CN1261579C - 形成异源二聚体的杂种蛋白质 - Google Patents
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- CN1261579C CN1261579C CNB971924112A CN97192411A CN1261579C CN 1261579 C CN1261579 C CN 1261579C CN B971924112 A CNB971924112 A CN B971924112A CN 97192411 A CN97192411 A CN 97192411A CN 1261579 C CN1261579 C CN 1261579C
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Abstract
一种杂种蛋白质,它包括形成一种二聚体的两种共同表达的氨基酸序列。每一个序列含有一种受体,如TBP1或TBP2,或一种配体,如IL6,IFN-β和TPO的结合部分。它与一种异源二聚体蛋白质类激素如人CG的亚基连接。每一个共表示的序列含有一个相应的激素亚基从而在表达后形成一种异源二聚体。也公开了相应的DNA分子,表达载体,宿主细胞,药物组合物和生产这种蛋白质的方法。
Description
所属技术领域
本发明涉及一种含有两个共同表达的氨基酸序列并形成一个二聚体的杂种蛋白质,该两种氨基酸序列的每一种含有:
(a)至少一种选自一种同源聚体受体(homomeric recedtor),一种异源聚体的链,一种配体,及它们的片段的氨基酸序列;以及
(b)一种异源二聚体蛋白质类激素的亚单位或其片段;其中(a)和(b)是直接结合的或通过一种肽连接物结合的,以及在每一个双体中,两个亚单位(b)是不同的并能够聚集形成二聚复合体。
发明背景
蛋白质-蛋白质相互作用对于细胞和多细胞有机体的正常生理功能来说是必不可少的。自然界中的许多蛋白质当与一个或多个其它蛋白质链相复合时,会表现出新的或最适的功能。这可通过各种配体-受体结合而对细胞活动的调节来说明。一些配体,如肿瘤坏死因子α(TNFα),TNFβ,或人类绒毛膜促性腺激素(hCG),是以多亚基复合体存在的。一些这种复合体含有多拷贝的同一种亚基。TNFα和TNFβ(下文统称为TNF)是由3个相同的亚单位形成的同源三聚体(1-4)。其它配体是由非同种亚基组成的。如,hCG是一种异源二聚体(5-7),受体也可以多链复合体的形式存在或发生功能。如,TNF的受体在聚集形成二聚体时可传递一个信号(8,9)。这些受体的配体促进两个受体链的聚集,从而提供一种受体活化的机制。例如,TNF介导的聚集激活TNF受体(10-12)。
蛋白质-蛋白质相互作用的调节可以是各种疾病和病理中治疗介入的有用机制。可与配体相互作用的可溶性结合蛋白质可能会将配体与受体分离,从而减弱特定的受体途径的活化。或者,配体的分离可能推迟它的清除或降解,从而增长它的效应期间,并可能增强它在体内的表观活性。对于TNF,可溶性TNF受体已主要与TNF活性的抑制相联系(13-17)。
可溶性结合蛋白质对于治疗人类疾病来说可能有用。例如,可溶性TNF受体已显示出在关节炎的动物模型中有效用(18,19)。
由于TNF对其受体具有3个结合位点(10-12),并且该细胞表面受体的二聚体化对于生物活性来说是足够的(8,9),很有可能单个可溶性受体与TNF的结合将带来一种可能性,即可溶性受体:TNF(三聚体)的这种1∶3复合体仍可结合和活化一对细胞表面TNF受体。为获得一种抑制效应,期望TNF三聚体上的受体结合位点中的两个必须由可溶性结合蛋白质占据或封闭。或者,该结合蛋白质可在细胞表面上封闭适当定向的TNF。
总而言之,人们觉得需要合成含有两个受体(或配体)链的作为二聚体杂种蛋白质的蛋白质。参见Wallach等,美国专利5478925。用于产生含有来自胞外受体结合区域的二聚体或多聚体杂种蛋白质的策略为将这些蛋白质与抗体重链的恒定区段相融合。
这一策略导致了,例如,CD4免疫粘附素的构建(20)。它们是由与抗体的重链和轻链的恒定区段融合的CD4的前面两个(或全部四个)免疫球蛋白类区域组成的。这个创建杂种分子的策略被调整用于TNF的受体(10,16,21),并导致了与单体可溶性结合蛋白质相比具有较高体外活性的构建体的产生。
人们普通认为,二聚体融合蛋白质较高的体外活性应当转化为较高的体内活性。一个研究的结果支持了这一论点,p75(TBP2)-Ig融合蛋白质在保护小鼠免于产生静脉内LPS注射的不良后果上显示出至少50倍高的活性。
但是,虽然免疫球蛋白融合蛋白质已得到广泛应用,这一策略具有几个缺点。一个是一些免疫球蛋白Fc区域参与免疫系统的效应物功能。这些功能在特定的治疗情形中可能是不期望的(22)。
第二个限制与期望产生异源聚体融合蛋白质,如异源聚体IL-6或I型干扰素受体的可溶性类似物的特定情形相关。虽然有很多方法产生双功能性抗体(如,通过共转染或杂交瘤融合),但通常情况下产生的同源二聚体和异源二聚体的混合物大大损害了合成的有效性(23)。最近已有几个报道描述了使用亮氨酸拉链基序引导异源二聚体的组装(24-26)。这好象是一个用于研究目的的有前途的方法,但所使用的这种非自身的或胞内序列由于其抗原性而不适合于临床上的长期使用。组装的效率和组装后的稳定性也可能是限制因素。
另一方面,在TNF受体的特定情况下,已发现对p55TNF受体的某些修饰有利于同源二聚体化和不存在配体时的信号(27,28)。已经发现,该受体的胞质区段,被称作“死亡区段”,可充当同源二聚体的基序(28,30)。作为免疫球蛋白杂种蛋白质的替代物,可以想象TNF受体的胞外区域与其胞质死亡区域的融合可产生一种分泌蛋白质,它能在没有TNF时二聚体化。这种融合蛋白质已在国际专利申请WO 95/31544中公开并要求专利保护。
用于产生可溶性TNF受体的二聚体的第三个策略是将该单体蛋白质与聚乙二醇进行化学交联(31)。
发明概述
用于获得这种蛋白质的另一种方式为本发明之一,它具有一些重要的优点,该方式在于使用相应于循环系统中具有长半衰期的非免疫球蛋白的天然异源二聚体支架。一个优选的例子是hCG,它是一种分泌良好,且有高的稳定性并具有长的半衰期的蛋白质(32-33)。由于hCG作为怀孕标记物的突出的作用,已开发出多种试剂以在体外和体内对该蛋白质进行定量和研究。此外,已对hCG使用诱变方法进行了广泛的研究,并且已知该蛋白质的小的缺失,例如去除α亚基最羧基末端的五个残基,可有效地消除其生物活性而同时保留其形成异源二聚体的能力(34,35)。多至30个氨基酸的小的插入已显示出在α亚基的氨基和羧基末端具有可容忍性(36),同时α亚基与β亚基的羧基末端的融合对异源二聚体的形成也具有很小的影响(37)。
也已报道了一种hCG的类似物,其中免疫球蛋白Fc区域与hCGβ亚基的C-末端融合;但该构建体没有被分泌,并还没有进行将其与α亚基相联合的实验(38)。
因而本发明的主要目的为含有形成一种二聚体的两种共同表达的氨基酸序列的杂种蛋白质,每一种氨基酸序列含有:
(a)至少一种选自一种同源聚体受体,一种异源聚体受体的链,一种配体,和它们的片段的氨基酸序列;和
(b)一种异源二聚体蛋白质类激素的亚基或其片段;其中(a)和(b)是直接结合的或通过一种肽连接物结合的,并且在每一个双体中,两个亚基(b)是不同的并且能够聚集形成一个二聚体复合体。
根据本发明,该连接物可被酶切。
序列(a)最好选自:TNF受体1(55KDa,也称作TBP1)的胞外区域,TNF受体2(75KDa,也称作TBP2)的胞外区域,或者仍含有配体结合区域的它们的片段;IL-6受体(也称作gp80和gp130)的胞外区域;IFNα/β受体或IFNγ受体的胞外区域;促性腺素受体或其胞外区域;抗体轻链,或其片段,任选地与相应的重链相联;抗体重链,或其片段,任选地与相应的轻链相联;抗体Fab区域;或配体蛋白质,如细胞因子,生长因子或除促性腺素外的激素,其特定的例子包括IL-6,TFN-β,TPO,或它们的片段。
序列(b)最好选自hCG,FSH,LH,TSH,抑制素亚基,或它们的片段。
可使用对这些蛋白质的修饰,如对蛋白质骨架的化学或者酶切割,或对一些氨基酸侧链的化学或酶修饰,使本发明的杂种蛋白质的组分失活。这种对活性的限制也可通过使用重组DNA技术,改变杂种蛋白质的编码序列以直接导致一个组分的活性受到限制的方式来完成,或者使该蛋白质更易于接受随后的化学或酶修饰来完成。
基于与(b)结合的氨基酸序列(a),上述杂种蛋白质将产生单功能,双功能或多功能分子。在每一个双体中,(a)可以与(b)的氨基端或羧基端连接,或与两者都相连。
本发明的一个单克隆杂种蛋白质可以,例如,含有一种促性腺素受体的胞外区域,它与相应的受体-结合促性腺素亚基之一相连。根据这一实施方案,本发明的杂种蛋白质可以是这样一种分子,其中,例如,FSH受体胞外区域与FSH相连,以提高血浆中的半衰期并改善生物活性。
这种制剂在有助繁殖方法中,例如诱导排卵或体外受精,可被用于诱导滤泡成熟,并充当剧烈放大对这一过程的成功所必需的激素的生物活性的工具,从而降低为达到排卵所需要的激素本身的量和需要注射的次数。
FSH受体以及人类FSH受体胞外区域的生产已被分别描述于WO92/16620和WO96/38575。
按照一个特定的实施方案,FSH受体的胞外区域(ECD)可与含有凝血酶识别/切割位点的肽连接物同框融合(29)并呈现一种“被栓住的”长臂。该肽连接物将FSH的胞外区域与FSH亚基相连,这样,当这种分子在循环系统中与凝血酶相接触时可使FSH受体的胞外区域通过在凝血酶切割位点切割而被除去。
在另一个实施方案中,使用的是大量存在于卵巢中的酶的酶识别位点,而不是凝血酶识别位点。这样,当ECD-FSH分子迁移到卵巢中时,它将暴露于该组织中高浓度存在的酶,ECD将被除去,从而使FSH可与膜结合受体相互作用。
在另一个实施方案中,不是使用酶识别位点,而是将一种柔性绞链区段克隆到ECD和FSH之间,从而ECD不会被酶从该激素上除去。这样,当ECD-FSH分子抵达卵巢后,绞链连接的ECD和卵巢细胞膜上存在的FSH受体的ECD之间将出现竞争。
在本发明的又一个优选实施方案中,该杂种蛋白质是由一对氨基酸序列的聚集体组成的,其中之一含有TBP1(或从氨基酸20至氨基酸161或至氨基酸190的一些片段)作为(a),和hCG的α亚基,作为(b),其它的总含有TBP1(或与上述相同的片段)作为(a),和hCG的β亚基或其片段,作为(b)。按照这一实施方案,基于被选作(b)的特定的序列(hCG的β全亚基,或其片段或修饰物),所产生的杂种蛋白将具有一种活性(只有TBP1的活性)或几种活性的组合(TNP1的活性与hCG的活性)。在后一种情况下,该杂种蛋白质可被用于,例如,联合治疗Kaposi氏肉瘤和艾滋病(AIDS)中的代谢损耗。
在本发明的再一个实施方案中,在两个亚基(b)之间加入一个或多个共价键,以提高所产生的杂种蛋白质的稳定性。这可通过例如加入一个或多个非天然的链间二硫键来进行。这些交联的位点可从已知的异源二聚体激素的结构推导出来。例如,hCG中的一个适当的位点为可将半胱氨酸置于α亚基的残基Lys45和β亚基的残基Glu21,将一个盐桥键(非共价键)用一个二硫键(共价键)替换。本发明的另一个目的是这些杂种蛋白质的PEG化或其它的化学修饰的形式。
本发明的又一个目的是一种DNA分子,它含有编码上述杂种蛋白质的DNA序列,以及基本上相同的核苷酸序列。“基本上相同的核苷序列”包括其它的由于遗传密码的简并性也编码给定的氨基酸序列的核酸序列。
对于生产本发明的杂种蛋白质来说,DNA序列(a)得自存在的克隆,如(b)那样。将编码所需的序列(a)的DNA序列与编码所需序列(b)的DNA序列相连。将这两种融合产物插入并连接进适当的质粒或将每一种连接进不同的质粒,完成后,将表达载体或两种表达载体导入适当的宿主细胞,然后宿主细胞对载体进行表达,产生如上定义的本发明的杂种蛋白质。
制备本发明杂种蛋白的优选方法是通过PCR技术,使用的寡核苷酸是对需要从编码序列(a)和(b)的克隆复制的期望序列具有特异性的寡核苷酸。
本文所述的本发明的任一个重组蛋白质的表达可使用适当的表达载体在真核细胞(如酵母,昆虫或哺乳动物细胞)或原核细胞中进行,可使用现有技术已知的任何一种方法。
例如,将编码通过上述任意一种方法得到的蛋白质的DNA分子用现有技术公知的技术插入适当构建的表达载体中(参见Sambrook等1989)。将双链cDNA与质粒载体连接的技术有同源多聚体加尾,涉及使用合成的DNA连接物的限制性连接和平端连接技术:使用DNA连接酶连接DNA分子,并且通过用碱性磷酸酶处理来避免不期望的连接。
为了能够表达所需的蛋白质,一个表达载体应该也包括含有转录和转译调节信息的特殊的核苷酸序列,并且它与编码期望蛋白质的DNA相连,其连接方式应允许基因的表达和该蛋白质的产生。对于要转录的基因来说最为重要的是它必须在其前面有一个可被RNA聚合酶识别的启动子,该聚合酶与启动子结合从而起始转录过程。已有多种这种启动子被投入使用,它们以不同的效率起作用(强和弱启动子)。
对于真核宿主来说,根据宿主的性质可以使用不同的转录和转译调节序列。它们可以来自病毒,如腺病毒,小牛乳头瘤病毒,猿猴病毒等,其中,该调节信号与具有很高的表达水平的特定基因相联。它们的例子有疱疹病毒的TK启动子,SV40早期启动子,酵母ga14基因启动子等。被选用的转录起始调节信号可以是允许抑制和激活的,从而可对基团的表示进行调控。
将含有编码本发明的杂种蛋白质的核苷酸序列插入载体中,该载体具有可操纵连接的转录和转译调节信号,并能够将期望基因序列整合进宿主细胞。已被导入的DNA稳定转化的细胞可通过也导入一个或多个标记物来选择,该标记物可允许对含有表达载体的宿主细胞进行选择。该标记物也可以对营养缺陷型宿主提供光营养,杀生物抗性,如抗生素,或重金属,如铜等。可选择的标记基因可直接与待表达的DNA基因序列相连,也可通过共转染导入同一个细胞中。对于本发明蛋白质的最适合成来说也可能需要其它的要素。
在选择特定的质粒或病毒载体中的重要因素包括:从那些不含有载体的受体细胞中识别和选出含有载体的受体细胞的难易性;在特定宿主中期望载体的拷贝数;以及是否需要它能够在不同种类的宿主细胞之间“穿梭”。
一旦制备出用于表达的含有所述构建体的载体或DNA序列,该DNA构建体可通过多种适当的方式中的任意一种被导入适当的宿主细胞:转化,转染,接合,原生质体融合,电穿孔,磷酸钙沉淀,直接微注射等。
宿主细胞可以是原核细胞或真核细胞。优选真核细胞,如哺乳动物细胞,如人、猴、小鼠、和中国仓鼠卵巢(CHO)细胞,这是因为它们对蛋白质分子提供转译后修饰,包括正确折叠和在正确的位点糖基化。酵母细胞也进行转译后肽修饰,包括糖基化。已有一些使用强启动子序列和高拷贝数质粒的重组DNA策略,它可用于在酵母中生产所需的蛋白质。酵母识别克隆的哺乳动物基因产物上的引导序列,并分泌携带引导序列的肽(即前肽)。
在载体引入之后,将宿主细胞培养于选择培养基中,它对含有载体的细胞的生长进行选择,克隆的基因序列的表达导致所需蛋白质的产生。
重组蛋白质的纯化是通过已知用于这一目的的任意一种方法,即涉及提取,沉淀,层折,电泳等的任一种常规方法来进行的。一种可优选用于纯化本发明的蛋白质进一步纯化方法为使用与靶蛋白质结合的单克隆抗体的亲和层折方法,该单克隆抗体产生和被固定在包含于一个柱中的凝胶基质上。将含有该重组蛋白质的不纯净的制剂流经该柱。这种蛋白质将通过特定的抗体与柱结合,而杂质将流出去。洗涤之后,通过改变pH值或离子强度将该蛋白质洗脱出来。
本文所使用的术语“杂种蛋白质”一般表示含有两种或多种不同的蛋白质或其片段的蛋白质。
本文使用的术语“融合蛋白质”是指一种杂种蛋白质,它是由共价相连的两种或多种蛋白或其片段组成的。
本文使用的术语“聚集体”是指两个多肽链之间形成强的特异性非共价相互作用,从而形成一个复合体,例如在异源二聚体激素(如FSH,LH,hCG或TSH)的α和β亚基之间形成的相互作用。
本文使用的术语“配体”或“配体蛋白质”是指一种除抗体或免疫球蛋白之外的分子,它可被受体的配体结合区域结合;这种分子可以是自然界中存在的,或者可以是化学修饰的或化学合成的。
本文使用的术语“配体结合区域”是指一种参与配体结合的受体部分,它通常是部分的或基本上全部的胞外区域。
本文使用的术语“受体”是指一种膜蛋白,它与相应的配体的结合引发二级细胞响应,导致胞内过程的激活或抑制。
另一方面,本发明提供了这种杂种蛋白质作为药物的应用。这种药品最好以含有本发明的蛋白质以及一种或多种药学上可接受的载体和/或赋形剂的药物组合物的形式提供。这种药物组合物代表了本发明的再一个方面。
附图简要说明
参阅附图可更好地理解本发明,其中:
图1(a)和1(b)分别表示了TBP(20-161)-hCGα以及TBP(20-161)-hCGβ构建体,以及相应的序列(SEQID NO:I-4)。图1(a)中显示的连接物序列为Ala-Gly-Ala-Ala-Pro-Gly(SEQ ID NO:9)。图1(b)中显示的连接物序列为Ala-Gly-Ala-Gly(SEQ ID NO:10)。
图2(a)和2(b)分别表示了TBP(20-190)-hCGα以及TBP(20-190)-hCGβ构建体,以及相应的序列(SEQID NO:5-8)。
图3表明CHO细胞表达的TBP-hCG(20-190)对TNFα-诱导的对BT-20细胞的细胞毒性的保护效应的剂量依赖性,以及各种对照。
图4表示COS细胞表达的TBP-hCG(20-190)对TNFα-诱导的对BT-20细胞毒性的保护效应的剂量依赖性,以及各种对照。
图5表明亲和纯化的CHO细胞表达的TBP-hCG(20-161)对TNFα-诱导的对BT-20细胞毒性的保护效应的剂量依赖性,以及各种对照。
优选实施方案的详细描述。
本发明将通过下文的实施例进行描述,不能解释为以任何方式限制本发明。
实施例
材料和方法
用于本研究的细胞系是得自美国典型培养物保藏中心(ATCC),Rockville,Maryland,除非另有说明。CHO-DUKX细胞系通过MIT的D,Houseman得自哥伦比亚大学的Chasin(39)。将缺乏功能性二氢叶酸还原酶基因的CHO-DUKX细胞常规地保持在补充了10%小牛血清(FBS)的完全α(+)改良的Eagles培养基(α(+)MEM)中。将COS-7细胞按照常规保持在补充有10%FBS的Dulbecco改良的Eagles培养基(DMEM)中。除非特别说明,将细胞打散使它们保持在对数生长期,培养试剂得自GIBCO(Grand Island,New York)。
1、编码杂种蛋白质的遗传构建体的组装
对p55TNF受体的编号基于Wallach的克隆论文(40),而对人hCG亚基的编号是基于Fichdes克隆论文的编号(41,42)。TBP,或TNF结合蛋白质这一称谓表示能够结合TNF的TNF受体的腹外区域部分。在这些实施例中,该DNA构建体将被命名为TBP-杂种蛋白质,加上构建体命名法中指明的TBP的参与物和区段。所有的TBP-hCG构建体均含有替换天然p55信号序列的人生长激素(hGH)信号肽。此外,hGH信号肽的定位使其紧接在TBP残基Asp20的前面,期望在成熟的分泌蛋白质中它成为第一个残基。这些修饰对于使用hCG作为该杂种蛋白质的参与物这一基本思想来说不是必需的。
编码该杂种蛋白质的DNA是使用PCR方法构建的(43)。
a.TBP1(20-161)-hCG
起初的TBP-hCG构建体被设计成含有来自p55TNF受体的胞外区域(从Asp20开始包括残基Cys161)的配体结合区域,它通过一个短的连接物与hCGα和β亚基(分别从残基αCys7或βPro7开始)融合。这一构建体,下文称作TBP1(20-161)-hCG,是一种两个被修饰的hCG亚基,TBP1(20-161)-hCGα和TBP1(20-161)-hCGβ,的异源二聚体。
用于TBP1(20-161)-hCGα构建体的寡核苷酸引物为:
引物1(αβ)TTT TCT CGA GAT GGC TAC AGG TAA GCG CCC(SEQ ID NO:11)
引物2(α)ACC TGG GGC ACG ACC GGC ACA GGA GAC ACA CTC GTT TTC
(SEQ ID NO:12)
引物3(α)TGT GCC GGT GCT GCC CCA GGT TGC CCA GAA TGC ACG CTA CAG
(SEQ ID NO:13)
引物4(α)TTT TGG ATC CTT AAG ATT TGT GAT AAT AAC AAG TAC
(SEQ ID NO:14)
这些引物和这些实施例中描述的其它引物是使用亚磷酰胺化学方法在Applied Biosysrem Model 392DNA合成仪上合成的(ABI,Foster市,加里福尼亚)。
由于两个TBP-hCG亚基构建体具有相同的5’一端(即hCH/TBP构建体的5’一端),两个TBP-hCG亚基构建体均使用引物1(αβ)。用于TBP1(20-161)-hCGβ构建体的其它引物为:
引物2(β)CCG TGG ACC AGC ACC AGC ACA GGA GAC ACA CTC GTT TTC
(SEQ ID NO:15)
引物3(β)TGT GCT GGT GCT GGT CCA CGG TGC CGC CCC ATC AAT
(SEQ ID NO:16)
引物4(β)TTT TGG ATC CTT ATT GTG GGA GGA TCG GGG TG(SEQ ID NO:17)
引物2(α)和3(α)为反向互补,并覆盖p55胞外区域编码区段的3’一端,以及hCGα亚基的5’一端。同样,引物2(β)和3(β)也为反向互补,并覆盖p55胞外区域编码区段的3’一端,和hCGβ亚基的5’一端。
对两个TBP-hCG亚基构建体的每一个进行两个PCR反应。第一个使用了引物1(αβ)和2(α或β),并使用了作为模板的编码可溶性p55残基20-180并在其前面存在hGH信号肽的质粒(pCMVhGHspcDNA.pA4质粒)。第二个使用了引物3(α或β)和4(α或β),并使用了作为模板的质粒pSVL-hCGα或pSVL-hCGβ两者之一(44)。PCR是使用得自New England Biolabs(Beverly,Massachusetts)的Vent(TM)聚合酶,并按照制造商推荐的方法进行的。每一反应进行25个循环,反应条件为:
模板DNA 100μg
每种引物1μg
Vent(TM)聚合酶(New Englandl Biolabs)2U
99℃变性30秒,
退火:对于引物1(αβ)和2(α)为59℃30秒;
引物3(α)和4(α)为59℃30秒;
引物1(αβ)和2(β)为57℃30秒;
引物3(β)和4(β)为63℃ 30秒;
在75℃延伸75秒。
将PCR产物在2%琼脂糖凝胶上电泳并用溴化乙锭(ethidiumbromide)染色,证实了其为期望的大小。然后将这些片段通过一个Wizard柱(Promega)并按照制造商的说明进行纯化。
TBP1(20-161)-hCGα的最后一个编码序列是通过使用引物1(αβ)和引物4(α)的融合PCR进行组装的,并且使用了从第一个PCR反应得到的从p55和hCGα片段纯化的产物作为模板。首先将两个模板,它们由于引物2(α)和3(α)之间的重叠而变性后退火到一起,在不加入任何引物的情况下进行10个循环的PCR。这些循环的条件基本上与先前使用的条件相同,只是退火是在67℃下进行的,延伸进行了2分钟。在这10个循环结束时加入引物1(αβ)和4(α),再进行另外10个循环。这后一轮反应的条件与先前使用的相同,只是所用的退火温度为59℃,延伸进行了75秒。
在1%琼脂糖凝胶上电泳对这一反应的产物的分析结果证实,我们获得了约1100bp的期望片段。将产物通过一个Wizard柱以纯化该片段,然后将它用XbaI和BamHI消化,并在0.7%低熔点琼脂糖凝胶上重新纯化。将纯化的片段亚克隆进质粒pSVL(pharmacia),该质粒已首先用XbaI和BamHI消化并在0.8%低熔点琼脂糖凝胶上重新纯化。在用T4连接酶连接之后,将混合物用于转化AG1大肠杆菌,然后将其平铺在LB/氨苄青霉素平板上,37℃过夜培养。将得自氨苄毒霉素抗性的菌落的质粒DNA通过用Xho和BamHI消化分析,以证实插入物(它在这一消化中被切去)的存在。发现6个克隆含有插入物,选择其中的一个(克隆7)用于进步一的研究并命名为pSVLTBPhCGα(含有TBP1(20-161)-hCGα)。对这一载体中插入物的双脱氧DNA测序(使用SeguenaseTM,U.S.Biochemicals.Cleveland,Ohio)的结果证实了该构建体是正确的,并且没有引入非期望的变化。
最终编码TBP1(20-161)-hCGβ的序列是以对TBP1(20-161)-hCGα所述相同的方式使用引物1(αβ)和4(β)的融合PCR组装的,并使用了从得自第一个PCR反应的p55和hCGβ片段纯化的产物作为模板。所产生含有期望的插入物的pSVL被命名为pSVLTBPhCGβ。
b。TBP(20-190)-hCG
通过对TBP(20-161)-hCG构建体进行修饰制备出第二组TBP-hCG蛋白质,以产生含有横跨Asp20至Thr 190的TBP的类似物,以代替原生类似物中的20-161区段。将通过将在质粒pSVLTBPhCGα中的BglII和XbaI位点之间的片段用含有该变化的PCR片段替换来进行的,这一PCR片段是使用融合PCR产生的。这些引物为:
引物1 TTT TAG ATC TCT TCT TGC ACA GTG GAC(SEQ ID NO:18)
引物2 TGT GGT GCC TGA GTC CTC AGT(SEQ ID NO:19)
引物3 ACT GAG GAC TCA GGC ACC ACA GCC GGT GCT GCC CCA GGT TG
(SEQ ID NO:20)
引物 4TTT TTC TAG AGA AGC AGC AGC AGC CCA TG(SEQ ID NO:21)
引物1和2被用于产生编从161-190的额外的p55残基的序列。该PCR反应基本上是如前所述的条件进行的,使用1μg的每种引物和pUC-p55作为模板。同样,引物3和3被用于通过PCR产生TBP编码区段的3’一端和编码hCGα亚基5’端之间的连接物,使用的是质粒pSVLTBPhCGα作为模板。在8%凝胶上的聚丙烯酰胺凝胶电泳(PAGE)证实了这些PCR反应的产物为正确大小(分别为约296bp和121bp),然后使用Wizard柱将其纯化。引物2和3的设计是这样的,它们含有重叠区段,从而使这两种PCR产物(得自引物1和2和得自引物3和4)可退火用于使用引物1和4的融合PCR。融合反应之后,将给400bp的期望产物用1.5%琼脂糖凝胶和Wizard柱进行证实和纯化。然后将这种DNA用BglII和XbaI进行消化,并与BglII/Xba I消化的pSVLTBPhCGα进行连接。在从转化的AG1大肠杆菌中分离的质粒中插入物的存在通过用BglII和XbaI的消化得到了证实。该新的构建体被命名为pSVLTBP(20-190)-hCGα。
同样,将质粒pSVLTBPhCGβ通过BglII和XcmI片段的替换来修饰,但是,这是通过单个PCR产物的亚克隆来进行的,而不是融合PCR产物。使用了引物1和2b(见下文),以及作为模板的pUC-p55。
引物2b TTT TCC ACA GCC AGG GTG GCA TTG ATG GGG CGG CAC CGT GGA CCAGCA CCA GCT GTG GTG CCT GAG TCC TCA GTG(SEQ ID NO:22)
将产生的PCR产物(约3376p)用上述方法进行证实和纯化,用BglII和XcmI消化,然后连接进用BglII/Xba I消化的pSVLTBPhCGβ中。对从得自转化的AG1大肠杆菌分离的质粒中插入物的存在通过用BglII和XcmI的消化来证实,该新构建体被命名为pSVLTBP(20-190)-hCGβ。
随后对该新构建体用DNA测序来证实。
除了产生这些新的以pSVL为基的质粒外,这些构建体也被亚克隆进其它可能更适于在CHO中稳定表达的其它表达载体中,特别是载体D2,以前被描述为质粒CLH3AXSV2DHFR(45)。这是通过将侧接基于pSVL的载体中的插入物的BamHI位点转换成XhoI位点,然后用XhoI切除该插入物并将它克隆进Xhol消化的Dα中。
2、杂种蛋白质的瞬间和稳定表达
用于瞬间表达TBP-hCG杂种蛋白质的COS-7细胞(ATCCCRL 1651,ref.46)的转染是使用电穿孔进行的(47)。将指数生长的COS-7细胞通过胰蛋白酶消化除去,通过轻微离心(800rpm,4分钟)收集,用冷的磷酸盐缓冲盐水(PBS),pH7.3-7.4洗涤,然后通过离心重新团聚。将细胞以每400μl冷PBS为5×106细胞的浓度重新悬浮并在预冷冻的2mm缺口电穿孔小槽中与10μg质粒DNA混合。对于共转染来说使用35μg的每种质粒。将小槽和细胞在冰上再冷冻10分钟,然后使用BTX600型仪器和125V,950μF和R=8的条件进行电穿孔。之后将细胞在冰上放置10分钟使之冷冻,转移至一个15ml含有9.5ml室温温度的完全培养基中(补充了10%小牛血清(FBS)的1%L-谷氨酰胺的Dulbecco’s改良Eagle’s培养基(DMEM)),并在室温下放置5分钟。在该15ml试管中轻微混合后,将全部内含物接种到两个P100平板上并置于一个37℃,5%CO2温箱中。18小时后,改换培养基,并且在一些情况下该新的培养基仅含有1%或0%FBS。再过72小时后,收获该条件培养基,离心除去细胞,然后在-70℃冷冻贮存。
用于瞬间或稳定表达的CHO-DUKX(CHO)细胞的转染是使用DNA的磷酸钙沉淀法进行的,转染前24小时,将指数生长的CHO细胞以每平板7.5×105细胞的密度铺布在100mm培养板上。在转染的当天,将10μg质粒DNA放入0.5ml转染缓冲液中(见下文),加入31μl的2MCaCl2,将该DNA-CaCl2溶液进行涡旋混合,并在室温下放置45分钟。在此之后,将培养基从平板上吸去,使用一个无菌塑料管将DNA加到细胞中,并将这些细胞在室温下放置20分钟,在此期间结束时,向平板中加入5ml含有10%FBS的完全α(+)MEM,将其在37℃温育4-6小时。然后将培养基从平板上吸去,将这些细胞在37℃用转染缓冲液中的15%甘油液温育3.5分钟而进行甘油冲击。在将该甘油溶液除去后,将细胞用PBS洗两次,再加入10ml完全α(+)MEM,10%FBS,并放回37℃培养箱。为了稳定转染,48小时后将这些细胞1∶10分开用选择培养基(完全α(-)MEM(缺乏核苷),10%透析的FBS,和0.02μM氨甲蝶呤)喂养。非转染的(非抗性的)细胞一般情况下在3-4周中被除去,剩下了转染的,氨甲蝶呤抗性的细胞群。
3、表达的定量分析
转染细胞的杂种蛋白质的分泌是使用检测可溶性p55的商品试剂盒(R&D Systems;Minneapolis,Minnesota),按照制造商的说明进行检测的。这一检测也提供了一种对条件化的和处理的培养基中杂种蛋白质的估测,以此为基础选择用于生物检测中的剂量。
4、对异源二聚体形成的评估
为评估TBP-hCG亚基融合体结合和形成异源二聚体的能力,进行了使用针对hCG亚基的抗体的夹心面包免疫检测(Sandwichimmunoassay)。在这一检测则中,将针对hCGβ亚基的单克隆抗体涂敷在微滴定板上并用于分析物捕获。主要的检测抗体是一种山羊多克隆抗体,它是针对人TSHα亚基培育的(#082422G-BiodesignInternational;Kennenbunkport,Maine),反过来使用辣根过氧化物酶偶联的兔抗羊多克隆抗体(Cappel;Durham,North Carolina)检测该山羊抗体。
在这一工作中使用几种不同的抗hCGβ亚基抗体,它们对自由α亚基均设有显示出可检测的交叉反应性。这些抗体中的一个(3/6)被用于商品MAI Aclone hCG检测试剂盒(Biodata;Romt,Italy)。
将高-蛋白质结合微滴板(Costar#3590)涂布以捕获抗体,通过温育(2小时,37℃)用100μl/孔涂敷缓冲液(PBS,pH7.4,01mMCa++,0.1mM Mg++)中的5μg/ml抗体溶液进行涂敷。用洗涤溶液(PBS,pH7.4+0.1%Tween20)洗过一次之后,通过用封闭溶液(PBS中3%小牛血清白蛋白(BSA;级分V-A-4503Sigma),pH7.4)将孔完全充满(约400μl/孔)并在37℃温育1小时或者在4℃过夜温育将板封闭,然后将板用洗涤缓冲液洗涤两次,并将参照和实验样品,在用稀释液(PBS中的5mg/ml BSA,pH7.4)稀释至100μl体积后加入其中。在将板和样品在37℃温育2小时后,将板用洗涤溶液再洗涤两次。加入在稀释液中稀释1∶5000的第一检测抗体(100μl/孔)并在37℃温育1小时。加入在稀释液中稀释1∶5000的第二检测抗体(HRP偶联的免抗羊Ig)(100μl/孔),并在37℃温育1小时后,用洗涤溶液将板洗涤三次。加入100μl TMB底物溶液(Kirkegaard和Perry实验室),将板在黑暗中室温下温育20分钟,然后通过加入50μl/孔,0.3M硫酸终止酶反应。然后使用微滴板读出器在波长450nm处对板进行分析。
5、部分纯化
为了更好地对这些杂种蛋质的活性进行定量,用免疫吸附层析法将TBP-hCG杂种蛋白质进行部分纯化。所用的抗体是一种可通过商业途径从R&D System(MAB#225)得到的单克隆抗体。柱为CNBr活化的琼脂糖,按照制造商(Pharmacia)的说明进行装填。
条件培养基是使用每天收获的50μl SFMII培养基(GIBC0)从每一细胞系的汇合T-175瓶中收集的,每一细胞系有5个收获物。将收集物进行离心(1000RPM)以除去细胞碎片。然后使用商品免疫检测试验对该材料进行TBP含量检测并进行浓缩(Centricon un:ts by Arnicon;Beverly,Massachusetts),使表观TBP浓度为约50ng/ml。
将10ml浓缩的TBP-hCG(样品#188737)通过加入NaCl并将该溶液的导电率调整至给85mS/cm而使其具有约1M NaCl。将它通过0.5ml的抗-TBP免疫亲和柱。收集流出物并使其第二次通过该柱。之后用PBS中的1M NaCl洗涤该柱。在用50mM柠檬酸(pH2.5)洗脱之后收集结合的TBP(20-161)-hCG。将洗脱液(约7ml)通过使用Amicon Centricon-10’s并按照制造商(Amicon)的说明进行的过滤进行浓缩,使其达到约200μl的体积。加入约800μl的PBS使样品体积达到1ml,4℃储存直至用生物检测法进行测试。
6、抗-TNF活性的评估。
多种用于评估可溶性TNF受体的类似物的体外TNF诱导的细胞毒性检测方法已有描述。我们使用了一种使用人乳腺癌细胞系。BT-20细胞(ATCC HTB19)的检测方法。这些细胞作为TNF生物检测的基础的使用已有描述(48)。将这些细胞在37℃培养在补充了10%热失活TBS的RPMI1640培养基中,将细胞生长至最大80-90%汇合,这样每3-4天限制了分裂,种子密度为每T175cm2瓶约3×106个细胞。
BT-20检测使用了一种细胞染料,结晶紫,的着色,以此作为用TNF处理后对存活细胞评估的检测方法,死亡细胞不能摄取并保持该染料。
简言之,用于检测抗TNF活性的方法如下。将重组人TNFα(R&D systems)和实验样品配制于培养基中(RPMI 1640和5%热灭活的FBS,并加到96孔培养板上的孔中。然后将这些细胞以1×105细胞/孔的密度平铺到这些孔中。加入的TNFα的量是早先在滴定研究中测定的,并代表一种约50%的细胞被杀死的剂量。
样品加入之后,将细胞在39℃培养48小时,之后,使用结晶紫染色和微滴板读出器(570nm)确定活细胞的比例。
结果
1、被研究的构建体
对研究的杂种蛋白质的设计总结于下;对两种对照蛋白质,一种单体可溶性p55(r-hTBP-1)和一种二聚体TBP-免疫球蛋白融合蛋白质(TBP-IgG3)(基本上如(10)所述制备的)进行了研究,以进行比较。
构建体 TNPN-末端 TBPC-末端 融合对应物
r-hTBP-1 9和20的混合物 180 无
TBP-IgG3 9和20的混合物 190 IgG3重链恒定区
TBP(20-161)-hCG 20 161 hCGα和hCGβ(异源二聚体)
(异源二聚体)(异源二聚体)
TBP(20-190)-hCG 20 190 hCGα和hCGβ
编码TBP(20-190)-hCG和TBP(20-161)-hCG的DNA的序列分别示于图1和2。
2、TBP-hCG蛋白质的分泌
发现所有这些构建体被转染的哺乳动物细胞产生并分泌至培养基中。表示这些结果的数据示于表1和表2。
3、组装成异源二聚体的TBP-hCG(αβ)融合蛋白质
TBP-hCGα和TBP-hCGβ的结合是使用hCG异源二聚体的夹心饼检测法得证实的,只有α和β亚基融合物的联合转染产生异源二聚体检测(表3)。
4、TBP-hCG杂种蛋白质与TBP单体相比显示出提高了的活性。
在COS-7或CHO细胞中产生的染种蛋白质被发现为在BT-20生和检测中为TNFα的强抑制物。一些样品的检测结果总结于表4。
负对照(来自模拟转染的条件培养基)被包括在1X培养基样品中。
如图3-5所示(Y轴上的点),TNF的加入(2.5ng/ml)导致活细胞数的明显降低(由OD570测得)。在每一种情况下,活性样品所具有的最大保护效能是将细胞存活性恢复到在不加入TNF时所看到的水平(即,标有“只有细胞”的对照)
正对照r-hTBP-1和TBP-IgG3均为保护性的,显示出明显的剂量依赖性,并且r-hTBP-1的ED50为约100ng/ml(图3-5),TBP-TgG3的ED50为约1.5ng/ml(图3)。
来自1X培养基或来自免疫纯化物的TBP-hCG构建体(CHO或COS)显示出剂量依赖性的保护,其ED50在约2-11ng/ml的范围内(图3-5)。
体外生物检测的结果描述于表5。这些结果表明,该杂种蛋白质抑制了TNF细胞毒性,并且它们基本上比TBP单体更有效力。负对照没有保护活性。
除了TBP的二聚体化可增强效力的可能性外,也有可能该杂种蛋白质的活性与TBP的二聚体相互作用没有关系,而是与空间位阻有关,这是由于该杂种蛋白质的参与物干扰可溶性TBP/TNF与细胞表面TNF受体的结合。
本文引用的所有参考文献,包括期刊论文或摘要,公开的或相应的美国或外国专利申请,已颁发的美国或外国专利,或其它任一种参考文献,合部引入本文作为参考,包括这些参考文献中全部的数据,表、图和正文。此外,本文参考文献中所引用的参考文献内容也全部引入本文。
对已知的方法步骤、常规的方法步骤、已知的方法或常规的方法的参考并不意味着我们承认本发明的任一方面,描述或实施方案已在这些相关文献中被公开,教导或提示。
前文对特定实施例的描述对本发明的总体的性质进行了完整的披露,其他人应用本领域技术的知识(包括本文引用的参考文献)可容易地对这些特定实施例进行修改和调整,而不需要进行过多的实验,但却不偏离本发明的总体内容。因而,基于本文的教导和指示,这些调整和修改应包含于本文公开的实施例等同意思和范围之内。本文的措辞和用语应被理解为为了更清楚描述的目的,而不限定其范围。本发明书的诸词和用语应由本技术领域人普通技术人员根据本文的教导和指示并结合本领域普通技术人员的现有知识进行解释。
表1:COS-7瞬间表达(TBP ELISA) | |
杂种蛋白质 | 浓度(pg/ml) |
TBP1 | 66 |
TBP-hCGα(20-161) | 5.1 |
TBP-hCGβ(20-161) | 0.5 |
TBP-hCG(20-161) | 2.7 |
对照 | <0.25 |
构建体是使用pSVL(Pharmacia)表达的
表2:COS-7瞬间表达(TBP ELISA) | |
杂种蛋白质 | 浓度(ng/ml) |
TBP1 | 131 |
TBP-hCGα(20-190) | 81 |
TBP-hCGβ(20-190) | 9 |
TBP-hCG(20-190) | 62 |
对照 | <1 |
构建体是使用含有小鼠金属硫蛋白启动子的载体-pDα表达的
表3:COS-7瞬间表达(hCG二聚体检测) | |
杂种蛋白质 | 浓度(ng/ml) |
TBP1 | <0.2 |
TBP-hCGα(20-190) | <0.2 |
TBP-hCGβ(20-190) | <0.2 |
TBP-hCG(20-190) | 38 |
对照 | <0.2 |
构建体是使用小鼠金属硫蛋白启动子的载体-pDα表达的
表4:对样品检测抗-TNF活性 | ||
构建体 | 细胞来源 | 样品性质 |
r-hTBP-1 | CHO | 纯化的 |
TBP-1gG3 | CHO | IX条件培养基 |
TBP(20-161)-hCG | CHO | 免疫纯化的(抗-TBP) |
TBP(20-190)-hCG | CHO | IX条件培养基 |
TBP(20-190)-hCG | COS | IX条件培养基 |
表5:对杂种蛋白质在TNF细胞毒性检测中的初步评估 | ||
构建体 | 融合参与物 | 在BT-20生物检测中的抗TNF活性(ED50)** |
r-hTBP-1 | 无 | 100ng/ml |
TBP-lgG3 | 1gG3重链恒定区 | 1.5ng/ml |
TBP(20-161)-hCG | hCGα和hCGβ(异源二聚体) | 2ng/ml |
TBP(20-190)-hCG | hCGα和hCGβ(异源二聚体) | 8-11ng/ml |
**用于材料的剂量的定量和ED50的估测是使用TBPELISA进行的
参考文献
1.Smith,R.A.et al.,J.Biol.Chem.262:6951-6954,1987.
2.Eck,M.J.et al.,J.Biol.Chem.264:17595-17605,1989.
3.Jones,E.Y.et al ,Nature 338:225-228,1989.
4.Eck,M.J.et al.,J.Biol.Chem.267:2119-2122,1992.
5.Pierce,J.G.et al.,Annu.Rev.Biochem.50:465-495,1981.
6.Lapthorn,A.J.et al.,Nature 369:455-461,1994.
7.Wu,H.,et al.,Structure 2:545-550,1994.
8.Engelmann,H.,et al.,J.Biol.Chem.265:14497-14504,1990.
9.Adam,D.et al.,J.Biol.Chem.270:17482-17487,1995.
10.Loetscher,H.R.,et al.,J.Biol.Chem.266:18324-18329,1991.
11.Banner,D.W.,et al.,Cell 73:431-445,1993.
12.Pennica,D.,et al.,Biochemistry 32:3131-3138,1993.
13.Engelmann,H.et al.,J.Biol.Chem.265:1531-1536,1990.
14.Van Zee,K.J.et al.,Proc.Natl.Acad.Sci.USA 89:4845-4849,1992.
15.Aderka,D.et al.,J.Exp.Med.175:323-329,1992.
16.Mohler,K.M.,et al.,J.Immunol.151:1548-1561,1993.
17.Bertini,R.,et al.,Eur.Cytokine Netw.,1993.
18.Piguet,P.F.,et al.,Immunology 77:510-514,1992.
19.Williams,R.O.,et al.,Immunology 84:433-439,1995.
20.Capon,D.J.,et al.,Nature 337:525-531,1989.
21.Ashkenazi,A.,et al.,Proc.Natl.Acad.Sci.88:10535-10539,1991.
22.Suitters,A.J.,et al.J.Exp.Med.179:849-856,1994.
23.Nolan,O.et al.,Biochim.Biophys.Acta 1040:1-11,1990.
24.Rodrigues,M.L.,et al.,J.Immunol.151:6954-6961,1993.
25.Chang,H.-C.,et al.,Proc.Natl.Acad.Sci.USA 91:11408-11412,1994.
26.Wu,Z.,et al.,J.Biol.Chem.270:16039-16044,1995.
27.Bazzoni,F.et al,Proc.Natl.Acad.Sci.USA92:5376-5380,1995.
28.Boldin,M.P.,et al.,J.Biol.Chem.270:387-391,1995.
29.Vu,T.-K.H.,et al.,Cell,64:1057-1068,1991.
30Song,H.Y.,et al.,J.Biol.Chem.269:22492-22495,1994.
31.Russell,D.A.,et al.,J.Infectious Diseases171:1528-1538,1995.
32.Rao C.V.et al.,Am.J.Obstet.Gynecol.,146,65-68,1983.
33.Damewood M.D.et al.,Fertil.Steril.52,398-400,1989.
34.Chen,F.,et al.,Mol.Endocrinol.6:914-919,1992.
35.Bielinska,M.,et al.,J.Cell Biol.111:330a,1990.
36.Furuhashi,M.,et al.,Mol Endocrinol.9:54-63,1995.
37.Sugahara,T.,et al.,Proc.Natl.Acad.Sci.USA 92:2041-2045,1995.
38.Johnson,G.A.,et al.,Biol.Reprod.52:68-73,1995.
39.Urlaub,G.and Chasin,L.Proc.Natl.Acad.Sci.USA 77:4216-4220,1980.
40.Nophar,Y.,et al.,EMBO J.9:3269-3278,1990.
41.Fiddes,J.C.et al.,Nature 281:351-356,1979.
42.Fiddes,J.C.et al.,Nature 286:684-687,1980.
43.Elion,E.A.,in Current Protocols in MolecularBiology,eds.Ausuble,FM.et al.,John Wiley & Sons,1993.
44.Campbell,R.,Proc.Natl.Acad.Sci.USA 88:760-764,1991.
45.Cole E.S.et al.,Biotechnology,11,1014-1024,1993.
46.Gluzman,Y.,Cell 23:175-182,1981.
47.Chu,G.et al.,Nucl.Acid Res.15:1311-1326,1987.
48.Yen,J.et al.,J.Immunotherapy 10:174-181,1991.
序列表
(1)总信息:
(i)申请人:
(A)名称:应用研究系统ARS控股公司
(B)街道:14John B.Gorsiraweg
(C)城市:Curacao
(D)国家:Netherlands Antilles
(E)邮政编码:
(A)姓名:CAMPBELL,Robert C.
(B)街道:25Meadowbrood Drive
(C)城市:Wrentham
(D)州:Massachusetts
(E)国家:USA
(A)姓名:JAMESON,Bradford A.
(B)街道:76Robbins Street
(C)城市:Milton
(D)州:Massachusetts
(E)国家:USA
(A)姓名:CHAPPEL,Scott C.
(B)街道:125Canton Avenue
(C)城市:Milton
(D)州:Massachusetts
(E)国家:USA
(ii)发明名称:杂种蛋白质
(iii)序列数目:22
(iv)通信地址:
(A)收信人:BROWDY AND NEIMARK
(B)街道:419Seventh Street N.W.,Ste.300
(C)城市:Washington
(D)州:D.C.
(E)国家:USA
(F)邮政编码:22207
(v)计算机可读形式:
(A)介质类型:软盘
(B)计算机:IBM PC兼容
(C)操作系统:PC-DOS/MS-DOS
(D)软件:PatentIn Release#1.0,Version#1.30
(vi)在先申请资料:
(A)申请号:60/011936
(B)申请日:1996年2月20日
(C)分类:
(viii)代理人信息:
(A)姓名:Browdy,RogerL.
(B)登记号:25618
(C)案号:CAMPBELL=2A PCT
(ix)通讯资料:
(A)电话:(202)628-5197
(B)传真:(202)737-3528
(2)SEQ ID NO:1的信息:
(i)序列特征:
(A)长度:1049碱基对
(B)类型:核酸
(C)链型:单链
(D)拓扑结构:线性
(ii)分子类型:cDNA
(ix)特点:
(A)名称/关键词:CDS
(B)位置:278..1047
(xi)序列描述:SEQ ID NO:1
TCCACATGGC TACAGGTAAG CGCCCCTAAA ATCCCTTTGG GCACAATGTG TCCTGAGGGG 60
AGAGGCAGCG ACCTGTAGAT GGGACGGGGG CACTAACCCT CAGGTTTGGG GCTTCTCAAT 120
CTCACTATCG CCATGTAAGC CCAGTATTTG GCCAATCTCA GAAAGCTCCT CCTCCCTGGA 180
GGGATGGAGA GAGAAAAACA AACAGCTCCT GGAGCAGGGA GAGTGCTGGC CTCTTGCTCT 240
CCGGCTCCCT CTGTTGCCCT CTGGTTTCTC CCCAGGC TCC CGG ACG TCC CTG CTC 295
Ser Arg Thr Ser Leu Leu
1 5
CTG GCT TTT GGC CTG CTC TGC CTG CCC TGG CTT CAA GAG GGC AGT GCC 343
Leu Ala Phe Gly Leu Leu Cys Leu Pro Trp Leu Gln Glu Gly Ser Ala
10 15 20
GAT AGT GTG TGT CCC CAA GGA AAA TAT ATC CAC CCT CAA AAT AAT TCC 391
Asp Ser Val Cys Pro Gln Gly Lys Tyr Ile His Pro Gln Asn Asn Ser
25 30 35
ATT TGC TGT ACC AAG TGC CAC AAA GGA ACC TAC TTG TAC AAT GAC TGT 439
Ile Cys Cys Thr Lys Cys His Lys Gly Thr Tyr Leu Tyr Asn Asp Cys
40 45 50
CCA GGC CCG GGG CAG GAT ACG GAC TGC AGG GAG TGT GAG AGC GGC TCC 487
Pro Gly Pro Gly Gln Asp Thr Asp Cys Arg Glu Cys Glu Ser Gly Ser
55 60 c 65 70
TTC ACC GCT TCA GAA AAC CAC CTC AGA CAC TGC CTC AGC TGC TCC AAA 535
Phe Thr Ala Ser Glu Asn His Leu Arg His Cys Leu Ser Cys Ser Lys
75 80 85
TGC CGA AAG GAA ATG GGT CAG GTG GAG ATC TCT TCT TGC ACA GTG GAC 583
Cys Arg Lys Glu Met Gly Gln Val Glu Ile Ser Ser Cys Thr Val Asp
90 95 100
CGG GAC ACC GTG TGT GGC TGC AGG AAG AAC CAG TAC CGG CAT TAT TGG 631
Arg Asp Thr Val Cys Gly Cys Arg Lys Asn Gln Tyr Arg His Tyr Trp
105 110 115
AGT GAA AAC CTT TTC CAG TGC TTC AAT TGC AGC CTC TGC CTC AAT GGG 679
Ser Glu Asn Leu Phe Gln Cys Phe Asn Cys Ser Leu Cys Leu Asn Gly
120 125 130
ACC GTG CAC CTC TCC TGC CAG GAG AAA CAG AAC ACC GTG TGC ACC TGC 727
Thr Val His Leu Ser Cys Gln Glu Lys Gln Asn Thr Val Cys Thr Cys
135 140 145 150
CAT GCA GGT TTC TTT CTA AGA GAA AAC GAG TGT GTC TCC TGT GCC GGT 775
His Ala Gly Phe Phe Leu Arg Glu Asn Glu Cys Val Ser Cys Ala Gly
155 160 165
GCT GCC CCA GGT TGC CCA GAA TGC ACG CTA CAG GAA AAC CCA TTC TTC 823
Ala Ala Pro Gly Cys Pro Glu Cys Thr Leu Gln Glu Asn Pro Phe Phe
170 175 180
TCC CAG CCG GGT GCC CCA ATA CTT CAG TGC ATG GGC TGC TGC TTC TCT 871
Ser Gln Pro Gly Ala Pro Ile Leu Gln Cys Met Gly Cys Cys Phe Ser
185 190 195
AGA GCA TAT CCC ACT CCA CTA AGG TCC AAG AAG ACG ATG TTG GTC CAA 919
Arg Ala Tyr Pro Thr Pro Leu Arg Ser Lys Lys Thr Met Leu Val Gln
200 205 210
AAG AAC GTC ACC TCA GAG TCC ACT TGC TGT GTA GCT AAA TCA TAT AAC 967
Lys Asn Val Thr Ser Glu Ser Thr Cys Cys Val Ala Lys Ser Tyr Asn
215 220 225 230
AGG GTC ACA GTC ATG GGG GGT TTC AAA GTG GAG AAC CAC ACG GGG TGC 1015
Arg Val Thr Val Met Gly Gly Phe Lys Val Glu Asn His Thr Gly Cys
235 240 245
CAC TGC AGT ACT TGT TAT TAT CAC AAA TCT TA AG 1049
His Cys Ser Thr Cys Tyr Tyr His Lys Ser
250 255
(2)SEQ ID NO:2的信息:
(i)序列特征:
(A)长度:256氨基酸
(B)类型:氨基酸
(C)拓扑结构:线性
(ii)分子类型:蛋白质
(xi)序列描述:SEQ ID NO:2
Ser Arg Thr Ser Leu Leu Leu Ala Phe Gly Leu Leu Cys Leu Pro Trp
1 5 10 15
Leu Gln Glu Gly Ser Ala Asp Ser Val Cys ProGln Gly Lys Tyr Ile
20 25 30
His Pro Gln Asn Asn Ser Ile Cys Cys Thr Lys Cys His Lys Gly Thr
35 40 45
Tyr Leu Tyr Asn Asp Cys Pro Gly Pro Gly Gln Asp Thr Asp Cys Arg
50 55 60
Glu Cys Glu Ser Gly Ser Phe Thr Ala Ser Glu Asn His Leu Arg His
65 70 75 80
Cys Leu Ser Cys Ser Lys Cys Arg Lys Glu Met Gly Gln Val Glu Ile
85 90 95
Ser Ser Cys Thr Val Asp Arg Asp Thr Val Cys Gly Cys Arg Lys Asn
100 105 110
Gln Tyr Arg His TVr Trp Ser Glu Asn Leu Phe Gln Cys Phe Asn Cys
115 120 125
Ser Leu Cys Leu Asn Gly Thr Val His Leu Ser Cys Gln Glu Lys Gln
130 135 140
Asn Thr Val Cys Thr Cys His Ala Gly Phe Phe Leu Arg Glu Asn Glu
145 150 155 160
Cys Val Ser Cys Ala Gly Ala Ala Pro Gly Cys Pro Glu Cys Thr Leu
165 170 175
Gln Glu Asn Pro Phe Phe Ser Gln Pro Gly Ala Pro Ile Leu Gln Cys
180 185 190
Met Gly Cys Cys Phe Ser Arg Ala Tyr Pro Thr Pro Leu Arg Ser Lys
195 200 205
Lys Thr Met Leu Val Gln Lys Asn Val Thr Ser Glu Ser Thr Cys Cys
210 215 220
Val Ala Lys Ser Tyr Asn Arg Val Thr Val Met Gly Gly Phe Lys Val
225 230 235 240
Glu Asn His Thr Gly Cys His Cys Ser Thr Cys Tyr Tyr His Lys Ser
245 250 255
(2)SEQ ID NO:3的信息:
(i)序列特征:
(A)长度:1202碱基对
(B)类型:核酸
(C)链型:单链
(D)拓扑结构:线性
(ii)分子类型:cDNA
(ix)特点:
(A)名称/关键词:CDS
(B)位置:279..1199
(xi)序列描述:SEQ ID NO:3
CTCGAGATGG CTACAGGTAA GCGCCCCTAA AATCCCTTTG GGCACAATGT GTCCTGAGGG 60
GAGAGGTAGC GACCTGTAGA TGGGACGGGG GCACTAACCC TGAGGTTTGG GGCTTCTGAA 120
TGTGAGTATC GCCATGTAAG CCCAGTATTT GGCCAATGTC AGAAAGCTCC TGGTCCCTGG 180
AGGGATGGAG AGAGAAAAAC AAACAGCTCC TGGAGCAGGG AGAGTGCTGG CCTCTTGCTC 240
TCCGGCTCCC TCTGTTGCCC TGTGGTTTCT CCCCAGGC TCC CGG ACG TCC CTG 293
Ser Arg Thr Ser Leu
260
CTC CTG GCT TTT GGC CTG CTC TGC CTG CCC TGG CTT CAA GAG GGC AGT 341
Leu Leu Ala Phe Gly Leu Leu Cys Leu Pro Trp Leu Gln Glu Gly Ser
265 270 275
GCC GAT AGT GTG TGT CCC CAA GGA AAA TAT ATC CAC CCT CAA AAT AAT 389
Ala Asp Ser Val Cys Pro Gln Gly Lys Tyr Ile His Pro Gln Asn Asn
280 285 290
TCG ATT TGC TGT ACC AAG TGC CAC AAA GGA ACC TAC TTG TAC AAT GAC 437
Ser Ile Cys Cys Thr Lys Cys His Lys Gly Thr Tyr Leu Tyr Asn Asp
295 300 305
TGT CCA GGC CCG GGG CAG GAT ACG GAC TGC AGG GAG TGT GAG AGC GGC 485
Cys Pro Gly Pro Gly Gln Asp Thr Asp Cys Arg Glu Cys Glu Ser Gly
310 315 320 325
TCT TTC ACC GCT TCA GAA AAC CAC CTC AGA CAC TGC CTC AGC TGC TCC 533
Ser Phe Thr Ala Ser Glu Asn His Leu Arg His Cys Leu Ser Cys Ser
330 335 340
AAA TGC CGA AAG GAA ATG GGT CAG GTG GAG ATC TCT TCT TGC ACA GTG 581
Lys Cys Arg Lys Glu Met Gly Gln Val Glu Ile Ser Ser Cys Thr Val
345 350 355
GAC CGG GAC ACC GTG TGT GGC TGC AGG AAG AAC CAG TAC CGG CAT TAT 629
Asp Arg Asp Thr Val Cys Gly Cys Arg Lys Asn Gln Tyr Arg His Tyr
360 365 370
TGG AGT GAA AAC CTT TTC CAG TGC TTC AAT TGC AGC CTC TGC CTC AAT 677
Trp Ser Glu Asn Leu Phe Gln Cys Phe Asn Cys Ser Leu Cys Leu Asn
375 380 385
GGG ACC GTG CAC CTC TCC TGC CAG GAG AAA CAG AAC ACC GTG TGC ACC 725
Gly Thr Val His Leu Ser Cys Gln Glu Lys Gln Asn Thr Val Cys Thr
390 395 400 405
TGC CAT GCA GGT TTC TTT CTA AGA GAA AAC GAG TGT GTC TCC TGT GCT 773
Cys His Ala Gly Phe Phe Leu Arg Glu Asn Glu Cys Val Ser Cys Ala
410 415 420
GGT GCT GGT CCA CGG TGC CGC CCC ATC AAT GCC ACC CTG GCT GTG GAG 821
Gly Ala Gly Pro Arg Cys Arg Pro Ile Asn Ala Thr Leu Ala Val Glu
425 430 435
AAG GAG GGC TGC CCC GTG TGC ATC ACC GTC AAC ACC ACC ATC TGT GCC 869
Lys Glu Gly Cys Pro Val Cys Ile Thr Val Asn Thr Thr Ile Cys Ala
440 445 450
GGC TAC TGC CCC ACC ATG ACC CGC GTG CTG CAG GGG GTC CTC CCC GCC 917
Gly Tyr Cys Pro Thr Met Thr Arg Val Leu Gln Gly Val Leu Pro Ala
455 460 465
CTG CCT CAG GTG GTG TGC AAC TAC CGC GAT GTG CGC TTC GAG TCC ATC 965
Leu Pro Gln Val Val Cys Asn Tyr Arg Asp Val Arg Phe Glu Ser Ile
470 475 480 485
CGG CTC CCT GGC TGC CCG CGC GGC GTG AAC CCC GTG GTC TCC TAC GCT 1013
Arg Leu Pro Gly Cys Pro Arg Gly Val Asn Pro Val Val Ser Tyr Ala
490 495 500
GTG GCT CTC AGC TGT CAA TGT GCA CTC TGC CGC CGC AGC ACC ACT GAC 1061
Val Ala Leu Ser Cys Gln Cys Ala Leu Cys Arg Arg Ser Thr Thr Asp
505 510 515
TGC GGG GGT CCC AAG GAC CAC CCC TTG ACC TGT GAT GAC CCC CGC TTC 1109
Cys Gly Gly Pro Lys Asp His Pro Leu Thr Cys Asp Asp Pro Arg Phe
520 525 530
CAG GAC TCC TCT TCC TCA AAG GCC CCT CCC CCC AGC CTT CCA AGC CCA 1157
Gln Asp Ser Ser Ser Ser Lys Ala Pro Pro Pro Ser Leu Pro Ser Pro
535 540 545
TCC CGA CTC CCG GGG CCC TCG GAC ACC CCG ATC CTC CCA CAA TAA 1202
Ser Arg Leu Pro Gly Pro Ser Asp Thr Pro Ile Leu Pro Gln
550 555 560
(2)SEQ ID NO:4的信息:
(i)序列特征:
(A)长度:307氨基酸
(B)类型:氨基酸
(C)拓扑结构:线性
(ii)分子类型:蛋白质
(xi)序列描述:SEQ ID NO:4
Ser Arg Thr Ser Leu Leu Leu Ala Phe Gly Leu Leu Cys Lau Pro Trp
1 5 10 15
Leu Gln Glu Gly Ser Ala Asp Ser Val Cys Pro Gln Gly Lys Tyr Ile
20 25 30
His Pro Gln Asn Asn Ser Ile Cys Cys Thr Lys Cys His Lys Gly Thr
35 40 45
Tyr Leu Tyr Asn Asp Cys Pro Gly Pro Gly Gln Asp Thr Asp Cys Arg
50 55 60
Glu Cys Glu Ser Gly Ser Phe Thr Ala Ser Glu Asn His Leu Arg His
65 70 75 80
Cys Leu Ser Cys Ser Lys Cys Arg Lys Glu Met Gly Gln Val Glu Ile
85 90 95
Ser Ser Cys Thr Val Asp Arg Asp Thr Val Cys Gly Cys Arg Lys Asn
100 105 110
Gln Tyr Arg His Tyr Trp Ser Glu Asn Leu Phe Gln Cys Phe Asn Cys
115 120 125
Ser Leu Cys Leu Asn Gly Thr Val His Leu Ser Cys Gln Glu Lys Gln
130 135 140
Asn Thr Val Cys Thr Cys His Ala Gly Phe Phe Leu Arg Glu Asn Glu
145 150 155 160
Cys Val Ser Cys Ala Gly Ala Gly Pro Arg Cys Arg Pro Ile Asn Ala
165 170 175
Thr Leu Ala Val Glu Lys Glu Gly Cys Pro Val Cys Ile Thr Val Asn
180 185 190
Thr Thr Ile Cys Ala Gly Tyr Cys Pro Thr Met Thr Arg Val Leu Gln
195 200 205
Gly Val Leu Pro Ala Leu Pro Gln Val Val Cys Asn Tyr Arg Asp Val
210 215 220
Arg Phe Glu Ser Ile Arg Leu Pro Gly Cys Pro Arg Gly Val Asn Pro
225 230 235 240
Val Val Ser Tyr Ala Val Ala Leu Ser Cys Gln Cys Ala Leu Cys Arg
245 250 255
Arg Ser Thr Thr Asp Cys Gly Gly Pro Lys Asp His Pro Leu Thr Cys
260 265 270
Asp Asp Pro Arg Phe Gln Asp Ser Ser Ser Ser Lys Ala Pro Pro Pro
275 280 285
Ser Leu Pro Ser Pro Ser Arg Leu Pro Gly Pro Ser Asp Thr Pro Ile
290 295 300
Leu Pro Gln
305
(2)SEQ ID NO:5的信息:
(i)序列特征:
(A)长度:1147碱基对
(B)类型:核酸
(C)链型:单链
(D)拓扑结构:线性
(ii)分子类型:cDNA
(ix)特点:
(A)名称/关键词:CDS
(B)位置:278..1132
(xi)序列描述:SEQ ID NO:5
TCGAGATGGC TACAGGTAAG CGCCCCTAAA ATCCCTTTGG GCACAATGTG TCCTGAGGGG 60
AGAGGCAGCG ACCTGTAGAT GGGACGGGGG CACTAACCCT CAGGTTTGGG GCTTTTGAAT 120
GTGAGTATGG CCATGTAAGC CCAGTATTTG CCCAATCTCA GAAAGCTCCT GGTCCCTGGA 180
GGGATGGAGA GAGAAAAACA AACAGCTCCT GGAGCAGGGA CACTCCTGGC CTCTTGCTCT 240
GCGGCTCCGT GTGTTGCCCT GTGGTTTCTC CCCACGC TCC CGG ACG TCC CTG CTC 295
Ser Arg Thr Ser Leu Leu
310
CTG GCT TTT GGC CTG CTC TGC CTG CCC TGG CTT CAA GAG GGC AGT GCC 343
Leu Ala Phe Gly Leu Leu Cys Leu Pro Trp Leu Gln Glu Gly Ser Ala
315 320 325
GAT AGT GTG TGT CCC CAA GGA AAA TAT ATC CAC CCT CAA AAT AAT TCG 391
Asp Ser Val Cys Pro Gln Gly Lys Tyr Ile His Pro Gln Asn Asn Ser
330 335 340 345
ATT TGC TGT ACC AAG TGC CAC AAA GGA ACC TAC TTG TAC AAT GAC TGT 439
Ile Cys Cys Thr Lys Cys His Lys Gly Thr Tyr Leu Tyr Asn Asp Cys
350 355 360
CCA GGC CCG GGG CAG GAT ACC GAC TGC AGG GAG TGT GAG AGC GGC TCC 487
Pro Gly Pro Gly Gln Asp Thr Asp Cys Arg Glu Cys Glu Ser Gly Ser
365 370 375
TTC ACC GCT TCA GAA AAC CAC CTC AGA CAC TGC CTC AGC TGC TCC AAA 535
Phe Thr Ala Ser Glu Asn His Leu Arg His Cys Leu Ser Cys Ser Lys
380 385 390
TGC CGA AAG GAA ATG GGT CAG GTG GAG ATC TCT TCT TGC ACA GTG GAC 583
Cys Arg Lys Glu Met Gly Gln Val Glu Ile Ser Ser Cys Thr Val Asp
395 400 405
CGG GAC ACC GTG TGT GGC TGC AGG AAG AAC CAG TAC CGG CAT TAT TGG 631
Arg Asp Thr Val Cys Gly Cys Arg Lys Asn Gln Tyr Arg His Tyr Trp
410 415 420 425
AGT GAA AAC CTT TTC CAG TGC TTC AAT TGC ACC CTC TGC CTC AAT GGG 679
Ser Glu Asn Leu Phe Gln Cys Phe Asn Cys Thr Leu Cys Leu Asn Gly
430 435 440
ACC GTG CAC CTC TCC TGT CAG GAG AAA CAG AAC ACC GTC TGC ACC TGC 727
Thr Val His Leu Ser Cys Gln Glu Lys Gln Asn Thr Val Cys Thr Cys
445 450 455
CAT GCA GGT TTC TTT CTA AGA GAA AAC GAG TGT GTC TCC TGT AGT AAC 775
His Ala Gly Phe Phe Leu Arg Glu Asn Glu Cys Val Ser Cys Ser Asn
460 465 470
TGT AAG AAA AGC CTG GAG TGC ACG AAG TTG TCC CTA CCC CAG ATT GAG 823
Cys Lys Lys Ser Leu Glu Cys Thr Lys Leu Ser Leu Pro Gln Ile Glu
475 480 485
AAT GTT AAG GGC ACT GAG GAC TCA GGC ACC ACA GCC GGT GCT GCC CCA 871
Asn Val Lys Gly Thr Glu Asp Ser Gly Thr Thr Ala Gly Ala Ala Pro
490 495 500 505
GGT TGC CCA GAA TGC ACG CTA CAG GAA AAC CCA TTC TTC TCC CAG CCG 919
Gly Cys Pro Glu Cys Thr Leu Gln Glu Asn Pro Phe Phe Ser Gln Pro
510 515 520
GGT GCC CCA ATA CTT CAG TGC ATG GGC TGC TGC TTC TCT AGA GCA TAT 967
Gly Ala Pro Ile Leu Gln Cys Met Gly Cys Cys Phe Ser Arg Ala Tyr
525 530 535
CCC ACT CCA CTA AGG TCC AAG AAG ACG ATG TTG GTC CAA AAG AAC GTC 1015
Pro Thr Pro Leu Arg Ser Lys Lys Thr Met Leu Val Gln Lys Asn Val
540 545 550
ACC TCA GAG TCC ACT TGC TGT GTA GCT AAA TCA TAT AAC AGG GTC ACA 1063
Thr Ser Glu Ser Thr Cys Cys Val Ala Lys Ser Tyr Asn Arg Val Thr
555 560 565
GTA ATG GGG GGT TTC AAA GTG GAG AAC CAC ACG GCG TGC CAC TGC AGT 1111
Val Met Gly Gly Phe Lys Val Glu Asn His Thr Ala Cys His Cys Ser
570 575 580 585
ACT TGT TAT TAT CAC AAA TCT TAAGGATCCC TCGAG 1147
Thr Cys Tyr Tyr His Lys Ser
590
(2)SEQ ID NO:6的信息:
(i)序列特征:
(A)长度:285氨基酸
(B)类型:氨基酸
(D)拓扑结构:线性
(ii)分子类型:蛋白质
(xi)序列描述:SEQ ID NO:6
Ser Arg Thr Ser Leu Leu Leu Ala Phe Gly Leu Leu Cys Leu Pro Trp
1 5 10 15
Leu Gln Glu Gly Ser Ala Asp Ser Val Cys Pro Gln Gly Lys Tyr Ile
20 25 30
His Pro Gln Asn Asn Ser Ile Cys Cys Thr Lys Cys His Lys Gly Thr
35 40 45
Tyr Leu Tyr Asn Asp Cys Pro Gly Pro Gly Gln Asp Thr Asp Cys Arg
50 55 60
Glu Cys Glu Ser Gly Ser Phe Thr Ala Ser Glu Asn His Leu Arg His
65 70 75 80
Cys Leu Ser Cys Ser Lys Cys Arg Lys Glu Met Gly Gln Val Glu Ile
85 90 95
Ser Ser Cys Thr Val Asp Arg Asp Thr Val Cys Gly Cys Arg Lys Asn
100 105 110
Gln Tyr Arg His Tyr Trp Ser Glu Asn Leu Phe Gln Cys Phe Asn Cys
115 120 125
Thr Leu Cys Leu Asn Gly Thr Val His Leu Ser Cys Gln Glu Lys Gln
130 135 140
Asn Thr Val Cys Thr Cys His Ala Gly Phe Phe Leu Arg Glu Asn Glu
145 150 155 160
Cys Val Ser Cys Ser Asn Cys Lys Lys Ser Leu Glu Cys Thr Lys Leu
165 170 175
Ser Leu Pro Gln Ile Glu Asn Val Lys Gly Thr Glu Asp Ser Gly Thr
180 185 190
Thr Ala Gly Ala Ala Pro Gly Cys Pro Glu Cys Thr Leu Gln Glu Asn
195 200 205
Pro Phe Phe Ser Gln Pro Gly Ala Pro Ile Leu Gln Cys Met Gly Cys
210 215 220
Cys Phe Ser Arg Ala Tyr Pro Thr Pro Leu Arg Ser Lys Lys Thr Met
225 230 235 240
Leu Val Gln Lys Asn Val Thr Ser Glu Ser Thr Cys Cys Val Ala Lys
245 250 255
Ser Tyr Asn Arg Val Thr Val Met Gly Gly Phe Lys Val Glu Asn His
260 265 270
Thr Ala Cys His Cys Ser Thr Cys Tyr Tyr His Lys Ser
275 280 285
(2)SEQ ID NO:7的信息:
(i)序列特征:
(A)长度:1301碱基对
(B)类型:核酸
(C)链型:单链
(D)拓扑结构:线性
(ii)分子类型:cDNA
(ix)特点:
(A)名称/关键词:CDS
(B)位置:279..1287
(xi))序列描述:SEQ ID NO:7
CTCGAGATGG CTACAGGTAA GCGCCCCTAA AATCCCTTTG GGCACAATGT GTCCTGAGGG 60
GAGAGGCAGC GACCTGTAGA TGGGACGGGG GCACTAACCC TCAGGTTTGG GGCTTCTGAA 120
TGTGAGTATC GCCATGTAAG CCCAGTATTT GGCCAATGTC AGAAAGCTCC TGGTCCCTGG 180
AGGGATGGAG AGAGAAAAAC AAACACCTCC TGGAGCAGGG AGAGTGCTGC CCTCTTGCTC 240
TCCGGCTCCC TCTGTTGCCC TCTGGTTTCT CCCCAGGC TCC CGG ACG TCC CTG 293
Ser Arg Thr Ser Leu
290
CTC CTG GCT TTT GGC CTG CTC TGC CTG CCC TGG CTT CAA GAG GGC AGT 341
Leu Leu Ala Phe Gly Leu Leu Cys Leu Pro Trp Leu Gln Glu Gly Ser
295 300 305
GCC GAT AGT GTG TGT CCC CAA GGA AAA TAT ATC CAC CCT CAA AAT AAT 389
Ala Asp Ser Val Cys Pro Gln Gly Lys Tyr Ile His Pro Gln Asn Asn
310 315 320
TCG ATT TGC TGT ACC AAG TGC CAC AAA GGA ACC TAC TTG TAC AAT GAC 437
Ser Ile Cys Cys Thr Lys Cys His Lys Gly Thr Tyr Leu Tyr Asn Asp
325 330 335
TGT CCA GGC CCG GGG CAG GAT ACG GAC TGC AGG GAG TGT GAG AGC GGC 485
Cys Pro Gly Pro Gly Gln Asp Thr Asp Cys Arg Glu Cys Glu Ser Gly
340 345 350
TCC TTC ACC GCT TCA GAA AAC CAC CTC AGA CAC TGC CTC AGC TGC TCC 533
Ser Phe Thr Ala Ser Glu Asn His Leu Arg His Cys Leu Ser Cys Ser
355 360 365 370
AAA TGC CGA AAG GAA ATG GGT CAG GTG GAG ATC TCT TCT TGC ACA GTG 581
Lys Cys Arg Lys Glu Met Gly Gln Val Glu Ile Ser Ser Cys Thr Val
375 380 385
GAC CGG GAC ACC GTG TGT GGC TGC AGG AAG AAC CAG TAC CGG CAT TAT 629
Asp Arg Asp Thr Val Cys Gly Cys Arg Lys Asn Gln Tyr Arg His Tyr
390 395 400
TGG AGT GAA AAC CTT TTC CAG TGC TTC AAT TGC AGC CTC TGC CTC AAT 677
Trp Ser Glu Asn Leu Phe Gln Cys Phe Asn Cys Ser Leu Cys Leu Asn
405 410 415
GGG ACC GTG CAC CTC TCC TGC CAG GAG AAA CAG AAC ACC GTG TGC ACC 725
Gly Thr Val His Leu Ser Cys Gln Glu Lys Gln Asn Thr Va1 Cys Thr
420 425 430
TGC CAT GCA GGT TTC TTT CTA AGA GAA AAC GAG TGT GTC TCC TGT AGT 773
Cys His Ala Gly Phe Phe Leu Arg Glu Asn Glu Cys Val Ser Cys Ser
435 440 445 450
AAC TGT AAG AAA AGC CTG GAG TGC ACG AAG TTG TGC CTA CCC CAG ATT 821
Asn Cys Lys Lys Ser Leu Glu Cys Thr Lys Leu Cys Leu Pro Gln Ile
455 460 465
GAG AAT GTT AAG GGC ACT GAG GAC TCA GGC ACC ACA GCT GGT GCT GGT 869
Glu Asn Val Lys Gly Thr Glu Asp Ser Gly Thr Thr Ala Gly Ala Gly
470 475 480
CCA CGG TGC CGC CCC ATC AAT GCC ACC CTG GCT GTG GAG AAG GAG GGC 917
Pro Arg Cys Arg Pro Ile Asn Ala Thr Leu Ala Val Glu Lys Glu Gly
485 490 495
TGC CCC GTG TGC ATC ACC GTC AAC ACC ACC ATC TGT GCC GGC TAC TGC 965
Cys Pro Val Cys Ile Thr Val Asn Thr Thr Ile Cys Ala Gly Tyr Cys
500 505 510
CCC ACC ATG ACC CGC GTG CTG CAG GGG GTC CTG CCG GCC CTG CCT CAG 1013
Pro Thr Met Thr Arg Val Leu Gln Gly Val Leu Pro Ala Leu Pro Gln
515 520 525 530
GTG GTG TGC AAC TAC CGC GAT GTG CGC TTC GAG TCC ATC CGG CTC CCT 1061
Val Val Cys Asn Tyr Arg Asp Val Arg Phe Glu Ser Ile Arg Leu Pro
535 540 545
GGC TGC CCG CGC GGC GTG AAC CCC GTG GTC TCC TAC GCC GTG GCT CTC 1109
Gly Cys Pro Arg Gly Val Asn Pro Val Val Ser Tyr Ala Val Ala Leu
550 555 560
AGC TGT CAA TGT GCA CTC TGC CGC CGC AGC ACC ACT GAC TGC GGG GGT 1157
Ser Cys Gln Cys Ala Leu Cys Arg Arg Ser Thr Thr Asp Cys Gly Gly
565 570 575
CCC AAG GAC CAC CCC TTG ACC TGT GAT GAC CCC CGC TTC CAG GAC TCC 1205
Pro Lys Asp His Pro Leu Thr Cys Asp Asp Pro Arg Phe Gln Asp Ser
580 585 590
TCT TCC TCA AAG GCC CCT CCC CCC AGC CTT CCA AGC CCA TCC CGA CTC 1253
Ser Ser Ser Lys Ala Pro Pro Pro Ser Leu Pro Ser Pro Ser Arg Leu
595 600 605 610
CCG GGG CCC TCG GAC ACC CCG ATC CTC CCA CAA T AAGGATCCCT CGAG 1301
Pro Gly Pro Ser Asp Thr Pro Ile Leu Pro Gln
615 620
(2)SEQ ID NO:8的信息:
(i)序列特征:
(A)长度:336氨基酸
(B)类型:氨基酸
(D)拓扑结构:线性
(ii)分子类型:蛋白质
(xi)序列描述:SEQ ID NO:8
Ser Arg Thr Ser Leu Leu Leu Ala Phe Gly Leu Leu Cys Leu Pro Trp
1 5 10 15
Leu Gln Glu Gly Ser Ala Asp Ser Val Cys Pro Gln Gly Lys Tyr Ile
20 25 30
His Pro Gln Asn Asn Ser Ile Cys Cys Thr Lys Cys His Lys Gly Thr
35 40 45
Tyr Leu Tyr Asn Asp Cys Pro Gly Pro Gly Gln Asp Thr Asp Cys Arg
50 55 60
Glu Cys Glu Ser Gly Ser Phe Thr Ala Ser Glu Asn His Leu Arg His
65 70 75 80
Cys Leu Ser Cys Ser Lys Cys Arg Lys Glu Met Gly Gln Val Glu Ile
85 90 95
Ser Ser Cys Thr Val Asp Arg Asp Thr Val Cys Gly Cys Arg Lys Asn
100 105 110
Gln Tyr Arg His Tyr Trp Ser Glu Asn Leu Phe Gln Cys Phe Asn Cys
115 120 125
Ser Leu Cys Leu Asn Gly Thr Val His Leu Ser Cys Gln Glu Lys Gln
130 135 140
Asn Thr Val Cys Thr Cys His Ala Gly Phe Phe Leu Arg Glu Asn Glu
145 150 155 160
Cys Val Ser Cys Ser Asn Cys Lys Lys Ser Leu Glu Cys Thr Lys Leu
165 170 175
Cys Leu Pro Gln Ile Glu Asn Val Lys Gly Thr Glu Asp Ser Gly Thr
180 185 190
Thr Ala Gly Ala Gly Pro Arg Cys Arg Pro Ile Asn Ala Thr Leu Ala
195 200 205
Val Glu Lys Glu Gly Cys Pro Val Cys Ile Thr Val Asn Thr Thr Ile
210 215 220
Cys Ala Gly Tyr Cys Pro Thr Met Thr Arg Val Leu Gln Gly Val Leu
225 230 235 240
Pro Ala Leu Pro Gln Val Val Cys Asn Tyr Arg Asp Val Arg Phe Glu
245 250 255
Ser Ile Arg Leu Pro Gly Cys Pro Arg Gly Val Asn Pro Val Val Ser
260 265 270
Tyr Ala Val Ala Leu Ser Cys Gln Cys Ala Leu Cys Arg Arg Ser Thr
275 280 285
Thr Asp Cys Gly Gly Pro Lys Asp His Pro Leu Thr Cys Asp Asp Pro
290 295 300
Arg Phe Gln Asp Ser Ser Ser Ser Lys Ala Pro Pro Pro Ser Leu Pro
305 310 315 320
Ser Pro Ser Arg Leu Pro Gly Pro Ser Asp Thr Pro Ile Leu Pro Gln
325 330 335
(2)SEQ ID NO:9的信息:
(i)序列特征:
(A)长度:6氨基酸
(B)类型:氨基酸
(C)链型:单链
(D)拓扑结构:线性
(ii)分子类型:肽
(xi)序列描述:SEQ ID NO:9
Ala Gly Ala Ala Pro Gly
1 5
(2)SEQ ID NO:10的信息:
(i)序列特征:
(A)长度:4氨基酸
(B)类型:氨基酸
(C)链型:单链
(D)拓扑结构:线性
(ii)分子类型:肽
(xi)序列描述:SEQ ID NO:10
Ala Gly Ala Gly
1
(2)SEQ ID NO:11的信息:
(i)序列特征:
(A)长度:30碱基对
(B)类型:核酸
(C)链型:单链
(D)拓扑结构:线性
(ii)分子类型:cDNA
(xi)序列描述:SEQ ID NO:11
TTTTCTCGAG ATGGCTACAG GTAAGCGCCC
(2)SEQ ID NO:12的信息:
(i)序列特征:
(A)长度:39碱基对
(B)类型:核酸
(C)链型:单链
(D)拓扑结构:线性
(ii)分子类型:cDNA
(xi)序列描述:SEQ ID NO:12
ACCTGGGGCA GCACCGGCAC AGGAGACACA CTCGTTTTC
(2)SEQ ID NO:13的信息:
(i)序列特征:
(A)长度:42碱基对
(B)类型:核酸
(C)链型:单链
(D)拓扑结构:线性
(ii)分子类型:cDNA
(xi)序列描述:SEQ ID NO:13
TGTGCCGGTG CTGCCCCAGG TTGCCCAGAA TGCACGCTAC AG 42
(2)SEQ ID NO:14的信息:
(i)序列特征:
(A)长度:36碱基对
(B)类型:核酸
(C)链型:单链
(D)拓扑结构:线性
(ii)分子类型:cDNA
(xi)序列描述:SEQ ID NO:14
TTTTGGATCC TTAAGATTTG TGATAATAAC AAGTAC 36
(2)SEQ ID NO:15的信息:
(i)序列特征:
(A)长度:39碱基对
(B)类型:核酸
(C)链型:单链
(D)拓扑结构:线性
(ii)分子类型:cDNA
(xi)序列描述:SEQ ID NO:15
CCGTGGACCA GCACCAGCAC AGGAGACACA CTCGTTTTC 39
(2)SEQ ID NO:16的信息:
(i)序列特征:
(A)长度:36碱基对
(B)类型:核酸
(C)链型:单链
(D)拓扑结构:线性
(ii)分子类型:cDNA
(xi)序列描述:SEQ ID NO:16
TGTGCTGGTG CTGGTCCACG GTGCCGCCCC ATCAAT 36
(2)SEQ ID NO:17的信息:
(i)序列特征:
(A)长度:32碱基对
(B)类型:核酸
(C)链型:单链
(D)拓扑结构:线性
(ii)分子类型:cDNA
(xi)序列描述:SEQ ID NO:17
TTTTGGATCC TTATTGTGGG AGGATCGGGG TG 32
(2)SEQ ID NO:18的信息:
(i)序列特征:
(A)长度:27碱基对
(B)类型:核酸
(C)链型:单链
(D)拓扑结构:线性
(ii)分子类型:cDNA
(xi)序列描述:SEQ ID NO:18
TTTTAGATCT CTTCTTGCAC AGTGGAC 27
(2)SEQ ID NO:19的信息:
(i)序列特征:
(A)长度:21碱基对
(B)类型:核酸
(C)链型:单链
(D)拓扑结构:线性
(ii)分子类型:cDNA
(xi)序列描述:SEQ ID NO:19
TGTGGTGCCT GAGTCCTCAG T 21
(2)SEQ ID NO:20的信息:
(i)序列特征:
(A)长度:41碱基对
(B)类型:核酸
(C)链型:单链
(D)拓扑结构:线性
(ii)分子类型:cDNA
(xi)序列描述:SEQ ID NO:20
ACTGAGGACT CAGGCACCAC AGCCGGTGCT GCCCCAGGTT G 1
(2)SEQ ID NO:21的信息:
(i)序列特征:
(A)长度:29碱基对
(B)类型:核酸
(C)链型:单链
(D)拓扑结构:线性
(ii)分子类型:cDNA
(xi)序列描述:SEQ ID NO:21
TTTTTCTAGA GAAGCAGCAG CAGCCCATG 29
(2)SEQ ID NO:22的信息:
(i)序列特征:
(A)长度:75碱基对
(B)类型:核酸
(C)链型:单链
(D)拓扑结构:线性
(ii)分子类型:cDNA
(xi)序列描述:SEQ ID NO:22
TTTTCCACAG CCAGGGTGGC ATTGATGGGG CGGCACCGTG GACCAGCACC AGCTGTGGTG 60
CCTGAGTCCT CAGTG 75
Claims (18)
1、一种杂种蛋白质,它含有形成一种二聚体的两种共表达的氨基酸序列,每种氨基酸序列含有:
(a)一种选自TBP1,TBP2,IFNα/β受体或IFNγ受体的胞外区域,促性腺素受体,抗体轻链,抗体重链,抗体Fab区域,IL-6,IFN-β,TPO的氨基酸序列;以及
(b)一种选自hCG,FSH,LH,TSH或抑制素的亚基;
其中序列(a)和(b)是直接结合的或通过一种肽连接物结合的,并且其中每一个所说的两种共表达的序列中的序列(b)能够聚集形成一种二聚体复合体。
2、按照权利要求1所述的杂种蛋白质,其中序列(a)是与序列(b)的氨基末端相连接。
3、按照权利要求1所述的杂种蛋白质,其中序列(a)是与序列(b)的羧基末端相连接。
4、按照权利要求1所述的杂种蛋白质,其中所说的两种共表达的氨基酸序列的每一种包括TBP1的序列或者它的片段,该片段相应于TBP1的氨基酸残基20-161或20-190作为序列(a),以及各自的hCG的α和β亚基作为序列(b)。
5、按照权利要求1所述的杂种蛋白质,其中所说的两种共表达的氨基酸序列的每一种包括促性腺素受体的胞外区域,作为序列(a),以及各自的促性腺素的α和β亚基,作为序列(b)。
6、按照权利要求5所述的杂种蛋白质,其中所说的序列(a)是FSH受体胞外区域和序列(b)是FSH亚基。
7、按照权利要求5所述的杂种蛋白质,其中所说的序列(a)和(b)由一个肽连接物连接。
8、按照权利要求7所述的杂种蛋白质,其中所说的肽连接物具有一种酶切位点。
9、按照权利要求8所述的杂种蛋白质,其中所说的酶切位点是一种凝血酶切割位点。
10、按照权利要求8所述的杂种蛋白质,其中所说的酶切位点被一种发现于卵巢中的酶识别和切割。
11、按照权利要求7所述的杂种蛋白质,其中所说的肽连接物充当一种柔性绞链。
12、按照权利要求1所述的杂种蛋白质,其中在两个亚基(b)之间加入了一个或多个共价键。
13、一种编码权利要求1所述的杂种蛋白质的DNA分子。
14、含有权利要求13所述DNA分子的表达载体。
15、一种宿主细胞,它含有权利要求14所述的表达载体并能够表达所说的杂质蛋白质。
16、一种生产杂种蛋白质的方法,包括培养权利要求15所述的宿主细胞并回收由其表达的杂种蛋白质。
17、一种药物组合物,它含有权利要求1所述的杂种蛋白质和药学上可接受的载体和/或赋形剂。
18、权利要求1-12中任一项所述的杂种蛋白质在制备诱导滤泡成熟的药物中的用途。
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Cited By (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN102002495A (zh) * | 2009-08-31 | 2011-04-06 | 成都蓉生药业有限责任公司 | 用于表达异二聚体糖蛋白激素的表达框、表达载体及制备方法 |
Families Citing this family (17)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
HUP9900619A3 (en) * | 1996-02-20 | 2001-11-28 | Applied Research Systems | Hybrid proteins which form heterodimers |
US6635740B1 (en) * | 1997-03-27 | 2003-10-21 | Board Of Supervisors Of Louisiana State University And Agricultural And Mechanical College | Ligand/lytic peptide compositions and methods of use |
CA2301846A1 (en) | 1997-09-04 | 1999-03-11 | Gryphon Sciences | Modular protein libraries and methods of preparation |
EA006605B1 (ru) * | 2000-02-22 | 2006-02-24 | Апплайд Резеч Системз Арс Холдинг Н.В. | СПОСОБ ОЧИСТКИ РЕКОМБИНАНТНОГО чХГ |
US20030228600A1 (en) * | 2000-07-14 | 2003-12-11 | Eppendorf 5 Prime, Inc. | DNA isolation method and kit |
IL147414A0 (en) * | 2001-12-31 | 2002-08-14 | Yeda Res & Dev | Ifnar2 mutants, their production and use |
US20030157091A1 (en) * | 2002-02-14 | 2003-08-21 | Dyax Corporation | Multi-functional proteins |
DE10247755B4 (de) * | 2002-10-14 | 2006-01-19 | Pfizenmaier, Klaus, Prof. Dr. | Selektive, lokale Aktivierung von Mitgliedern der TNF-Rezeptorfamilie durch systemisch inaktive nicht-Antikörper-TNF-Liganden-Fusionsproteine |
BRPI0507174A (pt) * | 2004-01-28 | 2008-04-01 | Syntonix Pharmaceuticals Inc | proteìnas de fusão hormÈnio-fc (fsh-fc) heterodiméricas estimuladoras de folìculo para o tratamento da infertilidade |
WO2006067210A1 (en) | 2004-12-23 | 2006-06-29 | Laboratoires Serono S.A. | Bcma polypeptides and uses thereof |
US7691611B2 (en) | 2005-06-03 | 2010-04-06 | Ares Trading S.A. | Production of recombinant IL-18 binding protein |
EP2200634B1 (en) * | 2007-09-21 | 2015-02-11 | The Regents of The University of California | Targeted interferon demonstrates potent apoptotic and anti-tumor activities |
CN101960014B (zh) * | 2008-02-01 | 2013-10-16 | 克罗莫塞尔公司 | 细胞系以及制备和使用其的方法 |
CN107840894A (zh) * | 2011-03-25 | 2018-03-27 | 格兰马克药品股份有限公司 | 异二聚体免疫球蛋白 |
CA2901226C (en) | 2013-02-18 | 2020-11-17 | Vegenics Pty Limited | Vascular endothelial growth factor binding proteins |
JP7439372B2 (ja) * | 2018-06-21 | 2024-02-28 | シャタック ラボ,インコーポレイテッド | ヘテロ二量体タンパク質及びその使用 |
KR20220012256A (ko) * | 2019-05-24 | 2022-02-03 | 프로비바 테라퓨틱스 (홍콩) 리미티드 | Il-2 조성물 및 이의 사용 방법 |
Family Cites Families (13)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
EP0325224B1 (en) * | 1988-01-22 | 1996-07-31 | ZymoGenetics, Inc. | Methods of producing secreted receptor analogs |
US6225449B1 (en) * | 1991-10-04 | 2001-05-01 | Washington University | Hormone analogs with multiple CTP extensions |
US5705478A (en) * | 1989-02-21 | 1998-01-06 | Washington University | Covalently linked β subunits of the glycoprotein hormones as antagonists |
US5116964A (en) * | 1989-02-23 | 1992-05-26 | Genentech, Inc. | Hybrid immunoglobulins |
AU5355790A (en) * | 1989-04-19 | 1990-11-16 | Cetus Corporation | Multifunctional m-csf proteins and genes encoding therefor |
US5650150A (en) * | 1990-11-09 | 1997-07-22 | Gillies; Stephen D. | Recombinant antibody cytokine fusion proteins |
IL99120A0 (en) * | 1991-08-07 | 1992-07-15 | Yeda Res & Dev | Multimers of the soluble forms of tnf receptors,their preparation and pharmaceutical compositions containing them |
US5932448A (en) * | 1991-11-29 | 1999-08-03 | Protein Design Labs., Inc. | Bispecific antibody heterodimers |
NZ251820A (en) | 1992-03-30 | 1996-07-26 | Immunex Corp | Fusion proteins with tnf-r (tumour necrosis factor-receptor) peptide linked to another tnf-r or an il-1r (interleukin-1-receptor) peptide |
US5447851B1 (en) * | 1992-04-02 | 1999-07-06 | Univ Texas System Board Of | Dna encoding a chimeric polypeptide comprising the extracellular domain of tnf receptor fused to igg vectors and host cells |
IL111125A0 (en) * | 1994-05-11 | 1994-12-29 | Yeda Res & Dev | Soluble oligomeric tnf/ngf super family ligand receptors and their use |
HUP9900619A3 (en) * | 1996-02-20 | 2001-11-28 | Applied Research Systems | Hybrid proteins which form heterodimers |
US6194177B1 (en) * | 1996-02-20 | 2001-02-27 | Applied Research Systems Ars Holding N.V. | DNA encoding a hybrid heterodimeric protein |
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Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
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CN102002495A (zh) * | 2009-08-31 | 2011-04-06 | 成都蓉生药业有限责任公司 | 用于表达异二聚体糖蛋白激素的表达框、表达载体及制备方法 |
CN102002495B (zh) * | 2009-08-31 | 2012-07-18 | 成都蓉生药业有限责任公司 | 用于表达异二聚体糖蛋白激素的表达框、表达载体及制备方法 |
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