CN1289525C - T2r,一类味觉受体 - Google Patents
T2r,一类味觉受体 Download PDFInfo
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- CN1289525C CN1289525C CNB00815435XA CN00815435A CN1289525C CN 1289525 C CN1289525 C CN 1289525C CN B00815435X A CNB00815435X A CN B00815435XA CN 00815435 A CN00815435 A CN 00815435A CN 1289525 C CN1289525 C CN 1289525C
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Abstract
本发明提供了味觉转导G-蛋白偶联受体新家族的核酸和氨基酸序列、这些受体的抗体、检测这些核酸和受体的方法,和筛选味觉转导G-蛋白偶联受体的调节蛋白的方法。
Description
相关申请的交叉引用
本申请要求1999年9月10日提交的USSN 09/393,634的优先权,引入本文供参考。
关于联邦政府资助的研究和开发的声明
本发明得到政府资助进行,基金号5R01DC03160,由国立卫生研究院授予。故政府对本发明有一定权利。
发明领域
本发明提供了味觉细胞特异性G-蛋白偶联受体的分离的核酸和氨基酸序列,这些受体的抗体,检测这些核酸和受体的方法,和筛选味觉细胞特异性G-蛋白偶联受体的调节剂的方法。
发明背景
味觉转导是动物化学转导的最复杂形式之一(见例如,Margolskee,BioEssays15:645-650(1993);Avenet&Lindemann,J.Membrane Biol.112:1-8(1989))。整个动物界,从简单的多细胞动物到最复杂的脊椎动物都发现了味觉信号传递;其主要目的是提供对不挥发配体的可靠信号传递反应。这些感觉的每一种是由受体或通道介导的独特信号传导途径所介导的,导致受体细胞去极化,产生受体或动作电位,并在味觉传入神经元突触处释放神经递质(见例如,Roper,Ann.Rev.Neurosci.12:329-353(1989))。
据信哺乳动物具有5种基本的味觉:甜、苦、酸、咸和鲜(谷氨酸钠的味道)(见例如,Kawamura&Kare,鲜味的介绍:一种基本味觉(1987);Kinnamon&Cummings,Ann.Rev.Physiol.54:715-731(1992);Lindermann,Physiol.Rev.76:718-766(1996);Steward等,Am.J.Physiol.272:1-26(1997))。对人的广泛精神生理学研究已报道,舌头的不同区域显示不同的味偏好(见例如,Hoffmann,Menchen.Arch.Path.Anat.Physiol.62:516-530(1875);Bradley等,AnatomicalRecord 212:246-249(1985);Miley & Reedy,Physiol Behav.47:1213-1219(1990))。对动物的许多生理学研究也已显示了味觉受体细胞可选择性的对不同的味道作出反应(见例如Akabas等,Science 242:1047-1050(1988);Gilbertson等,J.Gen.Physiol.100:803-24(1992);Bernhardt等,J.Physiol.490:325-336(1996);Cummings等,J.Neurophysiol.75:1256-1263(1996))。
在哺乳动物中,味觉受体细胞集中于分布在舌上皮不同乳头中的味蕾中。舌最后部发现的周缘乳头含有成百(小鼠)上千(人)个味蕾,而且对苦味物质特别敏感。叶状乳头(位于舌后侧两边缘)含有几十到几百个味蕾,对酸和苦味物质特别敏感。含有一个或几个味蕾的蕈状乳头位于舌前部,据认为介导大部分甜味感觉。
每个味蕾视物种的不同,含有50-150个细胞,包括前体细胞、支持细胞和味觉受体细胞(见例如Lindermann,Physiol.Rev.76:718-766(1996))。受体细胞受其基底处传入神经末梢支配,这些神经末梢通过脑干和丘脑的突触将信息传递给大脑皮质的味觉中枢。阐明味觉细胞信号传导和信息处理的机制,对于味觉功能的理解、调节和“感受”是关键的。
虽然对于味觉细胞功能的精神生理学和生理学已有了不少了解,但对于介导这些感觉信号传递反应的分子和途径所知很少(Gilbertson综述,Current Opin.Neurobiol.3:532-539(1993))。电生理学研究提出,酸味和咸味由H+和Na+离子通过细胞尖端表面的特殊膜通道直接进入细胞来调节味觉细胞功能。就酸性化合物而言,假定H+封闭了K+通道(见例如,Kinnamon等,Proc.Natl.Acad.Sci.USA85:7023-7027(1988))或pH敏感性通道的活化(见例如,Gilbertson等,J.Gen.Physiol.100:803-24(1992))导致味觉细胞去极化;盐转导可能是由Na+通过阿米洛利敏感性Na+通道进入而部分介导的(见例如Heck等,Science,223:403-405(1984);Brand等,Brain Res.207-214(1985);Avenet等,Nature 331:351-354(1988))。
据信甜、苦和鲜味觉转导是由G-蛋白偶联受体(GPCR)信号传递途径介导的(见例如,Striem等,Biochem.J.260:121-126(1989);Chaudhari等,J.Neuros.16:3817-3826(1996);Wong等,Nature 381:796-800(1996))。令人不解的是,甜味觉和苦味觉转导的信号传递途径模型和GPCR级连反应的效应物酶(如G蛋白亚基、cGMP磷酸二酯酶、磷脂酶C、腺苷酸环化酶;见例如,Kinnamon & Margolskee,Curr.Opin.Neurobiol.6:506-513(1996))几乎一样多。然而,对于参与味觉传导的特异性膜受体,或许多受单种味觉转导途径活化的单种胞内信号传递分子知之甚少。鉴定这些分子对苦味拮抗剂、甜味促效剂和咸与酸味调节剂在许多制药工业和食品工业中的应用是重要的。
已被鉴定出的一种味觉细胞特异性G蛋白称为Gustducin(McLaugin等,Nature 357:563-569(1992))。该蛋白被认为与某些苦味和甜味的检测有关,因为gustducin敲除的小鼠显示对某些甜味和苦味物(tastant)敏感性降低(Wong等,Nature,381:796-800(1996)),而且由于gustducin在所有类型的味觉乳头中一组重要的细胞亚组中表达(McLaughin等,Nature 357:563-569(1992))。另外,gustducin可在体外通过用苦味化合物刺激味觉膜,相似的通过激活苦味受体来激活(Ming等,PNAS 95:8933-8938(1998))。
近来,鉴定出了两种新GPCR,并发现它们在味觉细胞中特异性表达。虽然这些受体蛋白(称为TR1和TR2)似乎直接与味觉感受有关(Hoon等,Cell 96:541-551(1999)),它们仅在哺乳动物味觉受体细胞的一部分中表达。例如,两种基因都不在表达Gustducin的细胞中广泛表达。因此,清楚的是仍需要发现其它与味觉有关的GPCR。
哺乳动物中的基因研究鉴定出了许多与味觉检测有关的基因座。例如,精神生理学尝味研究已显示人可以分成苦味物质PROP(6-n-丙基硫脲嘧啶)的品尝者、非品尝者和超品尝者,尝出PROP的味道可由显性等位基因赋予,而非品尝者具有两个隐性等位基因,品尝者具有至少一个显性等位基因(见Bartoshuk等,PhysiolBehav 56(6):1165-71;58:203-204(1994))。近来,对PROP品尝相关的一个基因座已被定位于5p15(Reed等,Am.J.Hum.Genet.64(5):1478-80(1999))。然而尚未见存在于5p15座位的PROP尝味基因的描述。
另外,已在小鼠中绘出了许多与味觉有关的基因。例如,已在小鼠第6号染色体的一个区域中绘出了与苦味检测有关的一簇基因(Lush等,GenetRes.66:167-174(1995))。
新味觉受体和味觉信号传递分子的鉴定和分离,将获得对味觉转导途径作药物学和遗传学调节的方法。例如,受体和通道分子的获得将允许我们筛选高亲和力的促效剂、拮抗剂、反向促效剂和味觉细胞活性的调节剂。可将这些味觉调节化合物用于制药工业和食品工业,来按需要改变味道。另外,这些味觉细胞特异性分子在产生味觉局部解剖图上起着无价工具的作用,以阐明舌的味觉细胞和连接大脑味觉中枢的味觉神经元之间的关系。
发明简述
本发明提供了编码味觉细胞特异性G-蛋白偶联受体的新核酸。这些核酸及其编码的多肽称为G-蛋白偶联味觉受体的“T2R”家族。这些受体还称为G-蛋白偶联味觉受体的“SF”家族。该GPCR新家族包括味觉转导途径的成分。特别是,该家族的成员与检测苦味有关。
一方面,本发明提供了一种调节味觉细胞中味觉信号传递的化合物的鉴定方法,该方法包括步骤如下:(i)使该化合物和味觉转导G-蛋白偶联受体多肽接触,该多肽含有与选自SEQ ID NO:166、SEQ ID NO:167、SEQ ID NO:168、SEQ ID NO:169、SEQ ID NO:170和或SEQ ID NO:171的序列至少约50%以上相同的氨基酸序列;和(ii)确定该化合物对多肽的功能作用。
另一方面,本发明提供了一种调节味觉细胞中味觉信号传递的化合物的鉴定方法,该方法包括步骤:(i)使该化合物和味觉转导G-蛋白偶联受体多肽接触,该多肽与以下序列具有约60%以上相同的氨基酸序列,序列选自:SEQ ID NO:1、SEQID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ IDNO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ IDNO:22、SEQ ID NO:24、SEQ ID NO:26、SEQ ID NO:28、SEQ ID NO:30、SEQ IDNO:32、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQ ID NO:39、SEQ IDNO:40、SEQ ID NO:42、SEQ ID NO:44、SEQ ID NO:46、SEQ ID NO:47、SEQ IDNO:48、SEQ ID NO:49、SEQ ID NO:50、SEQ ID NO:51、SEQ ID NO:53、SEQ IDNO:55、SEQ ID NO:56、SEQ ID NO:58、SEQ ID NO:59、SEQ ID NO:60、SEQ IDNO:62、SEQ ID NO:64、SEQ ID NO:65、SEQ ID NO:66、SEQ ID NO:67、SEQ IDNO:68、SEQ ID NO:69、SEQ ID NO:70、SEQ ID NO:71、SEQ ID NO:72、SEQ IDNO:73、SEQ ID NO:74、SEQ ID NO:75、SEQ ID NO:76、SEQ ID NO:77、SEQ IDNO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ ID NO:87、SEQ IDNO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ ID NO:97、SEQ IDNO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQ ID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQ ID NO:123、SEQ ID NO:125、SEQID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQ ID NO:151、SEQ ID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ ID NO:162、SEQID NO:164;和(ii)确定该化合物对多肽的功能作用。
另一方面,本发明提供了一种调节味觉细胞中味觉信号传递的化合物的鉴定方法,该方法包括步骤:(i)使含有味觉转导G-蛋白偶联受体的胞外功能域或跨膜区域、或其片段的多肽与该化合物接触,胞外功能域或跨膜区域与下列多肽的胞外功能域或跨膜区域具有大于60%的氨基酸序列相同性,所述多肽选自:SEQ ID NO:1、SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ ID NO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ ID NO:22、SEQ ID NO:24、SEQ ID NO:26、SEQ ID NO:28、SEQ ID NO:30、SEQ ID NO:32、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQ ID NO:39、SEQ ID NO:40、SEQ ID NO:42、SEQ ID NO:44、SEQ ID NO:46、SEQ ID NO:47、SEQ ID NO:48、SEQ ID NO:49、SEQ ID NO:50、SEQ ID NO:51、SEQ ID NO:53、SEQ ID NO:55、SEQ ID NO:56、SEQ ID NO:58、SEQ ID NO:59、SEQ ID NO:60、SEQ ID NO:62、SEQ ID NO:64、SEQ ID NO:65、SEQ ID NO:66、SEQ ID NO:67、SEQ ID NO:68、SEQ ID NO:69、SEQ ID NO:70、SEQ ID NO:71、SEQ ID NO:72、SEQ ID NO:73、SEQ ID NO:74、SEQ ID NO:75、SEQ ID NO:76、SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ ID NO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ ID NO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ IDNO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQ ID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQ ID NO:123、SEQ ID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQ ID NO:151、SEQID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ ID NO:162、SEQ ID NO:164;和(ii)确定该化合物对胞外功能域或跨膜区域的功能作用。
在一个实施例中,该多肽具有G-蛋白偶联受体活性。在另一个实施例中,该功能作用是化学作用。在另一个实施例中,通过测量化合物与多肽的胞外功能域的结合确定功能作用。在另一个实施例中,通过测量与多肽结合的放射性GTP确定功能作用。在另一个实施例中,该多肽含有与异源多肽共价连接,形成嵌合多肽的胞外功能域或跨膜区域、或胞外功能域和跨膜区域的组合。在另一个实施例中,该胞外功能域与固相共价或非共价连接。在另一个实施例中,该多肽来自大鼠、小鼠或人类。
在另一个实施例中,该多肽在细胞或细胞膜中表达。在另一个实施例中,该细胞是真核细胞。在另一个实施例中,通过测定表达该多肽的细胞的电活动确定化合物对多肽的功能作用。在另一个实施例中,通过测量表达该多肽的细胞中胞内cAMP、cGMP、IP3或Ca2+的变化确定化合物对多肽的功能作用。在另一个实施例中,检测细胞中的FURA-2依赖性荧光检测了细胞中胞内Ca2+的变化。在另一个实施例中,细胞是真核细胞。在另一个实施例中,细胞是HEK-293细胞。在另一个实施例中,多肽是含有视紫红质蛋白N-末端至少约20个连续氨基酸的融合蛋白。在另一个实施例中,该视紫红质蛋白是牛视紫红质蛋白。在另一个实施例中,细胞含有Gα15。在另一个实施例中,多肽在细胞中表达,多肽在苦味物的存在下与化合物接触,其中化合物存在下苦味物对细胞的功能作用差异和化合物不存在下苦味物对细胞的功能作用差异表明,该化合物能调节味觉细胞中的味觉传递。
在另一个实施例中,多肽具有选自下列序列的氨基酸序列,这些多肽是:SEQID NO:1、SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ IDNO:11、SEQ ID NO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ IDNO:21、SEQ ID NO:22、SEQ ID NO:24、SEQ ID NO:26、SEQ ID NO:28、SEQ IDNO:30、SEQ ID NO:32、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQ IDNO:39、SEQ ID NO:40、SEQ ID NO:42、SEQ ID NO:44、SEQ ID NO:46、SEQ IDNO:47、SEQ ID NO:48、SEQ ID NO:49、SEQ ID NO:50、SEQ ID NO:51、SEQ IDNO:53、SEQ ID NO:55、SEQ ID NO:56、SEQ ID NO:58、SEQ ID NO:59、SEQ IDNO:60、SEQ ID NO:62、SEQ ID NO:64、SEQ ID NO:65、SEQ ID NO:66、SEQ IDNO:67、SEQ ID NO:68、SEQ ID NO:69、SEQ ID NO:70、SEQ ID NO:71、SEQ IDNO:72、SEQ ID NO:73、SEQ ID NO:74、SEQ ID NO:75、SEQ ID NO:76、SEQ IDNO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ IDNO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ IDNO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQ ID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQ ID NO:123、SEQID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQID NO:143、SEQ ID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQ ID NO:151、SEQ ID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQID NO:162、SEQ ID NO:164。
在另一个方面,本发明提供了编码味觉转导G-蛋白受体的分离的核酸,该受体与选自SEQ ID NO:166、SEQ ID NO:167、SEQ ID NO:168、SEQ ID NO:169、SEQID NO:170和SEQ ID NO:171的序列具有大于约50%的氨基酸序列相同性。
在另一个方面,本发明提供了编码味觉转导G-蛋白偶联受体的分离的核酸,其中用引物扩增了核酸,这些引物是简并引物,该核酸与编码选自SEQ ID NO:166、SEQ ID NO:167、SEQ ID NO:168、SEQ ID NO:169、SEQ ID NO:170和SEQ ID NO:171的氨基酸序列的相同序列选择性杂交。
在另一个方面,本发明提供了编码味觉转导G-蛋白偶联受体的分离的核酸,受体含有与选自下列的序列具有大于约60%氨基酸序列相同性,这些序列是:SEQ IDNO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ IDNO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ IDNO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQ ID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQ ID NO:123、SEQID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQ ID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQ ID NO:151、SEQID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ ID NO:162、SEQ ID NO:164。
在另一个方面,本发明提供了一种编码味觉转导G-蛋白偶联受体的分离的核酸,其中核酸在高严格条件下与具有选自SEQ ID NO:78、SEQ ID NO:80、SEQ IDNO:82、SEQ ID NO:84、SEQ ID NO:86、SEQ ID NO:88、SEQ ID NO:90、SEQ IDNO:92、SEQ ID NO:94、SEQ ID NO:96、SEQ ID NO:98、SEQ ID NO:100、SEQID NO:102、SEQ ID NO:104、SEQ ID NO:106、SEQ ID NO:108、SEQ ID NO:110、SEQ ID NO:112、SEQ ID NO:114、SEQ ID NO:116、SEQ ID NO:118、SEQ ID NO:120、SEQ ID NO:122、SEQ ID NO:124、SEQ ID NO:126、SEQ ID NO:128、SEQID NO:130、SEQ ID NO:132、SEQ ID NO:134、SEQ ID NO:136、SEQ ID NO:138、SEQ ID NO:140、SEQ ID NO:142、SEQ ID NO:144、SEQ ID NO:146、SEQ ID NO:148、SEQ ID NO:150、SEQ ID NO:152、SEQ ID NO:154、SEQ ID NO:156、SEQID NO:157、SEQ ID NO:159、SEQ ID NO:161、SEQ ID NO:163、SEQ ID NO:165的核酸序列的核酸杂交,但不和具有选自SEQ ID NO:2、SEQ ID NO:4、SEQ ID NO:6、SEQ ID NO:8、SEQ ID NO:10、SEQ ID NO:12、SEQ ID NO:14、SEQ ID NO:16、SEQ ID NO:18、SEQ ID NO:20、SEQ ID NO:23、SEQ ID NO:25、SEQ ID NO:27、SEQ ID NO:29、SEQ ID NO:31、SEQ ID NO:34、SEQ ID NO:36、SEQ ID NO:38、SEQ ID NO:41、SEQ ID NO:43、SEQ ID NO:45、SEQ ID NO:51、SEQ ID NO:52、SEQ ID NO:54、SEQ ID NO:57、SEQ ID NO:61和SEQ ID NO:63的核苷酸序列的核酸杂交。
在另一个方面,本发明提供了一种编码味觉转导G-蛋白偶联受体的分离的核酸,该受体含有与具有下列氨基酸序列的多肽大于约60%的氨基酸相同性,这些序列是:SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQID NO:85、SEQ ID NO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQID NO:95、SEQ ID NO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQ ID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQID NO:123、SEQ ID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQ ID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQID NO:151、SEQ ID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ ID NO:162和SEQ ID NO:164,其中核酸在中度严格杂交条件下和核苷酸序列选择性杂交,这些核苷酸序列选自:SEQ ID NO:78、SEQ ID NO:80、SEQ ID NO:82、SEQ ID NO:84、SEQ ID NO:86、SEQ ID NO:88、SEQ ID NO:90、SEQ ID NO:92、SEQ ID NO:94、SEQ ID NO:96、SEQ ID NO:98、SEQ ID NO:100、SEQ ID NO:102、SEQ ID NO:104、SEQ ID NO:106、SEQ ID NO:108、SEQ ID NO:110、SEQID NO:112、SEQ ID NO:114、SEQ ID NO:116、SEQ ID NO:118、SEQ ID NO:120、SEQ ID NO:122、SEQ ID NO:124、SEQ ID NO:126、SEQ ID NO:128、SEQ ID NO:130、SEQ ID NO:132、SEQ ID NO:134、SEQ ID NO:136、SEQ ID NO:138、SEQID NO:140、SEQ ID NO:142、SEQ ID NO:144、SEQ ID NO:146、SEQ ID NO:148、SEQ ID NO:150、SEQ ID NO:152、SEQ ID NO:154、SEQ ID NO:156、SEQ ID NO:157、SEQ ID NO:159、SEQ ID NO:161、SEQ ID NO:163和SEQ ID NO:165,但不和具有下列核苷酸序列的核酸杂交:SEQ ID NO:2、SEQ ID NO:4、SEQ ID NO:6、SEQ ID NO:8、SEQ ID NO:10、SEQ ID NO:12、SEQ ID NO:14、SEQ ID NO:16、SEQ ID NO:18、SEQ ID NO:20、SEQ ID NO:23、SEQ ID NO:25、SEQ ID NO:27、SEQ ID NO:29、SEQ ID NO:31、SEQ ID NO:34、SEQ ID NO:36、SEQ ID NO:38、SEQ ID NO:41、SEQ ID NO:43、SEQ ID NO:45、SEQ ID NO:51、SEQ ID NO:52、SEQ ID NO:54、SEQ ID NO:57、SEQ ID NO:61和SEQ ID NO:63。
在另一方面,本发明提供了编码味觉转导G-蛋白偶联受体的胞外结构域或跨膜区或其组合的分离的核酸,该胞外结构域或跨膜区与含有下列氨基酸序列的多肽的胞外结构域和跨膜区具有60%以上的氨基酸序列相同性:SEQ ID NO:77、SEQID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ ID NO:87、SEQID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ ID NO:97、SEQID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQ ID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQ ID NO:123、SEQ ID NO:125、SEQID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQ ID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQ ID NO:151、SEQ ID NO:153、SEQID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ ID NO:162和SEQ ID NO:164。
在一个实施例中,核酸编码一种受体,该受体与针对具有下列氨基酸序列的多肽的多克隆抗体特异性结合:SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ ID NO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ ID NO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQ ID NO:117、SEQ ID NO:119、SEQID NO:121、SEQ ID NO:123、SEQ ID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQ ID NO:145、SEQ ID NO:147、SEQID NO:149、SEQ ID NO:151、SEQ ID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ ID NO:162和SEQ ID NO:164,但不和针对具有下列氨基酸序列的多肽产生的多克隆抗体结合:SEQ ID NO:1、SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ ID NO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ ID NO:22、SEQ ID NO:24、SEQ ID NO:26、SEQ ID NO:28、SEQ ID NO:30、SEQ ID NO:32、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQ ID NO:39、SEQ ID NO:40、SEQ ID NO:42、SEQ ID NO:44、SEQ ID NO:46、SEQ ID NO:47、SEQ ID NO:48、SEQ ID NO:49、SEQ ID NO:50、SEQ ID NO:51、SEQ ID NO:53、SEQ ID NO:55、SEQ ID NO:56、SEQ ID NO:58、SEQ ID NO:59、SEQ ID NO:60、SEQ ID NO:62、SEQ ID NO:64、SEQ ID NO:65、SEQ ID NO:66、SEQ ID NO:67、SEQ ID NO:68、SEQ ID NO:69、SEQ ID NO:70、SEQ ID NO:71、SEQ ID NO:72、SEQ ID NO:73、SEQ ID NO:74、SEQ ID NO:75和SEQ ID NO:76。
在另一个实施例中,核酸编码含有选自下列的一个氨基酸序列的受体:SEQ IDNO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ IDNO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ IDNO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQ ID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQ ID NO:123、SEQID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQ ID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQ ID NO:151、SEQID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ ID NO:162、SEQ ID NO:164、SEQ ID NO:166、SEQ ID NO:167、SEQ ID NO:168、SEQ ID NO:169、SEQ ID NO:170和SEQ ID NO:171。
在另一个实施例中,核酸是含有选自:SEQ ID NO:78、SEQ ID NO:80、SEQID NO:82、SEQ ID NO:84、SEQ ID NO:86、SEQ ID NO:88、SEQ ID NO:90、SEQID NO:92、SEQ ID NO:94、SEQ ID NO:96、SEQ ID NO:98、SEQ ID NO:100、SEQ ID NO:102、SEQ ID NO:104、SEQ ID NO:106、SEQ ID NO:108、SEQ ID NO:11O、SEQ ID NO:112、SEQ ID NO:114、SEQ ID NO:116、SEQ ID NO:118、SEQID NO:120、SEQ ID NO:122、SEQ ID NO:124、SEQ ID NO:126、SEQ ID NO:128、SEQ ID NO:130、SEQ ID NO:132、SEQ ID NO:134、SEQ ID NO:136、SEQ ID NO:138、SEQ ID NO:140、SEQ ID NO:142、SEQ ID NO:144、SEQ ID NO:146、SEQID NO:148、SEQ ID NO:150、SEQ ID NO:152、SEQ ID NO:154、SEQ ID NO:156、SEQ ID NO:157、SEQ ID NO:159、SEQ ID NO:161、SEQ ID NO:163和SEQ ID NO:165的序列。
在另一个实施例中,核酸编码具有G-蛋白偶联受体活性的受体。在另一个实施例中,核酸来自大鼠或小鼠。
在另一个实施例中,核酸编码与异源多肽连接的胞外结构域或跨膜区或其组合,形成嵌合多肽。在另一个实施例中,核酸编码多肽的胞外结构域,具有选自下列的氨基酸序列:SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ ID NO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ ID NO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQID NO:113、SEQ ID NO:115、SEQ ID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQ ID NO:123、SEQ ID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQID NO:141、SEQ ID NO:143、SEQ ID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQ ID NO:151、SEQ ID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ ID NO:162、SEQ ID NO:164、SEQ ID NO:166、SEQ ID NO:167、SEQID NO:168、SEQ ID NO:169、SEQ ID NO:170和SEQ ID NO:171。
在另一个方面,本发明提供了一种表达载体,含有上述任何核酸。在另一个方面,本发明提供了含有表达载体的分离的细胞。
在另一个方面,本发明提供了一种分离的味觉转导G-蛋白偶联受体,受体与选自SEQ ID NO:166、SEQ ID NO:167、SEQ ID NO:168、SEQ ID NO:169、SEQID NO:170和SEQ ID NO:171的序列具有大于约50%的氨基酸相同性。
在另一个方面,本发明提供了一种分离的味觉转导G-蛋白偶联受体,受体与选自SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID N0:85、SEQ ID NO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ ID NO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ IDNO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQ ID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQ ID NO:123、SEQ ID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQ ID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQ ID NO:151、SEQ ID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ
ID NO:162和SEQ ID NO:164的序列具有约60%以上的氨基酸序列相同性。
在一个实施例中,受体含有选自SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ ID NO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ ID NO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ ID NO:107、SEQ ID NO:109、SEQID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQ ID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQ ID NO:123、SEQ ID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQ ID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQ ID NO:151、SEQ ID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ ID NO:162、SEQ ID NO:164、SEQ ID NO:166、SEQID NO:167、SEQ ID NO:168、SEQ ID NO:169、SEQ ID NO:170和SEQ ID NO:171的氨基酸序列。
在另一个实施例中,受体与多克隆抗体特异性结合,这些多克隆抗体是针对具有选自SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQID NO:85、SEQ ID NO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQID NO:95、SEQ ID NO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQ ID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQID NO:123、SEQ ID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQ ID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQID NO:151、SEQ ID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ ID NO:162和SEQ ID NO:164的氨基酸序列的多肽产生的,但不和针对含有选自SEQ ID NO:1、SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ ID NO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ ID NO:22、SEQ ID NO:24、SEQ ID NO:26、SEQ ID NO:28、SEQ ID NO:30、SEQ ID NO:32、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQ ID NO:39、SEQ ID NO:40、SEQ ID NO:42、SEQ ID NO:44、SEQ ID NO:46、SEQ ID NO:47、SEQ ID NO:48、SEQ ID NO:49、SEQ ID NO:50、SEQ ID NO:51、SEQ ID NO:53、SEQ ID NO:55、SEQ ID NO:56、SEQ ID NO:58、SEQ ID NO:59、SEQ ID NO:60、SEQ ID NO:62、SEQ ID NO:64、SEQ ID NO:65、SEQ ID NO:66、SEQ ID NO:67、SEQ ID NO:68、SEQ ID NO:69、SEQ ID NO:70、SEQ ID NO:71、SEQ ID NO:72、SEQ ID NO:73、SEQ ID NO:74、SEQ ID NO:75和SEQ ID NO:76的氨基酸序列的多肽产生的多克隆抗体杂交。在另一个实施例中,受体具有G-蛋白偶联受体活性。在另一个实施例中,受体来自大鼠或小鼠。
在另一个实施例中,本发明提供了分离的多肽,它含有味觉转导G-蛋白偶联受体的胞外结构域或跨膜区或其组合,胞外结构域或跨膜区与具有下列氨基酸序列的多肽的胞外结构域或跨膜区具有约60%以上的氨基酸序列相同性:SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ ID NO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ ID NO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQ ID NO:123、SEQ ID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQ ID NO:151、SEQ ID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ ID NO:162和SEQ ID NO:164。
在一个实施例中,多肽编码多肽的胞外结构域或跨膜区,含有选自:SEQ IDNO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ IDNO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ IDNO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQ ID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQ ID NO:123、SEQID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQ ID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQ ID NO:151、SEQID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ ID NO:162和SEQ ID NO:164、SEQ IDNO:166、SEQ IDNO:167、SEQ ID NO:168、SEQ IDNO:169、SEQ ID NO:170和SEQ ID NO:171的氨基酸序列。在另一个实施例中,胞外结构域或跨膜区与异源多肽共价连接,形成嵌合多肽。
在一个方面,本发明提供了一种与受体选择性结合的抗体,该受体含有与选自:SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ ID NO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ ID NO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ IDNO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQ ID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQ ID NO:123、SEQ ID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQ ID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQ ID NO:151、SEQ ID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQID NO:162和SEQ ID NO:164的序列约60%以上的氨基酸序列相同性。
在另一个方面,本发明提供了含有编码味觉转导G-蛋白偶联受体的核酸的表达载体,其中在味觉细胞中表达受体,该受体含有与选自:SEQ ID NO:77、SEQ IDNO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ ID NO:87、SEQ IDNO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ ID NO:97、SEQ IDNO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQ ID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQ ID NO:123、SEQ ID NO:125、SEQID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQ ID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQ ID NO:151、SEQ ID NO:153、SEQID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ ID NO:162和SEQ ID NO:164的序列约60%以上的氨基酸序列相同性。
在另一个方面,本发明提供了用该表达载体转染的宿主细胞。
在另一个方面,本发明提供了含有编码人味觉转导G-蛋白偶联受体的多核苷酸序列与异源启动子可操纵性连接的表达盒,其中受体含有与选自SEQ ID NO:1、SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ ID NO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ ID NO:22、SEQ ID NO:24、SEQ ID NO:26、SEQ ID NO:28、SEQ ID NO:30、SEQ ID NO:32、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQ ID NO:39、SEQ ID NO:40、SEQ ID NO:42、SEQ ID NO:44、SEQ ID NO:46、SEQ ID NO:47、SEQ ID NO:48、SEQ ID NO:49、SEQ ID NO:50、SEQ ID NO:51、SEQ ID NO:53、SEQ ID NO:55、SEQ ID NO:56、SEQ ID NO:58、SEQ ID NO:59、SEQ ID NO:60、SEQ ID NO:62、SEQ ID NO:64、SEQ ID NO:65、SEQ ID NO:66、SEQ ID NO:67、SEQ ID NO:68、SEQ ID NO:69、SEQ ID NO:70、SEQ ID NO:71、SEQ ID NO:72、SEQ ID NO:73、SEQ ID NO:74、SEQ ID NO:75和SEQ ID NO:76的序列具有约60%以上氨基酸序列相同性的氨基酸序列。
在一个实施例中,受体含有选自SEQ ID NO:1、SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ ID NO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ ID NO:22、SEQ ID NO:24、SEQ ID NO:26、SEQ ID NO:28、SEQ ID NO:30、SEQ ID NO:32、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQ ID NO:39、SEQ ID NO:40、SEQ ID NO:42、SEQ ID NO:44、SEQ ID NO:46、SEQ ID NO:47、SEQ ID NO:48、SEQ ID NO:49、SEQ ID NO:50、SEQ ID NO:51、SEQ ID NO:53、SEQ ID NO:55、SEQ ID NO:56、SEQ ID NO:58、SEQ ID NO:59、SEQ ID NO:60、SEQ ID NO:62、SEQ ID NO:64、SEQ ID NO:65、SEQ ID NO:66、SEQ ID NO:67、SEQ ID NO:68、SEQ ID NO:69、SEQ ID NO:70、SEQ ID NO:71、SEQ ID NO:72、SEQ ID NO:73、SEQ ID NO:74、SEQ ID NO:75和SEQ ID NO:76的氨基酸序列。
在另一个方面,本发明提供了一种含有表达盒的分离的真核细胞。
附图简述
图1显示了Gα15将μ阿片样肽受体和mGluR1受体的激活与胞内钙的释放偶联起来。用Gαi偶联的μ阿片样肽受体或Gαq偶联的mGluR1受体瞬时转染HEK-293细胞。试验含Gα15的转染细胞在加入受体促效剂:(c)10μM DAMGO和(d)20μM反式(±)1-氨基-1,3-环戊烷二甲酸(ACPD)之前(a,b)和之后(c,d)[Ca2+]的增加。[Ca2+]i依赖于Gα15(e,f栏)。量度表示FURA-2发射比确定的[Ca2+]i(nM)。
图2显示牛视紫红质头39个氨基酸有效将T2R靶向到HEK-293细胞的质膜。用抗-视紫红质mAb B6-30检测了代表性rho-T2R融合蛋白转染的非通透化细胞的免疫荧光染色。
图3显示苦味化合物刺激了T2R受体。用rho-mT2R5(a,d,g)、rho-hT2R4(b,e,h)和rho-mT2R8(c,f,I)转染HEK-293细胞。用1.5μM环己酰亚胺(d,g)刺激表达mT2R5的细胞,用1.5mM苯甲酸盐刺激表达hT2R4和hT2R8的细胞(e,f,h,i)。Gα-15不存在时,观察不到[Ca2+]i的增加(g-i);相反在存在味道物(d-f)时观察到强Gα15依赖性反应;量度表示FURA-2发射比确定的[Ca2+]i(nM)。直线痕迹(j-1)显示栏(d-f)的代表性细胞[Ca2+]i改变的动力学;箭头表示加入味道。
图4显示mT2R5是环己酰亚胺的味觉受体。(a)用2μM环己酰亚胺(CYX)、3mM 6-正丙基硫脲(PROP)或5mM地那铵(DEN)的多次脉冲攻击表达Gα15和rho-mT2R5的HEK-293细胞;上方的点和水平条表示味道脉冲的时间和持续时间。环己酰亚胺触发了强受体激活。该实验还说明为了重复刺激或在持续施加刺激过程中的脱敏作用。(b)对环己烯亚胺的反应是高度特异性的,在加入缓冲液(CON)或高浓度的其它味道后未观察不到。使用的缩写和浓度是:环己酰亚胺,CYX(5μM);阿托品,ATR(5mM);番木鳖碱,BRU(5mM);咖啡酸,CAFF(2mM);地那铵,DEN(5mM);表儿茶精,(-)EPI(3mM);苯基硫脲,PTC(3mM);6-正丙基硫脲嘧啶,PROP(10mM);糖精,SAC(10mM);马钱子碱,STR(5mM);蔗糖八乙酸酯,SOA(3mM)。柱表示至少6个独立实验的平均±标准偏差。(c)品尝者(DBA/2-等位基因)和非品尝者(C57BL/6-等位基因)品系的mT2R5基因介导对环己酰亚胺脉冲的不同[Ca2+]i变化。水平条描述刺激的时间和持续时间。在刺激之间经过200秒,确保细胞不会由于持续施加环己酰亚胺去极化。(d)mT2R5的环己酰亚胺剂量应答。[Ca2+]i的改变表示成标准化成30μM环己酰亚胺时的反应的FURA-2(F340/F380)比。非品尝者等位基因显示相对于品尝者等位基因,对环己酰亚胺敏感度的显著下降(分别是约2.3μM的EC50对0.5μM)。
图5显示hT2R4和mT2R8对地那铵的反应。用hT2R4或mT2R8受体瞬时转染表达Gα15的HEK-293细胞,如图3所示监测[Ca2+]i。(a)[Ca2+]i中的增加可由地那铵的刺激诱导,但不能被各种其它苦味化合物诱导。显示了(b)hT2R4和(c)mT2R8对一组55种不同苦味和甜味味道中的9个的反应情况(见实验方法)。CON指对照缓冲液加入,NAR是2mM柚苷,LYS是5mM赖氨酸。其它缩写和浓度如图4所报道。加入配体前后的平均FURA-2荧光比(F340/380)是从100个大小相等的含有全部反应细胞的区域中获得的。值代表至少6个实验的平均值±标准误差。
图6显示环己酰亚胺品尝者和非品尝者品系表达不同的们T2R5等位基因。(a)mT2R5预测的跨膜形态;标出非品尝者品系的等位基因中氨基酸的取代(SEQ IDNO:172)。由于具有相同多形性的品系衍生自同一发现者(Beck等,Nat Genet24:23-55(2000)),可以预测该诸位仅存在两种等位基因。原位杂交显示在(b)环己酰亚胺品尝者品系(DBM2)和(c)非品尝者品系(C57BL/6)的周缘乳头的细胞亚组中mT2R5的表达;在口腔其它区域的味蕾中检测不到表达模式中的品系特异性差异。
图7显示了mT2R5激活gustducin对环己酰亚胺的反应。(a)在300μM环己酰亚胺存在下,但在没有配体(对照),或1mM阿托品、番木鳖碱、咖啡因、地那铵、苯基硫脲、6-正丙基硫脲嘧啶、奎宁、糖精、马钱子碱、蔗糖八乙酸酯下的含有mT2R5的昆虫幼虫细胞膜。(b)用单位点结合(Kd=14.8+0.9μM)配合mT2R5gustducin激活依赖性的环己酰亚胺浓度。
图8提供了一张表,包括许多人、大鼠和小鼠T2R家族成员的核酸和蛋白质序列(SEQ ID NO:1-165)。
发明详述
I.介绍
本发明提供了编码味觉细胞特异性G-蛋白偶联受体新家族的核酸。这些核酸及其编码的受体称为味觉细胞特异性G蛋白偶联受体的“T2R”家族成员。这些味觉细胞特异性GPCR是味觉转导途径,如苦味转导途径的成员,而且与物质,如苦味物质6-正丙基硫脲嘧啶(PROP)、蔗糖八乙酸酯(soa)、棉子糖十一乙酸酯(roa)、环己酰亚胺(cyx)、地那铵、甘氨酸铜(Glb)和奎宁(qui)的味道检测有关。
这些核酸提供了用于鉴定味觉细胞的有价值探针,因为这些核酸在味觉细胞中特异性表达。例如,可用T2R多肽和蛋白质的探针鉴定叶状细胞、周缘细胞、和蕈状乳头,以及存在于geschmackstreifen和会厌中的味觉细胞。特别是,T2R探针可用于鉴定苦味感测、表达gustducin的细胞。它们也可作为工具,产生味觉局部解剖图,阐明舌头的味觉细胞和与大脑味觉中枢连接的味觉神经元之间的关系。另外,可用这些核酸及其编码的蛋白作为探针来研究味觉诱导行为。
本发明还提供了筛选这些新颖味觉细胞GPCR的调节剂,如活化剂、抑制剂、刺激剂、增强剂、促效剂和拮抗剂的方法。这些味觉转导调节剂可用于味觉信号传递途径的药物学和遗传学调节。可用这些筛选方法鉴定味觉细胞活性的高亲和力促效剂和拮抗剂。然后可将这些调节性化合物用于食品和制药工业,来按要求改变味道,例如减少食物或药物的苦味。因此,本发明提供了味觉调节的试验方法,其中T2R家族的成员作为调节剂对味觉转导作用的直接或间接报告分子。可在一些实验,如测量配体结合、离子浓度、膜电势、电流、离子流、转录、信号转导、受体配体相互反应、体外、体内和活体外第二信使浓度的变化中使用GPCR。在一个实施例中,可通过T2R家族的成员与第二报道分子,如绿色荧光蛋白结合(见例如,Mistili & Spector,自然生物技术15:961-964(1997))作为间接报道物。在另一个实施例中,在细胞中重组表达T2R家族的成员,并测量Ca2+水平和其它细胞内信使,如cAMP、cGMP和IP3的变化检测通过GPCR作用的味觉转导的调节。
在一个优选例中,在真核细胞中用异源嵌合体序列表达作为嵌合受体的T2R多肽,该异源嵌合体序列促进其成熟和通过分泌途径靶向。在一个优选例中,异源序列是视紫红质序列,如视紫红质的N-末端片段。可在任何真核细胞,如HEK-293细胞中表达这些嵌合的T2R受体。优选细胞含有功能性G蛋白,如Gα15,它能将嵌合受体与胞内信号传递途径,或与信号传递蛋白如磷脂酶Cβ偶联。可用任何标准方法,如通过检测细胞中FURA-2依赖性荧光检测胞内钙水平的变化,来检测这些细胞中这些嵌合受体的活化。
检测味觉转导的调节剂的方法包括用T2R多肽、其部分如胞外结构域或跨膜区或其组合、或含有T2R家族成员的一个或多个结构域的嵌合蛋白的体外配体结合试验;卵母细胞或组织培养细胞的T2R基因表达,或T2R片段或融合蛋白,如视紫红质融合蛋白的表达;T2R基因的转录活化;T2R家族成员的磷酸化和去磷酸化;G-蛋白和GPCR的结合;配体结合试验;电压、膜电势和电导变化;离子流试验;胞内第二信使如cGMP、cAMP和三磷酸肌醇的变化;胞内钙水平的变化;和神经递质释放的体外配体结合试验。
最后,本发明提供了检测T2R核酸和蛋白质表达的方法,可实现对味觉转导调控的研究和对味觉受体细胞的特异性鉴定。T2R家族成员还提供了可用于亲子鉴定和法医调查的核酸探针。T2R基因作为核酸探针,可鉴定味觉受体细胞,如叶状、蕈状、周缘、geschmackstreifen和会厌味觉受体细胞,特别是可感受苦味、表达gustducin的细胞。还可用T2R受体产生单克隆和多克隆抗体,用于鉴定味觉受体细胞。可用反转录和mRNA扩增,分离总RNA或聚腺苷酸RNA,RNA印迹、斑点印迹、原位杂交、RNase保护、S1消化、探测DNA微芯片阵列、蛋白质印迹等技术鉴定味觉受体细胞。
T2R基因包括相关味觉细胞特异性G-蛋白偶联受体的一个大家族。在基因组内,这些基因单独存在或存在于几个基因簇之一内。位于人基因组区12p13的一个基因簇含有至少9个基因,而第二个位于7q31的基因簇含有至少4个基因。总的说已鉴定了50个以上不同的T2R家族成员,含有80-120个之多的不同T2R基因,编码40-80个之多的功能性人受体。
T2R基因中的一些已与之前作图的味觉特异性座位联系起来。例如,人T2R01被精确的定位于人基因间隔区5p15,其中之前作图确定具有尝出PROP物质的能力的座位。另外,在基因组区12p13发现的人基因簇对应于小鼠6号染色体的一个区域,该区域显示含有许多尝苦味的基因,包括蔗糖八乙酸酯、视紫红质乙酸酯、环己酰亚胺和奎宁(见例如Lush等,Genet.Res.6:167-174(1995))。这些联系表明T2R基因与各种物质的味觉检测,特别是苦味物质的味觉检测有关。另外,如实施例7(见下)所示,小鼠T2R5对环己酰亚胺特别具有感受性,mT2R5基因中的突变产生Cyx表型。类似的,人T2R4和小鼠T2R8特别对地那铵和PROP具有感受性。
功能上,T2R基因代表与味觉转导有关的7个跨膜G-蛋白偶联受体的家族,其与G-蛋白相互作用,介导味觉信号转导(见例如,Fong,Cell Signal 8:217(1996);Baldwin,Curr.Opin.Cell.Biol.6:180(1994))。特别是T2R以配体特异性方式与G蛋白Gustducin相互作用。
结构上,T2R家族成员的核苷酸序列(见例如SEQ ID NO:2、4、6、8、10、12、14、16、18、20、23、25、27、29、31、34、36、38、41、43、45、51、52、54、57、61、63、78、80、82、84、86、88、90、92、94、96、98、100、102、104、106、108、110、112、114、116、118、120、122、124、126、128、130、132、134、136、138、140、142、144、146、148、150、152、154、156、157、159、161、163和165,分别分离自大鼠、小鼠和人)编码相关的多肽(含有一个胞外结构域、7个跨膜结构域和一个胞质结构域)家族。其它动物的相关T2R家族基因在长度至少约50个氨基酸,可任选在长度为100、200、500或更多个氨基酸的区域中与SEQ IDNO:2、4、6、8、10、12、14、16、18、20、23、25、27、29、31、34、36、38、41、43、45、51、52、54、57、61、63、78、80、82、84、86、88、90、92、94、96、98、100、102、104、106、108、110、112、114、116、118、120、122、124、126、128、130、132、134、136、138、140、142、144、146、148、150、152、154、156、157、159、161、163或165具有至少约60%的核苷酸相同性,或编码与1、3、5、7、9、11、13、15、17、19、21、22、24、26、28、30、32、33、35、37、39、40、42、44、46、47、48、49、50、51、53、55、56、58、59、60、62、64、65、66、67、68、69、70、71、72、73、74、75、76、77、79、81、83、85、87、89、91、93、95、97、99、101、103、105、107、109、111、113、115、117、119、121、123、125、127、129、131、133、135、137、139、141、143、145、147、149、151、153、155、158、160、162或164在长度至少约25个氨基酸,可任选50-100个氨基酸的氨基酸区域内具有至少约60%的核苷酸序列相同性的多肽。T2R基因在味觉细胞中特异性表达。
还鉴定出几种T2R家族成员特有的共有氨基酸序列或结构域。例如。T2R家族通常具有与SEQ ID NO:166(对应于例如SEQ ID NO:1中的氨基酸位置16-35和T2R跨膜区1)、SEQ ID NO:167(对应于例如SEQ ID NO:1中的氨基酸位置45-58和T2R跨膜区2)、SEQ ID NO:168(对应于例如SEQ ID NO:1中的氨基酸位置89-101和T2R跨膜区3)、SEQ ID NO:169(对应于例如SEQ ID NO:1中的氨基酸位置102-119和T2R跨膜区3)、SEQ ID NO:170(对应于例如SEQ ID NO:1中的氨基酸位置196-209和T2R跨膜区5)、或SEQ ID NO:171(对应于例如SEQ ID NO:35中的氨基酸位置273-286和T2R跨膜区7)具有至少约50%、可任选55%、60%、65%、70%、75%、80%、85%、90%、95%或更高的相同性的序列。因此,可用这些保守结构域通过%相同性鉴定、特异性杂交或扩增,或用对一个结构域产生的抗体特异性结合鉴定T2R家族的成员。
几个T2R基因代表彼此的明显直向同源物。例如,人T2R01(SEQ ID NO:1、2)、大鼠T2R01(SEQ ID NO:77、78)和小鼠T2R19(SEQ ID NO:141、142)是明显的直向同源物。另外,大鼠T2R08(SEQ ID NO:91、92)和小鼠T2R02(SEQ ID NO:107、108)在氨基酸序列水平大约有74%的相同性,彼此对人T2R13(SEQ ID NO:24、25)具有至少约50%的相同性。大鼠T2R03(SEQ ID NO:81、82)和小鼠T2R18(SEQ IDNO:139、140)大约有92%的相同性,分别与人T2R16(SEQ ID NO:30、31)具有至少约50%的相同性。最后,人T2R04(SEQ ID NO:7、8)和小鼠T2R08(SEQ ID NO:119、120)彼此具有至少约67%的相同性。
本发明还提供了本文描述的T2R蛋白的多态变体。例如,SEQ ID NO:77中描述的大鼠T2R中:变体#1,其中氨基酸位置7的亮氨酸残基被异亮氨酸残基取代;变体#2,其中氨基酸位置20的甘氨酸残基被丙氨酸残基取代。
本发明还提供了SEQ ID NO:79中描述的T2R蛋白的多态体:变体#1,其中氨基酸位置2的苯丙氨酸残基被酪氨酸残基取代;变体#2,其中氨基酸位置62的异亮氨酸残基被缬氨酸残基取代。
本发明还提供了SEQ ID NO:81中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置179的天冬酰胺残基被谷氨酰胺残基取代;变体#2,其中氨基酸位置183的甲硫氨酸残基被半胱氨酸残基取代。
本发明还提供了SEQ ID NO:83中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置4的丙氨酸残基被甘氨酸残基取代;变体#2,其中氨基酸位置63的异亮氨酸残基被亮氨酸残基取代。
本发明还提供了SEQ ID NO:85中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置56的异亮氨酸残基被缬氨酸残基取代;变体#2,其中氨基酸位置57的半胱氨酸残基被甲硫氨酸残基取代。
本发明还提供了SEQ ID NO:87中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置4的缬氨酸残基被异亮氨酸残基取代;变体#2,其中氨基酸位置5的甘氨酸残基被丙氨酸残基取代。
本发明还提供了SEQ ID NO:89中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置79的甘氨酸残基被丙氨酸残基取代;变体#2,其中氨基酸位置127的赖氨酸残基被精氨酸残基取代。
本发明还提供了SEQ ID NO:91中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置28的缬氨酸残基被亮氨酸残基取代;变体#2,其中氨基酸位置80的赖氨酸残基被精氨酸残基取代。
本发明还提供了SEQ ID NO:93中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置75的赖氨酸残基被精氨酸残基取代;变体#2,其中氨基酸位置251的半胱氨酸残基被甲硫氨酸残基取代。
本发明还提供了SEQ ID NO:95中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置48的丝氨酸残基被苏氨酸残基取代;变体#2,其中氨基酸位置49的缬氨酸残基被异亮氨酸残基取代。
本发明还提供了SEQ ID NO:97中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置25的天冬氨酸残基被谷氨酸残基取代;变体#2,其中氨基酸位置100的亮氨酸残基被异亮氨酸残基取代。
本发明还提供了SEQ ID NO:99中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置4的苏氨酸残基被丝氨酸残基取代;变体#2,其中氨基酸位置74的缬氨酸残基被异亮氨酸残基取代。
本发明还提供了SEQ ID NO:101中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置9的谷氨酰胺残基被天冬酰胺残基取代;变体#2,其中氨基酸位置18的酪氨酸残基被色氨酸残基取代。
本发明还提供了SEQ ID NO:103中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置26的丝氨酸残基被苏氨酸残基取代;变体#2,其中氨基酸位置8的缬氨酸残基被异亮氨酸残基取代。
本发明还提供了SEQ ID NO:105中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置4的亮氨酸残基被异亮氨酸残基取代;变体#2,其中氨基酸位置46的赖氨酸残基被精氨酸残基取代。
本发明还提供了SEQ ID NO:107中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置3的丝氨酸残基被苏氨酸残基取代;变体#2,其中氨基酸位置28的缬氨酸残基被异亮氨酸残基取代。
本发明还提供了SEQ ID NO:109中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置26的亮氨酸残基被异亮氨酸残基取代;变体#2,其中氨基酸位置50的赖氨酸残基被精氨酸残基取代。
本发明还提供了SEQ ID NO:111中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置4的丙氨酸残基被甘氨酸残基取代;变体#2,其中氨基酸位置60的色氨酸残基被苯丙氨酸残基取代。
本发明还提供了SEQ ID NO:113中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置62的亮氨酸残基被异亮氨酸残基取代:变体#2,其中氨基酸位置244的甘氨酸残基被丙氨酸残基取代。
本发明还提供了SEQ ID NO:115中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置3的苏氨酸残基被丝氨酸残基取代;变体#2,其中氨基酸位置123的精氨酸残基被赖氨酸残基取代。
本发明还提供了SEQ ID NO:117中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置65的谷氨酰胺残基被天冬酰胺残基取代;变体#2,其中氨基酸位置68的异亮氨酸残基被亮氨酸残基取代。
本发明还提供了SEQ ID NO:119中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置2的亮氨酸残基被异亮氨酸残基取代;变体#2,其中氨基酸位置4的谷氨酸残基被天冬氨酸残基取代。
本发明还提供了SEQ ID NO:121中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置16的亮氨酸残基被异亮氨酸残基取代;变体#2,其中氨基酸位置46的赖氨酸残基被精氨酸残基取代。
本发明还提供了SEQ ID NO:123中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置9的丝氨酸残基被苏氨酸残基取代;变体#2,其中氨基酸位置14的苯丙氨酸残基被色氨酸残基取代。
本发明还提供了SEQ ID NO:125中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置24的亮氨酸残基被异亮氨酸残基取代;变体#2,其中氨基酸位置53的赖氨酸残基被精氨酸残基取代。
本发明还提供了SEQ ID NO:127中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置51的色氨酸残基被苯丙氨酸残基取代;变体#2,其中氨基酸位置101的赖氨酸残基被精氨酸残基取代。
本发明还提供了SEQ ID NO:129中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置4的缬氨酸残基被异亮氨酸残基取代;变体#2,其中氨基酸位置52的丙氨酸残基被甘氨酸残基取代。
本发明还提供了SEQ ID NO:131中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置150的赖氨酸残基被精氨酸残基取代;变体#2,其中氨基酸位置225的缬氨酸残基被亮氨酸残基取代。
本发明还提供了SEQ ID NO:133中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置27的异亮氨酸残基被亮氨酸残基取代;变体#2,其中氨基酸位置127的精氨酸残基被赖氨酸残基取代。
本发明还提供了SEQ ID NO:135中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置102的丝氨酸残基被苏氨酸残基取代;变体#2,其中氨基酸位置220的天冬氨酸残基被谷氨酸残基取代。
本发明还提供了SEQ ID NO:137中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置24的亮氨酸残基被异亮氨酸残基取代;变体#2,其中氨基酸位置45的赖氨酸残基被精氨酸残基取代。
本发明还提供了SEQ ID NO:139中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置50的异亮氨酸残基被亮氨酸残基取代;变体#2,其中氨基酸位置53的甘氨酸残基被丙氨酸残基取代。
本发明还提供了SEQ ID NO:141中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置76的苏氨酸残基被丝氨酸残基取代;变体#2,其中氨基酸位置131的亮氨酸残基被异亮氨酸残基取代。
本发明还提供了SEQ ID NO:143中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置98的甘氨酸残基被丙氨酸残基取代;变体#2,其中氨基酸位置153的色氨酸残基被苯丙氨酸残基取代。
本发明还提供了SEQ ID NO:145中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置8的异亮氨酸残基被亮氨酸残基取代;变体#2,其中氨基酸位置100的丙氨酸残基被甘氨酸残基取代。
本发明还提供了SEQ ID NO:147中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置52的丙氨酸残基被甘氨酸残基取代;变体#2,其中氨基酸位置75的亮氨酸残基被缬氨酸残基取代。
本发明还提供了SEQ ID NO:149中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置44的精氨酸残基被赖氨酸残基取代;变体#2,其中氨基酸位置49的缬氨酸残基被亮氨酸残基取代。
本发明还提供了SEQ ID NO:151中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置5的亮氨酸残基被异亮氨酸残基取代;变体#2,其中氨基酸位置25的甘氨酸残基被丙氨酸残基取代。
本发明还提供了SEQ ID NO:153中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置7的天冬氨酸残基被谷氨酸残基取代;变体#2,其中氨基酸位置60的亮氨酸残基被异亮氨酸残基取代。
本发明还提供了SEQ ID NO:155中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置7的缬氨酸残基被异亮氨酸残基取代;变体#2,其中氨基酸位置23的丙氨酸残基被甘氨酸残基取代。
本发明还提供了SEQ ID NO:158中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置5的亮氨酸残基被异亮氨酸残基取代;变体#2,其中氨基酸位置21的甘氨酸残基被丙氨酸残基取代。
本发明还提供了SEQ ID NO:160中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置5的缬氨酸残基被亮氨酸残基取代;变体#2,其中氨基酸位置23的甘氨酸残基被丙氨酸残基取代。
本发明还提供了SEQ ID NO:162中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置22的亮氨酸残基被异亮氨酸残基取代;变体#2,其中氨基酸位置34的甘氨酸残基被丙氨酸残基取代。
本发明还提供了SEQ ID NO:164中描述的T2R蛋白的多态变体:变体#1,其中氨基酸位置49的异亮氨酸残基被亮氨酸残基取代;变体#2,其中氨基酸位置76的赖氨酸残基被精氨酸残基取代。
可用T2R核苷酸和氨基酸序列的特定区域来鉴定T2R家族成员的多态变体、种间同系物和等位基因。可在体外,如严格杂交条件下或PCR(用编码SEQ ID NO:166-171的引物)和测序,或使用计算机系统中的序列信息与其它核苷酸序列比较,进行该鉴定。通常,通过比较约25个氨基酸或更多,如50-100个氨基酸的氨基酸序列来进行对T2R家族成员的多态变体和等位基因的鉴定。至少约60%或以上,可任选65%、70%、75%、80%、85%或90-95%或以上相同的氨基酸通常证明一个蛋白质是T2R家族成员的多态变体、种间同系物、或等位基因。可用任何一种下文讨论的序列比较算法进行序列比较。还可用特异性结合T2R多肽或其保守区域的抗体鉴定等位基因、种间同系物和多态变体。
通过检测推定T2R多肽的味觉细胞特异性表达证实了T2R基因的多态变体、种间同系物和等位基因。通常,用具有SEQ ID NO:1、SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ ID NO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ ID NO:22、SEQ ID NO:24、SEQ ID NO:26、SEQ ID NO:28、SEQ ID NO:30、SEQ ID NO:32、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQ ID NO:39、SEQ ID NO:40、SEQ ID NO:42、SEQ ID NO:44、SEQ ID NO:46、SEQ ID NO:47、SEQ ID NO:48、SEQ ID NO:49、SEQ ID NO:50、SEQ ID NO:51、SEQ ID NO:53、SEQ ID NO:55、SEQ ID NO:56、SEQ ID NO:58、SEQ ID NO:59、SEQ ID NO:60、SEQ ID NO:62、SEQ ID NO:64、SEQ ID NO:65、SEQ ID NO:66、SEQ ID NO:67、SEQ ID NO:68、SEQ ID NO:69、SEQ ID NO:70、SEQ ID NO:71、SEQ ID NO:72、SEQ ID NO:73、SEQ ID NO:74、SEQ ID NO:75、SEQ ID NO:76、SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ ID NO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ ID NO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQ ID NO:117、SEQ ID NO:119、SEQID NO:121、SEQ ID NO:123、SEQ ID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQ ID NO:145、SEQ ID NO:147、SEQID NO:149、SEQ ID NO:151、SEQ ID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ ID NO:162或SEQ ID NO:164的氨基酸序列的T2R多肽作为与推定T2R蛋白比较的阳性对照,来证明对T2R家族成员的多态变体或等位基因的鉴定。多态变体、等位基因和种间同系物将预期保留G-蛋白偶联受体的7次跨膜结构。
本发明还提供了T2R受体的核苷酸序列,可用于驱动多核苷酸的味觉细胞特异性表达,尤其在感受苦味物质,表达gustducin的味觉细胞中。
还可用T2R家族成员的核苷酸和氨基酸序列信息在计算机系统中构建味觉细胞特异性多肽的模型。然后用这些模型来鉴定能活化或抑制T2R受体蛋白的化合物。可用这些调节T2R家族成员活性的化合物研究T2R基因在味觉转导中的作用。
T2R家族成员的分离首次提供了检验G-蛋白偶联受体味觉转导的抑制剂和激活物的方法。生物学活性的T2R蛋白用于测试作为味觉转导物的T2R的抑制剂和激活物,使用体内和体外表达,测量如:T2R依赖性基因的转录活化;配体结合;磷酸化和去磷酸化;与G-蛋白的结合;G-蛋白活化;调控分子结合;电压、膜电势和电导改变;离子流;胞内第二信使,如cAMP和三磷酸肌醇;胞内钙水平;和神经递质释放。用T2R家族成员鉴定的这些激活剂和抑制剂可用于进一步研究味觉转导和鉴定特异性味觉促效剂和拮抗剂。这些激活剂和抑制剂可用于药物和食品制剂来按要求改变味觉,例如,减少食物或药品中的苦味。
本发明还提供了试验方法,优选高通量试验方法,来鉴定与T2R多肽相互作用和/或调节它的分子。在许多试验中,用T2R家族成员的一个特定结构域,如胞外、跨膜、或胞内结构域或区域。在许多实施例中,一个胞外结构域或跨膜结构域或其组合与固相基质结合,用于例如分离配体、促效剂、拮抗剂或任何其它可以与T2R多肽的胞外结构域或跨膜区域结合和/或调节其活性的分子。在一些实施例中,T2R多肽的结构域,如胞外、跨膜或胞内结构域与异源多肽融合,从而形成嵌合多肽,如具有G蛋白偶联受体活性的嵌合多肽。这些嵌合多肽可用于例如鉴定配体,促效剂、拮抗剂或其它T2R多肽的调节剂。另外,这些嵌合多肽可用于产生具新配体结合特异性、调节模式、信号转导途径等或其它这些性质的新味觉受体,或产生具有新的配体结合特异性、调控模式、信号转导途径等的组合的新味觉受体。
检测T2R核酸和T2R多肽表达的方法也可用来鉴定味觉细胞,并建立舌的局部解剖图和舌味觉受体细胞与大脑中味觉神经元之间的关系。特别是检测T2R的方法可用于鉴定对苦味物质敏感的味觉细胞。可用编码人T2R基因的基因染色体定位,来鉴定由T2R家族成员导致或相关的疾病、突变和性状。
II.定义
如本文所用,下列术语(除非另外指定)具有赋予它们的意义。
“味觉细胞”是神经上皮细胞,其组成若干组,形成舌头的味蕾,如叶状、蕈状和周缘细胞(见例如,Roper等,Ann.Rev.Neurosci.12:329-353(1989))。味觉细胞还包括上颚和其它可能含有味觉细胞的组织,如食道和胃的细胞。
“T2R”指G-蛋白偶联受体家族的一个或多个成员,其在味觉受体细胞,如叶状、蕈状和周缘细胞,以及上颚、食道和胃的细胞中表达(见例如Hoon等,Cell 96:541-551(1999),在此完全引入以供参考)。该家族还可称为“SF家族”(见例如,USSN 09/393,634)。可鉴定这些味觉细胞,因为其表达特异性分子,如Gustducin味转导素,一种味觉细胞特异性G蛋白(McLaughin等,Nature 357:563-569(1992))。还可根据形态学鉴定味觉受体细胞(见例如Roper,见上)。T2R家族成员能起味觉转导受体的作用,T2R家族成员对于味觉受体还称为“GR”家族,,或“SF”家族。
“T2R”核酸编码具有7个跨膜区的,具有“G-蛋白偶联受体活性”的GPCR家族,例如,它们在对胞外刺激的反应中,与G-蛋白结合,并通过磷脂酶C和腺苷酸环化酶等酶的刺激促使产生第二信使,如IP3、cAMP、cGMP和Ca2+(对于GPCR的结构和功能的描述,见例如Fong,见上,和Baldwin,见上)。图2提供了提供某些T2R家族成员之间的关系的分枝图。这些核酸编码在味觉细胞中表达的蛋白质,特别是对苦味物质有反应的表达Gustducin的味觉细胞。一个味觉细胞可含有许多不同的T2R多肽。
因此,术语“T2R”家族指多态变体、等位基因、突变物和种间同系物,它们:(1)在一个约25个氨基酸,可任选50-100个氨基酸的窗口上具有和SEQ ID NO:1、SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ ID NO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ ID NO:22、SEQ ID NO:24、SEQ ID NO:26、SEQ ID NO:28、SEQ ID NO:30、SEQ ID NO:32、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQ ID NO:39、SEQ ID NO:40、SEQ ID NO:42、SEQ ID NO:44、SEQ ID NO:46、SEQ ID NO:47、SEQ ID NO:48、SEQ ID NO:49、SEQ ID NO:50、SEQ ID NO:51、SEQ ID NO:53、SEQ ID NO:55、SEQ ID NO:56、SEQ ID NO:58、SEQ ID NO:59、SEQ ID NO:60、SEQ ID NO:62、SEQ ID NO:64、SEQ ID NO:65、SEQ ID NO:66、SEQ ID NO:67、SEQ ID NO:68、SEQ ID NO:69、SEQ ID NO:70、SEQ ID NO:71、SEQ ID NO:72、SEQ ID NO:73、SEQ ID NO:74、SEQ ID NO:75、SEQ ID NO:76、SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ ID NO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ ID NO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQ ID NO:123、SEQ ID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQ ID NO:151、SEQ ID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ ID NO:162或SEQ ID NO:164约60%,可任选的约75、80、85、90或95%相同的氨基酸序列;(2)能与针对含有选自SEQ ID NO:1、SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ ID NO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ ID NO:22、SEQ ID NO:24、SEQ ID NO:26、SEQ ID NO:28、SEQ ID NO:30、SEQ ID NO:32、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQ ID NO:39、SEQ ID NO:40、SEQ ID NO:42、SEQ ID NO:44、SEQ ID NO:46、SEQ ID NO:47、SEQ ID NO:48、SEQ ID NO:49、SEQ ID NO:50、SEQ ID NO:51、SEQ ID NO:53、SEQ ID NO:55、SEQ ID NO:56、SEQ ID NO:58、SEQ ID NO:59、SEQ ID NO:60、SEQ ID NO:62、SEQ ID NO:64、SEQ ID NO:65、SEQ ID NO:66、SEQ ID NO:67、SEQ ID NO:68、SEQ ID NO:69、SEQ ID NO:70、SEQ ID NO:71、SEQ ID NO:72、SEQ ID NO:73、SEQ ID NO:74、SEQ ID NO:75、SEQ ID NO:76、SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ ID NO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ ID NO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ IDNO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQ ID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQ ID NO:123、SEQ ID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQ ID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQ ID NO:151、SEQ ID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQID NO:162和SEQ ID NO:164的氨基酸序列及其保守修饰变化的免疫原的抗体结合;(3)在严格杂交条件下,与选自SEQ ID NO:2、SEQ ID NO:4、SEQ ID NO:6、SEQ ID NO:8、SEQ ID NO:10、SEQ ID NO:12、SEQ ID NO:14、SEQ ID NO:16、SEQ ID NO:18、SEQ ID NO:20、SEQ ID NO:23、SEQ ID NO:25、SEQ ID NO:27、SEQ ID NO:29、SEQ ID NO:31、SEQ ID NO:34、SEQ ID NO:36、SEQ ID NO:38、SEQ ID NO:41、SEQ ID NO:43、SEQ ID NO:45、SEQ ID NO:51、SEQ ID NO:52、SEQ ID NO:54、SEQ ID NO:57、SEQ ID NO:61、SEQ ID NO:63、SEQ ID NO:78、SEQ ID NO:80、SEQ ID NO:82、SEQ ID NO:84、SEQ ID NO:86、SEQ ID NO:88、SEQ ID NO:90、SEQ ID NO:92、SEQ ID NO:94、SEQ ID NO:96、SEQ ID NO:98、SEQ ID NO:100、SEQ ID NO:102、SEQ ID NO:104、SEQ ID NO:106、SEQ ID NO:108、SEQ ID NO:110、SEQ ID NO:112、SEQ ID NO:114、SEQ ID NO:116、SEQID NO:118、SEQ ID NO:120、SEQ ID NO:122、SEQ ID NO:124、SEQ ID NO:126、SEQ ID NO:128、SEQ ID NO:130、SEQ ID NO:132、SEQ ID NO:134、SEQ ID NO:136、SEQ ID NO:138、SEQ ID NO:140、SEQ ID NO:142、SEQ ID NO:144、SEQID NO:146、SEQ ID NO:148、SEQ ID NO:150、SEQ ID NO:152、SEQ ID NO:154、SEQ ID NO:156、SEQ ID NO:157、SEQ ID NO:159、SEQ ID NO:161、SEQ ID NO:163和SEQ ID NO:165及其保守修饰变化的序列特异性杂交(大小至少约100,500,可任选至少约500-1000个核苷酸);(4)含有至少与选自SEQ ID NO:166、SEQ IDNO:167、SEQ ID NO:168、SEQ ID NO:169、SEQ ID NO:170和SEQ ID NO:171的氨基酸序列具有至少约50%相同性的序列;或(5)用在严格条件下与编码SEQ IDNO:166、SEQ ID NO:167、SEQ ID NO:168、SEQ ID NO:169、SEQ ID NO:170和SEQ ID NO:171的简并引物相同的序列特异性杂交的引物扩增出来。
拓扑形态上,感受GPCR具有“N-末端结构域”、“胞外结构域”、含有7个跨膜区和对应胞质和胞外环的“跨膜结构域”、“胞质结构域”和“C-末端结构域”(见例如Hoon等,Cell 96:541-551(1999);Buck & Axel,Cell 65:175-187(1991))。可用本领域技术人员已知的方法从结构上鉴定这些功能域,如鉴定疏水和亲水结构域的序列分析程序(见例如Stryer,Biochemistry(第三版,1998);另见许多基于互联网的序列分析程序,如dot.imgen.bcm.tmc.edu)。这些功能域对于制备本发明的嵌合蛋白和体外试验,如配体结合试验是有用的。
因此,“胞外功能域”指T2R多肽的突出于细胞膜并暴露于细胞胞外面的结构域。该结构域将包括暴露于细胞胞外表面的“N-末端结构域”和暴露于细胞胞外表面的跨膜结构域的胞外环,即跨膜区域2和3,和跨膜区域4和5之间的环。“N末端结构域”区域从N-末端开始,延伸到紧靠跨膜结构域起始位置的地方。这些胞外结构域用于液相和固相的体外配体结合试验。另外,下文所述的跨膜区域还与胞外结构域组合或单独与配体结合,因此也用于体外结合试验。
包含7个跨膜“区域”的“跨膜功能域”指T2R多肽的结构域位于质膜内,还包括对应的胞质(胞内)和胞外环,也称为跨膜结构域“区域”。可用标准方法,如(见例如Kyte & Doolittle,分子生物学杂志157:105-132(1982))所述的方法鉴定7个跨膜区域和胞外和胞质环。
“胞质结构域”指T2R蛋白面向细胞内侧的结构域,如“C末端结构域”和跨膜结构域的胞内环,如跨膜区域1和2之间的胞内环和跨膜区域3和4之间的胞内环。“C-末端结构域”指跨越蛋白质最后一个跨膜结构域的终点和C-末端的区域,通常位于胞质内。
本文所用的“生物学样品”是含有编码一种或多种的T2R蛋白的一种或多种T2R核酸的生物组织或液体样品。这些样品包括但不限于:分离自人、小鼠和大鼠,具体是舌、上颚或其它可能含有味觉细胞的组织,如食道和胃。生物学样品还包括供组织学研究目的的组织切片,如冰冻切片。通常生物学样品获自真核生物,如昆虫、原生动物、鸟类、鱼类、爬行类和优选哺乳动物,如大鼠、小鼠、牛、狗、豚鼠、或家兔,和最优选的是灵长类,如黑猩猩或人。
“GPCR活性”指GPCR转导信号的活性。可在异源细胞中,通过将GPCR(或嵌合性GPCR)与G-蛋白或混杂G-蛋白(如Gα15)和PLC等酶偶联,并通过(Offermans& SimonJ.Biol.Chem.270:15175-15180(1995))测量胞内钙的升高,来测量这种活性。可用荧光Ca2+-指示剂染料和荧光显像记录配体诱导的[Ca2+]i变化,有效测量受体活性。可任选的,本发明的多肽与感觉转导,可任选的,味觉细胞中的味觉转导有关。
在测试调节T2R家族成员介导的味觉转导的化合物试验中,词组“功能作用”包括测定任何在该受体直接或间接影响下的参数,如功能、物理和化学作用。它包括配体结合、离子流变化、膜电势、电流、转录、G-蛋白结合、GPCR磷酸化或去磷酸化,信号转导、受体-配体相互作用,第二信使浓度(如cAMP、cGMP、IP3、或胞内Ca2+)体外、体内、和活体外的变化,还包括增加或减少神经递质或激素释放等的其它生理学作用。
通过“测定功能作用”指测试能提高或降低间接或直接受T2R家族成员影响的,如功能性、物理和化学作用的参数的化合物。这些功能性作用可通过本领域技术人员已知的任一方法测量,如光谱特征(如荧光、吸光度、折射指标)、流体动力学(如形状)、色谱分析、或可溶性能的变化,膜片钳、电压敏感性染料、全细胞电流、放射性同位素射流、可诱导标记、卵母细胞T2R基因表达;配体结合试验;电压、膜电势和电导的变化;离子流试验;胞内第二信使,如cAMP、cGMP和三磷酸肌醇(IP3)的变化;胞内钙水平的变化;神经递质释放等。
可互换的使用T2R基因或蛋白质的“抑制剂”、“激活物”和“调节剂”,指抑制、活化或调节味觉转导用体外和体内试验鉴定的分子,如配体、促效剂、拮抗剂及其同类物和模仿物。抑制剂是能与刺激物结合,而部分或完全封闭刺激,降低,防止,延缓活化,灭活,脱敏,或下调味觉转导的化合物,如拮抗剂。激活物是例如:结合、刺激、提高、打开、活化、促进、增强活化、致敏或上调味觉转导的化合物,如促效剂。调节剂包括例如:改变受体与胞外蛋白质(其与激活物或抑制剂结合)(如舌腺蛋白和其它疏水载体家族的成员)相互反应;G-蛋白;激酶(如视紫红质激酶和β-肾上腺素受体激酶,其与受体灭活和脱敏有关);和视紫红质样抑制蛋白(它也灭活和脱敏受体)的相互作用。调节剂包括T2R的遗传修饰形式,如改变的活性,以及天然存在和合成的配体,拮抗剂,促效剂,小化学分子等。抑制剂和激活剂的这些试验包括例如:在细胞或细胞膜中表达T2R,应用推定的调节剂化合物,然后如上所述测定对味觉转导的功能性作用。将包含用可能的激活剂、抑制剂或调节剂处理的T2R的试验样品,与不含抑制剂、激活剂或调节剂的对照样品比较,来检测抑制程度。将对照样品(未用抑制剂处理)指定为相对T2R活性值100%。当相对对照的T2R活性值是大约80%时,可任选50%或25-0%,即实现了T2R的抑制。当相对于对照的T2R活性值是约110%,可任选150%或200-500%,或1000-3000%以上时,即实现了T2R的活化。
“生物学活性”T2R,指具有如上所述GPCR活性的T2R,与味觉受体细胞的味觉转导,特别是苦味味觉转导有关。
术语“分离的”、“纯化的”或“生物学纯的”指某物质基本或根本没有其天然状态下通常伴随它的成分。通常用分析化学技术,如聚丙烯酰胺凝胶电泳或高效液相层析测定纯度或均一性。在制备物中是主要物质的某蛋白质是基本纯化的。具体说,分离的T2R核酸是与T2R基因旁侧编码T2R以外的蛋白质的开放阅读框相分开。术语“纯化的”指核酸或蛋白在电泳凝胶上基本只产生一条条带。特别是,它指核酸或蛋白质是至少85%纯的,可任选至少95%纯的,和可任选至少99%纯的。
“核酸”指脱氧核糖核苷酸或核糖核苷及其单链或双链形式的聚合物。术语包括含有已知核苷酸同类物或修饰的骨架残基或连接键,或合成的、天然存在的、和非天然存在的核酸,它们具有参比核酸相似的结合性能。它们以参比核苷酸相似的方式代谢。这些同类物的例子包括但不限于:硫代磷酸酯、磷酰胺、膦酸甲酯、手性膦酸甲酯、2-O-甲基核糖核苷酸、肽-核酸(PNA)。
除非另有注明,特定核酸序列还包含其保守性修饰的变体(如简并密码子取代物)和互补序列,以及明确标出的序列。特别是,可通过产生在一个或多个选择的(或全部)密码子的第三个位置以混合的碱基和/或脱氧次黄嘌呤残基取代的序列,而实现简并密码子取代(Batzer等,Nucleic Acid Res.19:5081(1991);Ohtsuka等,J.Biol.Chem.260:2605-2608(1995);Rossolini等,Mol.Cell.Probes 8:91-98(1994))。术语核酸与基因,cDNA,mRNA,寡核苷酸和多核苷酸可互换使用。
本文可互换使用术语“多肽”、“肽”和“蛋白质”,指氨基酸残基的聚合物。该术语用于以下氨基酸聚合物,即其中一个或多个氨基酸残基是对应的天然存在的氨基酸的人工化学模拟物,以及天然存在的氨基酸聚合物和非天然存在的氨基酸聚合物。
术语“氨基酸”指天然存在和合成的氨基酸,以及氨基酸同类物和氨基酸模拟物,它们的功能在某方面和天然存在的氨基酸相似。天然存在的氨基酸是那些由遗传密码编码的,以及后来被修饰的氨基酸,如羟基脯氨酸、γ-羰基谷氨酸、和O-磷酸丝氨酸。氨基酸同类物指和天然存在的氨基酸具有相同基础化学结构的化合物,即,一个与氢结合的α碳,一个羧基、一个氨基和一个R基团,例如同丝氨酸、正亮氨酸、甲硫氨酸亚砜、甲硫氨酸甲基锍。这些同类物具有修饰的R基团(如正亮氨酸)或修饰的肽骨架,但保留天然存在的氨基酸的相同基础化学结构。氨基酸模拟物指具有和氨基酸的一般化学结构不同的结构,但功能与天然存在的氨基酸相似的化合物。
本文的氨基酸可用它们的公知三字母符号或用IUPAC-IUB生物化学命名委员会推荐的单字母符号相称。同样,核苷酸可用它们普遍接受的单字母密码相称。
“保守性修饰变体”同时用于氨基酸和核酸序列。对于特定核酸序列,保守性修饰变体指那些编码相同或基本相同的氨基酸序列的核酸,或其中核酸不编码基本相同序列的一个氨基酸序列。由于遗传密码的简并性,许多功能相同的核酸编码了一给定的蛋白质。例如,密码子GCA,GCC,GCG和GCU都编码氨基酸丙氨酸。因此,在丙氨酸密码子的每一个位置,可将该密码子变成上述相应密码子的任何一个而不改变编码的多肽。这种核酸变化是“沉默变化”,它是一种保守性修饰变化。对本文编码多肽的每一核酸序列还描述了该核酸的每个可能的沉默变体。技术人员将认识到,核酸中的每个密码子(除了AUG,它通常是甲硫氨酸的唯一密码子,和TGG,它是色氨酸的唯一密码子)可被修饰,得到功能相同的分子。因此,编码多肽的核酸中的每个沉默变体隐含在各所述序列中。
对于氨基酸序列,技术人员将认识到对核酸、肽、多肽或蛋白质序列的单个取代、缺失或添加(在编码的序列中改变、添加或除去一个氨基酸或一小百分比的氨基酸)是“保守性修饰变体”,其中改变的结果是用化学上相似的氨基酸取代氨基酸。提供功能相似的氨基酸的保守取代表是本领域熟知的。这类保守性修饰变化是补充,而不排除本发明的多态变体、种间同系物、和等位基因。
下列8组各含有彼此可保守性取代的氨基酸:
1)丙氨酸(A),甘氨酸(G);
2)天门冬氨酸(D),谷氨酸(E);
3)天门冬酰胺(N),谷氨酰胺(Q);
4)精氨酸(R),赖氨酸(K);
5)异亮氨酸(I),亮氨酸(L),甲硫氨酸(M),缬氨酸(V);
6)苯丙氨酸(P),酪氨酸(Y),色氨酸(W);
7)丝氨酸(S),苏氨酸(T);和
8)半胱氨酸(C),甲硫氨酸(M)
(见例如,Creighton,Proteins(1984))。
多肽结构等大分子结构可用多种结构水平的术语描述。关于结构的一般描述,见例如,Alberts等,Molecular Biology of the Cell(第3版,1994)和Cantor和Schimmel,Biophysical Chemistry第I部分:生物大分子的构象(1980)。“一级结构”指某特定肽的氨基酸序列。“二级结构”指某肽中局部排序的三维结构。这些结构一般称为功能域。功能域是多肽的部分,形成该多肽的紧密单元,通常长50-350个氨基酸。典型的功能域是由较短的结构,如β-折叠和α-螺旋伸展片段构成。“三级结构”指多肽单体的完整三维结构。“四级结构”指独立的三级单元非共价结合形成的三维结构。各向异性的术语也被称为能量术语。
“标记”或“可检测分子”指可被分光光度、光化学、生物化学、免疫化学或化学方法检测的组合物、例如,有用的标记包括32P、荧光染料、电子密度试剂、酶(如ELISA中常用的)、生物素、毛地黄毒苷、或半抗原,而且可通过在该肽中掺入放射性标记,并检测和该肽特异性反应的抗体,来检测蛋白质。
“标记的核酸探针或寡核苷酸”是共价通过接头或化学键、或非共价通过离子,范德华力、静电或氢键与标记结合的核酸探针或寡核苷酸,从而可通过检测与探针结合的标记的存在,来检测该探针的存在。
本文所用的“核酸探针或寡核苷酸”指能与互补序列的靶核酸通过一种或多种化学键,通常通过互补碱基形成氢键配对而结合的核酸。本文所用的探针可包括天然(即A、G、C或T)或修饰碱基(7-脱氮鸟嘌呤、次黄嘌呤核苷等)。另外,探针内的碱基可通过除磷酸二酯键外的键连接,只要其不受杂交干扰。因此,例如探针可以是肽核酸,其中组成碱基通过肽键连接,而不是通过磷酸二酯键连接。本领域技术人员可理解,取决于杂交条件的严格度,探针可结合与序列不完全互补的靶序列。探针可任选的直接用放射性同位素、发色团、发光团、色原标记,或间接用生物素标记,然后与链霉亲和素复合物结合。通过检验探针的存在与否,可检测所选择序列或亚序列的存在与否。
当对细胞或核酸、蛋白或载体使用术语“重组”时,指该细胞、核酸、蛋白质或载体已通过引入异源核酸或蛋白,或改变天然核酸或蛋白质而被修饰,或该细胞衍生自如此修饰的细胞。因此例如,重组细胞表达了在细胞的天然(非重组)形式中未发现的基因,或异常表达、表达低下或根本不表达天然基因。
术语“异源”和核酸部分相关使用时,指该核酸含有的两个或多个亚序列,发现它们在自然情况下相互之间无相同关系。例如,通常重组产生的核酸具有两个或多个来自无关基因的序列,排列形成新的功能性核酸,如来自一种来源的启动子和来自另一种来源的编码区。类似的,异源蛋白质表明蛋白质含有两种或多种在天然情况下彼此无相同关系的亚序列(如融合蛋白)。
“启动子”定义为核酸控制序列的阵列,它直接转录一种核酸。本文所用的启动子包括转录起始位点附近的必需核酸序列,如就聚合酶II型启动子而言,是一TATA元件。启动子还可任选的包括远端增强子或抑制子元件,它们可位于距离转录起始位点多达几千个碱基对。“组成型”启动子是一种在大部分环境和发育条件下有活性的启动子。“可诱导”启动子是在环境或发育调控下活性的启动子。术语“可操纵连接”指核酸表达控制序列(如启动子,或转录因子结合位点的阵列)和第二核酸序列之间的功能性键连接,其中表达控制序列指导了对应于第二序列的核酸的转录。
“表达载体”是重组或合成产生的核酸构建体,具有一系列允许特定核酸在宿主细胞中转录的特定核酸元件。表达载体可以是质粒、病毒、或核酸片段的一部分。通常,表达载体包含与启动子可操纵性连接的待转录的核酸。
关于两种或多种核酸或多肽序列,术语“相同”或百分数“相同性”指两条或多条序列或亚序列,当在比较窗口或指定区域上如用下列序列比较算法之一或手工排列和目测,比较并排列对比寻找最大对应性时它们是相同的,或具有特定百分比的相同氨基酸残基或核苷酸(即在一特定区域内有50%相同性,可任选55%、60%、65%、70%、75%、80%、85%、90%、95%或更高的相同性)。这样的序列称为“基本相同”。该定义还指对测试序列的评价。可任选的,相同性存在于至少长度为50个氨基酸和核苷酸的区域中,或更优选的存在于长度为75-100个氨基酸和核苷酸的区域中。
对于序列比较,通常将一条序列作为参照序列,与测试序列比较。当使用序列比较算法时,将测试序列和参照序列输入计算机,如需要,指定亚序列坐标,并指定序列算法程序参数。可使用默认程序参数,如下文的BLASTN和BLASTP程序所述的,或指定另外的参数。然后序列比较算法根据程序参数计算出测试序列相对于参照序列的序列相同性百分数。
本文所用的“比较窗”包括参比选自包含20-600、一般约50-200,更一般约100-150个毗邻位置数之一的片段,其中在将两条序列最佳排列后,可将一条序列和毗邻位置数相同的参比序列作比较。排列序列作比较的方法是本领域熟知的。可通过例如,Simth & Waterman,Adv.Appl.Math.,2:482(1981)的局部同源性算法,Needleman & Wunsch,J.Mol.Biol.,48:443(1970)的同源排列算法,Pearson& Lipman,Proc.Natl.Acad.Sci.USA 85:2444(1988)的类似方法,这些算法的计算机化操作(Wisconsin Genetics Software Package的GAP,BESTFIT,FASTA,和TFASTA,Genetics Computer Group,575 Science Dr.,Madison,WI),或手工排列和目测(见例如分子生物学现有方案(Ausubel等,编1995增订))来进行序列的最佳排列,作比较。
一种适用于测定序列相同性百分数和序列相似性的算法例子是BLAST和BLAST2.0算法,Altschul等,Nuc.Acids Res.25:3389-3402(1977)和Altschul等,J.Mol.Biol.215:403-410(1990)分别描述了它们。通过生物技术信息国家中心(
http://www.ncbi.nlm.nih.gov/)公众可获得进行BLAST分析的软件。该算法包括首先通过鉴定查询序列中长度为W的字,以鉴定高评分的配对(HSP),当与数据库序列中同样长度的字排列对比时,匹配或满足某个正值的阈评分T。T称为相邻字评分阈值(Altschul等,见上)。找到这些起始的相邻字作为引发寻找含有它们的更长HSP的搜索的出发点。将找到的字沿着各序列双向延伸,直到能增加累计的排列对比评分。对于核苷酸序列,用参数M(一对匹配残基的加分;总是>0)和N(错配残基的减分,总是<0)计算累计评分。对于氨基酸序列,用评分矩阵来计算累计评分。当:累计的排列对比评分从其最大达到值处下落数量X;由于一个或多个负评分残基排列对比的累计,该累计评分达到0或以下;或达到两序列之一的末端时,找到的字延伸被停止。BLAST算法参数W、T和X确定了排列对比的灵敏度和速度。BLASTN程序(用于核苷酸序列)用作默认值的句长(W)为11,预期值(E)为10,M=5,N=-4,比较两条链。对于氨基酸序列,BLASTP程序用作默认值的句长为3,预期值(E)为10,和BLOSUM62评分矩阵(见Henikoff & Henikoff,Proc.Natl.Acad.Sci.USA 89:10915(1989))排列(B)为50,预期值(E)为10,M=5,N=-4,并比较双链。
另一种有用算法的例子是PILEUP。PILEUP创立了一组相关序列的多条序列排列,使用推进的、成对排列对比来显示相互关系和序列相同性百分数。它还绘出了树状图或枝状图来显示用于建立排列对比的分类归并关系。PILEUP使用Feng &Doolittle,J.Mol.Evol.35:351-360(1987)的推进排列方法的简化形式。使用的方法和Higgins & Sharp,CABIOS 5:151-153(1989)描述的方法相似。该程序可排列多达300条序列,各自最大长度为5,000个核苷酸或氨基酸。该多重排列程序始于两条最相似序列的成对排列对比,产生一个两条排列序列的组。然后将该组和下一条最相关的序列或排列序列的组排列对比。通过简单延伸两条独立序列的配对排列,对两组序列进行排列对比。通过一系列推进的成对排列来实现最终的排列对比。通过指定特定序列及其与序列比较区域同等的氨基酸或核苷酸,并指定程序参数来运行该程序。使用PILEUP,将参照序列和其它测试序列作比较,用下列参数测定序列相同性百分数:默认缺口加权值(3.00)、默认缺口长度加权值(0.10)、和加权的末端缺口。PILEUP可从GCG序列分析软件包,如7.0版(Devereaux等,Nuc.Acids Res.12:387-395(1984))获得。
两条核酸序列或多肽基本相同的指标是,第一条核酸编码的多肽与针对第二条核酸编码的多肽产生的抗体发生免疫性交叉反应(如下所述)。因此,如果两条肽仅在保守取代上有不同,该多肽通常与第二条多肽基本相同。另一个两条核酸序列基本相同的指示是两个分子或其互补分子在严格条件下相互杂交(如下所述)。而另一个两条核酸序列基本相同的指示是可用同样的引物扩增该序列。
词组“与…选择性(或特异性)杂交”指当某序列存在于复杂混合物中(如总细胞或文库DNA或RNA)时,该分子仅和特定的核苷酸序列在严格条件下结合、形成双体或杂交。
词组“严格杂交条件”指在该条件下,通常在核酸的复杂混合物中,一种探针将与其靶亚序列杂交,但不和其它序列杂交。严格条件是序列依赖性的,而且视不同环境而不同。越长的序列在越高的温度下特异性杂交。在Tijssen,生物化学和分子生物学技术与核酸探针杂交,“杂交原理纵览和核酸试验策略”(1993)中可找到核酸杂交的一般指南。一般,所选的严格条件比在限定离子强度的pH下,特定序列的热解链温度(Tm)低约5-10℃。Tm是在平衡时,有50%与靶互补的探针与靶序列杂交的(特定离子浓度、pH和核酸的浓度下)温度(当靶序列过量,在Tm下平衡时50%探针被使用)。严格条件将是盐浓度小于约1.0M钠离子,通常约0.01-1.0M钠离子浓度(或其它盐),pH7.0-8.3,对于短探针(如10-50个核苷酸)温度至少约30℃,和对于长探针(如50个核苷酸以上)至少约60℃的条件。可加入去稳定剂,如甲酰胺来达到严格条件。对于选择性或特异性杂交,阳性信号是至少两倍于背景,可任选10倍于背景杂交。示范性严格条件可如下:50%甲酰胺,5xSSC,和1%SDS,42℃培育,或5x SSC,1%SDS,65℃培育,用0.2xSSC和0.1%SDS 65℃洗涤。这些杂交和洗涤步骤可以在例如1、2、5、10、15、30、60或更多时间内进行。
如果核酸编码的肽基本相同,在严格条件下不互相杂交的核酸仍是基本相同的。例如这发生在当用基因密码允许的最大密码子简并性建立核酸拷贝时。对此,核酸通常在中等严格杂交条件下杂交。示范性“中等严格杂交条件”包括在40%甲酰胺、1M NaCl、1%SDS的缓冲液中37℃杂交,并以1XSSC 45℃洗涤。该杂交和洗涤步骤可进行1、2、5、10、15、30、60或更多分钟。阳性杂交是至少背景的两倍。本领域普通技术人员将轻易认识到,可用其它杂交和洗涤条件来提供严格性相似的条件。
“抗体”指含有免疫球蛋白基因编码的框架区的多肽或其片段,其能特异性结合并识别一种抗原。已知的免疫球蛋白基因包括κ、λ、α、γ、δ、ε和μ恒定区基因,以及无数免疫球蛋白可变区基因。轻链分类为κ或λ。重链分类为γ、μ、α、δ或ε,其进而分别定义为免疫球蛋白,IgG、IgM、IgA、IgD和IgE类。
典型的免疫球蛋白(抗体)的结构单元是四聚体。各四聚体由相同的两对多肽链构成,每对具有一条“轻”链(约25kDa)和一条“重”链(约50-70kDa)。每条链的N-末端定义为约100-110或以上的氨基酸的可变区,主要负责抗原识别。术语可变轻链(VL)和可变重链(VH)分别指这些轻链和重链。
抗体可以完整的免疫球蛋白或各种肽酶消化产生的,特性明确的片段存在。因此,例如胃蛋白酶在铰链区二硫键下方消化抗体,产生F(ab)’2,一种Fab的二聚物,它自身是一条通过二硫键与VH-CH1结合的轻链。F(ab)’2可在温和条件下还原而打断铰链区中的二硫键,从而将F(ab)’2二聚物转化成Fab’单体。Fab’单体本质上是带有部分铰链区的Fab(见Fundamental Immunology(Paul编,第三版,1993))。虽然各种抗体片段被定义为完整抗体的消化物,本领域技术人员将理解,这些片段可用化学方法或用重组DNA方法合成。因此,本文使用的术语抗体还包括,修饰完整抗体产生的,或用重组DNA方法(如单链Fv)从头合成的,或用噬菌体展示文库鉴定的抗体片段(见例如,McCafferty等,Nature 348:552-554(1990))。
为了制备单克隆或多克隆抗体,可使用任何本领域已知的技术(见例如,Kohler & Milstein,Nature 256:495-497(1975);Kozbor等,Immunology Today4:72(1983);Cole等,Monoclonal Anitibodies and Cancer Therapy 77-96页(1985))。可使用产生单链抗体的技术(美国专利4,946,778),来产生抗本发明多肽的抗体。还可用转基因小鼠,或其它生物体,如其它哺乳动物来表达人源化抗体。另外,可用噬菌体展示技术来鉴定能与所选抗原特异性结合的抗体和异聚Fab片段(见例如McCafferty等,Nature 348:552-554(1990);Marks等,Biotechnology10:779-783(1992))。
“嵌合抗体”是一种抗体分子,其中(a)恒定区或其部分被改变、置换或交换,使抗原结合位点(可变区)与不同的或另一类抗体的有效应器功能的和/或动物的恒定区连接,或与赋予嵌合抗体新性能的完全不同的分子,如酶、毒素、激素、生长因子、药物等连接;或(b)用具有不同的或抗原特异性改变的可变区来改变、置换或交换其可变区或其部分。
“抗-T2R”抗体是能与T2R基因、cDNA、或其亚序列编码的多肽特异性结合的抗体或抗体片段。
术语“免疫试验”是用抗体与抗原特异性结合的试验。免疫试验的特征是用特定抗体的特异性结合性能来分离、靶向、和/或定量测定抗原。
当指蛋白质或肽时,词组“特异性(或选择性)结合”于抗体或“与抗体起特异性(或选择性)免疫反应”指一种结合反应,其对于蛋白质或其它生物制剂的异质性群体中该蛋白质的存在有确定作用。因此,在指定的免疫试验条件下,特定抗体与一特定蛋白质的结合至少是背景的两倍,而且和样品中存在的其它蛋白质基本上不以显著量结合。在这种条件下与抗体特异性结合需要选出一种抗体,其对与具体蛋白质有特异性。例如,可选用特殊动物,如大鼠、小鼠或人产生的抗T2R家族成员的多克隆抗体,来获得只与T2R蛋白质或其免疫原性部分有特异性免疫反应性,但不和其它蛋白质(除了T2R蛋白的直向同源物或多态变体和等位基因)起反应的多克隆抗体。可通过除去与来自其它物种的T2R分子或其它T2R分子起交叉反应的抗体,来实现该选择。还可选择仅识别T2R GPCR家族成员,但不识别其它家族的GPCR的抗体。可用各种形式免疫试验选择能与特定蛋白质特异性免疫反应的抗体。例如,常规使用固相ELISA免疫试验选择能与蛋白质起特异性免疫反应的抗体(见例如,Harlow & Lane,Antibodies,A Laboratory Manual(1988),对用于测定特异免疫反应性的免疫试验形式和条件的描述)。通常特异性或选择性反应将至少是背景信号或噪声的两倍,更通常是背景的10-100倍。
在一个实施例中,可用对应于SEQ ID NO:166-171的免疫原性结构域产生与T2R家族的多肽特异性结合的抗体。
词组“与…选择性结合”指一核酸与以上定义的另一核酸“选择性杂交”的能力,或一抗体与以上定义的一种蛋白质“选择性(或特异性)结合”的能力。
“宿主细胞”意味觉着含有一表达载体并支持该表达载体复制或表达的细胞。宿主细胞可以是大肠杆菌等原核细胞,或真核细胞,例如酵母、昆虫、两栖类或CHO、HeLa、HEK-293等哺乳动物细胞,如培养的细胞、外植体和体内细胞。
III.编码T2R家族成员的核酸的分离
A.一般重组DNA方法
本发明依据重组遗传学领域中的常规技术。公开本发明所用一般方法的基础文章包括Sambrook等,Molecular Cloning,A Laboratory Manual(第二版,1989);Kriegler,Gene Transfer and Expression:A Laboratory Manual(1990);和CurrentProtocols in Molecular Biology(Ausubel等编,1994)。
对于核酸,以千碱基(kb)或碱基对(bp)给出了大小。这些是从琼脂糖或丙烯酰胺凝胶电泳,从测序的核酸,或从出版的DNA序列估计得到的。对于蛋白质,以千道尔顿(kDa)或氨基酸残基数给出了大小。可从凝胶电泳,从测序的蛋白质,从衍生的氨基酸序列,或从出版的蛋白质序列估计蛋白质大小。
不能商品购得的寡核苷酸可根据Beaucage & Caruthers,Tetrahedron letts.22:1859-1862(1981)首选描述的固相亚磷酰胺三酯方法,使用自动合成仪,如VanDevanter等,Nucleic Acids Res.,12:6159-6168(1984)所述来化学合成。寡核苷酸的纯化可采用天然丙烯酰胺凝胶电泳或Pearson & Reanier,J.Chrom255:137-149(1983)所述的阴离子交换HPLC。
克隆后,可用例如Wallace等,Gene16:21-26(1981)的用于测序双链模板的链终止方法验证克隆的基因和合成的寡核苷酸序列。
B.分离编码T2R家族成员核苷酸序列的克隆方法。
一般从cDNA和基因组DNA文库,通过和探针杂交来克隆,或用寡核苷酸探针通过扩增技术来分离编码T2R家族成员的核酸序列和相关核酸序列同系物。例如,通常从哺乳动物核酸(基因组或cDNA)文库,和核酸探针(其序列可衍生自SEQ IDNO:2、SEQ ID NO:4、SEQ ID NO:6、SEQ ID NO:8、SEQ ID NO:10、SEQ ID NO:12、SEQ ID NO:14、SEQ ID NO:16、SEQ ID NO:18、SEQ ID NO:20、SEQ ID NO:23、SEQ ID NO:25、SEQ ID NO:27、SEQ ID NO:29、SEQ ID NO:31、SEQ ID NO:34、SEQ ID NO:36、SEQ ID NO:38、SEQ ID NO:41、SEQ ID NO:43、SEQ ID NO:45、SEQ ID NO:51、SEQ ID NO:52、SEQ ID NO:54、SEQ ID NO:57、SEQ ID NO:61、SEQ ID NO:63、SEQ ID NO:78、SEQ ID NO:80、SEQ ID NO:82、SEQ ID NO:84、SEQ ID NO:86、SEQ ID NO:88、SEQ ID NO:90、SEQ ID NO:92、SEQ ID NO:94、SEQ ID NO:96、SEQ ID NO:98、SEQ ID NO:100、SEQ ID NO:102、SEQ IDNO:104、SEQ ID NO:106、SEQ ID NO:108、SEQ ID NO:110、SEQ ID NO:112、SEQ ID NO:114、SEQ ID NO:116、SEQ ID NO:118、SEQ ID NO:120、SEQ ID NO:122、SEQ ID NO:124、SEQ ID NO:126、SEQ ID NO:128、SEQ ID NO:130、SEQID NO:132、SEQ ID NO:134、SEQ ID NO:136、SEQ ID NO:138、SEQ ID NO:140、SEQ ID NO:142、SEQ ID NO:144、SEQ ID NO:146、SEQ ID NO:148、SEQ ID NO:150、SEQ ID NO:152、SEQ ID NO:154、SEQ ID NO:156、SEQ ID NO:157、SEQID NO:159、SEQ ID NO:161、SEQ ID NO:163或SEQ ID NO:165)杂交,来分离T2R序列。可分离得到T2R家族成员的RNA和cDNA的合适组织是舌组织,可任选味蕾组织或单个味觉细胞。
还可用使用引物的扩增技术,从DNA或RNA扩增并分离T2R序列。还可用编码下列氨基酸序列的简并引物扩增T2R基因的序列:SEQ ID NO:166、167、168、169、170或171(见例如,Dieffenfach & Dveksler,PCR Primer:A LaboratoryManual(1995))。可用这些引物来扩增全长序列或一到数百个核苷酸的探针,然后可用其筛选哺乳动物文库中的全长T2R克隆。如上所述,可用这些探针分离全长序列,或例如从文库分离用于分离全长序列的探针。
还可用抗体作为探针从表达文库中分离编码T2R的核酸。可用SEQ ID NO:1、SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ ID NO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ ID NO:22、SEQ ID NO:24、SEQ ID NO:26、SEQ ID NO:28、SEQ ID NO:30、SEQ ID NO:32、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQ ID NO:39、SEQ ID NO:40、SEQ ID NO:42、SEQ ID NO:44、SEQ ID NO:46、SEQ ID NO:47、SEQ ID NO:48、SEQ ID NO:49、SEQ ID NO:50、SEQ ID NO:51、SEQ ID NO:53、SEQ ID NO:55、SEQ ID NO:56、SEQ ID NO:58、SEQ ID NO:59、SEQ ID NO:60、SEQ ID NO:62、SEQ ID NO:64、SEQ ID NO:65、SEQ ID NO:66、SEQ ID NO:67、SEQ ID NO:68、SEQ ID NO:69、SEQ ID NO:70、SEQ ID NO:71、SEQ ID NO:72、SEQ ID NO:73、SEQ ID NO:74、SEQ ID NO:75、SEQ ID NO:76、SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ ID NO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ ID NO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQ ID NO:123、SEQ ID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQ ID NO:151、SEQ ID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ ID NO:162或SEQ ID NO:164的序列来产生这类多克隆和单克隆抗体。
可用T2R核酸探针和寡核苷酸在严格条件下,通过筛选文库来分离与T2R家族成员基本相同的多态变体、等位基因和种间同系物。另外,可用表达文库通过检测表达的同系物(其能和针对T2R多肽制备的抗血清或纯化的抗体(它们也识别并与T2R同系物选择性结合)起免疫反应),来克隆T2R家族成员和T2R家族成员的多态变体、等位基因和种间同系物。
为了制备cDNA文库,应选择富含T2R mRNA的来源,如舌组织或分离的味蕾。然后用反转录酶将mRNA变成cDNA,连接入重组载体,转染入重组宿主来增殖、筛选并克隆。制备和筛选cDNA的方法是熟知的(见例如,Gubler & Hoffmann,Gene 25:263-269(1983);Sambrook等,见上;Ausubel等,见上)。
对于基因组文库,从组织中抽提DNA,机械剪切或用酶消化,得到约12-20kb的片段。然后用梯度离心将该片段与不需要的片段分开,构建入λ噬菌体载体。体外包装这些载体和噬菌体。用Benton & Davis,Science 196:180-182(1977)所述的噬斑杂交分析重组噬菌体。如Grunstein等,Proc.Natl.Acad.Sci.USA 72:3961-3965(1975)中所述进行集落杂交。
另一种分离T2R核酸及其同系物的方法,联合使用合成的寡核苷酸引物和RNA或DNA模板扩增(见美国专利4,683,195和4,683,202;PCR Protocols:A Guideto Methods and Applications(Innis等编,1990))。可用聚合酶链式反应(PCR)和连接酶链式反应(LCR)直接从mRNA、cDNA、基因组文库或cDNA文库扩增T2R基因的核酸序列。可设计简并寡核苷酸用本文提供的序列扩增T2R家族成员的同系物。可将限制性内切酶位点掺入引物,聚合酶链式反应或其它体外扩增的方法也可用来,例如,克隆编码待表达蛋白质的核酸序列,来制备核酸,用作检测生理学样品中编码T2R的mRNA的存在与否,用于核酸测序或其它目的。可从琼脂糖凝胶中纯化PCR反应扩增的基因,并克隆入合适的载体。
还可用本领域已知的技术,如反转录和mRNA扩增,总RNA或poly A+RNA分离,RNA印迹法、斑点印迹法、原位杂交、RNase保护、探测DNA微芯片阵列等,来分析T2R家族成员基因的表达。在一个实施例中,用高密度寡核苷酸分析技术(如GeneChipTM)来鉴定本发明GPCR的同系物和多态变体。就被鉴定的同系物与已知疾病的联系而言,可用它们和GeneChipTM作为诊断工具,来检测生物学样品中的疾病,见例如,Gunthand等,AIDS Res.Hum.Retroviruses 14:869-876(1998);Kozal等,Nat.Med.2:753-759(1996);Matson等,Anal.Biochem.224:110-106(1995);Lockhart等,Nat.Biotechnol.14:1675-1680(1996);Gingeras等,Genome Res.8:435-448(1998);Hacia等,Nucleic Acids.Res.26:3865-3866(1988)。
可用合成的寡核苷酸构建重组T2R基因,用作探针或表达蛋白质。用一系列重叠寡核苷酸,一般长40-120bp,代表基因的有义和反义链来进行该方法。然后将这些DNA片段退火、连接并克隆。另外,可用扩增技术和精密引物来扩增T2R核酸的特定亚序列。然后将该特异性亚序列连接入表达载体。
通常在转化入原核或真核细胞中,用于复制和/或表达前,将编码T2R基因的核酸克隆入中间载体。这些中间载体通常是原核载体,如质粒或穿梭载体。
可任选的,根据标准技术制备编码含有T2R多肽或其结构域的嵌合蛋白质的核酸。例如,可将配体结合域等功能域、胞外功能域、跨膜功能域(如含有7个跨膜区域和对应的胞外和胞质环)、跨膜功能域和细胞质功能域、活性位点、亚基结合区域等与异源蛋白质共价连接。例如,可将胞外功能域与异源GPCR跨膜功能域连接,或异源GPCR胞外功能域可与跨膜功能域连接。其它所选异源蛋白质包括例如,绿色荧光蛋白质,β-半乳糖苷酶、谷氨酸受体和视紫红质前序列。
C.原核细胞和真核细胞中的表达
为了获得一克隆的基因或核苷酸,如编码T2R家族成员的cDNA的高水平表达,通常将T2R序列亚克隆入含有指导转录的强启动子、转录/翻译终止子,和(如果核酸编码蛋白)引发翻译的核糖体结合位点的表达载体。合适的细菌启动子是本领域熟知的,而且在Sambrook等和Ausubel等中有所描述。表达T2R蛋白质的细菌表达系统是在如大肠杆菌,芽胞杆菌亚种和沙门氏菌中可得到的(Palva等,Gene 22:229-235(1983);Mosbach等,Nature 302:543-545(1983))。这些表达系统的试剂盒是可商业购得的。哺乳动物细胞、酵母和昆虫细胞的真核表达系统是本领域熟知的,而且也可商业购得。在一个实施例中,真核表达载体是腺病毒载体,一种腺伴随病毒载体或反转录病毒载体。
用来指导异源核酸表达的启动子由特定用途决定。可将启动子置于距离异源转录起始位点与离其天然位置中转录起始位点相同部位。然而如本领域已知,该距离可容纳一些变化,而启动子功能不丧失。
除了启动子,通常表达载体还含有一种转录单元或表达盒,其含有编码T2R的核酸在宿主细胞中表达所需的所有其它元件。因此,典型的表达盒含有与编码T2R的核酸序列可操纵性连接的启动子,和转录物有效聚腺苷酸化,核糖体结合位点,和翻译终止所需的信号。编码T2R的核酸序列通常与可切割信号肽序列连接,以促进转化细胞分泌编码蛋白质。这些信号肽将包括:组织纤维蛋白溶酶原激活物、胰岛素和神经生长因子、和烟芽夜蛾(Heliothis virescens)的保幼激素酯酶的信号肽。其它盒元件可包括增强子,和(如果用基因组DNA作为结构基因)带有功能性剪接供体和受体位点的内含子。
除了启动子序列,表达盒还应含有在结构基因下游的转录终止区,以提供有效终止。可从启动子序列的相同基因或从不同基因获得该终止区。
用来将遗传信息转运入细胞的特定表达载体不是特别关键的。可用任何用于在真核或原核细胞中表达的常规载体。标准的细菌表达载体包括质粒,如pBR322为基础的质粒、pSKF、pET23D和融合表达系统,如GST和LacZ。还可将表位标记加到重组蛋白质中,以提供便利的分离方法,如c-myc。
通常在真核表达载体,如SV40载体、乳头瘤病毒载体,和衍生自Epstein-Barr病毒的载体中使用真核病毒的,含有调控元件的表达载体。其它示范性真核载体包括:pMSG、pAV009/A+、pMT010/A+、pMAMneo-5、杆状病毒pDSVE、和其它使蛋白质在SV40早启动子、SV40晚启动子、金属硫蛋白启动子、小鼠乳腺肿瘤病毒启动子、Rous肉瘤病毒启动子、多角体启动子或其它在真核细胞中显示表达效果的启动子的指导下,使蛋白表达的载体。
某些表达系统含有提供基因扩增的标记,如胸腺嘧啶激酶、潮霉素B磷酸转移酶和二氢叶酸还原酶。另外,与基因扩增无关的高产量表达系统也是适合的,如在昆虫细胞中使用杆状病毒载体,和在多角体启动子或其它强杆状病毒启动子指导下的编码T2R家族成员的序列。
通常包含于表达载体中的元件还包括在大肠杆菌中起作用的复制子、能选择含有重组质粒的细菌的编码抗生素抗性的基因、和质粒非必需区中能插入真核序列的独特限制性位点。所选的特定抗生素抗性基因不是关键的,许多本领域已知的抗性基因中的任一种都是适合的。可选择原核序列,如需要,使它们不干涉DNA在真核细胞中的复制。
用标准转染方法来产生表达大量T2R蛋白质(其然后可用标准技术纯化)的细菌、哺乳动物、酵母或昆虫细胞系(见例如,Colley等,J.Biol.Chem.,264:17619-17622(1989);Guide to Protein Purification,in Methods in Enzymology,182卷(Deutscher编,1990))。根据标准技术进行真核和原核细胞的转化(见例如,Morrison,J.Bact.132:349-351(1977);Clark-Curtiss & Curtiss,Methods inEnzymology 101:347-362(Wu等编,1983))。
可使用任何将外源核苷酸引入宿主细胞的熟知程序。这些包括使用磷酸钙转染、Polybrene,原生质体融合、电穿孔、脂质体、显微注射、血浆载体、病毒载体和其它用于将克隆的基因组DNA、cDNA、合成DNA或其它外源遗传材料引入宿主细胞的熟知方法(见例如,Sambrook等,见上)。只是需要所用的特定基因工程程序能将至少一个基因成功引入能表达T2R基因的宿主细胞。
在一个优选例中,例如,用pEAK10哺乳动物表达载体(Edge Biosystems,MD)编码T2R的多核苷酸与EF-1α启动子可操纵性连接。这可用任何标准方法,如用LipofectAMINE(Lifetechnologies)将这些载体引入细胞,如HEK-293细胞。
表达载体被引入细胞后,在适合表达T2R家族成员的条件下培养转染细胞,用下文所述的标准技术从培养物中回收。
IV.T2R多肽的纯化
可纯化天然存在或重组的T2R多肽,用于功能性试验。可任选的纯化重组的T2R多肽。从哺乳动物组织,如舌组织,或其它T2R同系物来源中纯化天然存在的T2R多肽。从任何合适细菌或真核表达系统,如CHO细胞或昆虫细胞中纯化重组T2R多肽。
可用标准技术,包括用硫酸铵等物质选择性沉淀;柱层析;免疫纯化方法等将T2R蛋白纯化至基本纯(见例如,Scopes,Protein Purification:Principal andPractice(1982);美国专利号4,673,641;Ausubel等,见上;和Sambrook,见上)。
当纯化重组T2R家族成员时,可使用许多程序。例如,可将具有分子粘合性能的蛋白质与T2R多肽可逆融合。用合适配体,可将T2R选择性吸附于纯化柱,然后以相对纯的形式从柱上脱离。然后可用酶作用除去融合的蛋白质。最后可用免疫亲和柱纯化T2R蛋白。
A.从重组细胞中纯化T2R蛋白
通常在启动子诱导后,用转化的细菌或真核细胞,如CHO细胞或昆虫细胞大量表达重组蛋白质;但表达可以是组成性的。用IPTG启动子诱导是可诱导启动子系统的一个例子。按本领域标准程序培养细胞。用新鲜或冻结的细胞分离蛋白质。
在细菌中表达的蛋白可能形成可溶性积聚物(“包含体”)。几种方案适用于纯化T2R包含体。例如,包含体的纯化包括破坏细菌细胞,如通过在50mM TRIS/HClpH 7.5,50mM NaCl,5mM MgCl2,1mM DTT,0.1mM ATP,和1mM PMSF的缓冲液中培育,抽提、分离和/或纯化包含体。可通过弗氏压碎器2-3次,裂解细胞悬液,用Polyton(Brinkman Instruments)或冰上超声进行匀浆。其它裂解细菌的方法本领域技术人员是清楚的(见例如,Sambrook等,见上;Ausubel等,见上)。
如需要,溶解包含体,通常离心裂解的细胞悬液,来除去不需要的不溶物。形成包含体的蛋白质可用兼容缓冲液稀释或透析复性。合适的溶剂包括但不限于:尿素(约4M-8M)、甲酰胺(至少约80%体积/体积成分)和胍基盐酸(约4M-8M)。能溶解形成的积聚蛋白质的某些溶剂,如SDS(十二烷基磺酸钠),70%甲酸在该程序中由于可能使蛋白质不可逆变性而伴有免疫原性和/或活性的缺乏,所以是不适合用于该程序的。虽然盐酸胍和相似的试剂是变性剂,但该变性不是不可逆的,而且可在除去(例如通过透析)或稀释变性剂后复性,重新形成免疫原性和/或生物学活性的蛋白质。其它合适的缓冲液是本领域技术人员已知的。可从其它细菌蛋白质中,通过标准分离技术,如Ni-NTA琼脂糖树脂分离T2R多肽。
另外,可从细菌周质中纯化T2R多肽。细菌裂解后,当T2R蛋白被输入到细菌周质中时,可通过冷低渗休克和其它本领域已知的技术来分离细菌的周质组分。为了从周质中分离重组蛋白质,离心细菌细胞,形成沉淀。将沉淀重新悬浮在含有20%蔗糖的缓冲液中。为了裂解细胞,离心细菌,将沉淀重新悬浮在冰冷的5mM MgSO4中,保存在冰浴中约10分钟。离心细胞悬液,倒出上清液并保存。可从宿主蛋白质中通过本领域技术人员熟知的标准分离技术分离出上清液中的重组蛋白质。
B.纯化T2R多肽的标准蛋白质分离技术
溶解度分级
通常作为起始步骤,尤其如果蛋白质混合物是复杂的,初步盐分级可从感兴趣的重组蛋白中分离除去许多不需要的宿主细胞蛋白质(或衍生自细胞培养液的蛋白质)。在一个实施例中,该盐是硫酸铵。硫酸铵通过有效减少蛋白质混合物中的水量来沉淀蛋白质。然后依据其溶解度沉淀蛋白质。蛋白质的疏水性越强,它就更可能在较低的硫酸盐浓度下沉淀出来。典型的方案包括在蛋白质溶液中加入饱和硫酸铵,从而使得到的硫酸铵浓度在20-30%之间。该浓度将沉淀最疏水的蛋白质。然后丢弃沉淀物(除非感兴趣的蛋白质是疏水的),在上清液中加入硫酸铵,至能沉淀感兴趣的蛋白质的浓度。然后将沉淀溶于缓冲液,如需要,通过透析或渗滤除去过量的盐。其它依靠蛋白质溶解度的方法,如冷乙醇沉淀,是本领域技术人员已知的,可用于级分复杂的蛋白质混合物。
大小差异性过滤
可利用T2R蛋白的分子量从更大或更小尺寸的蛋白质中,通过不同孔径的膜(例如Amicon或Millipore膜)超过滤分离该蛋白质。作为第一步,通过孔径能截断比感兴趣的蛋白质分子量小的膜超滤该蛋白质混合物。然后通过孔径能截断比感兴趣的蛋白质分子量大的膜超滤上次超滤的保留物。重组蛋白质将穿过膜进入滤液。然后可如下层析滤液。
柱层析
可从其它蛋白质中,依据其大小、净表面电荷、疏水性、和对配体的亲和性分离T2R蛋白。另外,可将针对蛋白质产生的抗体与柱基质偶联,免疫纯化该蛋白质。所有这些方法都是本领域熟知的。技术人员清楚的是,可以任何规模,使用许多不同厂商(如Pharmacia Biotech)的器具进行层析技术。
V.T2R多肽的免疫学检测
除了用核酸杂交技术检测T2R基因和基因表达,还可用免疫试验法来检测T2R,如鉴定味觉受体细胞,特别是苦味受体细胞,以及T2R家族成员的变体。可用免疫试验来定性或定量分析T2R。可使用的技术的综述可见Harlow & Lane,Antibodies:A Laboratory Manual(1988)。
A.T2R家族成员的抗体
产生与T2R家族成员特异性反应的多克隆和单克隆抗体的方法是本领域技术人员已知的(见例如,Coligan,Current Protocols in Immunology(1991);Harlow& Lane,见上;和Kohler & Milstein,Nature 256:495-497(1975))。这些技术包括从噬菌体或类似载体中的重组抗体文库选择抗体,以及免疫家兔或小鼠制备多克隆和单克隆抗体,来制备抗体(见例如Huse等,Science 246:1275-1281(1989);Ward等,Nature 341:544-546(1989))。
可用许多含有T2R的免疫原来产生与T2R家族成员特异性反应的抗体。例如,如本文所述分离了重组T2R蛋白或其抗原性片段。合适的抗原性区域包括例如:用于鉴定T2R家族成员的保守基序,即SEQ ID NO:166、167、168、169、170和171。可如上所述在真核细胞或原核细胞中表达重组蛋白质,并如上所述纯化。重组蛋白质是用于产生单克隆和多克隆抗体的免疫原的例子。另外,衍生自本文公开的序列,并与载体蛋白质偶联的合成肽可用作免疫原。还可使用天然存在的蛋白质,无论纯或不纯的形式。然后将产物注射入能产生抗体的动物。可产生单克隆或多克隆的抗体,随后用于免疫试验,以测定该蛋白质。
产生多克隆抗体的方法是本领域技术人员已知的。用蛋白质和一种标准佐剂,如Freund’s佐剂,和标准免疫方案免疫近交系小鼠(如BALB/C小鼠)或家兔。抽取测试血样,并测定对T2R的反应性滴度,监测动物对免疫原制备物的免疫应答。当获得对免疫原的合适的抗体高滴度时,从动物收集血液,并制备抗血清。如需要,可进一步级分该抗血清,来浓缩与该蛋白质起反应的抗体(见Harlow & Lane,见上)。
可通过各种与本领域技术人员相似的技术获得单克隆抗体。简单说,无限增殖用所需抗原免疫的动物脾细胞(常通过与骨髓瘤细胞融合)(见Kohler &Milstein,Eur J.Immunol.6:511-519(1976))。其它无限增殖的方法包括用Epstein Barr病毒、癌基因或反转录病毒转化或其它本领域熟知的方法。筛选从单个无限增殖细胞产生的集落,以产生对抗原具有所需特异性和亲和性的抗体,而且这些细胞产生的单克隆抗体产量可用各种技术增强,包括注射入脊椎动物宿主的腹腔。另外,可根据Huse等,Science 246:1275-1281(1989)描述的通用方法,从人B细胞中筛选DNA文库,分离编码单克隆抗体或其结合片段的DNA序列。
在免疫试验中,例如免疫原固定在固相载体上的固相免疫试验中收集单克隆抗体和多克隆血清,滴定其对免疫原蛋白质的滴度。通常,选择滴度为104或更大的多克隆抗血清,并使用竞争性结合免疫试验测试其对非T2R蛋白质或甚至其它T2R家族成员,或其它生物的其它相关蛋白质的交叉反应性。特异性多克隆抗血清和单克隆抗体结合的Kd值一般至少约0.1mM,更通常至少约1μM,可任选至少约0.1μM或更低,或可任选0.01μM或更低。
一旦得到T2R家族成员特异性抗体,可通过各种免疫试验方法检测T2R蛋白。对于免疫学和免疫试验程序的综述,见基础和临床免疫学(Stites & Terr编,第7版,1991)。另外,可用数种配置中的任一种进行本发明的免疫试验,在“酶免疫试验”一节中广泛描述(Maggio编,1980);和Harlow & Lane,见上。
B.免疫结合试验
可用许多熟知的免疫学结合试验来检测和定量测定T2R蛋白(见例如,美国专利4,366,241;4,376,110;4,517,288;和4,837,168)。对于一般免疫试验的综述,也见“细胞生物学方法:细胞生物学中的抗体”,37卷(Asai编,1993);“基础和临床免疫学”(Stitier & Terr编,第7版,1991)。免疫学结合试验(或免疫试验)通常使用与所选蛋白质或抗原特异性结合的抗体(就此T2R家族成员或其抗原性亚序列)。可通过许多本领域技术人员熟知的,或如上所述的方法产生该抗体(如抗T2R)。
免疫试验也常用标记试剂,来特异性结合或标记抗体和抗原形成的复合物。标记试剂自身可以是含有抗体/抗原复合物的分子之一。因此,标记试剂可以是标记的T2R多肽,或标记的抗T2R抗体。另外,标记试剂可以是第三种分子,例如第二抗体,其与抗体/T2R复合物特异性结合(第二抗体通常对产生第一抗体的动物的抗体是特异性的)。其它能与免疫球蛋白恒定区结合的蛋白质,如蛋白质A或蛋白质G也可用作标记试剂。这些蛋白质显示对不同动物的免疫球蛋白恒定区的非免疫性强反应性(见例如Kronval等,J.Immunol.111:1401-1406(1973);Akerstrom等,J.Immunol.135:2589-2542(1985))。可用可检测分子,如生物素(可与另一种分子特异性结合,如链霉亲和素)修饰标记试剂。各种可检测分子是本领域技术人员熟知的。
在试验中,在各试剂结合后可能需要培育和/或洗涤步骤。培育步骤可从5秒到数小时而不同,可任选从约5分钟到约24小时。然而,培育时间将由试验方案、抗原、溶液体积、浓度等决定。通常,试验将在环境温度下进行,虽然可在一定温度范围内进行,如10℃到40℃。
非竞争性试验
检测样品中T2R蛋白的免疫试验可以是竞争性的或非竞争性的。非竞争性免疫试验是直接测量抗原量的试验。在一优选的“夹心”试验中,例如抗T2R抗体可与固相基质直接结合,固定在其上。然后这些固定化抗体可捕获存在于测试样品中的T2R蛋白。因此固定的T2R蛋白被标记剂,如带有标记的第二T2R抗体结合。另外,第二抗体可缺少标记,但它可以被标记的第三抗体结合,该第三抗体对产生第二抗体的动物的抗体是特异性的。第二或第三抗体通常被可检测分子,如生物素(另一种分子与其特异性结合,如链霉亲和素)修饰,来提供可检测分子。
竞争性试验
在竞争性试验中,测量已知加入的(外源)T2R蛋白被样品中存在的未知T2R蛋白从抗T2R抗体上取代(竞争)的量,来间接测量样品中存在的T2R蛋白量。在一竞争性试验中,在一样品中加入已知量的T2R蛋白,然后使该样品与特异性结合T2R的抗体接触。与抗体结合的外源T2R蛋白量和样品中存在的T2R蛋白浓度成反比。在一实施例中,抗体被固定在固相基质上。与抗体结合的T2R蛋白量可通过测量在T2R/抗体复合物中存在的T2R蛋白量,或通过测量剩余未复合的蛋白质量来确定。可提供标记的T2R分子来检测T2R蛋白量。
半抗原抑制试验是另一种竞争性试验。在该试验中,将已知的T2R固定在固相基质上。在样品中加入已知量的抗T2R抗体,然后使该样品与固定化的T2R接触。与已知固定化T2R蛋白结合的抗T2R抗体量,与样品中存在的T2R蛋白量成反比。另外,可通过检测抗体的固定化组分或剩余在溶液中的抗体组分来检测固定化抗体的量。抗体被标记时,检测可以是直接的,或随后加入与上述抗体特异性结合的分子间接检测。
交叉反应性决定簇
还可将竞争性结合形式中的免疫试验用于交叉反应性决定簇。例如,可将至少部分由SEQ ID NO:2、SEQ ID NO:4、SEQ ID NO:6、SEQ ID NO:8、SEQ ID NO:10、SEQ ID NO:12、SEQ ID NO:14、SEQ ID NO:16、SEQ ID NO:18、SEQ ID NO:20、SEQ ID NO:23、SEQ ID NO:25、SEQ ID NO:27、SEQ ID NO:29、SEQ ID NO:31、SEQ ID NO:34、SEQ ID NO:36、SEQ ID NO:38、SEQ ID NO:41、SEQ ID NO:43、SEQ ID NO:45、SEQ ID NO:51、SEQ ID NO:52、SEQ ID NO:54、SEQ ID NO:57、SEQ ID NO:61、SEQ ID NO:63、SEQ ID NO:78、SEQ ID NO:80、SEQ ID NO:82、SEQ ID NO:84、SEQ ID NO:86、SEQ ID NO:88、SEQ ID NO:90、SEQ ID NO:92、SEQ ID NO:94、SEQ ID NO:96、SEQ ID NO:98、SEQ ID NO:100、SEQ ID NO:102、SEQ ID NO:104、SEQ ID NO:106、SEQ ID NO:108、SEQ ID NO:110、SEQID NO:112、SEQ ID NO:114、SEQ ID NO:116、SEQ ID NO:118、SEQ ID NO:120、SEQ ID NO:122、SEQ ID NO:124、SEQ ID NO:126、SEQ ID NO:128、SEQ ID NO:130、SEQ ID NO:132、SEQ ID NO:134、SEQ ID NO:136、SEQ ID NO:138、SEQID NO:140、SEQ ID NO:142、SEQ ID NO:144、SEQ ID NO:146、SEQ ID NO:148、SEQ ID NO:150、SEQ ID NO:152、SEQ ID NO:154、SEQ ID NO:156、SEQ ID NO:157、SEQ ID NO:159、SEQ ID NO:161、SEQ ID NO:163或SEQ ID NO:165编码的蛋白质固定在固相载体上。在试验中加入与固定化抗原竞争结合抗血清的蛋白质(如T2R蛋白质和同源物)。比较加入的蛋白质与固定的蛋白质竞争结合抗血清的能力与SEQ ID NO:2、SEQ ID NO:4、SEQ ID NO:6、SEQ ID NO:8、SEQ ID NO:10、SEQ ID NO:12、SEQ ID NO:14、SEQ ID NO:16、SEQ ID NO:18、SEQ ID NO:20、SEQ ID NO:23、SEQ ID NO:25、SEQ ID NO:27、SEQ ID NO:29、SEQ ID NO:31、SEQ ID NO:34、SEQ ID NO:36、SEQ ID NO:38、SEQ ID NO:41、SEQ ID NO:43、SEQ ID NO:45、SEQ ID NO:51、SEQ ID NO:52、SEQ ID NO:54、SEQ ID NO:57、SEQ ID NO:61、SEQ ID NO:63、SEQ ID NO:78、SEQ ID NO:80、SEQ ID NO:82、SEQ ID NO:84、SEQ ID NO:86、SEQ ID NO:88、SEQ ID NO:90、SEQ ID NO:92、SEQ ID NO:94、SEQ ID NO:96、SEQ ID NO:98、SEQ ID NO:100、SEQ ID NO:102、SEQ ID NO:104、SEQ ID NO:106、SEQ ID NO:108、SEQ ID NO:110、SEQID NO:112、SEQ ID NO:114、SEQ ID NO:116、SEQ ID NO:118、SEQ ID NO:120、SEQ ID NO:122、SEQ ID NO:124、SEQ ID NO:126、SEQ ID NO:128、SEQ ID NO:130、SEQ ID NO:132、SEQ ID NO:134、SEQ ID NO:136、SEQ ID NO:138、SEQID NO:140、SEQ ID NO:142、SEQ ID NO:144、SEQ ID NO:146、SEQ ID NO:148、SEQ ID NO:150、SEQ ID NO:152、SEQ ID NO:154、SEQ ID NO:156、SEQ ID NO:157、SEQ ID NO:159、SEQ ID NO:161、SEQ ID NO:163或SEQ ID NO:165编码的T2R多肽与其自身竞争的能力。使用标准算法计算上述蛋白质的交叉反应性百分数。选择并合并与各种上述加入的蛋白质具有小于10%的交叉反应性的抗血清。可任选的通过加入所考虑的蛋白质(如远亲同系物),来免疫吸附,以除去合并抗血清中的交叉反应抗体。另外,可在交叉反应性确定中,使用含有代表用于鉴定T2R家族成员的保守基序的氨基酸序列的肽,即SEQ ID NO:166、SEQ ID NO:167、SEQ ID NO:168、SEQ ID NO:169、SEQ ID NO:170或SEQ ID NO:171。
然后将免疫吸附的合并抗血清用于上述竞争性结合免疫试验,来比较第二蛋白质(考虑其可能是T2R家族成员的等位或多态变体)和免疫原蛋白质(即SEQ ID NO:2、SEQ ID NO:4、SEQ ID NO:6、SEQ ID NO:8、SEQ ID NO:10、SEQ ID NO:12、SEQ ID NO:14、SEQ ID NO:16、SEQ ID NO:18、SEQ ID NO:20、SEQ ID NO:23、SEQ ID NO:25、SEQ ID NO:27、SEQ ID NO:29、SEQ ID NO:31、SEQ ID NO:34、SEQ ID NO:36、SEQ ID NO:38、SEQ ID NO:41、SEQ ID NO:43、SEQ ID NO:45、SEQ ID NO:51、SEQ ID NO:52、SEQ ID NO:54、SEQ ID NO:57、SEQ ID NO:61、SEQ ID NO:63、SEQ ID NO:78、SEQ ID NO:80、SEQ ID NO:82、SEQ ID NO:84、SEQ ID NO:86、SEQ ID NO:88、SEQ ID NO:90、SEQ ID NO:92、SEQ ID NO:94、SEQ ID NO:96、SEQ ID NO:98、SEQ ID NO:100、SEQ ID NO:102、SEQ ID NO:104、SEQ ID NO:106、SEQ ID NO:108、SEQ ID NO:110、SEQ ID NO:112、SEQID NO:114、SEQ ID NO:116、SEQ ID NO:118、SEQ ID NO:120、SEQ ID NO:122、SEQ ID NO:124、SEQ ID NO:126、SEQ ID NO:128、SEQ ID NO:130、SEQ ID NO:132、SEQ ID NO:134、SEQ ID NO:136、SEQ ID NO:138、SEQ ID NO:140、SEQID NO:142、SEQ ID NO:144、SEQ ID NO:146、SEQ ID NO:148、SEQ ID NO:150、SEQ ID NO:152、SEQ ID NO:154、SEQ ID NO:156、SEQ ID NO:157、SEQ ID NO:159、SEQ ID NO:161、SEQ ID NO:163或SEQ ID NO:165的T2R蛋白)。为了进行该比较,以广泛的浓度范围各检测了两种蛋白质,并确定抑制50%抗血清与固定化蛋白质结合所需的各蛋白质量。如果抑制50%结合所需的第二种蛋白质的量比SEQ ID NO:2、SEQ ID NO:4、SEQ ID NO:6、SEQ ID NO:8、SEQ ID NO:10、SEQ ID NO:12、SEQ ID NO:14、SEQ ID NO:16、SEQ ID NO:18、SEQ ID NO:20、SEQ ID NO:23、SEQ ID NO:25、SEQ ID NO:27、SEQ ID NO:29、SEQ ID NO:31、SEQ ID NO:34、SEQ ID NO:36、SEQ ID NO:38、SEQ ID NO:41、SEQ ID NO:43、SEQ ID NO:45、SEQ ID NO:51、SEQ ID NO:52、SEQ ID NO:54、SEQ ID NO:57、SEQ ID NO:61、SEQ ID NO:63、SEQ ID NO:78、SEQ ID NO:80、SEQ ID NO:82、SEQ ID NO:84、SEQ ID NO:86、SEQ ID NO:88、SEQ ID NO:90、SEQ ID NO:92、SEQ ID NO:94、SEQ ID NO:96、SEQ ID NO:98、SEQ ID NO:100、SEQ ID NO:102、SEQ ID NO:104、SEQ ID NO:106、SEQ ID NO:108、SEQ ID NO:110、SEQ ID NO:112、SEQ ID NO:114、SEQ ID NO:116、SEQ ID NO:118、SEQ ID NO:120、SEQID NO:122、SEQ ID NO:124、SEQ ID NO:126、SEQ ID NO:128、SEQ ID NO:130、SEQ ID NO:132、SEQ ID NO:134、SEQ ID NO:136、SEQ ID NO:138、SEQ ID NO:140、SEQ ID NO:142、SEQ ID NO:144、SEQ ID NO:146、SEQ ID NO:148、SEQID NO:150、SEQ ID NO:152、SEQ ID NO:154、SEQ ID NO:156、SEQ ID NO:157、SEQ ID NO:159、SEQ ID NO:161、SEQ ID NO:163或SEQ ID NO:165编码的蛋白质抑制50%结合所需的量少1O倍,则说第二种蛋白质能与针对T2R免疫原产生的多克隆抗体特异性结合。
还可用针对SEQ ID NO:166-171产生的抗体制备仅与T2R家族的GPCR,而不和其它家族的6PCR特异性结合的抗体。
可通过用其它T2R家族成员从交叉反应性抗体中扣除,制备与T2R家族的特定成员(如T2R01)特异性结合的多克隆抗体。可用类似方法制备物种特异性多克隆抗体。例如,可从与直向同源序列,如大鼠T2R01或小鼠T2R19交叉反应的抗体中扣除,制备对人T2R01特异性的抗体。
其它试验形式
用蛋白质印迹(免疫印迹)分析检测并定量样品中T2R蛋白的存在。该技术一般包括用凝胶电泳根据分子量分离样品蛋白质,将分离的蛋白质转移到合适固相载体(如硝酸纤维素滤膜、尼龙滤膜、或衍生的尼龙滤膜)上,并孵育样品和与T2R蛋白特异性结合的抗体。抗T2R多肽抗体与固相载体上的T2R多肽特异性结合。这些抗体可直接标记或随后用标记抗体(如标记的绵羊抗小鼠抗体,其与抗T2R抗体特异性结合)检测。
其它试验形式包括脂质体免疫试验(LIA),它用设计的脂质体结合特异性分子(如抗体),并释放包裹的试剂或标记物。然后根据标准技术检测释放的化学物质(见Monroe等,Amer.Clin.Prod.Rev.5:34-41(1986))。
非特异性结合的还原
本领域技术人员将理解,常需要使免疫试验中非特异性结合最小化。特别是,当试验涉及固定在固相基质上的抗原或抗体时,需要使与基质的非特异性结合量最小化是理想的。减少这些非特异性结合的方法是本领域技术人员熟知的。通常该技术涉及用蛋白质组合物包被基质。具体说,广泛使用牛血清清蛋白(BSA)、脱脂粉末状牛奶和明胶等蛋白质组合物,而粉末状牛奶是最优选的。
标记
在试验中使用的特定标记或可检测基团不是本发明的关键部分,只要它不显著影响试验中使用的抗体的特异性结合。可检测基团可以是任何具有可检测的物理或化学性能的材料。这些可检测标记已在免疫试验领域中良好开发,而且,一般这些方法中的大部分标记都可用于本发明。因此,标记是分光光度、光化学、生物化学、免疫化学、电子学、光学或化学方法可检测的任何组合物。本发明中可用的标记包括磁珠(如DYNABEADTM)、荧光染料(如异硫氰酸荧光素、Texas red、罗丹明等)、放射性标记(如3H、125I、35S、14C或32P)、酶(辣根过氧化物酶、碱性磷酸酶和其它ELISA中常用的酶)、和显色标记,如胶体金或着色玻璃或塑料珠(如聚苯乙烯、聚丙烯、乳胶等)。
根据本领域熟知的方法,可直接或间接将标记与所需试验成分偶联。如上所示,可使用各种各样的标记,选择标记由所需灵敏度、和化合物偶联的容易度、稳定性要求、可得仪器和处理准备而定。
常用间接方法来结合非放射性标记。通常配体分子(如生物素)与该分子共价结合。然后该配体与另一种分子(如链霉亲和素)结合,另一种分子本身可被检测或可与一种信号系统,如可检测的酶、荧光化合物或化学发光化合物共价结合。可将配体及其靶与识别T2R蛋白的抗体,或识别抗T2R的第二抗体适当联合使用。
可直接将分子与产生信号的化合物偶联,如与酶或荧光蛋白偶联。作为标记的感兴趣的酶主要是水解酶,特别是磷酸酯酶,酯酶和糖苷酶,氧化物酶(oxidotase),特别是过氧化物酶。荧光化合物包括荧光黄及其衍生物,罗丹明及其衍生物,丹(磺)酰、伞形酮等。化学发光化合物包括荧光素和2,3-二氢酞嗪二酮,如鲁米诺。关于可使用的各种标记或信号产生系统的综述,见美国专利号4,391,904。
检测标记的方法是本领域技术人员熟知的。因此例如,当标记是放射性标记时,检测仪器包括闪烁计数器或放射性自显影中的照相胶片。当标记是荧光标记时,可用合适波长的光激发荧光色素,并检测产生的荧光来检测。荧光可以目测,通过感光胶片,用电子探测器如电荷偶联装置(CCD)或光电倍增管等。类似的,可提供合适的酶底物并检测产生的反应产物,来检测酶标记。最后,可观察与标记相关的颜色,来简单检测显色标记。因此,在各种测量尺试验中,偶联的金常呈粉红色,而各种偶联的珠呈珠的颜色。
一些试验不需要使用标记的组分。例如,可用凝集试验检测靶抗体的存在。就此而言,抗原包裹的颗粒被含有靶抗体的样品凝集。在该试验中,没有任何成分需要标记,可简单目测靶抗体的存在。
VI.T2R家族成员的调节剂的试验
A.T2R蛋白活性试验
T2R家族成员及其等位和多态变体是参与味觉转导,如苦味转导的G-蛋白质偶联受体。可用不同的体外和体内试验评估T2R多肽的活性,来测定功能、化学和物理作用,如测量配体结合(如放射性配体结合)、第二信使(如cAMP、cGMP、IP3、DAG、或Ca2+)、离子流、磷酸化水平、转录水平、神经递质水平等。另外,可用这些试验检测T2R家族成员的抑制物和激活物。调节剂还可以是T2R受体的遗传改变版本。这些味觉转导活性的调节剂可用来按要求改变味觉。
该试验的T2R蛋白通常选自具有SEQ ID NO:1、SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ ID NO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ ID NO:22、SEQ ID NO:24、SEQ ID NO:26、SEQ ID NO:28、SEQ ID NO:30、SEQ ID NO:32、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQ ID NO:39、SEQ ID NO:40、SEQ ID NO:42、SEQ ID NO:44、SEQ ID NO:46、SEQ ID NO:47、SEQ ID NO:48、SEQ ID NO:49、SEQ ID NO:50、SEQ ID NO:51、SEQ ID NO:53、SEQ ID NO:55、SEQ ID NO:56、SEQ ID NO:58、SEQ ID NO:59、SEQ ID NO:60、SEQ ID NO:62、SEQ ID NO:64、SEQ ID NO:65、SEQ ID NO:66、SEQ ID NO:67、SEQ ID NO:68、SEQ ID NO:69、SEQ ID NO:70、SEQ ID NO:71、SEQ ID NO:72、SEQ ID NO:73、SEQ ID NO:74、SEQ ID NO:75、SEQ ID NO:76、SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ ID NO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ ID NO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQ ID NO:117、SEQ ID NO:119、SEQID NO:121、SEQ ID NO:123、SEQ ID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQ ID NO:145、SEQ ID NO:147、SEQID NO:149、SEQ ID NO:151、SEQ ID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ ID NO:162或SEQ ID NO:164的序列,或其保守性修饰的变体。
另外,试验用的T2R蛋白可衍生自真核生物,并包括具有与SEQ ID NO:1、SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ ID NO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ ID NO:22、SEQ ID NO:24、SEQ ID NO:26、SEQ ID NO:28、SEQ ID NO:30、SEQ ID NO:32、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQ ID NO:39、SEQ ID NO:40、SEQ ID NO:42、SEQ ID NO:44、SEQ ID NO:46、SEQ ID NO:47、SEQ ID NO:48、SEQ ID NO:49、SEQ ID NO:50、SEQ ID NO:51、SEQ ID NO:53、SEQ ID NO:55、SEQ ID NO:56、SEQ ID NO:58、SEQ ID NO:59、SEQ ID NO:60、SEQ ID NO:62、SEQ ID NO:64、SEQ ID NO:65、SEQ ID NO:66、SEQ ID NO:67、SEQ ID NO:68、SEQ ID NO:69、SEQ ID NO:70、SEQ ID NO:71、SEQ ID NO:72、SEQ ID NO:73、SEQ ID NO:74、SEQ ID NO:75、SEQ ID NO:76、SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ ID NO:87、SEQ ID NO:89、SEQ ID NO:91、SEQ ID NO:93、SEQ ID NO:95、SEQ ID NO:97、SEQ ID NO:99、SEQ ID NO:101、SEQ ID NO:103、SEQ ID NO:105、SEQ ID NO:107、SEQ ID NO:109、SEQ ID NO:111、SEQ ID NO:113、SEQ ID NO:115、SEQID NO:117、SEQ ID NO:119、SEQ ID NO:121、SEQ ID NO:123、SEQ ID NO:125、SEQ ID NO:127、SEQ ID NO:129、SEQ ID NO:131、SEQ ID NO:133、SEQ ID NO:135、SEQ ID NO:137、SEQ ID NO:139、SEQ ID NO:141、SEQ ID NO:143、SEQID NO:145、SEQ ID NO:147、SEQ ID NO:149、SEQ ID NO:151、SEQ ID NO:153、SEQ ID NO:155、SEQ ID NO:158、SEQ ID NO:160、SEQ ID NO:162或SEQ ID NO:164的氨基酸序列相同性的氨基酸序列。一般氨基酸序列相同性是至少60%,可任选至少70%-85%,可任选至少90-95%。可任选的,试验用的多肽可含有T2R蛋白的一个功能域,如胞外功能域、跨膜功能域、细胞质功能域、配体结合功能域、亚基结合功能域、活化位点等。T2R蛋白或其功能域可与异源蛋白共价连接,产生用于本文所述试验的嵌合蛋白质。
可用上述T2R多肽,不论是重组的或天然的,测试T2R受体的调节剂。可以分离蛋白质,在细胞中表达,在衍生自细胞的膜中表达,在组织或动物中表达。例如,可使用舌切片、从舌上脱下的细胞,转化的细胞或膜。用本文所述的体外或体内试验之一检测调节。可在体外用可溶性或固态反应,用T2R受体的全长T2R-GPCR或嵌合分子(如胞外结构域或跨膜区域,或其组合)与异源信号转导结构域共价连接,或异源胞外结构域和/或跨膜区域与T2R受体的跨膜和/或胞质结构域共价连接来检测味觉转导。另外,可在体外可溶性或固态反应中使用感兴趣蛋白质的配体结合功能域来测定配体结合。在许多实施例中,将制备含有全部或部分T2R多肽,以及另一条促使T2R定位到膜的序列,如视紫红质,如视紫红质蛋白的N-末端片段的嵌合受体。
可在溶液中、与固相结合的双层膜中,脂单层中,或在囊泡中测试与T2R蛋白、结构域或嵌合蛋白质结合的配体。可用例如,分光光度特性的变化(荧光、吸光度、折射指数)水动力学(如形状)、色谱或溶解特性的变化测试调节剂的结合。
还可检测受体-G-蛋白相互反应。例如,可检测G蛋白与受体的结合或其从受体上的释放。例如,不存在GTP的情况下,活化剂将导致G蛋白(所有三个亚基)与受体形成紧密的复合物。可用上述不同方法检测该复合物。可修改这些试验,来搜索抑制剂。在GTP不存在时在受体和G蛋白中加入激活剂,形成紧密复合物,然后通过观察受体-G蛋白复合物的解离来筛选抑制剂。GTP存在下,G蛋白α亚基从其它两个G蛋白亚基上释放可作为活化的指标。
在特定的优选例中,监测作为T2R受体激活的T2R-gustducin相互作用。如实施例IX所示,小鼠T2R5显示与Gustducin的强环己酰亚胺依赖性偶联。可用这种T2R受体的配体依赖性偶联作为标记,鉴定任何T2R家族成员的修饰物。
活化的或抑制的G蛋白将进而改变靶酶、通道和其它效应蛋白的性能。经典的例子是在视觉系统中转导素活化cGMP磷酸二酯酶。刺激性G蛋白活化腺嘌呤核苷酸环化酶,Gq和其它同源G蛋白对磷脂酶的活化,和Gi与其它G蛋白对多种通道的调节。也可检测下游共有序列,如磷脂酶C产生二酰甘油和IP3,和进而IP3使钙迁移。
在一个优选例中,T2R多肽在真核细胞中作为嵌合受体,与异源、嵌合体序列一起表达,该序列促使其成熟并靶向通过分泌途径。在一个优选例中,异源序列是视紫红质序列,如视紫红质的N-末端片段。该嵌合T2R序列可在任何真核细胞,如HEK-293细胞中表达。优选细胞含有功能性G蛋白,如Gα15,它能将嵌合受体与胞内信号途径偶联,或与信号传递蛋白,如磷脂酶Cβ偶联。这些嵌合受体在细胞中的激活可用任何标准方法检测,如通过检测细胞中FURA-2依赖性底物,检测胞内钙水平的改变。
活化的GPCR受体成为激酶的底物,激酶磷酸化受体的C-端尾(以及其它可能的位点)。因此,激活剂将启动32P从γ-标记的GTP向受体转移,这可用闪烁计数器测定。C-末端尾的磷酸化将启动视紫红质抑制蛋白样蛋白的结合,并将干扰G-蛋白的结合。该激酶/视紫红质抑制蛋白通路在许多GPCR受体的脱敏中起了关键作用。例如,调节味觉受体保持活化持续时间的化合物,可被用作延长所需味道或切断不好味道的工具。关于GPCR信号转导和检验信号转导的方法的综述,见例如,Methods in Enzymology,237卷和238卷(1994)和96卷(1983);Bourne等,Nature10:349:117-27(1991);Bourne等,Nature 348:125-32(1990);Pitcher等,Annu.Rev.Biochem.67:653-92(1998)。
将用潜在的T2R蛋白抑制剂或激活剂处理的样品或试验,与不含测试化合物的对照样品比较,来检测调节的程度。对照样品(未用激活剂或抑制剂)被指定相对T2R活性值为100。当与对照比较,T2R活性值是约90%,可任选50%,可任选25-0%时,实现对T2R蛋白的抑制。当T2R活性值相对于对照是110%,可任选150%,200-500%,或1000-2000%时,实现了对T2R蛋白的活化。
通过测定表达T2R蛋白的细胞或膜的极化(即电势)的变化,来评估离子流的变化。一种测定细胞极化中的变化的方法是用电压钳和膜片钳技术,如“细胞结合”模式,“外翻”模式,和“全细胞”模式测量电流的变化(从而测量极化的变化)(见例如,Ackerman等,New Engl.J.Med.336:1575-1595(1997))。用标准方法可方便的测定全细胞电流(见例如,Hamil等,PFlugers.Archiv.391:85(1981))。其它已知试验包括:放射性标记离子流试验和使用电压-敏感性染料的荧光试验(见例如Vestergarrd-Bogind等,J.Memberane Biol.88:67-75(1988);Gonzales& Tsien,Chem.Biol.4:269-277(1997);Daniel等,J.Pharmacol.Meth.25:185-193(1991);Holevinsky等,J.Membrane Biol.137:59-70(1994))。一般要测试的化合物存在范围是1pM到100mM。
可检测上述任一参数来测量测试化合物对该多肽功能的作用。可用任何合适的影响GPCR活性的生理学变化来评估测试化合物对本发明多肽的影响。当用完整细胞或动物测定功能性结果时,还可测量不同作用,如递质释放、激素释放、已知的和未确定特征的基因标记物(如RNA印迹)的转录变化,细胞代谢的变化,如细胞生长或pH的变化,和胞内第二信使,如Ca2+、IP3、cGMP和cAMP的变化。
G蛋白偶联受体的优选试验包括负载有离子或电压敏感性染料,能报告受体活性的细胞。测定这些受体活性的试验还可使用已知的作为阴性或阳性对照的G-蛋白偶联受体的刺激剂和拮抗剂,来评估测试化合物的活性。在鉴定调节性化合物(如刺激剂、拮抗剂)的试验中,细胞质离子水平或膜电压的变化将分别用离子敏感性指示剂或膜电压荧光指示剂监测。在可使用的离子敏感性指示剂和电压探针中,可使用“分子探针1997”目录中公开的那些。对于G-蛋白偶联受体,可在所选试验中使用混合G-蛋白如Gα15和Gα16(Wilkie等,Proc.Natl.Acad.Sci.USA 88:10049-10053(1991))。这些混合G-蛋白能偶联大量受体。
受体活化通常引发随后的胞内活动,如第二信使,如IP3的增加,其释放胞内钙离子的储藏。某些G-蛋白偶联受体的活化通过磷脂酶C-介导的磷脂酰肌醇的水解,刺激肌醇三磷酸(IP3)的形成(Berridge & Irvine,Nautre 312:315-21(1984))。IP3进而刺激胞内钙离子储藏的释放。因此,细胞质中钙离子水平的变化,或第二信使水平,如IP3的变化,可用来评估G-蛋白偶联受体的功能。表达这些G-蛋白偶联受体的细胞可能是显示提高的细胞质钙水平,是胞内钙储藏和离子通道活化产生的结果,此时虽然不一定要在无钙缓冲液中进行这种试验,但理想的是可在任选补充了EGTA等螯合剂的缓冲液中进行这些试验,来甄别从内部储藏释放的钙产生的荧光反应。
其它试验可能涉及测定受体活性,其当被活化时,通过活化或抑制腺苷酸环化酶等酶,导致胞内环化核苷酸,如cAMP或cGMP水平的变化。存在环化核苷酸阀门离子通道,如视杆光受体细胞通道和嗅神经元通道,它们在与cAMP或cGMP结合活化后,对阳离子是可通透的(见例如,Altenhofen等,Proc.Natl.Acad.Sci.USA88:9868-9872(1991)和Dhallan等,Nature 347:184-187(1990))。就受体的活化导致环化核苷酸水平降低的情况而言,优选使细胞暴露于能提高胞内环化核苷酸水平的试剂,如毛喉素,然后在试验的细胞中加入受体激活化合物。可用编码环化核苷酸阀门离子通道、GPCR磷酸酶的DNA和编码(如某些谷氨酸受体、毒蝇碱性乙酰胆碱受体、多巴胺受体、血清紧张素受体等)受体(当被活化时,导致胞质环化核苷酸水平的变化)的DNA共转染宿主细胞,来制备用于该类试验的细胞。
在一个实施例中,在具有将受体与磷脂酶C信号转导途径连接的混杂G-蛋白的异源细胞中,表达T2R基因,测量了T2R蛋白活性(见Offermanns & Simon,J.Biol.Chem.270:15175-15180(1995);还见实施例II)。可任选的细胞系是HEK-293(它不天然表达T2R基因)而混杂G-蛋白是Gα15(Offermanns & Simon,见上)。测量胞内Ca2+水平的变化(该变化是对给予与T2R蛋白结合的分子,调节了T2R信号转导途径的反应),以检测味觉转导的调节。用荧光Ca2+指示染料和荧光显影任选的测量了Ca2+水平的变化。
在一实施例中,可用免疫试验测量胞内cAMP或cGMP的变化。Offermann &Simon,J.Biol.Chem.270:15175-15180(1995)描述的方法可用来测定cAMP的水平。另外,可用Felley-Bosco等,Am.J.Resp.Cell and Mol.Biol.11:159-164(1994)描述的方法测定cGMP的水平。另外,美国专利4,115,538中描述了一种用于测量cAMP和/或cGMP的试剂盒,在此引入以供参考。
在另一个实施例中,可根据美国专利5,436,128分析磷脂酰肌醇(PI)的水解,在此引入以供参考。简单说,试验涉及用3H-肌醇标记细胞48小时或以上。用测试化合物处理细胞1小时。溶解处理的细胞,用氯仿-甲醇-水抽提,然后用离子交换层析分离肌醇磷酸酯,并用闪烁计数定量。计算促效剂存在时的cpm与缓冲对照cpm的比值,确定刺激倍数。同样,计算拮抗剂存在时的cpm与缓冲液对照(可含可不含促效剂)cpm的比值,确定抑制倍数。
在另一个实施例中,可测量转录水平,来评估测试化合物对信号转导的作用。使测试化合物与含有感兴趣的T2R蛋白的宿主细胞接触足够时间,来影响任何相互反应,然后测量基因表达的水平。可凭经验确定影响这种相互反应的时间量,如通过运行一段时间并测量作为时间的转录水平。可用任何本领域技术人员已知的合适方法测量转录量。例如,可用RNA印迹检测感兴趣的蛋白质的mRNA表达,或用免疫试验鉴定其多肽产物。另外,可如美国专利5,436,128描述的那样,用报道基因进行基于转录的试验。报道基因可以是,如氯霉素乙酰转移酶、萤火虫荧光素酶、细菌荧光素酶、β-半乳糖苷酶和碱性磷酸酶。另外,可将感兴趣的蛋白质通过与第二报道子,如绿色荧光蛋白结合,用作间接报道子(见例如,Mistili & Spector,Nature Biotechnology 15:961-964(1997))。
然后将转录量与不存在测试化合物的相同细胞中的转录量作比较,或与缺乏感兴趣的蛋白质的基本相同的细胞中的转录量作比较。基本相同的细胞可衍生自这样的相同细胞,即从它制备了重组细胞但未因引入异源DNA而改变。任何转录量的差异表明测试化合物以某种方式改变了固相蛋白质的活性。
B.调节剂
测试的作为T2R家族成员的调节剂的化合物可以是任何小化合物或生物分子,如蛋白质、糖、核酸或脂类。另外,调节剂可以是T2R基因的遗传学改变形式。通常测试化合物将是小化学分子和肽。实质上任何化合物可被用作本发明试验潜在的调节剂或配体,虽然大多数化合物可溶于水或有机(尤其是基于DMSO的)溶液。设计一种试验,其可通过自动化检验步骤,并为试验提供来源方便的化合物(通常平行进行,如在机器人试验中的微量滴定板上的微量滴定试验),来筛选庞大的化学药品文库。可理解,有许多化学药品供应商,包括Sigma(St.Louis,MO)、Aldrich(St.Louis,MO)、Sigma-Aldrich(St.Louis,MO)、Fluka Chemika-Biochemica Analytika(Buchs Switzerland)等。
在一实施例中,高通量筛选方法涉及提供一种含有许多潜在治疗化合物(可能的调节剂或配体化合物)的组合性化学药品或肽文库。然后在一个或多个本文所述的试验中筛选这些“组合性化学药品文库”或“配体文库”,来鉴定那些显示理想特征活性的文库成员(具体化学药品种类或亚类)。如此鉴定的化合物可作为常规“前导化合物”或本身能作为潜在的或实际的治疗剂。
组合性化学药品文库是由化学合成或生物学合成,通过组合许多化学“建筑构件”如试剂,产生的多样化合物的集合。例如,线性组合性化学药品文库,如多肽文库是通过以各种可能方式针对给定化合物长度(即多肽化合物中的氨基酸数)组合一组化学建筑构件(氨基酸)形成的。可通过这种化学建筑构件的组合性混合合成数以百万计的化合物。
制备和筛选组合性化学药品文库是本领域技术人员熟知的。这些组合性化学药品文库包括但不限于:肽文库(见例如,美国专利5,010,175,Furka,Int.J.Pept.Prot.Res.37:487-493(1991)和Houghton等,Nature 354:84-88(1991))。也可采用其它产生化学多样性文库的化学。这些化学包括但不限于:肽(如PCT出版号WO91/19735)、编码的肽(如PCT出版号WO 93/20242)、随机生物寡聚物(如PCT出版号WO 92/00091)、苯并二吖庚因(如美国专利号5,288,514)、多样体(diversome)如乙内酰脲、苯并二吖庚因和二肽(Hobbs等,Proc.Natl.Acad.Sci.USA 90:6909-6913(1993))、乙烯基类多肽(Hagihara等,Proc.Natl.Acad.Sci.USA 114:6568(1992))、具有葡萄糖骨架的非肽肽模拟物(Hirschmann等,Proc.Natl.Acad.Sci.USA 114:9217-9218(1992))、小化合物文库的类似有机合成(Chan等,Proc.Natl.Acad.Sci.USA 116:2661(1994))、寡氨基甲酸盐(Cho等,Science 261:1303(1993)),和/或肽基磷酸酯(Campbell等,J.Org.Chem.59:658(1994))、核酸文库(见Ausubel Berger和Sambrook,同上)、肽核酸文库(见专利5,539,083)、抗体文库(见例如Vaughn等,Nature Biotechnology,14(3):309-314(1996)和PCT/US96/10287)、糖类文库(见例如Liang等,Science 274:1520-1522(1996)和美国专利5,593,853)、小有机分子文库(见例如苯并二吖庚因,Baum C&EN,an 18,33页(1993);异戊二烯化合物,美国专利5,569,588;噻唑啉酮和间噻唑酮,美国专利5,549,974;吡咯啉酮,美国专利5,525,735和5,519,134;吗啉代化合物,美国专利5,506,337;苯并二吖庚因,5,288,514等)。
制备组合文库的装置是可商业购得的(见例如,357MPS,390MPS,Advanced ChemTech,Louisville KY,Symphony,Rainin,Woburn,MA,433A Applied Biosystems,Foster City,CA,9050 Plus,Millipore,Bedford,MA)。另外,许多组合文库自身是可商业购得的(见例如,ComGenex,Princeton,N.J.Tripos,Inc.,St.Louis,MO,3D Pharmaceuticals,Exton,PA,Martek Biosciences,Columbia,MD,等)。
C.固态和可溶性高通量试验
在一个优选例中本发明提供了的可溶性试验采用分子,如功能域,配体结合功能域、胞外功能域、跨膜功能域(如含有含有7个跨膜区和胞质环),跨膜功能域和胞质功能域、活化位点、亚基结合区等;与异源蛋白质共价连接,形成嵌合分子的功能域;T2R蛋白;表达天然存在或重组T2R蛋白的细胞或组织。在另一个实施例中,本发明提供了基于体外试验的高通量形式的固相,其中功能域、嵌合分子、T2R蛋白或表达T2R的细胞或组织与固相底物结合。
在本发明的高通量试验中,可能在一天中筛选上千种不同的调节剂或配体。具体说,可用微量滴定板的各孔进行针对所选潜在调节剂的独立试验,或如果要观察浓度或培育时间的作用,每5-10个孔可测试一个调节剂。因此,一个标准微量滴定板可检验约100(如96)种调节剂。如果使用1536孔板,则一块板可容易的检验从约100-1500种不同化合物。每天检验几块不同的板是可能的;用本发明的整合系统试验筛选约6000-20,000种不同化合物是可能的。最近,已由例如CaliperTechnologies(Palo alto,CA)开发了试剂操纵的微流体方法。
可将感兴趣的分子与固态组分直接或间接通过共价或非共价键,如通过标记结合。标记可以是不同组分的任一种。一般与标记结合(标记粘合剂)的分子固定于固相载体,而感兴趣的加标记的分子(如感兴趣的味觉转导分子)通过标记和标记粘合剂的相互作用结合于固相载体。
可采用许多标记和标记粘合剂,可根据文献中详述的已知分子的相互作用。例如,当标记是天然粘合剂,如生物素、蛋白A、蛋白G时,可与合适的标记粘合剂联用(亲和素、链霉亲和素、中性亲和素(neotravidin)、免疫球蛋白的Fc区等)。具有天然粘合剂如生物素的分子的抗体也是广泛可得的;合适的标记粘合剂,见SIGMA immunochemicals 1998 catagolue SIGMA,St.Louis MO)。
类似的,可联合合适的抗体使用任何半抗原或抗原化合物形成标记/标记粘合剂对。数以千计的特异性抗体是商品可购得的,而且许多其它抗体在文献中有所描述。例如,在一种常用配置中,标记是第一抗体,而标记结合剂是第二抗体,它识别第一抗体。除了抗体-抗原相互作用外,受体-配体相互作用也适合作为标记和标记-结合剂配对。例如,细胞膜受体促效剂和拮抗剂(如细胞受体-配体相互作用,如转铁蛋白、c-kit、病毒受体配体、细胞因子受体、趋化因子受体、白细胞介素受体、免疫球蛋白受体和抗体、钙粘着蛋自家族、整合素家族、选择素家族等;见如Pigott & Power,The Adhesion Molecule Facts Book I(1993))。类似的,毒素和毒物、病毒表位、激素(如鸦片制剂、类固醇等)、胞内受体(如介导各种小配体,包括:类固醇、甲状腺激素、类视色素和维生素D;肽的作用)、药物、凝集素、糖、核酸(线性和环化聚合物构象的)、寡糖、蛋白质、磷脂和抗体都可与不同细胞受体相互作用。
合成聚合物,如聚氨基甲酸乙酯、聚酯、聚羧酸、聚脲、聚酰胺、聚乙亚胺、聚硫化丙烯、聚硅氧烷、聚亚酰胺和聚乙酸酯也可形成合适的标记或标记结合剂。许多其它标记/标记结合剂配对在本文描述的试验系统中也是有用的,而且对于技术人员参考本公开后将是清楚的。
常用接头如肽、聚酯等也可作为标记,并包括多肽序列,如约5-200个氨基酸之间的聚甘氨酸序列。这些柔性接头对于本领域技术人员来说是已知的。例如,聚(乙二醇)接头可得自Shearwater Polymers,Inc.Huntsville,Alabama。这些接头可任选具有酰胺键、巯基键、或杂官能键。
使用任何现有的不同方法将标记结合剂固定在固相基质上。通过将全部或部分基质与一化学试剂(该试剂能将与标记结合剂的部分反应的化学基团固定到表面上)接触,衍生或官能化固相基质。例如,适合与较长链部分结合的基团包括胺、羟基、巯基和羧基。可用氨烷基硅烷和羟烷基硅烷官能化各种表面,如玻璃表面。这些固相生物聚合物阵列的构建在文献中被详细描述。见例如,Merrifield,J.Am.Chem.Soc.85:2149-2154(1963)(描述了肽的固相合成);Geysen等,J.Immun.Meth.102:259-274(1987)(描述了固相组分在针上的合成);Frank & Doring,Tetrahedron 44:60316040(1988)(描述了不同肽序列在纤维素盘上的合成);Fodor等,Science,251:767-777(1991);Sheldon等,Clinical Chemistry 39(4):718-719(1993);和Kozal等,Nature Medicine 2(7):753759(1996)(全都描述了固定在固相基质上的生物聚合物阵列)。用于将标记结合剂固定在基质上的非化学方法包括其它常用方法,如加热、UV辐照交联等。
D.基于计算机的试验
还有另一种研究调节T2R蛋白活性的化合物的试验涉及计算机辅助药物设计,其中使用计算机系统来产生T2R蛋白的三维结构,根据其氨基酸序列编码的结构信息。输入的氨基酸序列直接和活跃的与计算机程序中预先设定的算法相互作用,得到蛋白质的二级、三级和四级结构模型。然后检测蛋白质结构模型,来鉴定具有结合例如配体的能力的结构区域。然后用这些区域鉴定与该蛋白质结合的配体。
将至少10个氨基酸残基的序列或编码T2R多肽的相应核酸序列输入计算机系统,产生了该蛋白质的三维结构模型。从含有SEQ ID NO:1-165及其保守修饰形式选出编码多肽的核酸或多肽的氨基酸序列。氨基酸序列代表该蛋白质的初级序列或亚序列,其编码该蛋白质的结构信息。用计算机键盘将该氨基酸序列(或编码10个氨基酸的核苷酸序列)的至少10个残基输入计算机系统,计算机可读底物包括但不限于:电子储存媒体(如磁盘、磁带、盒式磁盘和芯片),光学媒介(如CD ROM),因特网站点散布的信息,和RAM。然后使用本领域技术人员已知的软件从氨基酸序列和计算机系统的相互作用产生该蛋白质的三维结构模型。
氨基酸序列代表编码形成感兴趣的蛋白质的二级、三级和四级结构所需的信息的初级结构。软件观察某些一级结构编码的参数来产生结构模型。这些参数被称为“能量条款”,主要包括静电电势、疏水性能、溶剂可接近表面和氢键、二级能量条款包括范德华力。生物分子形成以累计形式使能量条款最小化的结构。因此,计算机程序用这些一级结构或氨基酸序列编码的条件建立二级结构模型。
然后根据二级结构的能量条款,形成二级结构编码的蛋白质的三级结构。在此时用户可输入其它变量,如是否蛋白质是膜结合的或可溶的,它在体内的位置,及其细胞位置,如细胞质、表面或核。用这些变量和二级结构的能量条款形成三级结构的模型。在三级结构建模中,计算机程序使二级结构的疏水表面和类似的匹配,使二级结构的亲水表面和类似的匹配。
一旦结构已产生,用计算机系统鉴定可能的配体结合区域。如上所述输入氨基酸或核苷酸序列,或化合物的化学式,产生可能的配体的三维结构。然后将该可能的配体的三维结构与T2R蛋白的结构比较,来鉴定与蛋白结合的配体。用能量条款确定蛋白质和配体之间的结合亲和力,来确定哪些配体具有增强的结合该蛋白质的可能性。
还可用计算机系统筛选T2R基因的突变、多态变体、等位基因和种间同系物。可将这些突变与疾病情况或基因性状相联系。如上所述,可用GeneChipTM和相关技术筛选突变、多态变体、等位基因和种间同系物。一旦鉴定到了变体,可用诊断试验来鉴定具有这些突变基因的病人。突变的T2R基因的鉴定涉及接受输入编码T2R基因的第一核酸或氨基酸序列(选自SEQ ID NO:1-165的任一及其保守修饰形式)。如上所述将该序列输入计算机系统。然后将第一核酸或氨基酸序列与第二核酸或氨基酸序列(与第一序列基本相同)比较。将第二序列以上述方式输入计算机系统。一旦比较第一和第二序列,鉴定了序列之间的核苷酸或氨基酸差异。这些序列可代表T2R基因中的等位基因差异,和与疾病状态和基因性状有关的突变。
IX.施药和药物组合物
可直接将味觉调节剂施给哺乳动物,在体内调节味道。施药可通过通常用于将调节剂化合物引入,待治疗的组织与其最终接触的任何途径,如舌或口接触途径施药。以任何合适方式,可任选的与药物学上可接受的载体一起施用味觉调节剂。合适的施用这些调节剂的方法是现成的,而且是本领域技术人员熟知的,并且,虽然可使用一种以上途径施用一种具体组合物,但一条特定途径常比其它途径提供更迅速和更有效的反应。
药物学上可接受的载体部分由待施用的具体组合物,以及施用该组合物使用的特定方法所确定。因此,有各式各样本发明药物组合物的合适制剂(见例如,Remington’s Pharmaceutical Sciences,17版,1985)。
味觉调节剂,单独或联合其它合适成分,可制成气雾制剂(即它们可以是“喷雾状”),以通过吸入施用。可将气雾制剂置于压缩的可接受喷雾剂中,如二氯甲烷、丙烷、氮气等。
适合施用的制剂包括水或非水溶剂、等渗无菌溶液,它们可含有抗氧化剂、缓冲液、抑菌剂和使制剂等渗的溶剂,和水和非水无菌悬液,它可包含悬浮剂、增溶剂、增稠剂、稳定剂和防腐剂。在本发明的实施中,可口腔、外用、静脉内、腹膜内、膀胱内或鞘内施用该组合物。可任选的,口腔或鼻内施用该组合物。化合物制剂可置于单剂或多剂密封容器,如安瓿和试管中。可从上述的无菌粉末、颗粒和片剂制备溶剂和悬液。还可作为制备食品或药物的一部分施用调节剂。
本发明产品施给病人的剂量应足够能在一段时间内在病人体内产生有益反应。该剂量将由所用的具体味觉调节剂和病人的情况,以及体重或待治疗区域的表面积决定。剂量大小还将由具体病人施用具体化合物或载体后伴随发生的不良副作用、其性质和程度决定。
在确定要施用的调节剂的有效量时,医师可评估调节剂的循环血浆水平,调节剂的毒性,以及抗-调节剂抗体的产生。一般,相当于调节剂的剂量对于一般个体是约1ng/kg-10mg/kg。
为了施用,本发明的味觉调节剂可以根据该调节剂的LD-50,和抑制剂在不同浓度的副作用(如对病人的质量和总体健康)来确定施药速度。可通过单剂或分剂量完成施药。
VIII.试剂盒
T2R基因及其同系物是鉴定味觉受体细胞,法医学和亲子鉴定,和测定味觉转导的有用工具。用与T2R核酸特异性杂交的T2R家族成员特异性试剂,如T2R探针和引物,和与T2R蛋白特异性结合的T2R抗体等T2R特异性试剂,检测味觉细胞表达和味觉转导调控。
检测样品中T2R家族成员的DNA和RNA的存在的核酸试验包括许多本领域技术人员已知的技术,如DNA分析、RNA分析、斑点印迹、RNase保护、S1分析、扩增技术,如PCR和LCR,和原位杂交。例如在原位杂交中,靶核酸是从其细胞周质中释放出来的,从而能在细胞内杂交,而同时保存细胞形态,利于随后翻译和分析。下列文章提供了对原位杂交技术的纵览:Singer等,Biotechniques 4:230-250(1986);Haase等,Methods in Virology,VII卷,189-226(1984);和NucleicAcid Hybridization:A Prctical Approach(Hames等编,1987)。另外,可用上述各种免疫试验技术检测T2R蛋白。通常将测试样品同时与阳性对照(如表达重组T2R蛋白的样品)和阴性对照比较。
本发明还提供了试剂盒,来筛选T2R家族成员的调节剂。可从易得的材料和试剂制备这些试剂盒。例如,这些试剂盒可包含下列任一或多种材料:T2R核酸、反应试管和测试T2R活性的说明书。可任选的,该试剂盒含有生物学活性T2R受体。可根据本发明,根据需要试剂盒的用户和用户的具体需求,来制备各式各样的试剂盒和组分。
实施例
提供下列实施例仅供说明,而不是为了限制。本领域技术人员将轻易理解,可改变或修改各种非关键性参数,而得到实质上相似的结果。
实施例I:T2R基因家族的鉴定
近来人类遗传连锁研究鉴定出5p15的一个基因座与对苦味物质6-正丙基-2-硫脲嘧啶的反应能力有关(PROP;Reed等,Am.J.Hum.Genet.64:1478-1480(1999))。为了测定PROP敏感性中的差异是否反映了苦味受体中的功能差异,检索DNA序列数据库中在该位置编码候选跨膜蛋白的基因。对跨越该基因间隔区6个测序的人基因组BAC克隆(见例如登录号AC003015)450kbDNA的开放阅读框的分析在5p15.2鉴定出新的GPCR(T2R1)。T2R1具有7个推测的跨膜区段,因为7个保守的残基常存在于GPCR中(Probst等,DNA Cell Biol.11:1-20(1992))。
使用T2R1,并用T2R1相关序列重复的计算机检索揭示了19个额外的人受体(12个全长,7个假基因)。用PCR扩增基因组DNA分离全长hT2R。在低严格性(50-55℃用1X SSC洗涤)下用全长hT2R探测大鼠周缘cDNA文库(Hoon等,Cell,96:541-551(1999))和小鼠BAC滤膜阵列(Genome Systems)。用DNA杂交实验鉴定一组非冗余阳性BAC,并对重叠BAC进行排序。
这些称为T2R(也称为“SF”)的新受体确定了一组新的GPCR家族,他们和V1R犁鼻器受体和视蛋白遥远相关。与属于特征为大N-末端结构域的GPCR超家族的T1R相反,T2R仅具有短胞外N-末端。T2R家族的各成员显示30-70%的氨基酸相同性,大部分在前三个和后三个跨膜区段,以及第二个胞质环中具有高度保守序列基序。T2R之间最趋异的区域是胞外区段,部分延伸入跨膜螺旋中。假定T2R之间的高度可变性反映了识别许多结构多样的配体的需要。与其它许多GPCR一样,T2R不含打断编码区的内含子。
实施例II-人T2R基因的组织
鉴定出的人T2R基因定位在三条染色体上,常常是头-尾顺序排列的。例如,4个受体基因集中在7q31的一个PAC克隆中,9个在12p13的一个BAC克隆中。在这些阵列中可能有更多人T2R,因为部分测序的BAC克隆中发现了7个额外的人T2R,重叠9基因T2R基因簇。在一个给定的阵列内,受体相似性是高度可变的,同时包括较关联(如hT2R13、hT2R14和hT2R15)和较分化的受体(如hT2R3和hT2R4)。在小鼠中监测该类组织(见下文),与对于人、小鼠、苍蝇和蚯蚓的嗅觉受体基因观察到的基因组组织相似(Rouquier等,Nat.Gent.18:243-250(1998);Sullivian等,PNAS 3:884-888(1996);Clyne等,Neuron 22:327-388(1999);Vosshall等,Cell 96:725-736(1999);Troemel等,Cell 83:207-218(1995))。
为了估计该基因大家族大小,检测了各种基因组来源。分析Genome SequenceSurvey数据库(gss)得到12个部分T2R序列。由于该数据库代表约14%人基因组的主要随机采样,该数据提示在人基因组中可能有约90个T2R基因。对于完成和未完成的高通量人基因组序列数据库(htgs)的类似研究产生了36个全长和15个部分T2R序列,这些数据占基因组序列的约50%,还指出基因组中约100个T2R基因。认识由于可得数据库的质量,人基因组中成簇、非随机分散,该分析可能是不准确的,估计T2R家族具有80-120个成员。然而,全长人T2R的1/3以上是假基因;因此,功能性人受体的最终数量可能小得多(即40-80)。这和对于人嗅觉受体观察到的相似;其中许多基因可能是假基因(Rouquier等,Nat.Gent.18:243-250(1998))。
实施例III-T2R基因与苦味有关的基因座有关。
在小鼠中已广泛研究了甜味和苦味品尝的遗传学,其中已用行为性味道选择试验对许多影响对甜味和苦味物质的反应的基因座进行了作图(Warren和Lewis,Nature 227:77-78(1970);Fuller,J.Hered.65:33-66(1974))。小鼠染色体6的远侧末端含有一簇苦味基因,包括Soa(对于蔗糖八乙酸酯;Capeless等,Behav.Genet.22:655-663(1992)),Rua(棉子糖十一乙酸酯,Lush,Gent.Res.47:117-123(1986)),Cyx(环己酰亚胺;Lush和Holland,Genet.Res.52:207-212(1988))和Qui(奎宁;Lush,Genet.Res.44:151-160(1984))。重组研究表明这四个基因座彼此紧密连接,并与Prp(唾液富含脯氨酸蛋白;Azen等,Trends Genet.2:199-200(1986))紧密连接。人的9基因T2R基因簇含有三个交替的PRP基因,对与小鼠染色体6基因簇同源的间隔区作图。
为了限定小鼠染色体6苦味基因簇和T2R之间的关系,分离了T2R基因,测定其基因组组织,物理和基因图定位。通过用人T2R筛选小鼠基因组文库,分离了含61个BAC克隆的28个小鼠T2R。小鼠和人受体显示显著的氨基酸序列趋异性,但都具有与该受体新家族成员相同的序列基序。用小鼠/仓鼠放射杂交板(ResearchGenetics),并通过检测C57BL/6J x DBA/2J重组近交系(Jackson Laboratory)的板中的单核苷酸多态性对小鼠T2R作图。这些研究显示小鼠基因仅在几个基因组座位成簇。含有小鼠T2R的各基因间隔区都与含有其最近的人对应部分的基因同源:mT2R8和hT2R4、mT2R18和hT2R16、和mT2R19和hT2RI。在这三组人/小鼠受体的可能的直向同源配对中,人T2R1和T2R16作图都在与人苦味感受有关的位置(Conneally等,Hum.Hered.26:267-271(1976);Reed等Am.J.Hum.Genet.64:1578-1480(1999))。剩余的25个mT2R都作图在6号染色体的远侧末端,并由3个至少400kb长的BAC重叠群代表。
由于Prp和苦味-基因簇也在小鼠6号染色体的远侧末端,确定它们是否在该T2R阵列中。分析DBA/2x C57BL/6重组近交板,揭示全部3个BAC重叠群内的受体和Prp和苦味基因簇共同分离。另外,分离了小鼠Prp基因(登录号M23236,含D6Milt13),并显示它位于大6号染色体T2R基因簇内。这些结果显示T2R与涉及苦味感受的基因座密切关联。
实施例IV-在味觉受体细胞中表达T2R
舌上皮含有三种乳头的味蕾:舌很后部的周缘乳头、舌后侧边缘的叶状乳头和分散在舌表面前半部的蕈状乳头。口腔的其它部分也具有味蕾;这些在称为geschmackstrifen的上颚表皮内和食道中尤其显著。为了检测T2R表达的模式,用各种味觉乳头的切片进行了原位杂交。为了确保所用的探针在味觉组织中表达,筛选了一个大鼠周缘cDNA文库,分离了14只大鼠T2R cDNA,分别是小鼠基因组克隆的直向同源物。
为了尝试原位杂交,从成年大鼠和小鼠获得了组织。观察到表达模式的非性别特异性差异,因此可互换使用雄性和雌性动物。将新鲜的冷冻切片(16微米)附着在硅烷化载玻片上,如前所述制备原位杂交(Hoon等,Cell 96:541-551(1999))。全部原位杂交在高严格性下进行(杂交,5X SSC,50%甲酰胺,65-72℃,洗涤,0.2X SSC,72℃)。用碱性磷酸酶偶联抗体与毛地黄毒苷以及标准显色底物(BoehringerMannheim)显示信号。如果可能,探针含有延伸的3’-非翻译序列,使在原位杂交的严格性下观察不到的T2R之间的可能交叉杂交最小化。
这些实验证明T2R在舌和上颚表皮的味觉受体细胞亚组中选择性表达。各受体与平均每切片每各味蕾2个细胞杂交。由于这些实验中所用的切片占味蕾深度的1/5-1/3,所以这反映了总的6-10个阳性细胞/味蕾/探针(或味蕾中约15%细胞)。系列切片的检测显示,周缘乳头的所有味蕾含有对于每个探针都是阳性的细胞。迄今,已用11个大鼠T2R和在用17个不同mT2R探针杂交的小鼠切片中观察到相当的结果。
在叶状、geschmackstreifen和食道味蕾中的类似研究证明,各受体探针还标记各味蕾中约15%的细胞。相反,T2R很少在蕈状乳头中表达。用11个不同T2R探针的几百个蕈状味蕾证明,全部蕈状乳头的不到10%含有表达T2R的细胞。有趣的是,几个确实表达T2R的蕈状味蕾往往含有多个阳性细胞。实际上,这些乳头中的阳性细胞数与口腔其它区域的味蕾中所见的无显著不同。另外,含有表达T2R的细胞的蕈状乳头常常是成簇出现的。这个未料到的发现提供了味道编码逻辑的重要线索。已知鼓索神经的一根纤维促动蕈状味蕾中多个细胞,而相同的纤维投射到相邻乳头(Miller,J.Comp.Neurol.158:155-166(1974))。可能T2R阳性味觉受体细胞和蕈状乳头的味蕾的非随机分布反映了类似细胞之间的关联图。
RNA分析和原位杂交证明T2R在味觉组织外表达不广泛。
实施例V-各受体细胞表达多个T2R受体
上述结果证明任何给定的T2R在周缘、叶状和上颚味蕾的约15%细胞中表达。确定在啮齿类基因组中有30种以上的T2R,一个味觉细胞必需表达一个以上的受体。为了确定在任一细胞中表达多少受体和味觉受体细胞的哪一部分表达T2R,将用2、5或10个受体的不同混合物标记的周缘细胞数量与用对应的各探针标记的比较。通过计算多个系列切片中阳性细胞的数量,确定用混合探针(约20%)标记的味觉细胞数量仅比用任何单独受体标记的大一点点(约15%)。不惊奇的是,信号强度在混合探针杂交中显著增加。在来自口腔其它区域,包括蕈状乳头的味蕾中观察到类似的结果。为了直接证明共表达,用一组不同的标记的cPNA探针进行双标记实验。对于双标记荧光检测,用荧光素或毛地黄毒苷标记探针。用碱性磷酸酶偶联的抗荧光素抗体(Amersham)和辣根过氧化物酶偶联的抗-毛地黄毒苷抗体与坚牢红和酪酰胺荧光底物(Boehringer Mannheim和New England Nuclear)联合使用。在这些实验中,发现大多数细胞表达多个受体。
实施例VI-T2R基因在表达gustducin的细胞中选择性表达
之前的结果显示T1R在约30%味觉受体细胞中表达。用不同标记的T1R和T2R探针原位杂交显示在这两类受体的表达中没有重叠。gustducin还在一个大味觉受体细胞亚组中表达,但大部分不和T1R共表达(Hoon等,Cell,96:541-551(1999))。为了确定T2R是否在gustducin细胞中表达,用不同标记的T2R和gustducin核糖探针进行原位杂交。这些实验证明T2R主要在舌和上颚味蕾gustducin阳性的细胞中表达。
周缘、叶状和上颚味蕾中约1/3gustducin细胞不被10种T2R探针的混合物标记,提出不是所有表达gustducin的细胞都表达T2R。这些细胞可以表达其它,可能是更遥远相关的受体,或可以是不同发育阶段的受体。在蕈状味蕾中情况非常不同。由于仅10%蕈状味蕾含有T2R阳性细胞,舌前部的大部分gustducin阳性细胞不共同表达受体的T2R家族成员。因此,似乎蕈状乳头的gustducin阳性细胞中表达另一组受体。
实施例VII-T2R的功能性表达
T2R与Gα15,一种显示与磷脂酶Cβ的大量受体偶联的G蛋白α亚基联合表达(Offermanns和Simon。J.Biol Chem 270:15175-80(1995);Krautwurst等,Cell95:917-926(1998))。在该系统中,受体激活导致胞内钙[Ca2+]i的提高,可在单细胞水平用FURA-2钙-指示剂染料监测(Tsien等,Cell Calcium 6:145-157(1985))。为了测试和优化Gα15偶联,使用了两种不同GPCR(一种是Gαi偶联的μ-阿片样肽受体(Reisine,Neuropharm.34:463-472(1995))和一种Gαq-偶联的mGluR1受体(Masu等,Nature 349:760-765(1991)))。将这些受体转染入HEK-239细胞产生促效剂选择性的、Gα15依赖性的强Ca2+反应(图1)。
许多研究已显示,许多GPCR,特别是感觉受体需要特定的“嵌合体”以成熟和通过分泌途径靶向(Baker等,Embo J 13:4886-4895(1994);Dwyer等,Cell93:455-466(1998))。近来,Krautwurst等(Cell 95:917-926(1998))产生了含有视紫红质前20个氨基酸和各种啮齿类嗅觉受体的嵌合受体。这些靶向质膜,在HEK-293细胞中起到嗅觉受体的作用。为了测定视紫红质序列是否也帮助靶向T2R到质膜,构建了视紫红质-T2R嵌合体(rho-T2R)。这些融合蛋白的表达证明牛视紫红质的前39个氨基酸对将T2R靶向到HEK-293细胞的质膜非常有效(图2)。用11种人和16种啮齿类T2R也获得了类似的结果(见下文)。为了进一步提高T2R表达的水平,将rho-T2R置于强EF-1α启动子的控制下,作为游离质粒引入表达Gα15的修饰的HEK-293细胞中(pEAKrapid细胞)。
用桥重叠PCR延伸技术产生rho-T2R嵌合体,它在阅读框内含有牛视紫红质前39个氨基酸和人与啮齿类T2R编码序列(Mehta和Singh,Biotechnologies26:1082-1086(1999))。所有受体克隆入pEAK10表达载体(Edge Biosystems,MD)。37℃在补充有5%胎牛血清、100微克/毫升庆大霉素硫酸酯(Fisher)、1微克/毫升两性霉素B和2mM GlutamaxI(Lifetechnologies)的UltraCulture培养基(BioWhittaker)中培养和维持修饰的HEK-293细胞(PEAKrapid细胞;Edge Biosystems,MD)。为了转染,将细胞接种在用matrigel包裹的24-孔培养平板或35毫升记录室中。37℃24小时后,用OptiMEM培养液(Lifetechnologies)洗涤细胞,用LipofectAMINE试剂(Lifetechnologies)转染。通过与GFP报道质粒共转染估计转染效率,通常>70%。转染后36-48小时进行免疫荧光染色和活性试验。
对于免疫染色,在包裹的玻璃盖玻片上培养转染细胞,在冰冷的2%聚甲醛中固定20分钟,用1%BSA封阻,并在4℃在含有1∶1000稀释的抗视紫红质mAb B6-30的封阻缓冲液中培育4-6小时(Hargrave等,Exp Eye Res 42:363-373(1986))。用FITC-偶联的驴抗小鼠二抗(Jackson Immunochemical)显示嵌合受体表达。
使用两个平行方法鉴定T2R的配体。一个是选择随机的人、大鼠和小鼠T2R受体组,分别测试对于55种苦味和甜味物质的集合,包括(标出了测试的最大浓度):5mM马兜铃酸、5mM阿托品、5mM番木鳖碱、5mM咖啡酸、10mM咖啡因、1mM氯喹、5mM环己酰亚胺、10mM苯甲酸地那铵、5mM(-)表儿茶精、10mM L-亮氨酸、10mM L-赖氨酸、10mM MgCl2、5mM柚苷。10mM尼古丁、2.5mM盐酸罂粟碱、3mM苯基硫脲、10mM 6-正丙基硫脲嘧啶、1mM奎阿因、1mM盐酸奎宁、800μM棉子糖十一乙酸酯、3mM水杨苷、5mM鹰爪豆碱、5mM硝酸马钱子碱、3mM蔗糖八乙酸酯、2mM盐酸四乙基铵、10mM L-酪氨酸、5mM育亨宾、各10mM的L-甘氨酸、L-丙氨酸、D-色氨酸、L-苯丙氨酸、L-精氨酸、糖精钠、阿斯巴甜、环磺酸钠、乙酰舒泛K、0.1%莫尼糖蛋白、0.1%竹芋蛋白。其它甜味物质是150μM天门冬酰丙氨酸酯、1.8mM卫茅醇、800μM甜叶菊甙、1.9mM cyanosusan、600μM二盐酸新橘皮苷、10mM木糖醇、9.7mM H-Asp-D-Ala-OTMCP、70μM N-Dmb-L-Asp-L-Phe-Ome和12μM N-Dmb-L-Asp-D-Val-(S)-α甲基苯甲酰胺。在这些试验中,基于4个标准评估了功能性偶联:味道选择性、瞬间比活力、和受体与G蛋白依赖性。第二个策略依靠小鼠中苦味感受的遗传学数据将候选受体与特定的味道结合起来。
约30年前,首次报道了小鼠的各种近交系对苦味化合物蔗糖八乙酸酯的敏感性不同(Warren和Lewis,Nature 227:77-78(1970))。随后,许多研究显示该品系的差异是由于一个基因组(Soa)的等位基因变化(Whitney和Harder,Behav Genet16:559-574(1986);Capeless等,Behav Genet 22:655-663(1992))。这些发现延伸到其它影响对各种苦味物质的敏感性的基因座,包括棉子糖十一乙酸酯(Rua)、环己酰亚胺(Cyx)、甘氨酸铜(Glb)和奎宁(Qui)(Lush,Genet.Res.44:151-160(1984);Lush,Genet.Res.47:117-123(1986),Lush和Holland(1988))。基因作图实验显示Soa、Rua、Cyx、Qui和Glb基因座在6号染色体远侧末端成簇(Lush和Holland,Genet.Res 52:207-212(1988);Capeless等,Behav.Genet22:655-663(1992))。根据上述各T2R基因到小鼠中与苦味有关的基因座定位,构建了来自该阵列的T2R基因,对应于rho-mT2R嵌合体,各自转染入表达滥交Gα15蛋白的HEK-293细胞。在细胞中加入FURA-2后,测试对蔗糖八乙酸酯、棉子糖十一乙酸酯、甘氨酸铜、奎宁和环己酰亚胺的反应。
用含有1mM丙酮酸钠和10mM HEPES的Hank’s平衡盐溶液,pH7.4(试验缓冲液)洗涤转染细胞一次,并在室温下加入2μM FURA-2 AM(Molecular Probes)1小时。除去加样溶液,在200微升试验缓冲液中培养1小时,清除AM酯。对于大部分实验,用200微升2x味道物质溶液(见下一节)刺激含有表达一种rho-T2R的细胞的24孔组织培养平板。用装有10x/0.5荧光物镜和TILL显影系统(T.I.L.LPhotonics GmbH)的Nikon Diaphot 200显微镜监测[Ca2+]i。荧光影像的获得和分析使用TILL-Vision软件。通常通过依次在340nm和380nm照射细胞200ms,测量[Ca2+]i80-120秒,并用冷却的CCD照相机监测510纳米的荧光发射。分析F340/F380测量[Ca2+]i。
用浴灌注系统测量激活和灭活的动力学。将细胞接种在150微升微量灌注室中,用picospritzer装置(General Valve,Inc.)压力注射测试溶液。调节流速,确保浴溶液在4-5秒内完全交换。就mT2R5而言,测量了整个照像视野,因为>70%的细胞对环己酰亚胺有反应。对于mT2R8和hT2R4,平均各实验中有感兴趣的100个区域。
表达mT2R5的细胞对环己酰亚胺特异性反应(图3)。反应常常发生在几乎所有转染的细胞中,并且是受体和Gα15依赖性的,因为缺乏这些成分中任一的细胞不能引发[Ca2+]i的变化,甚至在环己酰亚胺浓度高5000倍时也不行。如该偶联系统所预期的,[Ca2+]i中味道引起的增加是由于从内部储藏中释放,因为在正常是0[Ca2+]i输出中获得类似的结果。环己酰亚胺对mT2R5的激活选择性非常强,因为该受体对其它任何味道物质都不反应,甚至在远超过其生物学上可靠的作用范围的浓度下也不反应(Saori,Naturwissenschaften 71:428-9(1984);Glendinning,Behav Neurosci 113:840-854(1994))(图4a,b)。虽然环己酰亚胺对于人仅是微苦的物质,啮齿类却对它强烈厌恶,阈值是约0.25μM(Kusano等,Appl.Exptl.Zool.6:40-50(1971);Lush和Holland,Genet.Res.52:207-212(1988))。在本文所述的基于细胞的试验中,引发mT2R5半最大反应的环己酰亚胺浓度是0.5μM;阈值是约0.2μM(图4c,d)。值得注意的是,该剂量反应与在小鼠中测得的环己酰亚胺的敏感性相同。
为了测定环己酰亚胺反应的动力学,将rho-mT2R5转染的细胞置于微量灌注室中,在各种条件下与测试溶液超混合。细胞显示在施加刺激后(延迟小于1秒)对微摩尔浓度的环己酰亚胺的强瞬时反应。如预期的,当除去味道物质时,[Ca2+]i回到基线。延长对环己酰亚胺的接触(>10秒)导致适应:[Ca2+]i快速提高,然后逐渐但不完全下降到休眠水平(图4a)。类似的,持续施加环己酰亚胺导致显著下降的反应,表明脱敏作用(Lefkowitz等,Cold Spring Harb Symp Quant Biol57:127-133(1992))。这似乎发生于受体水平,由于对照的共转染mGluR1反应在环己酰亚胺脱敏作用期间不改变。
为了确定其它T2R是否也被苦味化合物激活,通过分别转染入表达Gα15的HEK-293细胞测试11种视紫红质-标记的人T2R受体。对苦味和甜味物质库,包括氨基酸、肽和其它天然和合成化合物测试各转染系。这些实验证明强苦味物质地那铵引起被一种人候选味觉受体hT2R4转染的细胞中[Ca2+]i的显著瞬间增加,但在对照的未转染细胞中不增加(图3),或在用其它hT2R转染的细胞中也不增加。地那铵反应具有强剂量依赖性,阈值约100μM。感兴趣的是,hT2R4显示有限范围的滥交性,因为它还对高浓度的苦味物质6-正丙基硫脲嘧啶(PROP)反应(图5)。
如果hT2R4的反应反映了该受体的体内功能,那么假定可在其它物种中发现相似调整的受体。小鼠受体mT2R8是hT2R4可能的直向同源物:它们具有约70%的相同性,而次最接近的受体仅40%相同性;这两个基因包含在同源的基因间隔区内。产生了rho-mT2R8嵌合受体,用各式各样的味道物质检测其反应。事实上,mT2R8和其人相应物一样,被地那铵和高浓度的PROP激活(图3和5)。没有其它味道物质引起表达mT2R8的细胞显著反应。由于这两个受体仅具有70%的相同性,其对于苦味化合物的反应的相似性证实了它们作为直向同源苦味味觉受体的作用。
实施例VIII-非环己酰亚胺品尝者小鼠在mT2R5味觉受体中具有突变
证明mT2R5作为环己酰亚胺的高亲和力受体提示,mT2R5基因可能对应于Cyx基因座。与组织切片原位杂交证明,mT2R5的表达情况在品尝者和非品尝者品系之间是不可分辨的(图6)。为了测定mT2R5和Cyx基因座之间的联系,鉴定了mT2R5的多态性及其在来自C57BL/6J(非品尝者)xDBA/2J(品尝者)的重组近交组杂交。发现在mT2R5和Cyx基因座之间紧密连锁。为了测试mT2R5基因中的突变负责Cyx表型的可能性,从几种额外的特征确定的环己酰亚胺品尝者(CBA/Ca、BALB/c、C3H/He)和非品尝者(129/Sv)品系分离了mT2R5基因,测定其核苷酸序列。事实上,如预期的,如果mT2R5在这些品系中起到环己酰亚胺受体的作用,全部品尝者具有与DBA/2J相同的mT2R5等位基因,而非品尝者具有C57BL/6等位基因,它携带错义突变(图6),包括3个非保守氨基酸取代(T44I、G155D和L294R)。
如果mT2R5C57BL/6等位基因引起Cyx突变体的味觉缺陷,其环己酰亚胺剂量依赖性可能概括了在Cyx突变品系中所见的敏感性漂移。双瓶优势试验显示,Cyx品尝者品系避免了环己酰亚胺,阈值为0.25μM,而非品尝者的敏感性具有约8倍的增加(如,它们在1μM是非品尝者,但在8μM时强烈避开环己酰亚胺)。用来自非品尝者品系的mT2R5基因构建rho-mT2R5融合蛋白,比较其剂量依赖性与品尝者品系的剂量依赖性。明显来自非品尝者品系的mT2R5显示环己酰亚胺敏感性漂移(图4d),代表这些品系对该苦味物质的敏感性。总的说,这些结果证实mT2R5是环己酰亚胺受体,并强烈提示mT2R5对应于Cyx基因座。
实施例IX-与gustducin偶联的T2R
上述证明了T2R与gustducin共表达,提示T2R对苦味物质反应,激活该G-蛋白。为了调查T2R-G蛋白偶联的选择性,选择mT2R5用于研究,因为其被环己酰亚胺的激活概括了口腔的味觉反应。用杆状病毒表达系统制备了rho-标记的mT2R5和gustducin。含mT2R5的膜与各种纯化的G-蛋白,包括gustducin一起培育,并测量味道物质引起的GTP-γS结合(Hoon等,Biochem J.30:629-636(1995))。特别是,用Bac-to-Bac系统(Lifetechnologies,MD)产生含视紫红质的感染性Bacmid触发mT2R5(DBA/2-等位基因)。用重组Bacmid感染昆虫幼虫细胞60小时,如前所述制备膜(Ryba和Tirindelli,J Biol Chem 270:6757-6767(1995))。用8M脲处理除去周围的蛋白,将膜重悬浮在10mM HEPES pH7.5、1mM EDTA和1MM DTT中。用蛋白质印迹,使用mAb B6-30评估rho-mT2R5表达,并用已知量的视紫红质评估。从2×108感染细胞获得300pmol rho-mT2R5。如前所述分离Gustducin和Gβ1γ8异源二聚体(Hoon等,Biochem J 309:629-636(1995);Ryba和Tirindelli,J BiolChem,270:6757-6767(1995))。在10nM rho-mT2R5、100μM GDP和20μM Gβ1γ8存在下,测量了gustducin和其它G-蛋白α亚基上受体催化的GDP交换GTPγS。所有测量在15分钟时间点中进行,并反映了GTPγS初始速率。
这些GTP-γS结合试验揭示了mT2R5对gustducin精细的环己酰亚胺依赖性偶联(图7)。相反,用GαS、Gαi、Gαq或Gαo未发现偶联。在受体、gustducin或βγ-异源二聚物不存在时未发现显著GTPγS结合。T2R5对gustducin的高敏感性,和T2R在含有gustducin的味觉受体细胞中的广泛表达,确定了T2R起到gustducin连接的味觉受体的作用。
本说明书中引用的所有出版物和专利申请在此引入以供参考,如同各单独出版物或专利申请具体和单独指明引入以供参考的那样。
虽然前面的发明已用一些说明性的细节和便于理解的例子进行了描述,本领域普通技术人员将根据本发明的精神容易的懂得,可对本发明作某些改变和修饰,而不违背权利要求的精神和范围。
序列表
<110>C.S.朱克(Zuker,Charles S)
J.E.阿德莱尔(Adler,Jon Elliot)
N.里巴(Ryba,Nick)
K.米勒(Mueller,Ken)
M.胡恩(Hoon,Mark)
加利福尼亚大学董事会(The Regents of the University of Cal ifornia)
美国政府健康及人类服务部(The Government of the United States of America,as
represented by the Secretary,Department of Health and Human Services)
<120>T2R,一类新味觉受体
<130>02307E-098010PC
<140>PCT/US00/24821
<141>2000-09-08
<150>US 09/393,634
<151>1999-09-10
<150>US 09/510,332
<151>2000-02-22
<160>172
<170>PatentIn Ver.2.1
<210>1
<211>299
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R01,hGR01
<400>1
Met Leu Glu Ser His Leu Ile Ile Tyr Phe Leu Leu Ala Val Ile Gln
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Phe Leu Leu Gly Ile Phe Thr Asn Gly Ile Ile Val Val Val Asn Gly
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Ile Asp Leu Ile Lys His Arg Lys Met Ala Pro Leu Asp Leu Leu Leu
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Ser Cys Leu Ala Val Ser Arg Ile Phe Leu Gln Leu Phe Ile Phe Tyr
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Val Asn Val Ile Val Ile Phe Phe Ile Glu Phe Ile Met Cys Ser Ala
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Asn Cys Ala Ile Leu Leu Phe Ile Asn Glu Leu Glu Leu Trp Leu Ala
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Thr Trp Leu Gly Val Phe Tyr Cys Ala Lys Val Ala Ser Val Arg His
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Pro Leu Phe Ile Trp Leu Lys Met Arg Ile Ser Lys Leu Val Pro Trp
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Met Ile Leu Gly Ser Leu Leu Tyr Val Ser Met Ile Cys Val Phe His
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Ser Lys Tyr Ala Gly Phe Met Val Pro Tyr Phe Leu Arg Lys Phe Phe
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Ser Gln Asn Ala Thr Ile Gln Lys Glu Asp Thr Leu Ala Ile Gln Ile
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Phe Ser Phe Val Ala Glu Phe Ser Val Pro Leu Leu Ile Phe Leu Phe
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Arg Asn Thr Val Ala Gly Ser Arg Val Pro Gly Arg Gly Ala Pro Ile
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Ser Ala Leu Leu Ser Ile Leu Ser Phe Leu Ile Leu Tyr Phe Ser His
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Cys Met Ile Lys Val Phe Leu Ser Ser Leu Lys Phe His Ile Arg Arg
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gttttctatt gtgccaaggt tgccagcgtc cgtcacccac tcttcatctg gttgaagatg 360
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tgtgttttcc atagcaaata tgcagggttt atggtcccat acttcctaag gaaatttttc 480
tcccaaaatg ccacaattca aaaagaagat acactggcta tacagatttt ctcttttgtt 540
gctgagttct cagtgccatt gcttatcttc ctttttgctg ttttgctctt gattttctct 600
ctggggaggc acacccggca aatgagaaac acagtggccg gcagcagggt tcctggcagg 660
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tgcatgataa aagtttttct ctcttctcta aagtttcaca tcagaaggtt catctttctg 780
ttcttcatcc ttgtgattgg tatataccct tctggacact ctctcatctt aattttagga 840
aatcctaaat tgaaacaaaa tgcaaaaaag ttcctcctcc acagtaagtg ctgtcagtga 900
<210>3
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<220>
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Met Ala Leu Ser Phe Ser Ala Ile Leu His Ile Ile Met Met Ser Ala
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Cys Asn Glu Leu Ile Lys His Arg Lys Leu Met Pro Ile Gln Ile Leu
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Ile Ser Glu Val Leu Leu Val Asn Leu Ala Leu Ile Phe Pro Leu Ala
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195 200 205
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Phe Ile Ser Tyr Phe Ala Ala Phe Met Thr Asn Met Thr Phe Ser Leu
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<220>
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gggatcacag taaatggatt tcttatcatt gttaactgta atgaattgat caaacataga 120
aagctaatgc caattcaaat cctcttaatg tgcataggga tgtctagatt tggtctgcag 180
atggtgttaa tggtacaaag ttttttctct gtgttctttc cactccttta cgtcaaaata 240
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ctttggttga aattcaggat cccaaagtta ataccttggc tgcttctggg aagcgttctg 420
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aaatgccaac acagaagccc atataaatgc attaaaaaca gtgataacat tcttttgctt 720
ctttatttct tattttgctg ccttcatgac aaatatgaca tttagtttac cttacagaag 780
tcaccagttc tttatgctga aggacataat ggcagcatat ccctctggcc actcggttat 840
aataatcttg agtaattcta agttccaaca atcatttaga agaattctct gcctcaaaaa 900
gaaactatga 910
<210>5
<211>316
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R03,hGR03
<400>5
Met Met Gly Leu Thr Glu Gly Val Phe Leu Ile Leu Ser Gly Thr Gln
1 5 10 15
Phe Thr Leu Gly Ile Leu Val Asn Cys Phe Ile Glu Leu Val Asn Gly
20 25 30
Ser Ser Trp Phe Lys Thr Lys Arg Met Ser Leu Ser Asp Phe Ile Ile
35 40 45
Thr Thr Leu Ala Leu Leu Arg Ile Ile Leu Leu Cys Ile Ile Leu Thr
50 55 60
Asp Ser Phe Leu Ile Glu Phe Ser Pro Asn Thr His Asp Ser Gly Ile
65 70 75 80
Ile Met Gln Ile Ile Asp Val Ser Trp Thr Phe Thr Asn His Leu Ser
85 90 95
Ile Trp Leu Ala Thr Cys Leu Gly Val Leu Tyr Cys Leu Lys Ile Ala
100 105 110
Ser Phe Ser His Pro Thr Phe Leu Trp Leu Lys Trp Arg Val Ser Arg
115 120 125
Val Met Val Trp Met Leu Leu Gly Ala Leu Leu Leu Ser Cys Gly Ser
130 135 140
Thr Ala Ser Leu Ile Asn Glu Phe Lys Leu Tyr Ser Val Phe Arg Gly
145 150 155 160
Ile Glu Ala Thr Arg Asn Val Thr Glu His Phe Arg Lys Lys Arg Ser
165 170 175
Glu Tyr Tyr Leu Ile His Val Leu Gly Thr Leu Trp Tyr Leu Pro Pro
180 185 190
Leu Ile Val Ser Leu Ala Ser Tyr Ser Leu Leu Ile Phe Ser Leu Gly
195 200 205
Arg His Thr Arg Gln Met Leu Gln Asn Gly Thr Ser Ser Arg Asp Pro
210 215 220
Thr Thr Glu Ala His Lys Arg Ala Ile Arg Ile Ile Leu Ser Phe Phe
225 230 235 240
Phe Leu Phe Leu Leu Tyr Phe Leu Ala Phe Leu Ile Ala Ser Phe Gly
245 250 255
Asn Phe Leu Pro Lys Thr Lys Met Ala Lys Met Ile Gly Glu Val Met
260 265 270
Thr Met Phe Tyr Pro Ala Gly His Ser Phe Ile Leu Ile Leu Gly Asn
275 280 285
Ser Lys Leu Lys Gln Thr Phe Val Val Met Leu Arg Cys Glu Ser Gly
290 295 300
His Leu Lys Pro Gly Ser Lys Gly Pro Ile Phe Ser
305 310 315
<210>6
<211>951
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R03,hGR03
<400>6
atgatgggac tcaccgaggg ggtgttcctg attctgtctg gcactcagtt cacactggga 60
attctggtca attgtttcat tgagttggtc aatggtagca gctggttcaa gaccaagaga 120
atgtctttgt ctgacttcat catcaccacc ctggcactct tgaggatcat tctgctgtgt 180
attatcttga ctgatagttt tttaatagaa ttctctccca acacacatga ttcagggata 240
ataatgcaaa ttattgatgt ttcctggaca tttacaaacc atctgagcat ttggcttgcc 300
acctgtcttg gtgtcctcta ctgcctgaaa atcgccagtt tctctcaccc cacattcctc 360
tggctcaagt ggagagtttc tagggtgatg gtatggatgc tgttgggtgc actgctctta 420
tcctgtggta gtaccgcatc tctgatcaat gagtttaagc tctattctgt ctttagggga 480
attgaggcca ccaggaatgt gactgaacac ttcagaaaga agaggagtga gtattatctg 540
atccatgttc ttgggactct gtggtacctg cctcccttaa ttgtgtccct ggcctcctac 600
tctttgctca tcttctccct ggggaggcac acacggcaga tgctgcaaaa tgggacaagc 660
tccagagatc caaccactga ggcccacaag agggccatca gaatcatcct ttccttcttc 720
tttctcttct tactttactt tcttgctttc ttaattgcat catttggtaa tttcctacca 780
aaaaccaaga tggctaagat gattggcgaa gtaatgacaa tgttttatcc tgctggccac 840
tcatttattc tcattctggg gaacagtaag ctgaagcaga catttgtagt gatgctccgg 900
tgtgagtctg gtcatctgaa gcctggatcc aagggaccca ttttctctta g 951
<210>7
<211>299
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R04,hGR04
<400>7
Met Leu Arg Leu Phe Tyr Phe Ser Ala Ile Ile Ala Ser Val Ile Leu
1 5 10 15
Asn Phe Val Gly Ile Ile Met Asn Leu Phe Ile Thr Val Val Asn Cys
20 25 30
Lys Thr Trp Val Lys Ser His Arg Ile Ser Ser Ser Asp Arg Ile Leu
35 40 45
Phe Ser Leu Gly Ile Thr Arg Phe Leu Met Leu Gly Leu Phe Leu Val
50 55 60
Asn Thr Ile Tyr Phe Val Ser Ser Asn Thr Glu Arg Ser Val Tyr Leu
65 70 75 80
Ser Ala Phe Phe Val Leu Cys Phe Met Phe Leu Asp Ser Ser Ser Val
85 90 95
Trp Phe Val Thr Leu Leu Asn Ile Leu Tyr Cys Val Lys Ile Thr Asn
100 105 110
Phe Gln His Ser Val Phe Leu Leu Leu Lys Arg Asn Ile Ser Pro Lys
115 120 125
Ile Pro Arg Leu Leu Leu Ala Cys Val Leu Ile Ser Ala Phe Thr Thr
130 135 140
Cys Leu Tyr Ile Thr Leu Ser Gln Ala Ser Pro Phe Pro Glu Leu Val
145 150 155 160
Thr Thr Arg Asn Asn Thr Ser Phe Asn Ile Ser Glu Gly Ile Leu Ser
165 170 175
Leu Val Val Ser Leu Val Leu Ser Ser Ser Leu Gln Phe Ile Ile Asn
180 185 190
Val Thr Ser Ala Ser Leu Leu Ile His Ser Leu Arg Arg His Ile Gln
195 200 205
Lys Met Gln Lys Asn Ala Thr Gly Phe Trp Asn Pro Gln Thr Glu Ala
210 215 220
His Val Gly Ala Met Lys Leu Met Val Tyr Phe Leu Ile Leu Tyr Ile
225 230 235 240
Pro Tyr Ser Val Ala Thr Leu Val Gln Tyr Leu Pro Phe Tyr Ala Gly
245 250 255
Met Asp Met Gly Thr Lys Ser Ile Cys Leu Ile Phe Ala Thr Leu Tyr
260 265 270
Ser Pro Gly His Ser Val Leu Ile Ile Ile Thr His Pro Lys Leu Lys
275 280 285
Thr Thr Ala Lys Lys Ile Leu Cys Phe Lys Lys
290 295
<210>8
<211>900
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R04,hGR04
<400>8
atgcttcggt tattctattt ctctgctatt attgcctcag ttattttaaa ttttgtagga 60
atcattatga atctgtttat tacagtggtc aattgcaaaa cttgggtcaa aagccataga 120
atctcctctt ctgataggat tctgttcagc ctgggcatca ccaggtttct tatgctggga 180
ctatttctgg tgaacaccat ctacttcgtc tcttcaaata cggaaaggtc agtctacctg 240
tctgcttttt ttgtgttgtg tttcatgttt ttggactcga gcagtgtctg gtttgtgacc 300
ttgctcaata tcttgtactg tgtgaagatt actaacttcc aacactcagt gtttctcctg 360
ctgaagcgga atatctcccc aaagatcccc aggctgctgc tggcctgtgt gctgatttct 420
gctttcacca cttgcctgta catcacgctt agccaggcat caccttttcc tgaacttgtg 480
actacgagaa ataacacatc atttaatatc agtgagggca tcttgtcttt agtggtttct 540
ttggtcttga gctcatctct ccagttcatc attaatgtga cttctgcttc cttgctaata 600
cactccttga ggagacatat acagaagatg cagaaaaatg ccactggttt ctggaatccc 660
cagacggaag ctcatgtagg tgctatgaag ctgatggtct atttcctcat cctctacatt 720
ccatattcag ttgctaccct ggtccagtat ctcccctttt atgcagggat ggatatgggg 780
accaaatcca tttgtctgat ttttgccacc ctttactctc caggacattc tgttctcatt 840
attatcacac atcctaaact gaaaacaaca gcaaagaaga ttctttgttt caaaaaatag 900
<210>9
<211>299
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R05,hGR05
<400>9
Met Leu Ser Ala Gly Leu Gly Leu Leu Met Leu Val Ala Val Val Glu
1 5 10 15
Phe Leu Ile Gly Leu Ile Gly Asn Gly Ser Leu Val Val Trp Ser Phe
20 25 30
Arg Glu Trp Ile Arg Lys Phe Asn Trp Ser Ser Tyr Asn Leu Ile Ile
35 40 45
Leu Gly Leu Ala Gly Cys Arg Phe Leu Leu Gln Trp Leu Ile Ile Leu
50 55 60
Asp Leu Ser Leu Phe Pro Leu Phe Gln Ser Ser Arg Trp Leu Arg Tyr
65 70 75 80
Leu Ser Ile Phe Trp Val Leu Val Ser Gln Ala Ser Leu Trp Phe Ala
85 90 95
Thr Phe Leu Ser Val Phe Tyr Cys Lys Lys Ile Thr Thr Phe Asp Arg
100 105 110
Pro Ala Tyr Leu Trp Leu Lys Gln Arg Ala Tyr Asn Leu Ser Leu Trp
115 120 125
Cys Leu Leu Gly Tyr Phe Ile Ile Asn Leu Leu Leu Thr Val Gln Ile
130 135 140
Gly Leu Thr Phe Tyr His Pro Pro Gln Gly Asn Ser Ser Ile Arg Tyr
145 150 155 160
Pro Phe Glu Ser Trp Gln Tyr Leu Tyr Ala Phe Gln Leu Asn Ser Gly
165 170 175
Ser Tyr Leu Pro Leu Val Val Phe Leu Val Ser Ser Gly Met Leu IIe
180 185 190
Val Ser Leu Tyr Thr His His Lys Lys Met Lys Val His Ser Ala Gly
195 200 205
Arg Arg Asp Val Arg Ala Lys Ala His Ile Thr Ala Leu Lys Ser Leu
210 215 220
Gly Cys Phe Leu Leu Leu His Leu Val Tyr Ile Met Ala Ser Pro Phe
225 230 235 240
Ser Ile Thr Ser Lys Thr Tyr Pro Pro Asp Leu Thr Ser Val Phe Ile
245 250 255
Trp Glu Thr Leu Met Ala Ala Tyr Pro Ser Leu His Ser Leu Ile Leu
260 265 270
Ile Met Gly Ile Pro Arg Val Lys Gln Thr Cys Gln Lys Ile Leu Trp
275 280 285
Lys Thr Val Cys Ala Arg Arg Cys Trp Gly Pro
290 295
<210>10
<211>900
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R05,hGR05
<400>10
atgctgagcg ctggcctagg actgctgatg ctggtggcag tggttgaatt tctcatcggt 60
ttaattggaa atggaagcct ggtggtctgg agttttagag aatggatcag aaaattcaac 120
tggtcctcat ataacctcat tatcctgggc ctggctggct gccgatttct cctgcagtgg 180
ctgatcattt tggacttaag cttgtttcca cttttccaga gcagccgttg gcttcgctat 240
cttagtatct tctgggtcct ggtaagccag gccagcttat ggtttgccac cttcctcagt 300
gtcttctatt gcaagaagat cacgaccttc gatcgcccgg cctacttgtg gctgaagcag 360
agggcctata acctgagtct ctggtgcctt ctgggctact ttataatcaa tttgttactt 420
acagtccaaa ttggcttaac attctatcat cctccccaag gaaacagcag cattcggtat 480
ccctttgaaa gctggcagta cctgtatgca tttcagctca attcaggaag ttatttgcct 540
ttagtggtgt ttcttgtttc ctctgggatg ctgattgtct ctttgtatac acaccacaag 600
aagatgaagg tccattcagc tggtaggagg gatgtccggg ccaaggctca catcactgcg 660
ctgaagtcct tgggctgctt cctcttactt cacctggttt atatcatggc cagccccttc 720
tccatcacct ccaagactta tcctcctgat ctcaccagtg tcttcatctg ggagacactc 780
atggcagcct atccttctct tcattctctc atattgatca tggggattcc tagggtgaag 840
cagacttgtc agaagatcct gtggaagaca gtgtgtgctc ggagatgctg gggcccatga 900
<210>11
<211>287
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R06,hGR06
<220>
<221>MOD_RES
<222>(1)..(287)
<223>Xaa=任何氨基酸
<400>11
Met Leu Ala Ala Ala Leu Gly Leu Leu Met Pro Ile Ala Gly Ala Glu
1 5 10 15
Phe Leu Ile Gly Leu Val Gly Asn Gly Val Pro Val Val Cys Ser Phe
20 25 30
Arg Gly Trp Val Lys Lys Met Xaa Gly Val Pro Ile Asn Ser His Asp
35 40 45
Ser Gly Lys Xaa Pro Leu Ser Pro Thr Gln Ala Asp His Val Gly His
50 55 60
Lys Ser Val Ser Thr Phe Pro Glu Gln Trp Leu Ala Leu Leu Ser Xaa
65 70 75 80
Cys Leu Arg Val Leu Val Ser Gln Ala Asn Met Xaa Phe Ala Thr Phe
85 90 95
Phe Ser Gly Phe Cys Cys Met Glu Ile Met Thr Phe Val Xaa Xaa Xaa
100 105 110
Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
115 120 125
Xaa Leu Leu Val Ser Phe Lys Ile Thr Phe Tyr Phe Ser Ala Leu Val
130 135 140
Gly Trp Thr Leu Xaa Lys Pro Leu Thr Gly Asn Ser Asn Ile Leu His
145 150 155 160
Pro Ile Leu Asn Leu Leu Phe Leu Xaa Ile Ala Val Gln Xaa Arg Arg
165 170 175
Leu Ile Ala Ile Cys Asp Val Ser Val Pro Leu Val Phe Leu Xaa Arg
180 185 190
His His Arg Lys Met Glu Asp His Thr Ala Val Arg Arg Arg Leu Lys
195 200 205
Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
210 215 220
Xaa Leu Tyr Met Val Ser Ala Leu Ala Arg His Phe Ser Met Thr Phe
225 230 235 240
Xaa Ser Pro Ser Asp Leu Thr Ile Leu Ala Ile Ser Ala Thr Leu Met
245 250 255
Ala Val Tyr Thr Ser Phe Pro Ser Ile Val Met Val Met Arg Asn Gln
260 265 270
Thr Cys Gln Arg Ile Leu Xaa Glu Met Ile Cys Thr Trp Lys Ser
275 280 285
<210>12
<211>823
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R06,hGR06
<400>12
atgttggcgg ctgccctagg attgctgatg cccattgcag gggctgaatt tctcattggc 60
ctggttggaa atggagtccc tgtggtctgc agttttagag gatgggtcaa aaaaatgtaa 120
ggagtcccta taaattctca tgattctggt aagtagccac tttctcctac tcaggccgat 180
catgttggac ataagtctgt ttccactttc ccagagcagt ggttggcttt actatcttaa 240
tgtcttcgag tcctggtaag ccaggccaac atgtagtttg ccactttctt cagtggcttc 300
tgctgcatgg agatcatgac ctttgtcccg ctgacttctt gtagctgaaa agactgggtt 360
tttgtttttt gctagtgtct ttcaagatca ctttttattt ctcagctctt gttggctgga 420
ccctttaaaa acccttaaca ggaaacagca acatcctgca tcccatttta aatctgttat 480
ttttatagat tgctgtccag tgaaggagac tgattgctat ttgtgatgtt tctgttccac 540
ttgtcttttt gtaaagacat cacaggaaga tggaggacca cacagctgtc aggaggaggc 600
tcaaaccaag gtgctcatcg ctctgaactt ccccctttac atggtttctg ccttggccag 660
acacttttcc atgaccttct aatctccctc tgatctcacc attcttgcca tctctgcaac 720
actcatggct gtttatactt catttccgtc tattgtaatg gttatgagga atcagacttg 780
tcagagaatt ctgtaggaga tgatatgtac atggaaatcc tag 823
<210>13
<211>318
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R07,hGR07
<400>13
Met Ala Asp Lys Val Gln Thr Thr Leu Leu Phe Leu Ala Val Gly Glu
1 5 10 15
Phe Ser Val Gly Ile Leu Gly Asn Ala Phe Ile Gly Leu Val Asn Cys
20 25 30
Met Asp Trp Val Lys Lys Arg Lys Ile Ala Ser Ile Asp Leu Ile Leu
35 40 45
Thr Ser Leu Ala Ile Ser Arg Ile Cys Leu Leu Cys Val Ile Leu Leu
50 55 60
Asp Cys Phe Ile Leu Val Leu Tyr Pro Asp Val Tyr Ala Thr Gly Lys
65 70 75 80
Glu Met Arg Ile Ile Asp Phe Phe Trp Thr Leu Thr Asn His Leu Ser
85 90 95
Ile Trp Phe Ala Thr Cys Leu Ser Ile Tyr Tyr Phe Phe Lys Ile Gly
100 105 110
Asn Phe Phe His Pro Leu Phe Leu Trp Met Lys Trp Arg Ile Asp Arg
115 120 125
Val Ile Ser Trp Ile Leu Leu Gly Cys Val Val Leu Ser Val Phe Ile
130 135 140
Ser Leu Pro Ala Thr Glu Asn Leu Asn Ala Asp Phe Arg Phe Cys Val
145 150 155 160
Lys Ala Lys Arg Lys Thr Asn Leu Thr Trp Ser Cys Arg Val Asn Lys
165 170 175
Thr Gln His Ala Ser Thr Lys Leu Phe Leu Asn Leu Ala Thr Leu Leu
180 185 190
Pro Phe Cys Val Cys Leu Met Ser Phe Phe Leu Leu Ile Leu Ser Leu
195 200 205
Arg Arg His Ile Arg Arg Met Gln Leu Ser Ala Thr Gly Cys Arg Asp
210 215 220
Pro Ser Thr Glu Ala His Val Arg Ala Leu Lys Ala Val Ile Ser Phe
225 230 235 240
Leu Leu Leu Phe Ile Ala Tyr Tyr Leu Ser Phe Leu Ile Ala Thr Ser
245 250 255
Ser Tyr Phe Met Pro Glu Thr Glu Leu Ala Val Ile Phe Gly Glu Ser
260 265 270
Ile Ala Leu Ile Tyr Pro Ser Ser His Ser Phe Ile Leu Ile Leu Gly
275 280 285
Asn Asn Lys Leu Arg His Ala Ser Leu Lys Val Ile Trp Lys Val Met
290 295 300
Ser Ile Leu Lys Gly Arg Lys Phe Gln Gln His Lys Gln Ile
305 310 315
<210>14
<211>957
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R07,hGR07
<400>14
atggcagata aagtgcagac tactttattg ttcttagcag ttggagagtt ttcagtgggg 60
atcttaggga atgcattcat tggattggta aactgcatgg actgggtcaa gaagaggaaa 120
attgcctcca ttgatttaat cctcacaagt ctggccatat ccagaatttg tctattgtgc 180
gtaatactat tagattgttt tatattggtg ctatatccag atgtctatgc cactggtaaa 240
gaaatgagaa tcattgactt cttctggaca ctaaccaatc atttaagtat ctggtttgca 300
acctgcctca gcatttacta tttcttcaag ataggtaatt tctttcaccc acttttcctc 360
tggatgaagt ggagaattga cagggtgatt tcctggattc tactggggtg cgtggttctc 420
tctgtgttta ttagccttcc agccactgag aatttgaacg ctgatttcag gttttgtgtg 480
aaggcaaaga ggaaaacaaa cttaacttgg agttgcagag taaataaaac tcaacatgct 540
tctaccaagt tatttctcaa cctggcaacg ctgctcccct tttgtgtgtg cctaatgtcc 600
tttttcctct tgatcctctc cctgcggaga catatcaggc gaatgcagct cagtgccaca 660
gggtgcagag accccagcac agaagcccat gtgagagccc tgaaagctgt catttccttc 720
cttctcctct ttattgccta ctatttgtcc tttctcattg ccacctccag ctactttatg 780
ccagagacgg aattagctgt gatttttggt gagtccatag ctctaatcta cccctcaagt 840
cattcattta tcctaatact ggggaacaat aaattaagac atgcatctct aaaggtgatt 900
tggaaagtaa tgtctattct aaaaggaaga aaattccaac aacataaaca aatctga 957
<210>15
<211>309
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R08,hGR08
<400>15
Met Phe Ser Pro Ala Asp Asn Ile Phe Ile Ile Leu Ile Thr Gly Glu
1 5 10 15
Phe Ile Leu Gly Ile Leu Gly Asn Gly Tyr Ile Ala Leu Val Asn Trp
20 25 30
Ile Asp Trp Ile Lys Lys Lys Lys Ile Ser Thr Val Asp Tyr Ile Leu
35 40 45
Thr Asn Leu Val Ile Ala Arg Ile Cys Leu Ile Ser Val Met Val Val
50 55 60
Asn Gly Ile Val Ile Val Leu Asn Pro Asp Val Tyr Thr Lys Asn Lys
65 70 75 80
Gln Gln Ile Val Ile Phe Thr Phe Trp Thr Phe Ala Asn Tyr Leu Asn
85 90 95
Met Trp Ile Thr Thr Cys Leu Asn Val Phe Tyr Phe Leu Lys Ile Ala
100 105 110
Ser Ser Ser His Pro Leu Phe Leu Trp Leu Lys Trp Lys Ile Asp Met
115 120 125
Val Val His Trp Ile Leu Leu Gly Cys Phe Ala Ile Ser Leu Leu Val
130 135 140
Ser Leu Ile Ala Ala Ile Val Leu Ser Cys Asp Tyr Arg Phe His Ala
145 150 155 160
Ile Ala Lys His Lys Arg Asn Ile Thr Glu Met Phe His Val Ser Lys
165 170 175
Ile Pro Tyr Phe Glu Pro Leu Thr Leu Phe Asn Leu Phe Ala Ile Val
180 185 190
Pro Phe Ile Val Ser Leu Ile Ser Phe Phe Leu Leu Val Arg Ser Leu
195 200 205
Trp Arg His Thr Lys Gln Ile Lys Leu Tyr Ala Thr Gly Ser Arg Asp
210 215 220
Pro Ser Thr Glu Val His Val Arg Ala Ile Lys Thr Met Thr Ser Phe
225 230 235 240
Ile Phe Phe Phe Phe Leu Tyr TyrIle Ser Ser Ile Leu Met Thr Phe
245 250 255
Ser Tyr Leu Met Thr Lys Tyr Lys Leu Ala Val Glu Phe Gly Glu Ile
260 265 270
Ala Ala Ile Leu Tyr Pro Leu Gly His Ser Leu Ile Leu Ile Val Leu
275 280 285
Asn Asn Lys Leu Arg Gln Thr Phe Val Arg Met Leu Thr Cys Arg Lys
290 295 300
Ile Ala Cys Met Ile
305
<210>16
<211>930
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R08,hGR08
<400>16
atgttcagtc ctgcagataa catctttata atcctaataa ctggagaatt catactagga 60
atattgggga atggatacat tgcactagtc aactggattg actggattaa gaagaaaaag 120
atttccacag ttgactacat ccttaccaat ttagttatcg ccagaatttg tttgatcagt 180
gtaatggttg taaatggcat tgtaatagta ctgaacccag atgtttatac aaaaaataaa 240
caacagatag tcatttttac cttctggaca tttgccaact acttaaatat gtggattacc 300
acctgcctta atgtcttcta ttttctgaag atagccagtt cctctcatcc actttttctc 360
tggctgaagt ggaaaattga tatggtggtg cactggatcc tgctgggatg ctttgccatt 420
tccttgttgg tcagccttat agcagcaata gtactgagtt gtgattatag gtttcatgca 480
attgccaaac ataaaagaaa cattactgaa atgttccatg tgagtaaaat accatacttt 540
gaacccttga ctctctttaa cctgtttgca attgtcccat ttattgtgtc actgatatca 600
tttttccttt tagtaagatc tttatggaga cataccaagc aaataaaact ctatgctacc 660
ggcagtagag accccagcac agaagttcat gtgagagcca ttaaaactat gacttcattt 720
atcttctttt ttttcctata ctatatttct tctattttga tgacctttag ctatcttatg 780
acaaaataca agttagctgt ggagtttgga gagattgcag caattctcta ccccttgggt 840
cactcactta ttttaattgt tttaaataat aaactgaggc agacatttgt cagaatgctg 900
acatgtagaa aaattgcctg catgatatga 930
<210>17
<211>312
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R09,hGR09
<400>17
Met Pro Ser Ala Ile Glu Ala Ile Tyr Ile Ile Leu Ile Ala Gly Glu
1 5 10 15
Leu Thr Ile Gly Ile Trp Gly Asn Gly Phe Ile Val Leu Val Asn Cys
20 25 30
Ile Asp Trp Leu Lys Arg Arg Asp Ile Ser Leu Ile Asp Ile Ile Leu
35 40 45
Ile Ser Leu Ala Ile Ser Arg Ile Cys Leu Leu Cys Val Ile Ser Leu
50 55 60
Asp Gly Phe Phe Met Leu Leu Phe Pro Gly Thr Tyr Gly Asn Ser Val
65 70 75 80
Leu Val Ser Ile Val Asn Val Val Trp Thr Phe Ala Asn Asn Ser Ser
85 90 95
Leu Trp Phe Thr Ser Cys Leu Ser Ile Phe Tyr Leu Leu Lys Ile Ala
100 105 110
Asn Ile Ser His Pro Phe Phe Phe Trp Leu Lys Leu Lys Ile Asn Lys
115 120 125
Val Met Leu Ala Ile Leu Leu Gly Ser Phe Leu Ile Ser Leu Ile Ile
130 135 140
Ser Val Pro Lys Asn Asp Asp Met Trp Tyr His Leu Phe Lys Val Ser
145 150 155 160
His Glu Glu Asn Ile Thr Trp Lys Phe Lys Val Ser Lys Ile Pro Gly
165 170 175
Thr Phe Lys Gln Leu Thr Leu Asn Leu Gly Val Met Val Pro Phe Ile
180 185 190
Leu Cys Leu Ile Ser Phe Phe Leu Leu Leu Phe Ser Leu Val Arg His
195 200 205
Thr Lys Gln Ile Arg Leu His Ala Thr Gly Phe Arg Asp Pro Ser Thr
210 215 220
Glu Ala His Met Arg Ala Ile Lys Ala Val Ile Ile Phe Leu Leu Leu
225 230 235 240
Leu Ile Val Tyr Tyr Pro Val Phe Leu Val Met Thr Ser Ser Ala Leu
245 250 255
Ile Pro Gln Gly Lys Leu Val Leu Met Ile Gly Asp Ile Val Thr Val
260 265 270
Ile Phe Pro Ser Ser His Ser Phe Ile Leu Ile Met Gly Asn Ser Lys
275 280 285
Leu Arg Glu Ala Phe Leu Lys Met Leu Arg Phe Val Lys Cys Phe Leu
290 295 300
Arg Arg Arg Lys Pro Phe Val Pro
305 310
<210>18
<211>939
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R09,hGR09
<400>18
atgccaagtg caatagaggc aatatatatt attttaattg ctggtgaatt gaccataggg 60
atttggggaa atggattcat tgtactagtt aactgcattg actggctcaa aagaagagat 120
atttccttga ttgacatcat cctgatcagc ttggccatct ccagaatctg tctgctgtgt 180
gtaatatcat tagatggctt ctttatgctg ctctttccag gtacatatgg caatagcgtg 240
ctagtaagca ttgtgaatgt tgtctggaca tttgccaata attcaagtct ctggtttact 300
tcttgcctca gtatcttcta tttactcaag atagccaata tatcgcaccc atttttcttc 360
tggctgaagc taaagatcaa caaggtcatg cttgcgattc ttctggggtc ctttcttatc 420
tctttaatta ttagtgttcc aaagaatgat gatatgtggt atcacctttt caaagtcagt 480
catgaagaaa acattacttg gaaattcaaa gtgagtaaaa ttccaggtac tttcaaacag 540
ttaaccctga acctgggggt gatggttccc tttatccttt gcctgatctc atttttcttg 600
ttacttttct ccctagttag acacaccaag cagattcgac tgcatgctac agggttcaga 660
gaccccagta cagaggccca catgagggcc ataaaggcag tgatcatctt tctgctcctc 720
ctcatcgtgt actacccagt ctttcttgtt atgacctcta gcgctctgat tcctcaggga 780
aaattagtgt tgatgattgg tgacatagta actgtcattt tcccatcaag ccattcattc 840
attctaatta tgggaaatag caagttgagg gaagcttttc tgaagatgtt aagatttgtg 900
aagtgtttcc ttagaagaag aaagcctttt gttccatag 939
<210>19
<211>307
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R10,hGR10
<400>19
Met Leu Arg Val Val Glu Gly Ile Phe Ile Phe Val Val Val Ser Glu
1 5 10 15
Ser Val Phe Gly Val Leu Gly Asn Gly Phe Ile Gly Leu Val Asn Cys
20 25 30
Ile Asp Cys Ala Lys Asn Lys Leu Ser Thr Ile Gly Phe Ile Leu Thr
35 40 45
Gly Leu Ala Ile Ser Arg Ile Phe Leu Ile Trp Ile Ile Ile Thr Asp
50 55 60
Gly Phe Ile Gln Ile Phe Ser Pro Asn Ile Tyr Ala Ser Gly Asn Leu
65 70 75 80
Ile Glu Tyr Ile Ser Tyr Phe Trp Val Ile Gly Asn Gln Ser Ser Met
85 90 95
Trp Phe Ala Thr Ser Leu Ser Ile Phe Tyr Phe Leu Lys Ile Ala Asn
100 105 110
Phe Ser Asn Tyr Ile Phe Leu Trp Leu Lys Ser Arg Thr Asn Met Val
115 120 125
Leu Pro Phe Met Ile Val Phe Leu Leu Ile Ser Ser Leu Leu Asn Phe
130 135 140
Ala Tyr Ile Ala Lys Ile Leu Asn Asp Tyr Lys Thr Lys Asn Asp Thr
145 150 155 160
Val Trp Asp Leu Asn Met Tyr Lys Ser Glu Tyr Phe Ile Lys Gln Ile
165 170 175
Leu Leu Asn Leu Gly Val Ile Phe Phe Phe Thr Leu Ser Leu Ile Thr
180 185 190
Cys Ile Phe Leu Ile Ile Ser Leu Trp Arg His Asn Arg Gln Met Gln
195 200 205
Ser Asn Val Thr Gly Leu Arg Asp Ser Asn Thr Glu Ala His Val Lys
210 215 220
Ala Met Lys Val Leu Ile Ser Phe Ile Ile Leu Phe Ile Leu Tyr Phe
225 230 235 240
Ile Gly Met Ala Ile Glu Ile Ser Cys Phe Thr Val Arg Glu Asn Lys
245 250 255
Leu Leu Leu Met Phe Gly Met Thr Thr Thr Ala Ile Tyr Pro Trp Gly
260 265 270
His Ser Phe Ile Leu Ile Leu Gly Asn Ser Lys Leu Lys Gln Ala Ser
275 280 285
Leu Arg Val Leu Gln Gln Leu Lys Cys Cys Glu Lys Arg Lys Asn Leu
290 295 300
Arg Val Thr
305
<210>20
<211>924
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R10,hGR10
<400>20
atgctacgtg tagtggaagg catcttcatt tttgttgtag ttagtgagtc agtgtttggg 60
gttttgggga atggatttat tggacttgta aactgcattg actgtgccaa gaataagtta 120
tctacgattg gctttattct caccggctta gctatttcaa gaatttttct gatatggata 180
ataattacag atggatttat acagatattc tctccaaata tatatgcctc cggtaaccta 240
attgaatata ttagttactt ttgggtaatt ggtaatcaat caagtatgtg gtttgccacc 300
agcctcagca tcttctattt cctgaagata gcaaattttt ccaactacat atttctctgg 360
ttgaagagca gaacaaatat ggttcttccc ttcatgatag tattcttact tatttcatcg 420
ttacttaatt ttgcatacat tgcgaagatt cttaatgatt ataaaacgaa gaatgacaca 480
gtctgggatc tcaacatgta taaaagtgaa tactttatta aacagatttt gctaaatctg 540
ggagtcattt tcttctttac actatcccta attacatgta tttttttaat catttccctt 600
tggagacaca acaggcagat gcaatcgaat gtgacaggat tgagagactc caacacagaa 660
gctcatgtga aggcaatgaa agttttgata tctttcatca tcctctttat cttgtatttt 720
ataggcatgg ccatagaaat atcatgtttt actgtgcgag aaaacaaact gctgcttatg 780
tttggaatga caaccacagc catctatccc tggggtcact catttatctt aattctagga 840
aacagcaagc taaagcaagc ctctttgagg gtactgcagc aattgaagtg ctgtgagaaa 900
aggaaaaatc tcagagtcac atag 924
<210>21
<211>245
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R11,hGR11
<220>
<221>MOD_RES
<222>(1)..(245)
<223>Xaa=任何氨基酸
<400>21
Met Ala Asn Met Leu Lys Asn Met Leu Thr Met Ile Ser Ala Ile Asp
1 5 10 15
Phe Ile Met Gly Ile Gln Arg Ser Arg Val Met Val Leu Val His Cys
20 25 30
Ile Asp Trp Ile Arg Arg Trp Lys Leu Ser Leu Ile Asp Phe Ile Leu
35 40 45
Thr Cys Trp Ala Ile Ser Arg Ile Phe Xaa Xaa Xaa Xaa Xaa Xaa Xaa
50 55 60
Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
65 70 75 80
Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Asn His Leu Cys Thr Xaa Phe
85 90 95
Ala Thr Cys Leu Ala Val Phe Tyr Phe Leu Lys Ile Val Asn Phe Ser
100 105 110
Tyr Leu Phe Tyr Phe Trp Leu Lys Trp Arg Ile Asn Lys Val Ala Phe
115 120 125
Ile Leu Pro Leu Val Ser Ala Phe Ser Val Tyr Gln Leu Ser Phe Asp
130 135 140
Val His Phe Xaa Cys Leu Leu Val Ser Cys Pro Lys Lys Tyr Glu Arg
145 150 155 160
His Met Thr Gly Leu Leu Asn Val Ser Asn Asn Lys Asn Val Asn Asn
165 170 175
Ile Ile Ile Phe Phe Ile Gly Ser Leu Ser Ser Phe Ser Ile Ser Ser
180 185 190
Ile Phe Phe Leu Leu Leu Leu Leu Ser Ser Xaa Arg His Met Lys His
195 200 205
Ile Arg Phe Asn Phe Arg Asp Cys Arg Thr Pro Val Tyr Gly Pro Ile
210 215 220
Ser Glu Pro Arg Lys Arg Phe Ser Phe Phe Val Leu Leu Leu Tyr Lys
225 230 235 240
Asn Leu Pro Phe Ser
245
<210>22
<211>315
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R12,hGR12
<220>
<221>MOD_RES
<222>(1)..(315)
<223>Xaa=任何氨基酸
<400>22
Met Ser Ser Ile Trp Glu Thr Leu Phe Ile Arg Ile Leu Val Val Xaa
1 5 10 15
Phe Ile Met Gly Thr Val Gly Asn Xaa Phe Ile Val Leu Val Asn Ile
20 25 30
Ile Asp Xaa Ile Arg Asn Xaa Lys Val Ser Leu Ile Asp Phe Ile Leu
35 40 45
Asn Cys Leu Ala Ile Ser Arg Ile Cys Phe Leu Xaa Ile Thr Ile Leu
50 55 60
Ala Thr Ser Phe Asn Ile Gly Tyr Glu Lys Met Pro Asp Ser Lys Asn
65 70 75 80
Leu Ala Val Ser Phe Asp Ile Leu Trp Thr Gly Ser Ser Tyr Phe Cys
85 90 95
Leu Ser Cys Thr Thr Cys Leu Ser Val Phe Tyr Phe Leu Lys Val Ala
100 105 110
Asn Phe Ser Asn Pro Ile Phe Leu Trp Met Lys Trp Lys Ile His Lys
115 120 125
Val Leu Leu Phe Ile Val Leu Glu Ala Thr Ile Ser Phe Cys Thr Thr
130 135 140
Ser Ile Leu Lys Glu Ile Ile Ile Asn Ser Leu Ile Xaa Glu Arg Val
145 150 155 160
Thr Ile Lys Gly Asn Leu Thr Phe Asn Tyr Met Asp Thr Met His Asp
165 170 175
Phe Thr Ser Leu Phe Leu Leu Gln Met Met Phe Ile Leu Pro Phe Val
180 185 190
Glu Thr Leu Ala Ser Ile Leu Leu Leu Ile Leu Ser Leu Trp Ser His
195 200 205
Thr Arg Gln Met Lys Leu His Gly Ile Tyr Ser Arg Asp Pro Ser Thr
210 215 220
Glu Ala His Val Lys Pro Ile Lys Ala Ile Ile Ser Phe Leu Leu Leu
225 230 235 240
Phe Ile Val His Tyr Phe Ile Ser Ile Ile Leu Thr Leu Ala Cys Pro
245 250 255
Leu Leu Asp Phe Val Ala Ala Arg Thr Phe Ser Ser Val Leu Val Phe
260 265 270
Phe His Pro Ser Gly His Ser Phe Leu Leu Ile Leu Arg Asp Ser Lys
275 280 285
Leu Lys Gln Ala Ser Leu Cys Val Leu Lys Lys Met Lys Tyr Ala Lys
290 295 300
Lys Asp Ile Ile Ser His Phe Tyr Lys His Ala
305 310 315
<210>23
<211>948
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R12,hGR12
<400>23
atgtcaagca tttgggagac actgtttata agaattcttg tagtgtaatt cataatgggg 60
actgtgggaa attgattcat tgtattggtt aatatcattg actgaatcag gaactgaaag 120
gtctccctga ttgattttat tctcaactgc ttggccatct ccaggatatg tttcctgtag 180
ataacaattt tagctacctc tttcaatata ggctatgaga aaatgcctga ttctaagaat 240
cttgcagtaa gttttgacat tctctggaca ggatccagct atttctgcct gtcctgtacc 300
acttgcctca gtgtcttcta tttcctcaag gtagccaact tctccaatcc cattttcctc 360
tggatgaaat ggaaaattca caaggtgctt ctctttattg tactagaggc aacgatctct 420
ttctgcacaa cttccattct gaaggaaata ataattaata gtttaatcta agaacgggta 480
acaataaaag gcaacttgac atttaattat atggatacca tgcatgattt cacttctctg 540
tttctccttc agatgatgtt catccttcct tttgtggaaa cactggcttc cattcttctc 600
ttaatcctct ccttatggag ccacaccagg cagatgaagc tacatggtat ttattccagg 660
gatcccagca cagaagccca tgtaaaacct ataaaagcta taatttcatt tctactcctc 720
tttattgtgc attatttcat cagtatcata ctaacattgg cctgtcctct tctagacttc 780
gttgcggcaa ggacttttag tagtgtgctg gtatttttcc atccatctgg ccattcattt 840
cttctaattt tacgggacag caaactgaag caagcttctc tctgtgtcct gaagaagatg 900
aagtatgcca aaaaggacat aatctctcat ttttataaac atgcctga 948
<210>24
<211>303
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R13,hGR13
<400>24
Met Glu Ser Ala Leu Pro Ser Ile Phe Thr Leu Val Ile Ile Ala Glu
1 5 10 15
Phe Ile Ile Gly Asn Leu Ser Asn Gly Phe Ile Val Leu Ile Asn Cys
20 25 30
Ile Asp Trp Val Ser Lys Arg Glu Leu Ser Ser Val Asp Lys Leu Leu
35 40 45
Ile Ile Leu Ala Ile Ser Arg Ile Gly Leu Ile Trp Glu Ile Leu Val
50 55 60
Ser Trp Phe Leu Ala Leu His Tyr Leu Ala Ile Phe Val Ser Gly Thr
65 70 75 80
Gly Leu Arg Ile Met Ile Phe Ser Trp Ile Val Ser Asn His Phe Asn
85 90 95
Leu Trp Leu Ala Thr Ile Phe Ser Ile Phe Tyr Leu Leu Lys Ile Ala
100 105 110
Ser Phe Ser Ser Pro Ala Phe Leu Tyr Leu Lys Trp Arg Val Asn Lys
115 120 125
Val Ile Leu Met Ile Leu Leu Gly Thr Leu Val Phe Leu Phe Leu Asn
130 135 140
Leu Ile Gln Ile Asn Met His Ile Lys Asp Trp Leu Asp Arg Tyr Glu
145 150 155 160
Arg Asn Thr Thr Trp Asn Phe Ser Met Ser Asp Phe Glu Thr Phe Ser
165 170 175
Val Ser Val Lys Phe Thr Met Thr Met Phe Ser Leu Thr Pro Phe Thr
180 185 190
Val Ala Phe Ile Ser Phe Leu Leu Leu Ile Phe Ser Leu Gln Lys His
195 200 205
Leu Gln Lys Met Gln Leu Asn Tyr Lys Gly His Arg Asp Pro Arg Thr
210 215 220
Lys Val His Thr Asn Ala Leu Lys Ile Val Ile Ser Phe Leu Leu Phe
225 230 235 240
Tyr Ala Ser Phe Phe Leu Cys Val Leu Ile Ser Trp Ile Ser Glu Leu
245 250 255
Tyr Gln Asn Thr Val Ile Tyr Met Leu Cys Glu Thr Ile Gly Val Phe
260 265 270
Ser Pro Ser Ser His Ser Phe Leu Leu Ile Leu Gly Asn Ala Lys Leu
275 280 285
Arg Gln Ala Phe Leu Leu Val Ala Ala Lys Val Trp Ala Lys Arg
290 295 300
<210>25
<211>912
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R13,hGR13
<400>25
atggaaagtg ccctgccgag tatcttcact cttgtaataa ttgcagaatt cataattggg 60
aatttgagca atggatttat agtactgatc aactgcattg actgggtcag taaaagagag 120
ctgtcctcag tcgataaact cctcattatc ttggcaatct ccagaattgg gctgatctgg 180
gaaatattag taagttggtt tttagctctg cattatctag ccatatttgt gtctggaaca 240
ggattaagaa ttatgatttt tagctggata gtttctaatc acttcaatct ctggcttgct 300
acaatcttca gcatctttta tttgctcaaa atagcgagtt tctctagccc tgcttttctc 360
tatttgaagt ggagagtaaa caaagtgatt ctgatgatac tgctaggaac cttggtcttc 420
ttatttttaa atctgataca aataaacatg catataaaag actggctgga ccgatatgaa 480
agaaacacaa cttggaattt cagtatgagt gactttgaaa cattttcagt gtcggtcaaa 540
ttcactatga ctatgttcag tctaacacca tttactgtgg ccttcatctc ttttctcctg 600
ttaattttct ccctgcagaa acatctccag aaaatgcaac tcaattacaa aggacacaga 660
gaccccagga ccaaggtcca tacaaatgcc ttgaaaattg tgatctcatt ccttttattc 720
tatgctagtt tctttctatg tgttctcata tcatggattt ctgagctgta tcagaacaca 780
gtgatctaca tgctttgtga gacgattgga gtcttctctc cttcaagcca ctcctttctt 840
ctgattctag gaaacgctaa gttaagacag gcctttcttt tggtggcagc taaggtatgg 900
gctaaacgat ga 912
<210>26
<211>317
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R14,hGR14
<400>26
Met Gly Gly Val Ile Lys Ser Ile Phe Thr Phe Val Leu Ile Val Glu
1 5 10 15
Phe Ile Ile Gly Asn Leu Gly Asn Ser Phe Ile Ala Leu Val Asn Cys
20 25 30
Ile Asp Trp Val Lys Gly Arg Lys Ile Ser Ser Val Asp Arg Ile Leu
35 40 45
Thr Ala Leu Ala Ile Ser Arg Ile Ser Leu Val Trp Leu Ile Phe Gly
50 55 60
Ser Trp Cys Val Ser Val Phe Phe Pro Ala Leu Phe Ala Thr Glu Lys
65 70 75 80
Met Phe Arg Met Leu Thr Asn Ile Trp Thr Val Ile Asn His Phe Ser
85 90 95
Val Trp Leu Ala Thr Gly Leu Gly Thr Phe Tyr Phe Leu Lys Ile Ala
100 105 110
Asn Phe Ser Asn Ser Ile Phe Leu Tyr Leu Lys Trp Arg Val Lys Lys
115 120 125
Val Val Leu Val Leu Leu Leu Val Thr Ser Val Phe Leu Phe Leu Asn
130 135 140
Ile Ala Leu Ile Asn Ile His Ile Asn Ala Ser Ile Asn Gly Tyr Arg
145 150 155 160
Arg Asn Lys Thr Cys Ser Ser Asp Ser Ser Asn Phe Thr Arg Phe Ser
165 170 175
Ser Leu Ile Val Leu Thr Ser Thr Val Phe Ile Phe Ile Pro Phe Thr
180 185 190
Leu Ser Leu Ala Met Phe Leu Leu Leu Ile Phe Ser Met Trp Lys His
195 200 205
Arg Lys Lys Met Gln His Thr Val Lys Ile Ser Gly Asp Ala Ser Thr
210 215 220
Lys Ala His Arg Gly Val Lys Ser Val Ile Thr Phe Phe Leu Leu Tyr
225 230 235 240
Ala Ile Phe Ser Leu Ser Phe Phe Ile Ser Val Trp Thr Ser Glu Arg
245 250 255
Leu Glu Glu Asn Leu Ile Ile Leu Ser Gln Val Met Gly Met Ala Tyr
260 265 270
Pro Ser Cys His Ser Cys Val Leu Ile Leu Gly Asn Lys Lys Leu Arg
275 280 285
Gln Ala Ser Leu Ser Val Leu Leu Trp Leu Arg Tyr Met Phe Lys Asp
290 295 300
Gly Glu Pro Ser Gly His Lys Glu Phe Arg Glu Ser Ser
305 310 315
<210>27
<211>954
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R14,hGR14
<400>27
atgggtggtg tcataaagag catatttaca ttcgttttaa ttgtggaatt tataattgga 60
aatttaggaa atagtttcat agcactggtg aactgtattg actgggtcaa gggaagaaag 120
atctcttcgg ttgatcggat cctcactgct ttggcaatct ctcgaattag cctggtttgg 180
ttaatattcg gaagctggtg tgtgtctgtg tttttcccag ctttatttgc cactgaaaaa 240
atgttcagaa tgcttactaa tatctggaca gtgatcaatc attttagtgt ctggttagct 300
acaggcctcg gtacttttta ttttctcaag atagccaatt tttctaactc tatttttctc 360
tacctaaagt ggagggttaa aaaggtggtt ttggtgctgc ttcttgtgac ttcggtcttc 420
ttgtttttaa atattgcact gataaacatc catataaatg ccagtatcaa tggatacaga 480
agaaacaaga cttgcagttc tgattcaagt aactttacac gattttccag tcttattgta 540
ttaaccagca ctgtgttcat tttcataccc tttactttgt ccctggcaat gtttcttctc 600
ctcatcttct ccatgtggaa acatcgcaag aagatgcagc acactgtcaa aatatccgga 660
gacgccagca ccaaagccca cagaggagtt aaaagtgtga tcactttctt cctactctat 720
gccattttct ctctgtcttt tttcatatca gtttggacct ctgaaaggtt ggaggaaaat 780
ctaattattc tttcccaggt gatgggaatg gcttatcctt catgtcactc atgtgttctg 840
attcttggaa acaagaagct gagacaggcc tctctgtcag tgctactgtg gctgaggtac 900
atgttcaaag atggggagcc ctcaggtcac aaagaattta gagaatcatc ttga 954
<210>28
<211>300
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R15,hGR15
<220>
<221>MOD_RES
<222>(257)
<223>Xaa=任何氨基酸
<400>28
Met Ile Thr Phe Leu Pro Ile Ile Phe Ser Ile Leu Val Val Val Thr
1 5 10 15
Phe Val Leu Gly Asn Phe Ala Asn Gly Phe Ile Val Leu Val Asn Ser
20 25 30
Ile Glu Trp Val Lys Arg Gln Lys Ile Ser Phe Ala Asp Gln Ile Leu
35 40 45
Thr Ala Leu Ala Val Ser Arg Val Gly Leu Leu Trp Val Ile Leu Leu
50 55 60
His Trp Tyr Ala Thr Val Leu Asn Pro Gly Ser Tyr Ser Leu Gly Val
65 70 75 80
Arg Ile Thr Thr Ile Asn Ala Trp Ala Val Thr Asn His Phe Ser Ile
85 90 95
Trp Val Ala Thr Ser Leu Ser Ile Phe Tyr Phe Leu Lys Ile Ala Asn
100 105 110
Phe Ser Asn Phe Ile Phe Leu His Leu Lys Arg Arg Ile Lys Ser Val
115 120 125
Ile Pro Val Ile Leu Leu Gly Ser Leu Leu Phe Leu Val Cys His Leu
130 135 140
Val Val Val Asn Met Asp Glu Ser Met Trp Thr Lys Glu Tyr Glu Gly
145 150 155 160
Asn Val Ser Trp Glu Ile Lys Leu Ser Asp Pro Thr His Leu Ser Asp
165 170 175
Met Thr Val Thr Thr Leu Ala Asn Leu Ile Pro Phe Thr Leu Ser Leu
180 185 190
Leu Ser Phe Leu Leu Leu Ile Cys Ser Leu Cys Lys His Leu Lys Lys
195 200 205
Met Gln Phe His Gly Lys Gly Ser Pro Asp Ser Asn Thr Lys Val His
210 215 220
Ile Lys Ala Leu Gln Thr Val Thr Ser Phe Leu Leu Leu Phe Ala Val
225 230 235 240
Tyr Phe Leu Ser Leu Ile Thr Ser Ile Trp Asn Phe Arg Arg Arg Leu
245 250 255
Xaa Asn Glu Pro Val Leu Met Leu Ser Gln Thr Thr Ala Ile Ile Tyr
260 265 270
Pro Ser Phe His Ser Phe Ile Leu Ile Trp Gly Ser Lys Lys Leu Lys
275 280 285
Gln Thr Phe Leu Leu Ile Leu Cys Gln Ile Lys Cys
290 295 300
<210>29
<211>903
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R15,hGR15
<400>29
atgataactt ttctacccat cattttttcc attctagtag tggttacatt tgttcttggg 60
aattttgcta atggcttcat agtgttggta aattccattg agtgggtcaa gagacaaaag 120
atctcctttg ctgaccaaat tctcactgct ctggcagtct ccagagttgg tttgctctgg 180
gtaatattat tacattggta tgcaactgtt ttgaatccag gttcatatag tttaggagta 240
agaattacta ctattaatgc ctgggctgta accaaccatt tcagcatctg ggttgctact 300
agcctcagca tattttattt cctcaagatt gccaatttct ccaactttat ttttcttcac 360
ttaaaaagga gaattaagag tgtcattcca gtgatactat tggggtcttt gttatttttg 420
gtttgtcatc ttgttgtggt aaacatggat gagagtatgt ggacaaaaga atatgaagga 480
aacgtgagtt gggagatcaa attgagtgat ccgacgcacc tttcagatat gactgtaacc 540
acgcttgcaa acttaatacc ctttactctg tccctgttat cttttctgct cttaatctgt 600
tctttgtgta aacatctcaa gaagatgcag ttccatggca aaggatctcc agattccaac 660
accaaggtcc acataaaagc tttgcaaacg gtgacctcct tcctcttgtt atttgctgtt 720
tactttctgt ccctaatcac atcgatttgg aattttagga ggaggctgta gaacgaacct 780
gtcctcatgc tcagccaaac tactgcaatt atataccctt catttcattc attcatccta 840
atttggggaa gcaagaagct gaaacagacc tttcttttga ttttgtgtca gattaagtgc 900
tga 903
<210>30
<211>291
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R16,hGR16
<400>30
Met Ile Pro Ile Gln Leu Thr Val Phe Phe Met Ile Ile Tyr Val Leu
1 5 10 15
Glu Ser Leu Thr Ile Ile Val Gln Ser Ser Leu Ile Val Ala Val Leu
20 25 30
Gly Arg Glu Trp Leu Gln Val Arg Arg Leu Met Pro Val Asp Met Ile
35 40 45
Leu Ile Ser Leu Gly Ile Ser Arg Phe Cys Leu Gln Trp Ala Ser Met
50 55 60
Leu Asn Asn Phe Cys Ser Tyr Phe Asn Leu Asn Tyr Val Leu Cys Asn
65 70 75 80
Leu Thr Ile Thr Trp Glu Phe Phe Asn Ile Leu Thr Phe Trp Leu Asn
85 90 95
Ser Leu Leu Thr Val Phe Tyr Cys Ile Lys Val Ser Ser Phe Thr His
100 105 110
His Ile Phe Leu Trp Leu Arg Trp Arg Ile Leu Arg Leu Phe Pro Trp
115 120 125
Ile Leu Leu Gly Ser Leu Met Ile Thr Cys Val Thr Ile Ile Pro Ser
130 135 140
Ala Ile Gly Asn Tyr Ile Gln Ile Gln Leu Leu Thr Met Glu His Leu
145 150 155 160
Pro Arg Asn Ser Thr Val Thr Asp Lys Leu Glu Asn Phe His Gln Tyr
165 170 175
Gln Phe Gln Ala His Thr Val Ala Leu Val Ile Pro Phe Ile Leu Phe
180 185 190
Leu Ala Ser Thr Ile Phe Leu Met Ala Ser Leu Thr Lys Gln Ile Gln
195 200 205
His His Ser Thr Gly His Cys Asn Pro Ser Met Lys Ala Arg Phe Thr
210 215 220
Ala Leu Arg Ser Leu Ala Val Leu Phe Ile Val Phe Thr Ser Tyr Phe
225 230 235 240
Leu Thr Ile Leu Ile Thr Ile Ile Gly Thr Leu Phe Asp Lys Arg Cys
245 250 255
Trp Leu Trp Val Trp Glu Ala Phe Val Tyr Ala Phe Ile Leu Met His
260 265 270
Ser Thr Ser Leu Met Leu Ser Ser Pro Thr Leu Lys Arg Ile Leu Lys
275 280 285
Gly Lys Cys
290
<210>31
<211>876
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R16,hGR16
<400>31
atgataccca tccaactcac tgtcttcttc atgatcatct atgtgcttga gtccttgaca 60
attattgtgc agagcagcct aattgttgca gtgctgggca gagaatggct gcaagtcaga 120
aggctgatgc ctgtggacat gattctcatc agcctgggca tctctcgctt ctgtctacag 180
tgggcatcaa tgctgaacaa tttttgctcc tattttaatt tgaattatgt actttgcaac 240
ttaacaatca cctgggaatt ttttaatatc cttacattct ggttaaacag cttgcttacc 300
gtgttctact gcatcaaggt ctcttctttc acccatcaca tctttctctg gctgaggtgg 360
agaattttga ggttgtttcc ctggatatta ctgggttctc tgatgattac ttgtgtaaca 420
atcatccctt cagctattgg gaattacatt caaattcagt tactcaccat ggagcatcta 480
ccaagaaaca gcactgtaac tgacaaactt gaaaattttc atcagtatca gttccaggct 540
catacagttg cattggttat tcctttcatc ctgttcctgg cctccaccat ctttctcatg 600
gcatcactga ccaagcagat acaacatcat agcactggtc actgcaatcc aagcatgaaa 660
gcgcgcttca ctgccctgag gtcccttgcc gtcttattta ttgtgtttac ctcttacttt 720
ctaaccatac tcatcaccat tataggtact ctatttgata agagatgttg gttatgggtc 780
tgggaagctt ttgtctatgc tttcatctta atgcattcca cttcactgat gctgagcagc 840
cctacgttga aaaggattct aaagggaaag tgctag 876
<210>32
<211>330
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R17,hGR17
<220>
<221>MOD_RES
<222>(1)..(330)
<223>Xaa=任何氨基酸
<400>32
Met Cys Ser Ala Xaa Leu Leu Ile Ile Leu Ser Ile Leu Val Val Phe
1 5 10 15
Ala Phe Val Leu Gly Asn Val Ala Asn Gly Phe Ile Ala Leu Ile Asn
20 25 30
Val Asn Asp Trp Val Lys Thr Gln Lys Ile Ser Ser Thr Asp Gln Ile
35 40 45
Val Thr Ala Leu Ala Phe Ser Arg Ile Gly Leu Leu Xaa Thr Leu Ile
50 55 60
Ile Leu Leu His Trp Tyr Ala Thr Val Phe Asn Ser Ala Leu Tyr Ser
65 70 75 80
Leu Glu Val Arg Ile Val Pro Ser Asn Val Ser Ala Ile Ile Asn His
85 90 95
Phe Ser Ile Trp Leu Ala Thr Ser Leu Ser Ile Phe Tyr Leu Phe Lys
100 105 110
Ile Ala Asn Phe Ser Asn Phe Ile Phe Leu His Leu Lys Lys Arg Ile
115 120 125
Lys Ser Val Leu Leu Val Ile Leu Leu Gly Ser Leu Val Phe Leu Ile
130 135 140
Cys Asn Leu Ala Val Val Thr Met Asp Asp Ser Val Trp Thr Lys Glu
145 150 155 160
Phe Glu Gly Asn Val Thr Trp Lys Ile Glu Leu Arg Asn Ala Ile His
165 170 175
Leu Ser Asn Met Thr Ile Thr Asn His Ala Ser Lys Leu His Thr Val
180 185 190
His Ser Asp Ser Asn Ile Phe Ser Ala Val Ser Leu Phe Ser Xaa Thr
195 200 205
Met Leu Ala Asn Phe Thr Leu Phe Ile Leu Thr Leu Ile Ser Phe Leu
210 215 220
Leu Leu Val Cys Ser Pro Cys Lys His Leu Lys Met Met Gln Leu His
225 230 235 240
Gly Lys Gly Ser Gln Asp Leu Ser Thr Lys Val His Ile Lys Pro Leu
245 250 255
Gln Thr Val Ile Ser Phe Arg Met Leu Phe Ala Ile Tyr Phe Leu Cys
260 265 270
Ile Ile Thr Ser Thr Trp Asn Pro Arg Thr Gln Gln Ser Asn Leu Val
275 280 285
Phe Leu Leu Tyr Gln Thr Leu Ala Ile Met Tyr Pro Ser Phe His Ser
290 295 300
Phe Ile Leu Ile Met Arg Ser Arg Lys Leu Lys Gln Thr Ser Leu Ser
305 310 315 320
Val Leu Cys Gln Val Thr Cys Trp Val Lys
325 330
<210>33
<211>314
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R18,hGR18
<220>
<221>MOD_RES
<222>(98)
<223>Xaa=任何氨基酸
<400>33
Met Phe Val Gly Ile Asn Ile Phe Phe Leu Val Val Ala Thr Arg Gly
1 5 10 15
Leu Val Leu Gly Met Leu Gly Asn Gly Leu Ile Gly Leu Val Asn Cys
20 25 30
Ile Glu Trp Ala Lys Ser Trp Lys Val Ser Ser Ala Asp Phe Ile Leu
35 40 45
Thr Ser Leu Ala Ile Val Arg Ile Ile Arg Leu Tyr Leu Ile Leu Phe
50 55 60
Asp Ser Phe Ile Met Val Leu Ser Pro His Leu Tyr Thr Ile Arg Lys
65 70 75 80
Leu Val Lys Leu Phe Thr Ile Leu Trp Ala Leu Ile Asn Gln Leu Ser
85 90 95
Ile Xaa Phe Ala Thr Cys Leu Ser Ile Phe Tyr Leu Leu Lys Ile Ala
100 105 110
Asn Phe Ser His Ser Leu Phe Leu Trp Leu Lys Trp Arg Met Asn Gly
115 120 125
Met Ile Val Met Leu Leu Ile Leu Ser Leu Phe Leu Leu Ile Phe Asp
130 135 140
Ser Leu Val Leu Glu Ile Phe Ile Asp Ile Ser Leu Asn Ile Ile Asp
145 150 155 160
Lys Ser Asn Leu Thr Leu Tyr Leu Asp Glu Ser Lys Thr Leu Tyr Asp
165 170 175
Lys Leu Ser Ile Leu Lys Thr Leu Leu Ser Leu Thr Tyr Val Ile Pro
180 185 190
Phe Leu Leu Thr Leu Thr Ser Leu Leu Leu Leu Phe Ile Ser Leu Val
195 200 205
Arg His Thr Lys Asn Leu Gln Leu Asn Ser Leu Gly Ser Arg Asp Ser
210 215 220
Ser Thr Glu Ala His Lys Arg Ala Met Lys Met Val Ile Ala Phe Leu
225 230 235 240
Leu Leu Phe Ile Ile Asn Phe Ile Ser Thr Leu Ile Gly Asp Trp Ile
245 250 255
Phe Leu Glu Val Glu Asn Tyr Gln Val Met Met Phe Ile Met Met Ile
260 265 270
Leu Leu Ala Phe Pro Ser Gly His Ser Phe Ile Ile Ile Leu Gly Asn
275 280 285
Asn Lys Leu Arg Gln Ser Ser Leu Arg Leu Leu Trp His Leu Lys Phe
290 295 300
Ser Leu Lys Lys Ala Lys Pro Leu Thr Ser
305 310
<210>34
<211>945
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R18,hGR18
<400>34
atgttcgttg gaattaatat tttctttctg gtggtggcaa caagaggact tgtcttagga 60
atgctgggaa acgggctcat tggactggta aactgcattg agtgggccaa gagttggaag 120
gtctcatcag ctgatttcat cctcaccagc ttggctatag tcagaatcat tcgactgtat 180
ttaatactat ttgattcatt tataatggta ttgtcccctc atctatatac catccgtaaa 240
ctagtaaaac tgtttactat tctttgggca ttaattaatc agttaagtat ctagtttgcc 300
acctgcctaa gcattttcta cttgcttaag atagccaatt tctcccactc ccttttcctc 360
tggctgaagt ggagaatgaa cggaatgatt gttatgcttc ttatattgtc tttgttctta 420
ctgatttttg acagtttagt gctagaaata tttattgata tctcactcaa tataatagat 480
aaaagtaatc tgactttata tttagatgaa agtaaaactc tctatgataa actctctatt 540
ttaaaaactc ttctcagctt gacatacgtt attccctttc ttctgactct gacctctttg 600
ctccttttat ttatatcctt agtgagacac accaagaatt tgcagctcaa ctctctgggc 660
tcaagggact ccagcacaga ggcccataaa agggccatga aaatggtgat agccttcctc 720
ctccttttta ttattaactt tatttccact ttaataggag attggatctt ccttgaggta 780
gagaattatc aggtcatgat gtttattatg atgattttac ttgcctttcc ctcaggccac 840
tcatttatta taattttggg aaacaacaag ctaagacaga gctccttgag actactgtgg 900
catcttaaat tctctctgaa aaaagcaaaa cctttaactt catag 945
<210>35
<211>121
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R19,hGR19
<400>35
Val Thr Thr Leu Ala Asn Leu Ile Pro Phe Thr Leu Ser Leu Ile Cys
1 5 10 15
Phe Leu Leu Leu Ile Cys Ser Leu Cys Lys His Leu Lys Lys Met Arg
20 25 30
Leu His Ser Lys Gly Ser Gln Asp Pro Ser Thr Lys Val His Ile Lys
35 40 45
Ala Leu Gln Thr Val Thr Ser Phe Leu Met Leu Phe Ala Ile Tyr Phe
50 55 60
Leu Cys Ile Ile Thr Ser Thr Trp Asn Leu Arg Thr Gln Gln Ser Lys
65 70 75 80
Leu Val Leu Leu Leu Cys Gln Thr Val Ala Ile Met Tyr Pro Ser Phe
85 90 95
His Ser Phe Ile Leu Ile Met Gly Ser Arg Lys Leu Lys Gln Thr Phe
100 105 110
Leu Ser Val Leu Trp Gln Met Thr Cys
115 120
<210>36
<211>466
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R19,hGR19
<400>36
ctgtaactac tctagcaaac ctcataccct ttactctgag cctaatatgt tttctgctgt 60
taatctgttc tctttgtaaa catctcaaga agatgcggct ccatagcaaa ggatctcaag 120
atcccagcac caaggtccat ataaaagctt tgcaaactgt gacctccttc ctcatgttat 180
ttgccattta ctttctgtgt ataatcacat caacttggaa tcttaggaca cagcagagca 240
aacttgtact cctgctttgc caaactgttg caatcatgta tccttcattc cactcattca 300
tcctgattat gggaagtagg aagctaaaac agacctttct ttcagttttg tggcagatga 360
catgctgagt gaaagaagag aaaccctcaa ctccatagat tcacaagggg agcatcgtgg 420
gtcttctagc agaaaacaaa ctgatggtgt ctggaacatt ttatat 466
<210>37
<211>129
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R20,hGR20
<220>
<221>MOD_RES
<222>(3)
<223>Xaa=任何氨基酸
<400>37
His Leu Xaa Arg Lys Ala Lys Ser Val Val Leu Val Ile Val Leu Gly
1 5 10 15
Ser Leu Phe Phe Leu Val Cys Gln Leu Val Met Lys Asn Thr Tyr Ile
20 25 30
Asn Val Trp Thr Glu Glu Cys Glu Gly Asn Val Thr Trp Lys Ile Lys
35 40 45
Leu Arg Asn Ala Met His Leu Ser Asn Leu Thr Val Ala Met Leu Ala
50 55 60
Asn Leu Ile Pro Phe Thr Leu Thr Val Ile Ser Phe Leu Leu Leu Ile
65 70 75 80
Tyr Ser Leu Cys Lys His Leu Lys Lys Met Gln Leu His Gly Lys Gly
85 90 95
Ser Gln Asp Pro Ser Thr Lys Ile His Ile Lys Ala Leu Gln Thr Val
100 105 110
Thr Ser Phe Leu Val Leu Leu Ala Ile Tyr Phe Leu Cys Leu Ile Ile
115 120 125
Ser
<210>38
<211>397
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R20,hGR20
<220>
<221>modified_base
<222>(12)
<223>n=g,a,c或t
<400>38
ttcatcactt anaaaggaag gctaagagtg tagttctggt gatagtgttg gggtctttgt 60
tctttttggt ttgtcaactt gtgatgaaaa acacgtatat aaatgtgtgg acagaagaat 120
gtgaaggaaa cgtaacttgg aagatcaaac tgaggaatgc aatgcacctt tccaacttga 180
ctgtagccat gctagcaaac ttgataccat tcactctgac cgtgatatct tttctgctgt 240
taatctactc tctgtgtaaa catctgaaga agatgcagct ccatggcaaa ggatctcaag 300
atcccagcac caagatccac ataaaagctc tgcaaactgt gacctccttc ctcgtattac 360
ttgccattta ctttctgtgt ctaatcatat ccttttg 397
<210>39
<211>312
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R21,hGR21
<220>
<221>MOD_RES
<222>(1)..(312)
<223>Xaa=任何氨基酸
<400>39
Met Pro Pro Gly Ile Gly Asn Thr Phe Leu Ile Val Met Met Gly Glu
1 5 10 15
Phe Ile Ile Xaa Met Leu Gly Asn Gly Phe Ile Val Leu Val Asn Cys
20 25 30
Ile Asp Trp Xaa Gly Val Lys Xaa Ser Tyr Xaa Thr Thr Ala Ser Ser
35 40 45
Pro Ala Trp Leu Ser Pro Gln Ser Val Asn Phe Gly Xaa Tyr Tyr Leu
50 55 60
Ile His Leu Xaa Gln His Tyr Gly His Ile Tyr Met Pro Ser Ile Asn
65 70 75 80
Xaa Xaa Asn Leu Phe Ile Phe Phe Gly His Xaa Pro Ile Thr Xaa Leu
85 90 95
Pro Gly Leu Leu Pro Xaa Cys Phe Leu Leu Leu Xaa Asn Thr Tyr Phe
100 105 110
Ser His Pro Cys Phe Ile Trp Leu Arg Trp Arg Ile Ser Arg Thr Leu
115 120 125
Leu Glu Leu Pro Leu Gly Ser Leu Leu Leu Leu Phe Phe Asn Leu Ala
130 135 140
Leu Thr Gly Gly Leu Ser Asp Leu Trp Ile Asn Ile Tyr Thr Ile Tyr
145 150 155 160
Glu Arg Asn Ser Thr Trp Ser Leu Asp Val Ser Lys Ile Leu Tyr Cys
165 170 175
Ser Leu Trp Ile Leu Val Ser Leu Ile Tyr Leu Ile Ser Phe Leu Leu
180 185 190
Ser Leu Ile Ser Leu Leu Leu Leu Ile Leu Ser Leu Met Arg His Ile
195 200 205
Arg Asn Leu Gln Leu Asn Thr Met Gly Pro Arg Asp Leu Arg Met Lys
210 215 220
Ala His Lys Arg Ala Met Lys Met Lys MetLys Met Met Val Ser Phe
225 230 235 240
Leu Leu Phe Phe Leu Val His Phe Ser Ser Leu Leu Pro Thr Gly Trp
245 250 255
Ile Phe Leu Ile Gln Gln Lys Xaa Gln Ala Asn Phe Phe Val Leu Leu
260 265 270
Thr Ser Ile Ile Phe Pro Ser Ser His Ser Phe Val Leu Ile Leu Glu
275 280 285
Asn Cys Lys Leu Arg Gln Thr Ala Val Gly Pro Leu Trp His Leu Lys
290 295 300
Cys His Leu Lys Arg Val Lys Leu
305 310
<210>40
<211>90
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R22,hGR22
<220>
<221>MOD_RES
<222>(1)..(90)
<223>Xaa=任何氨基酸
<400>40
Met Ala Thr Glu Ser Asp Thr Asn Leu Leu Ile Leu Ala Ile Ala Glu
1 5 10 15
Phe Ile Ile Ser Met Leu Gly Asn Val Phe Ile Gly Leu Val Asn Cys
20 25 30
Ser Glu Xaa Ile Lys Asn Xaa Lys Val Phe Ser Ala Asp Phe Ile Leu
35 40 45
Thr Cys Leu Ala Ile Ser His Asn Gly Gln Leu Leu Val Ile Leu Phe
50 55 60
Asp Ser Phe Leu Val Gly Leu Ala Ser His Leu Tyr Thr Thr Tyr Arg
65 70 75 80
Leu Xaa Lys Asn Cys Ile Met Leu Trp Thr
85 90
<210>41
<211>656
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R22,hGR22
<220>
<221>modified_base
<222>(1)..(656)
<223>n=g,a,c或t
<400>41
tatagggacn gtgatgcttc gtacactctc caagaagaaa cactccgtga ggtatgtgag 60
actgcatncc ttagtagatc tnttgggata tatattcata atatagaaaa anaggcaaag 120
acttncttaa gtatatgaga ctctatccaa cagcagaagg ttctgatcaa gactggaagt 180
gcaatanaag caatgaagat aagtatcaga tatgaatgct cttctgcaat ggtctgattg 240
tnacattatt aatgatacan agtattaaaa acttggattt tnttgtctct ggagatggcc 300
accgaatcgg acacaaatct tctgattctg gcaatagcag aattcatcat cagcatgctg 360
gggaatgtgt tcattggact ggtaaactgc tctgaangga tcaagaacca naaggtcttc 420
tcagctgact tcatcctcac ctgcttggct atctctcaca atggacaact gttggtgata 480
ctgtttgatt catttctagt gggacttgct tcacatctat ataccacata tagactanga 540
aaaaactgta ttatgctttg gacatgacta atcacttgac acactgcttc gcacgtgcta 600
gcatattcta ttcttagata gccacttcnc actccttgtc tctgctgaag tgggat 656
<210>42
<211>72
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R23,hGR23
<220>
<221>MOD_RES
<222>(1)..(72)
<223>Xaa=任何氨基酸
<400>42
Val Ala Phe Val Leu Gly Asn Val Ala Asn Gly Phe Ile Ala Leu Val
1 5 10 15
Asn Val Ile Asp Xaa Val Asn Thr Arg Lys Ile Ser Ser Ala Glu Gln
20 25 30
Ile Leu Thr Ala Leu Val Val Ser Arg Ile Gly Xaa Thr Leu Xaa His
35 40 45
Ser Ile Pro Xaa Asp Ala Thr Arg Cys Xaa Ser Ala Leu Tyr Arg Xaa
50 55 60
Glu Val Arg Ile Val Ala Ser Asn
65 70
<210>43
<211>589
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R23,hGR23
<220>
<221>modified_base
<222>(1)..(589)
<223>n=g,a,c或t
<400>43
agggttgagt cgtgcttatc ttcacttaac ctagtatana antacagcat atagcaagga 60
gagaatgtat atgaagagga gtgaatttga gtctgtttga gaataatgac cttttctatt 120
tctataaaga cagttttgaa ttcatctatt agcatatgct ggtgcttgcc tgttgacact 180
agtcactgaa tttaaaggca gaaaatgtta ttgcacattt agtaatcaag tgttcatcga 240
agttaacatc tggatgttaa aggactcaga acaagtgtta ctaagcctgc atttttttat 300
ctgttcaaac atgatgtgtt ntctgctcat catttcatca attctggtag agttgcattt 360
gttcttggaa atgtngccaa tggcttcata gctctagtaa atgtcattga ctgngttaac 420
acacgaaaga tctcctcagc tgagcaaatt ctcactgctc tggtggtctc cagaattggt 480
nntactctgn gtcatagtat tccttgagat gcaactagat gttaatctgc tctatatagg 540
ntagaagtaa gaattgttgc ttctaatgcc tgagctcgta cgaaccatt 589
<210>44
<211>314
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R24,hGR24
<400>44
Met Ala Thr Glu Leu Asp Lys Ile Phe Leu Ile Leu Ala Ile Ala Glu
1 5 10 15
Phe Ile Ile Ser Met Leu Gly Asn Val Phe Ile Gly Leu Val Asn Cys
20 25 30
Ser Glu Gly Ile Lys Asn Gln Lys Val Phe Ser Ala Asp Phe Ile Leu
35 40 45
Thr Cys Leu Ala Ile Ser Thr Ile Gly Gln Leu Leu Val Ile Leu Phe
50 55 60
Asp Ser Phe Leu Val Gly Leu Ala Ser His Leu Tyr Thr Thr Tyr Arg
65 70 75 80
Leu Gly Lys Thr Val Ile Met Leu Trp His Met Thr Asn His Leu Thr
85 90 95
Thr Trp Leu Ala Thr Cys Leu Ser Ile Phe Tyr Phe Phe Lys Ile Ala
100 105 110
His Phe Pro His Ser Leu Phe Leu Trp Leu Arg Trp Arg Met Asn Gly
115 120 125
Met Ile Val Met Leu Leu Ile Leu Ser Leu Phe Leu Leu Ile Phe Asp
130 135 140
Ser Leu Val Leu Glu Ile Phe Ile Asp Ile Ser Leu Asn Ile Ile Asp
145 150 155 160
Lys Ser Asn Leu Thr Leu Tyr Leu Asp Glu Ser Lys Thr Leu Tyr Asp
165 170 175
Lys Leu Ser Ile Leu Lys Thr Leu Leu Ser Leu Thr Ser Phe Ile Pro
180 185 190
Phe Ser Leu Phe Leu Thr Ser Leu Leu Phe Leu Phe Leu Ser Leu Val
195 200 205
Arg His Thr Arg Asn Leu Lys Leu Ser Ser Leu Gly Ser Arg Asp Ser
210 215 220
Ser Thr Glu Ala His Arg Arg Ala Met Lys Met Val Met Ser Phe Leu
225 230 235 240
Phe Leu Phe Ile Val His Phe Phe Ser Leu Gln Val Ala Asn Gly Ile
245 250 255
Phe Phe Met Leu Trp Asn Asn Lys Tyr Ile Lys Phe Val Met Leu Ala
260 265 270
Leu Asn Ala Phe Pro Ser Cys His Ser Phe Ile Leu Ile Leu Gly Asn
275 280 285
Ser Lys Leu Arg Gln Thr Ala Val Arg Leu Leu Trp His Leu Arg Asn
290 295 300
Tyr Thr Lys Thr Pro Asn A1a Leu Pro Leu
305 310
<210>45
<21l>945
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R24,hGR24
<400>45
atggccaccg aattggacaa aatctttctg attctggcaa tagcagaatt catcatcagc 60
atgctgggga atgtgttcat tggactggta aactgctctg aagggatcaa gaaccaaaag 120
gtcttctcag ctgacttcat cctcacctgc ttggctatct ccacaattgg acaactgttg 180
gtgatactgt ttgattcatt tctagtggga cttgcttcac atttatatac cacatataga 240
ctaggaaaaa ctgttattat gctttggcac atgactaatc acttgacaac ctggcttgcc 300
acctgcctaa gcattttcta tttctttaag atagcccact tcccccactc ccttttcctc 360
tggctgaggt ggaggatgaa cggaatgatt gttatgcttc ttatattgtc tttgttctta 420
ctgatttttg acagtttagt gctagaaata tttattgata tctcactcaa tataatagat 480
aaaagtaatc tgactttata tttagatgaa agtaaaactc tctatgataa actctctatt 540
ttaaaaactc ttctcagctt aaccagtttt atcccctttt ctctgttcct gacctccttg 600
ctttttttat ttctgtcctt ggtgagacat actagaaatt tgaagctcag ttccttgggc 660
tctagagact ccagcacaga ggcccatagg agggccatga aaatggtgat gtctttcctt 720
ttcctcttca tagttcattt tttttcctta caagtggcca atgggatatt ttttatgttg 780
tggaacaaca agtacataaa gtttgtcatg ttagccttaa atgcctttcc ctcgtgccac 840
tcatttattc tcattctggg aaacagcaag ctgcgacaga cagctgtgag gctactgtgg 900
catcttagga actatacaaa aacaccaaat gctttacctt tgtag 945
<210>46
<211>72
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R25,hGR25
<400>46
Leu Ser Pro Phe Arg Met Leu Phe Ala Ile Tyr Phe Leu Cys Ile Ile
1 5 10 15
Thr Ser Thr Trp Asn Pro Arg Thr Gln Gln Ser Asn Leu Val Phe Leu
20 25 30
Leu Tyr Gln Thr Leu Ala Ile Met Tyr Pro Ser Phe His Ser Phe Ile
35 40 45
Leu Ile Met Arg Ser Arg Lys Leu Lys Gln Thr Ser Leu Ser Val Leu
50 55 60
Cys Gln Val Thr Cys Trp Val Lys
65 70
<210>47
<211>264
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R26,hGR26
<220>
<221>MOD_RES
<222>(20)
<223>Xaa=任何氨基酸
<400>47
Met Pro Pro Gly Ile Gly Asn Thr Phe Leu Ile Val Met Met Gly Glu
1 5 10 15
Phe Ile Ile Xaa Met Leu Gly Asn Gly Phe Ile Val Leu Val Asn Cys
20 25 30
Ile Asp Val Arg Ser Gln Met Ile Leu Leu Asp Asn Cys Ile Leu Thr
35 40 45
Ser Leu Ala Ile Ser Thr Ile Ser Gln Leu Trp Ile Ile Leu Leu Asp
50 55 60
Ser Phe Val Thr Ala Leu Trp Pro His Leu Tyr Ala Phe Asn Lys Leu
65 70 75 80
Ile Lys Phe Ile His Ile Phe Trp Ala Leu Thr Asn His Leu Val Thr
85 90 95
Trp Leu Ala Cys Cys Leu Ser Val Phe Tyr Phe Phe Lys Ile Ala Tyr
100 105 110
Phe Ser His Pro Cys Phe Ile Trp Leu Arg Trp Arg Ile Ser Arg Thr
115 120 125
Leu Leu Glu Leu Pro Leu Gly Ser Leu Leu Leu Leu Phe Phe Asn Leu
130 135 140
Ala Leu Thr Gly Gly Leu Ser Asp Leu Trp Ile Asn Ile Tyr Thr Met
145 150 155 160
Tyr Glu Arg Asn Ser Thr Trp Ser Leu Asp Val Ser Lys Ile Leu Tyr
165 170 175
Cys Ser Leu Trp Ile Leu Val Ser Leu Ile Tyr Leu Ile Ser Phe Leu
180 185 190
Leu Ser Leu Ile Ser Leu Leu Leu Leu Ile Leu Ser Leu Met Arg His
195 200 205
Ile Arg Asn Leu Gln Leu Asn Thr Met Gly Pro Arg Asp Leu Arg Met
210 215 220
Lys Ala His Lys Arg Ala Met Lys Met Lys Met Lys Met Met Val Ser
225 230 235 240
Phe Leu Leu Phe Phe Leu Val His Phe Ser Ser Leu Leu Pro Thr Gly
245 250 255
Trp Ile Phe Leu Ile Gln Gln Lys
260
<210>48
<211>264
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R27,hGR27
<220>
<221>MOD_RES
<222>(1)..(264)
<223>Xaa=任何氨基酸
<400>48
Leu Ala Asn Leu Ile Asp Trp Ala Glu Asn Gln Ile Cys Leu Met Asp
1 5 10 15
Phe Ile Leu Ser Ser Leu Ala Ile Cys Arg Thr Leu Leu Leu Gly Cys
20 25 30
Cys Val Ala Ile Arg Cys Thr Tyr Asn Asp Tyr Pro Asn Ile Asp Ala
35 40 45
Val Asn His Asn Leu Ile Lys Ile Ile Thr Ile Phe Asp Ile Leu Arg
50 55 60
Leu Val Ser Lys Xaa Leu Gly Ile Trp Phe Ala Ser Tyr Leu Ser Ile
65 70 75 80
Phe Tyr Leu Leu Lys Val Ala Leu Phe His His Ala Ile Phe Leu Trp
85 90 95
Leu Lys Trp Arg Ile Ser Arg Ala Val Phe Thr Phe Leu Met Ile Phe
100 105 110
Leu Phe Phe Tyr Ile Ser Ile Ile Ser Met Ile Lys Ile Lys Leu Phe
115 120 125
Leu Asp Gln Cys Xaa Tyr Lys Ile Xaa Glu Lys Leu Leu Leu Glu Gly
130 135 140
Arg Cys Glu Xaa Ser Pro Pro Ser Cys Xaa Pro Asp Ala His Xaa Pro
145 150 155 160
Gly Val Val Tyr Ser Leu Tyr His Phe Ser Tyr Leu Met Phe Leu Val
165 170 175
Cys Tyr Leu Pro Lys Gly Lys His Cys Thr Ala Val Val Ile Gly Asp
180 185 190
Trp Leu Gln Arg Pro Arg Thr Glu Ala Tyr Val Arg Ala Met Asn Ile
195 200 205
Met Ile Ala Phe Phe Phe His Leu Leu Tyr Ser Leu Gly Thr Ser Leu
210 215 220
Ser Ser Val Ser Tyr Phe Leu Cys Lys Arg Lys Ile Val Ala Leu Gly
225 230 235 240
Ala Tyr Leu Ser Tyr Pro Leu Ser His Ser Phe Ile Leu Ile Met Glu
245 250 255
Asn Asn Lys Val Arg Lys Ala Leu
260
<210>49
<211>35
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R28,hGR28
<400>49
Asn Ile Cys Val Leu Leu Ile Ile Leu Ser Ile Leu Val Val Ser Ala
1 5 10 15
Phe Val Leu Gly Asn Val Ala Asn Gly Phe Ile Ala Leu Ile Asn Val
20 25 30
Asn Asp Trp
35
<210>50
<211>36
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R29,hGR29
<400>50
Met Gln Ala Ala Leu Thr Ala Phe Phe Val Leu Leu Phe Ser Leu Leu
1 5 10 15
Ser Leu Leu Gly Ile Ala Ala Asn Gly Phe Ile Val Leu Val Leu Gly
20 25 30
Lys Glu Trp Leu
35
<210>51
<211>319
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R30,hGR30
<400>51
Met Ile Thr Phe Leu Pro Ile Ile Phe Ser Ile Leu Val Val Val Thr
1 5 10 15
Phe Val Leu Gly Asn Phe Ser Asn Gly Phe Ile Ala Leu Val Asn Ser
20 25 30
Ile Glu Trp Val Lys Thr Arg Lys Ile Ser Ser Ala Asp Gln Ile Leu
35 40 45
Thr Ala Leu Val Val Ser Arg Val Gly Leu Leu Trp Val Ile Leu Leu
50 55 60
His Trp Tyr Ala Asn Val Phe Asn Ser Ala Leu Tyr Ser Ser Glu Val
65 70 75 80
Gly Ala Val Ala Ser Asn Ile Ser Ala Ile Ile Asn His Phe Ser Ile
85 90 95
Trp Leu Ala Thr Ser Leu Ser Ile Phe Tyr Leu Leu Lys Ile Ala Asn
100 105 110
Phe Ser Asn Leu Ile Phe Leu His Leu Lys Lys Arg Ile Arg Ser Val
115 120 125
Val Leu Val Ile Leu Leu Gly Pro Leu Val Phe Leu Ile Cys Asn Leu
130 135 140
Ala Val Ile Thr Met Asp Asp Ser Val Trp Thr Lys Glu Tyr Glu Gly
145 150 155 160
Asn Val Thr Trp Lys Ile Lys Leu Arg Asn Ala Ile His Leu Ser Asn
165 170 175
Met Thr Val Ser Thr Leu Ala Asn Leu Ile Pro Phe Ile Leu Thr Leu
180 185 190
Ile Cys Phe Leu Leu Leu Ile Cys Ser Leu Cys Lys His Leu Lys Lys
195 200 205
Met Gln Leu His Gly Lys Gly Ser Gln Asp Pro Ser Thr Lys Val His
210 215 220
Ile Lys Ala Leu Gln Thr Val Thr Ser Phe Leu Leu Leu Cys Ala Ile
225 230 235 240
Tyr Phe Leu Ser Met Ile Ile Ser Val Cys Asn Phe Gly Arg Leu Glu
245 250 255
Lys Gln Pro Val Phe Met Phe Cys Gln Ala Ile Ile Phe Ser Tyr Pro
260 265 270
Ser Thr His Pro Phe Ile Leu Ile Leu Gly Asn Lys Lys Leu Lys Gln
275 280 285
Ile Phe Leu Ser Val Leu Arg His Val Arg Tyr Trp Val Lys Asp Arg
290 295 300
Ser Leu Arg Leu His Arg Phe Thr Arg Gly Ala Leu Cys Val Phe
305 310 315
<210>52
<211>960
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R30,hGR30
<400>52
atgataactt ttctacccat cattttttcc attctggtag tggttacatt tgttcttgga 60
aatttttcca atggcttcat agctctagta aattccattg agtgggtcaa gacacgaaag 120
atctcctcag ctgaccaaat cctcactgct ctggtggtct ccagagttgg tttactctgg 180
gtcatattat tacattggta tgcaaatgtg tttaattcag ctttatatag ttcagaagta 240
ggagctgttg cttctaatat ctcagcaata atcaaccatt tcagcatctg gcttgctact 300
agcctcagca tattttattt gctcaagatt gccaatttct ccaaccttat ttttctccac 360
ttaaagaaga gaattaggag tgttgttctg gtgatactgt tgggtccctt ggtatttttg 420
atttgtaatc ttgctgtgat aaccatggat gacagtgtgt ggacaaaaga atatgaagga 480
aatgtgactt ggaagatcaa attgaggaat gcaatacacc tttcaaatat gactgtaagc 540
acactagcaa acctcatacc cttcattctg accctaatat gttttctgct gttaatctgt 600
tctctgtgta aacatctcaa gaagatgcag ctccatggca aaggatctca agatcccagc 660
accaaggtcc acataaaagc tttgcaaact gtgacctcct ttcttctgtt atgtgccatt 720
tactttctgt ccatgatcat atcagtttgt aattttggga ggctggaaaa gcaacctgtc 780
ttcatgttct gccaagctat tatattcagc tatccttcaa cccacccatt catcctgatt 840
ttgggaaaca agaagctaaa gcagattttt ctttcagttt tgcggcatgt gaggtactgg 900
gtgaaagaca gaagccttcg tctccataga ttcacaagag gggcattgtg tgtcttctag 960
<210>53
<211>299
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R31,hGR31
<220>
<221>MOD_RES
<222>(1)..(299)
<223>Xaa=任何氨基酸
<400>53
Met Thr Thr Phe Ile Pro Ile Ile Phe Ser Ser Val Val Val Val Leu
1 5 10 15
Phe Val Ile Gly Asn Phe Ala Asn Gly Phe Ile Ala Leu Val Asn Ser
20 25 30
Ile Glu Arg Val Lys Arg Gln Lys Ile Ser Phe Ala Asp Gln Ile Leu
35 40 45
Thr Ala Leu Ala Val Ser Arg Val Gly Leu Leu Trp Val Leu Leu Leu
50 55 60
Asn Trp Tyr Ser Thr Val Phe Asn Pro Ala Phe Tyr Ser Val Glu Val
65 70 75 80
Arg Thr Thr Ala Tyr Asn Val Trp Ala Val Thr Gly His Phe Ser Asn
85 90 95
Trp Leu Ala Thr Ser Leu Ser Ile Phe Tyr Leu Leu Lys Ile Ala Asn
100 105 110
Phe Ser Asn Leu Ile Phe Leu His Leu Lys Arg Arg Val Lys Ser Val
115 120 125
Ile Leu Val Met Leu Leu Gly Pro Leu Leu Phe Leu Ala Cys Gln Leu
130 135 140
Phe Val Ile Asn Met Lys Glu Ile Val Arg Thr Lys Glu Phe Glu Gly
145 150 155 160
Asn Met Thr Trp Lys Ile Lys Leu Lys Ser Ala Met Tyr Phe Ser Xaa
165 170 175
Met Thr Val Thr Ile Gly Ala Xaa Leu Val Pro Phe Thr Leu Ser Leu
180 185 190
Ile Ser Phe Leu Met Leu Ile Cys Ser Leu Cys Lys His Leu Lys Lys
195 200 205
Met Gln Leu His Gly Glu Gly Ser Gln Asp Leu Ser Thr Lys Val His
210 215 220
Ile Lys Ala Leu Gln Thr Leu Ile Ser Phe Leu Leu Leu Cys Ala Ile
225 230 235 240
Phe Phe Leu Phe Leu Ile Val Ser Val Trp Ser Pro Arg Arg Leu Arg
245 250 255
Asn Asp Pro Val Val Met Val Ser Lys Ala Val Gly Asn Ile Tyr Leu
260 265 270
Ala Phe Asp Ser Phe Ile Leu Ile Trp Arg Thr Lys Lys Leu Lys His
275 280 285
Thr Phe Leu Leu Ile Leu Cys Gln Ile Arg Cys
290 295
<210>54
<211>900
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R31,hGR31
<220>
<221>modified_base
<222>(1)..(900)
<223>n=g,a,c或t
<400>54
atgacaactt ttatacccat cattttttcc agtgtggtag tggttctatt tgttattgga 60
aattttgcta atggcttcat agcattggta aattccattg agcgggtcaa gagacaaaag 120
atctcttttg ctgaccagat tctcactgct ctggcggtct ccagagttgg tttgctctgg 180
gtattattat taaattggta ttcaactgtg tttaatccag ctttttatag tgtagaagta 240
agaactactg cttataatgt ctgggcagta accggccatt tcagcaactg gcttgctact 300
agcctcagca tattttattt gctcaagatt gccaatttct ccaaccttat ttttcttcac 360
ttaaagagga gagttaagag tgtcattctg gtgatgctgt tggggccttt actatttttg 420
gcttgtcaac tttttgtgat aaacatgaaa gagattgtac ggacaaaaga atttgaagga 480
aacatgactt ggaagatcaa attgaagagt gcaatgtact tttcanatat gactgtaacc 540
attggagcan acttagtacc ctttactctg tccctgatat cttttctgat gctaatctgt 600
tctctgtgta aacatctcaa gaagatgcag ctccatggag aaggatcgca agatctcagc 660
accaaggtcc acataaaagc tttgcaaact ctgatctcct tcctcttgtt atgtgccatt 720
ttctttctat tcctaatcgt ttcggtttgg agtcctagga ggctgcggaa tgacccggtt 780
gtcatggtta gcaaggctgt tggaaacata tatcttgcat tcgactcatt catcctaatt 840
tggagaacca agaagctaaa acacaccttt cttttgattt tgtgtcagat taggtgctga 900
<210>55
<211>20
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R32,hGR32
<400>55
His Ser Phe Met Leu Thr Met Gly Ser Arg Lys Pro Lys Gln Thr Phe
1 5 10 15
Leu Ser Ala Leu
20
<210>56
<211>309
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R33,hGR33
<400>56
Met Val Tyr Phe Leu Pro Ile Ile Phe Ser Ile Leu Val Val Phe Ala
1 5 10 15
Phe Val Leu Gly Asn Phe Ser Asn Gly Phe Ile Ala Leu Val Asn Val
20 25 30
Ile Asp Trp Val Lys Arg Gln Lys Ile Ser Ser Ala Asp Gln Ile Leu
35 40 45
Thr Ala Leu Val Val Ser Arg Val Gly Leu Leu Trp Val Ile Leu Leu
50 55 60
His Trp Tyr Ala Asn Val Phe Asn Ser Ala Leu Tyr Ser Leu Glu Val
65 70 75 80
Arg Ile Val Ala Ser Asn Ile Ser Ala Val Ile Asn His Phe Ser Ile
85 90 95
Trp Leu Ala Ala Ser Leu Ser Ile Phe Tyr Leu Leu Lys Ile Ala Asn
100 105 110
Phe Ser Asn Leu Ile Phe Leu His Leu Lys Lys Arg Ile Lys Ser Val
115 120 125
Val Leu Val Ile Leu Leu Gly Pro Leu Val Phe Leu Ile Cys Asn Leu
130 135 140
Ala Val Ile Thr Met Asp Glu Arg Val Trp Thr Lys Glu Tyr Glu Gly
145 150 155 160
Asn Val Thr Trp Lys Ile Lys Leu Arg Asn Ala Ile His Leu Ser Ser
165 170 175
Leu Thr Val Thr Thr Leu Ala Asn Leu Ile Pro Phe Thr Leu Ser Leu
180 185 190
Ile Cys Phe Leu Leu Leu Ile Cys Ser Leu Cys Lys His Leu Lys Lys
195 200 205
Met Gln Leu His Ser Lys Gly Ser Gln Asp Pro Ser Thr Lys Val His
210 215 220
Ile Lys Ala Leu Gln Thr Val Ile Ser Phe Leu Met Leu Cys Ala Ile
225 230 235 240
Tyr Phe Leu Ser Ile Met Ile Ser Val Trp Asn Leu Arg Ser Leu Glu
245 250 255
Asn Lys Pro Val Phe Met Phe Cys Lys Ala Ile Arg Phe Ser Tyr Pro
260 265 270
Ser Ile His Pro Phe Ile Leu Ile Trp Gly Asn Lys Lys Leu Lys Gln
275 280 285
Thr Phe Leu Ser Val Phe Trp Gln Val Arg Tyr Trp Val Lys Gly Glu
290 295 300
Lys Pro Ser Ser Pro
305
<210>57
<211>930
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R33,hGR33
<400>57
atggtatatt ttctgcccat cattttttcc attctggtag tgtttgcatt tgttcttgga 60
aatttttcca atggcttcat agctctagta aatgtcattg actgggttaa gagacaaaag 120
atctcctcag ctgaccaaat tctcactgct ctggtggtct ccagagttgg tttactctgg 180
gtcatattat tacattggta tgcaaatgtg tttaattcag ctttatatag tttagaagta 240
agaattgttg cttctaatat ctcagcagta atcaaccatt tcagcatctg gcttgctgct 300
agcctcagca tattttattt gctcaagatt gccaatttct ccaaccttat ttttctccac 360
ctaaagaaga gaattaagag tgttgttctg gtgatactgt tggggccctt ggtatttctg 420
atttgtaatc ttgctgtgat aaccatggat gagagagtgt ggacaaaaga atatgaagga 480
aatgtgactt ggaagatcaa attgaggaat gcaatacacc tttcaagctt gactgtaact 540
actctagcaa acctcatacc ctttactctg agcctaatat gttttctgct gttaatctgt 600
tctctttgta aacatctcaa gaagatgcag ctccatagca aaggatctca agatcccagc 660
accaaggtcc acataaaagc tttgcaaact gtgatctcct tcctcatgtt atgtgccatt 720
tactttctgt ccataatgat atcagtttgg aatcttagga gtctggaaaa caaacctgtc 780
ttcatgttct gcaaagctat tagattcagc tatccttcaa tccacccatt catcctgatt 840
tggggaaaca agaagctaaa gcagactttt ctttcagttt tttggcaagt gaggtactgg 900
gtgaaaggag agaagccttc atctccatag 930
<210>58
<211>100
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R34,hGR34
<220>
<221>MOD_RES
<222>(1)..(100)
<223>Xaa=任何氨基酸
<400>58
Gly Ser Ser Arg Xaa Lys Pro Pro Arg Ile Pro His Lys Lys Leu Cys
1 5 10 15
Lys Leu Gly Pro Ser Phe Pro His Asn Asn Leu Pro Ile Tyr Phe Leu
20 25 30
Cys Xaa Asn His Ile Val Leu Glu Phe Leu Lys Met Arg Pro Lys Lys
35 40 45
Lys Cys Ser Leu Met Leu Cys Gln Ala Phe Gly Ile Ile Tyr Pro Ser
50 55 60
Phe His Ser Phe Ile Leu Xaa Trp Gly Asn Lys Thr Leu Lys Gln Thr
65 70 75 80
Phe Leu Ser Val Xaa Trp Gln Val Thr Cys Trp Ala Lys Gly Gln Asn
85 90 95
Gln Ser Thr Pro
100
<210>59
<211>128
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R35,hGR35
<220>
<221>MOD_RES
<222>(1)..(128)
<223>Xaa=任何氨基酸
<400>59
Asn Ala Ile Arg Pro Ser Lys Leu Trp Thr Val Thr Glu Ala Asp Lys
1 5 10 15
Thr Ser Gln Pro Gly Thr Ser Ala Asn Lys Ile Phe Ser Ala Gly Asn
20 25 30
Leu Ile Ser His Val Asn Met Ser Arg Arg Met Gln Leu His Gly Lys
35 40 45
Gly Ser Gln His Leu Ser Thr Arg Val His Ile Lys Ala Xaa Gln Thr
50 55 60
Val Ile Ser Phe Leu Met Leu Xaa Ala Ile Tyr Phe Leu Cys Leu Ile
65 70 75 80
Thr Ser Thr Trp Asn Pro Arg Thr Gln Gln Ser Lys Leu Val Phe Leu
85 90 95
Leu Tyr Gln Thr Leu Gly Phe Met Tyr Leu Leu Phe His Ser Phe Ile
100 105 110
Leu Thr Met Gly Ser Arg Lys Pro Lys Gln Thr Phe Leu Ser Ala Leu
115 120 125
<210>60
<211>309
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R36,hGR36
<400>60
Met Ile Cys Phe Leu Leu Ile Ile Leu Ser Ile Leu Val Val Phe Ala
1 5 10 15
Phe Val Leu Gly Asn Phe Ser Asn Gly Phe Ile Ala Leu Val Asn Val
20 25 30
Ile Asp Trp Val Lys Arg Gln Lys Ile Ser Ser Ala Asp Gln Ile Leu
35 40 45
Thr Ala Leu Val Val Ser Arg Val Gly Leu Leu Trp Val Ile Leu Leu
50 55 60
His Trp Tyr Ser Asn Val Leu Asn Ser Ala Leu Tyr Ser Ser Glu Val
65 70 75 80
Ile Ile Phe Ile Ser Asn Ala Trp Ala Ile Ile Asn His Phe Ser Ile
85 90 95
Trp Leu Ala Thr Ser Leu Ser Ile Phe Tyr Leu Leu Lys Ile Val Asn
100 105 110
Phe Ser Arg Leu Ile Phe His His Leu Lys Arg Lys Ala Lys Ser Val
115 120 125
Val Leu Val Ile Val Leu Gly Pro Leu Val Phe Leu Val Cys His Leu
130 135 140
Val Met Lys His Thr Tyr Ile Asn Val Trp Thr Lys Glu Tyr Glu Gly
145 150 155 160
Asn Val Thr Trp Lys Ile Lys Leu Arg Asn Ala Ile His Leu Ser Asn
165 170 175
Leu Thr Val Ser Thr Leu Ala Asn Leu Ile Pro Phe Thr Leu Thr Leu
180 185 190
Ile Ser Phe Leu Leu Leu Ile Tyr Ser Leu Cys Lys His Leu Lys Lys
195 200 205
Met Gln Leu His Gly Lys Gly Ser Gln Asp Pro Ser Thr Lys Val His
210 215 220
Ile Lys Ala Leu Gln Thr Val Thr Ser Phe Leu Leu Leu Cys Ala Ile
225 230 235 240
Tyr Phe Leu Ser Met Ile Ile Ser Val Cys Asn Phe Gly Arg Leu Glu
245 250 255
Lys Gln Pro Val Phe Met Phe Cys Gln Ala Ile Ile Phe Ser Tyr Pro
260 265 270
Ser Thr His Pro Phe Ile Leu Ile Leu Gly Asn Lys Lys Leu Lys Gln
275 280 285
Ile Phe Leu Ser Val Phe Trp Gln Met Arg Tyr Trp Val Lys Gly Glu
290 295 300
Lys Pro Ser Ser Pro
305
<210>61
<211>930
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R36,hGR36
<400>61
atgatatgtt ttctgctcat cattttatca attctggtag tgtttgcatt tgttcttgga 60
aatttttcca atggcttcat agctctagta aatgtcattg actgggtcaa gagacaaaag 120
atctcctcag ctgaccaaat cctcactgct ctggtggtct ccagagttgg tttactctgg 180
gtaatattat tacattggta ttcaaatgtg ttgaattcag ctttatatag ttcagaagta 240
ataattttta tttctaatgc ctgggcaata atcaaccatt tcagcatctg gcttgctact 300
agcctcagca tattttattt gctcaagatc gtcaatttct ccagacttat ttttcatcac 360
ttaaaaagga aggctaagag tgtagttctg gtgatagtgt tgggtccctt ggtatttttg 420
gtttgtcacc ttgtgatgaa acacacgtat ataaatgtgt ggacaaaaga atatgaagga 480
aatgtgactt ggaagatcaa actgaggaat gcaatacacc tttcaaactt gactgtaagc 540
acactagcaa acttgatacc cttcactctg accctgatat cttttctgct gttaatctac 600
tctctgtgta aacatctcaa gaagatgcag ctccatggca aaggatctca agatcccagc 660
accaaggtcc acataaaagc tttgcaaact gtgacctcct ttcttctgtt atgtgccatt 720
tactttctgt ccatgatcat atcagtttgt aattttggga ggctggaaaa gcaacctgtc 780
ttcatgttct gccaagctat tatattcagc tatccttcaa cccacccatt catcctgatt 840
ttgggaaaca agaagctaaa gcagattttt ctttcagttt tttggcaaat gaggtactgg 900
gtgaaaggag agaagccttc atctccatag 930
<210>62
<211>309
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R37,hGR37
<220>
<221>MOD_RES
<222>(1)..(309)
<223>Xaa=任何氨基酸
<400>62
Met Ile Thr Phe Leu Pro Ile Ile Phe Ser Ile Leu Ile Val Val Thr
1 5 10 15
Phe Val Ile Gly Asn Phe Ala Asn Gly Phe Ile Ala Leu Val Asn Ser
20 25 30
Ile Glu Trp Val Lys Arg Gln Lys Ile Ser Ser Ala Asp Gln Ile Ser
35 40 45
His Cys Ser Gly Gly Val Gln Asn Trp Phe Thr Leu Gly His Ile Ile
50 55 60
Thr Leu Val Cys Asn Cys Val Xaa Phe Gly Phe Ile Xaa Ile Arg Ser
65 70 75 80
Lys Asn Phe Trp Phe Xaa Cys Leu Ser Asn Asn Gln Ala Phe Gln His
85 90 95
Val Gly Val Thr Ser Leu Ser Ile Phe His Leu Leu Lys Thr Ala Asn
100 105 110
Phe Ser Asn Leu Ile Phe Leu His Leu Lys Lys Arg Ile Lys Ser Val
115 120 125
Gly Leu Val Ile Leu Leu Gly Pro Leu Leu Phe Phe Ile Cys Asn Leu
130 135 140
Phe Val Ile Asn Met Asp Glu Ser Val Trp Thr Lys Glu Tyr Glu Gly
145 150 155 160
Asn Val Thr Trp Lys Ile Lys Leu Arg Ser Ala Met Tyr His Ser Asn
165 170 175
Met Thr Leu Thr Met Leu Ala Asn Phe Val Pro Phe Thr Leu Thr Leu
180 185 190
Ile Ser Phe Leu Leu Leu Ile Cys Ser Leu Cys Lys His Leu Lys Lys
195 200 205
Met Gln Leu His Gly Lys Gly Ser Gln Asp Pro Ser Thr Lys Val His
210 215 220
Ile Lys Ala Leu Gln Thr Val Thr Ser Phe Leu Leu Leu Cys Ala Ile
225 230 235 240
Tyr Phe Leu Ser Met Ile Ile Ser Val Cys Asn Leu Gly Arg Leu Glu
245 250 255
Lys Gln Pro Val Phe Met Phe Cys Glu Ala Ile Ile Phe Ser Tyr Pro
260 265 270
Ser Thr His Pro Phe Ile Leu Ile Leu Gly Asn Lys Lys Leu Lys Gln
275 280 285
Ile Phe Leu Ser Val Leu Arg His Val Arg Tyr Trp Val Lys Gly Glu
290 295 300
Lys Pro Ser Ser Ser
305
<210>63
<211>930
<212>DNA
<213>人类(Homo sapiens)
<220>
<223>人T2R37,hGR37
<400>63
atgataactt ttctgcccat cattttttcc attctaatag tggttacatt tgtgattgga 60
aattttgcta atggcttcat agctctagta aattccattg agtgggttaa gagacaaaag 120
atctcatcag ctgaccaaat ttctcactgc tctggtggtg tccagaattg gtttactctg 180
ggtcatatta ttacattggt atgcaactgt gtttaatttg gcttcatata gattagaagt 240
aagaattttt ggttctaatg tctcagcaat aaccaagcat ttcagcatgt gggtgttact 300
agcctcagca tatttcattt gctcaagact gccaatttct ccaaccttat ttttctccac 360
ctaaagaaga ggattaagag tgttggtttg gtgatactat tggggccttt gctatttttc 420
atttgtaatc tttttgtgat aaacatggat gagagtgtat ggacaaaaga atatgaagga 480
aacgtgactt ggaagatcaa attgaggagt gcaatgtacc attcaaatat gactctaacc 540
atgctagcaa actttgtacc cttcactctg accctgatat cttttctgct gttaatctgt 600
tctctgtgta aacatctcaa gaagatgcag ctccatggca aaggatctca agatcccagc 660
accaaggtcc acataaaagc tttgcaaact gtgacctcct ttcttctgtt atgtgccatt 720
tactttctgt ccatgatcat atcagtttgt aatttgggga ggctggaaaa gcaacctgtc 780
ttcatgttct gcgaagctat tatattcagc tatccttcaa cccacccatt catcctgatt 840
ttgggaaaca agaagctaaa gcagattttt ctttcagttt tgcggcatgt gaggtactgg 900
gtgaaaggag agaagccttc atcttcatag 930
<210>64
<211>144
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R38,hGR38
<400>64
Met Leu Thr Leu Thr Arg Ile Arg Thr Val Ser Tyr Glu Val Arg Ser
1 5 10 15
Thr Phe Leu Phe Ile Ser Val Leu Glu Phe Ala Val Gly Phe Leu Thr
20 25 30
Asn Ala Phe Val Phe Leu Val Asn Phe Trp Asp Val Val Lys Arg Gln
35 40 45
Pro Leu Ser Asn Ser Asp Cys Val Leu Leu Cys Leu Ser Ile Ser Arg
50 55 60
Leu Phe Leu His Gly Leu Leu Phe Leu Ser Ala Ile Gln Leu Thr His
65 70 75 80
Phe Gln Lys Leu Ser Glu Pro Leu Asn His Ser Tyr Gln Ala Ile Ile
85 90 95
Met Leu Trp Met Ile Ala Asn Gln Ala Asn Leu Trp Leu Ala Ala Cys
100 105 110
Leu Ser Leu Leu Tyr Cys Ser Lys Leu Ile Arg Phe Ser His Thr Phe
115 120 125
Leu Ile Cys Leu Ala Ser Trp Ser Pro Gly Arg Ser Pro Val Pro Ser
130 135 140
<210>65
<211>140
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R39,hGR39
<400>65
Leu Arg Asn Ala Gly Leu Asn Asp Ser Asn Ala Lys Leu Val Arg Asn
1 5 10 15
Asn Asp Leu Leu Leu Ile Asn Leu Ile Leu Leu Leu Pro Leu Ser Val
20 25 30
Phe Val Met Cys Thr Ser Met Leu Phe Val Ser Leu Tyr Lys His Met
35 40 45
His Trp Met Gln Ser Glu Ser His Lys Leu Ser Ser Ala Arg Thr Glu
50 55 60
Ala His Ile Asn Ala Leu Lys Thr Val Thr Thr Phe Phe Cys Phe Phe
65 70 75 80
Val Ser Tyr Phe Ala Ala Phe Met Ala Asn Met Thr Phe Arg Ile Pro
85 90 95
Tyr Arg Ser His Gin Phe Phe Val Val Lys Glu Ile Met Ala Ala Tyr
100 105 110
Pro Ala Gly His Ser Val Ile Ile Val Leu Ser Asn Ser Lys Phe Lys
115 120 125
Asp Leu Phe Arg Arg Met Ile Cys Leu Gln Lys Glu
130 135 140
<210>66
<211>71
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R40,hGR40
<400>66
Ser Gln Tyr Ser Leu Gly His Ser Tyr Val Val Ile Phe Gly Tyr Gly
1 5 10 15
Gln Met Lys Lys Thr Phe Leu Gly Ile Leu Trp His Leu Lys Cys Gly
20 25 30
Leu Lys Gly Arg Ala Leu Leu Ala Thr Gln Val Gly Leu Arg Glu Lys
35 40 45
Ser Thr Arg Ser Leu Gly Val Ile Phe Leu Ala Ser Ser Tyr Ser Phe
50 55 60
Phe Val Tyr Val Leu Cys His
65 70
<210>67
<211>309
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R41,hGR41
<220>
<221>MOD_RES
<222>(253)
<223>Xaa=任何氨基酸
<400>67
Met Ile Thr Phe Leu Leu Ile Ile Leu Ser Ile Leu Val Val Phe Ala
1 5 10 15
Phe Val Leu Gly Asn Phe Ser Asn Gly Phe Ile Ala Leu Val Asn Val
20 25 30
Ile Asp Trp Val Asn Thr Arg Lys Ile Ser Ser Ala Asp Gln Ile Leu
35 40 45
Thr Ala Leu Ala Val Ser Arg Val Gly Leu Leu Trp Val Ile Leu Leu
50 55 60
His Trp Tyr Ala Asn Val Leu Asn Pro Ala Leu Tyr Ser Ser Glu Val
65 70 75 80
Ile Ile Phe Ile Ser Asn Ile Ser Ala Ile Ile Asn His Phe Ser Ile
85 90 95
Trp Leu Ala Thr Ser Leu Ser Ile Phe Tyr Leu Leu Lys Ile Val Asn
100 105 110
Phe Ser Arg Leu Ile Phe His His Leu Lys Arg Lys Ala Lys Ser Val
115 120 125
Val Leu Val Ile Val Leu Gly Pro Leu Val Phe Leu Val Cys His Leu
130 135 140
Val Met Lys His Thr Tyr Ile Asn Val Trp Thr Lys Glu Tyr Glu Gly
145 150 155 160
Asn Val Thr Trp Lys Ile Lys Leu Arg Asn Ala Ile His Leu Ser Asn
165 170 175
Leu Thr Val Ser Thr Leu Ala Asn Leu Ile Pro Phe Thr Leu Thr Leu
180 185 190
Ile Ser Phe Leu Leu Leu Ile Cys Ser Leu Cys Lys His Leu Lys Lys
195 200 205
Met Gln Leu His Ser Lys Gly Ser Gln Asp Pro Ser Thr Lys Val His
210 215 220
Ile Lys Ala Leu Gln Thr Val Thr Ser Phe Leu Met Leu Phe Ala Ile
225 230 235 240
Tyr Phe Leu Tyr Leu Ile Thr Ser Thr Trp Asn Leu Xaa Thr Gln Gln
245 250 255
Ser Lys Leu Val Phe MetPhe Cys Gln Thr Leu Gly Ile Met Tyr Pro
260 265 270
Ser Phe His Ser Phe Ile Leu Ile Met Gly Ser Arg Lys Leu Lys Gln
275 280 285
Thr Phe Leu Ser Val Leu Cys Gln Val Thr Cys Leu Val Lys Gly Gln
290 295 300
Gln Pro Ser Thr Pro
305
<210>68
<211>34
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R42,hGR42
<400>68
Phe Ile Gly Leu Thr Asp Cys Ile Ala Trp Met Arg Asn Gln Lys Leu
1 5 10 15
Cys Met Val Gly Phe Ile Leu Thr Arg Met Ala Leu Ala Arg Ile Asn
20 25 30
Ile Leu
<210>69
<211>297
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R43,hGR43
<220>
<221>MOD_RES
<222>(1)..(297)
<223>Xaa=任何氨基酸
<400>69
Leu Glu Leu Ile Phe Ser Xaa Lys Val Val Ala Thr Arg Gly Leu Val
1 5 10 15
Leu Gly Met Leu Gly Asn Gly Leu Ile Gly Leu Val Asn Cys Ile Glu
20 25 30
Trp Ala Lys Ser Trp Lys Val Ser Ser Ala Asp Phe Ile Leu Thr Ser
35 40 45
Leu Ala Ile Val Arg Ile Ile Arg Leu Tyr Leu Ile Leu Phe Asp Ser
50 55 60
Phe Ile Met Val Leu Ser Pro His Leu Tyr Thr Xaa Xaa Xaa Xaa Xaa
65 70 75 80
Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
85 90 95
Xaa Xaa Ser Leu Ser Ile Phe His Trp Phe Lys Thr Ala Asn Phe Ser
100 105 110
Asn Leu Ile Phe Leu Pro Leu Lys Glu Glu Asp Xaa Asn Val Trp Leu
115 120 125
Gly Asp Ala Val Gly Ala Leu Gly Ile Phe His Leu Xaa Ser Cys Ser
130 135 140
Glu Asn His Gly Xaa Glu Val Cys Gly Gln Lys Asn Met Lys Glu Phe
145 150 155 160
Cys Ser Gly Met Ile Lys Leu Arg Asn Ala Ile Gln Leu Ser Asn Leu
165 170 175
Thr Val Thr Met Pro Ala Asn Val Thr Pro Cys Thr Leu Thr Leu Ile
180 185 190
Ser Phe Leu Leu Leu Ile Tyr Ser Pro Cys Lys His Val Lys Lys Met
195 200 205
Gln Leu His Gly Lys Gly Ser Gln His Leu Ser Thr Lys Val His Ile
210 215 220
Lys Val Leu Gln Thr Val Ile Ser Phe Phe Leu Leu Cys Ala Ile Tyr
225 230 235 240
Phe Val Ser Val Ile Ile Ser Val Trp Ser Phe Lys Asn Leu Glu Asn
245 250 255
Lys Pro Val Phe Met Phe Cys Gln Ala Ile Gly Phe Ser Cys Ser Ser
260 265 270
Ala His Pro Phe Ile Leu Thr Met Gly Asn Lys Lys Leu Lys Gln Thr
275 280 285
Tyr Leu Ser Val Leu Trp Gln Met Arg
290 295
<210>70
<211>319
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R44,hGR44
<400>70
Met Ile Thr Phe Leu Pro Ile Ile Phe Ser Ile Leu Ile Val Val Ile
1 5 10 15
Phe Val Ile Gly Asn Phe Ala Asn Gly Phe Ile Ala Leu Val Asn Ser
20 25 30
Ile Glu Trp Val Lys Arg Gln Lys Ile Ser Phe Val Asp Gln Ile Leu
35 40 45
Thr Ala Leu Ala Val Ser Arg Val Gly Leu Leu Trp Val Leu Leu Leu
50 55 60
His Trp Tyr Ala Thr Gln Leu Asn Pro Ala Phe Tyr Ser Val Glu Val
65 70 75 80
Arg Ile Thr Ala Tyr Asn Val Trp Ala Val Thr Asn His Phe Ser Ser
85 90 95
Trp Leu Ala Thr Ser Leu Ser Met Phe Tyr Leu Leu Arg Ile Ala Asn
100 105 110
Phe Ser Asn Leu Ile Phe Leu Arg Ile Lys Arg Arg Val Lys Ser Val
115 120 125
Val Leu Val Ile Leu Leu Gly Pro Leu Leu Phe Leu Val Cys His Leu
130 135 140
Phe Val Ile Asn Met Asp Glu Thr Val Trp Thr Lys Glu Tyr Glu Gly
145 150 155 160
Asn Val Thr Trp Lys Ile Lys Leu Arg Ser Ala Met Tyr His Ser Asn
165 170 175
Met Thr Leu Thr Met Leu Ala Asn Phe Val Pro Leu Thr Leu Thr Leu
180 185 190
Ile Ser Phe Leu Leu Leu Ile Cys Ser Leu Cys Lys His Leu Lys Lys
195 200 205
Met Gln Leu His Gly Lys Gly Ser Gln Asp Pro Ser Thr Lys Val His
210 215 220
Ile Lys Ala Leu Gln Thr Val Thr Ser Phe Leu Leu Leu Cys Ala Ile
225 230 235 240
Tyr Phe Leu Ser Met Ile Ile Ser Val Cys Asn Leu Gly Arg Leu Glu
245 250 255
Lys Gln Pro Val Phe Met Phe Cys Gln Ala Ile Ile Phe Ser Tyr Pro
260 265 270
Ser Thr His Pro Phe Ile Leu Ile Leu Gly Asn Lys Lys Leu Lys Gln
275 280 285
Ile Phe Leu Ser Val Leu Arg His Val Arg Tyr Trp Val Lys Asp Arg
290 295 300
Ser Leu Arg Leu His Arg Phe Thr Arg Gly Ala Leu Cys Val Phe
305 310 315
<210>71
<211>314
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R45,hGR45
<400>71
Met Ala Thr Glu Leu Asp Lys Ile Phe Leu Ile Leu Ala Ile Ala Glu
1 5 10 15
Phe Ile Ile Ser Met Leu Gly Asn Val Phe Ile Gly Leu Val Asn Cys
20 25 30
Ser Glu Gly Ile Lys Asn Gln Lys Val Phe Ser Ala Asp Phe Ile Leu
35 40 45
Thr Cys Leu Ala Ile Ser Thr Ile Gly Gln Leu Leu Val Ile Leu Phe
50 55 60
Asp Ser Phe Leu Val Gly Leu Ala Ser His Leu Tyr Thr Thr Tyr Arg
65 70 75 80
Leu Gly Lys Thr Val Ile Met Leu Trp His Met Thr Asn His Leu Thr
85 90 95
Thr Trp Leu Ala Thr Cys Leu Ser Ile Phe Tyr Phe Phe Lys Ile Ala
100 105 110
His Phe Pro His Ser Leu Phe Leu Trp Leu Arg Trp Arg Met Asn Gly
115 120 125
Met Ile Val Met Leu Leu Ile Leu Ser Leu Phe Leu Leu Ile Phe Asp
130 135 140
Ser Leu Val Leu Glu Ile Phe Ile Asp Ile Ser Leu Asn Ile Ile Asp
145 150 155 160
Lys Ser Asn Leu Thr Leu Tyr Leu Asp Glu Ser Lys Thr Leu Tyr Asp
165 170 175
Lys Leu Ser Ile Leu Lys Thr Leu Leu Ser Leu Thr Ser Phe Ile Pro
180 185 190
Phe Ser Leu Phe Leu Thr Ser Leu Leu Phe Leu Phe Leu Ser Leu Val
195 200 205
Arg His Thr Arg Asn Leu Lys Leu Ser Ser Leu Gly Ser Arg Asp Ser
210 215 220
Ser Thr Glu Ala His Arg Arg Ala Met Lys Met Val Met Ser Phe Leu
225 230 235 240
Phe Leu Phe Ile Val His Phe Phe Ser Leu Gln Val Ala Asn Trp Ile
245 250 255
Phe Phe Met Leu Trp Asn Asn Lys Cys Ile Lys Phe Val Met Leu Ala
260 265 270
Leu Asn Ala Phe Pro Ser Cys His Ser Phe Ile Leu Ile Leu Gly Asn
275 280 285
Ser Lys Leu Gln Gln Thr Ala Val Arg Leu Leu Trp His Leu Arg Asn
290 295 300
Tyr Thr Lys Thr Pro Asn Pro Leu Pro Leu
305 310
<210>72
<211>59
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R46,hGR46
<400>72
Met Ser Phe Leu His Ile Val Phe Ser Ile Leu Val Val Val Ala Phe
1 5 10 15
Ile Leu Gly Asn Phe Ala Asn Gly Phe Ile Ala Leu Ile Asn Phe Ile
20 25 30
Ala Trp Val Lys Lys Gln Lys Ile Ser Ser Ala Asp Gln Ile Ile Ala
35 40 45
Asp Lys Gln Ser Pro Glu Leu Val Cys Ser Gly
50 55
<210>73
<211>65
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R47,hGR47
<220>
<221>MOD_RES
<222>(1)..(65)
<223>Xaa=任何氨基酸
<400>73
Met Leu Asn Ala Leu Tyr Ser Ile Leu Ile Ile Ile Ile Asn Ile Xaa
1 5 10 15
Phe Leu Ile Gly Ile Leu Gly Asn Gly Phe Ile Thr Leu Val Asn Gly
20 25 30
Ile Asp Trp Val Lys Met Xaa Lys Arg Ser Ser Ile Leu Thr Ala Leu
35 40 45
Thr Ile Ser Arg Ile Cys Leu Ile Ser Val Ile Met Val Arg Trp Phe
50 55 60
Ile
65
<210>74
<211>60
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R48,hGR48
<400>74
Val Ser Arg Val Gly Leu Leu Trp Val Ile Leu Leu His Trp Tyr Ser
1 5 10 15
Thr Val Leu Asn Pro Thr Ser Ser Asn Leu Lys Val Ile Ile Phe Ile
20 25 30
Ser Asn Ala Trp Ala Val Thr Asn His Phe Ser Ile Trp Leu Ala Thr
35 40 45
Ser Leu Ser Ile Phe Tyr Leu Leu Lys Ile Val Asn
50 55 60
<210>75
<211>72
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R49,hGR49
<400>75
Thr Val Thr Met Leu Ala Asn Leu Val Pro Phe Thr Val Thr Leu Ile
1 5 10 15
Ser Phe Leu Leu Leu Val Cys Ser Leu Cys Lys His Leu Lys Lys Met
20 25 30
His Leu His Gly Lys Gly Ser Gln Asp Pro Ser Thr Lys Val His Ile
35 40 45
Lys Val Leu Gln Thr Val Ile Ser Phe Leu Leu Leu Cys Ala Ile Tyr
50 55 60
Phe Val Ser Val Ile Ile Ser Ser
65 70
<210>76
<211>299
<212>PRT
<213>人类(Homo sapiens)
<220>
<223>人T2R50,hGR50
<400>76
Met Ile Thr Phe Leu Pro Ile Ile Phe Ser Ile Leu Val Val Val Thr
1 5 10 15
Phe Val Ile Gly Asn Phe Ala Asn Gly Phe Ile Ala Leu Val Asn Ser
20 25 30
Thr Glu Trp Val Lys Arg Gln Lys Ile Ser Phe Ala Asp Gln Ile Val
35 40 45
Thr Ala Leu Ala Val Ser Arg Val Gly Leu Leu Trp Val Leu Leu Leu
50 55 60
Asn Trp Tyr Ser Thr Val Leu Asn Pro Ala Phe Tyr Ser Val Glu Leu
65 70 75 80
Arg Thr Thr Ala Tyr Asn Ile Trp Ala Val Thr Gly His Phe Ser Asn
85 90 95
Trp Pro Ala Thr Ser Leu Ser Ile Phe Tyr Leu Leu Lys Ile Ala Asn
100 105 110
Phe Ser Asn Leu Ile Phe Leu Arg Leu Lys Arg Arg Val Lys Ser Val
115 120 125
Ile Leu Val Val Leu Leu Gly Pro Leu Leu Phe Leu Ala Cys His Leu
130 135 140
Phe Val Val Asn Met Asn Gln Ile Val Trp Thr Lys Glu Tyr Glu Gly
145 150 155 160
Asn Met Thr Trp Lys Ile Lys Leu Arg Arg Ala Met Tyr Leu Ser Asp
165 170 175
Thr Thr Val Thr Met Leu Ala Asn Leu Val Pro Phe Thr Val Thr Leu
180 185 190
Ile Ser Phe Leu Leu Leu Val Cys Ser Leu Cys Lys His Leu Lys Lys
195 200 205
Met Gln Leu His Gly Lys Gly Ser Gln Asp Pro Ser Thr Lys Val His
210 215 220
Ile Lys Val Leu Gln Thr Val Ile Ser Phe Phe Leu Leu Cys Ala Ile
225 230 235 240
Tyr Phe Val Ser Val Ile Ile Ser Val Trp Ser Phe Lys Asn Leu Glu
245 250 255
Asn Lys Pro Val Phe Met Phe Cys Gln Ala Ile Gly Phe Ser Cys Ser
260 265 270
Ser Ala His Pro Phe Ile Leu Ile Trp Gly Asn Lys Lys Leu Lys Gln
275 280 285
Thr Tyr Leu Ser Val Leu Trp Gln Met Arg Tyr
290 295
<210>77
<211>335
<212>PRT
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R01,rGR01
<400>77
Met Met Glu Gly His Ile Leu Phe Phe Phe Leu Val Val Met Val Gln
1 5 10 15
Phe Val Thr Gly Val Leu Ala Asn Gly Leu Ile Val Val Val His Ala
20 25 30
Ile Asp Leu Ile Met Trp Lys Lys Met Ala Pro Leu Asp Leu Leu Leu
35 40 45
Phe Cys Leu Ala Thr Ser Arg Ile Ile Leu Gln Leu Cys Ile Leu Phe
50 55 60
Ala Gln Leu Cys Leu Phe Ser Leu Val Arg His Thr Leu Phe Glu Asp
65 70 75 80
Asn Ile Thr Phe Val Phe Ile Ile Asn Glu Leu Ser Leu Trp Phe Ala
85 90 95
Thr Trp Leu Gly Val Phe Tyr Cys Ala Lys Ile Ala Thr Ile Pro His
100 105 110
Pro Leu Phe Leu Trp Leu Lys Met Arg Ile Ser Arg Leu Val Pro Trp
115 120 125
Leu Ile Leu Gly Ser Val Leu Tyr Val Ile Ile Thr Thr Phe Ile His
130 135 140
Ser Arg Glu Thr Ser Ala Ile Leu Lys Pro Ile Phe Ile Ser Leu Phe
145 150 155 160
Pro Lys Asn Ala Thr Gln Val Gly Thr Gly His Ala Thr Leu Leu Ser
165 170 175
Val Leu Val Leu Gly Leu Thr Leu Pro Leu Phe Ile Phe Thr Val Ala
180 185 190
Val Leu Leu Leu Ile Tyr Ser Leu Trp Asn Tyr Ser Arg Gln Met Arg
195 200 205
Thr Met Val Gly Thr Arg Glu Tyr Ser Gly His Ala His Ile Ser Ala
210 215 220
Met Leu Ser Ile Leu Ser Phe Leu Ile Leu Tyr Leu Ser His Tyr Met
225 230 235 240
Val Ala Val Leu Ile Ser Thr Gln Val Leu Tyr Leu Gly Ser Arg Thr
245 250 255
Phe Val Phe Cys Leu Leu Val Ile Gly Met Tyr Pro Ser Ile His Ser
260 265 270
Ile Val Leu Ile Leu Gly Asn Pro Lys Leu Lys Arg Asn Ala Lys Met
275 280 285
Phe Ile Val His Cys Lys Cys Cys His Cys Thr Arg Ala Trp Val Thr
290 295 300
Ser Arg Ser Pro Arg Leu Ser Asp Leu Pro Val Pro Pro Thr His Pro
305 310 315 320
Ser Ala Asn Lys Thr Ser Cys Ser Glu Ala Cys Ile Met Pro Ser
325 330 335
<210>78
<211>1331
<212>DNA
<213>大鼠
<220>
<223>大鼠T2R01,rGR01
<400>78
caggaatcat aaatggctga aactgggcag aactctatgc attatttaaa gaagtcattg 60
gtttgtcatt cttaaaatga tggaagggca tatactcttc ttctttttgg ttgtgatggt 120
gcagtttgtc actggggtct tggcaaatgg cctcattgtg gttgtccatg ctattgactt 180
gatcatgtgg aagaaaatgg ccccgttgga tctgcttcta ttttgcctgg cgacttctcg 240
gatcattctg cagttatgta tattgtttgc acaattgtgt ctattctctt tggtgagaca 300
cactttattt gaggacaata ttacctttgt cttcatcata aatgaactga gtctttggtt 360
tgctacatgg ctcggtgttt tctactgtgc caagattgct accattcctc acccactctt 420
tctgtggctg aagatgagga tatccaggtt ggtaccatgg ctgatcctgg gatctgtgct 480
ctatgtaatt attactactt tcatccatag cagagagact tcagcaatcc ttaaaccaat 540
ttttataagc ctttttccta aaaatgcaac tcaagtcgga acagggcatg ccacactact 600
ctcagtcctg gtccttgggc tcacactgcc gttgttcatc tttactgttg ctgttctgct 660
cttgatatac tccctgtgga attatagcag gcagatgagg actatggtag gcaccaggga 720
gtatagcgga catgctcaca tcagtgcaat gctgtccatt ctatcattcc tcatcctcta 780
tctctcccac tacatggtgg ctgttctgat ctctactcaa gtcctctacc ttggaagcag 840
aacctttgta ttctgcttac tggttattgg tatgtacccc tcaatacact cgattgtctt 900
aattttagga aatcctaagc tgaaacgaaa tgcaaaaatg ttcattgtcc attgtaagtg 960
ttgtcattgt acaagagctt gggtcacctc aaggagccca agactcagtg acttgccagt 1020
gcctcctact catccctcag ccaacaagac atcctgctca gaagcctgta taatgccatc 1080
ctaattgtcc agcctgaggt ttaatcctag gtttggtact atttcaaaga gtaaagttga 1140
tcattaaagc acaacatatg ttggtggatg acatcaaggt ccatatccca gttgtcaatt 1200
gtaaacctca ccttgcaaga tgatgtcact gagaaagcag gacaaatgga gtctaggtcc 1260
ttctgtatga cttgctgcag tatatgtgaa tctataattt tctccaaaaa aacaaaaaaa 1320
aaaaaaaaaa a 1331
<210>79
<211>333
<212>PRT
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R02,rGR02
<400>79
Met Phe Ser Gln Lys Thr Asn Tyr Ser His Leu Phe Thr Phe Ser Ile
1 5 10 15
Ile Phe Tyr Val Glu Ile Val Thr Gly Ile Leu Gly Asn Gly Phe Ile
20 25 30
Ala Leu Val Asn Ile Met Asp Trp Leu Lys Arg Arg Arg Ile Ser Thr
35 40 45
Ala Asp Gln Ile Leu Thr Ala Leu Ala Leu Thr Arg Leu Ile Tyr Val
50 55 60
Trp Ser Val Leu Ile Cys Ile Leu Leu Leu Phe Leu Cys Pro His Leu
65 70 75 80
Ser Met Arg Pro Glu Met Phe Thr Ala Ile Gly Val Ile Trp Val Val
85 90 95
Asp Asn His Phe Ser Ile Trp Leu Ala Thr Cys Leu Gly Val Phe Tyr
100 105 110
Phe Leu Lys Ile Ala Ser Phe Ser Asn Ser Leu Phe Leu Tyr Leu Lys
115 120 125
Trp Arg Val Lys Lys Val Val Leu Met Ile Ile Leu Ile Ser Leu Ile
130 135 140
Phe Leu Met Leu Asn Ile Ser Ser Leu Gly Met Tyr Asp His Phe Ser
145 150 155 160
Ile Asp Val Tyr Glu Gly Asn Met Ser Tyr Asn Leu Val Asp Ser Thr
165 170 175
His Phe Pro Arg Ile Phe Leu Phe Thr Asn Ser Ser Lys Val Phe Leu
180 185 190
Ile Ala Asn Ser Ser His Val Phe Leu Pro Ile Asn Ser Leu Phe Met
195 200 205
Leu Ile Pro Phe Thr Val Ser Leu Val Ala Phe Phe Val Leu Phe Leu
210 215 220
Ser Leu Trp Lys His His Lys Lys Met Gln Val Asn Ala Lys Gly Pro
225 230 235 240
Arg Asp Ala Ser Thr Met Ala His Thr Lys Ala Leu Gln Ile Gly Phe
245 250 255
Ser Phe Leu Leu Leu Tyr Ala Ile Tyr Leu Leu Phe Ile Ile Thr Gly
260 265 270
Ile Leu Asn Leu Asp Leu Met Arg Cys Ile Val Ile Leu Leu Phe Asp
275 280 285
His Ile Ser Gly Ala Val Phe Ser Ile Ser His Ser Phe Val Leu Ile
290 295 300
Leu Gly Asn Ser Lys Leu Arg Gln Ala Thr Leu Ser Val Leu Pro Cys
305 310 315 320
Leu Arg Cys Arg Ser Lys Asp Met Asp Thr Val Val Phe
325 330
<210>80
<211>2438
<212>DNA
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R02,rGR02
<400>80
attttgctcc actattttgc tcttctgcag taacacagac cacaaaacaa tggagccaat 60
gggtcaagag ctgaaacttc aggaagtggg agccaaattt tctttgtgat aggttggcat 120
atgagaattc attatttgat gcagcttctg aaaactggat gtgaaatact ggatgaagca 180
gaggtgatga cccctttgaa attaaaaagc caagatgttc atggagaaat tataaaacaa 240
tatctgggaa atttgatgct tcctaatcgg gtgtaaatgg gattttaaat gatgaacatt 300
ttgaatttcc aatgaccatt atgtaaagtt tttaaacaca gtagagacat cataaattga 360
agcatgttct cacagaaaac aaactacagc catttgttta ctttttcaat tattttttat 420
gtggaaatag taacaggaat cttaggaaat ggattcatag cactagtgaa tatcatggac 480
tggctcaaga ggaggaggat ctctactgca gatcagattc tcactgcttt ggcccttacc 540
agactcattt atgtgtggtc tgtactcatt tgtatattgt tactatttct gtgcccacat 600
ttgtctatga gaccagaaat gtttacagcg ataggtgtta tctgggtagt ggataaccac 660
ttcagcatct ggcttgctac atgtcttggt gtcttttatt tcctcaaaat agccagtttt 720
tctaactctt tgtttcttta cctaaagtgg agagttaaaa aagtggtttt aatgataata 780
ctgatatcac tgattttctt gatgttaaac atttcatcat tagggatgta tgatcatttc 840
tcaattgatg tttatgaagg taatatgtct tataatttgg tggattcaac acattttccc 900
agaattttct tattcacaaa ctcatctaag gtcttcttaa tcgccaattc atcccatgtt 960
ttcttaccca tcaactcact cttcatgctc atacccttca cagtttccct ggtagctttt 1020
ttcgtgctct ttctctcact gtggaagcat cacaagaaga tgcaggtcaa tgccaaagga 1080
cccagagatg ccagcaccat ggcccacaca aaagccttgc aaattgggtt ctccttcctc 1140
ctgctgtatg caatatactt acttttcatt atcacaggaa ttttgaacct tgacttgatg 1200
agatgtatag taatactttt atttgaccac atatctggag cagttttttc tataagccac 1260
tcatttgtgc tgattctggg aaacagtaag ctgagacaag ccactctttc tgtgctgcct 1320
tgtcttaggt gccggtccaa agatatggac actgtcgttt tctaataaat tccagagtac 1380
attatgcaaa atcttgaggg tgatcagttc atagaaaaag taatcttaga ggggaaaata 1440
aaatattggg gcttcaaatg ttggatgggt aatacatagg aaggcaggac aaggatgaag 1500
gagactagca ttatataagt gatttcacag gggaaatggg aaagagggct tttatataat 1560
gaagaagaag ataaatgatg aaggatgagg aagagttaaa tatgtaaaat gacaatagag 1620
atggcatcat gccgttttaa gaaatttgga atgcatatgt atgtttatat attttttaat 1680
ttttattgaa tatatttatt tacattttaa atgttatcct gtttccccca cccaacctcc 1740
cacctcttcc cacctccttg ccctgacatt cccctgcact ggggaatcca gccttgacag 1800
gaccaagggc ttctcctccc tttgttgcca acaaggccat tctttgctac atgtgcagca 1860
ggagccatgg atctgtctat gtgtactctt tggatggtgg tttagtccct gggagctctt 1920
gttggttggt attgttgttc ttatggtgtt gcaactccct tcagctcctt caatccttcc 1980
tgtaactcct ccaatgtgga ccctgttctc agtccaatgg ttgactatga gcattcacct 2040
ctgtgattgt catgctctgg cacagcttct cagaagacag ctacatcagt ctcctataag 2100
agtgcacttc atggcatcag caatgttgtc ttgatttggt gtctgtatgt atatgggctg 2160
gatcccaggt ggggcaggcg ctgaatggtc attccttcag tctttgctcc aaactttgtc 2220
tttatatctc ctatgaatat ttttgttccc ccttataaga atgactgaag tatccacact 2280
ttggccatcc ttcttcatga gcttcatgtg gtctgtgaat tgtacattgt gtaatccaag 2340
cttttgggct aatatccaat tatagtgagt gcataccaaa aaaaaaaaaa aaaaaaaaaa 2400
aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaa 2438
<210>81
<211>299
<212>PRT
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R03,rGR03
<400>81
Met Val Pro Thr Gln Val Thr Ile Phe Ser Ile Ile Met Tyr Val Leu
1 5 10 15
Glu Ser Leu Val Ile Ile Val Gln Ser Cys Thr Thr Val Ala Val Leu
20 25 30
Phe Arg Glu Trp Met His Phe Gln Arg Leu Ser Pro Val Glu Ile Ile
35 40 45
Leu Ile Ser Leu Gly Ile Ser His Phe Cys Leu Gln Trp Thr Ser Met
50 55 60
Leu Tyr Asn Phe Gly Thr Tyr Ser Arg Pro Val Leu Leu Phe Trp Lys
65 70 75 80
Val Ser Val Val Trp Glu Phe Met Asn Val Leu Thr Phe Trp Leu Thr
85 90 95
Ser Leu Leu Ala Val Leu Tyr Cys Val Lys Val Ser Ser Phe Ser His
100 105 110
Pro Val Phe Leu Trp Leu Arg Leu Lys Ile Leu Lys Leu Val Leu Trp
115 120 125
Leu Leu Leu Gly Ala Leu Ile Ala Ser Cys Leu Ser Ile Ile Pro Ser
130 135 140
Val Val Lys Tyr His Ile Gln Met Glu Leu Leu Thr Leu Asp His Leu
145 150 155 160
Pro Lys Asn Ser Ser Leu Ile Leu Arg Leu Gln Met Phe Glu Trp Tyr
165 170 175
Phe Ser Asn Pro Phe Lys Met Ile Gly Phe Gly Val Pro Phe Leu Val
180 185 190
Phe Leu Ile Ser Ile Ile Leu Leu Thr Val Ser Leu Val Gln His Trp
195 200 205
Gly Gln Met Lys His Tyr Ser Ser Ser Ser Ser Ser Leu Arg Ala Gln
210 215 220
Cys Thr Val Leu Lys Ser Leu Ala Thr Phe Phe Ile Phe Phe Thr Ser
225 230 235 240
Tyr Phe Leu Thr Ile Val Val Ser Phe Ile Gly Thr Val Phe Asp Lys
245 250 255
Lys Ser Trp Phe Trp Val Cys Glu Ala Val Ile Tyr Gly Leu Val Cys
260 265 270
Ile His Phe Thr Ser Leu Met Met Ser Asn Pro Thr Leu Lys Lys Ala
275 280 285
Leu Arg Leu Gln Phe Trp Ser Pro Glu Ser Ser
290 295
<210>82
<211>6552
<212>DNA
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R03,rGR03
<220>
<221>modified_base
<222>(1)..(6552)
<223>n=g,a,c或t
<400>82
gcatggtgcc aacccaagtc accatcttct ctatcatcat gtatgtgctt gagtccttag 60
tcataattgt gcaaagttgc acaacggttg cagtgctgtt cagagagtgg atgcactttc 120
aaagactgtc gccggtggaa ataattctca tcagcctggg catttcacat ttctgtctac 180
agtggacatc gatgctgtac aactttggta cctactctag gcctgtcctt ttattttgga 240
aggtatcggt cgtctgggag ttcatgaacg ttttgacatt ctggctaacc agtttgcttg 300
ctgtcctcta ctgtgtcaag gtctcttcct tctctcaccc cgtcttcctc tggctgaggt 360
tgaaaatttt gaaactggtt ctctggttgc tattgggcgc tctgatagct tcttgtttgt 420
caatcatccc ttctgttgtt aaatatcata tccagatgga attactcacc ctagatcatt 480
tacccaaaaa cagttctttg attctaagac tgcaaatgtt cgagtggtat ttttctaatc 540
ctttcaaaat gattgggttt ggcgttcctt tcctcgtgtt cctgatttct atcatcttac 600
tcacagtctc gctggtccag cattgggggc agatgaaaca ctacagcagc agcagctcca 660
gcctgagagc tcagtgcact gttctgaagt ctcttgccac cttcttcatc ttcttcacat 720
cctattttct gactatagtc gtctccttta ttggcaccgt gtttgataag aagtcatggt 780
tctgggtctg cgaagctgtc atctatggtt tagtctgtat tcacttcact tccctgatga 840
tgagcaaccc tacactgaaa aaagcactca ggttgcagtt ctggagccca gagtcttcct 900
aaggcaggga attcagtgaa gcctctgggg taaggaggct ttgcattggc acagttctta 960
gagtgaaatg caaacgtgga cacgaacttc attctctttc atgtccacag atggatggat 1020
ctataaatca tcaccaatct tccctgtatt ctgacccatc cttttcctgt cctatccata 1080
gtccccaggt tggttttgat ttttctcatg atcacacctt agctttagcc accgttgcaa 1140
tatcaaacat gatctatatg ttacagccaa aatcattctc acaattgtca attgcttcac 1200
aaattcagat aaatccccct tcctgtcagg aatgtattgt ctgtgcattc aatgctcacc 1260
atgctaagcc attcattccc ttcctaactt gagtttaaga agaaaatgtc ttactgttgc 1320
ccatgtccta ttgtgctgct tctggatgtt ttatgcagtg atttagacac acgcccttgc 1380
ctgtctccaa atactggccc tttattcctt tataagtcta gtagaaaatg aactcgtctt 1440
tacttcattg acgaagacat tgtattcttc cccaaaatag tgtttaacta ctctagtctc 1500
atccataata tccctaaata tcagtgattt cagtgagtaa aacctgacaa cagttattgc 1560
tttgactctt aattcaattg tgctgtaaca tagaggaaac attctagaac atttccatat 1620
taatttgtgc ttgtagcaaa ccaaaattct ccccagttgg gtaaaaatat caaaagcaca 1680
gagtaatcaa ttttgaaatc actcagaaga catcattgtt ctatatatgt tttttttaaa 1740
cttccctcta acaagtatca gatctttgcc tttacagggt ctggtcttac catgactata 1800
ttttatcacc atgacctatt ttctcttcat ctctttgttt tcactaactc agtagcaacc 1860
aaatatcaca ttaatagcta actctgggca cttatttctc agcctttatc tattccagac 1920
actttcaatg tatttctgct aaacacaatg acatctcttt ttgtgttcta acgacaagga 1980
atcataactt tccaactttt atacatggta gacatatttg gtgaacttaa cttctgactc 2040
tttctttaga agactgaaac tactccggaa agcaagcctt ctgatggaga aatagatacg 2100
ggtatcgtga ttcattgtga aagtgaattc cggtgcctgg aaagaaatgg atattttttt 2160
ttctcttgag tgtgtcactc tgacatatgt tccatgttga atccatattt gatactgata 2220
gcatgaatgt aagtaaagca tgtatgtaag taaagactgc taccaaaact tcgattcaac 2280
tttcctcagc agtatccctg atattgcata agaaagaaaa aacacgctgt cctacttgaa 2340
gaaggacgtg ttccatgcaa tgtggatgtg tcccaggcta cattggctca actgcagctg 2400
aaggtgggat gggaaatggt atagttagta atgtctgctg agctgtctca ctggaaagga 2460
ttctgagcag agtaaatgta agcaatgtgg ccaaggtctc ctaggaatgg gttgtaagct 2520
tgtaaggagt tgggttgtaa gagtttggga tcctttcaga atggattgag caagagccac 2580
tgaaacttgg actatacctt tgttatttgt atctaaatcc agaagggtct ttgcatgttc 2640
caaaatctca gatagctgga aggaagaagg actgttctct ttacaagtat ataaatagag 2700
aatgagctaa aaaggacccc ctcacccccg ccgtcacaca caggaatact attccagaaa 2760
ctagggagta tttttagtgt tctcactatt tccctttgaa aaaagtgcaa tggaaaactt 2820
atccatgaca tacatgaggt tggagtgata aaaacagctg aaggaagagg aagtctgaaa 2880
aaagatggaa acagcaatga tgcttgtcct atatatgtgt gacacccact agttcccaag 2940
gaaaccttac atccattatc tcatttcaag ctggaaggac aagtcaagat cactcaaccg 3000
acccagctgg aaaacagacc taagaatgtt aaactcatac tgatggttat ttctcactct 3060
aaagtcaatg caaatggata gcaaacaaag gggctatttt tttaagggac cagagggttt 3120
caatctagaa tcagagaaaa gataaaaagg gagatgctat agaaaaacaa tagagaagat 3180
gtggccaaga acaaggaaaa tctccagtta gcttggcact taggggccaa catgtttctg 3240
ttgttcggtc ttcaatactg tattgcatgt tgggctcact atgttttagt tgtgagtggg 3300
ttgtgcttcc tggaattaag aaaggtctgt ttctagattt caggtacaaa tgtttagaag 3360
cccattggta gcatcagtga aattaggaaa aaactgtgag cactgctggc tggacttggc 3420
aaagtcattc actatttaca catcaaatta ttagcaactt gaaagtaaat ctttgctcat 3480
catccagtgg cccccatgat cctggtgaat gacttgtaat actgtggaga ctggcaacga 3540
cggtgaattc ctagtaacac ttaccataga atctgttcat aattagactc gcccagattt 3600
tagttgctag agaacaatct ttctccttta cccacattcc tactgagtag gatgcatagg 3660
ttcggaaacc cccatggcat cgtttgactc ctcctggtag tcaagagagt ccagtcacca 3720
gtctccgaaa cacctgccaa gtcctaactc ccaacagtct acagtgtaaa cctcagtgtt 3780
tgcatgaggt ttatgtatct ccttaccatt tcctaaatgt caatacccgt gcacaggata 3840
tttgcatagg ctgcctccaa gcctgggaaa cactctcctc ctcgcatttg ctgggtttca 3900
cctttccaat tcagtgtgcc ctttaaaagg cactgctttt ctaggcccac cactattgct 3960
gctcacgcat gaacatcaaa tctaccacag gcttttgcct ctcagaatta ttcttctttc 4020
tactatgcaa tgtggtatcc atgagaactt tgtcacattg tcaaattcta cctttgtttt 4080
aatgngngcc tttgtaatag ngactatgcc cagaaattaa attatagtaa gatgggtaac 4140
aacncttcaa ttntggaatt tataattaaa taaatattat gtaatattat gacttattat 4200
aangtcaatc tactgtaccc tactcctact aggaatgcaa agacaaatag caatgtgatc 4260
agcatgtgct ctttcacaag atcatattgt gcatgttgct gatgatgccc acagtgcatc 4320
tatcagaata tctctgatca tttttttttt tttgcttttg agaagccccg gttggtgctg 4380
ggatgcttca tagcaggtcc accatagaca catgcttaga ggaaagctgc ctctctctct 4440
tcattcccaa ggaacagtaa aagcagaaaa ggctcttatg ttctaaagaa cagaaaatag 4500
cctgcatttc aactacctcc tgttcagaag gcaccgaaac acaccaccaa gcaagacacc 4560
cctttacttt ctcctgcttc cctcaatttg atgatcattt ggaaataaga agaaagaaaa 4620
agatgtggaa gccaattaaa aacagtcttg tctatctccc tggtgagctc tcaacttctt 4680
agtcagacca aagtaggtga aaaaataata atttttaatt tggtatgaga gtcatgttta 4740
ggctgaaaat cttaaaaaat cttagcataa aaacattttc ccctagaccc atgaaattta 4800
taatattatc tgtggttgag aaaggctagt tatagaaaaa tgtttagaat cagaatattt 4860
tgagggctct ttttttgttt tgcctaatca ttacatttgt tataagaagt ctaaaagttg 4920
gtatgctaca ggtcttgtca tattttctct gaggttgagt gccaagtagt ctgcattgtg 4980
tttaaatcct gcttaaaatt atcccaagac aatataactt ctcaggagct aagccaaggg 5040
cccctttcag actaccttag tcctctctca ccgttgtcac cgtggctcat acatcagaat 5100
cctgagggag catcatgaaa tctaaggctt tacaacagaa tctttctatc cctggtagaa 5160
atcttttaac cttgggtttt attctcatgc cattctgatg ctcgtattta aattttatgt 5220
gttttttcat atgttcttgc atttctatcg ttaaattatg gtgacatact ttcaaatgct 5280
ttgttatttt aaaaagggac aaagagagat agaaagacag ggaaagatag acagaggctt 5340
gcctaataca gtcaagaaag aagctatcaa aagtatttag caatacaaca tttatgatat 5400
attcataact gttaaccatt tttaatattc taaaatttca cttttgtttc agaaatgtat 5460
attaagagaa tctgagaaac atttttttct catagatgta gaaaaacaca caaaataagg 5520
tataacacat ttaagtgatt gaaaataaaa acaaaagctt gcaaacagga ggaaaagtac 5580
attgtaggct ttcgacatgg agctgctact aggacccagg acttgtttat catttatttg 5640
ccaagtccca caaactcagg gcaatacatc tctgagacag tttcctatat tttaataaaa 5700
cttccaaaat tgatactcag tgtgaattgg ctagctttaa tggcagtcat tggataaaca 5760
attccaatgc caaatttccc taagttgata tatttgatta atatgtatat taaaacatca 5820
ggctatccat cggttggatc aaatacattc tttagggatc cattcttttc cttaaatttg 5880
acttatatgt ggattctttt cacaataaat aagtaaatga gcatttattt taaaactatt 5940
ttagacggaa ctgaattaca gccaaggtag tcaaaatgac tgagaataat cacttacata 6000
tttacaaggg aaagtgactc ttcagattta agtttaaaat tagaagagag ataaatttca 6060
caagctttca ctcctaaggc taaagatagg ctgtgtaggt agttatttct gagcacattg 6120
gcacatcacc attgtcagta cttgagggtt tgaatgaagc tcactcaaag aacttggaaa 6180
gaaggtggtc ttctgacatc aatcaagaaa caagctttcc tccctacttc ttccctaaat 6240
gcaacaacct aagaattatc cacaagatgg atggcgcaag ggttcctcaa tcaatttcag 6300
gatgtacatc aatgcgcagc ctatactaca ccgaaaagga agcgcatggg tcttaaaaag 6360
taaaggggat atcaaaaaat tcgcaaccaa acaaaaagtg gcacacattt aagctaggtc 6420
tatgtttggt cagttacacc tggagaaggg ggacatttgg tcagctcatt cgaacactgt 6480
caagtcctac caacaattcc tctatgctat tacccattaa acctcaggtc tcatcgaaaa 6540
aaaaaaaaaa aa 6552
<210>83
<211>308
<212>PRT
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R04,rGR04
<400>83
Met Leu Ser Ala Ala Glu Gly Ile Leu Leu Cys Val Val Thr Ser Glu
1 5 10 15
Ala Val Leu Gly Val Leu Gly Asp Thr Phe Ile Ala Leu Ala Asn Cys
20 25 30
Met Glu Tyr Ala Lys Asn Lys Lys Leu Ser Lys Ile Gly Phe Ile Leu
35 40 45
Ile Gly Leu Ala Ile Ser Arg Ile Gly Val Val Trp Ile Ile Ile Leu
50 55 60
Gln Gly Tyr Met Gln Val Phe Phe Pro His Ile Leu Thr Phe Gly Asn
65 70 75 80
Ile Thr Glu Tyr Ile Thr Tyr Ile Trp Val Phe Leu Asn His Leu Ser
85 90 95
Val Trp Phe Ala Thr Asn Leu Asn Ile Leu Tyr Phe Leu Lys Ile Ala
100 105 110
Asn Phe Ser Asn Ser Val Phe Leu Trp Leu Lys Ser Arg Val Arg Val
115 120 125
Val Phe Ile Phe Leu Ser Gly Cys Leu Leu Thr Ser Trp Leu Leu Cys
130 135 140
Phe Pro Gln Phe Ser Lys Met Leu Asn Asn Ser Lys Met Tyr Trp Gly
145 150 155 160
Asn Thr Ser Trp Leu Gln Gln Gln Lys Asn Val Phe Leu Ile Asn Gln
165 170 175
Ser Leu Thr Asn Leu Gly Ile Phe Phe Phe Ile Ile Val Ser Leu Ile
180 185 190
Thr Cys Phe Leu Leu Ile Val Phe Leu Trp Arg His Ile Arg Gln Met
195 200 205
His Ser Asp Gly Ser Gly Leu Arg Asp Leu Asn Thr Glu Ala His Val
210 215 220
Lys Ala Met Arg Val Leu Ile Ser Phe Ala Val Leu Phe Ile Leu His
225 230 235 240
Phe Val Gly Leu Ser Ile Gln Val Leu Cys Phe Phe Leu Pro Gln Asn
245 250 255
Asn Leu Leu Phe Ile Thr Gly Leu Ile Ala Thr Cys Leu Tyr Pro Cys
260 265 270
Gly His Ser Ile Ile Leu Ile Leu Gly Asn Lys Gln Leu Lys Gln Ala
275 280 285
Ser Leu Lys Ala Leu Gln His Leu Thr Cys Cys Glu Thr Lys Arg Asn
290 295 300
Leu Ser Val Thr
305
<210>84
<211>3449
<212>DNA
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R04,rGR04
<400>84
tggttccatc acatgacaat aggcttgaaa aacttgcaga tagagaagac ataacccctc 60
caacaagaag ccaacatatg ggacattctc cagcagataa tttataacag atgcaacggg 120
agcaacttcg agatctgcaa agatgctgag tgcagcagaa ggcatcctcc tttgtgttgt 180
cactagtgag gcagtgctgg gggttttagg agacacattc attgcacttg caaactgcat 240
ggagtatgcc aagaacaaga agctctctaa gattggtttc attctcattg gcttggcgat 300
ttccagaatt ggtgtcgtat ggataataat tttacagggg tatatgcaag tattttttcc 360
acacatactt acctttggaa acataactga atatattact tacatatggg tgtttctcaa 420
tcacttaagt gtctggtttg ctaccaacct caatatcctc tactttctaa agatagcaaa 480
tttttccaac tctgtatttc tctggctgaa aagtagagtc cgtgtggttt ttatctttct 540
gtcaggatgc ttacttacct cgtggttact atgttttcca caattttcaa agatgcttaa 600
caacagtaaa atgtactggg gaaacacgtc ttggctccag cagcagaaaa atgtcttcct 660
tattaaccaa agtttaacca atctgggaat cttctttttc attattgtat ccctgattac 720
ctgcttcctg ttgattgttt tcctctggag acacatcagg caaatgcact cagatggttc 780
aggactcaga gacctcaaca cagaagctca tgtgaaagcc atgagagttc taatatcttt 840
tgcggtactc tttatcctgc atttcgtagg tctttccata caagtgctat gcttttttct 900
gccacaaaac aacctactct ttataactgg tttgatagcc acatgcctct atccctgtgg 960
tcactcaatc atcttaattc taggaaacaa gcagctgaag caagcctcct tgaaggcact 1020
gcagcactta acgtgctgtg agacaaaaag aaatctctca gtcacataaa tgggtttgcc 1080
aattaatatc tgccatgtta ttccactgat ttttacctgt tagtttctct gtgtctctgt 1140
ttagtttctg tttccatgat ctgtccattg atgagcgtgg ggtgttgaaa tctccgacta 1200
ttgttgtgtg agatgaaatg tgtgctttga gctttagtaa gatttctttt gtgaatgtag 1260
gtgcttttgc atttggtgca tagatattta agattgagag ttcagcttgg tggatttttc 1320
ctttgatgaa tatgaagtgt ccttgcttat cttttttgat gacttttgat tgaacgtcaa 1380
ttttattgga tattagattg gcaactcaag attgcttctt gaggtcattt gcttggaaag 1440
ttgtttttca gccatttact ctgaggtagt gtctgtcttt gtctctgagg tgtgtttcct 1500
gcattcagca aaatgctggg tcctctttac atatccagtt tgttagtcta tgtcttttta 1560
ttggggaatt gagtccattg atgttgagag atattaatga atagtgatca ttgcttcctg 1620
ttattttcgt tgttagatgt ggaattatgt ttgtttgtct ctcttttggt tttattgcaa 1680
ggaaattata tacttgcttt ctgtatggtg tagtttctct ccttgtgttg cagttttcct 1740
tctattatcc tttgtagggc tagatttgaa gaaagatatt gcataagctt ggttttgtca 1800
tgggatatct tggtttctcc atctatgtta attgagagtt ttgcaggata tagtagcctg 1860
ggatgacatt tgtgttctct tagggtctgt atgacatctg tccaaaatct tctggctttc 1920
atagtctctg gtgagaaatc ggatgtaatt ctcataagtc tgccattata tgtcacttga 1980
cctttttccc ttattgcttt ttatgttctt tctttgtttt gtgcatttgg tgttctgatt 2040
attatgtgat gtgaggtatt tctcttctgg tcaaatctat ttggagttct gtaggcttct 2100
tgtatgttta tgggcatctc tttctttagg ttatggatgt tttcttctat aattttgttg 2160
aatatatcta ctgtcccttt aagttaggag ccttcacttt cttctatacc tgttatcctt 2220
aggtttaatc ttctcactgg atttcctcga tgttttggac taggaacttt ttgcatttta 2280
cattatcttt gacaggtatt tcaatgtttt ctatggtatc ttctgccact gagattctct 2340
cttctagctc ttgtataatg ttggtgatgc ttgtacctgt gactccttgt ttcttcctta 2400
ggttttctat ctccagggtt gtctcccttt gtgctttttt tattgcttct atttccattc 2460
taaatcctgg atggttttgt tcaattcctt cacctctttg gttgtatttt cctgtaattc 2520
tttcagggat ttttgtgttt cctctttaag ggcttctact tgtttacttg tgttgtcctg 2580
tatttcttta aggtagttat ttatgtcctt cttgaagtcc tccatcatta tcaaaaaatg 2640
tgatttttaa atataaacct tgcttttctg gtgtgtttgg atgtcaagta ttttctttgc 2700
tgggagaact gggctctgat aatgccaagt tgtttgattt ctgttgctta gtttcctgtt 2760
cttgcctctc gccattgggt tttctctggt gtttgcttat cttgctgttt ctgagagtgg 2820
cttgacactc ttgtaggcat ctgtgtcagg cctcctgtag aactgtttcc ctgttttctt 2880
tcagcctttt ctgagaacag gtgctctgat ctcaggtgtg taggcattcc tggtgactat 2940
ctttcagctt taggagcagg caggaatcag aagggtcctg tccctgactg ctcctagatc 3000
cttgcaccca gggggcacag ttagcactag gcaattccct cttgtgtagg gaatgtgggt 3060
agaggatagt cgcctctgat ttctcaggaa tgtctgcact tctgaaagtc cagccctctc 3120
ccccacagga tttaggtgca gggagctgtt tgaccacttc aattcagtcc tgggtgtaga 3180
ccagaaccac aggtaaaaaa gaatgacttc attaaattag cagacaaatg ggtggaacta 3240
gaaaatgtca tcctgggctg gagagatggc tcagtggttc agaccactgg ctgctcttcc 3300
agaggtcctg agttcaattc ccaacaacta tatggtggct accaaccatt acaatgagat 3360
cagatgccct cctcttgtgt atctgaagag agtgacagtg tacttacata cataaaataa 3420
ataaataaat ctaaaaaaat gttaaaaaa 3449
<210>85
<211>309
<212>PRT
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R05,rGR05
<400>85
Met Leu Gly Ala Met Glu Gly Val Leu Leu Ser Val Ala Thr Ser Glu
1 5 10 15
Ala Leu Leu Gly Ile Val Gly Asn Thr Phe Ile Ala Leu Val Asn Cys
20 25 30
Met Asp Cys Thr Arg Asn Lys Asn Leu Tyr Asn Ile Gly Phe Ile Leu
35 40 45
Thr Gly Leu Ala Ile Ser Arg Ile Cys Leu Val Trp Ile Leu Ile Thr
50 55 60
Glu Ala Tyr Ile Lys Ile Phe Ser Pro Gln Leu Leu Ser Pro Ile Asn
65 70 75 80
Ile Ile Glu Leu Ile Ser Tyr Leu Trp Ile Ile Thr Ser Gln Leu Asn
85 90 95
Val Trp Phe Ala Thr Ser Leu Ser Ile Phe Tyr Phe Leu Lys Ile Ala
100 105 110
Asn Phe Ser His His Ile Phe Leu Trp Leu Lys Arg Arg Ile Asn Ile
115 120 125
Val Phe Ala Phe Leu Ile Gly Cys Leu Leu Met Ser Trp Leu Phe Ser
130 135 140
Phe Pro Val Val Val Lys Met Val Lys Asp Lys Lys Met Leu Tyr Ile
145 150 155 160
Asn Ser Ser Trp Gln Ile His Met Lys Lys Ser Glu Leu Ile Ile Asn
165 170 175
Tyr Val Phe Thr Asn Gly Gly Val Phe Leu Leu Phe Ile Ile Met Leu
180 185 190
Ile Val Cys Phe Leu Leu Ile Ile Ser Leu Trp Arg His Ser Lys Trp
195 200 205
Met Gln Ser Asn Glu Ser Gly Phe Arg Asp Leu Asn Thr Glu Val His
210 215 220
Val Lys Thr Ile Lys Val Leu Leu Ser Phe Ile Ile Leu Phe Ile Leu
225 230 235 240
His Leu Ile Gly Ile Thr Ile Asn Val Ile Cys Leu Leu Val Pro Glu
245 250 255
Asn Asn Leu Leu Phe Val Phe Gly Leu Thr Ile Ala Phe Leu Tyr Pro
260 265 270
Cys Cys His Ser Leu Ile Leu Ile Leu Ala Asn Ser Arg Leu Lys Arg
275 280 285
Cys Phe Val Arg Ile Leu Gln Gln Leu Met Cys Ser Glu Glu Gly Lys
290 295 300
Glu Phe Arg Asn Thr
305
<210>86
<211>1127
<212>DNA
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R05,rGR05
<400>86
aagagatttc agatactacc acaaacattt tttaaatata tgtaagtctt taaagaaaga 60
agggaaagcc actcctttat tgagcagcca atagattgcc atcttaaaat tctgtggcag 120
aagctatttt aaagatctgc gaagatgctg ggtgcaatgg aaggtgtcct cctttcagtt 180
gcaactagtg aggctttgct tggcattgta gggaacacat tcattgcact tgtgaactgc 240
atggactgta ccaggaacaa gaatctctat aatattggct tcattctcac tggcttggca 300
atttccagaa tctgcctcgt gtggatctta atcacagagg catacataaa aatattctct 360
ccacagttgc tgtctcctat caacataatt gaactcatca gttatctatg gataattacc 420
agtcaattga atgtttggtt tgctaccagc ctcagtatct tttatttcct caagatagca 480
aatttttccc accacatatt tctctggtta aaaagaagaa ttaatatagt ttttgccttc 540
ctgatagggt gcttacttat gtcatggcta ttttctttcc cagtagttgt gaagatggtt 600
aaagataaaa aaatgctgta tataaactca tcttggcaaa tccacatgaa gaaaagtgag 660
ttaatcatta actatgtttt caccaatggg ggagtatttt tactttttat aataatgtta 720
attgtatgtt ttctcttaat tatttccctt tggagacaca gcaagtggat gcaatcaaat 780
gaatcaggat tcagagatct caacacagaa gttcatgtga aaacaataaa agttttatta 840
tcttttatta tcctttttat attgcattta attggtatta ccatcaatgt catttgtctg 900
ttagtcccag aaaataactt gttattcgtg tttggtttga cgattgcatt cctctatccc 960
tgctgccact cacttatcct aattctagca aacagccggc tgaaacgatg ctttgtaagg 1020
atactgcaac aattaatgtg ctctgaggaa ggaaaagaat tcagaaacac atgacagtct 1080
ggaagacaaa caatcagaaa tagtaagtga aaaaaaaaaa aaaaaaa 1127
<210>87
<211>297
<212>PRT
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R06,rGR06
<400>87
Glu Ala Leu Val Gly Ile Leu Gly Asn Ala Phe Ile Ala Leu Val Asn
1 5 10 15
Phe Met Gly Trp Met Lys Asn Arg Lys Ile Thr Ala Ile Asp Leu Ile
20 25 30
Leu Ser Ser Leu Ala Met Ser Arg Ile Cys Leu Gln Cys Ile Ile Leu
35 40 45
Leu Asp Cys Ile Ile Leu Val Gln Tyr Pro Asp Thr Tyr Asn Arg Gly
50 55 60
Lys Glu Met Arg Ile Ile Asp Phe Phe Trp Thr Leu Thr Asn His Leu
65 70 75 80
Ser Val Trp Phe Ala Thr Cys Leu Ser Ile Phe Tyr Phe Phe Lys Ile
85 90 95
Ala Asn Phe Phe His Pro Leu Phe Leu Trp Ile Lys Trp Arg Ile Asp
100 105 110
Lys Leu Ile Leu Arg Thr Leu Leu Ala Cys Leu Ile Leu Ser Leu Cys
115 120 125
Phe Ser Leu Pro Val Thr Glu Asn Leu Ala Asp Asp Phe Arg Arg Cys
130 135 140
Val Lys Thr Lys Glu Arg Ile Asn Ser Thr Leu Arg Cys Lys Leu Asn
145 150 155 160
Lys Ala Gly Tyr Ala Ser Val Lys Val Asn Leu Asn Leu Val Met Leu
165 170 175
Phe Pro Phe Ser Val Ser Leu Val Ser Phe Leu Leu Leu Ile Leu Ser
180 185 190
Leu Trp Arg His Thr Arg Gln Met Gln Leu Asn Val Thr Gly Tyr Asn
195 200 205
Asp Pro Ser Thr Thr Ala His Val Lys Ala Thr Lys Ala Val Ile Ser
210 215 220
Phe Leu Val Leu Phe Ile Val Tyr Cys Leu Ala Phe Leu Ile Ala Thr
225 230 235 240
Ser Ser Tyr Phe Met Pro Glu Ser Glu Leu Ala Val Ile Trp Gly Glu
245 250 255
Leu Ile Ala Leu Ile Tyr Pro Ser Ser His Ser Phe Ile Leu Ile Leu
260 265 270
Gly Asn Ser Lys Leu Lys Gln Ala Ser Val Arg Val Leu Cys Arg Val
275 280 285
Lys Thr Met Leu Lys Gly Arg Lys Tyr
290 295
<210>88
<211>1304
<212>DNA
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R06,rGR06
<400>88
gtgaggcctt agtaggaatc ttaggaaatg cattcattgc attggtaaac ttcatgggct 60
ggatgaagaa taggaagatc actgctattg atttaatcct ctcaagtctg gctatgtcca 120
ggatttgtct acagtgtata attctattag attgtattat attggtgcag tatccagaca 180
cttacaacag gggtaaagaa atgaggatca ttgatttctt ctggacgctt accaaccatt 240
taagtgtctg gtttgccacc tgcctcagca ttttctattt cttcaagata gcaaacttct 300
tccatcctct tttcctctgg ataaagtgga gaattgacaa gctaattctg aggactctac 360
tggcatgctt gattctctcc ctatgcttta gcctcccagt cactgagaat ttggctgatg 420
atttcagacg ctgtgtcaag acaaaagaaa gaataaactc tactctgagg tgcaaattaa 480
ataaagctgg atatgcttct gtcaaggtaa atctcaactt ggtcatgctg ttcccctttt 540
ctgtgtccct tgtctcattc cttctcttga ttctctccct atggagacac accaggcaga 600
tgcaactcaa tgtaacaggg tacaatgatc ccagcacaac agctcatgtg aaagccacaa 660
aagcagtaat ttccttccta gttctgttta ttgtctactg cctggccttt cttatagcca 720
cttccagcta ctttatgcca gagagtgaat tagctgtaat ttggggtgag ctgatagctc 780
taatatatcc ctcaagccat tcatttatcc tgatccttgg gaacagtaaa ctaaaacagg 840
catctgtaag ggtgctttgt agagtaaaga ctatgttaaa gggaagaaaa tattagcatc 900
atggatatat ttgaagaaaa actatcactg tctaaagaaa aaggatgaca aatcattatc 960
tttcattctt atatgaatat tgctttcatg cggtaacatc ttttaacaaa cttaaatcaa 1020
atgttgggaa atctcatata cagcaacttt gcatgtctct ctgtctattt ccctctccct 1080
ttgtacatag ttgacataaa aaaagaattt tcatgacaaa attgtaataa atagctacag 1140
aggcagcaca ttttcatagt aagttctgaa tcactcttcc aaatgcaaag ctgcctgaca 1200
aattcaaaac aactgtaaca gtatttcact gctgtttgca ttctttggaa aagcaggtgg 1260
tttgttccta tgacctgact tggagttttc ttcttacatc actg 1304
<210>89
<211>305
<212>PRT
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R07,rGR07
<400>89
Met Gly Ser Ser Leu Tyr Asp Ile Leu Thr Ile Val Met Ile Ala Glu
1 5 10 15
Phe Ile Phe Gly Asn Val Thr Asn Gly Phe Ile Val Leu Thr Asn Cys
20 25 30
Ile Ala Trp Leu Ser Lys Arg Thr Leu Ser Phe Ile Gly Trp Ile Gln
35 40 45
Leu Phe Leu Ala Ile Ser Arg Val Val Leu Ile Trp Glu Met Leu Leu
50 55 60
Ala Trp Leu Lys Tyr Met Lys Tyr Ser Phe Ser Tyr Leu Ala Gly Thr
65 70 75 80
Glu Leu Arg Val Met Met Leu Thr Trp Val Val Ser Asn His Phe Ser
85 90 95
Leu Trp Leu Ala Thr Ile Leu Ser Ile Phe Tyr Leu Leu Lys Ile Ala
100 105 110
Ser Phe Ser Arg Pro Val Phe Leu Tyr Leu Lys Trp Arg Val Lys Lys
115 120 125
Val Leu Leu Leu Ile Leu Leu Gly Asn Leu Ile Phe Leu Met Phe Asn
130 135 140
Ile Leu Gln Ile Asn Thr His Ile Glu Asp Trp Met Asp Gln Tyr Lys
145 150 155 160
Arg Asn Ile Thr Trp Asp Ser Arg Val Asn Glu Phe Val Gly Phe Ser
165 170 175
Asn Leu Val Leu Leu Glu Met Ile Met Phe Ser Val Thr Pro Phe Thr
180 185 190
Val Ala Leu Val Ser Phe Ile Leu Leu Ile Phe Ser Leu Trp Lys His
195 200 205
Leu Gln Lys Met His Leu Ser Ser Arg Gly Glu Arg Asp Pro Ser Thr
210 215 220
Lys Ala His Val Asn Ala Leu Arg Ile Met Val Ser Phe Leu Leu Leu
225 230 235 240
Tyr Ala Thr Tyr Phe Ile Ser Phe Phe Ile Ser Leu Ile Pro Met Ala
245 250 255
His Lys Lys Gly Leu Asp Leu Met Phe Ser Leu Thr Val Gly Leu Phe
260 265 270
Tyr Pro Ser Ser His Ser Phe Ile Leu Ile Leu Gly His Ser Asn Leu
275 280 285
Arg His Ser Ser Cys Leu Val Ile Thr Tyr Leu Arg Cys Lys Glu Lys
290 295 300
Asp
305
<210>90
<211>3994
<212>DNA
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R07,rGR07
<220>
<221>modified_base
<222>(1)..(3994)
<223>n=g,a,c或t
<400>90
cagtagcaaa attttactat gttcattgat attatgtcan gncactacgt aagaaggaag 60
acttgaaaga aagcttatct gagtttttaa gaatacatgg acatttcagc ttggcaaatg 120
acgagctgtg aatttttgtc atctggacat gggaagcagc ctgtatgata tcttaactat 180
tgtcatgatt gcagagttta tattcggaaa tgtgaccaat ggattcatag tgctgacaaa 240
ctgtattgct tggctcagta aaagaactct ttctttcatt ggttggatcc agcttttctt 300
ggccatttcc agagtggttt tgatatggga aatgttacta gcatggctga aatatatgaa 360
gtattcattt tcatatttgg ctggcacaga attaagggtt atgatgttga cctgggtagt 420
ttccaatcac tttagtctct ggcttgccac cattctaagc atcttttatt tgctcaaaat 480
agctagtttc tccagacctg ttttcctgta tctgaagtgg agagtaaaaa aagtgctcct 540
gctgattctt ctcggaaatt taatcttcct gatgttcaat atattacaaa tcaacactca 600
catagaagac tggatggatc aatataagag aaatataacg tgggattcca gagtgaatga 660
atttgtgggg ttttcaaatc tggttttatt ggagatgatt atgttctctg taacaccatt 720
caccgtggct ctggtctcct tcatcctgtt aatcttctct ttatggaaac atctccagaa 780
gatgcatctc agttccagag gggaacgaga ccctagcaca aaagcccatg tgaatgccct 840
gagaattatg gtctccttcc tcttactcta tgccacttac ttcatatcct tttttatatc 900
attaattcct atggcacata aaaaaggact agatcttatg tttagcctaa ctgttggact 960
tttctaccct tcaagccact catttatctt gattttggga cattctaatc taaggcattc 1020
cagttgtctg gtgataacct atctgagatg taaggaaaag gattagaaat tcactattcc 1080
ataaggcagt taaaccacat gctattaggt atactcagtg ctagatccct aggcaagcat 1140
taacattaaa aatatataat ttctagattc ttctatttgt gataaaccac tcacttagaa 1200
taatgctaaa gtagcgtgat gttgtatata agtgtaagaa taaaatgtaa ttaatttagt 1260
ttaggcacaa taacatatgt ctactaagta aaaactaggc aggctgctac acgcatatta 1320
gaatccaggc tgaggtatat agactcaaga aatactgtgg aataaagatt ttaattttca 1380
ttctattgtg agttatgtga aatcaatgcc attaaaggca tacacaagat tttcacacac 1440
tgaaacaact tcttgcattt tgtcatattg tattggaagt aaattggaga taaacttaat 1500
atcaataaat tacaaaatgt aaacataaac agggtgatta aaaattagcc tctaggtcct 1560
ggggaaatga ttcaagtaaa gtgctttctt ttcaaatagg agaatctgat tgtaaatcat 1620
ctaaaagtct ggcataaaat gtcaatgaaa attgtatgta aaatatagct atggcmaaga 1680
gcaccmaaga aaagaaaatt tttgcctttg aaacccagta attgatatcc tttaaaaaag 1740
cagttacata tttttctgtt taagattttg tcaaagggta gctttgacaa ctaatataag 1800
ctgaggaagg tagcaagtgt gaagtcagct aatggggtca gtcaagtgct gttagcagca 1860
gatggaggcc actgctgaat ttagcaggca atttacaggg tgagcactgc tagtgctgac 1920
agaagaaaaa ctctgaaatt ttaactcttt agggtctggt gagaaagaaa aagagagaaa 1980
atcgcatata tatatatata tatatatata tatatatata tatatatata tatatatata 2040
tcatggaagc tctaacaagt tgactcaaac aactttatga tgtttttagg cccttttatt 2100
ttaatgtcag tgaattaggt gtggtacagc aatattgcta cttttaaatt caaagcagtt 2160
gttttatata ttattcatta tataagctaa ttataagttt aaatcaaaag gtttatttgt 2220
ccatgatttt actttatcat tgggcacacc tgtgctctca tccttgggct tgacctagaa 2280
tgaaagttta tccttgatca tatgtctgtc acaagactac ttctcttcct atagtagttt 2340
atgtacttac aatatacaaa agtttattga attcctttta tcacttatgc agccttttct 2400
tactattcta ttctattcta ttctattcta ttctattcta ttctattcta ttctattcta 2460
ttctattcta ttctattcta gaatctaacc tatacattca tttctggcaa aacaacttat 2520
atcatctcct taattatttt atcaattaat ctaacatcct gaagttattt aaatctaata 2580
taaggactct gtaaagtcac aaatttattt atacttcaca aaattcatta ttttatggaa 2640
ctgcagcatt gcctgggcca ggagtcacaa gagttccaga gttgacttta ttggcatctg 2700
cctggctaac tgaaggatca gttttctgtg tacaataatt ttgtgtatct cttttgatgc 2760
aagatatgaa aaataatttc agtctaaaag tgtccttaaa tttgaaactc tctggccaga 2820
atctaactat tgatgaccag tttgcaccat ggactcagtg tcttctattg ctttaaaata 2880
agcaacatct tgaatgcttt tcttgtgtat taggcaaata attaacaaca tgtttctatg 2940
attgtctcaa taacaatact atatttctca cagtttttaa tttttatggc aaagttggct 3000
aataagaatt tttttcaaat tatcaaacgt gaagaaaact tgacatttta tttcatggag 3060
attctaaatg ttttcttagc atattgcctt tttactaact tgatttttat catgttttgg 3120
tagtatttct aattttcctt tttttctaag tatgttatgt agtaacacca ggagaatgaa 3180
acaaatgaca tttatactaa ggatgtgaca aataaggccc aaagaaagtt ttgaaaatca 3240
tgatctcatt tctattcttc tttattaagt atagcataag caaaattctg atggtggtct 3300
tggcccatat ctttgaacac agtgtagtgg tgaagacttt ttcaaatatt atgtcatatt 3360
tgtacccatc tctgtaccta tttcttctga tttcatgagg aaaaaatgag gaagggtttg 3420
tttgtgtgct ggagcagctg aagtggacca aggggcagga attctctctg ttcggtccta 3480
gtgtgactga tgatgctctc attgaaaaac aggaagaaga agaaagactt tatatgcacc 3540
attcactcct tccccctcct acattccacc tccctcttga aagagtgtct atctatatag 3600
atatagctat cctgaaatcc attaagtaga cctgactggc ttaaatctca cagaaattca 3660
cctacctttt ccatgattgc tgaaattaaa gacatgtgcc gacatattgg gcacattcag 3720
accttttgcc aactgtcttt caactcattt ggacctactg agaagtattc aaaatatttg 3780
gttgttttaa ataaaaggaa agtgggtcta tattacttga attggataga gaaattttca 3840
cttacaagtg atattgaaaa tgggggagaa tgtattttag cataagcacc agaacacaaa 3900
gcaattcttg ttaaaacttt atcgataaat tggataaatg ttaaaaaaga aaaaataaaa 3960
tatacgaact attatgaaaa aaaaaaaaaa aaaa 3994
<210>91
<211>314
<212>PRT
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R08,rGR08
<400>91
Met Glu Pro Val Ile His Val Phe Ala Thr Leu Leu Ile His Val Glu
1 5 10 15
Phe Ile Phe Gly Asn Leu Ser Asn Gly Leu Ile Val Leu Ser Asn Phe
20 25 30
Trp Asp Trp Val Val Lys Arg Lys Leu Ser Thr Ile Asp Lys Ile Leu
35 40 45
Leu Thr Leu Ala Ile Ser Arg Ile Thr Leu Ile Trp Glu Met Tyr Ala
50 55 60
Cys Phe Lys Ile Val Tyr Gly Ser Ser Ser Phe Ile Phe Gly Met Lys
65 70 75 80
Leu Gln Ile Leu Tyr Phe Ala Trp Ile Leu Ser Ser His Phe Ser Leu
85 90 95
Trp Phe Ala Thr Ala Leu Ser Ile Phe Tyr Leu Leu Arg Ile Ala Asn
100 105 110
Cys Ser Trp Lys Ile Phe Leu Tyr Leu Lys Trp Arg Leu Lys Gln Val
115 120 125
Ile Val Gly Met Leu Leu Ala Ser Leu Val Phe Leu Pro Gly Ile Leu
130 135 140
Met Gln Arg Thr Leu Glu Glu Arg Pro Tyr Gln Tyr Gly Gly Asn Thr
145 150 155 160
Ser Glu Asp Ser Met Glu Thr Asp Phe Ala Lys Phe Thr Glu Leu Ile
165 170 175
Leu Phe Asn Met Thr Ile Phe Ser Val Ile Pro Phe Ser Leu Ala Leu
180 185 190
Ile Ser Phe Leu Leu Leu Ile Phe Ser Leu Trp Lys His Leu Gln Lys
195 200 205
Met Gln Leu Ser Ser Arg Gly His Gly Asp Pro Ser Thr Lys Ala His
210 215 220
Arg Asn Ala Leu Arg Ile Met Val Ser Phe Leu Leu Leu Tyr Thr Ser
225 230 235 240
Tyr Phe Leu Ser Leu Leu Ile Ser Trp Ile Ala Gln Lys His His Ser
245 250 255
Lys Leu Val Asp Ile Ile Gly Ile Ile Thr Glu Leu Met Tyr Pro Ser
260 265 270
Val His Ser Phe Ile Leu Ile Leu Gly Asn Ser Lys Leu Lys Gln Thr
275 280 285
Ser Leu Trp Ile Leu Ser His Leu Lys Cys Arg Leu Lys Gly Glu Asn
290 295 300
Ile Leu Thr Pro Ser Gly Lys Pro Ile Asn
305 310
<210>92
<211>1886
<212>DNA
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R08,rGR08
<220>
<221>modified_base
<222>(1351)
<223>n=g,a,c或t
<400>92
ctgcaggttg gtgatccagt aatgagcagc actgttatat ctcaggcttt ctaagatcat 60
ggaacctgtc attcacgtct ttgccactct actaatacat gtggagttca tttttgggaa 120
tctgagcaat ggattaatag tgttgtcaaa cttctgggac tgggtcgtta aacgaaaact 180
ttccacaatt gataaaattc ttcttacatt ggcaatttca agaatcactc tcatctggga 240
aatgtatgct tgttttaaaa ttgtatatgg ttcatcttca tttatatttg ggatgaagtt 300
acaaattctt tattttgcct ggatcctttc tagtcacttc agcctctggt ttgccacagc 360
tctcagcatc ttttacttac tcagaatagc taactgctcc tggaagatct tcctgtatct 420
gaaatggaga cttaaacaag tgattgtggg gatgttgctg gcaagcttgg tgttcttgcc 480
tggaatcctg atgcaaagga ctcttgaaga gaggccctat caatatggag gaaacacaag 540
tgaggattcc atggaaactg actttgcaaa gtttacagag ctgattcttt tcaacatgac 600
tatattctct gtaataccat tttcattggc cttgatttct tttctcctgc taatcttctc 660
tttgtggaaa catctccaga agatgcagct cagttccaga ggacatggag accctagcac 720
caaggcccac agaaatgctt tgagaattat ggtctccttc ctcttgctct acacttcata 780
tttcctgtct cttcttatat catggattgc tcagaagcat cacagtaaac tggttgacat 840
tattggtatt attactgaac tcatgtatcc ttcagtccac tcatttatcc tgattctagg 900
aaattctaaa ttaaagcaga cttctctttg gatactgagt catttgaaat gtagactgaa 960
aggagagaat attttaactc catctggcaa accaattaac tagctgttat atattctgta 1020
ttgcaaacaa atcagtgagt tagtggttca aggattccat ccttgactta ttgtatcatg 1080
gaagtcatat agggagaggc tgaacaagct atcttctgta aattggcaag ggttgcatat 1140
agtactggta ctgggacacc atccaaccat aaaaccttct aaccataacc tacctgactg 1200
caagatatgc tgggacaatg gtggctcaga gattttggga ctggccaacc aatgtctatt 1260
ctttcttgag gctcactcaa taaggaggcc atgcccaact cgtcctggat ggccaggaac 1320
cagaatctct gatggsccaa tgatctatgg nagaacccag cattactggg aaaaaagaat 1380
aatcactttg atgaatggtc aaatatttcc taaatatatt ctgatacact tgtacatcat 1440
ttctctttcc caatcatcat cacagggact tctccccagc acctgatggg aacagatacc 1500
aaaatctaca gccaaatact aaatgcaggt tggggaactc cacaaaagac tggaaggaag 1560
tactgtgaga gccagagtgg tccagaacac taggagaaca cagaacatcg aattaactaa 1620
gcagcactca tagggttaat gtaaaataaa gcagcagtca catagactgc acaggtgtac 1680
tctagatcct ctgcatatat gttgtggttg tcaaacttgg gagttttgtt ggactaataa 1740
caatgtgaat aagtaagtct ctgacactta ttcccgctct tggaaccctt ttccacattt 1800
tgtattgtct taccaccttg atatgaaggt ttctgaatag tccaaaaaaa aaaaaaaaaa 1860
aaaaaaaaaa aaaaaaaaaa aaaaaa 1886
<210>93
<211>309
<212>PRT
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R09,rGR09
<400>93
Met Leu Ser Ala Ala Glu Gly Ile Leu Leu Ser Ile Ala Thr Val Glu
1 5 10 15
Ala Gly Leu Gly Val Leu Gly Asn Thr Phe Ile Ala Leu Val Asn Cys
20 25 30
Met Asp Trp Ala Lys Asn Lys Lys Leu Ser Lys Ile Gly Phe Leu Leu
35 40 45
Phe Gly Leu Ala Thr Ser Arg Ile Phe Ile Val Trp Ile Leu Ile Leu
50 55 60
Asp Ala Tyr Ala Lys Leu Phe Phe Pro Gly Lys Tyr Leu Ser Lys Ser
65 70 75 80
Leu Thr Glu Ile Ile Ser Cys Ile Trp Met Thr Val Asn His Met Thr
85 90 95
Val Trp Phe Ala Thr Ser Leu Ser Ile Phe Tyr Phe Leu Lys Ile Ala
100 105 110
Asn Phe Ser His Tyr Ile Phe Leu Trp Leu Lys Arg Arg Thr Asp Lys
115 120 125
Val Phe Ala Phe Leu Leu Trp Cys Leu Leu Ile Ser Trp Ala Ile Ser
130 135 140
Phe Ser Phe Thr Val Lys Val Met Lys Ser Asn Pro Lys Asn His Gly
145 150 155 160
Asn Arg Thr Ser Gly Thr His Trp Glu Lys Arg Glu Phe Thr Ser Asn
165 170 175
Tyr Val Leu Ile Asn Ile Gly Val Ile Ser Leu Leu Ile Met Thr Leu
180 185 190
Thr Ala Cys Phe Leu Leu Ile Ile Ser Leu Trp Lys His Ser Arg Gln
195 200 205
Met Gln Ser Asn Val Ser Gly Phe Arg Asp Leu Asn Thr Glu Ala His
210 215 220
Val Lys Ala Ile Lys Phe Leu Ile Ser Phe Ile Ile Leu Phe Ile Leu
225 230 235 240
Tyr Phe Ile Gly Val Ala Val Glu Ile Ile Cys Met Phe Ile Pro Glu
245 250 255
Asn Lys Leu Leu Phe Ile Phe Gly Leu Thr Thr Ala Ser Val Tyr Pro
260 265 270
Cys Cys His Ser Val Ile Leu Ile Leu Thr Asn Ser Gln Leu Lys Gln
275 280 285
Ala Phe Val Lys Val Leu Glu Gly Leu Lys Phe Ser Glu Asn Gly Lys
290 295 300
Asp Leu Arg Ala Thr
305
<210>94
<211>2596
<212>DNA
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R09,rGR09
<400>94
ggacactgca gcagatctgc tatagaataa cagatacaaa catagcaacc tgcagagatg 60
ctcagtgcag cagaaggcat ccttctttcc attgcaactg ttgaagctgg gctgggagtt 120
ttagggaaca catttatcgc cctggttaac tgcatggatt gggccaagaa caagaagctc 180
tctaagattg gtttccttct ctttggctta gcaacttcca gaatttttat tgtatggata 240
ttaattttag acgcatatgc aaagctattc tttccgggga agtatttgtc taagagtctg 300
actgaaatca tctcttgtat atggatgact gtgaatcaca tgactgtctg gtttgccacc 360
agcctcagca tcttctattt cctaaaaata gcaaattttt cccactatat atttctctgg 420
ttaaagagga gaactgataa agtatttgcc tttctcttgt ggtgtttatt aatttcatgg 480
gcaatctcct tctcattcac tgtgaaagtg atgaagagca atccaaagaa tcatggaaac 540
aggaccagtg ggacacattg ggagaagaga gaattcacaa gtaactatgt tttaatcaat 600
attggagtca tttctctctt gatcatgacc ttaactgcat gtttcttgtt aattatttca 660
ctttggaaac acagcaggca gatgcagtct aatgtttcag gattcagaga tctcaacact 720
gaagctcatg tgaaagccat aaaattttta atttcattta tcatcctttt catcttgtac 780
tttataggtg ttgcagtaga aatcatctgc atgtttatcc cagaaaacaa actgctattt 840
atttttggtt tgacaactgc atccgtctat ccctgctgtc actcagtcat tctaattcta 900
acaaacagcc agctgaagca agcctttgta aaggtactgg agggattaaa gttctctgag 960
aacggaaaag atctcagggc cacatgagtc tggaacagaa atgggtagtc tggaataatt 1020
gtaaggaagt cgtagaaggt ctttttcatt tgtacagtgc tcttaccttg tttttgagga 1080
gatgtaaact tttttatttt tattttttat cctatgtgaa taagtgtgtg tgtgtgtgtg 1140
tgtgtttatg tgtgtgtgta tatatgtcta tgtgtgtttt aggaggttta agagggaaga 1200
gggaatagag gtatgttggt gtttttaaca tggatattca caggccaagg aacttgttct 1260
ctccttttac cttagggtag tgtcctttgt ggctgtcact ctgacagtct acactagttg 1320
aactaagagc ttttagccag ttcacttgtc taaacctccc ttctcatggt agcagtgttc 1380
tgattacaga atcatgctgt cacatacagc tttttaacaa ggttcccata gacagaattc 1440
atgtcaaacg gaatgcacag ctgtcactct tacccaccga tctctcttgc cagcccattc 1500
ctattgactt taaactgtag tattaaactt tactgaaatc ttctgcaacc agtctgacta 1560
tgtctcttga aatcacatga tatggtggaa ttttaatgcc atgtgaaaat ttgtttgttc 1620
agttagtttc ctactctgcc aaatcattct cttacacttg gcagaaaaaa accatcaact 1680
gtagactatt ttgtgtaaag actaatacag atagaataag tatcttaatc aagatgtcat 1740
tgtgattatc ctaatttccc cagagcactg gttccctttc cccagaaaga ctcacaaagg 1800
aactgaggca aacagttgtg gtcactcttg atatttacca gttgaaactg aagaacagtg 1860
tttcctttct gttcagtttt actacttaca gttactttat ttcatccatt aaatcccaaa 1920
gtgcttatta atagtagata tttgatgaag caacaatggt tataagagtg gatgtggatc 1980
tatgacaaag atctagagaa acagactatt tgtgaaagat ggatgaaagc cctgatgaaa 2040
ggattcttca tggtctttga ccccagggag ttttgaaatc aagcagccac agatcaaaga 2100
gagctgagaa gaggttctcc tgaagaaaat atccaaacac atggtgccag ccaaagcaga 2160
aaatagtgga caattcagtc caggacctga atgaggtaga caatgtcctg ttaagggttg 2220
gaacaaatat atagatatgg tcattcatat acagaaacct acaggcgtgt ttgaactctt 2280
ggtttctcag taatcaattc ttaaatcttt tttagaatgg attttttatc atcattcatg 2340
atctctcagc agagtctgca ggggctaaga gacacactaa gagtatctgg aggggggagt 2400
gtcttcctgc tctatcaacc cctaaagtca tatataacaa tacaaaattc cacattagtt 2460
aagttctttt ttttacatct ttattaaatt gggtatttct tatttacatt tcaaatgtga 2520
ttccctttcc tggtttccag gccaatatcc ccctaacctc tccccttcta tgtgggtatt 2580
ccctcgtgcc gaattc 2596
<210>95
<211>137
<212>PRT
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R10,rGR10
<400>95
Met Phe Leu His Thr Ile Lys Gln Arg Asp Ile Phe Thr Leu Ile Ile
1 5 10 15
Ile Phe Phe Val Glu Ile Thr Met Gly Ile Leu Gly Asn Gly Phe Ile
20 25 30
Ala Leu Val Asn Ile Val Asp Trp Ile Lys Arg Arg Arg Ile Ser Ser
35 40 45
Val Asp Lys Ile Leu Thr Thr Leu Ala Leu Thr Arg Leu Ile Tyr Ala
50 55 60
Trp Ser Met Leu Ile Phe Ile Leu Leu Phe Ile Leu Gly Pro His Leu
65 70 75 80
Ile Met Arg Ser Glu Ile Leu Thr Ser Met Gly Val Ile Trp Val Val
85 90 95
Asn Asn His Phe Ser Ile Trp Leu Ala Thr Cys Leu Gly Val Phe Tyr
100 105 110
Phe Leu Lys Ile Ala Asn Phe Ser Asn Ser Leu Phe Leu Tyr Leu Lys
115 120 125
Trp Arg Val Lys Lys Val Val Leu Met
130 135
<210>96
<211>818
<212>DNA
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R10,rGR10
<400>96
cccgggctgc aggattcggc acgagaatga aaacttttgc tctactattt tgctgttctg 60
tgataccaca gaccataaaa caatcgagcc aagggatcaa gagctgaaac ttcagaaagt 120
gggaatcaaa tttccttcct gataggttag cttatgagaa ttcagcatct tattcaactt 180
cagaaaattg gatataagat acagtgtctg gatgaagccg aattgatcta tttggggaga 240
aaaaacgcca acatttataa taaggtttta tgagacagtt cctgggaaat ttggatattt 300
cctagttagt aatgtgtaaa tgggatttta aaacatgatt attttgtatt tttaacaacc 360
aacatgagga gctttttaaa tgccacttag acattataaa ctgaagcatg ttcttacaca 420
caataaagca acgtgatatt tttactttga taatcatatt ttttgtggaa ataacaatgg 480
gaatcttagg aaatggattc atagcactag tgaacattgt ggactggatc aagagaagaa 540
ggatttcttc agtggataag attctcacta ccttggccct taccagactc atttatgcgt 600
ggtctatgct catttttata ttgttattca tactgggccc gcatttgatt atgagatcag 660
aaatacttac atcaatgggt gttatctggg tggtgaacaa tcacttcagc atctggcttg 720
ctacatgcct cggtgtcttt tattttctca agatagccaa tttttctaac tctttgtttc 780
tttacctaaa gtggagagtt aaaaaagtgg ttttaatg 818
<210>97
<211>105
<212>PRT
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R11,rGR11
<220>
<221>MOD_RES
<222>(101)
<223>Xaa=任何氨基酸
<400>97
Gly Ser Gly Asn Gly Phe Ile Val Ser Val Asn Gly Ser His Trp Phe
1 5 10 15
Lys Ser Lys Lys Ile Ser Leu Ser Asp Phe Ile Ile Thr Ser Leu Ala
20 25 30
Leu Phe Arg Ile Phe Leu Leu Trp Ile Ile Phe Thr Asp Ser Leu Ile
35 40 45
Ile Val Phe Ser Tyr His Ala His Asp Ser Gly Ile Arg Met Gln Leu
50 55 60
Ile Asp Val Phe Trp Thr Phe Thr Thr His Phe Ser Ile Trp Leu Ile
65 70 75 80
Ser Cys Leu Ser Val Phe Tyr Cys Leu Lys Ile Ala Thr Phe Ser His
85 90 95
Pro Ser Phe Leu Xaa Leu Lys Ser Arg
100 105
<210>98
<211>315
<212>DNA
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R11,rGR11
<400>98
ggatccggaa acggttttat cgtgtcagtc aatggcagcc attggttcaa gagcaagaag 60
atttctttgt ctgacttcat cattaccagc ttggccctct tcaggatctt tctgctgtgg 120
atcatcttta ctgatagcct cataatagtg ttctcttacc acgcccacga ctcagggata 180
aggatgcaac ttattgatgt tttctggaca tttacaaccc acttcagtat ttggcttatc 240
tcctgtctca gtgttttcta ctgcctgaaa atagccactt tctcccaccc ctcattcctg 300
tagctcaaat ctaga 315
<210>99
<211>308
<212>PRT
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R12,rGR12
<400>99
Met Leu Ser Thr Val Ser Val Phe Phe Met Ser Ile Phe Val Leu Leu
1 5 10 15
Cys Phe Leu Gly Ile Leu Ala Asn Gly Phe Ile Val Leu Met Leu Ser
20 25 30
Arg Glu Trp Leu Trp Arg Gly Arg Leu Leu Pro Ser Asp Met Ile Leu
35 40 45
Leu Ser Leu Gly Thr Ser Arg Phe Cys Gln Gln Cys Val Gly Leu Val
50 55 60
Asn Ser Phe Tyr Tyr Ser Leu His Leu Val Glu Tyr Ser Arg Ser Leu
65 70 75 80
Ala Arg Gln Leu Ile Ser Leu His Met Asp Phe Leu Asn Ser Ala Thr
85 90 95
Phe Trp Phe Gly Thr Trp Leu Ser Val Leu Phe Cys Ile Lys Ile Ala
100 105 110
Asn Phe Ser His Pro Ala Phe Leu Trp Leu Lys Trp Arg Phe Pro Ala
115 120 125
Leu Val Pro Trp Leu Leu Leu Gly Ser Ile Leu Val Ser Phe Ile Val
130 135 140
Thr Leu Met Phe Phe Trp Gly Asn His Thr Val Tyr Gln Ala Phe Leu
145 150 155 160
Arg Arg Lys Phe Ser Gly Asn Thr Thr Phe Lys Glu Trp Asn Arg Arg
165 170 175
Leu Glu Ile Asp Tyr Phe Met Pro Leu Lys Leu Val Thr Thr Ser Ile
180 185 190
Pro Cys Ser Leu Phe Leu Val Ser Ile Leu Leu Leu Ile Asn Ser Leu
195 200 205
Arg Arg His Ser Gln Arg Met Gln His Asn Ala His Ser Leu Gln Asp
210 215 220
Pro Asn Thr Gln Ala His Ser Arg Ala Leu Lys Ser Leu Ile Ser Phe
225 230 235 240
Leu Val Leu Tyr Ala Leu Ser Tyr Val Ser Met Val Ile Asp Ala Thr
245 250 255
Val Val Ile Ser Ser Asp Asn Val Trp Tyr Trp Pro Trp Gln Ile Ile
260 265 270
Leu Tyr Leu Cys Met Ser Val His Pro Phe Ile Leu Ile Thr Asn Asn
275 280 285
Leu Lys Phe Arg Gly Thr Phe Arg Gln Leu Leu Leu Leu Ala Arg Gly
290 295 300
Phe Trp Val Thr
305
<210>100
<211>1295
<212>DNA
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R12,rGR12
<400>100
gtgtgaggga ctgtgggtag gggctgggag gaggccagga accaaggcaa ccagtggtga 60
caggaggggc tgaaatgcta tcaactgtat cagttttctt catgtcgatc tttgttctgc 120
tctgtttcct gggaatcctg gcaaacggct tcattgtgct gatgctgagc agggaatggc 180
tatggcgcgg taggctgctc ccctcagaca tgatcctcct cagtttgggc acctcccgat 240
tctgccagca gtgcgttggg ctggtgaaca gtttctacta ttccctccac cttgttgagt 300
actccaggag ccttgcccgt caactcatta gtcttcacat ggacttcttg aactcagcca 360
ctttctggtt tggcacctgg ctcagcgtcc tgttctgtat caagattgct aacttctccc 420
atcctgcctt cctgtggttg aagtggagat tcccagcatt ggtgccttgg ctcctactgg 480
gctctatctt ggtgtccttc atcgtaactc tgatgttctt ttggggaaac cacactgtct 540
atcaggcatt cttaaggaga aagttttctg ggaacacaac ctttaaggag tggaacagaa 600
ggctggaaat agactatttc atgcctctga aacttgtcac cacgtcaatt ccttgctctc 660
tttttctagt ctcaattttg ctgttgatca attctctcag aaggcattca caaagaatgc 720
agcacaatgc tcacagcttg caagacccca acacccaggc tcacagcaga gccctgaagt 780
cactcatctc atttctggtt ctttacgcgc tgtcctatgt gtccatggtc attgacgcta 840
cagttgtcat ctcctcagat aacgtgtggt attggccctg gcaaattata ctttacttgt 900
gcatgtccgt acatccattt atccttatca ctaataatct caagttccga ggcaccttca 960
ggcagctact cctgttggcc aggggattct gggtgaccta gaaggtttgg tctctttatc 1020
tgtacccttt gaagagactt aggtgagggt gacttccctt ggaagtgatc tcatctacat 1080
ggaaatgtct ttgtaggctg acatggggtc atactatgtg gttcctcctt gggaaagagg 1140
agaagaaaat acagggattc tgagcgttct tccttatctt gggatattat gaaaatggac 1200
attctgaatc ctgaaccagt attgatctga agtgcaaagt acaatatgcc tgttcccttc 1260
atgtctgcta tcctcttggt acttattaat tccct 1295
<210>101
<211>332
<212>PRT
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R13,rGR13
<400>101
Met Cys Gly Phe Pro Leu Ser Ile Gln Leu Leu Thr Gly Leu Val Gln
1 5 10 15
Met Tyr Val Ile Leu Ile Ile Ala Val Phe Thr Pro Gly Met Leu Gly
20 25 30
Asn Val Phe Ile Gly Leu Val Asn Tyr Ser Asp Trp Val Lys Asn Lys
35 40 45
Lys Ile Thr Phe Ile Asn Phe Ile Leu Ile Cys Leu Ala Ala Ser Arg
50 55 60
Ile Ser Ser Val Leu Val Val Phe Ile Asp Ala Ile Ile Leu Glu Leu
65 70 75 80
Thr Pro His Val Tyr His Ser Tyr Ser Arg Val Lys Cys Ser Asp Ile
85 90 95
Phe Trp Val Ile Thr Asp Gln Leu Ser Thr Trp Leu Ala Thr Cys Leu
100 105 110
Ser Ile Phe Tyr Leu Leu Lys Ile Ala His Phe Ser His Pro Leu Phe
115 120 125
Leu Trp Leu Lys Trp Arg Leu Arg Gly Val Leu Val Gly Phe Leu Leu
130 135 140
Phe Ser Leu Phe Ser Leu Ile Val Tyr Phe Leu Leu Leu Glu Leu Leu
145 150 155 160
Ser Ile Trp Gly Asp Ile Tyr Val Ile Pro Lys Ser Asn Leu Thr Leu
165 170 175
Tyr Ser Glu Thr Ile Lys Thr Leu Ala Phe Gln Lys Ile Ile Val Phe
180 185 190
Asp Met Leu Tyr Leu Val Pro Phe Leu Val Ser Leu Ala Ser Leu Leu
195 200 205
Leu Leu Phe Leu Ser Leu Val Lys His Ser Gln Asn Leu Asp Arg Ile
210 215 220
Ser Thr Thr Ser Glu Asp Ser Arg Ala Lys Ile His Lys Lys Ala Met
225 230 235 240
Lys Met Leu Leu Ser Phe Leu Val Leu Phe Ile Ile His Ile Phe Cys
245 250 255
Met Gln Leu Ser Arg Trp Leu Phe Phe Leu Phe Pro Asn Asn Arg Ser
260 265 270
Thr Asn Phe Leu Leu Leu Thr Leu Asn Ile Phe Pro Leu Ser His Thr
275 280 285
Phe Ile Ile Ile Leu Gly Asn Ser Lys Leu Arg Gln Arg Ala Met Arg
290 295 300
Val Leu Gln His Leu Lys Ser Gln Leu Gln Glu Leu Ile Leu Ser Leu
305 310 315 320
His Arg Leu Ser Arg Val Phe Thr Met Glu Ile Ala
325 330
<210>102
<211>1287
<212>DNA
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R13,rGR13
<400>102
gggattcagt tggataagag aaaagtcaaa accctaagac taagaatttc cttaagtaga 60
tatcaatttc tatccattgg aaggagtttc caatcacact gaaattacaa taaaaaagga 120
gcaagataac tatgggaaag gatgattttc ggtggatgtt tgagaactga gcagcaaggc 180
aaattgatag atgtgtggat tccctctttc tattcaactg cttactggat tggttcaaat 240
gtacgtgata ttgataatag cagtgtttac acctggaatg ctggggaatg tgttcattgg 300
actggtaaac tactctgact gggtaaaaaa caagaaaatc accttcatca acttcatcct 360
gatctgtttg gcagcgtcca gaatcagctc tgtgttggtg gtatttattg atgcaatcat 420
cctagaacta actcctcatg tctatcattc ttacagtcga gtgaaatgct ctgatatatt 480
ctgggttata actgaccagc tgtcaacgtg gcttgccacc tgcctcagca ttttctactt 540
actcaaaata gcccacttct cccatcccct tttcctttgg ttgaagtgga gattgagagg 600
agtgcttgtt ggttttcttc tattttcttt gttctcattg attgtttatt ttctactcct 660
ggaattactg tctatttggg gagatattta tgtgatccct aaaagcaatc tgactttata 720
ttcagaaaca attaagaccc ttgcttttca aaagataatt gtttttgata tgctatattt 780
agtcccattt cttgtgtccc tagcctcatt gctcctttta tttttatcct tggtgaagca 840
ctcccaaaac cttgacagga tttctaccac ctctgaagat tccagagcca agatccacaa 900
gaaggccatg aaaatgctat tatctttcct cgttctcttt ataattcaca ttttttgcat 960
gcagttgtca cggtggttat tctttttgtt tccaaacaac aggtcaacta attttctttt 1020
gttaacatta aacatcttcc cattatctca tacattcatt atcatcctgg gaaacagcaa 1080
gcttcgacaa agagcaatga gggtcctgca acatcttaaa agccaacttc aagagttgat 1140
cctctccctt catagattgt ccagagtctt cactatggaa atagcttaaa ggggagactt 1200
ggaaggtcac tggtaacttg ttcttccgct gagttctgtt aagtaatgct ggacatatat 1260
gaactatccc tagtgcatac tgatatt 1287
<210>103
<211>68
<212>PRT
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R14,rGR14
<400>103
Val Ala Asn Ile Met Asp Trp Val Lys Arg Arg Lys Leu Ser Ala Val
1 5 10 15
Asp Gln Leu Leu Thr Val Leu Ala Ile Ser Arg Ile Thr Leu Leu Trp
20 25 30
Ser Leu Tyr Ile Leu Lys Ser Thr Phe Ser Met Val Pro Asn Phe Glu
35 40 45
Val Ala Ile Pro Ser Thr Arg Leu Thr Asn Leu Val Trp Ile Ile Ser
50 55 60
Asn His Phe Asn
65
<210>104
<211>206
<212>DNA
<213>大鼠(Rattus sp.)
<220>
<223>大鼠T2R14,rGR14
<400>104
ctgtggcaaa cataatggat tgggtcaaga gaaggaagct ctctgcagtg gatcagctcc 60
tcactgtgct ggccatctcc agaatcactc tgttgtggtc attgtacata ctgaaatcaa 120
cattttcaat ggtgccaaac tttgaggtag ctataccgtc aacaagacta actaatcttg 180
tctggataat ttctaaccat tttaat 206
<210>105
<211>327
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R01,mGR01
<400>105
Met Gln His Leu Leu Lys Thr Ile Phe Val Ile Cys His Ser Thr Leu
1 5 10 15
Ala Ile Ile Leu Ile Phe Glu Leu Ile Ile Gly Ile Leu Gly Asn Gly
20 25 30
Phe Met Ala Leu Val His Cys Met Asp Trp Val Lys Arg Lys Lys Met
35 40 45
Ser Leu Val Asn Lys Ile Leu Thr Ala Leu Ala Ile Ser Arg Ile Phe
50 55 60
His Leu Ser Leu Leu Leu Ile Ser Leu Val Ile Phe Phe Ser Tyr Ser
65 70 75 80
Asp Ile Pro Met Thr Ser Arg Met Thr Gln Val Ser Asn Asn Val Trp
85 90 95
Ile Ile Val Asn His Phe Ser Ile Trp Leu Ser Thr Cys Leu Ser Val
100 105 110
Leu Tyr Phe Leu Lys Ile Ser Asn Phe Ser Asn Ser Phe Phe Leu Tyr
115 120 125
Leu Lys Trp Arg Val Glu Lys Val Val Ser Val Thr Leu Leu Val Ser
130 135 140
Leu Leu Leu Leu Ile Leu Asn Ile Leu Leu Ile Asn Leu Glu Ile Ser
145 150 155 160
Ile Cys Ile Lys Glu Cys Gln Arg Asn Ile Ser Cys Ser Phe Ser Ser
165 170 175
His Tyr Tyr Ala Lys Cys His Arg Gln Val Ile Arg Leu His Ile Ile
180 185 190
Phe Leu Ser Val Pro Val Val Leu Ser Leu Ser Thr Phe Leu Leu Leu
195 200 205
Ile Phe Ser Leu Trp Thr Leu His Gln Arg Met Gln Gln His Val Gln
210 215 220
Gly Gly Arg Asp Ala Arg Thr Thr Ala His Phe Lys Ala Leu Gln Thr
225 230 235 240
Val Ile Ala Phe Phe Leu Leu Tyr Ser Ile Phe Ile Leu Ser Val Leu
245 250 255
Ile Gln Asn Glu Leu Leu Lys Lys Asn Leu Phe Val Val Phe Cys Glu
260 265 270
Val Val Tyr Ile Ala Phe Pro Thr Phe His Ser Tyr Ile Leu Ile Val
275 280 285
Gly Asp Met Lys Leu Arg Gln Ala Cys Leu Pro Leu Cys Ile Ile Ala
290 295 300
Ala Glu Ile Gln Thr Thr Leu Cys Arg Asn Phe Arg Ser Leu Lys Tyr
305 310 315 320
Phe Arg Leu Cys Cys Ile Phe
325
<210>106
<211>1374
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R01,mGR01
<400>106
agctgtgcgt gagcaaagca tttcttgtct gccacttctg agctgtgtga ggagacacat 60
tatcacggaa agagattcag actctgtcgc tgtcaaacct gtatgtttgc tcctctttta 120
ctgtgaaggc agagttacga aaaaaaatgt tatgagaacc aactcagaaa ttgacaaaaa 180
ttttctaaat gtcattttta aaaattatat ttcaaatgga aatgtgagca aatctttata 240
actaatatat aaaatgcagc atcttttaaa gacaatattt gttatctgcc atagcacact 300
tgcaatcatt ttaatctttg aattaataat tggaatttta ggaaatgggt tcatggccct 360
ggtgcactgt atggactggg ttaagagaaa gaaaatgtcc ttagttaata aaatcctcac 420
tgctttggca atctccagaa tttttcatct cagtttattg cttataagtt tagtcatatt 480
cttttcatat tctgatattc ctatgacttc aaggatgaca caagtcagta ataatgtttg 540
gattatagtc aatcatttca gtatctggct ttctacatgc ctcagtgtcc tttattttct 600
caagatatcc aatttttcta actctttttt tctttatcta aagtggagag ttgaaaaagt 660
agtttcagtt acactgttgg tgtcattgct cctcctgatt ttaaatattt tattaattaa 720
cttggaaatt agcatatgca taaaggaatg tcaaagaaac atatcatgca gcttcagttc 780
tcattactat gcaaagtgtc acaggcaggt gataaggctt cacattattt tcctgtctgt 840
ccccgttgtt ttgtccctgt caacttttct cctgctcatc ttctccctgt ggacacttca 900
ccagaggatg cagcagcatg ttcagggagg cagagatgcc agaaccacgg cccacttcaa 960
agccctacaa actgtgattg catttttcct actatattcc atttttattc tgtctgtctt 1020
aatacaaata tgaattactg aagaaaaatc ttttcgttgt attttgtgag gttgtatata 1080
tagcttttcc gacattccat tcatatattc tgattgtagg agacatgaag ctgagacagg 1140
cctgcctgcc tctctgtatt atcgcagctg aaattcagac tacactatgt agaaatttta 1200
gatcactaaa gtactttaga ttatgttgta tattctagac aaaaattaac tgatacaaat 1260
gtcttttgta tttttcattt taaatatcct ttaattttga ctgcatgaaa ttgatttctg 1320
cttgcaatta tcactgatta aaactattaa taatttaact agttgtatac aagg 1374
<210>107
<211>327
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R02,mGR02
<400>107
Met Glu Ser Val Leu His Asn Phe Ala Thr Val Leu Ile Tyr Val Glu
1 5 10 15
Phe Ile Phe Gly Asn Leu Ser Asn Gly Phe Ile Val Leu Ser Asn Phe
20 25 30
Leu Asp Trp Val Ile Lys Gln Lys Leu Ser Leu Ile Asp Lys Ile Leu
35 40 45
Leu Thr Leu Ala Ile Ser Arg Ile Thr Leu Ile Trp Glu Ile Tyr Ala
50 55 60
Trp Phe Lys Ser Leu Tyr Asp Pro Ser Ser Phe Leu Ile Gly Ile Glu
65 70 75 80
Phe Gln Ile Ile Tyr Phe Ser Trp Val Leu Ser Ser His Phe Ser Leu
85 90 95
Trp Leu Ala Thr Thr Leu Ser Val Phe Tyr Leu Leu Arg Ile Ala Asn
100 105 110
Cys Ser Trp Gln Ile Phe Leu Tyr Leu Lys Trp Arg Leu Lys Gln Leu
115 120 125
Ile Val Gly Met Leu Leu Gly Ser Leu Val Phe Leu Leu Gly Asn Leu
130 135 140
Met Gln Ser Met Leu Glu Glu Arg Phe Tyr Gln Tyr Gly Arg Asn Thr
145 150 155 160
Ser Val Asn Thr Met Ser Asn Asp Leu Ala Met Trp Thr Glu Leu Ile
165 170 175
Phe Phe Asn Met Ala Met Phe Ser Val Ile Pro Phe Thr Leu Ala Leu
180 185 190
Ile Ser Phe Leu Leu Leu Ile Phe Ser Leu Trp Lys His Leu Gln Lys
195 200 205
Met Gln Leu Ile Ser Arg Arg His Arg Asp Pro Ser Thr Lys Ala His
210 215 220
Met Asn Ala Leu Arg Ile Met Val Ser Phe Leu Leu Leu Tyr Thr Met
225 230 235 240
His Phe Leu Ser Leu Leu Ile Ser Trp Ile Ala Gln Lys His Gln Ser
245 250 255
Glu Leu Ala Asp Ile Ile Gly Met Ile Thr Glu Leu Met Tyr Pro Ser
260 265 270
Val His Ser Cys Ile Leu Ile Leu Gly Asn Ser Lys Leu Lys Gln Thr
275 280 285
Ser Leu Cys Mer Leu Arg His Leu Arg Cys Arg Leu Lys Gly Glu Asn
290 295 300
Ile Thr Ile Ala Tyr Ser Asn Gln Ile Thr Ser Phe Cys Val Phe Cys
305 310 315 320
Val Ala Asn Lys Ser Met Arg
325
<210>108
<211>1759
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R02,mGR02
<400>108
cagcacagtg aaaaactcat gggccacttg gtcacccagg gacaggcgac gctgttatat 60
gccaagcttt ctatgaacat ggaatctgtc cttcacaact ttgccactgt actaatatac 120
gtggagttta tttttgggaa tttgagcaat ggattcatag tgttgtcaaa cttcttggac 180
tgggtcatta aacaaaagct ttccttaata gataaaattc ttcttacatt ggcaatttca 240
agaatcactc tcatctggga aatatatgct tggtttaaaa gtttatatga tccatcttcc 300
tttttaattg gaatagaatt tcaaattatt tattttagct gggtcctttc tagtcacttc 360
agcctctggc ttgccacaac tctcagcgtc ttttatttac tcagaatagc taactgctcc 420
tggcagatct ttctctattt gaaatggaga cttaaacaac tgattgtggg gatgttgctg 480
ggaagcttgg tgttcttgct tggaaatctg atgcaaagca tgcttgaaga gaggttctat 540
caatatggaa ggaacacaag tgtgaatacc atgagcaatg accttgcaat gtggaccgag 600
ctgatctttt tcaacatggc tatgttctct gtaataccat ttacattggc cttgatttct 660
tttctcctgc taatcttctc tttgtggaaa catctccaga agatgcagct catttccaga 720
agacacagag accctagcac caaggcccac atgaatgcct tgagaattat ggtgtccttc 780
ctcttgctct ataccatgca tttcctgtct cttcttatat catggattgc tcaaaagcat 840
cagagtgaac tggctgatat tattggtatg ataactgaac tcatgtatcc ttcagtccat 900
tcatgtatcc tgattctagg aaattctaaa ttaaagcaga cttctctttg tatgctgagg 960
catttgagat gtaggctgaa aggagagaat atcacaattg catatagcaa ccaaataact 1020
agcttttgtg tattctgtgt tgcaaacaaa tctatgaggt agttgttcaa ggaatccttc 1080
cttgacttat tgtatcatgg aagtcatatg ggggagtctg aaagagctgt cttctgtaag 1140
caaggtttgt atacactagt ggggctggga caccaaccca agcacaaaac ctagctataa 1200
cctatcctgg ctgcaggata tgctggaaca atggtggctt ggaaattgtg ggactggcaa 1260
agcaatagct agtctaactt gaggcccatt ccacagcagg aagctcatgc ccacctctgc 1320
ctggatggcc aggaagcaaa atcttgatgg ccccaagacc tatggtaaac tgaacactac 1380
tggaaaaaga aagactcgtg ttaatgatct atcaaatatt tcctaatgat attctgataa 1440
actcatatat tagtccctgt cctaatcatc atcactggga ctccttccca gcacctgatg 1500
ggagcagata gagatctaca tccaaatagt aagtgtatct tggggaactc cacttaagaa 1560
tagaaggaac aattatgaga gccagagtga tccagaacac taggatcaca gaatcaacta 1620
agcagcatgc ataggggtta atggagactg aagtggcaat cacagagcct gcataggtct 1680
acactaagtc ctctgtgtat atactgtggc tgtttagctt aggaattttg ttggactcct 1740
aacaatggat aaggaattc 1759
<210>109
<211>312
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R03,mGR03
<400>109
Met Val Leu Thr Ile Arg Ala Ile Leu Trp Val Thr Leu Ile Thr Ile
1 5 10 15
Ile Ser Leu Glu Phe Ile Ile Gly Ile Leu Gly Asn Val Phe Ile Ala
20 25 30
Leu Val Asn Ile Ile Asp Trp Val Lys Arg Gly Lys Ile Ser Ala Val
35 40 45
Asp Lys Thr Tyr Met Ala Leu Ala Ile Ser Arg Thr Ala Phe Leu Leu
50 55 60
Ser Leu Ile Thr Gly Phe Leu Val Ser Leu Leu Asp Pro Ala Leu Leu
65 70 75 80
Gly Met Arg Thr Met Val Arg Leu Leu Thr Ile Ser Trp Met Val Thr
85 90 95
Asn His Phe Ser Val Trp Phe Ala Thr Cys Leu Ser Ile Phe Tyr Phe
100 105 110
Leu Lys Ile Ala Asn Phe Ser Asn Ser Ile Phe Leu Val Leu Lys Trp
115 120 125
Glu Ala Lys Lys Val Val Ser Val Thr Leu Val Val Ser Val Ile Ile
130 135 140
Leu Ile Met Asn Ile Ile Val Ile Asn Lys Phe Thr Asp Arg Leu Gln
145 150 155 160
Val Asn Thr Leu Gln Asn Cys Ser Thr Ser Asn Thr Leu Lys Asp Tyr
165 170 175
Gly Leu Phe Leu Phe Ile Ser Thr Gly Phe Thr Leu Thr Pro Phe Ala
180 185 190
Val Ser Leu Thr Met Phe Leu Leu Leu Ile Phe Ser Leu Trp Arg His
195 200 205
Leu Lys Asn Met Cys His Ser Ala Thr Gly Ser Arg Asp Val Ser Thr
210 215 220
Val Ala His Ile Lys Gly Leu Gln Thr Val Val Thr Phe Leu Leu Leu
225 230 235 240
Tyr Thr Ala Phe Val Met Ser Leu Leu Ser Glu Ser Leu Asn Ile Asn
245 250 255
Ile Gln His Thr Asn Leu Leu Ser His Phe Leu Arg Ser Ile Gly Val
260 265 270
Ala Phe Pro Thr Gly His Ser Cys Val Leu Ile Leu Gly Asn Ser Lys
275 280 285
Leu Arg Gln Ala Ser Leu Ser Val Ile Leu Trp Leu Arg Tyr Lys Tyr
290 295 300
Lys His Ile Glu Asn Trp Gly Pro
305 310
<210>110
<211>1484
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R03,mGR03
<400>110
ctttaatagc agggtgtgaa tatttaaatt ttctttctgc agcaactact gagggcttca 60
gactgctgta tacagggcat gaagcatctg gatgaagttc agctgtgctg cctttgacaa 120
caattttttg tgtatgtgtg gagaacataa accatttcat tagtgaaatt tggcttttgg 180
gtgacattgt ctatgatagt tctgaaagtg attatgttaa gaatcagaca cagccgtcta 240
gaagattgta ttaacacatc tttggtagtt cagaagaaat tagatcatca tggtgttgac 300
aataagggct attttatggg taacattgat aactattata agtctggagt ttatcatagg 360
aattttagga aatgtattca tagctctcgt gaacatcata gactgggtta aaagaggaaa 420
gatctctgca gtggataaga cctatatggc cctggccatc tccaggactg cttttttatt 480
gtcactaatc acagggttct tggtatcatt attggaccca gctttattgg gaatgagaac 540
gatggtaagg ctccttacta tttcctggat ggtgaccaat catttcagtg tctggtttgc 600
aacatgcctc agtatctttt attttctcaa gatagctaat ttctcaaatt ctattttcct 660
tgttctcaaa tgggaagcta aaaaagtggt atcagtgaca ttggtggtat ctgtgataat 720
cttgatcatg aacattatag tcataaacaa attcactgac agacttcaag taaacacact 780
ccagaactgt agtacaagta acactttaaa agattatggg ctctttttat tcattagcac 840
tgggtttaca ctcaccccat tcgctgtgtc tttgacaatg tttcttctgc tcatcttctc 900
cctgtggaga catctgaaga atatgtgtca cagtgccaca ggctccagag atgtcagcac 960
agtggcccac ataaaaggct tgcaaactgt ggtaaccttc ctgttactat atactgcttt 1020
tgttatgtca cttctttcag agtctttgaa tattaacatt caacatacaa atcttctttc 1080
tcatttttta cggagtatag gagtagcttt tcccacaggc cactcctgtg tactgattct 1140
tggaaacagt aagctgaggc aagcctctct ttctgtgata ttgtggctga ggtataagta 1200
caaacatata gagaattggg gcccctaaat catatcaggg atccttttcc acattctaga 1260
aaaaaatcag ttaataagaa caggaattta ggaaggaatc tgaaattatg aatctcatag 1320
gccatgaacc ttcagacaaa ggattcatta gagagataga gagagaacat tgttatctgt 1380
aactcgacag gcaacactgt agattatgaa aataaatgtc agtctgtaat ggaaagcaaa 1440
acatgctata ttttattaat tggttttggt ttaaggtcgg gata 1484
<210>111
<211>302
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R04,mGR04
<400>111
Met Leu Ser Ala Leu Glu Ser Ile Leu Leu Ser Val Ala Thr Ser Glu
1 5 10 15
Ala Met Leu Gly Val Leu Gly Asn Thr Phe Ile Val Leu Val Asn Tyr
20 25 30
Thr Asp Trp Val Arg Asn Lys Lys Leu Ser Lys Ile Asn Phe Ile Leu
35 40 45
Thr Gly Leu Ala Ile Ser Arg Ile Phe Thr Ile Trp Ile Ile Thr Leu
50 55 60
Asp Ala Tyr Thr Lys Val Phe Leu Leu Thr Met Leu Met Pro Ser Ser
65 70 75 80
Leu His Glu Cys Met Ser Tyr Ile Trp Val Ile Ile Asn His Leu Ser
85 90 95
Val Trp Phe Ser Thr Ser Leu Gly Ile Phe Tyr Phe Leu Lys Ile Ala
100 105 110
Asn Phe Ser His Tyr Ile Phe Leu Trp Met Lys Arg Arg Ala Asp Lys
115 120 125
Val Phe Val Phe Leu Ile Val Phe Leu Ile Ile Thr Trp Leu Ala Ser
130 135 140
Phe Pro Leu Ala Val Lys Val Ile Lys Asp Val Lys Ile Tyr Gln Ser
145 150 155 160
Asn Thr Ser Trp Leu Ile His Leu Glu Lys Ser Glu Leu Leu Ile Asn
165 170 175
Tyr Val Phe Ala Asn Met Gly Pro Ile Ser Leu Phe Ile Val Ala Ile
180 185 190
Ile Ala Cys Phe Leu Leu Thr Ile Ser Leu Trp Arg His Ser Arg Gln
195 200 205
Met Gln Ser Ile Gly Ser Gly Phe Arg Asp Leu Asn Thr Glu Ala His
210 215 220
Met Lys Ala Met Lys Val Leu Ile Ala Phe Ile Ile Leu Phe Ile Leu
225 230 235 240
Tyr Phe Leu Gly Ile Leu Ile Glu Thr Leu Cys Leu Phe Leu Thr Asn
245 250 255
Asn Lys Leu Leu Phe Ile Phe Gly Phe Thr Leu Ser Ala Met Tyr Pro
260 265 270
Cys Cys His Ser Phe Ile Leu Ile Leu Thr Ser Arg Glu Leu Lys Gln
275 280 285
Asp Thr Met Arg Ala Leu Gln Arg Leu Lys Cys Cys Glu Thr
290 295 300
<210>112
<211>1529
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R04,mGR04
<400>112
ctgcagcagg taaatcacac cagatccagc agaagccttc ttggaaattg gcagagatgc 60
tgagtgcact ggaaagcatc ctcctttctg ttgccactag tgaagccatg ctgggagttt 120
tagggaacac atttattgta cttgtaaact acacagactg ggtcaggaat aagaaactct 180
ctaagattaa ctttattctc actggcttag caatttccag gatttttacc atatggataa 240
taactttaga tgcatataca aaggttttcc ttctgactat gcttatgccg agcagtctac 300
atgaatgcat gagttacata tgggtaatta ttaaccatct gagcgtttgg tttagcacca 360
gcctcggcat cttttatttt ctgaagatag caaatttttc ccactacata tttctctgga 420
tgaagagaag agctgataaa gtttttgtct ttctaattgt attcttaatt ataacgtggc 480
tagcttcctt tccgctagct gtgaaggtca ttaaagatgt taaaatatat cagagcaaca 540
catcctggct gatccacctg gagaagagtg agttacttat aaactatgtt tttgccaata 600
tggggcccat ttccctcttt attgtagcca taattgcttg tttcttgtta accatttccc 660
tttggagaca cagcaggcag atgcaatcca ttggatcagg attcagagat ctcaacacag 720
aagctcacat gaaagccatg aaagttttaa ttgcatttat catcctcttt atcttatatt 780
ttttgggtat tctcatagaa acattatgct tgtttcttac aaacaataaa cttctcttta 840
tttttggctt cactttgtca gccatgtatc cctgttgcca ttcctttatc ctaattctaa 900
caagcaggga gctgaagcaa gacactatga gggcactgca gagattaaaa tgctgtgaga 960
cttgacagag aaatgaatgt tctggcacag ttcagcaggg aatccctgga gccctttcca 1020
ttcccactat gttctcacac tgtctttagt tgaattgtta aaagtttttg aaacctttgg 1080
caactgattg actgcagcta cgccagtgta agattttcat agtaagagca aacattgaaa 1140
ataagacttc tcagtcttat ttcattgagt ttctaaagca ttgacaccca ttcaccagaa 1200
aaaccaaagg ggaagagagg agttttcaga catgtgtgat gaatcttgat atttaggaca 1260
tggaattgag gagccagagg gatgctaccg tgtgtctaca gctttgtttg ttaaatagct 1320
acttttcctt tcccagttag ttaaagtaga tgcttggagt agtggtgaaa atcatggcag 1380
tagatgggat ctgtgggaag tggttgagga agcaggctgt ttctgaacga agagaccaga 1440
ggactgattg aactggtcat tgtgtatatc aaaaatagtg atttcagatg aagccaagtt 1500
gtagagcaaa gatatctgag gaagaattc 1529
<210>113
<211>300
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R05,mGR05
<400>113
Met Leu Ser Ala Ala Glu Gly Ile Leu Leu Ser Ile Ala Thr Val Glu
1 5 10 15
Ala Gly Leu Gly Val Leu Gly Asn Thr Phe Ile Ala Leu Val Asn Cys
20 25 30
Met Asp Trp Ala Lys Asn Asn Lys Leu Ser Met Thr Gly Phe Leu Leu
35 40 45
Ile Gly Leu Ala Thr Ser Arg Ile Phe Ile Val Trp Leu Leu Thr Leu
50 55 60
Asp Ala Tyr Ala Lys Leu Phe Tyr Pro Ser Lys Tyr Phe Ser Ser Ser
65 70 75 80
Leu Ile Glu Ile Ile Ser Tyr Ile Trp Met Thr Val Asn His Leu Thr
85 90 95
Val Trp Phe Ala Thr Ser Leu Ser Ile Phe Tyr Phe Leu Lys Ile Ala
100 105 110
Asn Phe Ser Asp Cys Val Phe Leu Trp Leu Lys Arg Arg Thr Asp Lys
115 120 125
Ala Phe Val Phe Leu Leu Gly Cys Leu Leu Thr Ser Trp Val Ile Ser
130 135 140
Phe Ser Phe Val Val Lys Val Met Lys Asp Gly Lys Val Asn His Arg
145 150 155 160
Asn Arg Thr Ser Glu Met Tyr Trp Glu Lys Arg Gln Phe Thr Ile Asn
165 170 175
Tyr Val Phe Leu Asn Ile Gly Val Ile Ser Leu Phe Met Met Thr Leu
180 185 190
Thr Ala Cys Phe Leu Leu Ile Met Ser Leu Trp Arg His Ser Arg Gln
195 200 205
Met Gln Ser Gly Val Ser Gly Phe Arg Asp Leu Asn Thr Glu Ala His
210 215 220
Val Lys Ala Ile Lys Phe Leu Ile Ser Phe Ile Ile Leu Phe Val Leu
225 230 235 240
Tyr Phe Ile Gly Val Ser Ile Glu Ile Ile Cys Ile Phe Ile Pro Glu
245 250 255
Asn Lys Leu Leu Phe Ile Phe Gly Phe Thr Thr Ala Ser Ile Tyr Pro
260 265 270
Cys Cys His Ser Phe Ile Leu Ile Leu Ser Asn Ser Gln Leu Lys Gln
275 280 285
Ala Phe Val Lys Val Leu Gln Gly Leu Lys Phe Phe
290 295 300
<210>114
<211>903
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R05,mGR05
<400>114
atgctgagtg cggcagaagg catcctcctt tccattgcaa ctgttgaagc tgggctggga 60
gttttaggga acacatttat tgcactggta aactgcatgg actgggccaa gaacaataag 120
ctttctatga ctggcttcct tctcatcggc ttagcaactt ccaggatttt tattgtgtgg 180
ctattaactt tagatgcata tgcaaagcta ttctatccaa gtaagtattt ttctagtagt 240
ctgattgaaa tcatctctta tatatggatg actgtgaatc acctgactgt ctggtttgcc 300
accagcctaa gcatcttcta tttcctgaag atagccaatt tttccgactg tgtatttctc 360
tggttgaaga ggagaactga taaagctttt gtttttctct tggggtgttt gctaacttca 420
tgggtaatct ccttctcatt tgttgtgaag gtgatgaagg acggtaaagt gaatcataga 480
aacaggacct cggagatgta ctgggagaaa aggcaattca ctattaacta cgttttcctc 540
aatattggag tcatttctct ctttatgatg accttaactg catgtttctt gttaattatg 600
tcactttgga gacacagcag gcagatgcag tctggtgttt caggattcag agacctcaac 660
acagaagctc atgtgaaagc cataaaattt ttaatttcat ttatcatcct tttcgtcttg 720
tattttatag gtgtttcaat agaaattatc tgcatattta taccagaaaa caaactgcta 780
tttatttttg gtttcacaac tgcatccata tatccttgct gtcactcatt tattctaatt 840
ctatctaaca gccagctaaa gcaagccttt gtaaaggtac tgcaaggatt aaagttcttt 900
tag 903
<210>115
<21l>308
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R06,mGR06
<400>115
Met Leu Thr Val Ala Glu Gly Ile Leu Leu Cys Phe Val Thr Ser Gly
1 5 10 15
Ser Val Leu Gly Val Leu Gly Asn Gly Phe Ile Leu His Ala Asn Tyr
20 25 30
Ile Asn Cys Val Arg Lys Lys Phe Ser Thr Ala Gly Phe Ile Leu Thr
35 40 45
Gly Leu Ala Ile Cys Arg Ile Phe Val Ile Cys Ile Ile Ile Ser Asp
50 55 60
Gly Tyr Leu Lys Leu Phe Ser Pro His Met Val Ala Ser Asp Ala His
65 70 75 80
Ile Ile Val Ile Ser Tyr Ile Trp Val Ile Ile Asn His Thr Ser Ile
85 90 95
Trp Phe Ala Thr Ser Leu Asn Leu Phe Tyr Leu Leu Lys Ile Ala Asn
100 105 110
Phe Ser His Tyr Ile Phe Phe Cys Leu Lys Arg Arg Ile Asn Thr Val
115 120 125
Phe Ile Phe Leu Leu Gly Cys Leu Phe Ile Ser Trp Ser Ile Ala Phe
130 135 140
Pro Gln Thr Val Lys Ile Phe Asn Val Lys Lys GIn His Arg Asn Val
145 150 155 160
Ser Trp Gln Val Tyr Leu Tyr Lys Asn Glu Phe Ile Val Set His Ile
165 170 175
Leu Leu Asn Leu Gly Val Ile Phe Phe Phe Met Val Ala Ile Ile Thr
180 185 190
Cys Phe Leu Leu Ile Ile Ser Leu Trp Lys His Asn Arg Lys Met Gln
195 200 205
Leu Tyr Ala Ser Arg Phe Lys Ser Leu Asn Thr Glu Val His Val Lys
210 215 220
Val Met Lys Val Leu Ile Ser Phe Ile Ile Leu Leu Ile Leu His Phe
225 230 235 240
Ile Gly Ile Leu Ile Glu Thr Leu Ser Phe Leu Lys Tyr Glu Asn Lys
245 250 255
Leu Leu Leu Ile Leu Gly Leu Ile Ile Ser Cys Met Tyr Pro Cys Cys
260 265 270
His Ser Phe Ile Leu Ile Leu Ala Asn Ser Gln Leu Lys Gln Ala Ser
275 280 285
Leu Lys Ala Leu Lys Gln Leu Lys Cys His Lys Lys Asp Lys Asp Val
290 295 300
Arg Val Thr Trp
305
<210>116
<211>1242
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R06,mGR06
<400>116
tatagttgca gcagaagcaa cgttagggat ctgtagagat gctgactgta gcagaaggaa 60
tcctcctttg ttttgtaact agtggttcag tcctgggagt tctaggaaat ggatttatcc 120
tgcatgcaaa ctacattaac tgtgtcagaa agaagttctc cacagctggc tttattctca 180
caggcttggc tatttgcaga atctttgtca tatgtataat aatctctgat ggatatttaa 240
aattgttttc tccacatatg gttgcctctg atgcccacat tatagtgatt tcttacatat 300
gggtaattat caatcataca agtatatggt ttgccaccag cctcaacctc ttctatctcc 360
tgaagatagc aaatttttct cactacatct tcttctgctt gaagagaaga atcaatacag 420
tatttatctt tctcctggga tgcttattta tatcatggtc aattgctttc ccacaaacag 480
tgaagatatt taatgttaaa aagcagcaca gaaatgtttc ctggcaggtt tacctctata 540
agaatgagtt cattgtaagc cacattcttc tcaacctggg agttatattc ttctttatgg 600
tggctatcat tacatgcttc ctattaatta tttcactttg gaaacataac agaaagatgc 660
agttgtatgc ctcaagattc aaaagcctta acacagaagt acatgtgaaa gtcatgaaag 720
ttttaatttc ttttattatc ctgttaatct tgcatttcat agggattttg atagaaacat 780
tgagcttttt aaaatatgaa aataaactgc tacttatttt gggtttgata atttcatgca 840
tgtatccttg ctgtcattca tttatcctaa ttctagcaaa cagtcagctg aagcaggctt 900
ctttgaaggc actgaagcaa ttaaaatgcc ataagaaaga caaggacgtc agagtgacat 960
ggtagactta tggagaaatg aatggtcaca agaaatagcc tggtgtggag atgttgatat 1020
ctctaaagac cgtttcactt ccaaattctt gcaattattt aaaaaaaaaa gtcttgctga 1080
tatcatggaa tcatgggaaa tgttgcaatt gtgttttggg gacagggtga ccagtgaagg 1140
tatggttaag cagcgaaaca ctcatacagc tcgttcgttc tttttgtatt ttattttgtg 1200
ttggtggcct tccaagacat gatttctcta tgtaagtttt gg 1242
<210>117
<211>308
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R07,mGR07
<400>117
Met Leu Asn Ser Ala Glu Gly Ile Leu Leu Cys Val Val Thr Ser Glu
1 5 10 15
Ala Val Leu Gly Val Leu Gly Asp Thr Tyr Ile Ala Leu Phe Asn Cys
20 25 30
Met Asp Tyr Ala Lys Asn Lys Lys Leu Ser Lys Ile Gly Phe Ile Leu
35 40 45
Ile Gly Leu Ala Ile Ser Arg Ile Gly Val Val Trp Ile Ile Ile Leu
50 55 60
Gln Gly Tyr Ile Gln Val Phe Phe Pro His Met Leu Thr Ser Gly Asn
65 70 75 80
Ile Thr Glu Tyr Ile Thr Tyr Ile Trp Val Phe Leu Asn His Leu Ser
85 90 95
Val Trp Phe Val Thr Asn Leu Asn Ile Leu Tyr Phe Leu Lys Ile Ala
100 105 110
Asn Phe Ser Asn Ser Val Phe Leu Trp Leu Lys Arg Arg Val Asn Ala
115 120 125
Val Phe Ile Phe Leu Ser Gly Cys Leu Leu Thr Ser Trp Leu Leu Cys
130 135 140
Phe Pro Gln Met Thr Lys Ile Leu Gln Asn Ser Lys Met His Gln Arg
145 150 155 160
Asn Thr Ser Trp Val His Gln Arg Lys Asn Tyr Phe Leu Ile Asn Gln
165 170 175
Ser Val Thr Asn Leu Gly Ile Phe Phe Phe Ile Ile Val Ser Leu Ile
180 185 190
Thr Cys Phe Leu Leu Ile Val Phe Leu Trp Arg His Val Arg Gln Met
195 200 205
His Ser Asp Val Ser Gly Phe Arg Asp His Ser Thr Lys Val His Val
210 215 220
Lys Ala Met Lys Phe Leu Ile Ser Phe Met Val Phe Phe Ile Leu His
225 230 235 240
Phe Val Gly Leu Ser Ile Glu Val Leu Cys Phe Ile Leu Pro Gln Asn
245 250 255
Lys Leu Leu Phe Ile Thr Gly Leu Thr Ala Thr Cys Leu Tyr Pro Cys
260 265 270
Gly His Ser Ile Ile Val Ile Leu Gly Asn Lys Gln Leu Lys Gln Ala
275 280 285
Ser Leu Lys Ala Leu Gln Gln Leu Lys Cys Cys Glu Thr Lys Gly Asn
290 295 300
Phe Arg Val Lys
305
<210>118
<211>1754
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R07,mGR07
<400>118
ttcataatga agaggaggca gggcaatgtt ggtttctgtt gtctgaccag tgtatttgac 60
agtgatacta cacatttgat tgctaaatgc aaatagttcc aaaggaacaa gtaaatttta 120
tgaaatagaa gcttctattt gcttattaac aaactgcaag caaacattag tctgcacaca 180
ttttatagac aagctaaatc ttcaaaagca ataaaaaaga gcacccataa agttctgact 240
ctatcacatg acaataggct tgaaaagatt gtctatgtag ataaagaaga tggcataact 300
tctccatcaa gaagccagta tatgggacat tctccagcag ataatttaca atagatgcag 360
cagaagtaac cttagagatc tgtaaagatg ctgaattcag cagaaggcat cctcctttgt 420
gttgtcacta gtgaggctgt gctcggagtt ttaggggaca catatattgc actttttaac 480
tgcatggact atgctaagaa caagaagctc tctaagatcg gtttcattct cattggcttg 540
gcgatttcca gaattggtgt tgtatggata ataattttac aagggtatat acaagtattt 600
tttccacaca tgcttacctc tggaaacata actgaatata ttacttacat atgggtattt 660
ctcaatcact taagtgtctg gtttgtcacc aacctcaaca tcctctactt tctaaagata 720
gctaattttt ccaactctgt atttctctgg ctgaaaagga gagtcaatgc agtttttatc 780
tttctgtcag gatgcttact tacctcatgg ttactatgtt ttccacaaat gacaaagata 840
cttcaaaata gtaaaatgca ccagagaaac acatcttggg tccaccagcg gaaaaattac 900
tttcttatta accaaagtgt gaccaatctg ggaatctttt tcttcattat tgtatccctg 960
attacctgct ttctgttgat tgttttcctc tggagacatg tcagacaaat gcactcagat 1020
gtttcaggat tcagagacca cagcacaaaa gtacatgtga aagctatgaa atttctaata 1080
tcttttatgg tcttctttat tctgcatttt gtaggccttt ccatagaagt gctatgcttt 1140
attctgccac aaaataaact gctctttata actggtttga cagccacatg cctctatccc 1200
tgcggtcact caatcatcgt aattttagga aataagcagt taaagcaagc ctctttgaag 1260
gcactgcagc aactaaaatg ctgtgagaca aaaggaaatt tcagagtcaa ataaatgggt 1320
ttgcaaataa atagctgcct tgttcttcca ctggttttta ccctgttagt tgatgttatg 1380
aaaagttcct gctatggttg atgacatctc aaggaatcta tttttctggt ggcatgttaa 1440
gtccacgtga agcctcactt catactgtga cttgactatg caaattcttt ccacaaaata 1500
accagataac attcagcctg gagataaatt catttaaagg cttttatggt gaggataaac 1560
aaaaaaaaaa aatcattttt ctgtgattca ctgtaactcc caggatgagt aaaagaaaac 1620
aagacaaatg gttgtgatca gcctttgtgt gtctagacag agctagggac cagatgttga 1680
tgcttgtgtg tggttttgag ttctttaaga agttattgcc tctctgccat tcggtattcc 1740
tcaggtgaga attc 1754
<210>119
<211>297
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R08,mGR08
<400>119
Met Leu Trp Glu Leu Tyr Val Phe Val Phe Ala Ala Ser Val Phe Leu
1 5 10 15
Asn Phe Val Gly Ile Ile Ala Asn Leu Phe Ile Ile Val Ile Ile Ile
20 25 30
Lys Thr Trp Val Asn Ser Arg Arg Ile Ala Ser Pro Asp Arg Ile Leu
35 40 45
Phe Ser Leu Ala Ile Thr Arg Phe Leu Thr Leu Gly Leu Phe Leu Leu
50 55 60
Asn Ser Val Tyr Ile Ala Thr Asn Thr Gly Arg Ser Val Tyr Phe Ser
65 70 75 80
Thr Phe Phe Leu Leu Cys Trp Lys Phe Leu Asp Ala Asn Ser Leu Trp
85 90 95
Leu Val Thr Ile Leu Asn Ser Leu Tyr Cys Val Lys Ile Thr Asn Phe
100 105 110
Gln His Pro Val Phe Leu Leu Leu Lys Arg Thr Ile Ser Met Lys Thr
115 120 125
Thr Ser Leu Leu Leu Ala Cys Leu Leu Ile Ser Ala Leu Thr Thr Leu
130 135 140
Leu Tyr Tyr Met Leu Ser Gln Ile Ser Arg Phe Pro Glu His Ile Ile
145 150 155 160
Gly Arg Asn Asp Thr Ser Phe Asp Leu Ser Asp Gly Ile Leu Thr Leu
165 170 175
Val Ala Ser Leu Val Leu Asn Ser Leu Leu Gln Phe Met Leu Asn Val
180 185 190
Thr Phe Ala Ser Leu Leu Ile His Ser Leu Arg Arg His Ile Gln Lys
195 200 205
Met Gln Arg Asn Arg Thr Ser Phe Trp Asn Pro Gln Thr Glu Ala His
210 215 220
Met Gly Ala Met Arg Leu Met Ile Cys Phe Leu Val Leu Tyr Ile Pro
225 230 235 240
Tyr Ser Ile Ala Thr Leu Leu Tyr Leu Pro Ser Tyr Met Arg Lys Asn
245 250 255
Leu Arg Ala Gln Ala Ile Cys Met Ile Ile Thr Ala Ala Tyr Pro Pro
260 265 270
Gly His Ser Val Leu Leu Ile Ile Thr His His Lys Leu Lys Ala Lys
275 280 285
Ala Lys Lys Ile Phe Cys Phe Tyr Lys
290 295
<210>120
<211>1475
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R08,mGR08
<400>120
aagcttgttt gtaattaggc attcctaaga aaataagaac aggagtgaag aaatagtaat 60
ttaatccttg aaagatttgc atctcagtaa aagcagctgc ctcttagacc agaaatggtg 120
tttgccatgc tggaaaataa aaaggagacc tctttccagg ctgcatcctg tgtctgctta 180
cttatttcag tttgttttca tcggcaccaa acgaggaaag atgctctggg aactgtatgt 240
atttgtgttt gctgcctcgg tttttttaaa ttttgtagga atcattgcaa atctatttat 300
tatagtgata attattaaga cttgggtcaa cagtcgcaga attgcctctc cggataggat 360
cctgttcagc ttggccatca ctagattcct gactttgggg ttgtttctac tgaacagtgt 420
ctacattgct acaaatactg gaaggtcagt ctacttttcc acattttttc tattgtgttg 480
gaagtttctg gatgcaaaca gtctctggtt agtgaccatt ctgaacagct tgtattgtgt 540
gaagattact aattttcaac acccagtgtt tctcctgttg aaacggacta tctctatgaa 600
gaccaccagc ctgctgttgg cctgtcttct gatttcagcc ctcaccactc tcctatatta 660
tatgctctca cagatatcac gttttcctga acacataatt gggagaaatg acacgtcatt 720
tgacctcagt gatggtatct tgacgttagt agcctctttg gtcctgaact cacttctaca 780
gtttatgctc aatgtgactt ttgcttcctt gttaatacat tccttgagaa gacatataca 840
gaagatgcag agaaacagga ccagcttttg gaatccccag acggaggctc acatgggtgc 900
tatgaggctg atgatctgtt tcctcgtgct ctacattcca tattcaattg ctaccctgct 960
ctatcttcct tcctatatga ggaagaatct gagagcccag gccatttgca tgattattac 1020
tgctgcttac cctccaggac attctgtcct cctcattatc acacatcata aactgaaagc 1080
taaagcaaag aagattttct gtttctacaa gtagcagaat ttcattagta gttaacagca 1140
tcaattcatg gtttggttgc attagaaatg tctcagtgat ctaaggactt aattttgtga 1200
tcttgtatct ggcatcctga ccctgagact aagtgcttat attttggtca atacagcatc 1260
ttttggctaa tattttaaag taaatcacat tccataagaa attgtttaag ggatttacgt 1320
atttttcatg gctatcacat tcctagacaa tggaaatcac catactgttt cgctagctac 1380
tgaagtacca ggggaaagtc catgaatgaa ggccacattg tgatgttctt ggttagcaca 1440
gattagagaa tttggcctca actgagcaag atatc 1475
<210>121
<211>316
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R09,mGR09
<400>121
Met Glu His Leu Leu Lys Arg Thr Phe Asp Ile Thr Glu Asn Ile Leu
1 5 10 15
Leu Ile Ile Leu Phe Ile Glu Leu Ile Ile Gly Leu Ile Gly Asn Gly
20 25 30
Phe Thr Ala Leu Val His Cys Met Asp Trp Val Lys Arg Lys Lys Met
35 40 45
Ser Leu Val Asn Lys Ile Leu Thr Ala Leu Ala Thr Ser Arg Ile Phe
50 55 60
Leu Leu Trp Phe Met Leu Val Gly Phe Pro Ile Ser Ser Leu Tyr Pro
65 70 75 80
Tyr Leu Val Thr Thr Arg Leu Met Ile Gln Phe Thr Ser Thr Leu Trp
85 90 95
Thr Ile Ala Asn His Ile Ser Val Trp Phe Ala Thr Cys Leu Ser Val
100 105 110
Phe Tyr Phe Leu Lys Ile Ala Asn Phe Ser Asn Ser Pro Phe Leu Tyr
115 120 125
Leu Lys Arg Arg Val Glu Lys Val Val Ser Val Thr Leu Leu Val Ser
130 135 140
Leu Val Leu Leu Phe Leu Asn Ile Leu Leu Leu Asn Leu Glu Ile Asn
145 150 155 160
Met Cys Ile Asn Glu Tyr His Gln Ile Asn Ile Ser Tyr Ile Phe Ile
165 170 175
Ser Tyr Tyr His Leu Ser Cys Gln Ile Gln Val Leu Gly Ser His Ile
180 185 190
Ile Phe Leu Ser Val Pro Val Val Leu Ser Leu Ser Thr Phe Leu Leu
195 200 205
Leu Ile Phe Ser Leu Trp Thr Leu His Lys Arg Met Gln Gln His Val
210 215 220
Gln Gly Gly Arg Asp Ala Arg Thr Thr Ala His Phe Lys Ala Leu Gln
225 230 235 240
Ala Val Ile Ala Phe Leu Leu Leu Tyr Ser Ile Phe Ile Leu Ser Leu
245 250 255
Leu Leu Gln Phe Trp Ile His Gly Leu Arg Lys Lys Pro Pro Phe Ile
260 265 270
Ala Phe Cys Gln Val Val Asp Thr Ala Phe Pro Ser Phe His Ser Tyr
275 280 285
Val Leu Ile Leu Arg Asp Arg Lys Leu Arg His Ala Ser Leu Ser Val
290 295 300
Leu Ser Trp Leu Lys Cys Arg Pro Asn Tyr Val Lys
305 310 315
<210>122
<211>1339
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R09,mGR09
<400>122
gaattcagaa atcatcaaaa aatcttcaaa actacatgtt taaaatagca cttcaaatga 60
atacatttgc aaatctttac aactaataca taaaatggag catcttttga agagaacatt 120
tgatatcacc gagaacatac ttctaattat tttattcatt gaattaataa ttggacttat 180
aggaaacgga ttcacagcct tggtgcactg catggactgg gttaagagaa aaaaaatgtc 240
attagttaat aaaatcctca ccgctttggc aacttctaga attttcctgc tctggttcat 300
gctagtaggt tttccaatta gctcactgta cccatattta gttactacta gactgatgat 360
acagttcact agtactctat ggactatagc taaccatatt agtgtctggt ttgctacatg 420
cctcagtgtc ttttattttc tcaagatagc caatttttct aattctcctt ttctctatct 480
aaagaggaga gttgaaaaag tagtttcagt tacattactg gtgtctctgg tcctcttgtt 540
tttaaatatt ttactactta atttggaaat taacatgtgt ataaatgaat atcatcaaat 600
aaacatatca tacatcttca tttcttatta ccatttaagt tgtcaaattc aggtgttagg 660
aagtcacatt attttcctgt ctgtccccgt tgttttgtcc ctgtcaactt ttctcctgct 720
catcttctcc ctgtggacac ttcacaagag gatgcagcag catgttcagg gaggcagaga 780
tgccagaacc acggcccact tcaaagcctt gcaagcagtg attgcctttc tcctactata 840
ctccattttt atcctgtcac tgttactaca attttggatc catggattaa ggaagaaacc 900
tcctttcatt gcattttgtc aggttgtaga tacagctttt ccttcattcc attcatatgt 960
cttgattctg agagacagga agctgagaca cgcctctctc tctgtgttgt cgtggctgaa 1020
atgcaggcca aattatgtga aataatattt ctttgtattt tcattttcaa ttttaaaata 1080
ttcttagaat ttgactgcat gtatttcatc ttttatttga aacaaccact aattaaagct 1140
attactaatt tagcaagtcg tatacaaggt tattttttaa tacacatatc aaaaactgac 1200
atgtttatgt tctacaaaaa cctgaatata tcaaaattat ataaattttg tatcaacgat 1260
taacaatgga gtttttttat ttatgacctg tcacgggact ccggtggagt cagcttgtca 1320
gatgaaagtc tgaaagctt 1339
<210>123
<211>333
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R10,mGR10
<400>123
Met Phe Ser Gln Ile Ile Ser Thr Ser Asp Ile Phe Thr Phe Thr Ile
1 5 10 15
Ile Leu Phe Val Glu Leu Val Ile Gly Ile Leu Gly Asn Gly Phe Ile
20 25 30
Ala Leu Val Asn Ile Met Asp Trp Thr Lys Arg Arg Ser Ile Ser Ser
35 40 45
Ala Asp Gln Ile Leu Thr Ala Leu Ala Ile Thr Arg Phe Leu Tyr Val
50 55 60
Trp Phe Met Ile Ile Cys Ile Leu Leu Phe Met Leu Cys Pro His Leu
65 70 75 80
Leu Thr Arg Ser Glu Ile Val Thr Ser Ile Gly Ile Ile Trp Ile Val
85 90 95
Asn Asn His Phe Ser Val Trp Leu Ala Thr Cys Leu Gly Val Phe Tyr
100 105 110
Phe Leu Lys Ile Ala Asn Phe Ser Asn Ser Leu Phe Leu Tyr Leu Lys
115 120 125
Trp Arg Val Lys Lys Val Val Leu Met Ile Ile Gln Val Ser Met Ile
130 135 140
Phe Leu Ile Leu Asn Leu Leu Ser Leu Ser Met Tyr Asp Gln Phe Ser
145 150 155 160
Ile Asp Val Tyr Glu Gly Asn Thr Ser Tyr Asn Leu Gly Asp Ser Thr
165 170 175
Pro Phe Pro Thr Ile Ser Leu Phe Ile Asn Ser Ser Lys Val Phe Val
180 185 190
Ile Thr Asn Ser Ser His Ile Phe Leu Pro Ile Asn Ser Leu Phe Met
195 200 205
Leu Ile Pro Phe Thr Val Ser Leu Val Ala Phe Leu Met Leu Ile Phe
210 215 220
Ser Leu Trp Lys His His Lys Lys Met Gln Val Asn Ala Lys Pro Pro
225 230 235 240
Arg Asp Ala Ser Thr Met Ala His Ile Lys Ala Leu Gln Thr Gly Phe
245 250 255
Ser Phe Leu Leu Leu Tyr Ala Val Tyr Leu Leu Phe Ile Val Ile Gly
260 265 270
Met Leu Ser Leu Arg Leu Ile Gly Gly Lys Leu Ile Leu Leu Phe Asp
275 280 285
His Ile Ser Gly Ile Gly Phe Pro Ile Ser His Ser Phe Val Leu Ile
290 295 300
Leu Gly Asn Asn Lys Leu Arg Gln Ala Ser Leu Ser Val Leu His Cys
305 310 315 320
Leu Arg Cys Arg Ser Lys Asp Met Asp Thr Met Gly Pro
325 330
<210>124
<211>1371
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R10,mGR10
<400>124
Gly Ala Ala Thr Thr Cys Ala Ala Cys Ala Thr Cys Thr Thr Ala Thr
1 5 10 15
Thr Cys Ala Ala Cys Thr Thr Cys Ala Gly Ala Ala Ala Ala Cys Thr
20 25 30
Gly Gly Ala Thr Ala Thr Thr Ala Gly Ala Cys Ala Cys Ala Gly Thr
35 40 45
Gly Thr Cys Thr Gly Gly Ala Thr Gly Ala Ala Gly Cys Ala Gly Ala
50 55 60
Gly Gly Thr Gly Ala Thr Cys Thr Cys Thr Thr Thr Gly Gly Gly Ala
65 70 75 80
Ala Ala Ala Ala Ala Ala Gly Cys Cys Ala Ala Gly Thr Ala Gly Thr
85 90 95
Cys Ala Thr Ala Ala Ala Gly Ala Ala Thr Thr Thr Ala Thr Gly Ala
100 105 110
Ala Ala Cys Ala Ala Thr Thr Cys Cys Thr Gly Gly Gly Ala Thr Thr
115 120 125
Gly Thr Thr Thr Ala Thr Ala Thr Thr Thr Gly Thr Thr Ala Cys Ala
130 135 140
Ala Ala Cys Ala Ala Ala Thr Thr Thr Ala Thr Ala Thr Gly Thr Thr
145 150 155 160
Thr Gly Thr Thr Ala Gly Thr Cys Ala Gly Thr Ala Ala Thr Gly Thr
165 170 175
Ala Thr Ala Ala Gly Thr Gly Gly Gly Ala Thr Thr Thr Thr Ala Ala
180 185 190
Ala Gly Cys Ala Thr Gly Ala Thr Thr Ala Thr Cys Thr Thr Gly Ala
195 200 205
Ala Thr Thr Thr Thr Thr Ala Ala Cys Ala Ala Ala Ala Ala Ala Cys
210 215 220
Ala Thr Gly Thr Ala Gly Thr Gly Cys Thr Thr Thr Thr Thr Ala Ala
225 230 235 240
Ala Thr Gly Thr Ala Gly Cys Ala Gly Ala Ala Ala Cys Ala Thr Thr
245 250 255
Ala Ala Ala Ala Ala Thr Thr Gly Ala Ala Gly Cys Ala Thr Gly Thr
260 265 270
Thr Cys Thr Cys Ala Cys Ala Gly Ala Thr Ala Ala Thr Ala Ala Gly
275 280 285
Cys Ala Cys Cys Ala Gly Thr Gly Ala Thr Ala Thr Thr Thr Thr Thr
290 295 300
Ala Cys Thr Thr Thr Thr Ala Cys Ala Ala Thr Ala Ala Thr Ala Thr
305 310 315 320
Thr Ala Thr Thr Thr Gly Thr Gly Gly Ala Ala Thr Thr Ala Gly Thr
325 330 335
Ala Ala Thr Ala Gly Gly Ala Ala Thr Thr Thr Thr Ala Gly Gly Ala
340 345 350
Ala Ala Thr Gly Gly Ala Thr Thr Cys Ala Thr Ala Gly Cys Ala Cys
355 360 365
Thr Ala Gly Thr Gly Ala Ala Thr Ala Thr Cys Ala Thr Gly Gly Ala
370 375 380
Cys Thr Gly Gly Ala Cys Cys Ala Ala Gly Ala Gly Ala Ala Gly Ala
385 390 395 400
Ala Gly Cys Ala Thr Thr Thr Cys Ala Thr Cys Ala Gly Cys Gly Gly
405 410 415
Ala Thr Cys Ala Gly Ala Thr Thr Cys Thr Cys Ala Cys Thr Gly Cys
420 425 430
Thr Thr Thr Gly Gly Cys Cys Ala Thr Thr Ala Cys Cys Ala Gly Ala
435 440 445
Thr Thr Thr Cys Thr Cys Thr Ala Thr Gly Thr Gly Thr Gly Gly Thr
450 455 460
Thr Thr Ala Thr Gly Ala Thr Cys Ala Thr Thr Thr Gly Thr Ala Thr
465 470 475 480
Ala Thr Thr Gly Thr Thr Ala Thr Thr Cys Ala Thr Gly Cys Thr Gly
485 490 495
Thr Gly Cys Cys Cys Ala Cys Ala Thr Thr Thr Gly Cys Thr Thr Ala
500 505 510
Cys Ala Ala Gly Ala Thr Cys Ala Gly Ala Ala Ala Thr Ala Gly Thr
515 520 525
Ala Ala Cys Ala Thr Cys Ala Ala Thr Thr Gly Gly Thr Ala Thr Thr
530 535 540
Ala Thr Thr Thr Gly Gly Ala Thr Ala Gly Thr Gly Ala Ala Thr Ala
545 550 555 560
Ala Cys Cys Ala Thr Thr Thr Cys Ala Gly Cys Gly Thr Thr Thr Gly
565 570 575
Gly Cys Thr Thr Gly Cys Cys Ala Cys Ala Thr Gly Cys Cys Thr Cys
580 585 590
Gly Gly Thr Gly Thr Cys Thr Thr Thr Thr Ala Thr Thr Thr Thr Cys
595 600 605
Thr Gly Ala Ala Gly Ala Thr Ala Gly Cys Cys Ala Ala Thr Thr Thr
610 615 620
Thr Thr Cys Thr Ala Ala Cys Thr Cys Thr Thr Thr Gly Thr Thr Thr
625 630 635 640
Cys Thr Thr Thr Ala Cys Cys Thr Ala Ala Ala Gly Thr Gly Gly Ala
645 650 655
Gly Ala Gly Thr Thr Ala Ala Ala Ala Ala Ala Gly Thr Ala Gly Thr
660 665 670
Thr Thr Thr Ala Ala Thr Gly Ala Thr Ala Ala Thr Ala Cys Ala Gly
675 680 685
Gly Thr Ala Thr Cys Ala Ala Thr Gly Ala Thr Thr Thr Thr Cys Thr
690 695 700
Thr Gly Ala Thr Thr Thr Thr Ala Ala Ala Cys Cys Thr Gly Thr Thr
705 710 715 720
Ala Thr Cys Thr Cys Thr Ala Ala Gly Cys Ala Thr Gly Thr Ala Thr
725 730 735
Gly Ala Thr Cys Ala Gly Thr Thr Cys Thr Cys Ala Ala Thr Thr Gly
740 745 750
Ala Thr Gly Thr Thr Thr Ala Thr Gly Ala Ala Gly Gly Ala Ala Ala
755 760 765
Thr Ala Cys Ala Thr Cys Thr Thr Ala Thr Ala Ala Thr Thr Thr Ala
770 775 780
Gly Gly Gly Gly Ala Thr Thr Cys Ala Ala Cys Cys Cys Cys Ala Thr
785 790 795 800
Thr Thr Cys Cys Cys Ala Cys Ala Ala Thr Thr Thr Cys Cys Thr Thr
805 810 815
Ala Thr Thr Cys Ala Thr Cys Ala Ala Thr Thr Cys Ala Thr Cys Ala
820 825 830
Ala Ala Ala Gly Thr Thr Thr Thr Cys Gly Thr Ala Ala Thr Cys Ala
835 840 845
Cys Cys Ala Ala Cys Thr Cys Ala Thr Cys Cys Cys Ala Thr Ala Thr
850 855 860
Thr Thr Thr Cys Thr Thr Ala Cys Cys Cys Ala Thr Cys Ala Ala Cys
865 870 875 880
Thr Cys Cys Cys Thr Gly Thr Thr Cys Ala Thr Gly Cys Thr Cys Ala
885 890 895
Thr Ala Cys Cys Cys Thr Thr Cys Ala Cys Ala Gly Thr Gly Thr Cys
900 905 910
Cys Cys Thr Gly Gly Thr Ala Gly Cys Cys Thr Thr Thr Cys Thr Cys
915 920 925
Ala Thr Gly Cys Thr Cys Ala Thr Cys Thr Thr Cys Thr Cys Ala Cys
930 935 940
Thr Gly Thr Gly Gly Ala Ala Gly Cys Ala Thr Cys Ala Cys Ala Ala
945 950 955 960
Ala Ala Ala Gly Ala Thr Gly Cys Ala Gly Gly Thr Cys Ala Ala Thr
965 970 975
Gly Cys Cys Ala Ala Ala Cys Cys Ala Cys Cys Thr Ala Gly Ala Gly
980 985 990
Ala Thr Gly Cys Cys Ala Gly Cys Ala Cys Cys Ala Thr Gly Gly Cys
995 1000 1005
Cys Cys Ala Cys Ala Thr Thr Ala Ala Ala Gly Cys Cys Thr Thr Gly
1010 1015 1020
Cys Ala Ala Ala Cys Ala Gly Gly Gly Thr Thr Cys Thr Cys Cys Thr
1025 1030 1035 1040
Thr Cys Cys Thr Gly Cys Thr Gly Cys Thr Gly Thr Ala Thr Gly Cys
1045 1050 1055
Ala Gly Thr Ala Thr Ala Cys Thr Thr Ala Cys Thr Thr Thr Thr Thr
1060 1065 1070
Ala Thr Thr Gly Thr Cys Ala Thr Ala Gly Gly Ala Ala Thr Gly Thr
1075 1080 1085
Thr Gly Ala Gly Cys Cys Thr Thr Ala Gly Gly Thr Thr Gly Ala Thr
1090 1095 1100
Ala Gly Gly Ala Gly Gly Ala Ala Ala Ala Thr Thr Ala Ala Thr Ala
1105 1110 1115 1120
Cys Thr Thr Thr Thr Ala Thr Thr Thr Gly Ala Cys Cys Ala Cys Ala
1125 1130 1135
Thr Thr Thr Cys Thr Gly Gly Ala Ala Thr Ala Gly Gly Thr Thr Thr
1140 1145 1150
Thr Cys Cys Thr Ala Thr Ala Ala Gly Cys Cys Ala Cys Thr Cys Ala
1155 1160 1165
Thr Thr Thr Gly Thr Gly Cys Thr Gly Ala Thr Thr Cys Thr Gly Gly
1170 1175 1180
Gly Ala Ala Ala Thr Ala Ala Cys Ala Ala Gly Cys Thr Gly Ala Gly
1185 1190 1195 1200
Ala Cys Ala Ala Gly Cys Cys Ala Gly Thr Cys Thr Thr Thr Cys Ala
1205 1210 1215
Gly Thr Gly Thr Thr Gly Cys Ala Thr Thr Gly Thr Cys Thr Gly Ala
1220 1225 1230
Gly Gly Thr Gly Cys Cys Gly Ala Thr Cys Cys Ala Ala Ala Gly Ala
1235 1240 1245
Thr Ala Thr Gly Gly Ala Cys Ala Cys Cys Ala Thr Gly Gly Gly Thr
1250 1255 1260
Cys Cys Ala Thr Ala Ala Ala Ala Ala Ala Thr Thr Thr Cys Ala Gly
1265 1270 1275 1280
Ala Gly Gly Thr Cys Ala Thr Thr Gly Gly Gly Ala Ala Ala Cys Ala
1285 1290 1295
Thr Thr Thr Thr Gly Ala Gly Ala Thr Cys Thr Thr Ala Thr Ala Gly
1300 1305 1310
Gly Gly Gly Ala Ala Ala Ala Ala Gly Ala Ala Ala Ala Thr Gly Thr
1315 1320 1325
Gly Gly Gly Gly Cys Thr Thr Cys Ala Ala Ala Gly Cys Thr Gly Gly
1330 1335 1340
Thr Ala Gly Gly Ala Gly Thr Ala Ala Thr Ala Thr Ala Gly Ala Gly
1345 1350 1355 1360
Ala Ala Gly Gly Ala Thr Ala Gly Gly Ala Gly
1365 1370
<210>125
<211>303
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R11,mGR11
<220>
<221>MOD_RES
<222>(169)
<223>Xaa=任何氨基酸
<400>125
Met Glu His Pro Leu Arg Arg Thr Phe Asp Phe Ser Gln Ser Ile Leu
1 5 10 15
Leu Thr Ile Leu Phe Ile Glu Leu Ile Ile Gly Leu Ile Arg Asn Gly
20 25 30
Leu Met Val Leu Val His Cys Ile Asp Trp Val Lys Arg Lys Lys Phe
35 40 45
His Leu Leu Ile Lys Ser Ser Pro Leu Trp Gln Thr Ser Arg Ile Cys
50 55 60
Leu Leu Trp Phe Met Leu Ile His Leu Leu Ile Thr Leu Leu Tyr Ala
65 70 75 80
Asp Leu Ala Ser Thr Arg Thr Met Met Gln Phe Ala Ser Asn Pro Trp
85 90 95
Thr Ile Ser Asn His Ile Ser Ile Trp Leu Ala Thr Cys Leu Gly Val
100 105 110
Phe Tyr Phe Leu Lys Ile Ala Asn Phe Ser Asn Ser Thr Phe Leu Tyr
115 120 125
Leu Lys Trp Arg Val Gln Phe Leu Leu Leu Asn Ile Leu Leu Val Lys
130 135 140
Phe Glu Ile Asn Met Trp Ile Asn Glu Tyr His Gln Ile Asn Ile Pro
145 150 155 160
Tyr Ser Phe Ile Ser Tyr Tyr Gln Xaa Cys Gln Ile Gln Val Leu Ser
165 170 175
Leu His Ile Ile Phe Leu Ser Val Pro Phe Ile Leu Ser Leu Ser Thr
180 185 190
Phe Leu Leu Leu Ile Phe Ser Leu Trp Thr Leu His Gln Arg Met Gln
195 200 205
Gln His Val Gln Gly Tyr Arg Asp Ala Ser Thr Met Ala His Phe Lys
210 215 220
Ala Leu Gln Ala Val Ile Ala Phe Leu Leu Ile His Ser Ile Phe Ile
225 230 235 240
Leu Ser Leu Leu Leu Gln Leu Trp Lys His Glu Leu Arg Lys Lys Pro
245 250 255
Pro Phe Val Val Phe Cys Gln Val Ala Tyr Ile Ala Phe Pro Ser Ser
260 265 270
His Ser Tyr Val Phe Ile Leu Gly Asp Arg Lys Leu Arg Gln Ala Cys
275 280 285
Leu Ser Val Leu Trp Arg Leu Lys Cys Arg Pro Asn Tyr Val Gly
290 295 300
<210>126
<211>1108
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R11,mGR11
<400>126
aataatgtat gtggaagagt taagtataaa tgttgtatga gaatgaactc agaaatcatc 60
aaaaatcttt aaaactgcat gttaaaaatc acacttcaaa tgaatatatt tgtaattctt 120
tagaactaat aaataaaatg gagcatcctt tgaggagaac atttgatttc tcccagagca 180
tacttctaac cattttattc attgaattaa taattggact tataagaaat ggattaatgg 240
tattggtgca ctgcatagat tgggttaaga gaaaaaaatt tcatttgtta atcaaatcct 300
caccactttg gcaaacttcc agaatttgtc tgctctggtt catgctaata catctcctga 360
ttactttatt gtatgcagat ttagctagta ctagaacgat gatgcaattc gctagcaatc 420
catggactat atctaaccat atcagcatct ggcttgctac atgccttggt gtcttttatt 480
ttctcaagat agccaatttt tctaactcta cttttctcta tctaaaatgg cgagttcagt 540
tcctcttgtt aaatatttta ctggttaaat ttgagattaa catgtggata aatgaatatc 600
atcaaataaa cataccatac agcttcattt cttattacca aattgtcaaa tacaggtgtt 660
aagtcttcac attattttcc tgtctgtccc ctttattttg tccctgtcaa cttttctcct 720
gctcatcttc tccctgtgga cacttcacca gaggatgcag cagcatgttc aaggatacag 780
agatgccagc acaatggccc acttcaaagc cttgcaagca gtgattgcct ttctcttaat 840
acactccatt tttatcctgt cactgttact acaactttgg aaacatgaat taaggaagaa 900
acctcctttt gttgtatttt gtcaggttgc atatatagct tttccttcat cccattcata 960
tgtcttcatt ctgggagaca gaaagctgag acaggcttgt ctctctgtgt tgtggaggct 1020
gaaatgcagg ccaaattatg tgggataaaa tctctttgtg ctttcatttc caattcttaa 1080
atattctttg attttgactg cataaatt 1108
<210>127
<211>150
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R12,mGR12
<400>127
Gly Ala Ile Val Asn Val Asp Phe Leu Ile Gly Asn Val Gly Asn Gly
1 5 10 15
Phe Ile Val Val Ala Asn Ile Met Asp Leu Val Lys Arg Arg Lys Leu
20 25 30
Ser Ser Val Asp Gln Leu Leu Thr Ala Leu Ala Val Ser Arg Ile Thr
35 40 45
Leu Leu Trp Tyr Leu Tyr Ile Met Lys Arg Thr Phe Leu Val Asp Pro
50 55 60
Asn Ile Gly Ala Ile Met Gln Ser Thr Arg Leu Thr Asn Val Ile Trp
65 70 75 80
Ile Ile Ser Asn His Phe Ser Ile Trp Leu Ala Thr Thr Leu Ser Ile
85 90 95
Phe Tyr Phe Leu Lys Ile Ala Asn Phe Ser Asn Ser Ile Phe Cys Tyr
100 105 110
Leu Arg Trp Arg Phe Glu Lys Val Ile Leu Met Ala Leu Leu Val Ser
115 120 125
Leu Val Leu Leu Phe Ile Asp Ile Leu Val Thr Asn Met Tyr Ile Asn
130 135 140
Ile Trp Thr Asp Glu Phe
145 150
<210>128
<211>520
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R12,mGR12
<400>128
ttttcagcag tgactttggg aagcagaacg tcctcttaga gacagtgggt gctgctatcc 60
tagttaatgt ggagcaatag ttaatgtgga tttcctaatt ggaaatgttg ggaatggatt 120
cattgttgtg gcaaacataa tggacttggt caagagaaga aagctttctt cagtggatca 180
gctgctcact gcactggccg tctccagaat cactttgctg tggtacctgt acataatgaa 240
acgaacattt ttagtggatc caaacattgg tgcaattatg caatcaacaa gactgactaa 300
tgttatctgg ataatttcta accattttag tatatggctg gccaccaccc tcagcatctt 360
ttattttctc aagatagcaa atttttctaa ctctattttc tgttacctga ggtggagatt 420
tgaaaaggtg attttgatgg cattgctggt gtccctggtc ctcttgttta tagatatttt 480
agtaacaaac atgtacatta atatttggac tgatgaattc 520
<210>129
<211>309
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R13,mGR13
<400>129
Met Val Ala Val Leu Gln Ser Thr Leu Pro Ile Ile Phe Ser Met Glu
1 5 10 15
Phe Ile Met Gly Thr Leu Gly Asn Gly Phe Ile Phe Leu Ile Val Cys
20 25 30
Ile Asp Trp Val Gln Arg Arg Lys Ile Ser Leu Val Asp Gln Ile Arg
35 40 45
Thr Ala Leu Ala Ile Ser Arg Ile Ala Leu Ile Trp Leu Ile Phe Leu
50 55 60
Asp Trp Trp Val Ser Val His Tyr Pro Ala Leu His Glu Thr Gly Lys
65 70 75 80
Met Leu Ser Thr Tyr Leu Ile Ser Trp Thr Val Ile Asn His Cys Asn
85 90 95
Phe Trp Leu Thr Ala Asn Leu Ser Ile Leu Tyr Phe Leu Lys Ile Ala
100 105 110
Asn Phe Ser Asn Ile Ile Phe Leu Tyr Leu Lys Phe Arg Ser Lys Asn
115 120 125
Val Val Leu Val Thr Leu Leu Val Ser Leu Phe Phe Leu Phe Leu Asn
130 135 140
Thr Val Ile Ile Lys Ile Phe Ser Asp Val Cys Phe Asp Ser Val Gln
145 150 155 160
Arg Asn Val Ser Gln Ile Phe Ile Met Tyr Asn His Glu Gln Ile Cys
165 170 175
Lys Phe Leu Ser Phe Thr Asn Pro Met Phe Thr Phe Ile Pro Phe Val
180 185 190
Met Ser Thr Val Met Phe Ser Leu Leu Ile Phe Ser Leu Trp Arg His
195 200 205
Leu Lys Asn Met Gln His Thr Ala Lys Gly Cys Arg Asp Ile Ser Thr
210 215 220
Thr Val His Ile Arg Ala Leu Gln Thr Ile Ile Val Ser Val Val Leu
225 230 235 240
Tyr Thr Ile Phe Phe Leu Ser Phe Phe Val Lys Val Trp Ser Phe Val
245 250 255
Ser Pro Glu Arg Tyr Leu Ile Phe Leu Phe Val Trp Ala Leu Gly Asn
260 265 270
Ala Val Phe Ser Ala His Pro Phe Val Met Ile Leu Val Asn Arg Arg
275 280 285
Leu Arg Leu Ala Ser Leu Ser Leu Ile Phe Trp Leu Trp Tyr Arg Phe
290 295 300
Lys Asn Ile Glu Val
305
<210>130
<211>1199
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R13,mGR13
<400>130
aagcttgttt gtgtttggat gaattctatt tatgtctatc aatttaagat tttcatatga 60
atcattaaga aatcttgata gttgtttgtg agatatcact tctgcaattt ttaaatgaaa 120
ttacactcat attttgaagg aacaatatgt tttaaaggaa tatattaaca aatcttcagc 180
agttacctca gaagtttggg tattgtttta cagaaaatgg tggcagttct acagagcaca 240
cttccaataa ttttcagtat ggaattcata atgggaacct taggaaatgg attcattttt 300
ctgatagtct gcatagactg ggtccaaaga agaaaaatct ctttagtgga tcaaatccgc 360
actgctctgg caattagcag aatcgctcta atttggttga tattcctaga ttggtgggtg 420
tctgttcatt acccagcatt acatgaaact ggtaagatgt tatcaacata tttgatttcc 480
tggacggtga tcaatcattg taacttttgg cttactgcaa acttgagcat cctttatttt 540
ctcaagatag ccaacttttc taacattatt tttctttatc taaagtttag atctaaaaat 600
gtggtattag tgaccctgtt agtgtctcta tttttcttgt tcttaaatac tgtaattata 660
aaaatatttt ctgatgtgtg ttttgatagt gttcaaagaa atgtgtctca aattttcata 720
atgtataacc atgaacaaat ttgtaaattt ctttccttta ctaaccctat gttcacattc 780
ataccttttg ttatgtccac ggtaatgttt tctttgctca tcttctccct gtggagacat 840
ctgaagaata tgcagcacac cgccaaagga tgcagagaca tcagcaccac agtgcacatc 900
agagccctgc aaaccatcat tgtgtctgta gtgctataca ctattttttt tctatcattt 960
tttgttaaag tttggagttt tgtgtcacca gagagatacc tgatcttttt gtttgtctgg 1020
gctctgggaa atgctgtttt ttctgctcac ccatttgtca tgattttggt aaacagaaga 1080
ttgagattgg cttctctctc tctgattttt tggctctggt acaggtttaa aaatatagaa 1140
gtatagggtc caaagaccac caaggaatca ttttccttat cctaaagaaa aatcaggag 1199
<210>131
<211>309
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R14,mGR14
<400>131
Met Leu Ser Thr Met Glu Gly Val Leu Leu Ser Val Ser Thr Ser Glu
1 5 10 15
Ala Val Leu Gly Ile Val Gly Asn Thr Phe Ile Ala Leu Val Asn Cys
20 25 30
Met Asp Tyr Asn Arg Asn Lys Lys Leu Ser Asa Ile Gly Phe Ile Leu
35 40 45
Thr Gly Leu Ala Ile Ser Arg Ile Cys Leu Val Leu Ile Leu Ile Thr
50 55 60
Glu Ala Tyr Ile Lys Ile Phe Tyr Pro Gln Leu Leu Ser Pro Val Asn
65 70 75 80
Ile Ile Glu Leu Ile Ser Tyr Leu Trp Ile Ile Ile Cys Gln Leu Asn
85 90 95
Val Trp Phe Ala Thr Ser Leu Ser Ile Phe Tyr Phe Leu Lys Ile Ala
100 105 110
Asn Phe Ser His Tyr Ile Phe Val Trp Leu Lys Arg Arg Ile Asp Leu
115 120 125
Val Phe Phe Phe Leu Ile Gly Cys Leu Leu Ile Ser Trp Leu Phe Ser
130 135 140
Phe Pro Val Val Ala Lys Met Val Lys Asp Asn Lys Met Leu Tyr Ile
145 150 155 160
Asn Thr Ser Trp Gln Ile His Met Lys Lys Ser Glu Leu Ile Ile Asn
165 170 175
Tyr Val Phe Thr Asn Gly Gly Val Phe Leu Phe Phe Met Ile Met Leu
180 185 190
Ile Val Cys Phe Leu Leu Ile Ile Ser Leu Trp Arg His Arg Arg Gln
195 200 205
Met Glu Ser Asn Lys Leu Gly Phe Arg Asp Leu Asn Thr Glu Val His
210 215 220
Val Arg Thr Ile Lys Val Leu Leu Ser Phe Ile Ile Leu Phe Ile Leu
225 230 235 240
His Phe Met Gly Ile Thr Ile Asn Val Ile Cys Leu Leu Ile Pro Glu
245 250 255
Ser Asn Leu Leu Phe Met Phe Gly Leu Thr Thr Ala Phe Ile Tyr Pro
260 265 270
Gly Cys His Ser Leu Ile Leu Ile Leu Ala Asn Ser Arg Leu Lys Gln
275 280 285
Cys Ser Val Met Ile Leu Gln Leu Leu Lys Cys Cys Glu Asn Gly Lys
290 295 300
Glu Leu Arg Asp Thr
305
<210>132
<211>1535
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R14,mGR14
<400>132
ctgcaggtat atacctaccc tgaaggcttc atctagagta aacaaagtag tctgtatagt 60
ctgccattcc tcagattctc ctcaacttcc caccctccag tgacctttct ccttttctac 120
agtcaaacta tggacctcac aacctgacac ttcttcagat gcaaaatatt ctcacagaga 180
caagtaaaac atacaaaaca aatactttaa tttgcctatt aacaaatggc aagaaaagat 240
tcaggcttga acatcctgta gacaagctaa ggacaggagc aactgaaggg atctccatga 300
agacctttca gatttctacc aaaagtaatt tttaactata tttaagtctt taaagaaaga 360
aagtaaagcc actcttttat tgaacagcaa tagattggaa tcttaaacaa ctgcaacaga 420
agccatttta aagatcaaca aagatgctga gcacaatgga aggtgtcctc ctttcagttt 480
caactagtga ggctgtgctg ggcattgtag ggaacacatt cattgcactt gtaaactgta 540
tggactataa caggaacaag aagctctcta atattggctt tattctcact ggcttggcaa 600
tttccagaat ttgccttgtg ttgatcttaa tcacagaggc atacataaaa atattctatc 660
cacagttgct gtctcctgtc aacataattg agctcatcag ttatctatgg ataattatct 720
gtcaattgaa tgtctggttt gccactagtc tcagtatttt ttatttcctg aagatagcaa 780
atttttccca ctacatattt gtctggttaa aaagaagaat tgatttagtt tttttcttcc 840
tgatagggtg cttgcttatc tcatggctat tttctttccc agttgttgcg aagatggtta 900
aagataataa aatgctgtat ataaacacat cttggcagat ccacatgaag aaaagtgagt 960
taatcattaa ctatgttttc accaatgggg gagtattttt attttttatg ataatgttaa 1020
ttgtatgttt cctgttaatc atttcacttt ggagacatcg caggcagatg gaatcaaata 1080
aattaggatt cagagatctc aacacagaag ttcatgtgag aacaataaaa gttttattgt 1140
cttttattat cctttttata ttgcatttca tgggtattac cataaatgta atttgtctgt 1200
taatcccaga aagcaacttg ttattcatgt ttggtttgac aactgcattc atctatcccg 1260
gctgccactc acttatccta attctagcaa acagtcggct gaagcagtgc tctgtaatga 1320
tactgcaact attaaagtgc tgtgagaatg gtaaagaact cagagacaca tgacagtctg 1380
gaacacatgc aatctggaat tgtcagtgga aaaagttact gaagatcttt tcacttgcac 1440
tatgctcttt tattgatttg gcatcattat caaacactgt tggagccttg tgaactcttg 1500
ttcagagtct tctgcctctc aaggaatcac actcc 1535
<210>133
<211>310
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R15,mGR15
<400>133
Met Cys Ala Val Leu Arg Ser Ile Leu Thr Ile Ile Phe Ile Leu Glu
1 5 10 15
Phe Phe Ile Gly Asn Leu Gly Asn Gly Phe Ile Ala Leu Val Gln Cys
20 25 30
Met Asp Leu Arg Lys Arg Arg Thr Phe Pro Ser Ala Asp His Phe Leu
35 40 45
Thr Ala Leu Ala Ile Ser Arg Leu Ala Leu Ile Trp Val Leu Phe Leu
50 55 60
Asp Ser Phe Leu Phe Ile Gln Ser Pro Leu Leu Met Thr Arg Asn Thr
65 70 75 80
Leu Arg Leu Ile Gln Thr Ala Trp Asn Ile Ser Asn His Phe Ser Ile
85 90 95
Trp Phe Ala Thr Ser Leu Ser Ile Phe Tyr Leu Phe Lys Ile Ala Ile
100 105 110
Phe Ser Asn Tyr Leu Phe Phe Tyr Leu Lys Arg Arg Val Lys Arg Val
115 120 125
Val Leu Val Ile Leu Leu Leu Ser Met Ile Leu Leu Phe Phe Asn Ile
130 135 140
Phe Leu Glu Ile Lys His Ile Asp Val Trp Ile Tyr Gly Thr Lys Arg
145 150 155 160
Asn Ile Thr Asn Gly Leu Ser Ser Asn Ser Phe Ser Glu Phe Ser Arg
165 170 175
Leu Ile Leu Ile Pro Ser Leu Met Phe Thr Leu Val Pro Phe Gly Val
180 185 190
Ser Leu Ile Ala Phe Leu Leu Leu Ile Phe Ser Leu Met Lys His Val
195 200 205
Arg Lys Met Gln Tyr Tyr Thr Lys Gly Cys Lys Asp Val Arg Thr Met
210 215 220
Ala His Thr Thr Ala Leu Gln Thr Val Val Ala Phe Leu Leu Leu Tyr
225 230 235 240
Thr Thr Phe Phe Leu Ser Leu Val Val Glu Val Ser Thr Leu Glu Met
245 250 255
Asp Glu Ser Leu Met Leu Leu Phe Ala Lys Val Thr Ile Met Ile Phe
260 265 270
Pro Ser Ile His Ser Cys Ile Phe Ile Leu Lys His Asn Lys Leu Arg
275 280 285
Gln Asp Leu Leu Ser Val Leu Lys Trp Leu Gln Tyr Trp Cys Lys Arg
290 295 300
Glu Lys Thr Leu Asp Ser
305 310
<210>134
<211>1482
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R15,mGR15
<400>134
aataatagat tttttaatat tcagaatttt taagtaatgt agtattgtta gcagcatagc 60
ttataggaaa agttccaagt aattttgatt ttgtaattct gattccccca aatcaagtat 120
caagtttacc tgcacagaca agggaagaag tggcaaaatg tgcaaatgag agcaacttta 180
tttgactgtc agtacgttga aattcagtgt ttccttaatc agttatggat tgacatttat 240
gtgcacagaa cctggaagaa tttcagccaa gctggaggta aaaatccaaa attctgatga 300
taaaaccaaa agtaaatcac aggtaaatct tctttatttt tcttttttaa tactgtatat 360
ggacattttt taatacagca tatttttttt ttgaaattta gaaaaaaacc actaagaaat 420
attcaccaat ggaatagact ttaaagtcac ttagagaatg tgtgctgttc tacgtagcat 480
actgacaatc attttcattt tggagttctt cattggaaat ctggggaatg gattcatagc 540
tctggtacaa tgcatggact tacgaaagag aagaacgttc ccttcagcag atcatttcct 600
cactgctctg gccatctcca ggcttgctct gatatgggtt ttatttctag attcatttct 660
gtttatacaa tccccattac tgatgactag aaatacatta agactgattc agactgcctg 720
gaatataagc aatcatttca gtatatggtt tgctaccagc ctcagcatct tttatctctt 780
caagatagcc attttttcta actatctttt cttctacctg aagcggagag ttaaaagggt 840
ggttttggtg atactgctgc tatccatgat ccttttgttt tttaatatat ttttagaaat 900
caaacatatt gatgtctgga tctatggaac caaaagaaac ataactaatg gtttgagttc 960
aaacagtttt tcagagtttt ccaggcttat tttaattcca agtttaatgt tcacattagt 1020
accctttggt gtatccttga tagctttcct cctcctaatc ttttccctta tgaaacatgt 1080
aaggaagatg cagtactaca ccaaaggatg caaagatgtc agaaccatgg cccacaccac 1140
agccctgcag actgtggttg ccttcctcct attatatact actttctttc tgtctctagt 1200
tgtggaagtt tcaacacttg aaatggatga aagtctgatg cttctgtttg caaaagttac 1260
tataatgatt tttccttcca tccactcctg tattttcatt ttgaaacata ataagttgag 1320
acaggacttg ctttcagtac tgaagtggct acagtattgg tgcaagcgtg agaaaacctt 1380
ggattcatag accattgtat gcatcacctt gaatattcta gaggggtgta ggttcatatg 1440
aaagtattga atttttaaat ttgagccttt tgtatatttt ct 1482
<210>135
<211>305
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R16,mGR16
<400>135
Met Asn Gly Val Leu Gln Val Thr Phe Ile Val Ile Leu Ser Val Glu
1 5 10 15
Phe Ile Ile Gly Ile Phe Gly Asn Gly Phe Ile Ala Val Val Asn Ile
20 25 30
Lys Asp Leu Val Lys Gly Arg Lys Ile Ser Ser Val Asp Gln Ile Leu
35 40 45
Thr Ala Leu Ala Ile Ser Arg Ile Ala Leu Leu Trp Leu Ile Leu Val
50 55 60
Ser Trp Trp Ile Phe Val Leu Tyr Pro Gly Gln Trp Met Thr Asp Arg
65 70 75 80
Arg Val Ser Ile Met His Ser Ile Trp Thr Thr Phe Asn Gln Ser Ser
85 90 95
Leu Trp Phe Ala Thr Ser Leu Ser Ile Phe Tyr Phe Phe Lys Ile Ala
100 105 110
Asn Phe Ser Asn Pro Ile Phe Leu Tyr Leu Lys Val Arg Leu Lys Lys
115 120 125
Val Met Ile Gly Thr Leu Ile Met Ser Leu Ile Leu Phe Cys Leu Asn
130 135 140
Ile Ile Ile Met Asn Ala Pro Glu Asn Ile Leu Ile Thr Glu Tyr Asn
145 150 155 160
Val Ser Met Ser Tyr Ser Leu Ile Leu Asn Asn Thr Gln Leu Ser Met
165 170 175
Leu Phe Pro Phe Ala Asn Thr Met Phe Gly Phe Ile Pro Phe Ala Val
180 185 190
Ser Leu Val Thr Phe Val Leu Leu Val Phe Ser Leu Trp Lys His Gln
195 200 205
Arg Lys Met Gln His Ser Ala His Gly Cys Arg Asp Ala Ser Thr Lys
210 215 220
Ala His Ile Arg Ala Leu Gln Thr Leu Ile Ala Ser Leu Leu Leu Tyr
225 230 235 240
Ser Ile Phe Phe Leu Ser His Val Met Lys Val Trp Ser Ala Leu Leu
245 250 255
Leu Glu Arg Thr Leu Leu Leu Leu Ile Thr Gln Val Ala Arg Thr Ala
260 265 270
Phe Pro Ser Val His Ser Trp Val Leu Ile Leu Gly Asn Ala Lys Met
275 280 285
Arg Lys Ala Ser Leu Tyr Val Phe Leu Trp Leu Arg Cys Arg His Lys
290 295 300
Glu
305
<210>136
<211>1316
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R16,mGR16
<400>136
tttatgatgg aaagaataaa accattagca aggcttaatg gcttgtttgg tattagacct 60
gtacattgtt tatggaacat gatatggagc tttgtttatt gaatatgcac aatattttag 120
aagcatgttt caaagaatct taagtaatta caatagaaat tgaagcatcc aagtgaagat 180
gaatggtgtc ctacaggtta catttatagt cattttgagt gtggaattta taattggcat 240
ctttggcaat ggattcatag cggtggtgaa cataaaggac ttggtcaagg gaaggaagat 300
ctcttcagtg gatcagatcc tcactgctct ggccatctcc agaattgcac tgctgtggtt 360
aatattagta agttggtgga tatttgtgct ttacccagga caatggatga ctgatagaag 420
agttagcata atgcacagta tatggacaac attcaaccag agtagtctct ggtttgctac 480
aagtctcagc atcttttatt ttttcaagat agcaaatttt tccaacccta tttttcttta 540
tttaaaggtc agacttaaaa aagtcatgat agggacattg ataatgtctt tgattctctt 600
ttgtttaaat attatcatta tgaatgcacc tgagaacatt ttaatcactg aatataatgt 660
atctatgtct tacagcttga ttttgaataa cacacagctt tctatgctgt ttccatttgc 720
caacaccatg tttgggttca taccttttgc tgtgtcactg gtcacttttg tccttcttgt 780
tttctccctg tggaaacatc agagaaagat gcaacacagt gcccatggat gcagagatgc 840
cagcactaag gcccacatca gagccttgca gacattgatt gcctccctcc tcctgtattc 900
cattttcttc ctgtctcatg ttatgaaggt ttggagtgct ctgcttctgg agaggacact 960
cctgcttttg atcacacagg ttgcaagaac agcttttccg tcagtgcact cctgggtcct 1020
gattctgggc aatgctaaga tgagaaaggc ttctctctat gtattcctgt ggctgaggtg 1080
caggcacaaa gaatgaaacc ctacagtgta cagacctggg gtatatttat gtggatgatc 1140
ttacatatct tagaggaaaa tggattaaaa gaaattctca tatttataaa tttttaggtc 1200
tgaattacat aaaaatgtat ataatatttt caaagtacaa gatagtagtt tataacttac 1260
atgataaata ctgtctatgc atcttctagt ctttgtagaa tatgtaaaaa catgtt 1316
<210>137
<211>330
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R17,mGR17
<400>137
Met Lys His Phe Trp Lys Ile Leu Ser Val Ile Ser Gln Ser Thr Leu
1 5 10 15
Ser Val Ile Leu Ile Val Glu Leu Val Ile Gly Ile Ile Gly Asn Gly
20 25 30
Phe Met Val Leu Val His Cys Met Asp Trp Val Lys Lys Lys Lys Met
35 40 45
Ser Leu Val Asn Gln Ile Leu Thr Ala Leu Ser Ile Ser Arg Ile Phe
50 55 60
Gln Leu Cys Leu Leu Phe Ile Ser Leu Val Ile Asn Phe Ser Tyr Thr
65 70 75 80
Asp Leu Thr Thr Ser Ser Arg Met Ile Gln Val Met Tyr Asn Ala Trp
85 90 95
Ile Leu Ala Asn His Phe Ser Ile Trp Ile Ala Thr Cys Leu Thr Val
100 105 110
Leu Tyr Phe Leu Lys Ile Ala Asn Phe Ser Asn Ser Phe Phe Leu Tyr
115 120 125
Leu Lys Trp Arg Val Glu Lys Val Val Ser Val Thr Leu Leu Val Ser
130 135 140
Leu Leu Leu Leu Ile Leu Asn Ile Leu Leu Thr Asn Leu Glu Thr Asp
145 150 155 160
Met Trp Thr Asn Glu Tyr Gln Arg Asn Ile Ser Cys Ser Phe Ser Ser
165 170 175
His Tyr Tyr Ala Lys Cys His Arg Gln Val Leu Arg Leu His Ile Ile
180 185 190
Phe Leu Ser Val Pro Val Val Leu Ser Leu Ser Thr Phe Leu Leu Leu
195 200 205
Ile Phe Ser Leu Trp Thr His His Lys Arg Met Gln Gln His Val Gln
210 215 220
Gly Gly Arg Asp Ala Arg Thr Thr Ala His Phe Lys Ala Leu Gln Thr
225 230 235 240
Val Ile Ala Phe Phe Leu Leu Tyr Ser Ile Phe Ile Leu Ser Val Leu
245 250 255
Ile Gln Ile Trp Lys Tyr Glu Leu Leu Lys Lys Asn Leu Phe Val Val
260 265 270
Phe Cys Glu Val Val Tyr Ile Ala Phe Pro Thr Phe His Ser Tyr Ile
275 280 285
Leu Ile Val Gly Asp Met Lys Leu Arg Gln Ala Cys Leu Pro Leu Cys
290 295 300
Ile Ile Ala Ala Glu Ile Gln Thr Thr Leu Cys Arg Asn Phe Arg Ser
305 310 315 320
Leu Lys Tyr Phe Arg Leu Cys Cys Ile Phe
325 330
<210>138
<211>1354
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R17,mGR17
<400>138
gaattctggt ctggcacccc tgagctgtgt gagtagacac attatcatgg aaagagattc 60
agaatctgtc actgtcaaaa ctgcatgttt gctcctctgt tagtgtgttg gggaaagtta 120
agaaaaatac attttatgag aatcaactca gaggttgtca gaaattgtcg aaacagcatt 180
ttaaaaattt acatctcaac tggatatatg agcaagtctt tataactgat atataaaatg 240
aagcactttt ggaagatatt atctgttatc tcccagagca cactttcagt cattttaatc 300
gtggaattag taattggaat tataggaaat gggttcatgg tcctggtcca ctgtatggac 360
tgggttaaga aaaagaaaat gtccctagtt aatcaaattc ttactgcttt gtcaatctcc 420
agaatttttc agctctgttt attgtttata agtttagtaa tcaacttttc atatacagat 480
ttaactacaa gttcaaggat gatacaagtc atgtacaatg cttggatttt agccaaccat 540
ttcagcatct ggattgctac atgcctcact gtcctttatt ttctaaagat agccaatttt 600
tctaactctt tttttcttta tctaaagtgg agagttgaaa aagtagtttc agttacactg 660
ttggtgtcat tgctcctcct gattttaaat attttactaa ctaacttgga aaccgacatg 720
tggacaaatg aatatcaaag aaacatatca tgcagcttca gttctcatta ctatgcaaag 780
tgtcacaggc aggtgttaag gcttcacatt attttcctgt ctgtccccgt tgttttgtcc 840
ctgtcaactt ttctcctgct catcttctcc ctgtggacac atcacaagag gatgcagcag 900
catgttcagg gaggcagaga tgccagaacc acggcccact tcaaagccct acaaactgtg 960
attgcatttt tcctactata ttccattttt attctgtctg tcttaataca aatttggaaa 1020
tatgaattac tgaagaaaaa tcttttcgtt gtattttgtg aggttgtata tatagctttt 1080
ccgacattcc attcatatat tctgattgta ggagacatga agctgagaca ggcctgcctg 1140
cctctctgta ttatcgcagc tgaaattcag actacactat gtagaaattt tagatcacta 1200
aagtacttta gattatgttg tatattctag acaaaaatta actgatacaa atgtcttttg 1260
tatttttcat tttaaatatc ctttaatttt gactgcatga aattgatttc tgcttgcaat 1320
tatcactgat taaaactatt aataatttaa ctag 1354
<210>139
<211>299
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R18,mGR18
<400>139
Met Val Pro Thr Gln Val Thr Ile Phe Ser Ile Ile Met Tyr Val Leu
1 5 10 15
Glu Ser Leu Val Ile Ile Val Gln Ser Cys Thr Thr Val Ala Val Leu
20 25 30
Phe Arg Glu Trp Met His Phe Gln Arg Leu Ser Pro Val Glu Thr Ile
35 40 45
Leu Ile Ser Leu Gly Ile Ser His Phe Cys Leu Gln Trp Thr Ser Met
50 55 60
Leu Tyr Asn Phe Gly Thr Tyr Ser Arg Pro Val Leu Leu Phe Trp Lys
65 70 75 80
Val Ser Val Val Trp Glu Phe Met Asn Ile Leu Thr Phe Trp Leu Thr
85 90 95
Ser Trp Leu Ala Val Leu Tyr Cys Val Lys Val Ser Ser Phe Thr His
100 105 110
Pro Ile Phe Leu Trp Leu Arg Met Lys Ile Leu Lys Leu Val Leu Trp
115 120 125
Leu Ile Leu Gly Ala Leu Ile Ala Ser Cys Leu Ser Ile Ile Pro Ser
130 135 140
Val Val Lys Tyr His Ile Gln Met Glu Leu Val Thr Leu Asp Asn Leu
145 150 155 160
Pro Lys Asn Asn Ser Leu Ile Leu Arg Leu Gln Gln Phe Glu Trp Tyr
165 170 175
Phe Ser Asn Pro Leu Lys Met Ile Gly Phe Gly Ile Pro Phe Phe Val
180 185 190
Phe Leu Ala Ser Ile Ile Leu Leu Thr Val Ser Leu Val Gln His Trp
195 200 205
Val Gln Met Lys His Tyr Ser Ser Ser Asn Ser Ser Leu Lys Ala Gln
210 215 220
Phe Thr Val Leu Lys Ser Leu Ala Thr Phe Phe Thr Phe Phe Thr Ser
225 230 235 240
Tyr Phe Leu Thr Ile Val Ile Ser Phe Ile Gly Thr Val Phe Asp Lys
245 250 255
Lys Ser Trp Phe Trp Val Cys Glu Ala Val Ile Tyr Gly Leu Val Cys
260 265 270
Ile His Phe Thr Ser Leu Met Met Ser Asn Pro Ala Leu Lys Lys Ala
275 280 285
Leu Lys Leu Gln Phe Trp Ser Pro Glu Pro Ser
290 295
<210>140
<211>2887
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R18,mGR18
<220>
<221>modified_base
<222>(1083)
<223>n=g,a,c或t
<400>140
gcgtgcttca cagagcagta tactacaaag caaatgtcat tgctgccatt gtatatttct 60
ctaaagacat ttcacatttt atctccctgt cccattgtgt gcagagccca cacttcaatc 120
aatcaattcc ttaattataa gctattgttt cattatttca tttcctacgt ttttttgcat 180
ttttactaaa actccaaagc agacattttc taattataat cctacatgta gttagaattt 240
taaaaattat atactatttt ctttgcacca ctgagttcag taggttttga aggtttatgc 300
ttaacaattg aacatttcat gttagattat tcctgccttc ctaatcttga ataattaaat 360
gtccatccag gcttagaatt cacagagtca acagctttca ccttgattct ctcactatct 420
atcaatgact agaatctgtc tgtcactttt gaaaccgcta attaaatagt tggtgcttat 480
ttaaagggtg ccccatgcca agagaaaatg tatttcttct ctagatgcct tcgtccttta 540
caagttacat gctttactga tggtgaattg gttttcttcc agttcatctg ggttaagtga 600
cctaagaacc tagccatgga aggagaaaca gaagcaaata ttaacgatac aagaacaagt 660
tccagaacat tggaaagtac ttagtaaagg cattggaatt agcaaaagaa tagtagcgaa 720
gcaaaaaata cttcatctcc attgggaggt caagaaagac tatgcagtgt ttttgatgca 780
acttgtcatc tctgagttag acgattcagc acacactttt gagattgaac ttcaacaggt 840
ggagccagca gacctgagct ttaggaatga tggtggaatt tccaagcaaa gacttccgtt 900
acctttttga tgtcccctaa caattcggtt gcaatgctca caccgcccaa ctgttgaaat 960
gcttgggaaa agggattctg agactggcat tagtatgtca tttgacagaa tggaaacatt 1020
gcccagggca ttaatgcaca gtaaaggatt caccttttct aagtgctcaa attttaaatt 1080
tgnatatttt tagaagacat tatttaaaag aaaggtggag aggatatcca aacagcacct 1140
tgagcagata aagaggtgaa gaagaaaaaa caacatgcgt acatgatgga tttctcttta 1200
tgaaaatgat caaatgatct taggatcaag aatccacacc tgaatgagat ttgcttgtat 1260
ccctgtgtga atttgaccta acaagcaaag cacagacaaa tgctgtagat agggaaatgt 1320
ctatgtcaaa tgtgtgtaag gaggatttgc atccacaaag aagtgccctc ttatactgag 1380
agtgctaaga acacatgtcc gtttcatatt cggaaagtgg tatagagctg ttgagtcttt 1440
ggctaggaag agacttcaga gtggaagcat ggtgccaacg caagtcacca tcttctccat 1500
catcatgtat gtgcttgagt ccttagtaat aattgtgcaa agttgcacaa cggttgcagt 1560
gctattcaga gagtggatgc actttcaaag actgtcaccg gtggagacga ttctcatcag 1620
cctgggcatc tcacatttct gtctacagtg gacatcaatg ctatacaact ttggtactta 1680
ttctaggcct gtccttttat tttggaaggt atcagtcgtc tgggagttca tgaacatttt 1740
gacattctgg ttaaccagtt ggcttgctgt cctctactgt gtcaaggtct cttccttcac 1800
tcaccccatc ttcctctggc tgaggatgaa aatcttgaaa ctggttctct ggttgatact 1860
gggtgctctg atagcttctt gtttgtcaat catcccttct gttgttaaat atcacatcca 1920
gatggaatta gtcaccctag ataatttacc caagaacaat tctttgattc taagactaca 1980
acagtttgaa tggtattttt ctaatccttt aaaaatgatt ggctttggta ttcctttctt 2040
cgtgttcctg gcttctatca tcttactcac agtctcattg gtccaacact gggtgcagat 2100
gaaacactac agcagcagca actccagcct gaaagctcag ttcactgttc tgaagtctct 2160
tgctaccttc ttcaccttct tcacatccta ttttctgact atagtcatct cctttattgg 2220
cactgtgttt gataagaaat cttggttctg ggtctgcgaa gctgtcatct atggtttagt 2280
ctgtattcac ttcacttcac tgatgatgag caaccctgca ttgaaaaagg cactgaagct 2340
gcagttctgg agcccagagc cttcctgagg caggaaacac agttaagcct ctagggtaag 2400
gagactttgc attggcacag tccctatagt gtaatgcaaa cttgaacaca aacttcatcc 2460
cttttcacat ccacaaatgg ctgcatctat acatcatcac cagtcttccc tgtattctga 2520
cccattctct tcctgtccta tccatagtcc ccaggttggt tttgattttt ctcatgatca 2580
caccaactct gcttagcttt tgccaccact gtaatagtaa acatggggtg ttctatatat 2640
tacagtcaaa atcattctca cattgttgat tgcctcacaa attcatataa atcccccttc 2700
ctgtcaggaa tttattgtct gctcacttaa tgctcaccat atattaaagc cattaattcc 2760
cccttcctac cttgagttta agaaggaaaa tgtcttacca ttgcccacaa cctattctgc 2820
tgcttctaga cttttatgca agtgatttat acacacacac acacacacac acacacatac 2880
aaacaac 2887
<210>141
<211>335
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R19,mGR19
<400>141
Met Met Glu Gly His Met Leu Phe Phe Leu Leu Val Val Val Val Gln
1 5 10 15
Phe Leu Thr Gly Val Leu Ala Asn Gly Leu Ile Val Val Val Asn Ala
20 25 30
Ile Asp Leu Ile Met Trp Lys Lys Met Ala Pro Leu Asp Leu Leu Leu
35 40 45
Phe Cys Leu Ala Thr Ser Arg Ile Ile Leu Gln Leu Cys Ile Leu Phe
50 55 60
Ala Gln Leu Gly Leu Ser Cys Leu Val Arg His Thr Leu Phe Ala Asp
65 70 75 80
Asn Val Thr Phe Val Tyr Ile Ile Asn Glu Leu Ser Leu Trp Phe Ala
85 90 95
Thr Trp Leu Gly Val Phe Tyr Cys Ala Lys Ile Ala Thr Ile Pro His
100 105 110
Pro Leu Phe Leu Trp Leu Lys Met Arg Ile Ser Arg Leu Val Pro Trp
115 120 125
Leu Ile Leu Ala Ser Val Val Tyr Val Thr Val Thr Thr Phe Ile His
130 135 140
Ser Arg Glu Thr Ser Glu Leu Pro Lys Gln Ile Phe Ile Ser Phe Phe
145 150 155 160
Ser Lys Asn Thr Thr Arg Val Arg Pro Ala His Ala Thr Leu Leu Ser
165 170 175
Val Phe Val Phe Gly Leu Thr Leu Pro Phe Leu Ile Phe Thr Val Ala
180 185 190
Val Leu Leu Leu Leu Ser Ser Leu Trp Asn His Ser Arg Gln Met Arg
195 200 205
Thr Met Val Gly Thr Arg Glu Pro Ser Arg His Ala Leu Val Ser Ala
210 215 220
Met Leu Ser Ile Leu Ser Phe Leu Ile Leu Tyr Leu Ser His Asp Met
225 230 235 240
Val Ala Val Leu Ile Cys Thr Gln Gly Leu His Phe Gly Ser Arg Thr
245 250 255
Phe Ala Phe Cys Leu Leu Val Ile Gly Met Tyr Pro Ser Leu His Ser
260 265 270
Ile Val Leu Ile Leu Gly Asn Pro Lys Leu Lys Arg Asn Ala Lys Thr
275 280 285
Phe Ile Val His Cys Lys Cys Cys His Cys Ala Arg Ala Trp Val Thr
290 295 300
Ser Arg Asn Pro Arg Leu Ser Asp Leu Pro Val Pro Ala Thr His His
305 310 315 320
Ser Ala Asn Lys Thr Ser Cys Ser Glu Ala Cys Ile Met Pro Ser
325 330 335
<210>142
<211>1698
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R19,mGR19
<400>142
ctgcagccta gagaactaat gcataggaaa cttatattcc cacctccgtg acgtcactct 60
gacagaagtg aacttatatt cccacctccg tgacgtcact ctgacagaag tgacttgttt 120
ttgtatgatg ctccaggatg cctcattagc attgaggaca atcataatta agtaaggcaa 180
ggcatgaagg tggtcctcac taggtacctg gaggcttctg gttgcatgat ttacttgtga 240
tgactctgac acttaagaag acctgaaaaa tgcaaaagct gtcataaggc acagttcgtt 300
tctatggtat ctcttcctta tttgactgac attgagttga gaaggcagca ctataaacaa 360
atgggcccca ccttcctctt ccattgtctt tgggttggca tcatctccaa aggaaccttg 420
gtctagttga aagaagccag aaatcataca tggctgagac tgtgcataac tctatgtatc 480
atttaaagaa gtcattggtt cttcttattt taaaatgatg gaaggtcata tgctcttctt 540
ccttctggtc gtggtagtgc agtttttaac tggggtcttg gcaaatggcc tcattgtggt 600
tgtcaatgcc atcgacttga tcatgtggaa gaaaatggcc ccactggatc tgcttctttt 660
ttgcctggcg acttctcgga tcattcttca attgtgtata ttgtttgcac agctgggtct 720
atcctgtttg gtgagacaca cgttatttgc tgacaatgtt acctttgtct acattataaa 780
cgaactgagt ctctggtttg ccacatggct tggtgttttc tactgtgcca agattgctac 840
catccctcac ccactctttc tgtggctgaa gatgaggata tccaggttgg tgccatggct 900
gatcctggca tctgtggtct atgtaactgt tactactttc atccatagca gagagacttc 960
agaacttcct aagcaaatct ttataagctt tttttctaaa aatacaactc gggtcagacc 1020
agcgcatgcc acactactct cagtctttgt ctttgggctc acactaccat ttctcatctt 1080
cactgttgct gttctgctct tgttgtcctc cctgtggaac cacagccggc agatgaggac 1140
tatggtggga actagggaac ctagcagaca tgccctcgtc agtgcgatgc tctccattct 1200
gtcattcctc atcctctatc tctcccatga catggtagct gttctgatct gtacccaagg 1260
cctccacttt ggaagcagaa cctttgcatt ctgcttattg gttattggta tgtacccctc 1320
cttacactcg attgtcttaa ttttaggaaa ccctaagctg aaacgaaatg caaaaacgtt 1380
cattgtccat tgtaagtgtt gtcattgtgc aagagcttgg gtcacctcaa ggaacccaag 1440
actcagcgac ttgccagtgc ctgctactca tcactcagcc aacaagacat cctgctcaga 1500
agcctgtata atgccatctt aattgtccaa cctgaggctt aatcatttca aagggtaaat 1560
tgatgatcaa agcccaacac atgatatgac atcaaggtcc atatcccagt agtcatgtgg 1620
aaataccacc ttgcaaaatg atgtcattga gaaaccaggg caaatggagt ctaggtcttt 1680
cagtatgatt tgctgcag 1698
<210>143
<211>295
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R20,mGR20
<400>143
Met Asn Leu Val Glu Trp Ile Val Thr Ile Ile Met Met Thr Glu Phe
1 5 10 15
Leu Leu Gly Asn Cys Ala Asn Val Phe Ile Thr Ile Val Asn Phe Ile
20 25 30
Asp Cys Val Lys Arg Arg Lys Ile Ser Ser Ala Asp Arg Ile Ile Thr
35 40 45
Ala Ile Ala Ile Phe Arg Ile Gly Leu Leu Trp Ala Met Leu Thr Asn
50 55 60
Trp His Ser His Val Phe Thr Pro Asp Thr Asp Asn Leu Gln Met Arg
65 70 75 80
Val Phe Gly Gly Ile Thr Trp Ala Ile Thr Asn His Phe Thr Thr Trp
85 90 95
Leu Gly Thr Ile Leu Ser Met Phe Tyr Leu Phe Lys Ile Ala Asn Phe
100 105 110
Ser Asn Ser Leu Phe Leu His Leu Lys Arg Lys Leu Asp Asn Val Leu
115 120 125
Leu Val Ile Phe Leu Gly Ser Ser Leu Phe Leu Val Ala Tyr Leu Gly
130 135 140
Met Val Asn Ile Lys Lys Ile Ala Trp Met Ser Ile His Glu Gly Asn
145 150 155 160
Val Thr Thr Lys Ser Lys Leu Lys His Val Thr Ser Ile Thr Asn Met
165 170 175
Leu Leu Phe Ser Leu Ile Asn Ile Val Pro Phe Gly Ile Ser Leu Asn
180 185 190
Cys Val Leu Leu Leu Ile Tyr Ser Leu Ser Lys His Leu Lys Asn Met
195 200 205
Lys Phe Tyr Gly Lys Gly Cys Gln Asp Gln Ser Thr Met Val His Ile
2l0 215 220
Lys Ala Leu Gln Thr Val Val Ser Phe Leu Leu Leu Tyr Ala Thr Tyr
225 230 235 240
Ser Ser Cys Val Ile Ile Ser Gly Trp Ser Leu Gln Asn Ala Pro Val
245 250 255
Phe Leu Phe Cys Val Thr Ile Gly Ser Phe Tyr Pro Ala Gly His Ser
260 265 270
Cys Ile Leu Ile Trp Gly Asn Gln Lys Leu Lys Gln Val Phe Leu Leu
275 280 285
Leu Leu Arg Gln Met Arg Cys
290 295
<210>144
<211>1394
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R20,mGR20
<400>144
ctagatgggc tgtttcatat aatgactgga actccctaca tgctccacgt cttgagttct 60
aaaatttcac taacaaattt ttgactgcca taaataatga aggtttaaag aaagaacaac 120
atttgaagca atggaccaga attcctcttt atttgactct tagcaaattg gaatgcagca 180
tcctttcaag agcagcactg aaatatacca gtcaatggca gagagtaaaa aagtatgcaa 240
ttggagacat tatggtaata taaatttcca ttaaaaatga gactgcattc acctattaca 300
acacattgct attctgctca acacagagtt aaaaagaaac aagaactctt gtatacattc 360
agttagtcac aagtataatt atgttcacat attttaaaaa aatgaatcat gatctgtgaa 420
ttgagcctgg ctttttttgt ctctctcttt ttattctttt cctttagaca gacacaatga 480
atttggtaga atggattgtt accatcataa tgatgacaga atttctctta ggaaactgtg 540
ccaatgtctt cataaccata gtgaacttca tcgactgtgt gaagagaaga aagatctcct 600
cagctgatcg aattataact gctattgcca tcttcagaat tggtttgttg tgggcaatgt 660
taacgaactg gcattcacat gtgtttactc cagacacaga caatttacaa atgagagttt 720
tcggtggaat tacctgggct ataaccaacc attttaccac ttggctgggg accatactga 780
gcatgtttta tttattcaag atagccaatt tttccaacag tctatttctt catctaaaaa 840
gaaaacttga caatgttcta cttgtgattt tcctgggatc gtctctgttt ttggttgcat 900
atcttgggat ggtgaacatc aagaagattg cttggatgag tattcatgaa ggaaatgtga 960
ccacaaagag caaactgaag catgtaacaa gcatcacaaa tatgcttctc ttcagcctga 1020
taaacattgt accatttggt atatcactga actgtgttct gctcttaatc tattccctga 1080
gtaaacatct caagaatatg aaattctatg gcaaaggatg tcaagatcag agcaccatgg 1140
tccacataaa ggccttgcaa actgtggtct cttttctctt gttatatgcc acatactctt 1200
cctgtgtcat tatatcaggt tggagtttgc aaaatgcacc agtcttcctg ttttgtgtga 1260
caattggatc cttctaccca gcaggtcatt cttgtatctt gatttgggga aaccagaaac 1320
ttaaacaggt ctttctgttg ttgctgaggc agatgagatg ctgactgaaa aaatgaaagt 1380
ccccctgtct ctag 1394
<210>145
<211>305
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R21,mGR21
<400>145
Met Gly Set Asn Val Tyr Gly Ile Leu Thr Met Val Met Ile Ala Glu
1 5 10 15
Phe Val Phe Gly Asn Met Ser Asn Gly Phe Ile Val Leu Ile Asn Cys
20 25 30
Ile Asp Trp Val Arg Lys Gly Thr Leu Ser Ser Ile Gly Trp Ile Leu
35 40 45
Leu Phe Leu Ala Ile Ser Arg MetVal Leu Ile Trp Glu Met Leu Ile
50 55 60
Thr Trp Ile Lys Tyr Met Lys Tyr Ser Phe Ser Phe Val Thr Gly Thr
65 70 75 80
Glu Leu Arg Gly Ile Met Phe Thr Trp Val Ile Ser Asn His Phe Ser
85 90 95
Leu Trp Leu Ala Thr Ile Leu Ser Ile Phe Tyr Leu Leu Lys Ile Ala
100 105 110
Ser Phe Ser Lys Pro Val Phe Leu Tyr Leu Lys Trp Arg Glu Lys Lys
115 120 125
Val Leu Leu Ile Val Leu Leu Gly Asn Leu Ile Phe Leu Met Leu Asn
130 135 140
Ile Leu Gln Ile Asn Lys His Ile Glu His Trp Met Tyr Gln Tyr Glu
145 150 155 160
Arg Asn Ile Thr Trp Ser Ser Arg Val Ser Asp Phe Ala Gly Phe Ser
165 170 175
Asn Leu Val Leu Leu Glu Met Ile Val Phe Ser Val Thr Pro Phe Thr
180 185 190
Val Ala Leu Val Ser Phe Ile Leu Leu Ile Phe Ser Leu Trp Lys His
195 200 205
Leu Gln Lys Met His Leu Asn Ser Arg Gly Glu Arg Asp Pro Ser Thr
210 215 220
Lys Ala His Val Asn Ala Leu Arg Ile Met Val Ser Phe Leu Leu Leu
225 230 235 240
Tyr Ala Thr Tyr Phe Ile Ser Phe Phe Leu Ser Leu Ile Pro Met Ala
245 250 255
His Lys Thr Arg Leu Gly Leu Met Phe Ser Ile Thr Val Gly Leu Phe
260 265 270
Tyr Pro Ser Ser His Ser Phe Ile Leu Ile Leu Gly His Ser Asn Leu
275 280 285
Arg Gln Ala Ser Leu Trp Val Met Thr Tyr Leu Lys Cys Gly Gln Lys
290 295 300
His
305
<210>146
<211>2567
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R21,mGR21
<400>146
ctcttttgaa gacaatagtt gttctactag ctattgatag catgtttaca tttgtcattt 60
tcaagtatgt tcagaaacaa agctacatat tgtggggagt atataaaata tgaaagcatg 120
ccattcccag gcatccaagg atccctgtgt attaaaaggc aacaaagcag aaccaaatgt 180
tctgttttgg acatgagctt cttccaattc aactgctgaa aaatttggat aactacatat 240
aaaactaaga acacagagtg tcacagagca gtctctgctc tccaattcac caggattaat 300
attgacagac ccaaaagatg tcatttaggt aaattttgga tgaatcatat tgttgtcacc 360
tttgtgctct agaacataag ctgatagaat caaattttct ttagcagaga caatgcaaat 420
tgatataaca gtgaaagaga atatatcttt atttgcatgt tagcaaatga cagctggatg 480
cacttcatga ttttctgcaa tctagttcag tctttagaag gatatatata tatatatata 540
tatatatata tatatatata tatatatata tataaacctt agtcttgaaa gatatcagaa 600
agaaggattt cacaagaatg tacagagcca ttagcaaaat tttaatatac tcatcgacat 660
taggtcagtc actacataag aaggacttga atgaaagctt atcttagttt ttgagactac 720
agggacattt caccttgcca aatgagaagc agtgagtctt ctttgtctgg acatgggaag 780
caatgtgtat ggtatcttaa ctatggttat gattgcagag tttgtatttg gaaatatgag 840
caatggattc atagtgctga taaactgcat tgattgggtc aggaaaggaa ctctttcttc 900
cattggttgg atcctgcttt tcttggccat ttcaagaatg gtgttgatat gggaaatgtt 960
aataacatgg ataaaatata tgaagtattc attttcattt gtgactggaa cagaattacg 1020
gggtatcatg tttacctggg taatttccaa tcacttcagt ctctggcttg ccactattct 1080
cagcatcttt tatttgctca aaatagccag tttctccaaa ccggtttttc tctatttgaa 1140
gtggagagag aagaaagtgc ttctgattgt ccttctggga aatttgatct tcttgatgct 1200
caacatatta caaataaaca aacatataga acactggatg tatcaatatg agagaaatat 1260
aacttggagt tctagagtga gtgactttgc agggttttca aatctggtct tattggagat 1320
gattgtgttc tctgtaacac cattcacagt ggccctggtc tccttcatcc tgttaatctt 1380
ctccttgtgg aaacatctac agaaaatgca tctcaattct agaggggaac gagaccccag 1440
cactaaagcc catgtgaatg ccttgagaat tatggtctcc ttcctcttac tctatgccac 1500
ttacttcata tctttttttc tatcattgat tcccatggca cataaaacac gactgggtct 1560
tatgtttagc ataactgttg ggcttttcta cccttcaagc cactcattta tcttaatttt 1620
gggacattct aatttaaggc aagccagtct ttgggtgatg acatatctta aatgtgggca 1680
aaagcattag aatttcacta ttccataagg cagccaaacc acgtgctact aggtatatga 1740
tactactcag tggtaaagcc ctaggcaaac attaacctta gaaaatatat aattttgtga 1800
ctcttctgta tttgataaat cactcacata tttagaagaa tgctacagta gtgtgatctt 1860
gtacatgatt gtaacaattc aattttatta atatagttca ggcatgataa catacccctg 1920
ataactgaaa agtaagtagg atgctacata tatatttaga tctagactta ggggcaaaga 1980
gagacccagc tgatagctgt gcaataaaga ttttaatttt catcctgttg tgagttatct 2040
gaaatctatg tcactgaagg cataagcaag attttcacac actgaaacaa tctcttatgc 2100
tttcttatat tgttttaaaa gtaaattaga aaatttaaat aaacttaatg gcaattgaaa 2160
ttacaaaagc taaacacatg tggttattag aaattagact gtatgtaggt cctaggggat 2220
ggcttagtaa agtgctttgt tgcaagcttc aggatatgat tctaaatccc tagattcaat 2280
taaaaacctg gcataaatag ccaatgtaaa atttgtctgt aaaatgtaac cagtgctaag 2340
agtaccaaga caacaaaatg tttactttta aaaccattta ttgatattct tttaaaaata 2400
ggtatgtatt ttactattta aataagattt tgtcaaaagc tagtcttgac accttaggta 2460
aacataggaa ggcaacaagt ttgaagtcag ctactgggga cagtgctgct agcagctgac 2520
agaggccact gctgactaca gcagatcatt tacaggttca gcactag 2567
<210>147
<211>309
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R22,mGR22
<400>147
Met Ser Ser Leu Leu Glu Ile Phe Phe Val Ile Ile Ser Val Val Glu
1 5 10 15
Phe Ile Ile Gly Thr Leu Gly Asn Gly Phe Ile Val Leu Ile Asn Ser
20 25 30
Thr Ser Trp Phe Lys Asn Gln Lys Ile Ser Val Ile Asp Phe Ile Leu
35 40 45
Thr Trp Leu Ala Ile Ser Arg Met Cys Val Leu Trp Thr Thr Ile Ala
50 55 60
Gly Ala Ser Leu Arg Lys Phe Tyr Lys Thr Leu Ser Tyr Ser Lys Asn
65 70 75 80
Phe Lys Phe Cys Phe Asp Ile Ile Trp Thr Gly Ser Asn Tyr Leu Cys
85 90 95
Ile Ala Cys Thr Thr Cys Ile Ser Val Phe Tyr Leu Phe Lys Ile Ala
100 105 110
Asn Phe Ser Asn Ser Ile Phe Phe Trp Ile Lys Gln Arg Ile His Ala
115 120 125
Val Leu Leu Ala Ile Val Leu Gly Thr Leu Met Tyr Phe Ile Leu Phe
130 135 140
Leu Ile Phe Met Lys Met Ile Ala Asn Asn Phe Ile Tyr Lys Trp Thr
145 150 155 160
Lys Leu Glu Gln Asn Thr Thr Phe Pro Val Leu Asp Thr Leu Ser Gly
165 170 175
Phe Leu Val Tyr His Ser Leu Tyr Asn Gly Ile Leu Ile Phe Phe Phe
180 185 190
Ile Val Ser Leu Thr Ser Phe Leu Leu Leu Ile Phe Ser Leu Trp Ser
195 200 205
His Leu Arg Arg Met Lys Leu Gln Gly Ile His Thr Lys Asp Ile Ser
210 215 220
Thr Glu Ala His Ile Lys Ala Met Lys Thr Met Met Ser Phe Leu Leu
225 230 235 240
Phe Phe Ile Ile Tyr Tyr Ile Ser Asn Ile Met Leu Ile Val Ala Ser
245 250 255
Ser Ile Leu Asp Asn Val Val Ala Gln Ile Phe Ser Tyr Asn Leu Ile
260 265 270
Phe Leu Tyr Leu Ser Val His Pro Phe Leu Leu Val Leu Trp Asn Ser
275 280 285
Lys Leu Lys Trp Thr Phe Gln His Val Leu Arg Lys Leu Val Cys His
290 295 300
Cys Gly Gly Tyr Ser
305
<210>148
<211>1488
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R22,mGR22
<400>148
aaatgaataa tttcatgcaa aggataccat tagaatatga tcactattta aattttagca 60
aatacatatt caaataccag cacaatgttt caaatttaaa atataaacat tataaaaccc 120
agcagagaac aaaatgatag ccttgataat tgttggtttg ctcaagaaaa atgggtgtat 180
actttaacat ttaattggga actcagttga gagcatacat ttagggtttt acagaggtat 240
tcattgccca tttaagattt ggattcacac atctacatca atgtggctgt aatccatttt 300
cccatgatga aataaggtag agactgccta ttaaacgaca tgtcgagcct actggagatt 360
ttctttgtga tcatttcggt tgtagaattc ataataggaa ctttgggaaa tggatttatt 420
gtcctgataa acagtacttc ttggttcaag aatcagaaaa tctctgtaat tgatttcatt 480
cttacttggt tggccatctc cagaatgtgt gttctatgga caacaattgc tggtgcctct 540
ctcaggaaat tctacaagac gttaagttac tctaagaatt tcaaattttg ttttgacatt 600
atctggacag gatccaacta tttatgcata gcctgtacaa cgtgcatcag tgtcttctac 660
ttgttcaaga ttgccaactt ttctaattcc attttcttct ggattaaaca gagaattcat 720
gcagtacttc tggctattgt cctaggcaca ctcatgtatt tcattttatt tctcattttt 780
atgaaaatga tagctaataa ttttatctac aaatggacaa aattggaaca aaacacaaca 840
ttccctgttt tagatactct aagtggtttc ttagtctacc atagcctcta caatgggatt 900
ctcattttct tttttatagt gtctctgacc tcatttcttc ttttaatctt ctctttatgg 960
agccacctta ggaggatgaa actacagggc atacatacca aagacataag cacagaagca 1020
cacataaaag ctatgaaaac tatgatgtca ttccttttgt tcttcatcat atattatatt 1080
agcaacatta tgcttattgt ggcaagctcc attcttgaca atgtggttgc acaaattttc 1140
tcttataacc taatatttct gtatttatct gttcatcctt ttcttctggt tttatggaac 1200
agcaaattga aatggacatt ccagcatgta ttgagaaagc tggtgtgtca ttgtggaggt 1260
tattcttgat ttcagtaaat acactcaata taactgatgg atttctaagg taagaaaaat 1320
ggaacaagga ataaagagga gaaatatatt ccttttcaga tcatctgctc tgtcattctg 1380
tccttagcat gctattaaga attgttgact aaatccagtc atttttaaca tgaggaaagg 1440
atgtttcaat ccaacttaga gagggtacaa aatagtccta ggaggcag 1488
<210>149
<211>333
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R23,mGR23
<400>149
Met Phe Ser Gln Lys Ile Asn Tyr Ser His Leu Phe Thr Phe Ser Ile
1 5 10 15
Thr Leu Tyr Val Glu Ile Val Thr Gly Ile Leu Gly His Gly Phe Ile
20 25 30
Ala Leu Val Asn Ile Met Asp Trp Val Lys Arg Arg Arg Ile Ser Ser
35 40 45
Val Asp Gln Ile Leu Thr Ala Leu Ala Leu Thr Arg Phe Ile Tyr Val
50 55 60
Leu Ser Met Leu Ile Cys Ile Leu Leu Phe Met Leu Cys Pro His Leu
65 70 75 80
Pro Arg Arg Ser Glu Met Leu Ser Ala MetGly Ile Phe Trp Val Val
85 90 95
Asn Ser His Phe Ser Ile Trp Leu Thr Thr Cys Leu Gly Val Phe Tyr
100 105 110
Phe Leu Lys Ile Ala Asn Phe Ser Asn Ser Phe Phe Leu Tyr Leu Lys
115 120 125
Trp Arg Val Lys Lys Val Ile Leu Ile Ile Ile Leu Ala Ser Leu Ile
130 135 140
Phe Leu Thr Leu His Ile Leu Ser Leu Gly Ile Tyr Asp Gln Phe Ser
145 150 155 160
Ile Ala Ala Tyr Val Gly Asn Met Ser Tyr Ser Leu Thr Asp Leu Thr
165 170 175
Gln Phe Ser Ser Thr Phe Leu Phe Ser Asn Ser Ser Asn Val Phe Leu
180 185 190
Ile Thr Asn Ser Ser His Val Phe Leu Pro Ile Asn Ser Leu Phe Met
195 200 205
Leu Ile Pro Phe Thr Val Ser Leu Val Ala Phe Leu Met Leu Ile Phe
210 215 220
Ser Leu Trp Lys His His Lys Lys Met Gln Val Asn Ala Lys Gln Pro
225 230 235 240
Arg Asp Val Ser Thr Met Ala His Ile Lys Ala Leu Gln Thr Val Phe
245 250 255
Ser Phe Leu Leu Leu Tyr Ala Ile Tyr Leu Leu Phe Leu Ile Ile Gly
260 265 270
Ile Leu Asn Leu Gly Leu Met Glu Lys Ile Val Ile Leu Ile Phe Asp
275 280 285
His Ile Ser Gly Ala Val Phe Pro Ile Ser His Ser Phe Val Leu Ile
290 295 300
Leu Gly Asn Ser Lys Leu Arg Gln Ala Ser Leu Ser Val Leu Pro Cys
305 310 315 320
Leu Arg Cys Gln Ser Lys Asp Met Asp Thr Met Gly Leu
325 330
<210>150
<211>1442
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R23,mGR23
<400>150
aattttcagc aaccaatatg tagactgctt aaatgcatca gaaacattat aaattgaagc 60
atgttttcac agaaaataaa ctacagccat ttgtttactt tttcaatcac cttgtatgtg 120
gaaatagtaa cgggaatctt aggacatgga ttcatagcat tagtgaacat catggactgg 180
gtcaaaagaa gaaggatctc ttcagtggat cagattctca ctgctttggc ccttaccaga 240
ttcatttatg tcttgtctat gctgatttgc atattgttat tcatgctgtg cccacatttg 300
cctaggagat cagaaatgct ttcagcaatg ggtattttct gggtagtcaa cagccatttt 360
agcatctggc ttactacatg cctcggtgtc ttttattttc tcaagatagc caatttttct 420
aactcttttt ttctttatct aaagtggaga gttaaaaaag tgattttaat aataatcctg 480
gcatcactga ttttcttgac tttacacatt ttatctttag ggatatatga tcagttctca 540
attgctgctt atgtaggaaa tatgtcttat agtttgacag atttaacaca attttccagt 600
actttcttat tctccaactc atccaatgtt ttcttaatca ccaactcatc ccatgttttc 660
ttacccatca actccctgtt catgctcata cccttcacag tgtccctggt agcctttctc 720
atgctcatct tctcactgtg gaagcatcac aaaaagatgc aggtcaatgc caaacaacct 780
agagatgtca gtactatggc ccacattaaa gccttgcaaa ctgtgttctc cttcctgctg 840
ctgtatgcca tatacttact tttccttatc ataggaattt tgaaccttgg attgatggag 900
aaaatagtga tactgatatt tgaccacatt tctggagcag tttttcctat aagccactca 960
tttgtactga ttctgggaaa cagtaagctg agacaagcca gtctttctgt gttgccttgt 1020
ctaaggtgcc agtccaaaga tatggacacc atgggtctct agtaaattcc agagtacatt 1080
ttgtaaaaat cttgaggatg atcagttcat agaaaaaagt taccttatgg gggaaaataa 1140
aaagtggggc ttcaatcctg ggagtaataa tacacaggag ggtaggacag catgaaggag 1200
actagcacta tataagtggt ctcatacagg atatgggaaa ggaaagattt atgcaataaa 1260
gagggagatc atattggagg atgaggaggc attacatatg taaaatgact ataagaatgg 1320
aatcatgcta atctaaaaaa atctgtaatg catttcattc agactatata catatatgcc 1380
tatatatgga tatatgggga tatatattct atacatattt taaaagaacc tttcttatat 1440
ag 1442
<210>151
<211>309
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R24,mGR24
<400>151
Met Val Pro Val Leu His Ser Leu Ser Thr Ile Ile Leu Ile Ala Glu
1 5 10 15
Phe Val Trp Gly Asn Leu Ser Asn Gly Leu Ile Val Leu Lys Asn Cys
20 25 30
Ile Asp Trp Ile Asn Lys Lys Glu Leu Ser Thr Val Asp Gln Ile Leu
35 40 45
Ile Val Leu Ala Ile Ser Arg Ile Ser Leu Ile Trp Glu Thr Leu Ile
50 55 60
Ile Trp Val Lys Asp Gln Leu Ile Ser Ser Ile Thr Ile Glu Glu Leu
65 70 75 80
Lys Ile Ile Val Phe Ser Phe Ile Leu Ser Ser His Phe Ser Leu Trp
85 90 95
Leu Ala Thr Ala Leu Ser Ile Phe Tyr Leu Phe Arg Ile Pro Asn Cys
100 105 110
Tyr Trp Gln Ile Phe Leu Tyr Leu Lys Trp Arg Ile Lys Gln Leu Ile
115 120 125
Val His Met Leu Leu Gly Ser Leu Val Phe Leu Val Ala Asn Met Ile
130 135 140
Gln Ile Thr Ile Thr Leu Glu Glu Arg Phe Tyr Gln Tyr Gly Gly Asn
145 150 155 160
Thr Ser Val Asn Ser Met Glu Thr Glu Phe Ser Ile Leu Ile Glu Leu
165 170 175
Met Leu Phe Asn Met Thr Met Phe Ser Ile Ile Pro Phe Ser Leu Ala
180 185 190
Leu Ile Ser Phe Leu Leu Leu Ile Phe Ser Leu Trp Lys His Leu Gln
195 200 205
Lys Met Pro Leu Asn Ser Arg Gly Asp Arg Asp Pro Ser Ala Thr Ala
210 215 220
His Arg Asn Ala Leu Arg Ile Leu Val Ser Phe Leu Leu Leu Tyr Thr
225 230 235 240
Ile Tyr Phe Leu Ser Leu Leu Ile Ser Trp Val Ala Gln Lys Asn Gln
245 250 255
Ser Glu Leu Val His Ile Ile Cys Met Ile Thr Ser Leu Val Tyr Pro
260 265 270
Ser Phe His Ser Tyr Ile Leu Ile Leu Gly Asn Tyr Lys Leu Lys Gln
275 280 285
Thr Ser Leu Trp Val Met Arg Gln Leu GIy Cys Arg Met Lys Arg Gln
290 295 300
Asn Thr Pro Thr Thr
305
<210>152
<211>1465
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R24,mGR24
<400>152
caaagaggag aaatatttag ctacacagtg taccacatac aagccgttca atcagtataa 60
ggggagcagt catatagaat ttgggctttc tttcttttaa tatggtacct gttctgcaca 120
gtctctccac catcatacta attgcagagt ttgtttgggg aaatttgagc aatggtttga 180
tagtgttgaa gaactgcatt gactggatca ataaaaaaga gctctccaca gttgatcaaa 240
tactcattgt cttggcaatt tcaagaatta gtctcatctg ggaaacacta attatatggg 300
ttaaagatca actaatttca tctattacta ttgaagaatt aaaaataatt gtgttcagct 360
ttatactatc tagccacttc agtctctggc ttgctacagc tctcagcatc ttctatttat 420
tcagaatacc taattgctac tggcagatct ttctctactt gaaatggaga ataaagcaac 480
tgattgtcca catgcttctg ggaagcttgg tgttcttggt tgcaaatatg atacagataa 540
ccatcactct tgaagagagg ttctatcaat atggaggaaa tacaagtgta aattccatgg 600
agactgagtt ctcaattttg atagagctga tgttatttaa catgactatg ttctccatta 660
taccattttc attggcctta atttcttttc ttctgctaat cttctcttta tggaaacatc 720
tccagaagat gccactcaat tctagaggag atagagaccc tagtgctacg gcccacagaa 780
atgccttgag aattttggtc tccttcctct tgctctatac tatatatttc ctgtctcttc 840
ttatatcatg ggttgctcag aagaatcaaa gtgaactggt tcacattatt tgtatgataa 900
cttcactcgt gtatccttca ttccactcat atatcctgat tctgggaaat tataaattaa 960
agcagacctc tctttgggta atgaggcagc tgggatgtag gatgaaaaga cagaatacac 1020
caactacata aggcagccaa acagtctatt gggttttaga taacaaatct aaatctatga 1080
ggaagtagtt caataacatt tttccccttg acatggagta gcagggtttt tttttattag 1140
atattttctt tacttacatt tcaaatgcta tcccgaaaat tccctgtacc ctctccctgt 1200
cctgttcccc tacccaccca ctcccacttc ttggccctgg cattcccctg gagtatcagt 1260
tttttattag tcaaactatc tcactgacta agggtcataa aacaagttat tttaacacta 1320
atttcaatta aatcaaaggt aaagtgtcag cacatgcctt taatcacaca attccatcaa 1380
attcagcact caggagaggg tgatctctgt gaattccagc acactggcgg ccgttactag 1440
tggatccgag ctcggtacca agctt 1465
<210>153
<211>311
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R25,mGR25
<400>153
Met Met Gly Ile Ala Ile Asp Ile Leu Trp Ala Ala Ile Ile Ile Val
1 5 10 15
Gln Phe Ile Ile Gly Asn Ile Ala Asn Gly Phe Ile Ala Leu Val Asn
20 25 30
Ile Ile Asp Trp Val Lys Arg Arg Lys Ile Ser Leu Met Asp Lys Ile
35 40 45
Ile Thr Ala Leu Ala Ile Ser Arg Ile Tyr Leu Leu Trp Ser Thr Phe
50 55 60
Leu Ile Thr Leu Thr Ser Ser Leu Asp Pro Asp Ile Lys Met Ala Val
65 70 75 80
Lys Ile Ile Arg Ile Ser Asn Asn Thr Trp Ile Ile Ala Asn His Phe
85 90 95
Ser Ile Trp Phe Ala Thr Cys Leu Ser Ile Phe Tyr Phe Leu Lys Ile
100 105 110
Ala Asn Phe Ser Asn Tyr Ile Phe Leu Tyr Leu Arg Trp Arg Phe Lys
115 120 125
Lys Val Val Ser Val Thr Leu Leu Ile Ser Leu Ile Phe Leu Leu Leu
130 135 140
Asn Ile Leu Leu Met Asn Met His Ile Asp Ile Trp Ser Asp Lys Ser
145 150 155 160
Lys Arg Asn Leu Ser Phe Ser Val Arg Ser Asn Asn Cys Thr Gln Phe
165 170 175
Pro Arg Leu Val Leu Leu Ile Asn Thr Met Phe Thr Ser Ile Pro Phe
180 185 190
Thr Val Ser Leu Leu Ala Phe Leu Leu Leu Ile Phe Ser Leu Trp Arg
195 200 205
His Leu Lys Thr Met Gln Tyr Tyr Ala Lys Gly Ser Glu Asp Thr Thr
210 215 220
Thr Ala Ala His Ile Lys Ala Leu His Met Val Val Ala Phe Leu Leu
225 230 235 240
Phe Tyr Thr Val Phe Phe Leu Ser Leu Ala Ile Gln Tyr Trp Thr Ser
245 250 255
Gly Ser Gln Glu Asn Asn Asn Leu Phe Tyr Ala Thr Ile Val Ile Thr
260 265 270
Phe Pro Ser Val His Ser Cys Ile Leu Ile Leu Arg Asn Ser Gln Leu
275 280 285
Arg Gln Ala Ser Leu Leu Val Leu Trp Trp Leu Leu Cys Lys Ser Lys
290 295 300
Asp Val Arg Met Leu Val Pro
305 310
<210>154
<211>1103
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R25,mGR25
<400>154
aaaactattc gaattgaaca cagtaaccaa ttcttcagcg gacttacaca aatcaagcta 60
ttatcttatg gatgatgggt attgccatag atatcttatg ggcagctatt atcattgtgc 120
aattcataat tgggaatatt gcaaatggat tcatagcatt ggtgaacatc atagactggg 180
tgaagagaag aaaaatctct ttaatggata agatcattac tgctttggca atctctagga 240
tttatctgct gtggtctaca ttcttaatta cactaacatc ttcactggat ccagatatta 300
aaatggctgt gaaaatcatt agaataagca ataacacctg gattattgca aatcatttca 360
gcatttggtt tgctacatgt ctcagcatct tttattttct caagatagcc aatttttcta 420
actatatttt tctctactta aggtggagat ttaagaaggt ggtttcagtg acattgctaa 480
tctctcttat cttcctgctt ttaaatattt tactgatgaa catgcatatt gatatctgga 540
gtgataagtc caaaagaaac ctttctttta gtgtcagatc aaataattgc actcagtttc 600
ccagacttgt ccttttaatc aacacaatgt tcacatcaat ccccttcact gtgtccctgt 660
tggcttttct gcttctcatc ttctccctgt ggagacacct gaaaaccatg caatactatg 720
ctaaaggctc cgaagacacc accacagctg cacatataaa ggccttgcac atggtagtgg 780
cctttctcct gttctacaca gttttctttt tgtctcttgc catacaatat tggacctctg 840
ggtctcaaga gaataacaac ctgttttatg ccacaattgt aattactttc ccttcagtcc 900
attcatgtat cctgattctg agaaacagcc agctgaggca ggcatctctg ttggtgctgt 960
ggtggctgct gtgcaagtcc aaagatgtac ggatgttggt tccctgaaat actctgtcaa 1020
tgctctttag tagtgaagaa gaaaatagct tagttaagga aattcttgtt cattaccgaa 1080
gtatactttc aagtttatgt ate 1103
<210>155
<211>308
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R26,mGR26
<400>155
Met Leu Pro Thr Leu Ser Val Phe Phe Met Leu Thr Phe Val Leu Leu
1 5 10 15
Cys Phe Leu Gly Ile Leu Ala Asn Gly Phe Ile Val Leu Met Leu Ser
20 25 30
Arg Glu Trp Leu Leu Arg Gly Arg Leu Leu Pro Ser Asp Met Ile Leu
35 40 45
Phe Ser Leu Gly Thr Ser Arg Phe Phe Gln Gln Cys Val Gly Leu Val
50 55 60
Asn Ser Phe Tyr Tyr Phe Leu His Leu Val Glu Tyr Ser Gly Ser Leu
65 70 75 80
Ala Arg Gln Leu Ile Ser Leu His Trp Asp Phe Leu Asn Ser Ala Thr
85 90 95
Phe Trp Phe Cys Thr Trp Leu Ser Val Leu Phe Cys Ile Lys Ile Ala
100 105 110
Asn Phe Ser His Pro Ala Phe Leu Trp Leu Lys Trp Arg Phe Pro Ala
115 120 125
Leu Val Pro Trp Phe Leu Leu Gly Ser Ile Leu Val Ser Val Ile Val
130 135 140
Thr Leu Leu Phe Phe Trp Gly Asn His Thr Ile Tyr Gln Ala Phe Leu
145 150 155 160
Arg Arg Lys Phe Thr Gly Asn Thr Thr Phe Lys Glu Trp Asn Arg Arg
165 170 175
Leu Glu Ile Asp Tyr Phe Met Pro Leu Lys Val Val Thr Met Ser Ile
180 185 190
Pro Cys Ser Leu Phe Leu Val Ser Ile Leu Leu Leu Ile Ser Ser Leu
195 200 205
Arg Arg His Ser Leu Arg Met Gln His Asn Thr His Ser Leu Gln Asp
210 215 220
Pro Asn Val Gln Ala His Ser Arg Ala Leu Lys Ser Leu Ile Ser Phe
225 230 235 240
Leu Val Leu Tyr Ala Val Ser Phe Val Ser Met Ile Ile Asp Ala Thr
245 250 255
Val Phe Ile Ser Ser Asp Asn Val Trp Tyr Trp Pro Trp Gln Ile Ile
260 265 270
Leu Tyr Phe Cys Met Ser Val His Pro Phe Ile Leu Ile Thr Asn Asn
275 280 285
Leu Arg Phe Arg Gly Thr Phe Arg Gln Leu Leu Leu Leu Ala Arg Gly
290 295 300
Phe Trp Val Ala
305
<210>156
<211>3437
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R26,mGR26
<400>156
gaattctaga caaggaaaga cacacactaa atgactttac ttgtgggacc taaaataacc 60
aaaataagtc aaaatcacag tgatgttact agggatctag gataagggaa tgaagagaaa 120
gatgttggtc atagagtaca aaaattcagc taagaactca gtcctggagg ctgaatgtat 180
agctgtgtga cagacagcag ctagccatac cagagtatac acttgcctct tgctgaaaga 240
gtagatctta tgtgtccttg tcacacataa aagtaattga aaaagtaact ctctgagatg 300
acagatacgt taaaatggtt ttacttttca acctgctcca gtaggggtcc ctttaatgtt 360
tgtgctagta gatgggggac tctcaagtat ctttgtggta gacaaatcta aggtggcctt 420
catgaatacc aacccagact tttgtgactt tgtgatcccc cacttttgaa gtggataaga 480
gctgtgactt gagtctaatc aaaggagtcc aacgtgttgt ttattctgta acagtgcttt 540
gtgtttctag ttaataacac aggcaaagaa ggctagggtg acattcctag gattgtgtta 600
tttctatctt gctcatgcct ccctctgctg gtctaatgaa ataagtcagt ggccatattt 660
aaatatgact acgtggcaaa tactgatgat agcctgtgtg ttccaacaaa tatccagtag 720
gagacctagg cattcagtcc tgcagccaca aggaaatagg ttctttcact ggaaaaagag 780
cagtttagat ggttataaat tacttaatcc atagaagcca taggggcttt atgtagagat 840
ttgggtagag aggtagacct agatattgac ttaggagtgg ctattcctga gtgggggtag 900
atatatggca gggaaactca gataagaaag acttctttag tgtcacgatt tttcctaggt 960
atctccttgt gccagatatc tatgcgtcta tgtacctacc tacctaccta cctacctacc 1020
tacctaccta cctactgaca cctaatagga agaggcaagt ggtcacaacc tgcaatgatg 1080
ggataagaat gatggaactc agttaccaag attaaaatac cttccccact gatgttattg 1140
caagcatggc agcatgtagg caaaatcaga gaaggcaaat catgagcagc tgctgcccca 1200
tggtacccga gcccgggaaa tatttgcatc atatctgagc caaaagcaca ccttttatct 1260
actgcctgag catttttcac attgaagttc tggctcacat gcagaatcca accatttatc 1320
tcctgtctcc agaagggagt gtcagggact gtgggtaggg gcagggagga ggccaggaac 1380
caaggcaatc agtggtgaca ggaggaggga ctgaaatgct accaacatta tcagttttct 1440
tcatgttgac ctttgttctg ctctgtttcc tggggatcct ggccaacggc ttcattgtgc 1500
tgatgctgag cagggaatgg ctactgcgtg gtaggctgct cccctcggac atgatcctct 1560
tcagtttggg cacctcccga ttcttccagc agtgtgtggg attggtcaac agtttctatt 1620
acttcctcca tctggttgag tactccggga gccttgcccg gcagctcatt agtcttcact 1680
gggacttctt gaactcagcc actttctggt tttgtacctg gctcagcgtc ctgttctgta 1740
tcaagattgc taacttctcc catcctgcct tcctgtggtt gaagtggaga ttcccagcgt 1800
tggtgccctg gttcttgttg ggctctatct tggtgtccgt cattgtaact ctgctgttct 1860
tttggggaaa ccacactata tatcaggcat tcttaaggag aaagtttact gggaacacaa 1920
cctttaagga gtggaacaga aggctggaaa tagactattt catgcctctg aaagttgtca 1980
ccatgtcaat tccttgttct ctttttctgg tctcaatttt gctgttgatc agttctctca 2040
gaaggcattc gctaagaatg cagcacaata cccacagctt gcaagacccc aacgtccagg 2100
ctcacagcag agccctgaag tcactcatct cattcctggt tctttatgcg gtgtcctttg 2160
tgtccatgat cattgatgct acagtcttca tctcctcaga taatgtgtgg tattggccct 2220
ggcaaattat actttacttt tgcatgtctg tacatccatt tatcctcatc accaataatc 2280
tcaggttccg cggcaccttc aggcagctac tcctgttggc caggggattc tgggtggcct 2340
agaaggcttg gtctctttat ctagagcctt tgaagagact caggtgaggg taacttcact 2400
tggaagtgag ctcatctacg tggaaatgtc tttgtaggca ggcatggggt catactgtga 2460
ggttcctcat tgggaaagag gagaagaaaa tacagagtgt ccttccttac cttaggatat 2520
tatgaaagtg gaaattccga atcctggacc agtattgatc taagtgcaaa gtacaatatg 2580
tcctgttcct ttcatgtctg ttttcctttt gttactgatt cattctctag ggaatagtct 2640
tgatcaactg aatcatctca tctggctggc cactggggag gtaaaagaac tttgtgtcac 2700
tgctgcattg ggatatacat gggtgggaag caagtgtccc tgaggcagag tagcactcag 2760
tatgagaacc tcaaagagca ggtggctgtg catgcagggg ctggggcaag gagtcctgat 2820
cactcttcac tgtatgggga ttatttgtct cttgccaaaa tttggagact ttggctttag 2880
ttttgtgaag atgactggaa aaattcttaa tgctaccctg tatcatttct caataatatt 2940
ttccttttcc tgcctttaat tttctcctat ctgcagcgcc ccttgcttgt tatccgtaaa 3000
taaataaata aataaataaa taagcccaat cctcattttc ctgtctttgg gaaccctttt 3060
acttccccag gtatacgcta caaagccact tctgcattga ataaacatta tctttcattc 3120
agaaaaagac ttaagaatct cacctttaca aaaaaaaaaa aaaaagaatc tcacttattt 3180
tatattcaaa ttccattttt aaaaagaaaa gcacagcatt aatttttcta aatactgttt 3240
ataaaaataa cttgctctaa gaattataca aatgttttga aaggtaactt tggaaaaaaa 3300
gtgtgattag acatggatgt ttgtaagaca gaacaaagag ctcttggaag tccatggcag 3360
ctcattggtc ttgccttcag tagagcctgt ctgaatcctg taacctctta tgcccttttg 3420
tagcttttct gcagatc 3437
<210>157
<211>340
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R27,mGR27
<400>157
gaattcgccc ttgcgggatc cgggaacgga ttcatagcac tggtaaactt catgggctgg 60
atgaagaata ggaagattgc ctccattgat ttaatcctca caagtctggc catatccaga 120
atttgtctat tgtgcgtaat actattagat tgttttatat tggtgctata tccagatgtc 180
tatgccactg gtaaagaaat gagaatcatt gacttcttct ggacactaac caatcactta 240
agtatctggt ttgcaacctg cctcagcatt tactatttct tcaagatagg taatttcttt 300
cacccacttt tcctatgcct caagtctaga cgccaagggc 340
<210>158
<211>82
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R28,mGR28
<400>158
Gly Arg Glu Trp Leu Arg Tyr Gly Arg Leu Leu Pro Leu Asp Met Ile
1 5 10 15
Leu Ile Ser Leu Gly Ala Ser Arg Phe Cys Leu Gln Leu Val Gly Thr
20 25 30
Val His Asn Phe Tyr Tyr Ser Ala Gln Lys Val Glu Tyr Ser Gly Gly
35 40 45
Leu Gly Arg Gln Phe Phe His Leu His Trp His Phe Leu Asn Ser Ala
50 55 60
Thr Phe Trp Phe Cys Ser Trp Leu Ser Val Leu Phe Cys Val Lys Ile
65 70 75 80
Ala Asn
<210>159
<211>341
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R28,mGR28
<400>159
gaattcgccc ttgcgggatc cgggaacggg tttattgtgc tggtgctggg cagggagtgg 60
ctgcgatatg gcaggttgct gcccttggat atgatcctca ttagcttggg tgcctcccgc 120
ttctgcctgc agttggttgg gacggtgcac aacttctact actctgccca gaaggtcgag 180
tactctgggg gtctcggccg acagttcttc catctacact ggcacttcct gaactcagcc 240
accttctggt tttgcagctg gctcagtgtc ctgttctgtg tgaagattgc taacatcaca 300
cactccacct tcctgtgtct caagtctaga cgccaagggc g 341
<210>160
<211>320
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R29,mGR29
<400>160
Met Asp Gly Ile Val Gln Asn Met Phe Thr Phe Ile Val Ile Val Glu
1 5 10 15
Ile Ile Ile Gly Trp Ile Gly Asn Gly Phe Ile Ala Leu Val Asn Cys
20 25 30
Ile His Trp Tyr Lys Arg Arg Lys Ile Ser Ala Leu Asn Gln Ile Leu
35 40 45
Thr Ala Leu Ala Phe Ser Arg Ile Tyr Leu Leu Leu Thr Val Phe Thr
50 55 60
Val Ile Ala Val Ser Thr Leu Tyr Thr His Val Leu Val Thr Arg Arg
65 70 75 80
Val Val Lys Leu Ile Asn Phe His Leu Leu Phe Ser Asn His Phe Ser
85 90 95
Met Trp Leu Ala Ala Cys Leu Gly Leu Tyr Tyr Phe Leu Lys Ile Ala
100 105 110
His Phe Pro Asn Ser Ile Phe Val Tyr Leu Lys Met Arg Ile Asn Gln
115 120 125
Val Val Ser Gly Thr Leu Leu Met Ser Leu Gly Leu Leu Phe Leu Asn
130 135 140
Thr Leu Leu Ile Asn Ser Tyr Ile Asp Thr Lys Ile Asp Asp Tyr Arg
145 150 155 160
Glu His Leu Leu Tyr Asp Phe Thr Ser Asn Asn Thr Ala Ser Phe Tyr
165 170 175
Arg Val Ile Leu Val Ile Asn Asn Cys Ile Phe Thr Ser Ile Pro Phe
180 185 190
Thr Leu Ser Gln Ser Thr Phe Leu Leu Leu Ile Phe Ser Leu Trp Arg
195 200 205
His Tyr Lys Lys Met Gln Gln His Ala Gln Arg Cys Arg Asp Val Leu
210 215 220
Ala Asp Ala His Ile Arg Val Leu Gln Thr Met Val Thr Tyr Val Leu
225 230 235 240
Leu Cys Ala Ile Phe Phe Leu Ser Leu Ser Met Gln Ile Leu Arg Ser
245 250 255
Glu Leu Leu Lys Asn Ile Leu Tyr Val Arg Phe Cys Glu Ile Val Ala
260 265 270
Ala Val Phe Pro Ser Gly His Ser Cys Val Leu Ile Cys Arg Asp Thr
275 280 285
Asn Leu Arg Gly Thr Phe Leu Ser Val Leu Ser Trp Leu Lys Gln Arg
290 295 300
Phe Thr Ser Trp Ile Pro Asn Ile Asn Cys Arg Ser Ser Cys Ile Phe
305 310 315 320
<210>161
<211>1108
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R29,mGR29
<400>161
agcttgatat ttcctatttg ttactgcaca gagttttttt taaaaattga gtttgttatg 60
tggattcaat actcagatag agctctttaa tttttttaca gtgacctcat gaatcataac 120
ttgccttaca gacaatggat ggaatcgtac agaacatgtt tacattcatt gtaattgtgg 180
aaataataat aggatggatt ggaaatggat tcatagctct ggtgaactgc atacactggt 240
acaagagaag aaagatctct gcactgaatc aaatactcac agccttggct ttctccagaa 300
tctaccttct tttaacagta ttcactgtta tagcagtgtc tacgctatac acacacgtgt 360
tggtaactag aagagtggta aaactgatta atttccattt gcttttcagc aatcatttta 420
gcatgtggct tgctgcatgc cttggccttt attattttct taaaatagct cattttccta 480
actctatttt tgtttactta aagatgagaa ttaaccaggt ggtttcaggg actttgctca 540
tgtctttggg cctcttgttt ctaaacactc tgctgataaa ctcatacatt gataccaaga 600
tagatgacta cagagaacat ctactgtatg atttcacttc gaataatact gcttcatttt 660
acagggttat tttagtcatt aacaactgta ttttcacatc tatacccttt acactttccc 720
agtccacttt tctcctgctc atcttctccc tgtggagaca ttacaagaag atgcaacagc 780
atgcacaaag atgcagagat gtccttgcag atgcccacat cagagtcttg caaaccatgg 840
tcacctatgt cctactctgt gccattttct ttctgtctct ttccatgcaa attttgagga 900
gtgagttgtt gaagaacatt ctttacgtta ggttctgcga gattgttgca gcagtttttc 960
cttcaggaca ctcctgtgtc ttaatctgta gagacacaaa cctgagaggg acctttcttt 1020
ctgtgctatc gtggctgaag cagaggttta catcatggat tcctaacata aattgcagat 1080
catcttgcat attctaaaag aaactgag 1108
<210>162
<211>312
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R30,mGR30
<400>162
Met Thr Tyr Glu Thr Asp Thr Thr Leu Met Leu Val Ala Val Gly Glu
1 5 10 15
Ala Leu Val Gly Ile Leu Gly Asn Ala Phe Ile Ala Leu Val Asn Phe
20 25 30
Met Gly Trp Met Lys Asn Arg Lys Ile Ala Ser Ile Asp Leu Ile Leu
35 40 45
Ser Ser Val Ala Met Ser Arg Ile Cys Leu Gln Cys Ile Ile Leu Leu
50 55 60
Asp Cys Ile Ile Leu Val Gln Tyr Pro Asp Thr Tyr Asn Arg Gly Lys
65 70 75 80
Glu Met Arg Thr Val Asp Phe Phe Trp Thr Leu Thr Asn His Leu Ser
85 90 95
Val Trp Phe Ala Thr Cys Leu Ser Ile Phe Tyr Leu Phe Lys Ile Ala
100 105 110
Asn Phe Phe His Pro Leu Phe Leu Trp Ile Lys Trp Arg Ile Asp Lys
115 120 125
Leu Ile Leu Arg Thr Leu Leu Ala Cys Val Ile Ile Ser Leu Cys Phe
130 135 140
Ser Leu Pro Val Thr Glu Asn Leu Ser Asp Asp Phe Arg Arg Cys Val
145 150 155 160
Lys Thr Lys Glu Arg Ile Asn Ser Thr Leu Arg Cys Lys Val Asn Lys
165 170 175
Ala Gly His Ala Ser Val Lys Val Asn Leu Asn Leu Val Met Leu Phe
180 185 190
Pro Phe Ser Val Ser Leu Val Ser Phe Leu Leu Leu Ile Leu Ser Leu
195 200 205
Trp Arg His Thr Arg Gln Ile Gln Leu Ser Val Thr Gly Tyr Lys Asp
210 215 220
Pro Ser Thr Thr Ala His Val Lys Ala Met Lys Ala Val Ile Ser Phe
225 230 235 240
Leu Ala Leu Phe Val Val Tyr Cys Leu Ala Phe Leu Ile Ala Thr Ser
245 250 255
Ser Tyr Phe Met Pro Glu Ser Glu Leu Ala Val Ile Trp Gly Glu Leu
260 265 270
Ile Ala Leu Ile Tyr Pro Ser Ser His Ser Phe Ile Leu Ile Leu Gly
275 280 285
Ser Ser Lys Leu Lys Gln Ala Ser Val Arg Val Leu Cys Arg Val Lys
290 295 300
Thr Met Leu Lys Gly Lys Lys Tyr
305 310
<210>163
<211>3775
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R30,mGR30
<400>163
aaaaatgttc attgtttatc taaaattcaa atttaactga gtgccctaca tttttattta 60
ttcaatctag tagctgtact gaggttatta gtgtgatttc tgaagcccaa atttgtaaaa 120
cttagcctca gataaacagc ttgagaccat ggaaagtaat ttggtaaatt tgcatcttag 180
caaatagtag ctcagcctaa attaactgtg tgtagaaaag aatgacctgc ggagaagata 240
aatggacata caatatccag gctaaggatt gccaaacaca ctgtttttaa gactaattga 300
gatttagata aactatctac agtcttcatg tataattctc atcttcatca caagacagac 360
ttcaacttaa ggaggtaaag acaaggacag cgaaccctaa acagccaagt gtagaaacca 420
aactgcatca aatcagccag aaactaattg gatacttctc tactttaaaa tgacatacga 480
aacagatact accttaatgc ttgtagctgt tggtgaggcc ttagtaggga ttttaggaaa 540
tgcattcatt gcactggtaa acttcatggg ctggatgaag aataggaaga ttgcctctat 600
tgatttaatc ctctcaagtg tggccatgtc cagaatttgt ctacagtgta taatcctatt 660
agattgtatt atattggtgc agtatccaga cacctacaac agaggtaaag aaatgaggac 720
cgttgacttc ttctggacac ttaccaacca tttaagtgtc tggtttgcca cctgcctcag 780
cattttctat ttattcaaga tagcaaactt cttccaccct cttttcctct ggataaagtg 840
gagaattgac aagctaattc tcagaactct actggcatgt gtgattatct ccctgtgttt 900
tagcctccca gtcactgaaa atctgagtga tgatttcaga cgttgtgtta agacaaagga 960
gagaataaac tctactttga gatgcaaagt aaataaagct ggacatgcct ctgtcaaggt 1020
aaatctcaac ttggtcatgc tgttcccctt ttctgtgtct ctggtctcct ttctcctctt 1080
gatcctctcc ctgtggagac acaccaggca gatacaactc agtgtaacag ggtacaaaga 1140
tcccagcaca acagctcatg tgaaagccat gaaagcagta atttccttcc tggccctgtt 1200
tgttgtctac tgcctagcct ttctcatagc cacctccagc tactttatgc cagagagtga 1260
attagctgta atatggggtg agctgatagc tctaatctat ccttcaagcc attcatttat 1320
cctcatcctg gggagtagta aactaaaaca agcatctgtg agggtgcttt gtagagtaaa 1380
gaccatgtta aagggaaaaa aatattagca tcatgagcat atctgaagaa aaactatcac 1440
tttctaagag aaaggaagac acgatcatta tccgtccttt tcacatgaat attgatttca 1500
tgcagtgaca tcctcttaac aaacttaaat tgaaccttga gaaatctcat atacagcaac 1560
tttgcatgtc tctatctctg ctttttctct ccttttcaat atgagttgac ataaaaaata 1620
attttcagaa caaattataa cagaagaaag ggcattttca taatcagttc tgaatcactc 1680
ctccaaatgc aaagctgcct gacaaattca aaacaattgt aacagcatct cactgtcgtt 1740
tgcattcttt ggaaaagcag gtggtttgtt cttggagcct ggcttagagt tttcttctta 1800
gaccattgaa ttatgttcat gattggagaa gagtcaagta ccaagtaaca atttttattg 1860
tgaagatggg tgttcatcat gtgattttgg ctggcctgga acttgttatg tagactagtc 1920
tgtcatcaaa cacacaaaga tctgcctgcc tcacctgcca gttctaggat tcaaggaatg 1980
caccaccaca gcttgttcaa gtgacaattc ttacaaatgt tttagaaata aataatatac 2040
tagaaattaa cactgaatgt aagtgctgtt taggtataaa ttatgattaa atgttatagt 2100
tagaaaatta tttaagatta tagatcagtg atgaaaatat tctagaataa gttttatgaa 2160
gaaactttta taaagaaact ggaaaaaaat ctcttgattg catattgaaa caaatttctc 2220
caaaaagaac acctacaaat ttgctctaga catctagact gtatcaaaca gtgaatatga 2280
aaatatcata acaggatata gcctttagta ttgaagacag gttcatctat attaaacctg 2340
catacatacc taaaagacta agtcaatatc ccacaaacat atttgcacta tcatgtctat 2400
tgaaacacta ttcatagtag ctaaaatatg gcacaaaact agacattcat caatagatga 2460
atcaataaag caaatgtaca tacacaagat gaaattgtat tcaggcataa agaagaatgc 2520
agtcatgtca ttagcaaaaa cataaacaga attggaggtc attgtgataa ttgaaataaa 2580
ccagacctgg aaaaaacaaa acctgtgtaa tttttctgaa gtagagaata tactcttgga 2640
tggatagatg ggtactgtta tagtataaaa tgtgtgtgtg tgtgtgtgtg tgtgtgtgtg 2700
tatttcatga aagcaagaat gggactgctt agagaaagaa aaggacaaac aggtgaaggg 2760
gtgaaagaaa aaggcaatga caaggagtaa tgatatgagc aaagtaccat tattaaacat 2820
gtgacaatat tatatagaaa cacatgattt tgtgtgccta ccaaaactgg ataataattt 2880
ttaaaatgta tctattaaaa ggaaagaaaa gaaagtgcaa gcccaggaaa gggagaaaag 2940
gaaacaatga gagagaaatg gaaaatggtg agaagtgaag agaacaaaaa gaaatggagt 3000
aagtgtggcc aggaatgaag gatctcagct atagttatcc cagtacggta atacaaatct 3060
gtgactccag cacttgacaa ggctgagaga tgtgagagag ggccagttaa caaccagtct 3120
gggcttattc caagagataa gaagattggg ggaaagtatg tagaagggtt tggagggaag 3180
agagagaaga gggaaatgat gtaatgatag tacaaatcaa aagttatttt ttctaaaaaa 3240
gcaatgggac aggaaaccaa cctaacaagt aaaggtgctt ggttcacaag accagcaacc 3300
tgagtgcatc cttgctagaa tgaaattggc cttactctgg aaagcttact tcctcagtgt 3360
attcattgtt aaaattcatg tggagatttt aaagaaaaaa ggaaaaaaaa agttaaatgg 3420
tagatttgtg taggggaata ttcccctaat taattgatta gataataaag atgacaagca 3480
aattgctgtg caaaaaggaa gacaaggtct aagaggggaa gaggggacac gggaggaaaa 3540
aaaacggccc tttttaaagc aaggtgggga gtgagggaag cgagatgtag acagggaact 3600
gttagacctg gtggcagctt ctgccacctg aagattttca acatagtata gttcatgagt 3660
ttaggaagat atgttccctg cccagcggtt gtatcatctg ttgattttaa actaagattg 3720
tctggtgttt tccatttgcg gagactcaag tagaccaaag ggaaagaatg aattc 3775
<210>164
<211>310
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R31,mGR31
<400>164
Met Tyr Met Ile Leu Val Arg Ala Val Phe Ile Thr Gly Met Leu Gly
1 5 10 15
Asn Met Phe Ile Gly Leu Ala Asn Cys Ser Asp Trp Val Lys Asn Gln
20 25 30
Lys Ile Thr Phe Ile Asn Phe Ile Met Val Cys Leu Ala Ala Ser Arg
35 40 45
Ile Ser Ser Val Leu Met Leu Phe Ile Asp Ala Thr Ile Gln Glu Leu
50 55 60
Ala Pro His Phe Tyr Tyr Ser Tyr Arg Leu Val Lys Cys Ser Asp Ile
65 70 75 80
Phe Trp Val Ile Thr Asp Gln Leu Ser Thr Trp Leu Ala Thr Cys Leu
85 90 95
Ser Ile Phe Tyr Leu Phe Lys Val Ala His Ile Ser His Pro Leu Phe
100 105 110
Leu Trp Leu Lys Trp Arg Leu Arg Gly Val Leu Val Val Phe Leu Val
115 120 125
Phe Ser Leu Phe Leu Leu Ile Ser Tyr Phe Leu Leu Leu Glu Thr Leu
130 135 140
Pro Ile Trp Gly Asp Ile Tyr Val Thr Leu Lys Asn Asn Leu Thr Leu
145 150 155 160
Phe Ser Gly Thr Ile Lys Thr Thr Ala Phe Gln Lys Ile Ile Val Phe
165 170 175
Asp Ile Ile Tyr Leu Val Pro Phe Leu Val Ser Leu Ala Ser Leu Leu
180 185 190
Leu Leu Phe Leu Ser Leu Val Lys His Ser Arg Ser Leu Asp Leu Ile
195 200 205
Ser Thr Thr Ser Glu Asp Ser Arg Thr Lys Ile His Lys Lys Ala Met
210 215 220
Lys Met Leu Val Ser Phe Leu Ile Leu Phe Ile Ile His Ile Phe Phe
225 230 235 240
Met Gln Leu Ala Arg Trp Leu Leu Phe Leu Phe Pro Met Ser Arg Pro
245 250 255
Ile Asn Phe Ile Leu Thr Leu Asn Ile Phe Ala Leu Thr His Ser Phe
260 265 270
Ile Leu Ile Leu Gly Asn Ser Asn Leu Arg Gln Arg Ala Met Arg Ile
275 280 285
Leu Gln His Leu Lys Ser Gln Leu Gln Glu Leu Ile Leu Ser Leu His
290 295 300
Arg Phe Ser Ser Leu Tyr
305 310
<210>165
<211>4675
<212>DNA
<213>小鼠(Mus sp.)
<220>
<223>小鼠T2R31,mGR31
<220>
<221>modified_base
<222>(1)..(4675)
<223>n=g,a,c或t
<400>165
ctgcagcttt ctagaaatct caccagaatg tctttgtgca gctttaatag ttcctggtta 60
taccttgtca cattataagc taagacatct ttggtgccac aatatactct cactaatcag 120
agagattaga cagaaaaata agtttcttaa caactgtttt agatagggtc atgaaatgac 180
ataaaacacc aatgctaagg caatccatta tgttttctca tgaggagccc atatgtacac 240
ttgagtgtgt cttattattt ccctgagtga ttttgtaatt ttattaaaca cttaactgtg 300
attcatacta gttagttctg aaattctttt cttcatcaaa gccattaatc ctggggtttt 360
ttaaatggag aaccccaaaa caaagtgaaa tgttgtgtgt ggagcaggct gtcttcccac 420
acactaccat gagatgctca ttctgtaatt gttccccgga ataggaaatg ccctgaattc 480
aggcacacaa gagctagtct gtgcaccatg tctggttctt gcattaatac ccacttttgt 540
cacgaagctt cattgattcg catcttcaga agctggtatc attattagtt tctttcctca 600
ggtgactctg gnccaaaata ttanggcgcc ctttaaaaaa gtaaaactac aaaatttctt 660
tataattttc tttaagtttg ttataatata gcatgaccta cacacacaca cacacacaca 720
cacacacaca cacacacaca agtatgcctc tcctttcctt ctaaaaatct cacttaaagc 780
aattgtttag ctgtcttcga agtctagact gccactgtcg tgcttctagc caaaacaaat 840
gcaacacata aaatgataga gctcaaaact taggaatcta tttaactgtg aagatcacgc 900
aagcaaacct gagaaacctc tagaaggaaa ccacagcaaa tcactggaga gaaggtgtta 960
atctagtaag aatagttttt attttgggta tccttttgta gattggttag ttcatccaaa 1020
atccaacttg ttagttcttc ataaattgta agtgtctcca acatcaaagc accacttctc 1080
tcttttcccc tgtatgaaga tgctttaagt acagagttac tctttttctg tactgacagt 1140
aatttaaaaa aattgttcac tcattctttt ttggtgttgt tattctgtgt tcctcaatgt 1200
tatctttttt ttttcaaaac tttcttttat aaaaagtcat acacatagca aatgcagtgc 1260
atgtttatgg aatccataac taacttattg agacttctcc tagtactttc tttgaacagt 1320
aacaaagata tctgcttcta cagagtgcag tgtttcaggt gaggaggaac atattataca 1380
aatcagtgaa aaaaaaatct gattcaaatt tgtattttaa tatatttgac tttatcactt 1440
cagatattac atcaatggga attttgaagg cacacaagtg atgatgtggg catagagact 1500
gtctgtacta gaatttaata tttcttttaa atatctttaa ataaaaatat gatgctgtat 1560
tcataaacag atctttatag attaagtatg agattaaagt tggaaaaaca aaagacaaaa 1620
acctaggact aagaatttcc ttaagtatgt gtgaatatca acctaatgga ggaagtttcc 1680
aatcaaagct gaaattacag taaaaaggag gaagataaat atggaaaagg atgattttct 1740
gtggaagttt gtttgagaac tgatccacga gacaaattgc tagaagtgtg gattcccttt 1800
tactattcaa ctgcttatag gactggatca aatgtatatg atactggtaa gagcagtatt 1860
tataactgga atgctgggaa atatgttcat tggactggca aactgctctg actgggtcaa 1920
gaaccagaaa atcaccttca tcaacttcat catggtctgt ttggcagctt ccagaatcag 1980
ctctgtgctg atgttattta ttgatgcaac catacaagaa ctagcgcctc atttctatta 2040
ttcttaccgt ctagtaaaat gctctgatat attctgggtt ataactgatc aactatcaac 2100
atggcttgcc acctgcctga gcatattcta cttattcaaa gtagcccaca tttcccatcc 2160
ccttttcctc tggttgaagt ggagattgag aggtgtgctt gttgtttttc ttgtattttc 2220
tttgttctta ttgatttctt attttctact gcttgaaaca cttcctattt ggggagatat 2280
ttatgtaacc cttaaaaaca atctgacctt attttcaggt acaattaaga ccactgcttt 2340
tcaaaagata attgtttttg atataatata tttagtccca tttcttgtgt ccctagcatc 2400
attgctcctt ttatttttgt ccttggtgaa acactcccga agccttgacc tgatttctac 2460
cacttctgaa gattccagaa ccaagattca taagaaggcc atgaaaatgc tggtgtcttt 2520
cctcattctc tttataattc acattttttt catgcagtta gcacggtggt tattattttt 2580
gtttccaatg agcaggccaa ttaatttcat cttaacatta aatatctttg ccttaactca 2640
ctcatttatt ctcatcctgg gaaatagcaa tcttcgacag agagcaatga ggatcctgca 2700
acatcttaaa agccagcttc aagagctgat cctctccctt catagattct ccagtcttta 2760
ctagaggaac agcttaacag ggagacttgg aaggtcactg gcaaattatt cttctttgat 2820
ttcttttaag tactgctgaa catatatgaa ctgtccccag agcatagtgc tatcttatga 2880
gaaggatatc atctcacagt ctggttataa aacacaaacc aatcttttta taatttcttt 2940
acagcattgc taataaaaga cttgtagtct caaatatttt aaagagaata attaatttta 3000
taggcaaaag gtatgaaatt acaattcaca gggaaggttc atgactcctt agatattaaa 3060
gttaattgta agccacaata ggcagaagat gagcaaaatg ttgataggag ataaataaaa 3120
tctaaagtta cggagaaaaa aaacatcaac ttgcctttta gattacttta aagctctctc 3180
tctcgctctc tctctctgta tctacttact ttatatatac aaatgttttg tctgcatgta 3240
tttctttgca ccatataaat gtctaagtat ccagaangtc agcagagggc atcaaattct 3300
ctggaaagag agttacaaat tgctgtgggt aacactgggt gctgggaact aacctgagtc 3360
ctctgccaca gcaactgctc ttccctgctg agtcatgttt taagtctcca caacttaaac 3420
tcattgttga tgtggtcatt gcataatgat gaatttacat tctaaggttt gtatcatagg 3480
taggagggct ggttttaatc atattctaat gttcttatac aaacccaggt tttgtaagag 3540
actgtattct atcatgagac tctttcccca caccgccaat gtaacatttt tattaatttt 3600
gaggggaatt ttatacagtg taccctgatc acccttgctt cccactcctt gcaggtctac 3660
cctcccacca ttgctcaatc ccccctaaaa gagagagaaa caaaccatgt ccaatttgtg 3720
ttggacacat actcagtgga acatggccaa acccctagtg agcagttcct taaagaaaac 3780
taagctgcct ccccaccact accaccatag ggcattaact gtgaagagct acactttagc 3840
tattttatca ccaatttaaa agactgtctt caatagcttc ctctatggac tgtttctggt 3900
tttagtggga cagggagaag gggtcaagag gttgtcacag aaacttttga tgtctcttat 3960
tctcagttaa agtccactgc aaaagaagtc tgctggctct aataaagctt gcaacagcat 4020
gggccagtga catcatcatg atttctggca acaatatgga ccacaaatat catggctcag 4080
gtggcattac ggaccacaga catcaacatg gtctctggca gcaagaacca gaatcttttg 4140
aggaggcttc attcagaaaa tgaatttttc ttcatcccag atatactgat gttgctcaat 4200
cagagtatta gtatggttgg gcaccatatt tggggacagg accttcaata tttccaggct 4260
gctgtgtaac acattatctt tagtgtcagg tgcccttagt gtcaggacat gaccatcatg 4320
tatgcgcctg tgggcagaaa tacatctttg tactttctta cacctagcag ggtgagtagc 4380
aggagcagcg gcattaatac ttccatacct ctgggcagcc tatcaggtat catctaggca 4440
aggtaagccc agtagtggcc caaggctcct ggtgtctact tggcaacaac atgctccttt 4500
gtctgcactg ccatatctat ggctggttct ccatccctag ttctgcttct ctcaggtttt 4560
atacgactct attccacatt ctatttttcc agttccatga aaccagtgtt taaaagtatc 4620
atcccataag accggccttt taaaggttat tctggagata ttgcagagtc tgcag 4675
<210>166
<211>20
<212>PRT
<213>人工序列
<220>
<223>人工序列的描述:T2R家族共有序列1,保守基序T2R跨膜区1
<220>
<221>MOD_RES
<222>(2)
<223>Xaa=Phe或Ala
<220>
<221>MOD_RES
<222>(3)
<223>Xaa=Ile,Val或Thr
<220>
<221>MOD_RES
<222>(4)
<223>Xaa=Val或Leu
<220>
<221>MOD_RES
<222>(6)
<223>Xaa=Ile或Val
<220>
<221>MOD_RES
<222>(7)
<223>Xaa=Leu或Val
<220>
<221>MOD_RES
<222>(10)
<223>Xaa=Gly或Thr
<220>
<221>MOD_RES
<222>(13)
<223>Xaa=Val或Ala
<220>
<221>MOD_RES
<222>(18)
<223>Xaa=Ile或Met
<400>166
Glu Xaa Xaa Xaa Gly Xaa Xaa Gly Asn Xaa Phe Ile Xaa Leu Val Asn
1 5 10 15
Cys Xaa Asp Trp
20
<210>167
<211>14
<212>PRT
<213>人工序列
<220>
<223>人工序列的描述:T2R家族共有序列2,保守基序T2R跨膜区2
<220>
<221>MOD_RES
<222>(1)
<223>Xaa=Asp或Gly
<220>
<221>MOD_RES
<222>(2)
<223>Xaa=Phe或Leu
<220>
<221>MOD_RES
<222>(3)
<223>Xaa=Ile或Leu
<220>
<221>MOD_RES
<222>(5)
<223>Xaa=Thr或Ile
<220>
<221>MOD_RES
<222>(6)
<223>Xaa=Gly,Ala或Ser
<220>
<221>MOD_RES
<222>(13)
<223>Xaa=Cys,Gly或Phe
<400>167
Xaa Xaa Xaa Leu Xaa Xaa Leu Ala Ile Ser Arg Ile Xaa Leu
1 5 10
<210>168
<211>13
<212>PRT
<213>人工序列
<220>
<223>人工序列的描述:T2R家族共有序列3,保守基序T2R跨膜区3
<220>
<221>MOD_RES
<222>(3)
<223>Xaa=Leu或Phe
<220>
<221>MOD_RES
<222>(4)
<223>Xaa=Ser,Thr或Asn
<220>
<221>MOD_RES
<222>(5)
<223>Xaa=Leu,Ile或Val
<220>
<221>MOD_RES
<222>(7)
<223>Xaa=Phe或Leu
<220>
<221>MOD_RES
<222>(8)
<223>Xaa=Ala或Thr
<220>
<221>MOD_RES
<222>(10)
<223>Xaa=Cys,Ser或Asn
<220>
<221>MOD_RES
<222>(12)
<223>Xaa=Ser,Asn或Gly
<220>
<221>MOD_RES
<222>(13)
<223>Xaa=Ile或Val
<400>168
Asn His Xaa Xaa Xaa Trp Xaa Xaa Thr Xaa Leu Xaa Xaa
1 5 10
<210>169
<211>18
<212>PRT
<213>人工序列
<220>
<223>人工序列的描述:T2R家族的共有序列4,保守基序T2R跨膜区4
<220>
<221>MOD_RES
<222>(3)
<223>Xaa=Phe或Cys
<220>
<221>MOD_RES
<222>(8)
<223>Xaa=Asn或Ser
<220>
<221>MOD_RES
<222>(11)
<223>Xaa=His或Asn
<220>
<221>MOD_RES
<222>(12)
<223>Xaa=Pro或Ser
<220>
<221>MOD_RES
<222>(13)
<223>Xaa=Leu,Ile或Val
<220>
<221>MOD_RES
<222>(16)
<223>Xaa=Trp或Tyr
<400>169
Phe Tyr Xaa Leu Lys Ile Ala Xaa Phe Ser Xaa Xaa Xaa Phe Leu Xaa
1 5 10 15
Leu Lys
<210>170
<211>14
<212>PRT
<213>人工序列
<220>
<223>人工序列的描述:T2R家族共有序列5,保守基序T2R跨膜区5
<220>
<221>MOD_RES
<222>(4)
<223>Xaa=Ile,Phe或Val
<220>
<221>MOD_RES
<222>(8)
<223>Xaa=Lys或Arg
<220>
<221>MOD_RES
<222>(10)
<223>Xaa=Ser或Thr
<220>
<221>MOD_RES
<222>(11)
<223>Xaa=Lys或Arg
<220>
<221>MOD_RES
<222>(12)
<223>Xaa=Gln或Lys
<220>
<221>MOD_RES
<222>(13)
<223>Xaa=Met或Ile
<220>
<221>MOD_RES
<222>(14)
<223>Xaa=Gln或Lys
<400>170
Leu Leu Ile Xaa Ser Leu Trp Xaa His Xaa Xaa Xaa Xaa Xaa
1 5 10
<210>171
<211>14
<212>PRT
<213>人工序列
<220>
<223>人工序列的描述:T2R家族共有序列6,保守基序T2R跨膜区6
<220>
<221>MOD_RES
<222>(3)
<223>Xaa=Phe或Leu
<220>
<221>MOD_RES
<222>(4)
<223>Xaa=Ile或Val
<220>
<221>MOD_RES
<222>(7)
<223>Xaa=Leu或Met
<220>
<221>MOD_RES
<222>(8)
<223>Xaa=Gly,Ser或Thr
<220>
<221>MOD_RES
<222>(10)
<223>Xaa=Pro,Ser或Asn
<220>
<221>MOD_RES
<222>(13)
<223>Xaa=Lys或Arg
<220>
<221>MOD_RES
<222>(14)
<223>Xaa=Gln或Arg
<400>171
His Ser Xaa Xaa Leu Ile Xaa Xaa Asn Xaa Lys Leu Xaa Xaa
1 5 10
<210>172
<211>300
<212>PRT
<213>小鼠(Mus sp.)
<220>
<223>在来自品尝者和非品尝者品系的等位基因中具有氨基酸取代的小鼠T2R05,mGR05
<220>
<221>MOD_RES
<222>(44)
<223>Xaa=Ile或Thr
<220>
<221>MOD_RES
<222>(85)
<223>Xaa=Val或Ile
<220>
<221>MOD_RES
<222>(101)
<223>Xaa=Ala或Thr
<220>
<221>MOD_RES
<222>(155)
<223>Xaa=Asp或Gly
<220>
<221>MOD_RES
<222>(294)
<223>Xaa=Arg或Leu
<400>172
Met Leu Ser Ala Ala Glu Gly Ile Leu Leu Ser Ile Ala Thr Val Glu
1 5 10 15
Ala Gly Leu Gly Val Leu Gly Asn Thr Phe Ile Ala Leu Val Asn Cys
20 25 30
Met Asp Trp Ala Lys Asn Asn Lys Leu Ser Met Xaa Gly Phe Leu Leu
35 40 45
Ile Gly Leu Ala Thr Ser Arg Ile Phe Ile Val Trp Leu Leu Thr Leu
50 55 60
Asp Ala Tyr Ala Lys Leu Phe Tyr Pro Ser Lys Tyr Phe Ser Ser Ser
65 70 75 80
Leu Ile Glu Ile Xaa Ser Tyr Ile Trp Met Thr Val Asn His Leu Thr
85 90 95
Val Trp Phe Ala Xaa Ser Leu Ser Ile Phe Tyr Phe Leu Lys Ile Ala
100 105 110
Asn Phe Ser Asp Cys Val Phe Leu Trp Leu Lys Arg Arg Thr Asp Lys
115 120 125
Ala Phe Val Phe Leu Leu Gly Cys Leu Leu Thr Ser Trp Val Ile Ser
130 135 140
Phe Ser Phe Val Val Lys Val Met Lys Asp Xaa Lys Val Asn His Arg
145 150 155 160
Asn Arg Thr Ser Glu Met Tyr Trp Glu Lys Arg Gln Phe Thr Ile Asn
165 170 175
Tyr Val Phe Leu Asn Ile Gly Val Ile Ser Leu Phe Met Met Thr Leu
180 185 190
Thr Ala Cys Phe Leu Leu Ile Met Ser Leu Trp Arg His Ser Arg Gln
195 200 205
Met Gln Ser Gly Val Ser Gly Phe Arg Asp Leu Asn Thr Glu Ala His
210 215 220
Val Lys Ala Ile Lys Phe Leu Ile Ser Phe Ile Ile Leu Phe Val Leu
225 230 235 240
Tyr Phe Ile Gly Val Ser Ile Glu Ile Ile Cys Ile Phe Ile Pro Glu
245 250 255
Asn Lys Leu Leu Phe Ile Phe Gly Phe Thr Thr Ala Ser Ile Tyr Pro
260 265 270
Cys Cys His Ser Phe Ile Leu Ile Leu Ser Asn Ser Gln Leu Lys Gln
275 280 285
Ala Phe Val Lys Val Xaa Gln Leu Leu Lys Phe Phe
290 295 300
Claims (25)
1.一种鉴定在味觉细胞中调节味觉信号传递的化合物的方法,其特征在于,该方法包括步骤:
(i)使味觉转导G-蛋白偶联受体多肽与化合物接触,所述多肽具有SEQ ID NO:7的氨基酸序列;和
(ii)确定该化合物对该多肽的功能性作用。
2.如权利要求1所述的方法,其特征在于,所述功能性作用是化学作用。
3.如权利要求1所述的方法,其特征在于,所述功能性作用是物理作用。
4.如权利要求1所述的方法,其特征在于,所述功能性作用是通过测量所述化合物与所述多肽的胞外结构域或跨膜区域的结合确定的。
5.如权利要求1所述的方法,其特征在于,所述功能性作用是通过测量放射性GTP与所述多肽的结合决定的。
6.如权利要求1所述的方法,其特征在于,所述多肽是重组的。
7.如权利要求1所述的方法,其特征在于,所述多肽在细胞或细胞膜中表达。
8.如权利要求7所述的方法,其特征在于,所述功能性作用是通过测定表达所述多肽的细胞中电活动的变化确定的。
9.如权利要求7所述的方法,其特征在于,所述功能性作用是通过测量胞内cAMP、cGMP、IP3或Ca2+的变化确定的。
10.如权利要求9所述的方法,其特征在于,所述胞内Ca2+的变化是通过检测细胞中FURA-2依赖性荧光的变化确定的。
11.如权利要求7所述的方法,其特征在于,所述细胞是真核细胞。
12.如权利要求11所述的方法,其特征在于,所述细胞是HEK-293细胞。
13.如权利要求7所述的方法,其特征在于,所述细胞含有Gα15。
14.如权利要求9所述的方法,其特征在于,所述多肽与化合物在苦味物质的存在下接触,在所述化合物存在下,苦味物质对细胞的功能性作用的变化和在所述化合物不存在下,苦味物质对细胞的功能性作用的变化表明所述化合物能调节味觉细胞中的味觉信号传递。
15.如权利要求1所述的方法,其特征在于,所述多肽与固相连接。
16.如权利要求15所述的方法,其特征在于,所述多肽与固相共价连接。
17.一种编码味觉转导G-蛋白偶联受体的分离的核酸,其特征在于,该受体具有SEQ ID NO:7的氨基酸序列。
18.如权利要求17所述的核酸,其特征在于,所述核酸具有SEQ ID NO:8的核苷酸序列。
19.一种表达盒,其特征在于,该表达盒含有权利要求17所述的核酸。
20.如权利要求19所述的表达盒,其特征在于,所述核酸与异源启动子可操纵性连接。
21.一种含有权利要求19所述的表达盒的分离的细胞。
22.如权利要求21所述的细胞,其特征在于,该细胞是真核细胞。
23.一种分离的味觉转导G-蛋白偶联受体,其特征在于,该受体具有SEQ IDNO:7的氨基酸序列。
24.权利要求23所述的受体的用途,其特征在于,用该受体与异源多肽共价连接,形成嵌合多肽。
25.权利要求23所述的受体的用途,其特征在于,用于制备与权利要求23的受体选择性结合的抗体。
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US09/393,634 US6558910B2 (en) | 1999-09-10 | 1999-09-10 | SF, a novel family of taste receptors |
US09/393,634 | 1999-09-10 | ||
US09/510,332 US7244584B2 (en) | 1999-09-10 | 2000-02-22 | T2R, a novel family of taste receptors |
US09/510,332 | 2000-02-22 |
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CNA2006101598520A Division CN101058605A (zh) | 1999-09-10 | 2000-09-08 | T2r,一类味觉受体 |
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EP (1) | EP1214343A2 (zh) |
JP (1) | JP4339542B2 (zh) |
CN (1) | CN1289525C (zh) |
AU (2) | AU773600B2 (zh) |
CA (1) | CA2384777A1 (zh) |
IL (1) | IL148593A0 (zh) |
MX (1) | MXPA02002538A (zh) |
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AU3665893A (en) | 1992-02-12 | 1993-09-03 | United States Of America, As Represented By The Secretary, Department Of Health And Human Services, The | Sequences characteristic of human gene transcription product |
JP2001502525A (ja) | 1996-09-04 | 2001-02-27 | モーヘン インターナショナル エヌ.ブイ. | 抗真菌タンパク質、これをコードするこのdna、およびこのdnaを組込む宿主 |
AU2685599A (en) | 1998-02-18 | 1999-09-06 | Genetics Institute Inc. | Secreted proteins and polynucleotides encoding them |
WO2000038536A2 (en) | 1998-12-23 | 2000-07-06 | Mount Sinai School Of Medicine Of New York University | Inhibitors of the bitter taste response |
EP1214343A2 (en) | 1999-09-10 | 2002-06-19 | The Regents Of The University Of California | T2r taste receptor family |
US6558910B2 (en) | 1999-09-10 | 2003-05-06 | The Regents Of The University Of California | SF, a novel family of taste receptors |
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Cited By (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US9525148B2 (en) | 2008-04-03 | 2016-12-20 | Qd Vision, Inc. | Device including quantum dots |
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AU773600B2 (en) | 2004-05-27 |
IL148593A0 (en) | 2002-09-12 |
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US7465550B2 (en) | 2008-12-16 |
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WO2001018050A2 (en) | 2001-03-15 |
CN1387536A (zh) | 2002-12-25 |
US8329885B2 (en) | 2012-12-11 |
NO20021164D0 (no) | 2002-03-08 |
AU2004203826A1 (en) | 2004-09-16 |
US8624012B2 (en) | 2014-01-07 |
CA2384777A1 (en) | 2001-03-15 |
WO2001018050A3 (en) | 2002-01-10 |
US20150260733A1 (en) | 2015-09-17 |
US20180128842A1 (en) | 2018-05-10 |
NO20021164L (no) | 2002-05-10 |
US20050048586A1 (en) | 2005-03-03 |
US20130189712A1 (en) | 2013-07-25 |
US9063124B2 (en) | 2015-06-23 |
US20120009598A1 (en) | 2012-01-12 |
WO2001018050A9 (en) | 2002-02-07 |
US7595166B2 (en) | 2009-09-29 |
EP1214343A2 (en) | 2002-06-19 |
JP4339542B2 (ja) | 2009-10-07 |
US20140120553A1 (en) | 2014-05-01 |
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