠体内。在施用细胞后1小时、2小时、3小时收集血液，并通过流式细胞仪进行分析。从每只小鼠的一个后肢收集脾细胞和BMC，并在BMC注射后4小时进行分析。
血清DSA检测测定
通过腹膜内注射20×106个FVB脾细胞使NOD小鼠敏化。收集敏化前和敏化后4至6周以及在imlifidase和EndoS处理后4小时的血清。用FcR阻断法(抗小鼠CD16/CD32大鼠IgG2b抗体，克隆2.4G2，BE0307，Bio X cell)处理FVB脾细胞(2×105个)5分钟，随后与100μL的滴定量血清孵育30分钟。清洗细胞两次，并与100μL的荧光染料缀合的二抗孵育30分钟。使用以下的二抗：FITC缀合的兔抗小鼠IgG Fc抗体的F(ab')2片段(1:200,315-096-046,Jackson ImmunoResearch)，APC缀合的山羊抗小鼠IgG1 Fc抗体(1:100,115-135-205,Jackson ImmunoResearch)，以及FITC缀合的山羊抗小鼠IgG3 Fc抗体(1:100,115-095-209,Jackson ImmunoResearch)。清洗细胞两次，并通过流式细胞术进行分析。使用含2％FBS的HBSS以清洗细胞和重构。
BMT方案和嵌合体的定义
为了确定供者BMC在敏化受者中的短期存活率，将已用B6.CD45.1脾细胞敏化的NOD小鼠进行T细胞耗竭(抗CD4，0.25mg；抗CD8，0.25mg；抗CD90 0.3mg；静脉内注射)，两天后进行BMT，以及静脉内注射EndoS和imlifidase且4小时后进行BMT(通过静脉内注射80×106个B6.CD45.2 BMC)。在BMC注射后4小时分析脾细胞和BMC。
为了长期诱导嵌合体，在基于BMT当日起往前算的第6天(即，第-6天)通过静脉内注射imlifidase和EndoS来处理用FVB脾细胞敏化的NOD小鼠。在BMT之前的第4天(即，第-4天)给予环磷酰胺(150mg/kg，腹膜内或静脉内)和硼替佐米(1mg/kg，静脉内)。在BMT之前的第2天(即，第-2天)、BMT的第2天、第6天、第11天和第16天静脉内注射T细胞耗竭性抗体。在第0天给予重复剂量的imlifidase和EndoS以及6Gy全身放射(TBI，Gammacell 1000Elite)，4小时后进行BMT。在第0天，通过尾静脉静脉内注射FVB骨髓细胞(80×106个)。在确定BMT进行之前环磷酰胺和硼替佐米对敏化受者的影响的实验中，给予较低剂量(20×106个)的骨髓细胞，以限制DSA对供者骨髓细胞的潜在吸附。在指定的时间点，收集外周血进行流式细胞术分析。对于长期的嵌合体，在BMT之后的第28天，如果淋巴细胞门中有至少5％的MHC-I+细胞是供者来源的，则认为受者产生嵌合体。
抗体和流式细胞术
针对小鼠H-2Kd(SF1-1.1.1)、H-2Kq(KH114)、H-2Kb(AF6-88.5)、CD45.2(104)、CD19(6D5)、CD138(281-2)、B220(RA3-6B2)、TCRβ(H57-597)、CD4(RM4-5或RM4-4)、CD8β(H35-17.2)、CD11b(M1/70)、CD11c(N418)、CD49b(DX5)、CD122(TM-β1)的荧光染料标记的抗体均购自BD Pharmingen(美国加州)、BioLegend(美国加州)或Thermo Fisher Scientific(美国加州)。使用LSR II(Becton Dickson，美国加州)的流式细胞术进行数据获取，并使用FlowJo(Treestar软件，美国俄勒冈州)进行数据分析。
统计分析
在适当的地方使用了曼-惠特尼U检验(Mann-Whitney U test)、比率配对t检验、具有霍尔姆-西达克(Holm-Sidak)多重比较检验的单因素方差分析和费舍尔(Fisher)精确检验，如所说明的。所有的统计分析都是用Prism(GraphPad软件，美国加州)完成的。
结果
EndoS抑制单克隆DSA介导的对供者BMC的杀伤
进行DSA被动转移实验以评估EndoS对于抑制抗体介导的对供者BMC的杀伤作用的影响。在所有DSA中，抗供者的MHC或HLA抗体在临床上最为重要。因此，幼稚(naive)NOD小鼠被注射小鼠IgG2b抗体(靶向表达MHC-I Kb的细胞)，经EndoS处理或未经EndoS处理，随后从B6小鼠进行骨髓转移。
如图1的A和B所示，在给予10μg抗Kb mAb单剂量的NOD受者中，与未接受酶处理的小鼠相比，在经EndoS处理的小鼠中在BMT后一小时血液中B6细胞与NOD细胞的比率显著增加。两组之间的血液中B6细胞与NOD细胞比率的这种差异在BMT后两小时和三小时保持稳定。同样，与用单独的100μg抗Kb mAb处理相比，给予100μg抗Kb mAb和EndoS的小鼠的血液中B6细胞与NOD细胞比率在1小时和2小时时增加。然而，该比率的增加持续不超过三小时，这表明剩余单抗的效应子功能随时间而积累。在BMT后四小时，与只接受10μg抗Kb mAb的小鼠相比，用EndoS和10μg抗Kb mAb处理的小鼠的BM和脾脏中的B6细胞与NOD细胞比率也观察到显著增加。
值得注意的是，NOD小鼠缺乏溶血性补体C5，该补体C5是补体依赖性细胞毒性必需的并且与MHC基因没有遗传联系。因此，与补体充足的宿主相比，NOD小鼠中DSA的效果可能会下降。因此，还研究了补体充足宿主中EndoS对DSA的作用。将NOD MHC同源的B6.H-2g7小鼠用作受者。与NOD宿主相比，EndoS以相似程度改善B6.H-2g7小鼠中供者细胞与受者细胞的比率(图1的C和D)。
简言之，无论受者是否补体充足，在抗MHC抗体的存在下，EndoS提高了供者细胞的存活率，这表明至少在这个模型系统中，对其他耗竭机制(例如，FcgR介导的)产生效果。
EndoS提高预敏化受者中供者BMC的存活率
接下来，研究了EndoS是否能够提高异体敏化的受者中供者BMC的存活率，这些异体敏化的受者具有多样化的针对供者抗原的抗体库。将EndoS与Imlifidase结合使用以测试这一点。Imlifidase切割鼠类IgG2c和IgG3，但不能切割鼠类IgG1和IgG2b。因此，共同使用EndoS来减弱未被imlifidase切割的鼠类IgG同种型的效应子功能。如图2A所示，imlifidase和EndoS共同导致用FVB脾细胞敏化的NOD小鼠中的DSA-IgG显著减少。供者细胞的IgG靶向性的下降可能是由于imlifidase，而不是EndoS，因为去糖基化仍然允许Fc特异性检测抗体的结合。鼠类IgG同种型的敏感性差异通过如下示出：DSA-IgG3减少约80％(IgG3是被imlifidase切割的亚型)(图2C)，而DSA-IgG1的水平没有变化(图2B)。虽然imlifidase对IgG3的降解仅导致完整IgG适度减少，但EndoS可以对imlifidase抗性IgG分子产生脱糖作用以进一步促进DSA-IgG效应子功能的减少。这两种酶的组合允许分析具有多克隆DSA的敏化受者中供者细胞的存活率。
除DSA之外，致敏(primed)的供者抗原特异性细胞毒性T细胞可以促进快速地杀死供者BMC。因此，将用同源性B6.CD45.1脾细胞敏化的CD45.1 NOD受者进行T细胞耗竭，两天后进行imlifidase和EndoS处理，以避免由敏化T细胞介导的急性细胞毒作用(图2D)。在给予T细胞耗竭性单抗两天后，受者的外周血中超过95％的T细胞被耗竭(数据未示出)。在本文中，将CD45.1/2系统用来协助鉴定存活的供者BMC，其中MHC染色可能被DSA干扰。如图2的E和F所示，在给予溶媒对照(BM 0.22％和脾脏0.27％)或仅仅给予imlifidase(BM 0.15％和脾脏0.46％)时，BMT后4小时在敏化NOD小鼠中B6.CD45.2供者细胞几乎完全被消除。相比之下，用EndoS和imlifidase处理的敏化NOD小鼠中，接近0.5％的BMC和约1.5％的脾细胞来自B6.CD45.2供者。因此，与用单独的溶媒或imlifidase处理的敏化受者相比，在施用imlifidase和EndoS四小时后进行BMT拯救了异体敏化受者中相当一部分的供者BMC(图2F)。有趣的是，在用imlifidase和EndoS处理的受者中的大多数残留供者细胞示出较低的MHC-I Kb染色，这表明供者MHC表位被去糖基化DSA或DSA的F(ab')2阻断(图2E)。另外，存活的供者细胞可能是那些表达较少MHC I类的细胞。
总之，这些数据表明imlifidase和EndoS的组合提高异体敏化受者中供者BMC的存活率。换言之，基本上所有血清IgG失活提高异体敏化受者中供者BMS的存活率。
硼替佐米和环磷酰胺处理后进行BMT减少BM中的B细胞
除了将imlifidase和/或EndoS用于BMT外，能减少产DSA的细胞的方法也可以为BMT后供者细胞的持续存活和进一步发展提供更长的低DSA环境的窗口。为了减少现有的浆细胞和可以在BMT后分化为浆细胞的B细胞，在对敏化小鼠进行BMT之前，采用硼替佐米来耗竭产生抗体的细胞，并采用环磷酰胺来减少B细胞(图3A)。硼替佐米和环磷酰胺(CyBor)的组合已被用于不符合移植条件的多发性骨髓瘤患者以及用于预防同种异体BMT后的移植体抗宿主病(GVHD)，但鲜有用于DSA脱敏的目的。
在BMT后五天，BM中BMC的细胞性在各组之间没有差异。有趣的是，用CyBor预处理小鼠的组中脾细胞的总数目增加。然而，与溶媒组相比，用CyBor处理的小鼠中的BM CD19+B细胞、CD19-CD138+B220+浆细胞和CD19-CD138+B220-浆细胞显著减少(图3B)。与BM中B细胞的减少相反，在检查时CyBor处理组中CD19+B脾细胞的减少并不显著。此外，CyBor处理小鼠的脾脏中CD19-CD138+B220+浆母细胞和CD19-CD138+B220-浆细胞显著增加(图3C)。
随后检查CyBor处理是否能预防由BMC注射刺激形成的DSA增加。如图3D所示，对照组的5只小鼠中有2只小鼠以及CyBor处理组的5只小鼠中有2只小鼠的DSA水平大幅增加，这表明CyBor不能降低DSA水平。然而，当比较BMT后5天(CyBor后9天)DSA水平的百分数变化时，用CyBor处理的小鼠中DSA增加趋于更少，这表明在进行BMT前进行CyBor处理抑制BMC注射刺激的DSA增加(图3E)。
总之，这些数据说明，CyBor减少BM中B细胞数量的效果很明显，CyBor可以限制由BMC注射引起的DSA增加。
用Imlifidase、EndoS、T细胞耗竭和CyBor的组合可在预敏化受者中实现植入
根据上述数据，假设的是将imlifidase和EndoS与T细胞耗竭抗体和CyBor骨髓浆细胞耗竭、连同非致命剂量的放射和较大剂量的BMC组合在一起，能够允许供者细胞在预敏化受者中植入。我们探讨了这种方案是否能诱导NOD小鼠以及B6.H-2g7小鼠(与NOD的MHC匹配，但不耐受嵌合体诱导)中的嵌合体。受者小鼠用FVB细胞敏化，四周后进行嵌合体诱导。对幼稚受者和致敏受者给予相同的调理方案，如方法部分和图4A所示。
正如所预期的，虽然所有的幼稚小鼠在BMT后四周几乎完全与FVB细胞嵌合，但在未经imlifidase和EndoS处理的致敏小鼠中，供者细胞被排斥。如图4B所示，在未接受酶处理的敏化NOD小鼠中，甚至在BMT后两天便检测不到供者细胞。相反，在接受酶处理的7只敏化NOD受者中，5只受者中的供者细胞在BMT后第4天或第9天超过5％。此外，在接受酶处理的四只敏化NOD小鼠中，嵌合体水平在BMT后第16天稳步上升至50％以上。最终，八只预敏化的NOD和B6.H-2g7小鼠中有5只小鼠在BMT后4周与供者细胞嵌合，其中2只预敏化的NOD小鼠具有稳定的混合型嵌合体，外周血中有多个谱系的供者细胞(表1和图4C)。在任何嵌合体中都观察不到GVHD的迹象。在试图不使用环磷酰胺或硼替佐米来简化该方案时，似乎这两者对诱导敏化受者中嵌合体的当前方案的成功至关重要(表1)。
总之，当与CyBor和标准调理剂组合起来时，imlifidase和EndoS的组合(即，使基本上所有的血清IgG失活)使供者BMC能够在预敏化受者小鼠中植入。
表1 EndoS-imlifidase允许预敏化受者具有造血嵌合体
Figure BDA0003925505260000191
嵌合体诱导方案的细节参见图4的图例。
Figure BDA0003925505260000201
代表B6.H-2g7受者(n＝2)和NOD受者(n＝4)。
Figure BDA0003925505260000202
代表B6.H-2g7受者(n＝2)和NOD受者(n＝5)。※代表一个B6.H-2g7受者(n＝1)和NOD受者(n＝7)。¤代表NOD受者。#示出的是BMT后四周的嵌合体水平。*通过双侧费舍尔精确检验，在与“CyBor”致敏组相比时p<0.05。
讨论
DSA是在敏化受者中进行同种异体骨髓移植的主要障碍。先前的工作表明，imlifidase可用于消除或减少DSA，EndoS可以在各种模型中抑制IgG介导的细胞毒性，但这两种酶都没有在HSPC移植/骨髓移植中使用，因为骨髓来源细胞上MHC的高表达可能增加对剩余功能性DSA的敏感性。
最近在敏化受者中肾脏移植的临床试验的先前结果与这些实验一起表明，imlifidase确实可以用于调理人类患者来接受HSPC移植/骨髓移植。目前的研究还表明，EndoS可以在这种情况下使用。发现了，在体内存在DSA下，单独的EndoS提高了供者细胞的存活率。考虑到如Maria Allhorn和Mattias Collin所报告的经EndoS处理的IgG降低了固定补体的能力，我们就EndoS以相似程度提高供者细胞在B6.H-2g7和NOD中的存活率的发现表明其他机制(如FcgR)是该BMT模型中DSA致病性的主要调理因子。给予低剂量或高剂量的单克隆DSA和EndoS在NOD和B6.H-2g7小鼠之间的差异表明，非MHC基因可以在不同个体中对EndoS的效果产生影响。NOD和B6.H-2g7之间的这种差异可能是由于NOD和B6背景的小鼠中IgG2b与各种Fc受体的结合能力不同。FcR多态性可能同样重要。结果还表明，EndoS对较低滴度DSA的作用更强。
发现了，imlifidase和EndoS的组合提高了供者BMC的存活率，并允许在已经用T细胞耗竭、CyBor和低于致死的放射调理的敏化小鼠中产生供者嵌合体。在测试的方案中，外周血中的T细胞耗竭的效果不受EndoS的影响。这表明，通过选择适当设计的时机，对血清IgG的酶耗竭可以与抗体类产品(如IVIG)和B细胞耗竭抗体(如利妥昔单抗)一起使用。换言之，可以用酶来失活DSA而不会不利地影响IgG类生物制剂的效应子功能，前提是仔细选择各自的给药时机。
关于环磷酰胺和硼替佐米的使用，除了针对B细胞或浆细胞外，两者都具有免疫调节作用。例如，环磷酰胺可以通过减少记忆性T细胞来促进敏化受者中嵌合体的诱导。关于硼替佐米，该发现与已发表的数据一致，显示在硼替佐米处理后脾脏B细胞代偿性增加，进而导致体液补偿。然而，在目前的研究中，BMT后脾脏B细胞的这种增加是否伴随DSA的反弹仍未知。重要的是，该方案中采用的T细胞耗竭可以抑制幼稚B细胞和记忆B细胞的恢复和成熟，以及抑制DSA从头生成。
最后，需要考虑到本研究的发现存在一些局限性。尽管imlifidase可以切割所有的人类IgG亚类，但它只能切割小鼠IgG的两个亚类，IgM不受影响。虽然IgM DSA水平与IgG相比很低，但它们可以降低所观察到的嵌合体水平。在临床上，IgM DSA可以通过血浆透析去除。为了在小鼠中对DSA实现最大效果，有必要将EndoS和imlifidase结合起来。已有研究表明，imlifidase通过在体外切割膜结合的BCR来暂时性抑制记忆B细胞的激活，这可以促进嵌合体的成功。然而，imlifidase只能切割小鼠IgG2c和IgG3，所以在这个模型中imlifidase对小鼠IgG的作用并不完全(图2A)。用imlifidase作为唯一脱敏剂的方案将在人类中更有效，在人类中imlifidase完全去除所有细胞外IgG/使所有细胞外IgG失活，因此使IgG DSA库完全失活。因此，这些发现可能低估了这些酶在临床环境上的潜力。
第二点限制涉及嵌合体诱导方案的毒性。然而，目前的研究是对原理研究的证明，表明调节IgG Fc在战略上可以用于对敏化受者进行BMT。此外，EndoS或imlifidase可以与其他脱敏方法结合使用。目前，还不清楚酶介导的DSA阻断是否能预防抗体的反弹。可能在阻断DSA功能(即，IgG Fc的去糖基化)的同时保持一定水平的DSA，这可能具有较小的可能性引发反弹而不是完全去除DSA。这些实验采用了较短的时间范围来重复注射酶(两次注射间隔6天)，以避免因宿主产生抗酶抗体而导致活性降低。imlifidase在人类环境中的功效更大，这可以允许酶被分开给予(例如，先给予imlifidase，再给予EndoS)，从而减轻可能因抗酶抗体出现的任何担忧。
最后，可以得出结论，imlifidase和EndoS的组合(即，使基本上所有血清IgG酶促失活)与其他脱敏策略相结合，可用于诱导异体敏化受者小鼠中的供者嵌合体。
实施例2-imlifidase和抗体类疗法的最佳间隔时间
背景
Imlifidase(在欧盟附有条件地被批准用于肾脏移植脱敏)是一种半胱氨酸蛋白酶，可将人和兔IgG的所有亚型切割为F(ab')2片段和二聚Fc片段。兔抗胸腺细胞球蛋白(rATG)是一种被批准用于诱导肾脏移植的抗体耗竭性疗法(它能使循环T淋巴细胞大量减少)。如果抗体类治疗(如rATG)与imlifidase一起给予，其可能会失活。本研究的目的是调查开始rATG治疗的最早时间点，同时以免影响其余imlifidase的大部分切割活性。
方法
用健康对象(n＝11)的血清来研究rATG的切割模式，该血清用0.25mg/kg的imlifidase(EudraCT编号：2019-002770-31)处理。将血清样本与50μg/mL(固定的且临床相关的浓度)的rATG(通常在给予1.5mg/kg的剂量后才观察到)在37℃下孵育2小时。收集从imlifidase处理前至imlifidase处理后14天的血清样本，使用SDS-PAGE和蛋白质印迹进行分析，用山羊抗兔IgG、F(ab')2特异性抗体显影，以评估rATG的切割。基于MSD技术，使用经验证的电致发光免疫测定来分析Imlifidase的浓度。
结果
对象中imlifidase血清浓度迅速下降，在96小时时的平均浓度为0.5μg/mL，但个体差异很大，<0.1μg/mL至1.8μg/mL(图5)。在该时间点上，11名对象中有8名对象的imlifidase活性水平已经下降到足以避免rATG被完全切割(图6)。
结论
考虑到有些患者中第一次施用的rATG的一部分可能会被切割，最早可在imlifidase后4天开始rATG。然而，由于rATG的剂量很高并且在几天内重复施用，预计在治疗开始时的这种切割不会对rATG疗效产生不利的整体影响。
实施例3-混合型嵌合体方案的特异性增强和毒性降低
将对实施例1中提出的混合型嵌合体方案进行逐步改变，旨在提高该方法的特异性并减少潜在的毒性，从而实现更大的临床转化潜力。具体为：
(i)将施用供者CD8-α细胞输注以增加敏化受者中稳定嵌合体的频率。供者T细胞输注可以通过减少宿主T细胞的存活率来促进BMT植入。
(ii)将与经酶(IdeS和/或EndoS)消除DSA和最大限度地耗竭T细胞和NK细胞一起，施用抗CD117/抗CD47。这将允许对预敏化受者进行首次非放射、非骨髓清除的嵌合体方案。抗CD117/抗CD47抗体有助于耗竭宿主的HSC。
序列表
<110> 汉莎生物制药股份有限公司
<120> 用于移植的调理方案
<130> N418790WO
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His Gln Ile Thr Ser Val Trp Thr Lys Gly Val Thr Pro Leu Thr Pro
20 25 30
Glu Gln Phe Thr Gln Gly Glu Asp Val Ile His Ala Pro Tyr Leu Ala
35 40 45
His Gln Gly Trp Tyr Asp Ile Thr Lys Ala Phe Asp Gly Lys Asp Asn
50 55 60
Leu Leu Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe
65 70 75 80
Asp Gln Asn Lys Thr Glu Ile Glu Ala Tyr Leu Ser Lys His Pro Glu
85 90 95
Lys Gln Lys Ile Ile Phe Asn Asn Gln Glu Leu Phe Asp Leu Lys Ala
100 105 110
Ala Ile Asp Thr Lys Asp Ser Gln Thr Asn Ser Gln Leu Phe Asn Tyr
115 120 125
Phe Arg Asp Lys Ala Phe Pro Asn Leu Ser Ala Arg Gln Leu Gly Val
130 135 140
Met Pro Asp Leu Val Leu Asp Met Phe Ile Asn Gly Tyr Tyr Leu Asn
145 150 155 160
Val Phe Lys Thr Gln Ser Thr Asp Val Asn Arg Pro Tyr Gln Asp Lys
165 170 175
Asp Lys Arg Gly Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asp Gln
180 185 190
Thr Thr Leu Leu Thr Ala Arg His Asp Leu Lys Asn Lys Gly Leu Asn
195 200 205
Asp Ile Ser Thr Ile Ile Lys Gln Glu Leu Thr Glu Gly Arg Ala Leu
210 215 220
Ala Leu Ser His Thr Tyr Ala Asn Val Ser Ile Ser His Val Ile Asn
225 230 235 240
Leu Trp Gly Ala Asp Phe Asn Ala Glu Gly Asn Leu Glu Ala Ile Tyr
245 250 255
Val Thr Asp Ser Asp Ala Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe
260 265 270
Val Gly Ile Asn Ala His Gly His Val Ala Ile Ser Ala Lys Lys Ile
275 280 285
Glu Gly Glu Asn Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser
290 295 300
Ser Gly Lys Asp Ile Trp Gln Lys Leu Ser
305 310
<210> 11
<211> 312
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 11
Asp Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val Pro
1 5 10 15
His Gln Ile Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Glu Gln
20 25 30
Phe Thr Gln Gly Glu Asp Val Ile His Ala Pro Tyr Leu Ala His Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Ala Phe Asp Gly Lys Asp Asn Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Thr Glu Ile Glu Ala Tyr Leu Ser Lys His Pro Glu Lys Gln
85 90 95
Lys Ile Ile Phe Asn Asn Gln Glu Leu Phe Asp Leu Lys Ala Ala Ile
100 105 110
Asp Thr Lys Asp Ser Gln Thr Asn Ser Gln Leu Phe Asn Tyr Phe Arg
115 120 125
Asp Lys Ala Phe Pro Asn Leu Ser Ala Arg Gln Leu Gly Val Met Pro
130 135 140
Asp Leu Val Leu Asp Met Phe Ile Asn Gly Tyr Tyr Leu Asn Val Phe
145 150 155 160
Lys Thr Gln Ser Thr Asp Val Asn Arg Pro Tyr Gln Asp Lys Asp Lys
165 170 175
Arg Gly Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asp Gln Thr Thr
180 185 190
Leu Leu Thr Ala Arg His Asp Leu Lys Asn Lys Gly Leu Asn Asp Ile
195 200 205
Ser Thr Ile Ile Lys Gln Glu Leu Thr Glu Gly Arg Ala Leu Ala Leu
210 215 220
Ser His Thr Tyr Ala Asn Val Ser Ile Ser His Val Ile Asn Leu Trp
225 230 235 240
Gly Ala Asp Phe Asn Ala Glu Gly Asn Leu Glu Ala Ile Tyr Val Thr
245 250 255
Asp Ser Asp Ala Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly
260 265 270
Ile Asn Ala His Gly His Val Ala Ile Ser Ala Lys Lys Ile Glu Gly
275 280 285
Glu Asn Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Ser Gly
290 295 300
Lys Asp Ile Trp Gln Lys Leu Ser
305 310
<210> 12
<211> 313
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 12
Asp Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val Pro
1 5 10 15
His Gln Ile Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Glu Gln
20 25 30
Phe Arg Tyr Asn Asn Glu Asp Val Ile His Ala Pro Tyr Leu Ala His
35 40 45
Gln Gly Trp Tyr Asp Ile Thr Lys Ala Phe Asp Gly Lys Asp Asn Leu
50 55 60
Leu Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp
65 70 75 80
Gln Asn Lys Thr Glu Ile Glu Ala Tyr Leu Ser Lys His Pro Glu Lys
85 90 95
Gln Lys Ile Ile Phe Asn Asn Gln Glu Leu Phe Asp Leu Lys Ala Ala
100 105 110
Ile Asp Thr Lys Asp Ser Gln Thr Asn Ser Gln Leu Phe Asn Tyr Phe
115 120 125
Arg Asp Lys Ala Phe Pro Asn Leu Ser Ala Arg Gln Leu Gly Val Met
130 135 140
Pro Asp Leu Val Leu Asp Met Phe Ile Asn Gly Tyr Tyr Leu Asn Val
145 150 155 160
Phe Lys Thr Gln Ser Thr Asp Val Asn Arg Pro Tyr Gln Asp Lys Asp
165 170 175
Lys Arg Gly Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asp Gln Thr
180 185 190
Thr Leu Leu Thr Ala Arg His Asp Leu Lys Asn Lys Gly Leu Asn Asp
195 200 205
Ile Ser Thr Ile Ile Lys Gln Glu Leu Thr Glu Gly Arg Ala Leu Ala
210 215 220
Leu Ser His Thr Tyr Ala Asn Val Ser Ile Ser His Val Ile Asn Leu
225 230 235 240
Trp Gly Ala Asp Phe Asn Ala Glu Gly Asn Leu Glu Ala Ile Tyr Val
245 250 255
Thr Asp Ser Asp Ala Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val
260 265 270
Gly Ile Asn Ala His Gly His Val Ala Ile Ser Ala Lys Lys Ile Glu
275 280 285
Gly Glu Asn Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Ser
290 295 300
Gly Lys Asp Ile Trp Gln Lys Leu Ser
305 310
<210> 13
<211> 312
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 13
Asp Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val Pro
1 5 10 15
His Gln Ile Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Glu Gln
20 25 30
Phe Thr Gln Gly Glu Asp Val Ile His Ala Pro Tyr Leu Ala His Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Ala Phe Asp Gly Lys Asp Asn Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Thr Glu Ile Glu Ala Tyr Leu Ser Lys His Pro Glu Lys Gln
85 90 95
Lys Ile Ile Ile Asn Asn Gln Glu Leu Phe Asp Leu Lys Ala Ala Ile
100 105 110
Asp Thr Lys Asp Ser Gln Thr Asn Ser Gln Leu Phe Asn Tyr Phe Arg
115 120 125
Asp Lys Ala Phe Pro Asn Leu Ser Ala Arg Gln Leu Gly Val Met Pro
130 135 140
Asp Leu Val Leu Asp Met Phe Ile Asn Gly Tyr Tyr Leu Asn Val Phe
145 150 155 160
Lys Thr Gln Ser Thr Asp Val Asn Arg Pro Tyr Gln Asp Lys Asp Lys
165 170 175
Arg Gly Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asp Gln Thr Thr
180 185 190
Leu Leu Thr Ala Arg His Asp Leu Lys Asn Lys Gly Leu Asn Asp Ile
195 200 205
Ser Thr Ile Ile Lys Gln Glu Leu Thr Glu Gly Arg Ala Leu Ala Leu
210 215 220
Ser His Thr Tyr Ala Asn Val Ser Ile Ser His Val Ile Asn Leu Trp
225 230 235 240
Gly Ala Asp Phe Asn Ala Glu Gly Asn Leu Glu Ala Ile Tyr Val Thr
245 250 255
Asp Ser Asp Ala Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly
260 265 270
Ile Asn Ala His Gly His Val Ala Ile Ser Ala Lys Lys Ile Glu Gly
275 280 285
Glu Asn Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Ser Gly
290 295 300
Lys Asp Ile Trp Gln Lys Leu Ser
305 310
<210> 14
<211> 312
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 14
Asp Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val Pro
1 5 10 15
His Gln Ile Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Glu Gln
20 25 30
Phe Thr Gln Gly Glu Asp Val Ile His Ala Pro Tyr Leu Ala His Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Ala Phe Asp Gly Lys Asp Asn Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Thr Glu Ile Glu Ala Tyr Leu Ser Lys His Pro Glu Lys Gln
85 90 95
Lys Ile Ile Phe Arg Asn Gln Glu Leu Phe Asp Leu Lys Ala Ala Ile
100 105 110
Asp Thr Lys Asp Ser Gln Thr Asn Ser Gln Leu Phe Asn Tyr Phe Arg
115 120 125
Asp Lys Ala Phe Pro Asn Leu Ser Ala Arg Gln Leu Gly Val Met Pro
130 135 140
Asp Leu Val Leu Asp Met Phe Ile Asn Gly Tyr Tyr Leu Asn Val Phe
145 150 155 160
Lys Thr Gln Ser Thr Asp Val Asn Arg Pro Tyr Gln Asp Lys Asp Lys
165 170 175
Arg Gly Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asp Gln Thr Thr
180 185 190
Leu Leu Thr Ala Arg His Asp Leu Lys Asn Lys Gly Leu Asn Asp Ile
195 200 205
Ser Thr Ile Ile Lys Gln Glu Leu Thr Glu Gly Arg Ala Leu Ala Leu
210 215 220
Ser His Thr Tyr Ala Asn Val Ser Ile Ser His Val Ile Asn Leu Trp
225 230 235 240
Gly Ala Asp Phe Asn Ala Glu Gly Asn Leu Glu Ala Ile Tyr Val Thr
245 250 255
Asp Ser Asp Ala Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly
260 265 270
Ile Asn Ala His Gly His Val Ala Ile Ser Ala Lys Lys Ile Glu Gly
275 280 285
Glu Asn Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Ser Gly
290 295 300
Lys Asp Ile Trp Gln Lys Leu Ser
305 310
<210> 15
<211> 312
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 15
Asp Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val Pro
1 5 10 15
His Gln Ile Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Glu Gln
20 25 30
Phe Thr Gln Gly Glu Asp Val Ile His Ala Pro Tyr Leu Ala His Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Ala Phe Asp Gly Lys Asp Asn Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Thr Glu Ile Glu Ala Tyr Leu Ser Lys His Pro Glu Lys Gln
85 90 95
Lys Ile Ile Ile Arg Asn Gln Glu Leu Phe Asp Leu Lys Ala Ala Ile
100 105 110
Asp Thr Lys Asp Ser Gln Thr Asn Ser Gln Leu Phe Asn Tyr Phe Arg
115 120 125
Asp Lys Ala Phe Pro Asn Leu Ser Ala Arg Gln Leu Gly Val Met Pro
130 135 140
Asp Leu Val Leu Asp Met Phe Ile Asn Gly Tyr Tyr Leu Asn Val Phe
145 150 155 160
Lys Thr Gln Ser Thr Asp Val Asn Arg Pro Tyr Gln Asp Lys Asp Lys
165 170 175
Arg Gly Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asp Gln Thr Thr
180 185 190
Leu Leu Thr Ala Arg His Asp Leu Lys Asn Lys Gly Leu Asn Asp Ile
195 200 205
Ser Thr Ile Ile Lys Gln Glu Leu Thr Glu Gly Arg Ala Leu Ala Leu
210 215 220
Ser His Thr Tyr Ala Asn Val Ser Ile Ser His Val Ile Asn Leu Trp
225 230 235 240
Gly Ala Asp Phe Asn Ala Glu Gly Asn Leu Glu Ala Ile Tyr Val Thr
245 250 255
Asp Ser Asp Ala Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly
260 265 270
Ile Asn Ala His Gly His Val Ala Ile Ser Ala Lys Lys Ile Glu Gly
275 280 285
Glu Asn Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Ser Gly
290 295 300
Lys Asp Ile Trp Gln Lys Leu Ser
305 310
<210> 16
<211> 312
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 16
Asp Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val Pro
1 5 10 15
His Gln Ile Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Glu Gln
20 25 30
Phe Thr Gln Gly Glu Asp Val Ile His Ala Pro Tyr Leu Ala Asn Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Ala Phe Asp Gly Lys Asp Asn Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Thr Glu Ile Glu Ala Tyr Leu Ser Lys His Pro Glu Lys Gln
85 90 95
Lys Ile Ile Phe Arg Asn Gln Glu Leu Phe Asp Leu Lys Glu Ala Ile
100 105 110
Arg Thr Lys Asp Ser Gln Thr Asn Ser Gln Leu Phe Glu Tyr Phe Arg
115 120 125
Asp Lys Ala Phe Pro Tyr Leu Ser Ala Arg Gln Leu Gly Val Met Pro
130 135 140
Asp Leu Val Leu Asp Met Phe Ile Asn Gly Tyr Tyr Leu Asn Val Phe
145 150 155 160
Lys Thr Gln Ser Thr Asp Val Lys Arg Pro Tyr Gln Asp Lys Asp Lys
165 170 175
Arg Gly Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Thr Thr
180 185 190
Leu Leu Thr Ala Arg His Asp Leu Lys Asn Lys Gly Leu Asn Asp Ile
195 200 205
Ser Thr Ile Ile Lys Glu Glu Leu Thr Lys Gly Arg Ala Leu Ala Leu
210 215 220
Ser His Thr Tyr Ala Asn Val Ser Ile Ser His Val Ile Asn Leu Trp
225 230 235 240
Gly Ala Asp Phe Asn Ala Glu Gly Asn Leu Glu Ala Ile Tyr Val Thr
245 250 255
Asp Ser Asp Ala Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly
260 265 270
Ile Asn Lys His Gly His Val Ala Ile Ser Ala Lys Lys Ile Glu Gly
275 280 285
Glu Asn Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Ser Gly
290 295 300
Lys Asp Ile Trp Gln Lys Leu Asn
305 310
<210> 17
<211> 292
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 17
Ser Val Trp Thr Lys Gly Val Thr Pro Pro Glu Gln Phe Thr Gln Gly
1 5 10 15
Glu Asp Val Ile His Ala Pro Tyr Leu Ala His Gln Gly Trp Tyr Asp
20 25 30
Ile Thr Lys Ala Phe Asp Gly Lys Asp Asn Leu Leu Cys Gly Ala Ala
35 40 45
Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln Asn Lys Thr Glu
50 55 60
Ile Glu Ala Tyr Leu Ser Lys His Pro Glu Lys Gln Lys Ile Ile Phe
65 70 75 80
Arg Asn Gln Glu Leu Phe Asp Leu Lys Ala Ala Ile Asp Thr Lys Asp
85 90 95
Ser Gln Thr Asn Ser Gln Leu Phe Asn Tyr Phe Arg Asp Lys Ala Phe
100 105 110
Pro Asn Leu Ser Ala Arg Gln Leu Gly Val Met Pro Asp Leu Val Leu
115 120 125
Asp Met Phe Ile Asn Gly Tyr Tyr Leu Asn Val Phe Lys Thr Gln Ser
130 135 140
Thr Asp Val Asn Arg Pro Tyr Gln Asp Lys Asp Lys Arg Gly Gly Ile
145 150 155 160
Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Thr Thr Leu Leu Thr Ala
165 170 175
Arg His Asp Leu Lys Asn Lys Gly Leu Asn Asp Ile Ser Thr Ile Ile
180 185 190
Lys Gln Glu Leu Thr Glu Gly Arg Ala Leu Ala Leu Ser His Thr Tyr
195 200 205
Ala Asn Val Ser Ile Ser His Val Ile Asn Leu Trp Gly Ala Asp Phe
210 215 220
Asn Ala Glu Gly Asn Leu Glu Ala Ile Tyr Val Thr Asp Ser Asp Ala
225 230 235 240
Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly Ile Asn Ala His
245 250 255
Gly His Val Ala Ile Ser Ala Lys Lys Ile Glu Gly Glu Asn Ile Gly
260 265 270
Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Ser Gly Lys Asp Ile Trp
275 280 285
Gln Lys Leu Ser
290
<210> 18
<211> 312
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 18
Asp Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val Pro
1 5 10 15
His Gln Ile Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Glu Gln
20 25 30
Phe Thr Gln Gly Glu Asp Val Ile His Ala Pro Tyr Leu Ala His Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Ala Phe Asp Gly Ala Asp Asn Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Thr Glu Ile Glu Ala Tyr Leu Ser Lys His Pro Glu Lys Gln
85 90 95
Lys Ile Ile Phe Arg Asn Gln Glu Leu Phe Asp Leu Lys Ala Ala Ile
100 105 110
Asp Thr Lys Asp Ser Gln Thr Asn Ser Gln Leu Phe Asn Tyr Phe Arg
115 120 125
Asp Lys Ala Phe Pro Asn Leu Ser Ala Arg Gln Leu Gly Val Met Pro
130 135 140
Asp Leu Val Leu Asp Met Phe Ile Asn Gly Tyr Tyr Leu Asn Val Phe
145 150 155 160
Lys Thr Gln Ser Thr Asp Val Asn Arg Pro Tyr Gln Asp Lys Asp Lys
165 170 175
Arg Gly Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Thr Thr
180 185 190
Leu Leu Thr Ala Arg His Asp Leu Lys Asn Lys Gly Leu Asn Asp Ile
195 200 205
Ser Thr Ile Ile Lys Gln Glu Leu Thr Glu Gly Arg Ala Leu Ala Leu
210 215 220
Ser His Thr Tyr Ala Asn Val Ser Ile Ser His Val Ile Asn Leu Trp
225 230 235 240
Gly Ala Asp Phe Asn Ala Glu Gly Asn Leu Glu Ala Ile Tyr Val Thr
245 250 255
Asp Ser Asp Ala Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly
260 265 270
Ile Asn Ala His Gly His Val Ala Ile Ser Ala Lys Lys Ile Glu Gly
275 280 285
Glu Asn Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Ser Gly
290 295 300
Lys Asp Ile Trp Gln Lys Leu Ser
305 310
<210> 19
<211> 292
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 19
Ser Val Trp Thr Lys Gly Val Thr Pro Pro Glu Gln Phe Thr Gln Gly
1 5 10 15
Glu Asp Val Ile His Ala Pro Tyr Leu Ala His Gln Gly Trp Tyr Asp
20 25 30
Ile Thr Lys Ala Phe Asp Gly Ala Asp Asn Leu Leu Cys Gly Ala Ala
35 40 45
Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln Asn Lys Thr Glu
50 55 60
Ile Glu Ala Tyr Leu Ser Lys His Pro Glu Lys Gln Lys Ile Ile Phe
65 70 75 80
Arg Asn Gln Glu Leu Phe Asp Leu Lys Ala Ala Ile Asp Thr Lys Asp
85 90 95
Ser Gln Thr Asn Ser Gln Leu Phe Asn Tyr Phe Arg Asp Lys Ala Phe
100 105 110
Pro Asn Leu Ser Ala Arg Gln Leu Gly Val Met Pro Asp Leu Val Leu
115 120 125
Asp Met Phe Ile Asn Gly Tyr Tyr Leu Asn Val Phe Lys Thr Gln Ser
130 135 140
Thr Asp Val Asn Arg Pro Tyr Gln Asp Lys Asp Lys Arg Gly Gly Ile
145 150 155 160
Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Thr Thr Leu Leu Thr Ala
165 170 175
Arg His Asp Leu Lys Asn Lys Gly Leu Asn Asp Ile Ser Thr Ile Ile
180 185 190
Lys Gln Glu Leu Thr Glu Gly Arg Ala Leu Ala Leu Ser His Thr Tyr
195 200 205
Ala Asn Val Ser Ile Ser His Val Ile Asn Leu Trp Gly Ala Asp Phe
210 215 220
Asn Ala Glu Gly Asn Leu Glu Ala Ile Tyr Val Thr Asp Ser Asp Ala
225 230 235 240
Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly Ile Asn Ala His
245 250 255
Gly His Val Ala Ile Ser Ala Lys Lys Ile Glu Gly Glu Asn Ile Gly
260 265 270
Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Ser Gly Lys Asp Ile Trp
275 280 285
Gln Lys Leu Ser
290
<210> 20
<211> 312
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 20
Asp Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val Pro
1 5 10 15
His Gln Ile Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Glu Gln
20 25 30
Phe Thr Gln Gly Glu Asp Val Ile His Ala Pro Tyr Leu Ala His Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Ala Phe Asp Gly Lys Asp Asn Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Thr Glu Ile Glu Ala Tyr Leu Ser Lys His Pro Glu Lys Gln
85 90 95
Lys Ile Ile Phe Arg Asn Gln Glu Leu Phe Asp Leu Lys Ala Ala Ile
100 105 110
Asp Thr Lys Asp Ser Gln Thr Asn Ser Gln Leu Phe Asn Tyr Phe Arg
115 120 125
Asp Lys Ala Phe Pro Asn Leu Ser Ala Arg Gln Leu Gly Val Met Pro
130 135 140
Asp Leu Val Leu Asp Met Phe Ile Asn Gly Tyr Tyr Leu Asn Val Phe
145 150 155 160
Lys Thr Gln Ser Thr Asp Val Asn Arg Pro Tyr Gln Asp Lys Asp Lys
165 170 175
Arg Gly Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Thr Thr
180 185 190
Leu Leu Thr Ala Arg His Asp Leu Lys Asn Lys Gly Leu Asn Asp Ile
195 200 205
Ser Thr Ile Ile Lys Gln Glu Leu Thr Glu Gly Arg Ala Leu Ala Leu
210 215 220
Ser His Thr Tyr Ala Asn Val Ser Ile Ser His Val Ile Asn Leu Trp
225 230 235 240
Gly Ala Asp Phe Asn Ala Glu Gly Asn Leu Glu Ala Ile Tyr Val Thr
245 250 255
Asp Ser Asp Ala Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly
260 265 270
Ile Asn Ala His Gly His Val Ala Ile Ser Ala Lys Lys Ile Glu Gly
275 280 285
Glu Asn Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Ser Gly
290 295 300
Lys Asp Ile Trp Gln Lys Leu Ser
305 310
<210> 21
<211> 312
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 21
Asp Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val Pro
1 5 10 15
His Gln Ile Thr Ser Val Trp Thr Lys Gly Val Thr Pro Leu Thr Pro
20 25 30
Glu Gln Phe Thr Gln Gly Glu Asp Val Phe His Ala Pro Tyr Val Ala
35 40 45
Asn Gln Gly Trp Tyr Asp Ile Thr Lys Ala Phe Asp Gly Lys Asp Asn
50 55 60
Leu Leu Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe
65 70 75 80
Asp Gln Asn Lys Asp Gln Ile Lys Arg Tyr Leu Glu Glu His Pro Glu
85 90 95
Lys Gln Lys Ile Asn Phe Asn Gly Glu Asn Met Phe Asp Val Lys Lys
100 105 110
Ala Ile Asp Thr Lys Asn His Gln Leu Asp Ser Lys Leu Phe Asn Tyr
115 120 125
Phe Lys Glu Lys Ala Phe Pro Tyr Leu Ser Ala Lys His Leu Gly Val
130 135 140
Phe Pro Asp His Val Ile Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser
145 150 155 160
Leu Thr Asn His Gly Pro Thr Pro Val Lys Glu Gly Ser Lys Asp Pro
165 170 175
Arg Gly Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Ser Lys
180 185 190
Leu Leu Thr Ser Arg His Asp Phe Lys Asn Lys Asn Leu Asn Asp Ile
195 200 205
Ser Thr Ile Ile Lys Gln Glu Leu Thr Lys Gly Lys Ala Leu Gly Leu
210 215 220
Ser His Thr Tyr Ala Asn Val Arg Ile Asn His Val Ile Asn Leu Trp
225 230 235 240
Gly Ala Asp Phe Asn Ala Glu Gly Asn Leu Glu Ala Ile Tyr Val Thr
245 250 255
Asp Ser Asp Ser Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly
260 265 270
Val Asn Ala His Gly His Val Ala Ile Ser Ala Lys Lys Ile Glu Gly
275 280 285
Glu Asn Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Thr Gly
290 295 300
Gln Asp Ser Trp Gln Lys Leu Ser
305 310
<210> 22
<211> 312
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 22
Asp Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val Pro
1 5 10 15
His Gln Ile Thr Ser Val Trp Thr Lys Gly Val Thr Pro Leu Thr Pro
20 25 30
Glu Gln Phe Thr Gln Gly Glu Asp Val Phe His Ala Pro Tyr Val Ala
35 40 45
Asn Gln Gly Trp Tyr Asp Ile Thr Lys Ala Phe Asp Gly Lys Asp Asn
50 55 60
Leu Leu Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe
65 70 75 80
Asp Gln Asn Lys Asp Gln Ile Lys Arg Tyr Leu Glu Glu His Pro Glu
85 90 95
Lys Gln Lys Ile Asn Phe Arg Gly Glu Asn Met Phe Asp Val Lys Glu
100 105 110
Ala Ile Arg Thr Lys Asn His Gln Leu Asp Ser Lys Leu Phe Glu Tyr
115 120 125
Phe Lys Glu Lys Ala Phe Pro Tyr Leu Ser Ala Lys His Leu Gly Val
130 135 140
Phe Pro Asp His Val Ile Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser
145 150 155 160
Leu Thr Asn His Gly Pro Thr Pro Val Lys Lys Gly Ser Lys Asp Pro
165 170 175
Arg Gly Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Ser Lys
180 185 190
Leu Leu Thr Ser Arg His Asp Phe Lys Asn Lys Asn Leu Asn Asp Ile
195 200 205
Ser Thr Ile Ile Lys Ser Glu Leu Thr Asn Gly Lys Ala Leu Gly Leu
210 215 220
Ser His Thr Tyr Ala Asn Val Arg Ile Asn His Val Ile Asn Leu Trp
225 230 235 240
Gly Ala Asp Phe Asn Ala Glu Gly Asn Leu Glu Ala Ile Tyr Val Thr
245 250 255
Asp Ser Asp Ser Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly
260 265 270
Val Asn Lys His Gly His Val Ala Ile Ser Ala Lys Lys Ile Glu Gly
275 280 285
Glu Asn Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Thr Gly
290 295 300
Gln Asp Ser Trp Gln Lys Leu Asn
305 310
<210> 23
<211> 312
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 23
Asp Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val Pro
1 5 10 15
His Gln Ile Thr Ser Val Trp Thr Lys Gly Val Thr Pro Leu Thr Pro
20 25 30
Glu Gln Phe Thr Gln Gly Glu Asp Val Phe His Ala Pro Tyr Val Ala
35 40 45
Asn Gln Gly Trp Tyr Asp Ile Thr Lys Thr Phe Asn Gly Lys Asp Asp
50 55 60
Leu Leu Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe
65 70 75 80
Asp Gln Asn Lys Asp Gln Ile Lys Arg Tyr Leu Glu Glu His Pro Glu
85 90 95
Lys Gln Lys Ile Asn Phe Asn Gly Glu Gln Met Phe Asp Val Lys Glu
100 105 110
Ala Ile Asp Thr Lys Asn His Gln Leu Asp Ser Lys Leu Phe Glu Tyr
115 120 125
Phe Lys Glu Lys Ala Phe Pro Tyr Leu Ser Thr Lys His Leu Gly Val
130 135 140
Phe Pro Asp His Val Ile Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser
145 150 155 160
Leu Thr Asn His Gly Pro Thr Pro Val Lys Glu Gly Ser Lys Asp Pro
165 170 175
Arg Gly Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Ser Lys
180 185 190
Leu Leu Thr Ser Arg His Asp Phe Lys Glu Lys Asn Leu Lys Glu Ile
195 200 205
Ser Asp Leu Ile Lys Gln Glu Leu Thr Glu Gly Lys Ala Leu Gly Leu
210 215 220
Ser His Thr Tyr Ala Asn Val Arg Ile Asn His Val Ile Asn Leu Trp
225 230 235 240
Gly Ala Asp Phe Asp Ala Glu Gly Asn Leu Lys Ala Ile Tyr Val Thr
245 250 255
Asp Ser Asp Ser Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly
260 265 270
Val Asn Ala Ala Gly Lys Val Ala Ile Ser Ala Lys Lys Ile Glu Gly
275 280 285
Glu Asn Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Thr Gly
290 295 300
Gln Asp Ser Trp Asn Gln Thr Ser
305 310
<210> 24
<211> 312
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 24
Asp Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val Pro
1 5 10 15
His Gln Ile Thr Ser Val Trp Thr Lys Gly Val Thr Pro Leu Thr Pro
20 25 30
Glu Gln Phe Thr Gln Gly Glu Asp Val Phe His Ala Pro Tyr Val Ala
35 40 45
Asn Gln Gly Trp Tyr Asp Ile Thr Lys Thr Phe Asn Gly Lys Asp Asp
50 55 60
Leu Leu Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe
65 70 75 80
Asp Gln Asn Lys Asp Gln Ile Lys Arg Tyr Leu Glu Glu His Pro Glu
85 90 95
Lys Gln Lys Ile Asn Phe Arg Gly Glu Gln Met Phe Asp Val Lys Glu
100 105 110
Ala Ile Arg Thr Lys Asn His Gln Leu Asp Ser Lys Leu Phe Glu Tyr
115 120 125
Phe Lys Glu Lys Ala Phe Pro Tyr Leu Ser Thr Lys His Leu Gly Val
130 135 140
Phe Pro Asp His Val Ile Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser
145 150 155 160
Leu Thr Asn His Gly Pro Thr Pro Val Lys Lys Gly Ser Lys Asp Pro
165 170 175
Arg Gly Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Ser Lys
180 185 190
Leu Leu Thr Ser Arg His Asp Phe Lys Glu Lys Asn Leu Lys Glu Ile
195 200 205
Ser Asp Leu Ile Lys Glu Glu Leu Thr Lys Gly Lys Ala Leu Gly Leu
210 215 220
Ser His Thr Tyr Ala Asn Val Arg Ile Asn His Val Ile Asn Leu Trp
225 230 235 240
Gly Ala Asp Phe Asp Ala Glu Gly Asn Leu Lys Ala Ile Tyr Val Thr
245 250 255
Asp Ser Asp Ser Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly
260 265 270
Val Asn Lys Ala Gly Lys Val Ala Ile Ser Ala Lys Lys Ile Glu Gly
275 280 285
Glu Asn Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Thr Gly
290 295 300
Gln Asp Ser Trp Asn Gln Thr Asn
305 310
<210> 25
<211> 310
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 25
Asp Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val Pro
1 5 10 15
His Gln Ile Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Glu Gln
20 25 30
Phe Thr Gln Gly Glu Asp Val Ile His Ala Pro Tyr Val Ala Asn Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Ala Phe Asp Gly Lys Asp Asn Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Asp Gln Ile Lys Arg Tyr Leu Glu Glu His Pro Glu Lys Gln
85 90 95
Lys Ile Asn Phe Arg Gly Glu Gln Met Phe Asp Val Lys Lys Ala Ile
100 105 110
Asp Thr Lys Asn His Gln Leu Asp Ser Lys Leu Phe Asn Tyr Phe Lys
115 120 125
Glu Lys Ala Phe Pro Gly Leu Ser Ala Arg Arg Ile Gly Val Phe Pro
130 135 140
Asp His Val Ile Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser Leu Thr
145 150 155 160
Asn His Gly Pro Thr Pro Val Lys Glu Gly Ser Lys Asp Pro Arg Gly
165 170 175
Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Ser Lys Leu Leu
180 185 190
Thr Ser Arg His Asp Phe Lys Asn Lys Asn Leu Asn Asp Ile Ser Thr
195 200 205
Ile Ile Lys Gln Glu Leu Thr Lys Gly Lys Ala Leu Gly Leu Ser His
210 215 220
Thr Tyr Ala Asn Val Ser Ile Asn His Val Ile Asn Leu Trp Gly Ala
225 230 235 240
Asp Phe Asn Ala Glu Gly Asn Leu Glu Ala Ile Tyr Val Thr Asp Ser
245 250 255
Asp Ser Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly Val Asn
260 265 270
Ala His Gly His Val Ala Ile Ser Ala Lys Lys Ile Glu Gly Glu Asn
275 280 285
Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Thr Gly Gln Asp
290 295 300
Ser Trp Gln Lys Leu Ser
305 310
<210> 26
<211> 320
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 26
Met Asp Ser Phe Ser Ala Asn Gln Glu Ile Arg Tyr Ser Glu Val Thr
1 5 10 15
Pro Tyr His Val Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Ala
20 25 30
Asn Phe Thr Gln Gly Glu Asp Val Phe His Ala Pro Tyr Val Ala Asn
35 40 45
Gln Gly Trp Tyr Asp Ile Thr Lys Thr Phe Asn Gly Lys Asp Asp Leu
50 55 60
Leu Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp
65 70 75 80
Gln Asn Lys Asp Gln Ile Lys Arg Tyr Leu Glu Glu His Pro Glu Lys
85 90 95
Gln Lys Ile Asn Phe Asn Gly Glu Gln Met Phe Asp Val Lys Glu Ala
100 105 110
Ile Asp Thr Lys Asn His Gln Leu Asp Ser Lys Leu Phe Glu Tyr Phe
115 120 125
Lys Glu Lys Ala Phe Pro Tyr Leu Ser Thr Lys His Leu Gly Val Phe
130 135 140
Pro Asp His Val Ile Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser Leu
145 150 155 160
Thr Asn His Gly Pro Thr Pro Val Lys Glu Gly Ser Lys Asp Pro Arg
165 170 175
Gly Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asp Gln Ser Lys Leu
180 185 190
Leu Thr Ser Arg His Asp Phe Lys Glu Lys Asn Leu Lys Glu Ile Ser
195 200 205
Asp Leu Ile Lys Lys Glu Leu Thr Glu Gly Lys Ala Leu Gly Leu Ser
210 215 220
His Thr Tyr Ala Asn Val Arg Ile Asn His Val Ile Asn Leu Trp Gly
225 230 235 240
Ala Asp Phe Asp Ser Asn Gly Asn Leu Lys Ala Ile Tyr Val Thr Asp
245 250 255
Ser Asp Ser Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly Val
260 265 270
Asn Ser Ala Gly Lys Val Ala Ile Ser Ala Lys Glu Ile Lys Glu Asp
275 280 285
Asn Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Thr Gly Gln
290 295 300
Asp Ser Trp Asn Gln Thr Asn Gly Gly Gly His His His His His His
305 310 315 320
<210> 27
<211> 325
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 27
Met Asp Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val
1 5 10 15
Pro His Gln Ile Thr Ser Val Trp Thr Lys Gly Val Thr Pro Leu Thr
20 25 30
Pro Glu Gln Phe Arg Tyr Asn Asn Glu Asp Val Ile His Ala Pro Tyr
35 40 45
Leu Ala His Gln Gly Trp Tyr Asp Ile Thr Lys Ala Phe Asp Gly Lys
50 55 60
Asp Asn Leu Leu Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp
65 70 75 80
Trp Phe Asp Gln Asn Lys Thr Glu Ile Glu Ala Tyr Leu Ser Lys His
85 90 95
Pro Glu Lys Gln Lys Ile Ile Phe Asn Asn Gln Glu Leu Phe Asp Leu
100 105 110
Lys Ala Ala Ile Asp Thr Lys Asp Ser Gln Thr Asn Ser Gln Leu Phe
115 120 125
Asn Tyr Phe Arg Asp Lys Ala Phe Pro Asn Leu Ser Ala Arg Gln Leu
130 135 140
Gly Val Met Pro Asp Leu Val Leu Asp Met Phe Ile Asn Gly Tyr Tyr
145 150 155 160
Leu Asn Val Phe Lys Thr Gln Ser Thr Asp Val Asn Arg Pro Tyr Gln
165 170 175
Asp Lys Asp Lys Arg Gly Gly Ile Phe Asp Ala Val Phe Thr Arg Gly
180 185 190
Asp Gln Thr Thr Leu Leu Thr Ala Arg His Asp Leu Lys Asn Lys Gly
195 200 205
Leu Asn Asp Ile Ser Thr Ile Ile Lys Gln Glu Leu Thr Glu Gly Arg
210 215 220
Ala Leu Ala Leu Ser His Thr Tyr Ala Asn Val Ser Ile Ser His Val
225 230 235 240
Ile Asn Leu Trp Gly Ala Asp Phe Asn Ala Glu Gly Asn Leu Glu Ala
245 250 255
Ile Tyr Val Thr Asp Ser Asp Ala Asn Ala Ser Ile Gly Met Lys Lys
260 265 270
Tyr Phe Val Gly Ile Asn Ala His Gly His Val Ala Ile Ser Ala Lys
275 280 285
Lys Ile Glu Gly Glu Asn Ile Gly Ala Gln Val Leu Gly Leu Phe Thr
290 295 300
Leu Ser Ser Gly Lys Asp Ile Trp Gln Lys Leu Ser Gly Gly Gly His
305 310 315 320
His His His His His
325
<210> 28
<211> 323
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 28
Met Asp Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val
1 5 10 15
Pro His Gln Ile Thr Ser Val Trp Thr Lys Gly Val Thr Pro Leu Thr
20 25 30
Pro Glu Gln Phe Arg Tyr Asn Asn Glu Asp Val Phe His Ala Pro Tyr
35 40 45
Val Ala Asn Gln Gly Trp Tyr Asp Ile Thr Lys Ala Phe Asp Gly Lys
50 55 60
Asp Asn Leu Leu Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp
65 70 75 80
Trp Phe Asp Gln Asn Lys Asp Gln Ile Lys Arg Tyr Leu Glu Glu His
85 90 95
Pro Glu Lys Gln Lys Ile Asn Phe Asn Gly Asp Asn Met Phe Asp Val
100 105 110
Lys Lys Ala Ile Asp Thr Lys Asn His Gln Leu Asp Ser Lys Leu Phe
115 120 125
Asn Tyr Phe Lys Glu Lys Ala Phe Pro Gly Leu Ser Ala Arg Arg Ile
130 135 140
Gly Val Phe Pro Asp His Val Ile Asp Met Phe Ile Asn Gly Tyr Arg
145 150 155 160
Leu Ser Leu Thr Asn His Gly Pro Thr Pro Val Lys Glu Gly Ser Lys
165 170 175
Asp Pro Arg Gly Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asn Gln
180 185 190
Ser Lys Leu Leu Thr Ser Arg His Asp Phe Lys Asn Lys Asn Leu Asn
195 200 205
Asp Ile Ser Thr Ile Ile Lys Gln Glu Leu Thr Lys Gly Lys Ala Leu
210 215 220
Gly Leu Ser His Thr Tyr Ala Asn Val Ser Ile Asn His Val Ile Asn
225 230 235 240
Leu Trp Gly Ala Asp Phe Asn Ala Glu Gly Asn Leu Glu Ala Ile Tyr
245 250 255
Val Thr Asp Ser Asp Ser Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe
260 265 270
Val Gly Val Asn Ala His Gly His Val Ala Ile Ser Ala Lys Lys Ile
275 280 285
Glu Gly Glu Asn Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser
290 295 300
Thr Gly Gln Asp Ser Trp Gln Lys Leu Ser Gly Gly Gly His His His
305 310 315 320
His His His
<210> 29
<211> 11
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 29
Pro Leu Thr Pro Glu Gln Phe Arg Tyr Asn Asn
1 5 10
<210> 30
<211> 8
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 30
Pro Pro Ala Asn Phe Thr Gln Gly
1 5
<210> 31
<211> 20
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 31
Asp Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val Pro
1 5 10 15
His Gln Ile Thr
20
<210> 32
<211> 20
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 32
Asp Ser Phe Ser Ala Asn Gln Glu Ile Arg Tyr Ser Glu Val Thr Pro
1 5 10 15
Tyr His Val Thr
20
<210> 33
<211> 963
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 33
atggatagtt tttctgctaa tcaagagatt agatattcgg aagtaacacc ttatcacgtt 60
acttccgttt ggaccaaagg agttactcct ccagcaaact tcactcaagg tgaagatgtt 120
tttcacgctc cttatgttgc taaccaagga tggtatgata ttaccaaaac attcaatgga 180
aaagacgatc ttctttgcgg ggctgccaca gcagggaata tgcttcactg gtggttcgat 240
caaaacaaag accaaattaa acgttatttg gaagagcatc cagaaaagca aaaaataaac 300
ttcaatggcg aacagatgtt tgacgtaaaa gaagctatcg acactaaaaa ccaccagcta 360
gatagtaaat tatttgaata ttttaaagaa aaagctttcc cttatctatc tactaaacac 420
ctaggagttt tccctgatca tgtaattgat atgttcatta acggctaccg ccttagtcta 480
actaaccacg gtccaacgcc agtaaaagaa ggtagtaaag atccccgagg tggtattttt 540
gacgccgtat ttacaagagg tgatcaaagt aagctattga caagtcgtca tgattttaaa 600
gaaaaaaatc tcaaagaaat cagtgatctc attaagaaag agttaaccga aggcaaggct 660
ctaggcctat cacacaccta cgctaacgta cgcatcaacc atgttataaa cctgtgggga 720
gctgactttg attctaacgg gaaccttaaa gctatttatg taacagactc tgatagtaat 780
gcatctattg gtatgaagaa atactttgtt ggtgttaatt ccgctggaaa agtagctatt 840
tctgctaaag aaataaaaga agataatata ggtgctcaag tactagggtt atttacactt 900
tcaacagggc aagatagttg gaatcagacc aatggcggtg gccatcatca ccatcaccac 960
taa 963
<210> 34
<211> 978
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 34
atggacgatt accaaaggaa tgctacggaa gcttatgcca aagaagtacc acatcagatc 60
acttctgtat ggaccaaagg tgttacacca ctaacacccg agcagtttcg atataataac 120
gaagatgtga tccatgcgcc atatcttgct catcaaggct ggtacgatat caccaaggcc 180
ttcgatggga aggataatct cttgtgtggc gcagcaacgg caggtaatat gctgcattgg 240
tggtttgatc aaaataaaac agagattgaa gcctatttaa gtaaacaccc tgaaaagcaa 300
aaaatcattt ttaacaacca agagctattt gatttgaaag ctgctatcga taccaaggac 360
agtcaaacca atagtcagct ttttaattat tttagagata aagcctttcc aaatctatca 420
gcacgtcaac tcggggttat gcctgatctt gttctagata tgtttatcaa tggttactac 480
ttaaatgtgt ttaaaacaca gtctactgat gtcaatcgac cttatcagga caaggacaaa 540
cgaggtggta ttttcgatgc tgttttcacc agaggagatc agacaacgct cttgacagct 600
cgtcatgatt taaaaaataa aggactaaat gacatcagca ccattatcaa gcaagaactg 660
actgaaggaa gagcccttgc tttatcacat acctacgcca atgttagcat tagccatgtg 720
attaacttgt ggggagctga ttttaatgct gaaggaaacc ttgaggccat ctatgtcaca 780
gactcagatg ctaatgcgtc tattggtatg aaaaaatatt ttgtcggcat taatgctcat 840
ggacatgtcg ccatttctgc caagaaaata gaaggagaaa acattggcgc tcaagtatta 900
ggcttattta cgctttccag tggcaaggac atttggcaga aactgagcgg cggtggccat 960
catcaccatc accactaa 978
<210> 35
<211> 972
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 35
atggatgatt atcagcgcaa cgcgaccgaa gcgtatgcga aagaagtgcc gcatcagatt 60
accagcgtgt ggaccaaagg cgtgaccccg ctgaccccgg aacagtttcg ctataacaac 120
gaagatgtgt ttcatgcgcc gtatgtggcg aaccagggct ggtatgatat taccaaagcg 180
tttgatggca aagataacct gctgtgcggc gcggcgaccg cgggcaacat gctgcattgg 240
tggtttgatc agaacaaaga tcagattaaa cgctatctgg aagaacatcc ggaaaaacag 300
aaaattaact ttaacggcga taacatgttt gatgtgaaaa aagcgattga taccaaaaac 360
catcagctgg atagcaaact gtttaactat tttaaagaaa aagcgtttcc gggcctgagc 420
gcgcgccgca ttggcgtgtt tccggatcat gtgattgata tgtttattaa cggctatcgc 480
ctgagcctga ccaaccatgg cccgaccccg gtgaaagaag gcagcaaaga tccgcgcggc 540
ggcatttttg atgcggtgtt tacccgcggc aaccagagca aactgctgac cagccgccat 600
gattttaaaa acaaaaacct gaacgatatt agcaccatta ttaaacagga actgaccaaa 660
ggcaaagcgc tgggcctgag ccatacctat gcgaacgtga gcattaacca tgtgattaac 720
ctgtggggcg cggattttaa cgcggaaggc aacctggaag cgatttatgt gaccgatagc 780
gatagcaacg cgagcattgg catgaaaaaa tattttgtgg gcgtgaacgc gcatggccat 840
gtggcgatta gcgcgaaaaa aattgaaggc gaaaacattg gcgcgcaggt gctgggcctg 900
tttaccctga gcaccggcca ggatagctgg cagaaactga gcggcggtgg ccatcatcac 960
catcaccact aa 972
<210> 36
<211> 978
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 36
atggatgatt atcagcgcaa cgcgaccgaa gcgtatgcga aagaagtgcc gcatcagatt 60
accagcgtgt ggaccaaagg cgtgaccccg ctgaccccgg aacagtttcg ctataacaac 120
gaagatgtga ttcatgcgcc gtatctggcg aaccagggct ggtatgatat taccaaagcg 180
tttgatggca aagataacct gctgtgcggc gcggcgaccg cgggcaacat gctgcattgg 240
tggtttgatc agaacaaaac cgaaattgaa gcgtatctga gcaaacatcc ggaaaaacag 300
aaaattattt ttcgcaacca ggaactgttt gatctgaaag aagcgattcg caccaaagat 360
agccagacca acagccagct gtttgaatat tttcgcgata aagcgtttcc gtatctgagc 420
gcgcgccagc tgggcgtgat gccggatctg gtgctggata tgtttattaa cggctattat 480
ctgaacgtgt ttaaaaccca gagcaccgat gtgaaacgcc cgtatcagga taaagataaa 540
cgcggcggca tttttgatgc ggtgtttacc cgcggcaacc agaccaccct gctgaccgcg 600
cgccatgatc tgaaaaacaa aggcctgaac gatattagca ccattattaa agaagaactg 660
accaaaggcc gcgcgctggc gctgagccat acctatgcga acgtgagcat tagccatgtg 720
attaacctgt ggggcgcgga ttttaacgcg gaaggcaacc tggaagcgat ttatgtgacc 780
gatagcgatg cgaacgcgag cattggcatg aaaaaatatt ttgtgggcat taacaaacat 840
ggccatgtgg cgattagcgc gaaaaaaatt gaaggcgaaa acattggcgc gcaggtgctg 900
ggcctgttta ccctgagcag cggcaaagat atttggcaga aactgaacgg cggtggccat 960
catcaccatc accactaa 978
<210> 37
<211> 978
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 37
atggatgatt atcagcgcaa cgcgaccgaa gcgtatgcga aagaagtgcc gcatcagatt 60
accagcgtgt ggaccaaagg cgtgaccccg ctgaccccgg aacagtttcg ctataacaac 120
gaagatgtga ttcatgcgcc gtatctggcg catcagggct ggtatgatat taccaaaacc 180
tttaacggca aagataacct gctgtgcggc gcggcgaccg cgggcaacat gctgcattgg 240
tggtttgatc agaacaaaac cgaaattgaa gcgtatctga gcaaacatcc ggaaaaacag 300
aaaattattt ttaacaacga agaactgttt gatctgaaag cggcgattga taccaaagat 360
agccagacca acagccagct gtttaactat tttaaagaaa aagcgtttcc gaacctgagc 420
acccgccagc tgggcgtgat gccggatctg gtgctggata tgtttattaa cggctattat 480
ctgaacgtgt ttaaaaccca gagcaccgat gtgaaccgcc cgtatcagga taaagataaa 540
cgcggcggca tttttgatgc ggtgtttacc cgcggcaacc agaccaccct gctgaccgcg 600
cgccatgatt ttaaagaaaa aggcctgaaa gatattagca ccattattaa acaggaactg 660
accgaaggcc gcgcgctggc gctgagccat acctatgcga acgtgagcat tagccatgtg 720
attaacctgt ggggcgcgga ttttgatgcg gaaggcaacc tgaaagcgat ttatgtgacc 780
gatagcgatg cgaacgcgag cattggcatg aaaaaatatt ttgtgggcat taacgcgcat 840
ggcaaagtgg cgattagcgc gaaaaaaatt gaaggcgaaa acattggcgc gcaggtgctg 900
ggcctgttta ccctgagcag cggcaaagat atttggcagc agctgagcgg cggtggccat 960
catcaccatc accactaa 978
<210> 38
<211> 978
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 38
atggatagct ttagcgcgaa ccaggaaatt cgctatagcg aagtgacccc gtatcatgtg 60
accagcgtgt ggaccaaagg cgtgaccccg ctgaccccgg aacagtttcg ctataacaac 120
gaagatgtga ttcatgcgcc gtatctggcg catcagggct ggtatgatat taccaaagcg 180
tttgatggca aagataacct gctgtgcggc gcggcgaccg cgggcaacat gctgcattgg 240
tggtttgatc agaacaaaac cgaaattgaa gcgtatctga gcaaacatcc ggaaaaacag 300
aaaattattt ttaacaacca ggaactgttt gatctgaaag cggcgattga taccaaagat 360
agccagacca acagccagct gtttaactat tttcgcgata aagcgtttcc gaacctgagc 420
gcgcgccagc tgggcgtgat gccggatctg gtgctggata tgtttattaa cggctattat 480
ctgaacgtgt ttaaaaccca gagcaccgat gtgaaccgcc cgtatcagga taaagataaa 540
cgcggcggca tttttgatgc ggtgtttacc cgcggcgatc agaccaccct gctgaccgcg 600
cgccatgatc tgaaaaacaa aggcctgaac gatattagca ccattattaa acaggaactg 660
accgaaggcc gcgcgctggc gctgagccat acctatgcga acgtgagcat tagccatgtg 720
attaacctgt ggggcgcgga ttttaacgcg gaaggcaacc tggaagcgat ttatgtgacc 780
gatagcgatg cgaacgcgag cattggcatg aaaaaatatt ttgtgggcat taacgcgcat 840
ggccatgtgg cgattagcgc gaaaaaaatt gaaggcgaaa acattggcgc gcaggtgctg 900
ggcctgttta ccctgagcag cggcaaagat atttggcaga aactgagcgg cggtggccat 960
catcaccatc accactaa 978
<210> 39
<211> 918
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 39
atgagcgtgt ggaccaaagg cgtgaccccg ctgaccccgg aacagtttcg ctataacaac 60
gaagatgtga ttcatgcgcc gtatctggcg catcagggct ggtatgatat taccaaagcg 120
tttgatggca aagataacct gctgtgcggc gcggcgaccg cgggcaacat gctgcattgg 180
tggtttgatc agaacaaaac cgaaattgaa gcgtatctga gcaaacatcc ggaaaaacag 240
aaaattattt ttaacaacca ggaactgttt gatctgaaag cggcgattga taccaaagat 300
agccagacca acagccagct gtttaactat tttcgcgata aagcgtttcc gaacctgagc 360
gcgcgccagc tgggcgtgat gccggatctg gtgctggata tgtttattaa cggctattat 420
ctgaacgtgt ttaaaaccca gagcaccgat gtgaaccgcc cgtatcagga taaagataaa 480
cgcggcggca tttttgatgc ggtgtttacc cgcggcgatc agaccaccct gctgaccgcg 540
cgccatgatc tgaaaaacaa aggcctgaac gatattagca ccattattaa acaggaactg 600
accgaaggcc gcgcgctggc gctgagccat acctatgcga acgtgagcat tagccatgtg 660
attaacctgt ggggcgcgga ttttaacgcg gaaggcaacc tggaagcgat ttatgtgacc 720
gatagcgatg cgaacgcgag cattggcatg aaaaaatatt ttgtgggcat taacgcgcat 780
ggccatgtgg cgattagcgc gaaaaaaatt gaaggcgaaa acattggcgc gcaggtgctg 840
ggcctgttta ccctgagcag cggcaaagat atttggcaga aactgagcgg cggtggccat 900
catcaccatc accactaa 918
<210> 40
<211> 975
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 40
atggacgatt accaaaggaa tgctacggaa gcttatgcca aagaagtacc acatcagatc 60
acttctgtat ggaccaaagg tgttacacca ctaacacccg agcagtttac tcaaggtgaa 120
gatgtgatcc atgcgccata tcttgctcat caaggctggt acgatatcac caaggccttc 180
gatgggaagg ataatctctt gtgtggcgca gcaacggcag gtaatatgct gcattggtgg 240
tttgatcaaa ataaaacaga gattgaagcc tatttaagta aacaccctga aaagcaaaaa 300
atcattttta acaaccaaga gctatttgat ttgaaagctg ctatcgatac caaggacagt 360
caaaccaata gtcagctttt taattatttt agagataaag cctttccaaa tctatcagca 420
cgtcaactcg gggttatgcc tgatcttgtt ctagatatgt ttatcaatgg ttactactta 480
aatgtgttta aaacacagtc tactgatgtc aatcgacctt atcaggacaa ggacaaacga 540
ggtggtattt tcgatgctgt tttcaccaga ggagatcaga caacgctctt gacagctcgt 600
catgatttaa aaaataaagg actaaatgac atcagcacca ttatcaagca agaactgact 660
gaaggaagag cccttgcttt atcacatacc tacgccaatg ttagcattag ccatgtgatt 720
aacttgtggg gagctgattt taatgctgaa ggaaaccttg aggccatcta tgtcacagac 780
tcagatgcta atgcgtctat tggtatgaaa aaatattttg tcggcattaa tgctcatgga 840
catgtcgcca tttctgccaa gaaaatagaa ggagaaaaca ttggcgctca agtattaggc 900
ttatttacgc tttccagtgg caaggacatt tggcagaaac tgagcggcgg tggccatcat 960
caccatcacc actaa 975
<210> 41
<211> 969
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 41
atggacgatt accaaaggaa tgctacggaa gcttatgcca aagaagtacc acatcagatc 60
acttctgtat ggaccaaagg tgttacacca cccgagcagt ttactcaagg tgaagatgtg 120
atccatgcgc catatcttgc tcatcaaggc tggtacgata tcaccaaggc cttcgatggg 180
aaggataatc tcttgtgtgg cgcagcaacg gcaggtaata tgctgcattg gtggtttgat 240
caaaataaaa cagagattga agcctattta agtaaacacc ctgaaaagca aaaaatcatt 300
tttaacaacc aagagctatt tgatttgaaa gctgctatcg ataccaagga cagtcaaacc 360
aatagtcagc tttttaatta ttttagagat aaagcctttc caaatctatc agcacgtcaa 420
ctcggggtta tgcctgatct tgttctagat atgtttatca atggttacta cttaaatgtg 480
tttaaaacac agtctactga tgtcaatcga ccttatcagg acaaggacaa acgaggtggt 540
attttcgatg ctgttttcac cagaggagat cagacaacgc tcttgacagc tcgtcatgat 600
ttaaaaaata aaggactaaa tgacatcagc accattatca agcaagaact gactgaagga 660
agagcccttg ctttatcaca tacctacgcc aatgttagca ttagccatgt gattaacttg 720
tggggagctg attttaatgc tgaaggaaac cttgaggcca tctatgtcac agactcagat 780
gctaatgcgt ctattggtat gaaaaaatat tttgtcggca ttaatgctca tggacatgtc 840
gccatttctg ccaagaaaat agaaggagaa aacattggcg ctcaagtatt aggcttattt 900
acgctttcca gtggcaagga catttggcag aaactgagcg gcggtggcca tcatcaccat 960
caccactaa 969
<210> 42
<211> 972
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 42
atggacgatt accaaaggaa tgctacggaa gcttatgcca aagaagtacc acatcagatc 60
acttctgtat ggaccaaagg tgttacacca cccgagcagt ttcgatataa taacgaagat 120
gtgatccatg cgccatatct tgctcatcaa ggctggtacg atatcaccaa ggccttcgat 180
gggaaggata atctcttgtg tggcgcagca acggcaggta atatgctgca ttggtggttt 240
gatcaaaata aaacagagat tgaagcctat ttaagtaaac accctgaaaa gcaaaaaatc 300
atttttaaca accaagagct atttgatttg aaagctgcta tcgataccaa ggacagtcaa 360
accaatagtc agctttttaa ttattttaga gataaagcct ttccaaatct atcagcacgt 420
caactcgggg ttatgcctga tcttgttcta gatatgttta tcaatggtta ctacttaaat 480
gtgtttaaaa cacagtctac tgatgtcaat cgaccttatc aggacaagga caaacgaggt 540
ggtattttcg atgctgtttt caccagagga gatcagacaa cgctcttgac agctcgtcat 600
gatttaaaaa ataaaggact aaatgacatc agcaccatta tcaagcaaga actgactgaa 660
ggaagagccc ttgctttatc acatacctac gccaatgtta gcattagcca tgtgattaac 720
ttgtggggag ctgattttaa tgctgaagga aaccttgagg ccatctatgt cacagactca 780
gatgctaatg cgtctattgg tatgaaaaaa tattttgtcg gcattaatgc tcatggacat 840
gtcgccattt ctgccaagaa aatagaagga gaaaacattg gcgctcaagt attaggctta 900
tttacgcttt ccagtggcaa ggacatttgg cagaaactga gcggcggtgg ccatcatcac 960
catcaccact aa 972
<210> 43
<211> 969
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 43
atggacgatt accaaaggaa tgctacggaa gcttatgcca aagaagtacc acatcagatc 60
acttctgtat ggaccaaagg tgttacacca cccgagcagt ttactcaagg tgaagatgtg 120
atccatgcgc catatcttgc tcatcaaggc tggtacgata tcaccaaggc cttcgatggg 180
aaggataatc tcttgtgtgg cgcagcaacg gcaggtaata tgctgcattg gtggtttgat 240
caaaataaaa cagagattga agcctattta agtaaacacc ctgaaaagca aaaaatcatt 300
attaacaacc aagagctatt tgatttgaaa gctgctatcg ataccaagga cagtcaaacc 360
aatagtcagc tttttaatta ttttagagat aaagcctttc caaatctatc agcacgtcaa 420
ctcggggtta tgcctgatct tgttctagat atgtttatca atggttacta cttaaatgtg 480
tttaaaacac agtctactga tgtcaatcga ccttatcagg acaaggacaa acgaggtggt 540
attttcgatg ctgttttcac cagaggagat cagacaacgc tcttgacagc tcgtcatgat 600
ttaaaaaata aaggactaaa tgacatcagc accattatca agcaagaact gactgaagga 660
agagcccttg ctttatcaca tacctacgcc aatgttagca ttagccatgt gattaacttg 720
tggggagctg attttaatgc tgaaggaaac cttgaggcca tctatgtcac agactcagat 780
gctaatgcgt ctattggtat gaaaaaatat tttgtcggca ttaatgctca tggacatgtc 840
gccatttctg ccaagaaaat agaaggagaa aacattggcg ctcaagtatt aggcttattt 900
acgctttcca gtggcaagga catttggcag aaactgagcg gcggtggcca tcatcaccat 960
caccactaa 969
<210> 44
<211> 969
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 44
atggacgatt accaaaggaa tgctacggaa gcttatgcca aagaagtacc acatcagatc 60
acttctgtat ggaccaaagg tgttacacca cccgagcagt ttactcaagg tgaagatgtg 120
atccatgcgc catatcttgc tcatcaaggc tggtacgata tcaccaaggc cttcgatggg 180
aaggataatc tcttgtgtgg cgcagcaacg gcaggtaata tgctgcattg gtggtttgat 240
caaaataaaa cagagattga agcctattta agtaaacacc ctgaaaagca aaaaatcatt 300
tttcgtaacc aagagctatt tgatttgaaa gctgctatcg ataccaagga cagtcaaacc 360
aatagtcagc tttttaatta ttttagagat aaagcctttc caaatctatc agcacgtcaa 420
ctcggggtta tgcctgatct tgttctagat atgtttatca atggttacta cttaaatgtg 480
tttaaaacac agtctactga tgtcaatcga ccttatcagg acaaggacaa acgaggtggt 540
attttcgatg ctgttttcac cagaggagat cagacaacgc tcttgacagc tcgtcatgat 600
ttaaaaaata aaggactaaa tgacatcagc accattatca agcaagaact gactgaagga 660
agagcccttg ctttatcaca tacctacgcc aatgttagca ttagccatgt gattaacttg 720
tggggagctg attttaatgc tgaaggaaac cttgaggcca tctatgtcac agactcagat 780
gctaatgcgt ctattggtat gaaaaaatat tttgtcggca ttaatgctca tggacatgtc 840
gccatttctg ccaagaaaat agaaggagaa aacattggcg ctcaagtatt aggcttattt 900
acgctttcca gtggcaagga catttggcag aaactgagcg gcggtggcca tcatcaccat 960
caccactaa 969
<210> 45
<211> 969
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 45
atggacgatt accaaaggaa tgctacggaa gcttatgcca aagaagtacc acatcagatc 60
acttctgtat ggaccaaagg tgttacacca cccgagcagt ttactcaagg tgaagatgtg 120
atccatgcgc catatcttgc tcatcaaggc tggtacgata tcaccaaggc cttcgatggg 180
aaggataatc tcttgtgtgg cgcagcaacg gcaggtaata tgctgcattg gtggtttgat 240
caaaataaaa cagagattga agcctattta agtaaacacc ctgaaaagca aaaaatcatt 300
attcgtaacc aagagctatt tgatttgaaa gctgctatcg ataccaagga cagtcaaacc 360
aatagtcagc tttttaatta ttttagagat aaagcctttc caaatctatc agcacgtcaa 420
ctcggggtta tgcctgatct tgttctagat atgtttatca atggttacta cttaaatgtg 480
tttaaaacac agtctactga tgtcaatcga ccttatcagg acaaggacaa acgaggtggt 540
attttcgatg ctgttttcac cagaggagat cagacaacgc tcttgacagc tcgtcatgat 600
ttaaaaaata aaggactaaa tgacatcagc accattatca agcaagaact gactgaagga 660
agagcccttg ctttatcaca tacctacgcc aatgttagca ttagccatgt gattaacttg 720
tggggagctg attttaatgc tgaaggaaac cttgaggcca tctatgtcac agactcagat 780
gctaatgcgt ctattggtat gaaaaaatat tttgtcggca ttaatgctca tggacatgtc 840
gccatttctg ccaagaaaat agaaggagaa aacattggcg ctcaagtatt aggcttattt 900
acgctttcca gtggcaagga catttggcag aaactgagcg gcggtggcca tcatcaccat 960
caccactaa 969
<210> 46
<211> 969
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 46
atggatgatt atcagcgcaa cgcgaccgaa gcgtatgcga aagaagtgcc gcatcagatt 60
accagcgtgt ggaccaaagg cgtgaccccg ccggaacagt ttactcaagg tgaagatgtg 120
attcatgcgc cgtatctggc gaaccagggc tggtatgata ttaccaaagc gtttgatggc 180
aaagataacc tgctgtgcgg cgcggcgacc gcgggcaaca tgctgcattg gtggtttgat 240
cagaacaaaa ccgaaattga agcgtatctg agcaaacatc cggaaaaaca gaaaattatt 300
tttcgcaacc aggaactgtt tgatctgaaa gaagcgattc gcaccaaaga tagccagacc 360
aacagccagc tgtttgaata ttttcgcgat aaagcgtttc cgtatctgag cgcgcgccag 420
ctgggcgtga tgccggatct ggtgctggat atgtttatta acggctatta tctgaacgtg 480
tttaaaaccc agagcaccga tgtgaaacgc ccgtatcagg ataaagataa acgcggcggc 540
atttttgatg cggtgtttac ccgcggcaac cagaccaccc tgctgaccgc gcgccatgat 600
ctgaaaaaca aaggcctgaa cgatattagc accattatta aagaagaact gaccaaaggc 660
cgcgcgctgg cgctgagcca tacctatgcg aacgtgagca ttagccatgt gattaacctg 720
tggggcgcgg attttaacgc ggaaggcaac ctggaagcga tttatgtgac cgatagcgat 780
gcgaacgcga gcattggcat gaaaaaatat tttgtgggca ttaacaaaca tggccatgtg 840
gcgattagcg cgaaaaaaat tgaaggcgaa aacattggcg cgcaggtgct gggcctgttt 900
accctgagca gcggcaaaga tatttggcag aaactgaacg gcggtggcca tcatcaccat 960
caccactaa 969
<210> 47
<211> 909
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 47
atgtctgtat ggaccaaagg tgttacacca cccgagcagt ttactcaagg tgaagatgtg 60
atccatgcgc catatcttgc tcatcaaggc tggtacgata tcaccaaggc cttcgatggg 120
aaggataatc tcttgtgtgg cgcagcaacg gcaggtaata tgctgcattg gtggtttgat 180
caaaataaaa cagagattga agcctattta agtaaacacc ctgaaaagca aaaaatcatt 240
tttcgtaacc aagagctatt tgatttgaaa gctgctatcg ataccaagga cagtcaaacc 300
aatagtcagc tttttaatta ttttagagat aaagcctttc caaatctatc agcacgtcaa 360
ctcggggtta tgcctgatct tgttctagat atgtttatca atggttacta cttaaatgtg 420
tttaaaacac agtctactga tgtcaatcga ccttatcagg acaaggacaa acgaggtggt 480
attttcgatg ctgttttcac cagaggaaac cagacaacgc tcttgacagc tcgtcatgat 540
ttaaaaaata aaggactaaa tgacatcagc accattatca agcaagaact gactgaagga 600
agagcccttg ctttatcaca tacctacgcc aatgttagca ttagccatgt gattaacttg 660
tggggagctg attttaatgc tgaaggaaac cttgaggcca tctatgtcac agactcagat 720
gctaatgcgt ctattggtat gaaaaaatat tttgtcggca ttaatgctca tggacatgtc 780
gccatttctg ccaagaaaat agaaggagaa aacattggcg ctcaagtatt aggcttattt 840
acgctttcca gtggcaagga catttggcag aaactgagcg gcggtggcca tcatcaccat 900
caccactaa 909
<210> 48
<211> 969
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 48
atggacgatt accaaaggaa tgctacggaa gcttatgcca aagaagtacc acatcagatc 60
acttctgtat ggaccaaagg tgttacacca cccgagcagt ttactcaagg tgaagatgtg 120
atccatgcgc catatcttgc tcatcaaggc tggtacgata tcaccaaggc cttcgatggg 180
gcggataatc tcttgtgtgg cgcagcaacg gcaggtaata tgctgcattg gtggtttgat 240
caaaataaaa cagagattga agcctattta agtaaacacc ctgaaaagca aaaaatcatt 300
tttcgtaacc aagagctatt tgatttgaaa gctgctatcg ataccaagga cagtcaaacc 360
aatagtcagc tttttaatta ttttagagat aaagcctttc caaatctatc agcacgtcaa 420
ctcggggtta tgcctgatct tgttctagat atgtttatca atggttacta cttaaatgtg 480
tttaaaacac agtctactga tgtcaatcga ccttatcagg acaaggacaa acgaggtggt 540
attttcgatg ctgttttcac cagaggaaat cagacaacgc tcttgacagc tcgtcatgat 600
ttaaaaaata aaggactaaa tgacatcagc accattatca agcaagaact gactgaagga 660
agagcccttg ctttatcaca tacctacgcc aatgttagca ttagccatgt gattaacttg 720
tggggagctg attttaatgc tgaaggaaac cttgaggcca tctatgtcac agactcagat 780
gctaatgcgt ctattggtat gaaaaaatat tttgtcggca ttaatgctca tggacatgtc 840
gccatttctg ccaagaaaat agaaggagaa aacattggcg ctcaagtatt aggcttattt 900
acgctttcca gtggcaagga catttggcag aaactgagcg gcggtggcca tcatcaccat 960
caccactaa 969
<210> 49
<211> 909
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 49
atgtctgtat ggaccaaagg tgttacacca cccgagcagt ttactcaagg tgaagatgtg 60
atccatgcgc catatcttgc tcatcaaggc tggtacgata tcaccaaggc cttcgatggg 120
gcggataatc tcttgtgtgg cgcagcaacg gcaggtaata tgctgcattg gtggtttgat 180
caaaataaaa cagagattga agcctattta agtaaacacc ctgaaaagca aaaaatcatt 240
tttcgtaacc aagagctatt tgatttgaaa gctgctatcg ataccaagga cagtcaaacc 300
aatagtcagc tttttaatta ttttagagat aaagcctttc caaatctatc agcacgtcaa 360
ctcggggtta tgcctgatct tgttctagat atgtttatca atggttacta cttaaatgtg 420
tttaaaacac agtctactga tgtcaatcga ccttatcagg acaaggacaa acgaggtggt 480
attttcgatg ctgttttcac cagaggaaat cagacaacgc tcttgacagc tcgtcatgat 540
ttaaaaaata aaggactaaa tgacatcagc accattatca agcaagaact gactgaagga 600
agagcccttg ctttatcaca tacctacgcc aatgttagca ttagccatgt gattaacttg 660
tggggagctg attttaatgc tgaaggaaac cttgaggcca tctatgtcac agactcagat 720
gctaatgcgt ctattggtat gaaaaaatat tttgtcggca ttaatgctca tggacatgtc 780
gccatttctg ccaagaaaat agaaggagaa aacattggcg ctcaagtatt aggcttattt 840
acgctttcca gtggcaagga catttggcag aaactgagcg gcggtggcca tcatcaccat 900
caccactaa 909
<210> 50
<211> 969
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 50
atggacgatt accaaaggaa tgctacggaa gcttatgcca aagaagtacc acatcagatc 60
acttctgtat ggaccaaagg tgttacacca cccgagcagt ttactcaagg tgaagatgtg 120
atccatgcgc catatcttgc tcatcaaggc tggtacgata tcaccaaggc cttcgatggg 180
aaggataatc tcttgtgtgg cgcagcaacg gcaggtaata tgctgcattg gtggtttgat 240
caaaataaaa cagagattga agcctattta agtaaacacc ctgaaaagca aaaaatcatt 300
tttcgtaacc aagagctatt tgatttgaaa gctgctatcg ataccaagga cagtcaaacc 360
aatagtcagc tttttaatta ttttagagat aaagcctttc caaatctatc agcacgtcaa 420
ctcggggtta tgcctgatct tgttctagat atgtttatca atggttacta cttaaatgtg 480
tttaaaacac agtctactga tgtcaatcga ccttatcagg acaaggacaa acgaggtggt 540
attttcgatg ctgttttcac cagaggaaac cagacaacgc tcttgacagc tcgtcatgat 600
ttaaaaaata aaggactaaa tgacatcagc accattatca agcaagaact gactgaagga 660
agagcccttg ctttatcaca tacctacgcc aatgttagca ttagccatgt gattaacttg 720
tggggagctg attttaatgc tgaaggaaac cttgaggcca tctatgtcac agactcagat 780
gctaatgcgt ctattggtat gaaaaaatat tttgtcggca ttaatgctca tggacatgtc 840
gccatttctg ccaagaaaat agaaggagaa aacattggcg ctcaagtatt aggcttattt 900
acgctttcca gtggcaagga catttggcag aaactgagcg gcggtggcca tcatcaccat 960
caccactaa 969
<210> 51
<211> 969
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 51
atggatgatt atcagcgcaa cgcgaccgaa gcgtatgcga aagaagtgcc gcatcagatt 60
accagcgtgt ggaccaaagg cgtgaccccg ctgaccccgg aacagtttac ccagggcgaa 120
gatgtgtttc atgcgccgta tgtggcgaac cagggctggt atgatattac caaagcgttt 180
gatggcaaag ataacctgct gtgcggcgcg gcgaccgcgg gcaacatgct gcattggtgg 240
tttgatcaga acaaagatca gattaaacgc tatctggaag aacatccgga aaaacagaaa 300
attaacttta acggcgaaaa catgtttgat gtgaaaaaag cgattgatac caaaaaccat 360
cagctggata gcaaactgtt taactatttt aaagaaaaag cgtttccgta tctgagcgcg 420
aaacatctgg gcgtgtttcc ggatcatgtg attgatatgt ttattaacgg ctatcgcctg 480
agcctgacca accatggccc gaccccggtg aaagaaggca gcaaagatcc gcgcggcggc 540
atttttgatg cggtgtttac ccgcggcaac cagagcaaac tgctgaccag ccgccatgat 600
tttaaaaaca aaaacctgaa cgatattagc accattatta aacaggaact gaccaaaggc 660
aaagcgctgg gcctgagcca tacctatgcg aacgtgcgca ttaaccatgt gattaacctg 720
tggggcgcgg attttaacgc ggaaggcaac ctggaagcga tttatgtgac cgatagcgat 780
agcaacgcga gcattggcat gaaaaaatat tttgtgggcg tgaacgcgca tggccatgtg 840
gcgattagcg cgaaaaaaat tgaaggcgaa aacattggcg cgcaggtgct gggcctgttt 900
accctgagca ccggccagga tagctggcag aaactgagcg gcggtggcca tcatcaccat 960
caccactaa 969
<210> 52
<211> 969
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 52
atggatgatt atcagcgcaa cgcgaccgaa gcgtatgcga aagaagtgcc gcatcagatt 60
accagcgtgt ggaccaaagg cgtgaccccg ctgaccccgg aacagtttac ccagggcgaa 120
gatgtgtttc atgcgccgta tgtggcgaac cagggctggt atgatattac caaagcgttt 180
gatggcaaag ataacctgct gtgcggcgcg gcgaccgcgg gcaacatgct gcattggtgg 240
tttgatcaga acaaagatca gattaaacgc tatctggaag aacatccgga aaaacagaaa 300
attaactttc gcggcgaaaa catgtttgat gtgaaagaag cgattcgcac caaaaaccat 360
cagctggata gcaaactgtt tgaatatttt aaagaaaaag cgtttccgta tctgagcgcg 420
aaacatctgg gcgtgtttcc ggatcatgtg attgatatgt ttattaacgg ctatcgcctg 480
agcctgacca accatggccc gaccccggtg aaaaaaggca gcaaagatcc gcgcggcggc 540
atttttgatg cggtgtttac ccgcggcaac cagagcaaac tgctgaccag ccgccatgat 600
tttaaaaaca aaaacctgaa cgatattagc accattatta aaagcgaact gaccaacggc 660
aaagcgctgg gcctgagcca tacctatgcg aacgtgcgca ttaaccatgt gattaacctg 720
tggggcgcgg attttaacgc ggaaggcaac ctggaagcga tttatgtgac cgatagcgat 780
agcaacgcga gcattggcat gaaaaaatat tttgtgggcg tgaacaaaca tggccatgtg 840
gcgattagcg cgaaaaaaat tgaaggcgaa aacattggcg cgcaggtgct gggcctgttt 900
accctgagca ccggccagga tagctggcag aaactgaacg gcggtggcca tcatcaccat 960
caccactaa 969
<210> 53
<211> 969
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 53
atggatgatt atcagcgcaa cgcgaccgaa gcgtatgcga aagaagtgcc gcatcagatt 60
accagcgtgt ggaccaaagg cgtgaccccg ctgaccccgg aacagtttac ccagggcgaa 120
gatgtgtttc atgcgccgta tgtggcgaac cagggctggt atgatattac caaaaccttt 180
aacggcaaag atgatctgct gtgcggcgcg gcgaccgcgg gcaacatgct gcattggtgg 240
tttgatcaga acaaagatca gattaaacgc tatctggaag aacatccgga aaaacagaaa 300
attaacttta acggcgaaca gatgtttgat gtgaaagaag cgattgatac caaaaaccat 360
cagctggata gcaaactgtt tgaatatttt aaagaaaaag cgtttccgta tctgagcacc 420
aaacatctgg gcgtgtttcc ggatcatgtg attgatatgt ttattaacgg ctatcgcctg 480
agcctgacca accatggccc gaccccggtg aaagaaggca gcaaagatcc gcgcggcggc 540
atttttgatg cggtgtttac ccgcggcaac cagagcaaac tgctgaccag ccgccatgat 600
tttaaagaaa aaaacctgaa agaaattagc gatctgatta aacaggaact gaccgaaggc 660
aaagcgctgg gcctgagcca tacctatgcg aacgtgcgca ttaaccatgt gattaacctg 720
tggggcgcgg attttgatgc ggaaggcaac ctgaaagcga tttatgtgac cgatagcgat 780
agcaacgcga gcattggcat gaaaaaatat tttgtgggcg tgaacgcggc gggcaaagtg 840
gcgattagcg cgaaaaaaat tgaaggcgaa aacattggcg cgcaggtgct gggcctgttt 900
accctgagca ccggccagga tagctggaac cagaccagcg gcggtggcca tcatcaccat 960
caccactaa 969
<210> 54
<211> 969
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 54
atggatgatt atcagcgcaa cgcgaccgaa gcgtatgcga aagaagtgcc gcatcagatt 60
accagcgtgt ggaccaaagg cgtgaccccg ctgaccccgg aacagtttac ccagggcgaa 120
gatgtgtttc atgcgccgta tgtggcgaac cagggctggt atgatattac caaaaccttt 180
aacggcaaag atgatctgct gtgcggcgcg gcgaccgcgg gcaacatgct gcattggtgg 240
tttgatcaga acaaagatca gattaaacgc tatctggaag aacatccgga aaaacagaaa 300
attaactttc gcggcgaaca gatgtttgat gtgaaagaag cgattcgcac caaaaaccat 360
cagctggata gcaaactgtt tgaatatttt aaagaaaaag cgtttccgta tctgagcacc 420
aaacatctgg gcgtgtttcc ggatcatgtg attgatatgt ttattaacgg ctatcgcctg 480
agcctgacca accatggccc gaccccggtg aaaaaaggca gcaaagatcc gcgcggcggc 540
atttttgatg cggtgtttac ccgcggcaac cagagcaaac tgctgaccag ccgccatgat 600
tttaaagaaa aaaacctgaa agaaattagc gatctgatta aagaagaact gaccaaaggc 660
aaagcgctgg gcctgagcca tacctatgcg aacgtgcgca ttaaccatgt gattaacctg 720
tggggcgcgg attttgatgc ggaaggcaac ctgaaagcga tttatgtgac cgatagcgat 780
agcaacgcga gcattggcat gaaaaaatat tttgtgggcg tgaacaaagc gggcaaagtg 840
gcgattagcg cgaaaaaaat tgaaggcgaa aacattggcg cgcaggtgct gggcctgttt 900
accctgagca ccggccagga tagctggaac cagaccaacg gcggtggcca tcatcaccat 960
caccactaa 969
<210> 55
<211> 963
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 55
atggatgatt atcagcgcaa cgcgaccgaa gcgtatgcga aagaagtgcc gcatcagatt 60
accagcgtgt ggaccaaagg cgtgaccccg ccggaacagt ttactcaagg tgaagatgtg 120
attcatgcgc cgtatgtggc gaaccagggc tggtatgata ttaccaaagc gtttgatggc 180
aaagataacc tgctgtgcgg cgcggcgacc gcgggcaaca tgctgcattg gtggtttgat 240
cagaacaaag atcagattaa acgctatctg gaagaacatc cggaaaaaca gaaaattaac 300
tttcgcggcg aacagatgtt tgatgtgaaa aaagcgattg ataccaaaaa ccatcagctg 360
gatagcaaac tgtttaacta ttttaaagaa aaagcgtttc cgggcctgag cgcgcgccgc 420
attggcgtgt ttccggatca tgtgattgat atgtttatta acggctatcg cctgagcctg 480
accaaccatg gcccgacccc ggtgaaagaa ggcagcaaag atccgcgcgg cggcattttt 540
gatgcggtgt ttacccgcgg caaccagagc aaactgctga ccagccgcca tgattttaaa 600
aacaaaaacc tgaacgatat tagcaccatt attaaacagg aactgaccaa aggcaaagcg 660
ctgggcctga gccataccta tgcgaacgtg agcattaacc atgtgattaa cctgtggggc 720
gcggatttta acgcggaagg caacctggaa gcgatttatg tgaccgatag cgatagcaac 780
gcgagcattg gcatgaaaaa atattttgtg ggcgtgaacg cgcatggcca tgtggcgatt 840
agcgcgaaaa aaattgaagg cgaaaacatt ggcgcgcagg tgctgggcct gtttaccctg 900
agcaccggcc aggatagctg gcagaaactg agcggcggtg gccatcatca ccatcaccac 960
taa 963
<210> 56
<211> 310
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 56
Asp Ser Phe Ser Ala Asn Gln Glu Ile Arg Tyr Ser Glu Val Thr Pro
1 5 10 15
Tyr His Val Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Ala Asn
20 25 30
Phe Thr Gln Gly Glu Asp Val Phe His Ala Pro Tyr Val Ala Asn Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Thr Phe Asn Gly Lys Asp Asp Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Asp Gln Ile Lys Arg Tyr Leu Glu Glu His Pro Glu Lys Gln
85 90 95
Lys Ile Asn Phe Arg Gly Glu Gln Met Phe Asp Val Lys Glu Ala Ile
100 105 110
Asp Thr Lys Asn His Gln Leu Asp Ser Lys Leu Phe Glu Tyr Phe Lys
115 120 125
Glu Lys Ala Phe Pro Tyr Leu Ser Thr Lys His Leu Gly Val Phe Pro
130 135 140
Asp His Val Ile Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser Leu Thr
145 150 155 160
Asn His Gly Pro Thr Pro Val Lys Glu Gly Ser Lys Asp Pro Arg Gly
165 170 175
Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asp Gln Ser Lys Leu Leu
180 185 190
Thr Ser Arg His Asp Phe Lys Glu Lys Asn Leu Lys Glu Ile Ser Asp
195 200 205
Leu Ile Lys Lys Glu Leu Thr Glu Gly Lys Ala Leu Gly Leu Ser His
210 215 220
Thr Tyr Ala Asn Val Arg Ile Asn His Val Ile Asn Leu Trp Gly Ala
225 230 235 240
Asp Phe Asp Ser Asn Gly Asn Leu Lys Ala Ile Tyr Val Thr Asp Ser
245 250 255
Asp Ser Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly Val Asn
260 265 270
Ser Ala Gly Lys Val Ala Ile Ser Ala Lys Glu Ile Lys Glu Asp Asn
275 280 285
Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Thr Gly Gln Asp
290 295 300
Ser Trp Asn Gln Thr Asn
305 310
<210> 57
<211> 310
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 57
Asp Ser Phe Ser Ala Asn Gln Glu Ile Arg Tyr Ser Glu Val Thr Pro
1 5 10 15
Tyr His Val Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Ala Asn
20 25 30
Phe Thr Gln Gly Glu Asp Val Phe His Ala Pro Tyr Val Ala Asn Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Thr Phe Asn Gly Lys Asp Asp Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Asp Gln Ile Lys Arg Tyr Leu Glu Glu His Pro Glu Lys Gln
85 90 95
Lys Ile Asn Phe Lys Gly Glu Gln Met Phe Asp Val Lys Glu Ala Ile
100 105 110
Asp Thr Lys Asn His Gln Leu Asp Ser Lys Leu Phe Glu Tyr Phe Lys
115 120 125
Glu Lys Ala Phe Pro Tyr Leu Ser Thr Lys His Leu Gly Val Phe Pro
130 135 140
Asp His Val Ile Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser Leu Thr
145 150 155 160
Asn His Gly Pro Thr Pro Val Lys Arg Gly Ser Lys Asp Pro Arg Gly
165 170 175
Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Ser Lys Leu Leu
180 185 190
Thr Ser Arg His Asp Phe Lys Glu Lys Asn Leu Lys Glu Ile Ser Asp
195 200 205
Leu Ile Lys Lys Glu Leu Thr Glu Gly Lys Ala Leu Gly Leu Ser His
210 215 220
Thr Tyr Ala Asn Val Arg Ile Asn His Val Ile Asn Leu Trp Gly Ala
225 230 235 240
Asp Phe Asp Ser Asn Gly Asn Leu Lys Ala Ile Tyr Val Thr Asp Ser
245 250 255
Asp Ser Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly Val Asn
260 265 270
Ser Ala Gly Lys Val Ala Ile Ser Ala Lys Glu Ile Lys Glu Asp Asn
275 280 285
Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Thr Gly Gln Asp
290 295 300
Ser Trp Asn Gln Thr Asn
305 310
<210> 58
<211> 310
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 58
Asp Ser Phe Ser Ala Asn Gln Glu Ile Arg Tyr Ser Glu Val Thr Pro
1 5 10 15
Tyr His Val Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Ala Asn
20 25 30
Phe Thr Gln Gly Glu Asp Val Phe His Ala Pro Tyr Val Ala Asn Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Thr Phe Asn Gly Lys Asp Asp Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Asp Gln Ile Lys Arg Tyr Leu Glu Glu His Pro Glu Lys Gln
85 90 95
Lys Ile Asn Phe Arg Gly Glu Gln Met Phe Asp Val Lys Glu Ala Ile
100 105 110
Asp Thr Lys Asn His Gln Leu Asp Ser Lys Leu Phe Glu Tyr Phe Lys
115 120 125
Glu Lys Ala Phe Pro Tyr Leu Ser Thr Lys His Leu Gly Val Phe Pro
130 135 140
Asp His Val Ile Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser Leu Thr
145 150 155 160
Asn His Gly Pro Thr Pro Val Lys Lys Gly Ser Lys Asp Pro Arg Gly
165 170 175
Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Ser Lys Leu Leu
180 185 190
Thr Ser Arg His Asp Phe Lys Glu Lys Asn Leu Lys Glu Ile Ser Asp
195 200 205
Leu Ile Lys Lys Glu Leu Thr Glu Gly Lys Ala Leu Gly Leu Ser His
210 215 220
Thr Tyr Ala Asn Val Arg Ile Asn His Val Ile Asn Leu Trp Gly Ala
225 230 235 240
Asp Phe Asp Ser Asn Gly Asn Leu Lys Ala Ile Tyr Val Thr Asp Ser
245 250 255
Asp Ser Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly Val Asn
260 265 270
Lys Ala Gly Lys Val Ala Ile Ser Ala Lys Glu Ile Lys Glu Asp Asn
275 280 285
Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Thr Gly Gln Asp
290 295 300
Ser Trp Asn Gln Thr Asn
305 310
<210> 59
<211> 310
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 59
Asp Ser Phe Ser Ala Asn Gln Glu Ile Arg Tyr Ser Glu Val Thr Pro
1 5 10 15
Tyr His Val Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Ala Asn
20 25 30
Phe Thr Gln Gly Glu Asp Val Phe His Ala Pro Tyr Val Ala Asn Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Thr Phe Asn Gly Lys Asp Asp Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Asp Gln Ile Lys Arg Tyr Leu Arg Glu His Pro Glu Lys Gln
85 90 95
Lys Ile Asn Phe Asn Gly Glu Gln Met Phe Asp Val Lys Glu Ala Ile
100 105 110
Asp Thr Lys Asn His Gln Leu Asp Ser Lys Leu Phe Glu Tyr Phe Lys
115 120 125
Glu Lys Ala Phe Pro Tyr Leu Ser Thr Lys His Leu Gly Val Phe Pro
130 135 140
Asp His Val Ile Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser Leu Thr
145 150 155 160
Asn His Gly Pro Thr Pro Val Lys Glu Gly Ser Lys Asp Pro Arg Gly
165 170 175
Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Ser Lys Leu Leu
180 185 190
Thr Ser Arg His Asp Phe Lys Glu Lys Asn Leu Lys Glu Ile Ser Asp
195 200 205
Leu Ile Lys Lys Glu Leu Asp Glu Gly Lys Ala Leu Gly Leu Ser His
210 215 220
Thr Tyr Ala Asn Val Arg Ile Asn His Val Ile Asn Leu Trp Gly Ala
225 230 235 240
Asp Phe Asp Ser Asn Gly Asn Leu Lys Ala Ile Tyr Val Thr Asp Ser
245 250 255
Asp Ser Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly Val Asn
260 265 270
Ser Ala Gly Lys Val Ala Ile Ser Ala Lys Glu Ile Lys Glu Asp Asn
275 280 285
Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Thr Gly Gln Asp
290 295 300
Ser Trp Asn Gln Thr Asn
305 310
<210> 60
<211> 310
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 60
Asp Ser Phe Ser Ala Asn Gln Glu Ile Arg Tyr Ser Glu Val Thr Pro
1 5 10 15
Tyr His Val Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Ala Asn
20 25 30
Phe Thr Gln Gly Glu Asp Val Phe His Ala Pro Tyr Val Ala Asn Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Thr Phe Asn Gly Lys Asp Asp Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Asp Gln Ile Lys Arg Tyr Leu Lys Glu His Pro Glu Lys Gln
85 90 95
Lys Ile Asn Phe Asn Gly Glu Gln Met Phe Asp Val Lys Glu Ala Ile
100 105 110
Arg Thr Lys Asn His Gln Leu Asp Ser Lys Leu Phe Glu Tyr Phe Lys
115 120 125
Glu Lys Ala Phe Pro Tyr Leu Ser Thr Lys His Leu Gly Val Phe Pro
130 135 140
Asp His Val Ile Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser Leu Thr
145 150 155 160
Asn His Gly Pro Thr Pro Val Lys Glu Gly Ser Lys Asp Pro Arg Gly
165 170 175
Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Ser Lys Leu Leu
180 185 190
Thr Ser Arg His Asp Phe Lys Glu Lys Asn Leu Lys Glu Ile Ser Asp
195 200 205
Leu Ile Lys Lys Glu Leu Glu Glu Gly Lys Ala Leu Gly Leu Ser His
210 215 220
Thr Tyr Ala Asn Val Arg Ile Asn His Val Ile Asn Leu Trp Gly Ala
225 230 235 240
Asp Phe Asp Ser Asn Gly Asn Leu Lys Ala Ile Tyr Val Thr Asp Ser
245 250 255
Asp Ser Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly Val Asn
260 265 270
Lys Ala Gly Lys Val Ala Ile Ser Ala Lys Glu Ile Lys Glu Asp Asn
275 280 285
Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Thr Gly Gln Asp
290 295 300
Ser Trp Asn Gln Thr Asn
305 310
<210> 61
<211> 310
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 61
Asp Ser Phe Ser Ala Asn Gln Glu Ile Arg Tyr Ser Glu Val Thr Pro
1 5 10 15
Tyr His Val Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Ala Asn
20 25 30
Phe Thr Gln Gly Glu Asp Val Phe His Ala Pro Tyr Val Ala Asn Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Thr Phe Asn Gly Lys Asp Asp Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Asp Gln Ile Glu Arg Tyr Leu Glu Glu His Pro Glu Lys Gln
85 90 95
Lys Ile Asn Phe Asn Gly Glu Gln Met Phe Asp Val Lys Glu Ala Ile
100 105 110
Asp Thr Lys Asn His Gln Leu Asp Ser Lys Leu Phe Glu Tyr Phe Lys
115 120 125
Glu Lys Ala Phe Pro Tyr Leu Ser Thr Lys His Leu Gly Val Phe Pro
130 135 140
Asp His Val Ile Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser Leu Thr
145 150 155 160
Asn His Gly Pro Thr Pro Val Lys Glu Gly Ser Lys Asp Pro Arg Gly
165 170 175
Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Ser Lys Leu Leu
180 185 190
Thr Ser Arg His Asp Phe Lys Glu Lys Asn Leu Lys Glu Ile Ser Asp
195 200 205
Leu Ile Lys Glu Glu Leu Thr Lys Gly Lys Ala Leu Gly Leu Ser His
210 215 220
Thr Tyr Ala Asn Val Arg Ile Asn His Val Ile Asn Leu Trp Gly Ala
225 230 235 240
Asp Phe Asp Ser Asn Gly Asn Leu Lys Ala Ile Tyr Val Thr Asp Ser
245 250 255
Asp Ser Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly Val Asn
260 265 270
Ser Ala Gly Lys Val Ala Ile Ser Ala Lys Glu Ile Lys Glu Lys Asn
275 280 285
Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Thr Gly Gln Lys
290 295 300
Ser Trp Asn Gln Thr Asn
305 310
<210> 62
<211> 310
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 62
Asp Ser Phe Ser Ala Asn Gln Glu Ile Arg Tyr Ser Glu Val Thr Pro
1 5 10 15
Tyr His Val Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Ala Asn
20 25 30
Phe Thr Gln Gly Glu Asp Val Phe His Ala Pro Tyr Val Ala Asn Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Thr Phe Asn Gly Lys Asp Asp Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Asp Gln Ile Lys Arg Tyr Leu Lys Glu His Pro Glu Lys Gln
85 90 95
Lys Ile Asn Phe Arg Gly Glu Gln Met Phe Asp Val Lys Glu Ala Ile
100 105 110
Arg Thr Lys Asn His Gln Leu Asp Ser Lys Leu Phe Glu Tyr Phe Lys
115 120 125
Glu Lys Ala Phe Pro Tyr Leu Ser Thr Lys His Leu Gly Val Phe Pro
130 135 140
Asp His Val Ile Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser Leu Thr
145 150 155 160
Asn His Gly Pro Thr Pro Val Lys Glu Gly Ser Lys Asp Pro Arg Gly
165 170 175
Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Ser Lys Leu Leu
180 185 190
Thr Ser Arg His Asp Phe Lys Glu Lys Asn Leu Lys Glu Ile Ser Asp
195 200 205
Leu Ile Lys Ser Glu Leu Glu Asn Gly Lys Ala Leu Gly Leu Ser His
210 215 220
Thr Tyr Ala Asn Val Arg Ile Asn His Val Ile Asn Leu Trp Gly Ala
225 230 235 240
Asp Phe Asp Ser Asn Gly Asn Leu Lys Ala Ile Tyr Val Thr Asp Ser
245 250 255
Asp Ser Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly Val Asn
260 265 270
Lys Ala Gly Lys Val Ala Ile Ser Ala Lys Glu Ile Lys Glu Asp Asn
275 280 285
Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Thr Gly Gln Asp
290 295 300
Ser Trp Asn Gln Thr Asn
305 310
<210> 63
<211> 310
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 63
Asp Ser Phe Ser Ala Asn Gln Glu Ile Arg Tyr Ser Glu Val Thr Pro
1 5 10 15
Tyr His Val Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Ala Asn
20 25 30
Phe Thr Gln Gly Glu Asp Val Phe His Ala Pro Tyr Val Ala Asn Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Thr Phe Asn Gly Lys Asp Asp Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Asp Gln Ile Lys Arg Tyr Leu Lys Glu His Pro Glu Lys Gln
85 90 95
Lys Ile Asn Phe Arg Gly Glu Gln Met Phe Asp Val Lys Glu Ala Ile
100 105 110
Arg Thr Lys Asn His Gln Leu Asp Ser Lys Leu Phe Glu Tyr Phe Lys
115 120 125
Glu Lys Ala Phe Pro Tyr Leu Ser Thr Lys His Leu Gly Val Phe Pro
130 135 140
Asp His Val Ile Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser Leu Thr
145 150 155 160
Asn His Gly Pro Thr Pro Val Lys Lys Gly Ser Lys Asp Pro Arg Gly
165 170 175
Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Ser Lys Leu Leu
180 185 190
Thr Ser Arg His Asp Phe Lys Glu Lys Asn Leu Lys Glu Ile Ser Asp
195 200 205
Leu Ile Lys Lys Glu Leu Glu Glu Gly Lys Ala Leu Gly Leu Ser His
210 215 220
Thr Tyr Ala Asn Val Arg Ile Asn His Val Ile Asn Leu Trp Gly Ala
225 230 235 240
Asp Phe Asp Ser Asn Gly Asn Leu Lys Ala Ile Tyr Val Thr Asp Ser
245 250 255
Asp Ser Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly Val Asn
260 265 270
Ser Ala Gly Lys Val Ala Ile Ser Ala Lys Glu Ile Lys Glu Asp Asn
275 280 285
Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Thr Gly Gln Asp
290 295 300
Ser Trp Asn Gln Thr Asn
305 310
<210> 64
<211> 310
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 64
Asp Ser Phe Ser Ala Asn Gln Glu Ile Arg Tyr Ser Glu Val Thr Pro
1 5 10 15
Tyr His Val Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Ala Asn
20 25 30
Phe Thr Gln Gly Glu Asp Val Phe His Ala Pro Tyr Val Ala Asn Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Thr Phe Asn Gly Lys Asp Asp Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Asp Gln Ile Glu Arg Tyr Leu Glu Glu His Pro Glu Lys Gln
85 90 95
Lys Ile Asn Phe Arg Gly Glu Gln Met Phe Asp Val Lys Glu Ala Ile
100 105 110
Asp Thr Lys Asn His Gln Leu Asp Ser Lys Leu Phe Glu Tyr Phe Lys
115 120 125
Glu Lys Ala Phe Pro Tyr Leu Ser Thr Lys His Leu Gly Val Phe Pro
130 135 140
Asp His Val Ile Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser Leu Thr
145 150 155 160
Asn His Gly Pro Thr Pro Val Lys Lys Gly Ser Lys Asp Pro Arg Gly
165 170 175
Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Ser Lys Leu Leu
180 185 190
Thr Ser Arg His Asp Phe Lys Glu Lys Asn Leu Lys Glu Ile Ser Asp
195 200 205
Leu Ile Lys Glu Glu Leu Thr Lys Gly Lys Ala Leu Gly Leu Ser His
210 215 220
Thr Tyr Ala Asn Val Arg Ile Asn His Val Ile Asn Leu Trp Gly Ala
225 230 235 240
Asp Phe Asp Ser Asn Gly Asn Leu Lys Ala Ile Tyr Val Thr Asp Ser
245 250 255
Asp Ser Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly Val Asn
260 265 270
Ser Ala Gly Lys Val Ala Ile Ser Ala Lys Glu Ile Lys Glu Asp Asn
275 280 285
Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Thr Gly Gln Lys
290 295 300
Ser Trp Asn Gln Thr Asn
305 310
<210> 65
<211> 306
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 65
Asp Ser Phe Ser Ala Asn Gln Glu Ile Arg Tyr Ser Glu Val Thr Pro
1 5 10 15
Tyr His Val Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Ala Asn
20 25 30
Phe Thr Gln Gly Glu Asp Val Phe His Ala Pro Tyr Val Ala Asn Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Thr Phe Asn Gly Lys Asp Asp Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Asp Gln Ile Lys Arg Tyr Leu Glu Glu His Pro Glu Lys Gln
85 90 95
Lys Ile Asn Phe Asn Gly Glu Gln Met Phe Asp Val Lys Glu Ala Ile
100 105 110
Asp Thr Lys Asn His Gln Leu Asp Ser Lys Leu Phe Glu Tyr Phe Lys
115 120 125
Glu Lys Ala Phe Pro Tyr Leu Ser Thr Lys His Leu Gly Val Phe Pro
130 135 140
Asp His Val Ile Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser Leu Thr
145 150 155 160
Asn His Gly Pro Thr Pro Val Lys Glu Gly Ser Lys Asp Pro Arg Gly
165 170 175
Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asp Gln Ser Lys Leu Leu
180 185 190
Thr Ser Arg His Asp Phe Lys Glu Lys Asn Leu Lys Glu Ile Ser Asp
195 200 205
Leu Ile Lys Lys Glu Leu Thr Glu Gly Lys Ala Leu Gly Leu Ser His
210 215 220
Thr Tyr Ala Asn Val Arg Ile Asn His Val Ile Asn Leu Trp Gly Ala
225 230 235 240
Asp Phe Asp Ser Asn Gly Asn Leu Lys Ala Ile Tyr Val Thr Asp Ser
245 250 255
Asp Ser Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly Val Asn
260 265 270
Ser Ala Gly Lys Val Ala Ile Ser Ala Lys Glu Ile Lys Glu Asp Asn
275 280 285
Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Thr Gly Gln Asp
290 295 300
Ser Trp
305
<210> 66
<211> 290
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 66
Ser Val Trp Thr Lys Gly Val Thr Pro Pro Ala Asn Phe Thr Gln Gly
1 5 10 15
Glu Asp Val Phe His Ala Pro Tyr Val Ala Asn Gln Gly Trp Tyr Asp
20 25 30
Ile Thr Lys Thr Phe Asn Gly Lys Asp Asp Leu Leu Cys Gly Ala Ala
35 40 45
Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln Asn Lys Asp Gln
50 55 60
Ile Lys Arg Tyr Leu Glu Glu His Pro Glu Lys Gln Lys Ile Asn Phe
65 70 75 80
Asn Gly Glu Gln Met Phe Asp Val Lys Glu Ala Ile Asp Thr Lys Asn
85 90 95
His Gln Leu Asp Ser Lys Leu Phe Glu Tyr Phe Lys Glu Lys Ala Phe
100 105 110
Pro Tyr Leu Ser Thr Lys His Leu Gly Val Phe Pro Asp His Val Ile
115 120 125
Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser Leu Thr Asn His Gly Pro
130 135 140
Thr Pro Val Lys Glu Gly Ser Lys Asp Pro Arg Gly Gly Ile Phe Asp
145 150 155 160
Ala Val Phe Thr Arg Gly Asp Gln Ser Lys Leu Leu Thr Ser Arg His
165 170 175
Asp Phe Lys Glu Lys Asn Leu Lys Glu Ile Ser Asp Leu Ile Lys Lys
180 185 190
Glu Leu Thr Glu Gly Lys Ala Leu Gly Leu Ser His Thr Tyr Ala Asn
195 200 205
Val Arg Ile Asn His Val Ile Asn Leu Trp Gly Ala Asp Phe Asp Ser
210 215 220
Asn Gly Asn Leu Lys Ala Ile Tyr Val Thr Asp Ser Asp Ser Asn Ala
225 230 235 240
Ser Ile Gly Met Lys Lys Tyr Phe Val Gly Val Asn Ser Ala Gly Lys
245 250 255
Val Ala Ile Ser Ala Lys Glu Ile Lys Glu Asp Asn Ile Gly Ala Gln
260 265 270
Val Leu Gly Leu Phe Thr Leu Ser Thr Gly Gln Asp Ser Trp Asn Gln
275 280 285
Thr Asn
290
<210> 67
<211> 290
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 67
Ser Val Trp Thr Lys Gly Val Thr Pro Pro Ala Asn Phe Thr Gln Gly
1 5 10 15
Glu Asp Val Phe His Ala Pro Tyr Val Ala Asn Gln Gly Trp Tyr Asp
20 25 30
Ile Thr Lys Thr Phe Asn Gly Lys Asp Asp Leu Leu Cys Gly Ala Ala
35 40 45
Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln Asn Lys Asp Gln
50 55 60
Ile Lys Arg Tyr Leu Glu Glu His Pro Glu Lys Gln Lys Ile Asn Phe
65 70 75 80
Lys Gly Glu Gln Met Phe Asp Val Lys Glu Ala Ile Asp Thr Lys Asn
85 90 95
His Gln Leu Asp Ser Lys Leu Phe Glu Tyr Phe Lys Glu Lys Ala Phe
100 105 110
Pro Tyr Leu Ser Thr Lys His Leu Gly Val Phe Pro Asp His Val Ile
115 120 125
Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser Leu Thr Asn His Gly Pro
130 135 140
Thr Pro Val Lys Glu Gly Ser Lys Asp Pro Arg Gly Gly Ile Phe Asp
145 150 155 160
Ala Val Phe Thr Arg Gly Asn Gln Ser Lys Leu Leu Thr Ser Arg His
165 170 175
Asp Phe Lys Glu Lys Asn Leu Lys Glu Ile Ser Asp Leu Ile Lys Lys
180 185 190
Glu Leu Thr Glu Gly Lys Ala Leu Gly Leu Ser His Thr Tyr Ala Asn
195 200 205
Val Arg Ile Asn His Val Ile Asn Leu Trp Gly Ala Asp Phe Asp Ser
210 215 220
Asn Gly Asn Leu Lys Ala Ile Tyr Val Thr Asp Ser Asp Ser Asn Ala
225 230 235 240
Ser Ile Gly Met Lys Lys Tyr Phe Val Gly Val Asn Ser Ala Gly Lys
245 250 255
Val Ala Ile Ser Ala Lys Glu Ile Lys Glu Asp Asn Ile Gly Ala Gln
260 265 270
Val Leu Gly Leu Phe Thr Leu Ser Thr Gly Gln Asp Ser Trp Asn Gln
275 280 285
Thr Asn
290
<210> 68
<211> 290
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 68
Ser Val Trp Thr Lys Gly Val Thr Pro Pro Ala Asn Phe Thr Gln Gly
1 5 10 15
Glu Asp Val Phe His Ala Pro Tyr Val Ala Asn Gln Gly Trp Tyr Asp
20 25 30
Ile Thr Lys Thr Phe Asn Gly Lys Asp Asp Leu Leu Cys Gly Ala Ala
35 40 45
Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln Asn Lys Asp Gln
50 55 60
Ile Glu Arg Tyr Leu Glu Glu His Pro Glu Lys Gln Lys Ile Asn Phe
65 70 75 80
Lys Gly Glu Gln Met Phe Asp Val Lys Lys Ala Ile Asp Thr Lys Asn
85 90 95
His Gln Leu Asp Ser Lys Leu Phe Glu Tyr Phe Lys Glu Lys Ala Phe
100 105 110
Pro Tyr Leu Ser Thr Lys His Leu Gly Val Phe Pro Asp His Val Ile
115 120 125
Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser Leu Thr Asn His Gly Pro
130 135 140
Thr Pro Val Lys Glu Gly Ser Lys Asp Pro Arg Gly Gly Ile Phe Asp
145 150 155 160
Ala Val Phe Thr Arg Gly Asn Gln Ser Lys Leu Leu Thr Ser Arg His
165 170 175
Asp Phe Lys Glu Lys Asn Leu Lys Glu Ile Ser Asp Leu Ile Lys Glu
180 185 190
Glu Leu Thr Lys Gly Lys Ala Leu Gly Leu Ser His Thr Tyr Ala Asn
195 200 205
Val Arg Ile Asn His Val Ile Asn Leu Trp Gly Ala Asp Phe Asp Ser
210 215 220
Asn Gly Asn Leu Lys Ala Ile Tyr Val Thr Asp Ser Asp Ser Asn Ala
225 230 235 240
Ser Ile Gly Met Lys Lys Tyr Phe Val Gly Val Asn Ser Ala Gly Lys
245 250 255
Val Ala Ile Ser Ala Lys Glu Ile Lys Glu Asp Asn Ile Gly Ala Gln
260 265 270
Val Leu Gly Leu Phe Thr Leu Ser Thr Gly Gln Lys Ser Trp Asn Gln
275 280 285
Thr Asn
290
<210> 69
<211> 319
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 69
Asp Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val Pro
1 5 10 15
His Gln Ile Thr Ser Val Trp Thr Lys Gly Val Thr Pro Pro Ala Asn
20 25 30
Phe Thr Gln Gly Glu Asp Val Phe His Ala Pro Tyr Val Ala Asn Gln
35 40 45
Gly Trp Tyr Asp Ile Thr Lys Thr Phe Asn Gly Lys Asp Asp Leu Leu
50 55 60
Cys Gly Ala Ala Thr Ala Gly Asn Met Leu His Trp Trp Phe Asp Gln
65 70 75 80
Asn Lys Asp Gln Ile Glu Arg Tyr Leu Glu Glu His Pro Glu Lys Gln
85 90 95
Lys Ile Asn Phe Lys Gly Glu Gln Met Phe Asp Val Lys Lys Ala Ile
100 105 110
Asp Thr Lys Asn His Gln Leu Asp Ser Lys Leu Phe Glu Tyr Phe Lys
115 120 125
Glu Lys Ala Phe Pro Tyr Leu Ser Thr Lys His Leu Gly Val Phe Pro
130 135 140
Asp His Val Ile Asp Met Phe Ile Asn Gly Tyr Arg Leu Ser Leu Thr
145 150 155 160
Asn His Gly Pro Thr Pro Val Lys Glu Gly Ser Lys Asp Pro Arg Gly
165 170 175
Gly Ile Phe Asp Ala Val Phe Thr Arg Gly Asn Gln Ser Lys Leu Leu
180 185 190
Thr Ser Arg His Asp Phe Lys Glu Lys Asn Leu Lys Glu Ile Ser Asp
195 200 205
Leu Ile Lys Glu Glu Leu Thr Lys Gly Lys Ala Leu Gly Leu Ser His
210 215 220
Thr Tyr Ala Asn Val Arg Ile Asn His Val Ile Asn Leu Trp Gly Ala
225 230 235 240
Asp Phe Asp Ser Asn Gly Asn Leu Lys Ala Ile Tyr Val Thr Asp Ser
245 250 255
Asp Ser Asn Ala Ser Ile Gly Met Lys Lys Tyr Phe Val Gly Val Asn
260 265 270
Ser Ala Gly Lys Val Ala Ile Ser Ala Lys Glu Ile Lys Glu Asp Asn
275 280 285
Ile Gly Ala Gln Val Leu Gly Leu Phe Thr Leu Ser Thr Gly Gln Lys
290 295 300
Ser Trp Asn Gln Thr Asn Gly Gly Gly His His His His His His
305 310 315
<210> 70
<211> 4
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 70
Asn Gln Thr Asn
1
<210> 71
<211> 20
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 71
Asp Ser Phe Ser Ala Asn Gln Glu Ile Arg Tyr Ser Glu Val Thr Pro
1 5 10 15
Tyr His Val Thr
20
<210> 72
<211> 963
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 72
atggatagtt tttctgctaa tcaagagatt agatattcgg aagtaacacc ttatcacgtt 60
acttccgttt ggaccaaagg agttactcct ccagcaaact tcactcaagg tgaagatgtt 120
tttcacgctc cttatgttgc taaccaagga tggtatgata ttaccaaaac attcaatgga 180
aaagacgatc ttctttgcgg ggctgccaca gcagggaata tgcttcactg gtggttcgat 240
caaaacaaag accaaattaa acgttatttg gaagagcatc cagaaaagca aaaaataaac 300
ttcaatggcg aacagatgtt tgacgtaaaa gaagctatcg acactaaaaa ccaccagcta 360
gatagtaaat tatttgaata ttttaaagaa aaagctttcc cttatctatc tactaaacac 420
ctaggagttt tccctgatca tgtaattgat atgttcatta acggctaccg ccttagtcta 480
actaaccacg gtccaacgcc agtaaaagaa ggtagtaaag atccccgagg tggtattttt 540
gacgccgtat ttacaagagg tgatcaaagt aagctattga caagtcgtca tgattttaaa 600
gaaaaaaatc tcaaagaaat cagtgatctc attaagaaag agttaaccga aggcaaggct 660
ctaggcctat cacacaccta cgctaacgta cgcatcaacc atgttataaa cctgtgggga 720
gctgactttg attctaacgg gaaccttaaa gctatttatg taacagactc tgatagtaat 780
gcatctattg gtatgaagaa atactttgtt ggtgttaatt ccgctggaaa agtagctatt 840
tctgctaaag aaataaaaga agataatata ggtgctcaag tactagggtt atttacactt 900
tcaacagggc aagatagttg gaatcagacc aatggcggtg gccatcatca ccatcaccac 960
taa 963
<210> 73
<211> 963
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 73
atggatagtt tttctgctaa tcaagagatt agatattcgg aagtaacacc ttatcacgtt 60
acttccgttt ggaccaaagg agttactcct ccagcaaact tcactcaagg tgaagatgtt 120
tttcacgctc cttatgttgc taaccaagga tggtatgata ttaccaaaac attcaatgga 180
aaagacgatc ttctttgcgg ggctgccaca gcagggaata tgcttcactg gtggttcgat 240
caaaacaaag accaaattaa acgttatttg gaagagcatc cagaaaagca aaaaataaac 300
ttccgtggcg aacagatgtt tgacgtaaaa gaagctatcg acactaaaaa ccaccagcta 360
gatagtaaat tatttgaata ttttaaagaa aaagctttcc cttatctatc tactaaacac 420
ctaggagttt tccctgatca tgtaattgat atgttcatta acggctaccg ccttagtcta 480
actaaccacg gtccaacgcc agtaaaagaa ggtagtaaag atccccgagg tggtattttt 540
gacgccgtat ttacaagagg tgatcaaagt aagctattga caagtcgtca tgattttaaa 600
gaaaaaaatc tcaaagaaat cagtgatctc attaagaaag agttaaccga aggcaaggct 660
ctaggcctat cacacaccta cgctaacgta cgcatcaacc atgttataaa cctgtgggga 720
gctgactttg attctaacgg gaaccttaaa gctatttatg taacagactc tgatagtaat 780
gcatctattg gtatgaagaa atactttgtt ggtgttaatt ccgctggaaa agtagctatt 840
tctgctaaag aaataaaaga agataatata ggtgctcaag tactagggtt atttacactt 900
tcaacagggc aagatagttg gaatcagacc aatggcggtg gccatcatca ccatcaccac 960
taa 963
<210> 74
<211> 963
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 74
atggatagtt tttctgctaa tcaagagatt agatattcgg aagtaacacc ttatcacgtt 60
acttccgttt ggaccaaagg agttactcct ccagcaaact tcactcaagg tgaagatgtt 120
tttcacgctc cttatgttgc taaccaagga tggtatgata ttaccaaaac attcaatgga 180
aaagacgatc ttctttgcgg ggctgccaca gcagggaata tgcttcactg gtggttcgat 240
caaaacaaag accaaattaa acgttatttg gaagagcatc cagaaaagca aaaaataaac 300
ttcaaaggcg aacagatgtt tgacgtaaaa gaagctatcg acactaaaaa ccaccagcta 360
gatagtaaat tatttgaata ttttaaagaa aaagctttcc cttatctatc tactaaacac 420
ctaggagttt tccctgatca tgtaattgat atgttcatta acggctaccg ccttagtcta 480
actaaccacg gtccaacgcc agtaaaacgt ggtagtaaag atccccgagg tggtattttt 540
gacgccgtat ttacaagagg taaccaaagt aagctattga caagtcgtca tgattttaaa 600
gaaaaaaatc tcaaagaaat cagtgatctc attaagaaag agttaaccga aggcaaggct 660
ctaggcctat cacacaccta cgctaacgta cgcatcaacc atgttataaa cctgtgggga 720
gctgactttg attctaacgg gaaccttaaa gctatttatg taacagactc tgatagtaat 780
gcatctattg gtatgaagaa atactttgtt ggtgttaatt ccgctggaaa agtagctatt 840
tctgctaaag aaataaaaga agataatata ggtgctcaag tactagggtt atttacactt 900
tcaacagggc aagatagttg gaatcagacc aatggcggtg gccatcatca ccatcaccac 960
taa 963
<210> 75
<211> 963
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 75
atggatagtt tttctgctaa tcaagagatt agatattcgg aagtaacacc ttatcacgtt 60
acttccgttt ggaccaaagg agttactcct ccagcaaact tcactcaagg tgaagatgtt 120
tttcacgctc cttatgttgc taaccaagga tggtatgata ttaccaaaac attcaatgga 180
aaagacgatc ttctttgcgg ggctgccaca gcagggaata tgcttcactg gtggttcgat 240
caaaacaaag accaaattaa acgttatttg gaagagcatc cagaaaagca aaaaataaac 300
ttccgtggcg aacagatgtt tgacgtaaaa gaagctatcg acactaaaaa ccaccagcta 360
gatagtaaat tatttgaata ttttaaagaa aaagctttcc cttatctatc tactaaacac 420
ctaggagttt tccctgatca tgtaattgat atgttcatta acggctaccg ccttagtcta 480
actaaccacg gtccaacgcc agtaaaaaaa ggtagtaaag atccccgagg tggtattttt 540
gacgccgtat ttacaagagg taaccaaagt aagctattga caagtcgtca tgattttaaa 600
gaaaaaaatc tcaaagaaat cagtgatctc attaagaaag agttaaccga aggcaaggct 660
ctaggcctat cacacaccta cgctaacgta cgcatcaacc atgttataaa cctgtgggga 720
gctgactttg attctaacgg gaaccttaaa gctatttatg taacagactc tgatagtaat 780
gcatctattg gtatgaagaa atactttgtt ggtgttaata aagctggaaa agtagctatt 840
tctgctaaag aaataaaaga agataatata ggtgctcaag tactagggtt atttacactt 900
tcaacagggc aagatagttg gaatcagacc aatggcggtg gccatcatca ccatcaccac 960
taa 963
<210> 76
<211> 963
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 76
atggatagtt tttctgctaa tcaagagatt agatattcgg aagtaacacc ttatcacgtt 60
acttccgttt ggaccaaagg agttactcct ccagcaaact tcactcaagg tgaagatgtt 120
tttcacgctc cttatgttgc taaccaagga tggtatgata ttaccaaaac attcaatgga 180
aaagacgatc ttctttgcgg ggctgccaca gcagggaata tgcttcactg gtggttcgat 240
caaaacaaag accaaattaa acgttatttg cgtgagcatc cagaaaagca aaaaataaac 300
ttcaatggcg aacagatgtt tgacgtaaaa gaagctatcg acactaaaaa ccaccagcta 360
gatagtaaat tatttgaata ttttaaagaa aaagctttcc cttatctatc tactaaacac 420
ctaggagttt tccctgatca tgtaattgat atgttcatta acggctaccg ccttagtcta 480
actaaccacg gtccaacgcc agtaaaagaa ggtagtaaag atccccgagg tggtattttt 540
gacgccgtat ttacaagagg taaccaaagt aagctattga caagtcgtca tgattttaaa 600
gaaaaaaatc tcaaagaaat cagtgatctc attaagaaag agttagatga aggcaaggct 660
ctaggcctat cacacaccta cgctaacgta cgcatcaacc atgttataaa cctgtgggga 720
gctgactttg attctaacgg gaaccttaaa gctatttatg taacagactc tgatagtaat 780
gcatctattg gtatgaagaa atactttgtt ggtgttaatt ccgctggaaa agtagctatt 840
tctgctaaag aaataaaaga agataatata ggtgctcaag tactagggtt atttacactt 900
tcaacagggc aagatagttg gaatcagacc aatggcggtg gccatcatca ccatcaccac 960
taa 963
<210> 77
<211> 963
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 77
atggatagtt tttctgctaa tcaagagatt agatattcgg aagtaacacc ttatcacgtt 60
acttccgttt ggaccaaagg agttactcct ccagcaaact tcactcaagg tgaagatgtt 120
tttcacgctc cttatgttgc taaccaagga tggtatgata ttaccaaaac attcaatgga 180
aaagacgatc ttctttgcgg ggctgccaca gcagggaata tgcttcactg gtggttcgat 240
caaaacaaag accaaattaa acgttatttg aaagagcatc cagaaaagca aaaaataaac 300
ttcaatggcg aacagatgtt tgacgtaaaa gaagctatcc gtactaaaaa ccaccagcta 360
gatagtaaat tatttgaata ttttaaagaa aaagctttcc cttatctatc tactaaacac 420
ctaggagttt tccctgatca tgtaattgat atgttcatta acggctaccg ccttagtcta 480
actaaccacg gtccaacgcc agtaaaagaa ggtagtaaag atccccgagg tggtattttt 540
gacgccgtat ttacaagagg taaccaaagt aagctattga caagtcgtca tgattttaaa 600
gaaaaaaatc tcaaagaaat cagtgatctc attaagaaag agttagaaga aggcaaggct 660
ctaggcctat cacacaccta cgctaacgta cgcatcaacc atgttataaa cctgtgggga 720
gctgactttg attctaacgg gaaccttaaa gctatttatg taacagactc tgatagtaat 780
gcatctattg gtatgaagaa atactttgtt ggtgttaata aagctggaaa agtagctatt 840
tctgctaaag aaataaaaga agataatata ggtgctcaag tactagggtt atttacactt 900
tcaacagggc aagatagttg gaatcagacc aatggcggtg gccatcatca ccatcaccac 960
taa 963
<210> 78
<211> 963
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 78
atggatagtt tttctgctaa tcaagagatt agatattcgg aagtaacacc ttatcacgtt 60
acttccgttt ggaccaaagg agttactcct ccagcaaact tcactcaagg tgaagatgtt 120
tttcacgctc cttatgttgc taaccaagga tggtatgata ttaccaaaac attcaatgga 180
aaagacgatc ttctttgcgg ggctgccaca gcagggaata tgcttcactg gtggttcgat 240
caaaacaaag accaaattga acgttatttg gaagagcatc cagaaaagca aaaaataaac 300
ttcaatggcg aacagatgtt tgacgtaaaa gaagctatcg acactaaaaa ccaccagcta 360
gatagtaaat tatttgaata ttttaaagaa aaagctttcc cttatctatc tactaaacac 420
ctaggagttt tccctgatca tgtaattgat atgttcatta acggctaccg ccttagtcta 480
actaaccacg gtccaacgcc agtaaaagaa ggtagtaaag atccccgagg tggtattttt 540
gacgccgtat ttacaagagg taaccaaagt aagctattga caagtcgtca tgattttaaa 600
gaaaaaaatc tcaaagaaat cagtgatctc attaaggaag agttaaccaa aggcaaggct 660
ctaggcctat cacacaccta cgctaacgta cgcatcaacc atgttataaa cctgtgggga 720
gctgactttg attctaacgg gaaccttaaa gctatttatg taacagactc tgatagtaat 780
gcatctattg gtatgaagaa atactttgtt ggtgttaatt ccgctggaaa agtagctatt 840
tctgctaaag aaataaaaga aaaaaatata ggtgctcaag tactagggtt atttacactt 900
tcaacagggc aaaaaagttg gaatcagacc aatggcggtg gccatcatca ccatcaccac 960
taa 963
<210> 79
<211> 963
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 79
atggatagtt tttctgctaa tcaagagatt agatattcgg aagtaacacc ttatcacgtt 60
acttccgttt ggaccaaagg agttactcct ccagcaaact tcactcaagg tgaagatgtt 120
tttcacgctc cttatgttgc taaccaagga tggtatgata ttaccaaaac attcaatgga 180
aaagacgatc ttctttgcgg ggctgccaca gcagggaata tgcttcactg gtggttcgat 240
caaaacaaag accaaattaa acgttatttg aaagagcatc cagaaaagca aaaaataaac 300
ttccgtggcg aacagatgtt tgacgtaaaa gaagctatcc gtactaaaaa ccaccagcta 360
gatagtaaat tatttgaata ttttaaagaa aaagctttcc cttatctatc tactaaacac 420
ctaggagttt tccctgatca tgtaattgat atgttcatta acggctaccg ccttagtcta 480
actaaccacg gtccaacgcc agtaaaagaa ggtagtaaag atccccgagg tggtattttt 540
gacgccgtat ttacaagagg taaccaaagt aagctattga caagtcgtca tgattttaaa 600
gaaaaaaatc tcaaagaaat cagtgatctc attaagagtg agttagaaaa cggcaaggct 660
ctaggcctat cacacaccta cgctaacgta cgcatcaacc atgttataaa cctgtgggga 720
gctgactttg attctaacgg gaaccttaaa gctatttatg taacagactc tgatagtaat 780
gcatctattg gtatgaagaa atactttgtt ggtgttaata aagctggaaa agtagctatt 840
tctgctaaag aaataaaaga agataatata ggtgctcaag tactagggtt atttacactt 900
tcaacagggc aagatagttg gaatcagacc aatggcggtg gccatcatca ccatcaccac 960
taa 963
<210> 80
<211> 963
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 80
atggatagtt tttctgctaa tcaagagatt agatattcgg aagtaacacc ttatcacgtt 60
acttccgttt ggaccaaagg agttactcct ccagcaaact tcactcaagg tgaagatgtt 120
tttcacgctc cttatgttgc taaccaagga tggtatgata ttaccaaaac attcaatgga 180
aaagacgatc ttctttgcgg ggctgccaca gcagggaata tgcttcactg gtggttcgat 240
caaaacaaag accaaattaa acgttatttg aaagagcatc cagaaaagca aaaaataaac 300
ttccgtggcg aacagatgtt tgacgtaaaa gaagctatcc gtactaaaaa ccaccagcta 360
gatagtaaat tatttgaata ttttaaagaa aaagctttcc cttatctatc tactaaacac 420
ctaggagttt tccctgatca tgtaattgat atgttcatta acggctaccg ccttagtcta 480
actaaccacg gtccaacgcc agtaaaaaaa ggtagtaaag atccccgagg tggtattttt 540
gacgccgtat ttacaagagg taaccaaagt aagctattga caagtcgtca tgattttaaa 600
gaaaaaaatc tcaaagaaat cagtgatctc attaagaaag agttagaaga aggcaaggct 660
ctaggcctat cacacaccta cgctaacgta cgcatcaacc atgttataaa cctgtgggga 720
gctgactttg attctaacgg gaaccttaaa gctatttatg taacagactc tgatagtaat 780
gcatctattg gtatgaagaa atactttgtt ggtgttaatt ccgctggaaa agtagctatt 840
tctgctaaag aaataaaaga agataatata ggtgctcaag tactagggtt atttacactt 900
tcaacagggc aagatagttg gaatcagacc aatggcggtg gccatcatca ccatcaccac 960
taa 963
<210> 81
<211> 963
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 81
atggatagtt tttctgctaa tcaagagatt agatattcgg aagtaacacc ttatcacgtt 60
acttccgttt ggaccaaagg agttactcct ccagcaaact tcactcaagg tgaagatgtt 120
tttcacgctc cttatgttgc taaccaagga tggtatgata ttaccaaaac attcaatgga 180
aaagacgatc ttctttgcgg ggctgccaca gcagggaata tgcttcactg gtggttcgat 240
caaaacaaag accaaattga acgttatttg gaagagcatc cagaaaagca aaaaataaac 300
ttccgtggcg aacagatgtt tgacgtaaaa gaagctatcg acactaaaaa ccaccagcta 360
gatagtaaat tatttgaata ttttaaagaa aaagctttcc cttatctatc tactaaacac 420
ctaggagttt tccctgatca tgtaattgat atgttcatta acggctaccg ccttagtcta 480
actaaccacg gtccaacgcc agtaaaaaaa ggtagtaaag atccccgagg tggtattttt 540
gacgccgtat ttacaagagg taaccaaagt aagctattga caagtcgtca tgattttaaa 600
gaaaaaaatc tcaaagaaat cagtgatctc attaaggaag agttaaccaa aggcaaggct 660
ctaggcctat cacacaccta cgctaacgta cgcatcaacc atgttataaa cctgtgggga 720
gctgactttg attctaacgg gaaccttaaa gctatttatg taacagactc tgatagtaat 780
gcatctattg gtatgaagaa atactttgtt ggtgttaatt ccgctggaaa agtagctatt 840
tctgctaaag aaataaaaga agataatata ggtgctcaag tactagggtt atttacactt 900
tcaacagggc aaaaaagttg gaatcagacc aatggcggtg gccatcatca ccatcaccac 960
taa 963
<210> 82
<211> 957
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 82
atggatagtt tttctgctaa tcaagagatt agatattcgg aagtaacacc ttatcacgtt 60
acttccgttt ggaccaaagg agttactcct ccagcaaact tcactcaagg tgaagatgtt 120
tttcacgctc cttatgttgc taaccaagga tggtatgata ttaccaaaac attcaatgga 180
aaagacgatc ttctttgcgg ggctgccaca gcagggaata tgcttcactg gtggttcgat 240
caaaacaaag accaaattaa acgttatttg gaagagcatc cagaaaagca aaaaataaac 300
ttcaatggcg aacagatgtt tgacgtaaaa gaagctatcg acactaaaaa ccaccagcta 360
gatagtaaat tatttgaata ttttaaagaa aaagctttcc cttatctatc tactaaacac 420
ctaggagttt tccctgatca tgtaattgat atgttcatta acggctaccg ccttagtcta 480
actaaccacg gtccaacgcc agtaaaagaa ggtagtaaag atccccgagg tggtattttt 540
gacgccgtat ttacaagagg tgatcaaagt aagctattga caagtcgtca tgattttaaa 600
gaaaaaaatc tcaaagaaat cagtgatctc attaagaaag agttaaccga aggcaaggct 660
ctaggcctat cacacaccta cgctaacgta cgcatcaacc atgttataaa cctgtgggga 720
gctgactttg attctaacgg gaaccttaaa gctatttatg taacagactc tgatagtaat 780
gcatctattg gtatgaagaa atactttgtt ggtgttaatt ccgctggaaa agtagctatt 840
tctgctaaag aaataaaaga agataatata ggtgctcaag tactagggtt atttacactt 900
tcaacagggc aagatagttg gggtggcggc ggtggccatc atcaccatca ccactaa 957
<210> 83
<211> 903
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 83
atgtccgttt ggaccaaagg agttactcct ccagcaaact tcactcaagg tgaagatgtt 60
tttcacgctc cttatgttgc taaccaagga tggtatgata ttaccaaaac attcaatgga 120
aaagacgatc ttctttgcgg ggctgccaca gcagggaata tgcttcactg gtggttcgat 180
caaaacaaag accaaattaa acgttatttg gaagagcatc cagaaaagca aaaaataaac 240
ttcaatggcg aacagatgtt tgacgtaaaa gaagctatcg acactaaaaa ccaccagcta 300
gatagtaaat tatttgaata ttttaaagaa aaagctttcc cttatctatc tactaaacac 360
ctaggagttt tccctgatca tgtaattgat atgttcatta acggctaccg ccttagtcta 420
actaaccacg gtccaacgcc agtaaaagaa ggtagtaaag atccccgagg tggtattttt 480
gacgccgtat ttacaagagg tgatcaaagt aagctattga caagtcgtca tgattttaaa 540
gaaaaaaatc tcaaagaaat cagtgatctc attaagaaag agttaaccga aggcaaggct 600
ctaggcctat cacacaccta cgctaacgta cgcatcaacc atgttataaa cctgtgggga 660
gctgactttg attctaacgg gaaccttaaa gctatttatg taacagactc tgatagtaat 720
gcatctattg gtatgaagaa atactttgtt ggtgttaatt ccgctggaaa agtagctatt 780
tctgctaaag aaataaaaga agataatata ggtgctcaag tactagggtt atttacactt 840
tcaacagggc aagatagttg gaatcagacc aatggcggtg gccatcatca ccatcaccac 900
taa 903
<210> 84
<211> 903
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 84
atgtccgttt ggaccaaagg agttactcct ccagcaaact tcactcaagg tgaagatgtt 60
tttcacgctc cttatgttgc taaccaagga tggtatgata ttaccaaaac attcaatgga 120
aaagacgatc ttctttgcgg ggctgccaca gcagggaata tgcttcactg gtggttcgat 180
caaaacaaag accaaattaa acgttatttg gaagagcatc cagaaaagca aaaaataaac 240
ttcaaaggcg aacagatgtt tgacgtaaaa gaagctatcg acactaaaaa ccaccagcta 300
gatagtaaat tatttgaata ttttaaagaa aaagctttcc cttatctatc tactaaacac 360
ctaggagttt tccctgatca tgtaattgat atgttcatta acggctaccg ccttagtcta 420
actaaccacg gtccaacgcc agtaaaagaa ggtagtaaag atccccgagg tggtattttt 480
gacgccgtat ttacaagagg taaccaaagt aagctattga caagtcgtca tgattttaaa 540
gaaaaaaatc tcaaagaaat cagtgatctc attaagaaag agttaaccga aggcaaggct 600
ctaggcctat cacacaccta cgctaacgta cgcatcaacc atgttataaa cctgtgggga 660
gctgactttg attctaacgg gaaccttaaa gctatttatg taacagactc tgatagtaat 720
gcatctattg gtatgaagaa atactttgtt ggtgttaatt ccgctggaaa agtagctatt 780
tctgctaaag aaataaaaga agataatata ggtgctcaag tactagggtt atttacactt 840
tcaacagggc aagatagttg gaatcagacc aatggcggtg gccatcatca ccatcaccac 900
taa 903
<210> 85
<211> 903
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 85
atgtccgttt ggaccaaagg agttactcct ccagcaaact tcactcaagg tgaagatgtt 60
tttcacgctc cttatgttgc taaccaagga tggtatgata ttaccaaaac attcaatgga 120
aaagacgatc ttctttgcgg ggctgccaca gcagggaata tgcttcactg gtggttcgat 180
caaaacaaag accaaattga acgttatttg gaagagcatc cagaaaagca aaaaataaac 240
ttcaaaggcg aacagatgtt tgacgtaaaa aaagctatcg acactaaaaa ccaccagcta 300
gatagtaaat tatttgaata ttttaaagaa aaagctttcc cttatctatc tactaaacac 360
ctaggagttt tccctgatca tgtaattgat atgttcatta acggctaccg ccttagtcta 420
actaaccacg gtccaacgcc agtaaaagaa ggtagtaaag atccccgagg tggtattttt 480
gacgccgtat ttacaagagg taaccaaagt aagctattga caagtcgtca tgattttaaa 540
gaaaaaaatc tcaaagaaat cagtgatctc attaaggaag agttaaccaa aggcaaggct 600
ctaggcctat cacacaccta cgctaacgta cgcatcaacc atgttataaa cctgtgggga 660
gctgactttg attctaacgg gaaccttaaa gctatttatg taacagactc tgatagtaat 720
gcatctattg gtatgaagaa atactttgtt ggtgttaatt ccgctggaaa agtagctatt 780
tctgctaaag aaataaaaga agataatata ggtgctcaag tactagggtt atttacactt 840
tcaacagggc aaaaaagttg gaatcagacc aatggcggtg gccatcatca ccatcaccac 900
taa 903
<210> 86
<211> 963
<212> DNA
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 86
atggacgatt accaaaggaa tgctacggaa gcttatgcca aagaagtacc acatcagatc 60
acttccgttt ggaccaaagg agttactcct ccagcaaact tcactcaagg tgaagatgtt 120
tttcacgctc cttatgttgc taaccaagga tggtatgata ttaccaaaac attcaatgga 180
aaagacgatc ttctttgcgg ggctgccaca gcagggaata tgcttcactg gtggttcgat 240
caaaacaaag accaaattga acgttatttg gaagagcatc cagaaaagca aaaaataaac 300
ttcaaaggcg aacagatgtt tgacgtaaaa aaagctatcg acactaaaaa ccaccagcta 360
gatagtaaat tatttgaata ttttaaagaa aaagctttcc cttatctatc tactaaacac 420
ctaggagttt tccctgatca tgtaattgat atgttcatta acggctaccg ccttagtcta 480
actaaccacg gtccaacgcc agtaaaagaa ggtagtaaag atccccgagg tggtattttt 540
gacgccgtat ttacaagagg taaccaaagt aagctattga caagtcgtca tgattttaaa 600
gaaaaaaatc tcaaagaaat cagtgatctc attaaggaag agttaaccaa aggcaaggct 660
ctaggcctat cacacaccta cgctaacgta cgcatcaacc atgttataaa cctgtgggga 720
gctgactttg attctaacgg gaaccttaaa gctatttatg taacagactc tgatagtaat 780
gcatctattg gtatgaagaa atactttgtt ggtgttaatt ccgctggaaa agtagctatt 840
tctgctaaag aaataaaaga agataatata ggtgctcaag tactagggtt atttacactt 900
tcaacagggc aaaaaagttg gaatcagacc aatggcggtg gccatcatca ccatcaccac 960
taa 963
<210> 87
<211> 19
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 87
Ser Phe Ser Ala Asn Gln Glu Ile Arg Tyr Ser Glu Val Thr Pro Tyr
1 5 10 15
His Val Thr
<210> 88
<211> 19
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 88
Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val Pro His
1 5 10 15
Gln Ile Thr
<210> 89
<211> 20
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 89
Asp Asp Tyr Gln Arg Asn Ala Thr Glu Ala Tyr Ala Lys Glu Val Pro
1 5 10 15
His Gln Ile Thr
20
<210> 90
<211> 959
<212> PRT
<213> 酿脓链球菌 (Streptococcus pyogenes)
<400> 90
Glu Glu Lys Thr Val Gln Val Gln Lys Gly Leu Pro Ser Ile Asp Ser
1 5 10 15
Leu His Tyr Leu Ser Glu Asn Ser Lys Lys Glu Phe Lys Glu Glu Leu
20 25 30
Ser Lys Ala Gly Gln Glu Ser Gln Lys Val Lys Glu Ile Leu Ala Lys
35 40 45
Ala Gln Gln Ala Asp Lys Gln Ala Gln Glu Leu Ala Lys Met Lys Ile
50 55 60
Pro Glu Lys Ile Pro Met Lys Pro Leu His Gly Pro Leu Tyr Gly Gly
65 70 75 80
Tyr Phe Arg Thr Trp His Asp Lys Thr Ser Asp Pro Thr Glu Lys Asp
85 90 95
Lys Val Asn Ser Met Gly Glu Leu Pro Lys Glu Val Asp Leu Ala Phe
100 105 110
Ile Phe His Asp Trp Thr Lys Asp Tyr Ser Leu Phe Trp Lys Glu Leu
115 120 125
Ala Thr Lys His Val Pro Lys Leu Asn Lys Gln Gly Thr Arg Val Ile
130 135 140
Arg Thr Ile Pro Trp Arg Phe Leu Ala Gly Gly Asp Asn Ser Gly Ile
145 150 155 160
Ala Glu Asp Thr Ser Lys Tyr Pro Asn Thr Pro Glu Gly Asn Lys Ala
165 170 175
Leu Ala Lys Ala Ile Val Asp Glu Tyr Val Tyr Lys Tyr Asn Leu Asp
180 185 190
Gly Leu Asp Val Asp Val Glu His Asp Ser Ile Pro Lys Val Asp Lys
195 200 205
Lys Glu Asp Thr Ala Gly Val Glu Arg Ser Ile Gln Val Phe Glu Glu
210 215 220
Ile Gly Lys Leu Ile Gly Pro Lys Gly Val Asp Lys Ser Arg Leu Phe
225 230 235 240
Ile Met Asp Ser Thr Tyr Met Ala Asp Lys Asn Pro Leu Ile Glu Arg
245 250 255
Gly Ala Pro Tyr Ile Asn Leu Leu Leu Val Gln Val Tyr Gly Ser Gln
260 265 270
Gly Glu Lys Gly Gly Trp Glu Pro Val Ser Asn Arg Pro Glu Lys Thr
275 280 285
Met Glu Glu Arg Trp Gln Gly Tyr Ser Lys Tyr Ile Arg Pro Glu Gln
290 295 300
Tyr Met Ile Gly Phe Ser Phe Tyr Glu Glu Asn Ala Gln Glu Gly Asn
305 310 315 320
Leu Trp Tyr Asp Ile Asn Ser Arg Lys Asp Glu Asp Lys Ala Asn Gly
325 330 335
Ile Asn Thr Asp Ile Thr Gly Thr Arg Ala Glu Arg Tyr Ala Arg Trp
340 345 350
Gln Pro Lys Thr Gly Gly Val Lys Gly Gly Ile Phe Ser Tyr Ala Ile
355 360 365
Asp Arg Asp Gly Val Ala His Gln Pro Lys Lys Tyr Ala Lys Gln Lys
370 375 380
Glu Phe Lys Asp Ala Thr Asp Asn Ile Phe His Ser Asp Tyr Ser Val
385 390 395 400
Ser Lys Ala Leu Lys Thr Val Met Leu Lys Asp Lys Ser Tyr Asp Leu
405 410 415
Ile Asp Glu Lys Asp Phe Pro Asp Lys Ala Leu Arg Glu Ala Val Met
420 425 430
Ala Gln Val Gly Thr Arg Lys Gly Asp Leu Glu Arg Phe Asn Gly Thr
435 440 445
Leu Arg Leu Asp Asn Pro Ala Ile Gln Ser Leu Glu Gly Leu Asn Lys
450 455 460
Phe Lys Lys Leu Ala Gln Leu Asp Leu Ile Gly Leu Ser Arg Ile Thr
465 470 475 480
Lys Leu Asp Arg Ser Val Leu Pro Ala Asn Met Lys Pro Gly Lys Asp
485 490 495
Thr Leu Glu Thr Val Leu Glu Thr Tyr Lys Lys Asp Asn Lys Glu Glu
500 505 510
Pro Ala Thr Ile Pro Pro Val Ser Leu Lys Val Ser Gly Leu Thr Gly
515 520 525
Leu Lys Glu Leu Asp Leu Ser Gly Phe Asp Arg Glu Thr Leu Ala Gly
530 535 540
Leu Asp Ala Ala Thr Leu Thr Ser Leu Glu Lys Val Asp Ile Ser Gly
545 550 555 560
Asn Lys Leu Asp Leu Ala Pro Gly Thr Glu Asn Arg Gln Ile Phe Asp
565 570 575
Thr Met Leu Ser Thr Ile Ser Asn His Val Gly Ser Asn Glu Gln Thr
580 585 590
Val Lys Phe Asp Lys Gln Lys Pro Thr Gly His Tyr Pro Asp Thr Tyr
595 600 605
Gly Lys Thr Ser Leu Arg Leu Pro Val Ala Asn Glu Lys Val Asp Leu
610 615 620
Gln Ser Gln Leu Leu Phe Gly Thr Val Thr Asn Gln Gly Thr Leu Ile
625 630 635 640
Asn Ser Glu Ala Asp Tyr Lys Ala Tyr Gln Asn His Lys Ile Ala Gly
645 650 655
Arg Ser Phe Val Asp Ser Asn Tyr His Tyr Asn Asn Phe Lys Val Ser
660 665 670
Tyr Glu Asn Tyr Thr Val Lys Val Thr Asp Ser Thr Leu Gly Thr Thr
675 680 685
Thr Asp Lys Thr Leu Ala Thr Asp Lys Glu Glu Thr Tyr Lys Val Asp
690 695 700
Phe Phe Ser Pro Ala Asp Lys Thr Lys Ala Val His Thr Ala Lys Val
705 710 715 720
Ile Val Gly Asp Glu Lys Thr Met Met Val Asn Leu Ala Glu Gly Ala
725 730 735
Thr Val Ile Gly Gly Ser Ala Asp Pro Val Asn Ala Arg Lys Val Phe
740 745 750
Asp Gly Gln Leu Gly Ser Glu Thr Asp Asn Ile Ser Leu Gly Trp Asp
755 760 765
Ser Lys Gln Ser Ile Ile Phe Lys Leu Lys Glu Asp Gly Leu Ile Lys
770 775 780
His Trp Arg Phe Phe Asn Asp Ser Ala Arg Asn Pro Glu Thr Thr Asn
785 790 795 800
Lys Pro Ile Gln Glu Ala Ser Leu Gln Ile Phe Asn Ile Lys Asp Tyr
805 810 815
Asn Leu Asp Asn Leu Leu Glu Asn Pro Asn Lys Phe Asp Asp Glu Lys
820 825 830
Tyr Trp Ile Thr Val Asp Thr Tyr Ser Ala Gln Gly Glu Arg Ala Thr
835 840 845
Ala Phe Ser Asn Thr Leu Asn Asn Ile Thr Ser Lys Tyr Trp Arg Val
850 855 860
Val Phe Asp Thr Lys Gly Asp Arg Tyr Ser Ser Pro Val Val Pro Glu
865 870 875 880
Leu Gln Ile Leu Gly Tyr Pro Leu Pro Asn Ala Asp Thr Ile Met Lys
885 890 895
Thr Val Thr Thr Ala Lys Glu Leu Ser Gln Gln Lys Asp Lys Phe Ser
900 905 910
Gln Lys Met Leu Asp Glu Leu Lys Ile Lys Glu Met Ala Leu Glu Thr
915 920 925
Ser Leu Asn Ser Lys Ile Phe Asp Val Thr Ala Ile Asn Ala Asn Ala
930 935 940
Gly Val Leu Lys Asp Cys Ile Glu Lys Arg Gln Leu Leu Lys Lys
945 950 955

Claims (13)

1.一种用于移植细胞、组织或器官至对象的调理方案，包括向所述对象施用使所述对象的血清IgG分子失活的酶，任选地其中所述移植是造血干细胞和祖细胞(HSPC)的移植。
2.根据权利要求1所述的调理方案，其中所述酶的施用量足以使所述对象的血清中存在的所有或基本上所有的IgG分子失活。
3.根据前述权利要求中任一项所述的调理方案，其中所述酶是IgG半胱氨酸蛋白酶或IgG内切糖苷酶。
4.根据权利要求3所述的调理方案，其中：
(i)所述IgG半胱氨酸蛋白酶来自链球菌属细菌，例如酿脓链球菌，任选地其中所述酶是IdeS、IdeZ或MAC2多肽，或
(ii)所述IgG内切糖苷酶来自链球菌属细菌，例如酿脓链球菌、马链球菌或兽疫链球菌，或者来自假结核棒杆菌，粪肠球菌，或脑膜脓毒性伊丽莎白金菌，任选地其中所述酶是EndoS、CP40、EndoE或EndoF2多肽。
5.根据权利要求4所述的调理方案，其中：
-所述IgG半胱氨酸蛋白酶是具有与SEQ ID NO：2、4或5至少80％相同，例如至少85％、90％、95％或99％相同的序列的多肽，或者其中所述IgG半胱氨酸蛋白酶包含以下或由以下组成：SEQ ID NO：6至25和55至69中任一个的序列，任选地其中所述序列包含N端的附加甲硫氨酸和/或C端的组氨酸标签；或者
-所述IgG内切糖苷酶是具有与SEQ ID NO：90至少80％相同，例如至少85％、90％、95％或99％相同的序列的多肽，任选地其中所述序列包含N端的附加甲硫氨酸和/或C端的组氨酸标签。
6.根据前述权利要求中任一项所述的调理方案，其中所述酶是imlifidase和/或EndoS。
7.根据前述权利要求中任一项所述的调理方案，包括以下中的一个或多个：
(a)向所述对象施用非致命剂量的放射和/或任何其他耗竭所述对象的HSPC的药剂；
(b)施用用于减少所述对象中淋巴细胞的数量和/或下调所述对象中淋巴细胞的活性的药剂，其中所述淋巴细胞包括：
i.T细胞；和/或
ii.B细胞(任选地包括产生抗体的细胞)；
(c)施用任何其他降低免疫系统活性的药剂或方案，例如补体抑制剂、细胞因子抑制剂、先天免疫细胞抑制剂、耐受性诱导剂。
8.根据权利要求7所述的调理方案，包括至少(a)和(b)，任选地其中：
-(a)另外包括施用供者CD8-α细胞输注；和/或
-(a)包括施用抗CD117抗体和/或抗CD47抗体；和/或
-(b)包括施用抗CD4抗体、抗CD8抗体和抗CD90抗体、硼替佐米和环磷酰胺，和/或施用rATG。
9.一种在对象中诱导造血嵌合体的方法，所述方法包括实施前述权利要求中任一项所述的调理方案，并且随后以足以在所述对象中诱导造血嵌合体的量和在适于在所述对象中诱导造血嵌合体的条件下向所述对象施用造血干细胞和祖细胞(HSPC)。
10.根据权利要求9所述的方法，其中所述HSPC是同种异体的、同基因的或自体的，并且任选地是基因改造的。
11.一种预防或治疗细胞、组织或器官移植体的免疫排斥的方法，所述方法包括实施权利要求9或10所述的方法，并且向所述对象施用与所述HSPC来自同一供者的细胞、组织或器官移植体。
12.根据权利要求11所述的方法，其中所述移植体是肾脏、肝脏、心脏、胰腺、肺、小肠、皮肤、血管/血管组织、面部、手臂、气管、眼睛的部分、胰岛、黑质、骨髓或干细胞，任选地包括用于权利要求9或10的方法中的所述HSPC，如此不再需要额外的移植体。
13.用于预防或治疗细胞、组织或器官移植体的免疫排斥的方法中的酶，所述酶使血清IgG分子失活，所述方法包括权利要求11或12所述的方法。
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