CN117050150A - 一种次黄嘌呤光学探针及其制备方法和应用 - Google Patents

一种次黄嘌呤光学探针及其制备方法和应用 Download PDF

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CN117050150A
CN117050150A CN202210481174.9A CN202210481174A CN117050150A CN 117050150 A CN117050150 A CN 117050150A CN 202210481174 A CN202210481174 A CN 202210481174A CN 117050150 A CN117050150 A CN 117050150A
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杨弋
赵玉政
李写
董文萍
张莉娟
王承诺
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East China University of Science and Technology
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Abstract

本发明涉及次黄嘌呤光学探针。具体地，本发明提供一种次黄嘌呤结合蛋白变体以及包含其的次黄嘌呤光学探针，所述次黄嘌呤结合蛋白变体具有SEQ ID NO:1所示的序列并且具有突变。本发明的荧光探针荧光动态变化大，特异性好，可在细胞内外高通量、定量检测次黄嘌呤。

Description

一种次黄嘌呤光学探针及其制备方法和应用
技术领域
本发明涉及光学探针技术领域，尤其涉及一种次黄嘌呤光学探针及其制备方法和应用。
背景技术
次黄嘌呤(HX)是一种天然嘌呤碱，在嘌呤分解代谢过程中产生，然后转化为黄嘌呤(X)和尿酸(UA)，并在黄嘌呤氧化酶(XO)的作用下产生活性氧(ROS)(Lawal等，Talanta2012，100，217-228)。由于其结构是极性的，而且金结构较小，因此易在体液和组织中积累。作为尿酸的前体和嘌呤降解的中间体，监测HX的含量变化在肉制品的质量控制和临床诊断中都具有巨大的意义(Dalkiran，B等，Sensors&Actuators：B.Chemical，2014，200：83-91；Muamer Dervisevic等，Sensors&Actuators:B.Chemical 2016，225：181-187)。
次黄嘌呤由肌苷通过核苷磷酸化酶衍生而来。次黄嘌呤鸟嘌呤磷酸核糖转移酶(HGPRT)是嘌呤补救途径中的一种酶，HGPRT通常催化次黄嘌呤形成IMP。未溶出的次黄嘌呤随后被氧化成黄嘌呤，黄嘌呤在黄嘌呤氧化酶的作用下进一步氧化成尿酸。在人体中，尿酸是嘌呤降解的最终产物，并在尿液中排泄。Lesch–Nyhan综合征是一种严重的神经系统疾病，由HGPRT缺乏引起，HGPRT的缺乏会导致尿酸的积累，从而导致痛风和肾脏疾病，以及认知障碍等(Philip G等，Wiley Interdiplinary Reviews:Membrane Transport andSignaling，2012，1)。
次黄嘌呤在临床诊断中引起广泛关注，它可以穿过细胞膜并在人体的体液，如血清、羊水、尿液、玻璃体液等积累。据报道，正常人血清中次黄嘌呤的浓度在14至38μM之间(Causs e等，Electrophoresis 2007，28(3)，381–387)。对基于HX水平上来诊断病理状况，科学家们已经进行了许多研究。包括脑缺血、高尿酸血症、缺氧、白血病、围产期窒息、肺炎、肾衰竭、黄尿症等。(Farthing,D等，Journal of Chromatography B 2007，854，158–164；Smith,E等，Best Practice&Research Clinical Rheumatology 2010，24(6)，811–827)。此外，通过检测次黄嘌呤还可以在早期诊断急性心脏缺血或心肌缺血(Farthing,D等，Experimental Biology and Medicine 2015，240,821–831),最近，Wang,Y等人(Wang,Y等，Journal of Clinical Laboratory Analysis 2019，33(5)，1–7)发现痛风患者即使在血尿酸水平正常的情况下，其次黄嘌呤水平亦高于正常人。此外，还发现肥胖患者的血浆HX水平高于瘦型患者(Furuhashi，Koyama，Higashiura，Murase，Nakamura，&Matsumoto，2020)。这些报道的研究强调了HX在临床上的重要性。
目前最常用的检测次黄嘌呤的方法有亲水相互作用液相色谱和紫外-可见光谱(HILIC-UV)(Greco，G等，Journal of Chromatographic Sciences 2013，51，684–693)，反相高效液相色谱法(Iqbal,J等，Electrophoresis 2008，29，3685–3693)，高效毛细管电泳法(Daniel Ana Paula等，Journal of food science 2014，79(6)：S1205-11)、质谱法(Khajehsharifi,H等，Foods and Raw Materials 2011，2(2)，161–171)分光光光度法(Rong,S等，Food Chemistry 2015，170，303–307)以及基于黄嘌呤氧化酶的生物传感器法(Homaei,A等，Journal of Chemical Biology 2013，6，185–205)等。这些检测分析方法或需要专业分析仪器设备，或步骤繁琐，无法区分次黄嘌呤和黄嘌呤，易出现人为误差，且仅适用于体外检测，无法实时监控活细胞内的次黄嘌呤浓度变化。因此，亟待开发新的检测方法，实现在细胞内、外，简便、快捷、特异性高的实时定位、定量、高通量检测次黄嘌呤。
发明内容
本发明的目的在于提供在细胞内、外实时定位、高通量、定量检测次黄嘌呤的探针和方法。
为了实现上述发明目的，本发明提供以下技术方案：
本发明第一方面提供一种次黄嘌呤结合蛋白变体，其：
(1)具有SEQ ID NO:1所示的序列并且在选自以下的1个、2个或3个、4个、5个、6个、7个或更多个位点具有突变：P129、L130、D146、W147、G148、E149、A150、K151、A152、N311、K312、R313、E314、E315、P316，所述氨基酸突变包括氨基酸的修饰、取代或缺失，
(2)是(1)的具有第58-341位氨基酸的截短变体，或
(3)是与(1)或(2)的序列具有至少70％序列相同性并具有(1)所述突变并保留对次黄嘌呤敏感性的序列。
在一个或多个实施方案中，所述突变的位点选自以下的1个、2个或3个、4个、5个、6个或7个：P129、L130、W147、G148、E149、A150、K151、A152、N311、K312、E315、P316。
在一个或多个实施方案中，所述突变包括选自以下任一组的位点处的突变：E149和A150，A150和K151，G148和E149，K151和A152，N311和K312，E315和P316。
在一个或多个实施方案中，所述突变至少包括K151和A152位点处的突变。在一些实施方案中，所述突变还包括选自P129、L130、W147、E149和A150中的1个、2个、3个、4个或5个位点处的突变。优选地，所述突变还包括W147、E149和A150位点处或P129、L130、W147、E149和A150位点处的突变。
在一个或多个实施方案中，P129突变为Q。在一个或多个实施方案中，L130突变为M。在一个或多个实施方案中，W147突变为R。在一个或多个实施方案中，G148突变为P。在一个或多个实施方案中，E149突变为G、L、I、S、T、C、Y、V、H、D或R。在一个或多个实施方案中，A150突变为V、L、I、C、R、S、W、M、F或K。在一个或多个实施方案中，K151突变为C、I、V、R、L、Q、A、F、Y或N。在一个或多个实施方案中，A152突变为AL、F、P、G、R、I或M。在一个或多个实施方案中，N311突变为K。在一个或多个实施方案中，K312突变为A或G。在一个或多个实施方案中，E315突变为V。在一个或多个实施方案中，P316突变为Q。
在一个或多个实施方案中，所述突变选自以下47项中的任一项：(1)E149L和A150F，(2)E149I、A150K，(3)A150M、K151C，(4)G148P、E149G，(5)K151L、A152L，(6)K151C、A152L，(7)K151Q、A152F，(8)N311K、K312A，(9)N311K、K312G，(10)E315V、P316Q，(11)K151R、A152P，(12)K151R、A152L，(13)K151A、A152G，(14)K151L、A152R，(15)K151F、A152L，(16)K151Y、A152I，(17)K151V、A152L，(18)K151N、A152M，(19)K151L、A152A，(20)K151I、A152L，(21)P129Q、L130M、W147R、K151V、A152L、E149S、A150I，(22)W147R、K151V、A152L、E149T、A150V，(23)W147R、K151V、A152L、E149G、A150V，(24)W147R、K151V、A152L、E149G、A150L，(25)W147R、K151V、A152L、E149C、A150L，(26)W147R、K151V、A152L、E149Y、A150I，(27)W147R、K151V、A152L、E149G、A150I，(28)W147R、K151V、A152L、E149C、A150I，(29)W147R、K151V、A152L、E149S、A150L，(30)W147R、K151V、A152L、E149V、A150I，(31)W147R、K151V、A152L、E149G、A150I，(32)W147R、K151V、A152L、E149G、A150C，(33)W147R、K151V、A152L、E149S、A150I，(34)W147R、K151V、A152L、E149S、A150V，(35)W147R、K151V、A152L、E149G、A150R，(36)W147R、K151V、A152L、E149G、A150F，(37)W147R、K151V、A152L、E149T、A150I，(38)W147R、K151V、A152L、E149I、A150I，(39)W147R、K151V、A152L、E149H、A150F，(40)W147R、K151V、A152L、E149S、A150F，(41)W147R、K151V、A152L、E149G、A150S，(42)W147R、K151V、A152L、E149D、A150F，(43)W147R、K151V、A152L、E149R、A150I，(44)W147R、K151V、A152L、E149G、A150W，(45)W147R、K151V、A152L、E149S、A150L，(46)W147R、K151V、A152L、E149R、A150F，(47)W147R、K151V、A152L、E149C、A150F。
本发明另一方面提供一种次黄嘌呤光学探针，包含次黄嘌呤敏感多肽和光学活性多肽，其中光学活性多肽位于次黄嘌呤敏感多肽的序列内。次黄嘌呤敏感多肽被光学活性多肽分为第一部分和第二部分。
在一个或多个实施方案中，所述次黄嘌呤光学探针，包括次黄嘌呤敏感多肽B和光学活性多肽A，其中光学活性多肽A位于次黄嘌呤敏感多肽B的序列内，将次黄嘌呤敏感多肽B分为第一部分B1和第二部分B2，形成B1-A-B2式的探针结构。
在一个实施方式中，次黄嘌呤敏感多肽具有：
(1)SEQ ID NO:1所示的序列或其具有第58-341位氨基酸的截短变体，或与它们有至少70％序列相同性并保留对次黄嘌呤敏感性的序列，
(2)本文第一方面任一实施方案所述的次黄嘌呤结合蛋白变体的序列，或
(3)与(2)所述序列具有至少70％序列相同性并具有(2)所述突变并保留对次黄嘌呤敏感性的序列。
在一个实施方式中，光学活性多肽是荧光蛋白或其功能变体。在一个实施方式中，荧光蛋白选自黄色荧光蛋白、橘黄色荧光蛋白、红色荧光蛋白、绿色荧光蛋白、蓝色荧光蛋白、苹果红荧光蛋白。优选地，光学活性多肽是cpYFP。
在一个实施方式中，荧光蛋白具有SEQ ID NO:2-9中任一所示的序列。在一个或多个实施方案中，荧光蛋白的功能变体具有SEQ ID NO:2所示的序列并在Y1位点具有突变。优选地，所述突变为Y1C、Y1M、Y1Q、Y1V、Y1R、Y1W、Y1L或Y1H。
在一个实施方式中，光学探针还包含侧接所述光学活性多肽的一个或多个接头。本发明所述接头可以是任何长度的任何氨基酸序列。在一个实施方式中，光学活性多肽侧翼包含不超过5个氨基酸的接头，例如0、1、2、3、4个氨基酸的接头。在一个实施方式中，光学活性多肽侧翼的接头包含氨基酸Y。在一个实施方式中，接头Y位于光学活性多肽的N端和/或C端。在一个实施方式中，光学探针如下所示：次黄嘌呤敏感多肽的第一部分B1-Y-光学活性多肽A-次黄嘌呤敏感多肽的第二部分B2。在一个实施方式中，本发明光学探针不包含接头。
在一个实施方式中，本发明光学探针还包含定位序列，用于将探针定位到例如细胞的特定细胞器。
在一个实施方式中，光学活性多肽位于次黄嘌呤敏感多肽的残基66-72，97-101，123-128，145-149，147-156，160-163，187-191，210-215，220-223，237-239，246-248，275-279，291-294，312-317或321-324之间，编号对应于次黄嘌呤敏感多肽的全长。在一个实施方式中，光学活性多肽置换次黄嘌呤敏感多肽的残基66-72，97-101，123-128，145-149，147-156，160-163，187-191，210-215，220-223，237-239，246-248，275-279，291-294，312-317和/或321-324中的一个或多个氨基酸，编号对应于次黄嘌呤敏感多肽的全长。
在一个实施方式中，光学活性多肽位于次黄嘌呤敏感多肽的选自下述位点中的任一个或多个：66/67、66/68、66/69,66/70、66/71、66/72、67/68、67/69、67/70、67/71、67/72、68/69、68/70、68/71、68/72、69/70、69/71、69/72、70/71、70/72、71/72、97/98、97/99、97/100、97/101、98/99、98/100、98/101、99/100、99/101、100/101、123/124、123/125、123/126、123/127、123/128、124/125、124/126、124/127、124/128、125/126、125/127、125/128、126/127、126/128、127/128、145/146、145/147、145/148、145/149、146/147、146/148、146/149、147/148、147/149、147/150、147/151、147/152、147/153、147/154、147/155、147/156、148/149、148/150、148/151、148/152、148/153、148/154、148/155、148/156、149/150、149/151、149/152、149/153、149/154、149/155、149/156、150/151、150/152、150/153、150/154、150/155、150/156、151/152、151/153、151/154、151/155、151/156、152/153、152/154、152/155、152/156、153/154、153/155、153/156、154/155、154/156、155/156、160/161、160/162,1 60/163、161/162、161/163、162/163、187/188、187/189、187/190、187/191、188/189、188/190、188/191、189/190、189/191、190/191、210/211、210/212、210/213、210/214、210/215、211/212、211/213、211/214、211/215、212/213、212/214、212/215、213/214、213/215、214/215、220/221、220/222、220/223、221/222、221/223、222/223、237/238、237/239、238/239、246/247、246/248、247/248、275/276、275/277、275/278、275/279、276/277、276/278、276/279、277/278、277/279、278/279、291/292、291/293、291/294、292/293、292/294、293/294、312/313、312/314、312/315、312/316、312/317、313/314,313/315、313/316、313/317、314/315、314/316、314/317、315/316、315/317、316/317、321/322、321/323、321/324、322/323、322/324、323/324。
在一个或多个实施方案中，次黄嘌呤敏感多肽是SEQ ID NO:1的具有第58-341位氨基酸的截短变体，并且光学活性多肽位于次黄嘌呤敏感多肽的选自下述位点中的任一个或多个：147/149，149/150，149/151，150/151，312/315，313/315或314/315。优选地，光学探针具有SEQ ID NO:10-16中任一项所示序列或由其组成。
在一个或多个实施方案中，次黄嘌呤敏感多肽具有本文第一方面任一实施方案所述的次黄嘌呤结合蛋白变体的序列，并且光学活性多肽位于次黄嘌呤敏感多肽的选自下述位点中的任一个或多个：147/149、149/150、149/151、150/151、312/315、314/315；优选地，光学活性多肽位于次黄嘌呤敏感多肽的150/151位点。
在一个或多个实施方案中，光学探针包含氨基酸序列SEQ ID NO:17-76中任一项所示的序列，或与其有至少70％序列相同性的序列。
本发明另一方面还提供融合多肽，包含本文所述光学探针和其它多肽。在一些实施方式中，其他多肽位于所述光学探针的N端和/或C端。在一些实施方式中，其他多肽包括将光学探针定位到不同细胞器或亚细胞器的多肽、用于纯化的标签或者用于免疫印迹的标签。
本发明另一方面还提供核酸分子，其包含：(a)本文任一实施方案所述的多肽或探针的编码序列，或(b)(a)的互补序列，或(c)(a)或(c)的片段。
在一个或多个实施方案中，所述片段是引物。
本发明还涉及上述核酸分子的变体，包括编码本发明光学探针或融合蛋白的片段、类似物、衍生物、可溶性片段和变体的核酸序列或其互补序列。
本发明另一方面还提供包含本文所述核酸分子的核酸构建物。该核酸序列编码本发明所述光学探针或融合多肽。
在一个或多个实施方案中，所述核酸构建物是克隆载体、表达载体或重组载体。
在一个或多个实施方案中，所述核酸分子与表达控制序列操作性连接。
在一些实施方案中，表达载体选自原核表达载体、真核表达载体和病毒载体。
本发明另一方面还提供一种宿主细胞，所述宿主细胞：(1)表达本发明任一实施方案所述的光学探针或融合多肽；(2)包含本发明任一实施方案所述的核酸分子；或(3)包含本发明任一实施方案所述的核酸构建物。所述宿主细胞优选大肠杆菌。
本发明另一方面还提供次黄嘌呤检测试剂盒，包括本文所述光学探针或融合多肽或多核苷酸或如本文所述方法制备的光学探针。
在一个或多个实施方案中，所述试剂盒还包含选自以下的一种或多种试剂：缓冲液、培养基、次黄嘌呤标准品。
本发明另一方面提供制备本文所述光学探针的方法，包括：提供表达本文所述光学探针或融合多肽的宿主细胞，在所述细胞表达的条件下培养所述宿主细胞，和分离光学探针或融合多肽。
在一个或多个实施方案中，所述方法包括包括以下步骤：1)将编码本文所述次黄嘌呤光学探针的核酸分子纳入表达载体；2)将表达载体转移到宿主细胞中；2)在适合所述表达载体表达的条件下培养所述宿主细胞，3)分离次黄嘌呤光学探针。
本发明另一方面还提供检测样品中次黄嘌呤的方法，包括：使本文所述光学探针或融合多肽或宿主细胞与样品接触，和检测光学活性多肽的变化。所述检测可以在体内、体外、亚细胞或原位进行。所述样品例如血液。
本文另一方面还提供定量样品中次黄嘌呤的方法，包括：使本文所述光学探针或融合多肽或宿主细胞与样品接触，检测光学活性多肽的光学变化，和根据光学活性多肽的光学变化定量样品中的次黄嘌呤。
本发明另一方面还提供筛选化合物(例如药物)的方法，包括：在含次黄嘌呤的体系中使本文所述光学探针或融合多肽或宿主细胞与候选化合物接触，检测光学活性多肽的光学变化，和根据光学活性多肽的光学变化筛选化合物。所述方法可以高通量地筛选化合物。
在一个或多个实施方案中，在含次黄嘌呤的体系中使本文所述宿主细胞与候选化合物接触，并且光学活性多肽的光学变化指示所述候选化合物是否能调节细胞对次黄嘌呤的摄取。
在一个或多个实施方案中，所述体系是溶液体系、细胞体系、亚细胞体系。
本发明另一方面还提供本文所述次黄嘌呤光学探针或融合多肽或宿主细胞在检测样品中的次黄嘌呤、筛选化合物或次黄嘌呤细胞内/外定位中的应用。在一个或多个实施方案中，所述定位是实时定位。
本发明的有益效果：本发明提供的次黄嘌呤光学探针易于成熟，荧光动态变化大，特异性好，并且能够通过基因操作的方法在细胞中表达，可在细胞内外实时定位、高通量、定量检测次黄嘌呤，省去了耗时的处理样品步骤。实验效果表明本申请所提供的次黄嘌呤光学探针对次黄嘌呤的最高响应达到对照的40倍以上，并且可以在细胞浆、线粒体、细胞核、内质网、溶酶体和高尔基体等亚细胞结构中对细胞进行定位、定性、定量检测，并且可以进行高通量的化合物筛选以及血液中次黄嘌呤定量检测。
附图说明
下面结合附图和实施例对本发明作进一步说明。
图1为实施例1所述的示例性次黄嘌呤光学探针的SDS-PAGE图；
图2为实施例7所述的示例性次黄嘌呤光学探针对不同浓度次黄嘌呤的响应的滴定曲线图；
图3为实施例7所述的示例性次黄嘌呤光学探针对4种次黄嘌呤类似物的特异性检测的柱状图；
图4为实施例8所述的示例性次黄嘌呤光学探针在哺乳动物细胞中的亚细胞器定位照片；
图5为实施例8所述的对示例性次黄嘌呤光学探针在哺乳动物细胞中对胞浆中的次黄嘌呤浓度进行动态监测的示意图；
图6为实施例9所述的示例性次黄嘌呤光学探针在活细胞水平进行高通量化合物筛选的点图；
图7为实施例10所述的示例性次黄嘌呤光学探针对小鼠和人血液中的次黄嘌呤进行定量的柱状图；
具体实施方式
在给出数值或范围时，本文所用术语“约”指该数值或范围在给定数值或范围的20％以内、10％以内和5％以内。
本文所用术语“包含”、“包括”和其等同形式包括“含有”以及“由……组成”的含义，例如“包含”X的组合物可仅由X组成或可含有其它物质，例如X+Y。
本文所用术语“次黄嘌呤敏感多肽”或“次黄嘌呤响应多肽”指对次黄嘌呤产生响应的多肽，所述响应包括与敏感多肽的相互作用相关的多肽的化学，生物学，电学或生理学参数的任何响应。响应包括小的变化，例如，多肽的氨基酸或肽片段的方向的变化以及例如多肽的一级，二级或三级结构的变化，包括例如质子化，电化学势和/或构象的变化。“构象”是分子中包含侧基的分子的一级，二级和三级结构的三维排列；当分子的三维结构发生变化时，构象发生变化。构象变化的实例包括从α-螺旋转变为β-折叠或从β-折叠转变为α-螺旋。可以理解的是，只要荧光蛋白部分的荧光被改变，可检测到的改变不需要是构象改变。本文所述次黄嘌呤敏感多肽还可包括其功能变体。次黄嘌呤敏感多肽的功能变体包括但不限于可以与次黄嘌呤相互作用从而发生与亲本次黄嘌呤敏感多肽相同或相似变化的变体。
本发明所述次黄嘌呤敏感多肽包括来源于大肠杆菌(Escherichia coli)的次黄嘌呤结合蛋白PurR或其截短变体，或与它们有90％以上同源性的变体。PurR是LacI/GalR转录阻遏物家族的成员，可以感应次黄嘌呤浓度的变化，在次黄嘌呤浓度动态变化的过程中次黄嘌呤结合蛋白的空间构象也会发生改变。示例性PurR蛋白如SEQ ID NO:1所示。示例性地，PurR蛋白的保留次黄嘌呤结合活性的截短变体是SEQ ID NO:1的包含第58-341位氨基酸的片段，优选为SEQ ID NO:1的第58-341位氨基酸。本文中，当描述光学活性多肽在次黄嘌呤敏感多肽中的插入位点或描述次黄嘌呤结合蛋白的突变位点时，氨基酸残基编号均参考SEQ ID NO:1。
本文所用术语“光学探针”是指与光学活性多肽融合的次黄嘌呤敏感多肽。发明人发现，次黄嘌呤敏感多肽例如次黄嘌呤结合蛋白专一性地对生理浓度的次黄嘌呤结合后所产生的构象变化会引起光学活性多肽(例如荧光蛋白)的构象变化，进而导致光学活性多肽的光学性质发生改变。借助不同次黄嘌呤浓度下测定的荧光蛋白的荧光绘制标准曲线，可以检测并分析次黄嘌呤的存在和/或水平。
在本发明的光学探针中，光学活性多肽(例如荧光蛋白)可操作地插入次黄嘌呤敏感多肽中。基于蛋白质的“光学活性多肽”是具有发射荧光能力的多肽。荧光是光学活性多肽的一种光学性质，其可用作检测本发明的光学探针的响应性的手段。如本文所用，术语“荧光性质”是指适当激发波长下的摩尔消光系数，荧光量子效率，激发光谱或发射光谱的形状，激发波长最大值和发射波长最大值，两个不同波长激发的振幅，两个不同波长的发射振幅比，激发态寿命或荧光各向异性。活性和无活性状态之间的这些性质中的任何一个的可测量的差异足以用于本发明的荧光蛋白底物在活性测定中的效用。可测量的差异可通过确定任何定量荧光性质的量来确定，例如，特定波长处的荧光量或荧光在发射光谱上的积分。优选地，选择蛋白质底物以具有在未激活和活化的构象状态下容易区分的荧光特性。本文所述光学活性多肽还可包括其功能变体。光学活性多肽的功能变体包括但不限于可以发生与亲本光学活性多肽相同或相似荧光性质变化的变体。
“接头”或“连接区”指在本发明多肽、蛋白质或核酸中连接两个部分的氨基酸或核苷酸序列。示例性地，本发明中次黄嘌呤敏感多肽与光学活性多肽的连接区氨基端的氨基酸数目选择的是0-3个，羧基端的氨基酸数目选择的是0-2个；当重组光学探针作为基本单元与功能蛋白连接时，可以融合在重组光学探针的氨基酸或羧基端。接头序列可为一个或多个柔性氨基酸组成的短肽链，如Y。
本文所用术语“生色团”、“荧光团”与“荧光蛋白”同义，指在激发光照射下发出荧光的蛋白质。荧光蛋白作为生物科学领域的基础检测手段，例如生物技术领域常用的绿色荧光蛋白GFP及由该蛋白突变衍生出的环状重排的蓝色荧光蛋白(cpBFP)、环状重排的绿色荧光蛋白(cpGFP)、环状重排的黄色荧光蛋白(cpYFP)等；还有本技术领域常用的红色荧光蛋白RFP，及由该蛋白衍生出来的环状重排的蛋白，如cpmApple，cpmOrange，cpmKate等。示例性cpYFP的序列如SEQ ID NO:2所示。示例性cpmOrange的序列如SEQ ID NO:3所示。示例性cpmKate如SEQ ID NO:4或8所示。示例性mCherry如SEQ ID NO:5所示。示例性cpGFP如SEQID NO:6所示。示例性cpBFP如SEQ ID NO:7所示。示例性cpmApple如SEQ ID NO:9所示。
本发明中的荧光蛋白也包括具有突变的功能变体。例如，cpYFP的功能变体具有SEQ ID NO:2所示的序列并在Y1位点具有突变。优选地，所述突变为Y1C、Y1M、Y1Q、Y1V、Y1R、Y1W、Y1L或Y1H。
本发明所述的次黄嘌呤光学探针包括次黄嘌呤敏感多肽B(例如次黄嘌呤结合蛋白或其变体)和光学活性多肽A(例如荧光蛋白)。光学活性多肽A插入到次黄嘌呤敏感多肽B中，将B分为B1和B2两个部分，形成B1-A-B2式的探针结构；次黄嘌呤敏感多肽B和次黄嘌呤相互作用导致光学活性多肽A的光学信号变强。
在本发明的光学探针中，光学活性多肽以N-C方向位于N-C方向的次黄嘌呤敏感多肽的如下区域，氨基酸残基66-72，97-101，123-128，145-149，147-156，160-163，187-191，210-215，220-223，237-239，246-248，275-279，291-294和312-317以及321-324区域。示例性地，光学活性多肽位于次黄嘌呤结合蛋白的氨基酸序列的如下位点：66/67、66/68、66/69,66/70、66/71、66/72、67/68、67/69、67/70、67/71、67/72、68/69、68/70、68/71、68/72、69/70、69/71、69/72、70/71、70/72、71/72,97/98、97/99、97/100、97/101、98/99、98/100、98/101、99/100、99/101、100/101、123/124、123/125、123/126、123/127、123/128、124/125、124/126、124/127、124/128、125/126、125/127、125/128、126/127、126/128、127/128、145/146、145/147、145/148、145/149、146/147、146/148、146/149、147/148、147/149、147/150、147/151、147/152、147/153、147/154、147/155、147/156、148/149、148/150、148/151、148/152、148/153、148/154、148/155、148/156、149/150、149/151、149/152、149/153、149/154、149/155、149/156、150/151、150/152、150/153、150/154、150/155、150/156、151/152、151/153、151/154、151/155、151/156、152/153、152/154、152/155、152/156、153/154、153/155、153/156、154/155、154/156、155/156、160/161、160/162,160/163、161/162、161/163、162/163、187/188、187/189、187/190、187/191、188/189、188/190、188/191、189/190、189/191、190/191、210/211、210/212、210/213、210/214、210/215、211/212、211/213、211/214、211/215、212/213、212/214、212/215、213/214、213/215、214/215、220/221、220/222、220/223、221/222、221/223、222/223、237/238、237/239、238/239、246/247、246/248、247/248、275/276、275/277、275/278、275/279、276/277、276/278、276/279、277/278、277/279、278/279、291/292、291/293、291/294、292/293、292/294、293/294、312/313、312/314、312/315、312/316、312/317、313/314,313/315、313/316、313/317、314/315、314/316、314/317、315/316、315/317、316/317、321/322、321/323、321/324、322/323、322/324、323/324。本文中，如果以“X/Y”形式表示的位点中的两个数字是连续的整数，则表示光学活性多肽位于该数字所述的氨基酸之间。例如插入位点147/148表示光学活性多肽位于次黄嘌呤敏感多肽的氨基酸147与148之间。如果以“X/Y”形式表示的位点中的两个数字不是连续的整数，则表示光学活性多肽置换该数字所示氨基酸之间的氨基酸。例如插入位点147/156表示光学活性多肽置换次黄嘌呤敏感多肽的氨基酸148-155。优选地，光学活性多肽插入次黄嘌呤敏感多肽的下述位点：147/149，149/150，149/151，150/151，312/315，313/315或314/315。在优选实施方式中，光学活性多肽位于次黄嘌呤结合蛋白的如下位点：147/149，149/150，149/151，150/151，312/315，313/315或314/315。在示例性实施方式中，本发明B1-A-B2式光学探针可为当cpYFP位于PurR的截短变体(例如SEQ ID NO:1的包含第58-341位氨基酸的片段)的147/149，149/150，149/151，150/151，312/315，313/315，314/315时形成的探针，优选如SEQ IDNO:10～16所示。
提到某多肽或蛋白时，本发明所用术语“变体”或“突变体”包括具有所述多肽或蛋白相同功能、但序列不同的变体。多肽或蛋白的变体可包括：同源序列、保守性变体、等位变体、天然突变体、诱导突变体。这些变体包括但并不限于：在所述多肽或蛋白的序列中缺失、插入和/或取代一个或多个(通常为1-30个，较佳地1-20个，更佳地1-10个，最佳地1-5个)氨基酸，以及在其羧基末端和/或氨基末端添加一个或数个(通常为20个以内，较佳地为10个以内，更佳地为5个以内)氨基酸获得的序列。这些变体还可包含与所述多肽或蛋白的序列相同性为至少约70％、至少约75％、至少约80％、至少约85％、至少约90％、至少约95％、至少约98％、至少约99％或100％的多肽或蛋白。不希望受理论限制，氨基酸残基发生改变而不改变多肽或蛋白质的总体构型和功能，即功能保守突变。例如，在本领域中，用性能相近或相似的氨基酸进行取代时，通常不会改变多肽或蛋白的功能。在本领域中，性能相似的氨基酸往往指具有相似侧链的氨基酸家族，在本领域已有明确定义。这些家族包括具有碱性侧链的氨基酸(例如赖氨酸、精氨酸、组氨酸)、具有酸性侧链的氨基酸(例如天冬氨酸、谷氨酸)、具有不带电荷的极性侧链的氨基酸(例如甘氨酸、天冬酰胺、谷氨酰胺、丝氨酸、苏氨酸、酪氨酸、半胱氨酸)、具有非极性侧链的氨基酸(例如丙氨酸、缬氨酸、亮氨酸、异亮氨酸、精氨酸、苯丙氨酸、甲硫氨酸、色氨酸)、具有β-分支侧链的氨基酸(例如苏氨酸、缬氨酸、异亮氨酸)和具有芳香侧链的氨基酸(例如酪氨酸、苯丙氨酸、色氨酸、组氨酸)。又比如，在氨基末端和/或羧基末端添加一个或数个氨基酸通常也不会改变多肽或蛋白的功能。对于许多常见已知非遗传性编码氨基酸的保守氨基酸取代本领域已知。其他非编码氨基酸的保守取代可基于其物理性质与遗传上编码的氨基酸的性质的比较来确定。
发明人发现，在SEQ ID NO:1或其截短变体的选自以下的位点具有突变的次黄嘌呤结合蛋白变体表现出与次黄嘌呤不同的结合活性：P129、L130、D146、W147、G148、E149、A150、K151、A152、N311、K312、R313、E314、E315、P316。所述氨基酸突变包括氨基酸的修饰、取代或缺失。特别地，在SEQ ID NO:1或其截短变体的选自以下的位点具有突变的次黄嘌呤结合蛋白变体更有利于用于本发明的光学探针以更高效检测次黄嘌呤：P129、L130、W147、G148、E149、A150、K151、A152、N311、K312、E315、P316。在较佳的实施方案中，次黄嘌呤结合蛋白变体的突变包括选自以下任一组的位点处的突变：E149和A150，A150和K151，G148和E149，K151和A152，N311和K312，E315和P316。优选地，所述突变至少包括K151和A152位点处的突变；进一步地，所述突变还包括选自P129、L130、W147、E149和A150中的1个、2个、3个、4个或5个位点处的突变。
其中，作为实施例中的示例，在SEQ ID NO:1或其截短变体中，P129突变为Q；L130突变为M；W147突变为R；G148突变为P；E149突变为G、L、I、S、T、C、Y、V、H、D或R；A150突变为V、L、I、C、R、S、W、M、F或K；K151突变为C、I、V、R、L、Q、A、F、Y或N；A152突变为AL、F、P、G、R、I或M；N311突变为K；K312突变为A或G；E315突变为V；P316突变为Q。
在优选实施方案中，针对次黄嘌呤结合蛋白变体(SEQ ID NO:1或其截短变体的)的突变选自以下47项中的任一项：(1)E149L和A150F，(2)E149I、A150K，(3)A150M、K151C，(4)G148P、E149G，(5)K151L、A152L，(6)K151C、A152L，(7)K151Q、A152F，(8)N311K、K312A，(9)N311K、K312G，(10)E315V、P316Q，(11)K151R、A152P，(12)K151R、A152L，(13)K151A、A152G，(14)K151L、A152R，(15)K151F、A152L，(16)K151Y、A152I，(17)K151V、A152L，(18)K151N、A152M，(19)K151L、A152A，(20)K151I、A152L，(21)P129Q、L130M、W147R、K151V、A152L、E149S、A150I，(22)W147R、K151V、A152L、E149T、A150V，(23)W147R、K151V、A152L、E149G、A150V，(24)W147R、K151V、A152L、E149G、A150L，(25)W147R、K151V、A152L、E149C、A150L，(26)W147R、K151V、A152L、E149Y、A150I，(27)W147R、K151V、A152L、E149G、A150I，(28)W147R、K151V、A152L、E149C、A150I，(29)W147R、K151V、A152L、E149S、A150L，(30)W147R、K151V、A152L、E149V、A150I，(31)W147R、K151V、A152L、E149G、A150I，(32)W147R、K151V、A152L、E149G、A150C，(33)W147R、K151V、A152L、E149S、A150I，(34)W147R、K151V、A152L、E149S、A150V，(35)W147R、K151V、A152L、E149G、A150R，(36)W147R、K151V、A152L、E149G、A150F，(37)W147R、K151V、A152L、E149T、A150I，(38)W147R、K151V、A152L、E149I、A150I，(39)W147R、K151V、A152L、E149H、A150F，(40)W147R、K151V、A152L、E149S、A150F，(41)W147R、K151V、A152L、E149G、A150S，(42)W147R、K151V、A152L、E149D、A150F，(43)W147R、K151V、A152L、E149R、A150I，(44)W147R、K151V、A152L、E149G、A150W，(45)W147R、K151V、A152L、E149S、A150L，(46)W147R、K151V、A152L、E149R、A150F，(47)W147R、K151V、A152L、E149C、A150F。
本发明提供具有这些突变的次黄嘌呤结合蛋白变体以及包含此类次黄嘌呤结合蛋白变体作为次黄嘌呤敏感多肽的光学探针。
在具体实施方案中，光学探针中的次黄嘌呤敏感多肽是本文任一实施方案所述的次黄嘌呤结合蛋白变体，并且光学探针中的荧光蛋白如SEQ ID NO:2所示并且具有选自以下的突变：Y1C、Y1M、Y1Q、Y1V、Y1R、Y1W、Y1L、Y1H。优选地，光学探针包含SEQ ID NO:17-76中任一所示的序列。
在两种或多种多肽或核酸分子序列中，术语“相同性”或“相同性百分数”指在比较窗口或指定区域上，采用本领域已知方法如序列比较算法，通过手工比对和目测检查来比较和比对最大对应性时，两个或多个序列或子序列相同或其中在指定区域有一定百分数的氨基酸残基或核苷酸相同(例如，60％、65％、70％、75％、80％、85％、90％、91％、92％、93％、94％、95％、96％、97％、98％、99％或100％相同)。例如，适合测定序列相同性百分数和序列相似性百分数的优选算法是BLAST和BLAST 2.0算法，分别可参见Altschul等(1977)Nucleic Acids Res.25:3389和Altschul等(1990)J.Mol.Biol.215:403。
本领域技术人员公知，在基因克隆操作中，常常需要设计合适的酶切位点，这势必在所表达的多肽或蛋白末端引入了一个或多个不相干的残基，而这并不影响目的多肽或蛋白的活性。又如为了构建融合蛋白、促进重组蛋白的表达、获得自动分泌到宿主细胞外的重组蛋白、或利于重组蛋白的纯化，常常需要将一些氨基酸添加至重组蛋白的N-末端、C-末端或该蛋白内的其它合适区域内，例如，包括但不限于，适合的接头肽、信号肽、前导肽、末端延伸、谷胱甘肽S-转移酶(GST)、麦芽糖E结合蛋白、蛋白A、如6His或Flag的标签，或Xa因子或凝血酶或肠激酶的蛋白水解酶位点。
本文所用术语“功能片段”、“衍生物”和“类似物”是指基本上保持与原始多肽或蛋白(例如次黄嘌呤结合蛋白或荧光蛋白)相同的生物学功能或活性的蛋白。本发明的多肽或蛋白(例如次黄嘌呤结合蛋白或荧光蛋白)的功能变体、衍生物或类似物可以是(i)有一个或多个保守或非保守性氨基酸残基(优选保守性氨基酸残基)被取代的蛋白，而这样的取代的氨基酸残基可以是也可以不是由遗传密码编码的，或(ii)在一个或多个氨基酸残基中具有取代基团的蛋白，或(iii)成熟蛋白与另一个化合物(比如延长蛋白半衰期的化合物，例如聚乙二醇)融合所形成的蛋白，或(iv)附加的氨基酸序列融合到此蛋白序列而形成的蛋白(如分泌序列或用来纯化此蛋白的序列或蛋白原序列，或与抗原IgG片段的形成的融合蛋白)。根据本文的教导，这些功能变体、衍生物和类似物属于本领域熟练技术人员公知的范围。所述类似物还包括具有不同于天然L-氨基酸的残基(如D-氨基酸)的类似物，以及具有非天然存在的或合成的氨基酸(如β、γ-氨基酸)的类似物。应理解，本发明的次黄嘌呤敏感多肽并不限于上述列举的代表性蛋白、变体、衍生物和类似物。修饰(通常不改变一级结构)形式包括：体内或体外的蛋白的化学衍生形式如乙酰化或羧基化。修饰还包括糖基化，如那些在蛋白的合成和加工中或进一步加工步骤中进行糖基化修饰而产生的蛋白。这种修饰可以通过将蛋白暴露于进行糖基化的酶(如哺乳动物的糖基化酶或去糖基化酶)而完成。修饰形式还包括具有磷酸化氨基酸残基(如磷酸酪氨酸，磷酸丝氨酸，磷酸苏氨酸)的序列。还包括被修饰从而提高了其抗蛋白水解性能或优化了溶解性能的蛋白。
本发明融合多肽包含本文所述光学探针和其它多肽。在一些实施方案中，本文所述光学探针还包含与之融合的其它多肽。本文所述其他多肽不影响光学探针的性质。其他多肽可位于所述光学探针的N端和/或C端。在一些实施方案中，其他多肽包括将光学探针定位到不同细胞器或亚细胞器的多肽、用于纯化的标签或者用于免疫印迹的标签。本文所述融合多肽中的光学探针和其它多肽之间可具有接头。
本文所述亚细胞器包括细胞浆、线粒体、细胞核、内质网、细胞膜、高尔基体、溶酶体和过氧化物酶体等。在一些实施方案中，用于纯化的标签或者用于免疫印迹的标签包括6组氨酸(6*His)、谷胱甘肽硫转移酶(GST)、Flag。
本发明包含编码本发明所述次黄嘌呤敏感多肽或光学探针的核酸分子。本发明所用术语“核酸”或“核苷酸”或“多核苷酸”或“核酸序列”可以是DNA形式或RNA形式。DNA形式包括cDNA、基因组DNA或人工合成的DNA。DNA可以是单链的或是双链的。DNA可以是编码链或非编码链。提到核酸时，本文所用术语“变体”可以是天然发生的等位变体或非天然发生的变体。这些核苷酸变体包括简并变体、取代变体、缺失变体和插入变体。如本领域所知的，等位变体是一个核酸的替换形式，它可能是一个或多个核苷酸的取代、缺失或插入，但不会从实质上改变其编码的蛋白的功能。本发明核酸可包含与所述核酸序列的序列相同性为至少约50％、至少约60％、至少约70％、至少约75％、至少约80％、至少约85％、至少约90％、至少约95％、至少约98％、至少约99％或100％的核苷酸序列。本发明还涉及与上述的序列杂交的核酸片段。如本文所用，“核酸片段”的长度至少含15个核苷酸，较好是至少30个核苷酸，更好是至少50个核苷酸，最好是至少100个核苷酸以上。核酸片段可用于核酸的扩增技术(如PCR)。
本发明光学探针或融合蛋白的全长序列或其片段通常可以用PCR扩增法、人工合成法或重组法获得。本领域知晓常规PCR、合成法、重组法的步骤和所用试剂。此外，可通过突变PCR或化学合成等方法将突变引入本发明蛋白序列中。
本发明也涉及核酸构建物，该核酸构建物含有本文所述的多核苷酸，以及与这些序列操作性连接的一个或多个调控序列。本发明所述的多核苷酸可以多种方式被操作以保证所述多肽或蛋白的表达。在将核酸构建物插入载体之前可根据表达载体的不同或要求而对核酸构建物进行操作。利用重组DNA方法来改变多核苷酸序列的技术是本领域已知的。
在某些实施方案中，所述核酸构建物是载体。载体可以是克隆载体，也可以是表达载体，或者是同源重组载体。本发明的多核苷酸可被克隆入许多类型的载体，例如，质粒、噬菌粒、噬菌体衍生物、动物病毒和粘粒。克隆载体可用于提供本发明蛋白或多肽的编码序列。表达载体可以以细菌载体或病毒载体形式提供给细胞。通常通过可操作地连接本发明的多核苷酸至启动子，并将构建体并入表达载体，实现本发明多核苷酸的表达。该载体对于复制和整合真核细胞可为合适的。在一个或多个实施方案中，克隆载体和表达载体是一种载体，即克隆表达载体。同源重组载体用于将本文所述的表达框整合到宿主基因组中。
典型的表达载体包含可用于调节期望核酸序列表达的表达控制序列，与本发明所述的核酸序列或其互补序列操作性连接。本文所用术语“表达控制序列”指调控目的基因的转录、翻译和表达的可以与目的基因操作性连接的元件，可以是复制起点、启动子、标记基因或翻译控制元件，包括增强子、操纵子、终止子、核糖体结合位点等，表达控制序列的选择取决于所用的宿主细胞。在重组表达载体中，“操作性连接”是指目的的核苷酸序列与调节序列以允许核苷酸序列表达的方式连接。本领域的技术人员熟知能用于构建含本发明融合蛋白编码序列和合适的转录/翻译控制信号的表达载体的方法。这些方法包括体外重组DNA技术、DNA合成技术、体内重组技术等。所述的DNA序列可有效连接到表达载体中的适当启动子上，以指导mRNA合成。这些启动子的代表性例子有：大肠杆菌的lac或trp启动子；λ噬菌体PL启动子；真核启动子包括CMV立即早期启动子、HSV胸苷激酶启动子、早期和晚期SV40启动子、反转录病毒的LTR和其他一些已知的可控制基因在原核或真核细胞或其病毒中表达的启动子。表达载体还包括翻译起始用的核糖体结合位点和转录终止子。在一个实施方案中，表达载体可采用市售的pCDF载体，无其他特殊要求。示例性地，采用BamHI和EcoRI分别对编码所述光学探针的核苷酸序列和表达载体进行双酶切，然后将二者的酶切产物连接得到重组表达载体。本发明对酶切和连接的具体步骤和参数没有特殊限定，采用本领域常规的步骤和参数即可。
在获得重组表达载体后，将该载体转化到宿主细胞中，以产生包括融合蛋白的蛋白或肽。此种转移过程可用转化或转染等本领域技术人员熟知的常规技术进行。本发明所述的宿主细胞是指能够接收和容纳重组DNA分子的细胞，是重组基因扩增的场所，理想的受体细胞应该满足易于获取和增殖两个条件。本发明的“宿主细胞”可包括原核细胞和真核细胞，具体包括细菌细胞、酵母细胞、昆虫细胞和哺乳动物细胞。具体的可为大肠杆菌，链霉菌属，鼠伤寒沙门氏菌的细菌细胞，真菌细胞如酵母，植物细胞，果蝇S2或Sf9的昆虫细胞，CHO、COS、HEK293、HeLa细胞、或Bowes黑素瘤细胞的动物细胞等，其中包括但不限于上述的那些宿主细胞。所述宿主细胞优选各种利于基因产物表达或发酵生产的细胞，此类细胞已为本领域熟知并常用。在本发明实施例中所用的示例性宿主细胞为大肠杆菌JM109-DE3菌株。本领域一般技术人员都清楚如何选择适当的载体、启动子、增强子和宿主细胞。
本发明所述的转移到宿主细胞的方法为本领域常规的方法，包括磷酸钙或氯化钙共沉淀、DEAE-甘露聚糖-介导的转染、脂转染、天然感受态、化学介导的转移或电穿孔。当宿主为原核生物如大肠杆菌时，所述方法优选的为CaCl2法或MgCl2法处理，所用的步骤为本领域公知。当宿主细胞是真核细胞时，可选用如下的DNA转染方法：磷酸钙共沉淀法，常规机械方法如显微注射、电穿孔、脂质体包装等。
本发明在将表达载体转入宿主细胞后，对转入表达载体的宿主细胞进行扩增表达培养，分离得到次黄嘌呤光学探针。所述宿主细胞扩增表达培养采用常规的方法即可。根据所用的宿主细胞种类，培养中所用的培养基可以是各种常规培养基。在适于宿主细胞生长的条件下进行培养。
在本发明中，光学探针在细胞内、细胞膜上表达、或分泌到细胞外。如果需要，可利用其物理的、化学的和其它特性通过各种分离方法分离或纯化重组的蛋白。本发明对分离所述次黄嘌呤荧光蛋白的方法没有特殊限定，采用本领域常规的融合蛋白的分离方法即可。这些方法是本领域技术人员所熟知的，包括但并不限于：常规的复性处理、盐析方法、离心、渗透破菌、超声处理、超离心、分子筛层析、吸附层析、离子交换层析、高效液相层析(HPLC)和其它各种液相层析技术及这些方法的结合。在一个实施方案中，利用His标签的亲和层析法进行光学探针的分离。
本发明还提供了所述次黄嘌呤光学探针在次黄嘌呤实时定位、定量检测以及高通量化合物筛选中的应用。在一个方面，所述的次黄嘌呤光学探针优选与细胞不同部位的信号肽连接，转入到细胞中，通过检测细胞中荧光信号的强弱，进行次黄嘌呤的实时定位；通过次黄嘌呤标准滴加曲线结合荧光信号的变化进行相应次黄嘌呤的定量检测。荧光信号的变化通过例如标准化后的荧光信号比值展示，在涉及cpYFP的实施方案中，所述比值是样品的485纳米荧光信号与420纳米荧光信号之比与对照的相应之比的比值。本发明所述的次黄嘌呤标准滴加曲线是根据次黄嘌呤光学探针在不同浓度次黄嘌呤的情况下的荧光信号绘制而成。本发明所述次黄嘌呤光学探针直接转入细胞中，在次黄嘌呤实时定位和定量检测过程中，不需要耗时的样品处理过程，更加准确。本发明次黄嘌呤光学探针在进行高通量化合物筛选时，将不同的化合物添加到细胞培养液中，测定次黄嘌呤含量的变化，从而筛选出对次黄嘌呤含量变化有影响的化合物。在本发明中所述的次黄嘌呤光学探针在次黄嘌呤实时定位、定量检测以及高通量化合物筛选中的应用，均是非诊断和治疗目的，不涉及疾病的诊断和治疗。
本发明还提供包括本文所述光学探针、核酸分子、核酸构建物和/或细胞的检测试剂盒。所述试剂盒还包含检测次黄嘌呤所需的其他试剂。所述其他试剂本领域周知，例如缓冲液、细胞培养基、次黄嘌呤标品。示例性缓冲液例如100mM HEPES和100mM NaCl，pH 7.4。
在本文中，浓度、含量、百分数和其它数值均可用范围的形式表示。也应理解，使用这种范围形式只是为了方便和简洁，应该被弹性地解读为包括范围上下限所明确提及的数值，还应包括该范围内包括的所有单个数值或子范围。
实施例
下面结合实施例对本发明提供的次黄嘌呤光学探针进行详细的说明，但是不能把它们理解为对本发明保护范围的限定。
I.实验材料和试剂
实施例中主要采用常规的基因工程分子生物学克隆方法和细胞培养以及成像方法等，这些方法是本领域普通技术人员所熟知的，例如：简·罗斯凯姆斯等的《分子生物学实验参考手册》，J.萨姆布鲁克，D.W.拉塞尔著,黄培堂等译：《分子克隆实验指南》(第三版，2002年8月，科学出版社出版，北京)；费雷谢尼等的《动物细胞培养:基本技术指南》(第五版)，章静波，徐存拴等译；J.S.博尼费斯农，M.达索等的《精编细胞生物学实验指南》，章静波等译。
实施例中所用的基于pCDF-cpYFP，pCDF-次黄嘌呤结合蛋白质粒由华东理工大学蛋白质实验室构建，pCDF质粒载体购自Invitrogen公司。所有用于PCR的引物均由上海捷瑞生物工程技术有限公司合成、纯化和经质谱法鉴定正确。实施例中构建的表达质粒都经过序列测定，序列测定由华大基因公司和杰李测序公司完成。各实施例所用的Taq DNA聚合酶购自东盛生物，pfu DNA聚合酶购自天根生化科技(北京)有限公司，primeSTAR DNA聚合酶购自TaKaRa公司，三种聚合酶购买时都附带赠送对应聚合酶缓冲液和dNTP。BamHI、BglII、HindIII、NdeI、XhoI、EcoRI、SpeI等限制性内切酶、T4连接酶、T4磷酸化酶(T4 PNK)购自Fermentas公司，购买时附带有相对应的缓冲液等。转染试剂Lip2000 Kit购于Invitrogen公司。次黄嘌呤等嘌呤类似物均购自Aladdin公司。除非特别声明，无机盐类等化学试剂均购自Sigma-Aldrich公司。HEPES盐，氨苄青霉素(Amp)和嘌呤霉素购自Ameresco公司。96孔检测黑板、384孔荧光检测黑板购自Grenier公司。
实施例中所用的DNA纯化试剂盒购自BBI公司(加拿大)，普通质粒小抽试剂盒购自天根生化科技(北京)有限公司。克隆菌株Mach1购自Invitrogen公司。镍柱亲和层析柱和脱盐柱填料均来自GE healthcare公司。
实施例中用到的主要仪器包括：Biotek Synergy 2多功能酶标仪(美国Bio-Tek公司)，X-15R高速冷冻离心机(美国Beckman公司)，Microfuge22R台式高速冷冻离心机(美国Beckman公司)，PCR扩增仪(德国Biometra公司)，超声破碎仪(宁波新芝公司)，核酸电泳仪(申能博彩公司)，荧光分光光度计(美国Varian公司)，CO2恒温细胞培养箱(SANYO)，倒置荧光显微镜(日本尼康公司)。
II.分子生物学方法和细胞实验方法
II.1聚合酶链式反应(PCR)：
1.目的片段扩增PCR：
该方法主要用于基因片段扩增和菌落PCR鉴定阳性克隆。所述PCR扩增的反应体系如下：模板序列0.5-1μL，正向引物(25μM)0.5μL，反向引物(25μM)0.5μL，10×pfu缓冲液5μL，pfu DNA聚合酶0.5μL，dNTP(10mM)1μL，灭菌超纯水(ddH2O)41.5-42μL，总体积50μL。PCR扩增程序如下：95℃变性2-10分钟，30轮循环(94-96℃持续30-45秒，50-65℃持续30-45秒，72℃持续一定时间(600bp/min))，72℃延伸10分钟。
2.长片段(>2500bp)扩增PCR：
本发明中使用的长片段扩增，主要是反向PCR扩增载体，在下述实施例中用于获得定点突变的一种技术。在变异部位设计反向PCR引物，其中一条引物的5’端包含变异的核苷酸序列。扩增后的产物就含有相应的突变位点。长片段扩增PCR反应体系如下：模板序列(10pg-1ng)1μL，正向引物(25μM)0.5μL，反向引物(25μM)0.5μL，5×PrimerSTAR缓冲液10μL，PrimerSTAR DNA聚合酶0.5μL，dNTP(2.5mM)4μL，灭菌超纯水(ddH2O)33.5μL，总体积50μL。PCR扩增程序如下：95℃变性5分钟，30轮循环(98℃持续10秒，50-68℃持续5-15秒，72℃持续一定时间(1000bp/min))，72℃延伸10分钟；或者95℃变性5分钟，30轮循环(98℃持续10秒，68℃持续一定时间(1000bp/min))，72℃延伸10分钟。
II.2核酸内切酶酶切反应：
对质粒载体进行双酶切的体系如下：质粒载体20μL(约1.5μg)，10×缓冲液5μL，限制性内切酶11-2μL，限制性内切酶21-2μL，用灭菌超纯水补至总体积50μL。反应条件37℃，1-7小时。
II.3DNA片段5’端磷酸化反应
从微生物中抽提出的质粒或者基因组末端都含有磷酸基团，而PCR产物没有，故需对PCR产物的5’端碱基进行磷酸基团加成反应，只有末端含有磷酸基团DNA分子才能发生连接反应。磷酸化反应体系如下：PCR产物片段DNA序列5-8μL，10×T4连接酶缓冲液1μL，T4多聚核苷酸激酶(T4 PNK)1μL，灭菌超纯水0-3μL，总体积10μL。反应条件37℃，30分钟-2小时后72℃灭活20分钟。
II.4目的片段和载体的连接反应
不同的片段和载体之间的连接方法有所差异，本发明中使用了三种连接方法
1.平末端短片段和线性化载体的平末端连接
该方法的原理是PCR获得的平末端产物在T4 PNK作用下对DNA片段的5’末端进行磷酸化反应后，与线性化的载体在PEG4000和T4 DNA连接酶的作用下连接获得重组质粒。同源重组连接体系如下：T4 PNK处理的DNA片段4μL，线性化载体片段4μL，PEG4000 1μL，10×T4连接酶缓冲液1μL，T4 DNA连接酶1μL，总计10μL。反应条件22℃，30分钟。
2.含有粘性末端的DNA片段和含有粘性末端载体片段的连接
通过限制性内切酶切割的DNA片段通常会产生突出的粘性末端，因此可以和含有序列互补的粘性末端载体片段连接，形成重组质粒。连接反应体系如下：酶切后的PCR产物片段DNA 1-7μL，酶切后的质粒0.5-7μL，10×T4连接酶缓冲液1μL，T4 DNA连接酶1μL，灭菌超纯水补至总体积10μL。反应条件16℃，4-8小时。
3.反向PCR引入定点突变后5’端磷酸化的DNA片段产物自身环化的连接反应
将5’端磷酸化的DNA片段通过自身环化连接反应将线性化载体的3’端和5’端连接反应得到重组质粒。自身环化连接反应体系如下：磷酸化反应体系10μL，T4连接酶(5U/μL)0.5μL，总体积10.5μL。反应条件16℃，4-16小时。
II.5感受态细胞的制备与转化
感受态细胞的制备：
1.挑取单菌落(如Mach1)接种于5mL LB培养基中，37℃摇床过夜。
2.取0.5-1mL过夜培养的菌液转种到50mL LB培养基中，37℃，220rpm培养3至5小时，直到OD600达到0.5。
3.冰浴预冷细胞2小时。
4. 4℃，4000rpm离心10分钟。
5.弃上清，用5mL预冷的缓冲液重悬细胞，待均匀后再加入重悬缓冲液至终体积为50mL。
6.冰浴45分钟。
7. 4℃ 4000rpm离心10分钟，用5mL冰预冷的储存缓冲液重悬细菌。
8.每个EP管中放100μL菌液，-80℃或液氮冻存。
重悬缓冲液:CaCl2(100mM)、MgCl2(70mM)、NaAc(40mM)
储存缓冲液:0.5mL DMSO、1.9mL 80％甘油、1mL 10×CaCl2(1M)、1mL10×MgCl2(700mM)、1mL 10×NaAc(400mM)、4.6mL ddH2O
感受态细胞的转化：
1.取100μL感受态细胞于冰浴上融化。
2.加入适当体积的连接产物，轻轻吹打混匀，冰浴30分钟。通常加入的连接产物的体积少于感受态细胞体积的1/10。
3.将菌液放入42℃水浴中热激90秒，迅速转移至冰浴中放置5分钟。
4.加入500μL LB，于37℃恒温摇床上200rpm培养1小时。
5.将菌液4000rpm离心3分钟，留200μL上清将菌体吹匀，均匀涂布于含适当抗生素的琼脂平板表面，平板于37℃恒温培养箱内倒置过夜。
II.6蛋白质的表达，纯化和荧光检测
1.将表达载体(例如以pCDF为基础的次黄嘌呤光学探针表达载体)转化到BL21(DE3)细胞中，倒置培养过夜，从平板上挑取克隆到250ml锥形瓶中，置于37℃摇床，220rpm培养至OD＝0.4-0.8，加入1/1000(v/v)的IPTG(1M)，18℃诱导表达24-36小时。
2.诱导表达完成后，4000rpm，30分钟离心收菌，加入50mM的磷酸盐缓冲液重悬菌体沉淀，超声破碎至菌体澄清。9600rpm，4℃离心20分钟。
3.离心上清通过自装的镍柱亲和层析柱纯化获得蛋白，镍柱亲和层析后的蛋白再通过自装的脱盐柱获得溶解在100mM HEPES缓冲液(pH 7.4)中的蛋白。
4.纯化的蛋白经过SDS-PAGE鉴定后，使用测定缓冲液(100mM HEPES，100mM NaCl，pH 7.4)稀释探针成终浓度为0.2-5μM的蛋白溶液。用测定缓冲液(100mM HEPES，100mMNaCl，pH 7.4)将次黄嘌呤配制成终浓度为50mM的储液。
5.取100μl 1μM的蛋白溶液，37℃温育10分钟，加入次黄嘌呤滴定，测定蛋白的420nm光激发后528nm发射和485nm光激发后528nm发射的荧光强度。对样品的荧光激发、发射测定利用多功能荧光酶标仪完成。
6.取100μl 1μM的蛋白溶液，37℃温育10分钟，加入次黄嘌呤，测定蛋白的吸收光谱和荧光光谱。对样品的吸收光谱和荧光光谱的测定通过分光光度计和荧光分光光度计完成。
II.7哺乳动物细胞的转染和荧光检测
1.将pCDNA3.1+为基础的次黄嘌呤光学探针质粒通过转染试剂Lipofectamine2000(Invitrogen)转染到HEK293中，置于37℃，5％CO2的细胞培养箱中培养。待外源基因充分表达24～36h后进行荧光检测。
2.诱导表达完成后，将贴壁的HEK293细胞，用PBS冲洗三次，置于HBSS溶液中分别进行荧光显微镜和酶标仪检测。
实施例1：次黄嘌呤结合蛋白质粒
通过PCR扩增大肠杆菌基因中的PurR基因片段(编码SEQ ID NO:1的第58-341位氨基酸)，PCR产物凝胶电泳后回收后用BamHI和XhoI酶切，同时对pCDF载体进行相应的双酶切。用T4 DNA连接酶连接后，用产物转化DH5α，转化的DH5α涂布于LB平板(链霉素100ug/mL)，置于37℃培养过夜。将生长DH5α转化子进行质粒抽提后，进行PCR鉴定。阳性质粒经过测序正确后进行后续的质粒构建。
实施例2：不同插入位点的cpYFP光学探针的表达和检测
本实施例中，以pCDF-PurR为基础选择了下述位点插入cpYFP(SEQ ID NO:2)，得到相应pCDF-PurR-cpYFP质粒：66/67、66/68、66/69、66/70、66/71、66/72、67/68、67/69、67/70、67/71、67/72、68/69、68/70、68/71、68/72、69/70、69/71、69/72、70/71、70/72、71/72、97/98、97/99、97/100、97/101、98/99、98/100、98/101、99/100、99/101、100/101、123/124、123/125、123/126、123/127、123/128、124/125、124/126、124/127、124/128、125/126、125/127、125/128、126/127、126/128、127/128、145/146、145/147、145/148、145/149、146/147、146/148、146/149、147/148、147/149、147/150、147/151、147/152、147/153、147/154、147/155、147/156、148/149、148/150、148/151、148/152、148/153、148/154、148/155、148/156、149/150、149/151、149/152、149/153、149/154、149/155、149/156、150/151、150/152、150/153、150/154、150/155、150/156、151/152、151/153、151/154、151/155、151/156、152/153、152/154、152/155、152/156、153/154、153/155、153/156、154/155、154/156、155/156、160/161、160/162,160/163、161/162、161/163、162/163、187/188、187/189、187/190、187/191、188/189、188/190、188/191、189/190、189/191、190/191、210/211、210/212、210/213、210/214、210/215、211/212、211/213、211/214、211/215、212/213、212/214、212/215、213/214、213/215、214/215、220/221、220/222、220/223、221/222、221/223、222/223、237/238、237/239、238/239、246/247、246/248、247/248、275/276、275/277、275/278、275/279、276/277、276/278、276/279、277/278、277/279、278/279、291/292、291/293、291/294、292/293、292/294、293/294、312/313、312/314、312/315、312/316、312/317、313/314,313/315、313/316、313/317、314/315、314/316、314/317、315/316、315/317、316/317、321/322、321/323、321/324、322/323、322/324、323/324。
利用PCR技术扩增获得cpYFP的DNA片段，和pCDF-PurR线性化载体，其5’和3’最末端分别带有和cpYFP两末端对应的完全一致的序列(15bp～20bp)。将线性化的pCDF-PurR和cpYFP片段在Hieff Clone Enzyme的作用下发生同源重组。产物转化DH5α，转化的DH5α涂布于LB平板(链霉素100ug/mL)，置于37℃培养过夜。PCR鉴定的阳性克隆抽质粒后测序。由杰李测序公司完成测序。
经过测序正确后，将重组质粒转化到BL21(DE3)中诱导表达，并纯化蛋白质，通过SDS-PAGE电泳大小在43Kda附近。该大小符合pCDF-PurR-cpYFP表达出的含His-tag纯化标签的PurR-cpYFP融合蛋白质的大小。结果如图1所示。
用表达PurR-cpYFP融合蛋白质的大肠杆菌的破碎上清进行次黄嘌呤响应筛选，将含有1mM次黄嘌呤的融合荧光蛋白质的检测信号除以无次黄嘌呤的融合荧光蛋白质的检测信号。结果如表1所示，检测结果显示对次黄嘌呤响应超过1.2倍的光学探针有7个，在147/149(SEQ ID NO:10)，149/150(SEQ ID NO:11)，149/151(SEQ ID NO:12)，150/151(SEQ IDNO:13)，312/315(SEQ ID NO:14)，313/315(SEQ ID NO:15)，314/315(SEQ ID NO:16)位点。
表1
插入位点 变化倍数 插入位点 变化倍数 插入位点 变化倍数 插入位点 变化倍数 插入位点 变化倍数
66/67 1.09 123/128 1.10 149/150 1.80 187/190 1.01 275/277 1.06
66/68 1.04 124/125 1.12 149/151 1.30 187/191 1.04 275/278 1.05
66/69 1.04 124/126 1.05 149/152 1.00 188/189 1.05 275/279 1.07
66/70 1.07 124/127 1.05 149/153 1.00 188/190 1.10 276/277 1.01
66/71 1.14 124/128 1.04 149/154 1.06 188/191 1.09 276/278 1.03
66/72 1.14 125/126 1.03 149/155 0.97 189/190 1.08 276/279 0.96
67/68 1.00 125/127 1.01 149/156 0.97 189/191 0.96 277/278 1.12
67/69 1.09 125/128 1.10 150/151 1.30 190/191 0.99 277/279 1.07
67/70 1.11 126/127 0.95 150/152 1.02 210/211 0.98 278/279 1.01
67/71 1.03 126/128 1.11 150/153 0.98 210/212 1.03 291/292 1.09
67/72 1.09 127/128 1.03 150/154 1.03 210/213 1.07 291/293 1.03
68/69 1.01 145/146 0.89 150/155 1.00 210/214 1.05 291/294 1.03
68/70 1.08 145/147 1.02 150/156 1.06 210/215 1.06 292/293 1.01
68/71 1.01 145/148 0.87 151/152 1.02 211/212 1.04 292/294 1.05
68/72 1.15 145/149 0.98 151/153 1.00 211/213 1.12 293/294 1.03
69/70 1.01 146/147 1.03 151/154 0.99 211/214 1.01 312/313 1.02
69/71 1.06 146/148 1.09 151/155 0.99 211/215 1.02 312/314 1.02
69/72 1.13 146/149 0.97 151/156 1.00 212/213 1.06 312/315 1.20
70/71 1.11 147/148 0.92 152/153 1.01 212/214 1.12 312/316 1.01
70/72 1.14 147/149 1.30 152/154 1.07 212/215 1.07 312/317 1.14
71/72 1.02 147/150 1.09 152/155 1.02 213/214 1.03 313/314 1.01
97/98 0.94 147/151 0.99 152/156 1.01 213/215 1.18 313/315 1.30
97/99 0.93 147/152 1.08 153/154 0.99 214/215 0.98 313/316 1.01
97/100 0.98 147/153 1.01 153/155 1.16 220/221 1.10 313/317 1.18
97/101 1.00 147/154 0.98 153/156 1.00 220/222 1.05 314/315 1.50
98/99 1.06 147/155 0.97 154/155 1.03 220/223 1.07 314/316 1.01
98/100 1.06 147/156 0.96 154/156 1.10 221/222 1.07 314317 1.08
98/101 0.94 148/149 1.09 155/156 1.04 221/223 1.01 315/316 1.14
99/100 0.98 148/150 1.03 160/161 1.03 222/223 1.06 315/317 1.08
99/101 1.06 148/151 0.97 160/162 1.04 237/238 1.06 316/317 1.05
100/101 1.02 148/152 1.01 160/163 1.03 237/239 1.12 321/322 1.03
123/124 1.04 148/153 1.01 161/162 1.07 238/239 1.07 321/323 1.10
123/125 1.09 148/154 0.96 161/163 1.07 246/247 1.01 321/324 1.04
123/126 1.08 148/155 1.09 162/163 1.02 246/248 1.05 322/323 1.10
123/127 1.18 148/156 0.97 187/188 1.01 247/248 1.06 322/324 1.04
cpYFP 1.00 187/189 1.09 275/276 1.07 323/324 1.08
实施例3：不同插入位点的cpGFP光学探针的表达和检测
按照实施例2中的方法将cpYFP替换为cpGFP(SEQ ID NO：6)，构建次黄嘌呤绿色荧光蛋白荧光探针。如表3所示，检测结果显示对次黄嘌呤响应超过1.2倍的光学探针有在147/149，149/150，149/151和150/151位点或者其家族蛋白的对应氨基酸位点实施插入的光学探针。
表2
插入位点 变化倍数 插入位点 变化倍数 插入位点 变化倍数 插入位点 变化倍数 插入位点 变化倍数
66/67 0.96 123/128 1.01 149/150 1.09 187/190 1.07 275/277 1.08
66/68 1.05 124/125 0.98 149/151 1.33 187/191 1.05 275/278 1.01
66/69 1.12 124/126 1.06 149/152 1.00 188/189 1.07 275/279 0.99
66/70 1.03 124/127 1.09 149/153 1.06 188/190 1.01 276/277 1.01
66/71 1.07 124/128 1.01 149/154 0.99 188/191 1.07 276/278 1.03
66/72 1.05 125/126 1.06 149/155 0.97 189/190 0.98 276/279 1.07
67/68 1.06 125/127 1.01 149/156 1.08 189/191 0.96 277/278 1.05
67/69 0.98 125/128 1.15 150/151 1.50 190/191 0.99 277/279 1.07
67/70 1.03 126/127 1.09 150/152 0.98 210/211 0.98 278/279 1.01
67/71 1.09 126/128 1.06 150/153 1.07 210/212 1.03 291/292 1.09
67/72 1.02 127/128 1.13 150/154 1.07 210/213 1.07 291/293 1.03
68/69 1.06 145/146 1.11 150/155 1.06 210/214 1.05 291/294 1.03
68/70 1.12 145/147 1.14 150/156 1.02 210/215 1.06 292/293 1.01
68/71 1.07 145/148 1.02 151/152 1.07 211/212 1.04 292/294 1.05
68/72 1.12 145/149 0.93 151/153 0.99 211/213 1.12 293/294 1.03
69/70 1.18 146/147 1.09 151/154 1.07 211/214 1.01 312/313 1.02
69/71 0.98 146/148 1.12 151/155 0.99 211/215 1.02 312/314 1.02
69/72 1.13 146/149 1.06 151/156 1.02 212/213 1.05 312/315 1.09
70/71 1.11 147/148 1.06 152/153 1.01 212/214 1.12 312/316 1.01
70/72 1.08 147/149 1.25 152/154 1.07 212/215 1.07 312/317 1.10
71/72 1.03 147/150 1.32 152/155 1.06 213/214 1.03 313/314 1.04
97/98 0.98 147/151 1.06 152/156 1.02 213/215 1.18 313/315 1.08
97/99 1.03 147/152 1.02 153/154 1.10 214/215 0.98 313/316 1.01
97/100 1.09 147/153 1.04 153/155 1.04 220/221 1.01 313/317 1.18
97/101 1.00 147/154 1.09 153/156 1.10 220/222 1.18 314/315 1.17
98/99 1.09 147/155 1.08 154/155 1.03 220/223 1.04 314/316 1.08
98/100 1.06 147/156 1.13 154/156 1.02 221/222 1.10 314/317 1.13
98/101 0.94 148/149 1.10 155/156 1.04 221/223 1.04 315/316 1.10
99/100 1.07 148/150 1.03 160/161 1.03 222/223 1.06 315/317 1.03
99/101 1.03 148/151 0.98 160/162 1.04 237/238 1.06 316/317 1.05
100/101 1.04 148/152 1.01 160/163 1.03 237/239 1.12 321/322 1.03
123/124 1.03 148/153 1.01 161/162 1.07 238/239 1.08 321/323 1.09
123/125 1.01 148/154 0.96 161/163 1.13 246/247 1.01 321/324 1.00
123/126 1.07 148/155 1.09 162/163 1.03 246/248 1.05 322/323 1.15
123/127 1.02 148/156 0.97 187/188 1.02 247/248 1.13 322/324 1.04
cpGFP 1.00 187/189 1.12 275/276 1.03 323/324 1.08
实施例4：不同插入位点的cpBFP光学探针的表达和检测
按照实施例2中的方法将cpYFP替换为cpBFP(SEQ ID NO:7)，构建次黄嘌呤蓝色荧光蛋白荧光探针。如表3所示，检测结果显示对次黄嘌呤响应超过1.3倍的光学探针有在150/151，312/315，313/315和314/315位点或者其家族蛋白的对应氨基酸位点实施插入的光学探针。
表3
插入位点 变化倍数 插入位点 变化倍数 插入位点 变化倍数 插入位点 变化倍数 插入位点 变化倍数
66/67 1.01 123/128 1.10 149/150 1.12 187/190 1.01 275/277 1.06
66/68 1.06 124/125 1.12 149/151 1.04 187/191 1.04 275/278 1.05
66/69 0.94 124/126 1.05 149/152 1.12 188/189 1.05 275/279 1.07
66/70 1.06 124/127 1.05 149/153 1.06 188/190 1.10 276/277 1.01
66/71 0.94 124/128 1.04 149/154 1.06 188/191 1.09 276/278 1.03
66/72 1.11 125/126 1.03 149/155 0.94 189/190 1.08 276/279 0.96
67/68 1.06 125/127 1.01 149/156 1.09 189/191 0.96 277/278 1.12
67/69 1.02 125/128 1.10 150/151 1.50 190/191 0.99 277/279 1.07
67/70 0.93 126/127 0.95 150/152 1.02 210/211 0.98 278/279 1.01
67/71 1.09 126/128 1.11 150/153 0.98 210/212 1.03 291/292 1.09
67/72 1.12 127/128 1.03 150/154 1.03 210/213 1.07 291/293 1.03
68/69 1.06 145/146 0.89 150/155 1.00 210/214 1.05 291/294 1.03
68/70 1.06 145/147 1.02 150/156 1.06 210/215 1.06 292/293 1.01
68/71 0.94 145/148 0.87 151/152 1.02 211/212 1.04 292/294 1.05
68/72 1.09 145/149 0.98 151/153 1.04 211/213 1.12 293/294 1.03
69/70 1.01 146/147 1.03 151/154 1.03 211/214 1.01 312/313 1.02
69/71 1.06 146/148 1.05 151/155 0.99 211/215 1.02 312/314 1.02
69/72 1.13 146/149 1.12 151/156 1.00 212/213 1.01 312/315 1.40
70/71 1.11 147/148 1.07 152/153 1.03 212/214 1.18 312/316 1.01
70/72 1.14 147/149 1.03 152/154 1.07 212/215 1.09 312/317 1.14
71/72 1.06 147/150 1.18 152/155 1.02 213/214 1.01 313/314 1.01
97/98 1.03 147/151 0.98 152/156 1.01 213/215 1.08 313/315 1.30
97/99 1.18 147/152 1.01 153/154 0.99 214/215 1.14 313/316 1.01
97/100 0.98 147/153 1.18 153/155 1.16 220/221 1.08 313/317 1.18
97/101 1.01 147/154 1.04 153/156 1.00 220/222 1.05 314/315 1.40
98/99 1.06 147/155 0.97 154/155 1.03 220/223 1.07 314/316 1.01
98/100 1.06 147/156 0.96 154/156 1.10 221/222 1.07 314/317 1.08
98/101 0.94 148/149 1.09 155/156 1.04 221/223 1.01 315/316 1.14
99/100 0.98 148/150 1.03 160/161 1.03 222/223 1.06 315/317 1.08
99/101 1.06 148/151 0.97 160/162 1.04 237/238 1.06 316/317 1.05
100/101 1.02 148/152 1.01 160/163 1.03 237/239 1.12 321/322 1.03
123/124 1.04 148/153 1.01 161/162 1.07 238/239 1.07 321/323 1.10
123/125 1.09 148/154 0.96 161/163 1.07 246/247 1.01 321/324 1.03
123/126 1.08 148/155 1.09 162/163 1.02 246/248 1.05 322/323 1.18
123/127 1.18 148/156 0.97 187/188 1.01 247/248 1.06 322/324 0.98
cpBFP 1.00 187/189 1.09 275/276 1.07 323/324 1.01
实施例5：不同插入位点的cpmApple光学探针的表达和检测
按照实施例2中的方法将cpYFP替换为cpmApple(SEQ ID NO:9)，构建次黄嘌呤红色荧光蛋白荧光探针。如表4所示，检测结果显示对次黄嘌呤响应超过1.2倍的光学探针有在312/315，313/315和314/315位点或者其家族蛋白的对应氨基酸位点实施插入的光学探针；
表4
插入位点 变化倍数 插入位点 变化倍数 插入位点 变化倍数 插入位点 变化倍数 插入位点 变化倍数
66/67 1.03 124/125 1.08 149/151 1.08 187/190 1.06 275/277 1.11
66/68 1.01 124/126 1.07 149/152 1.05 187/191 1.09 275/278 1.08
66/69 1.10 124/127 0.94 149/153 1.02 188/189 1.12 275/279 0.98
66/70 0.95 124/128 1.06 149/154 0.98 188/190 1.01 276/277 1.03
66/71 1.01 125/126 0.94 149/155 0.95 188/191 1.03 276/278 0.99
66/72 1.03 125/127 1.11 149/156 0.97 189/190 0.96 276/279 1.12
67/68 0.89 125/128 1.06 150/151 0.98 189/191 1.09 277/278 1.03
67/69 1.02 126/127 0.98 150/152 1.02 190/191 1.07 277/279 1.06
67/70 1.18 126/128 0.93 150/153 0.99 210/211 1.01 278/279 0.97
67/71 0.98 127/128 0.99 150/154 1.03 210/212 1.02 291/292 1.13
67/72 1.03 145/146 1.12 150/155 1.00 210/213 1.03 291/293 1.01
68/69 1.05 145/147 1.06 150/156 1.07 210/214 1.07 291/294 0.99
68/70 1.12 145/148 1.06 151/152 1.02 210/215 1.12 292/293 1.01
68/71 0.87 145/149 0.94 151/153 1.11 211/212 1.05 292/294 1.11
68/72 0.98 146/147 1.13 151/154 0.99 211/213 1.12 293/294 1.03
69/70 1.01 146/148 1.01 151/155 1.03 211/214 1.15 312/313 0.93
69/71 1.18 146/149 1.06 151/156 1.00 211/215 1.16 312/314 1.02
69/72 1.09 147/148 1.14 152/153 1.08 212/213 1.05 312/315 1.23
70/71 1.01 147/149 1.11 152/154 1.07 212/214 1.12 312/316 1.01
70/72 1.08 147/150 1.09 152/155 1.03 212/215 1.07 312/317 1.14
97/98 1.14 147/151 0.99 152/156 1.01 213/214 1.03 313/314 1.01
97/99 1.08 147/152 1.08 153/154 0.99 213/215 1.14 313/315 1.33
97/100 1.05 147/153 1.01 153/155 1.16 214/215 0.98 313/316 1.01
97/101 1.07 147/154 0.98 153/156 1.00 220/221 1.10 313/317 1.18
98/99 1.07 147/155 1.18 154/155 1.03 220/222 1.05 314/315 1.36
98/100 0.98 147/156 1.09 154/156 1.10 220/223 1.07 314/316 1.01
98/101 1.06 148/149 1.06 155/156 1.04 221/222 1.07 314/317 1.08
99/100 1.01 148/150 1.03 160/161 1.14 221/223 1.01 315/316 1.10
99/101 1.06 148/151 1.14 160/162 1.13 222/223 1.06 315/317 1.08
100/101 1.02 148/152 1.01 160/163 1.03 237/238 1.08 316/317 1.05
123/124 1.01 148/153 1.01 161/162 1.14 237/239 1.12 321/322 1.03
123/125 1.09 148/154 1.16 161/163 1.03 238/239 0.89 321/323 1.10
123/126 1.08 148/155 1.09 162/163 1.02 246/247 0.98 321/324 1.04
123/127 1.18 148/156 0.97 187/188 1.13 246/248 1.05 322/323 0.89
123/128 0.98 149/150 0.99 187/189 1.14 247/248 1.07 322/324 1.04
cpmApple 1.00 275/276 1.14 323/324 0.98
实施例6：突变的cpYFP光学探针的表达和检测
对于实施例2中所得的对次黄嘌呤响应超过1.2倍的光学探针，即在147/149，149/150，149/151，150/151，312/315，313/315，314/315位点实施插入的7种光学探针基础上通过反向PCR线性化探针，引物中引入突变位点的序列，对得到的PCR产物在Hieff CloneEnzyme的作用下进行同源重组，建立突变库。将突变库重组质粒转化到BL21(DE3)中诱导表达，用表达探针蛋白的大肠杆菌的破碎上清进行次黄嘌呤响应筛选，将含有1mM次黄嘌呤的融合荧光蛋白质的检测信号除以无次黄嘌呤的融合荧光蛋白质的检测信号。结果如表5所示，检测结果显示对次黄嘌呤响应超过或等于2倍的光学探针如下所示。
表5
在150/151-151V152L的基础上对PurR蛋白进行进一步突变，然后通过反向PCR线性化探针，引物中引入突变位点的序列，对得到的PCR产物进行同源重组，建立突变库。将突变库重组质粒转化到BL21(DE3)中诱导表达，用表达探针蛋白的大肠杆菌的破碎上清进行次黄嘌呤响应筛选，将含有1mM次黄嘌呤的融合荧光蛋白质的检测信号除以无次黄嘌呤的融合荧光蛋白质的检测信号。结果表6所示，检测结果显示对次黄嘌呤响应超过或等于2倍的光学探针如下所示。
表6
实施例7：光学探针突变体的性能
将实施例6所述的表5中部分次黄嘌呤光学探针以及表6中所有的次黄嘌呤光学探针分别进行浓度梯度(0～5mM)的次黄嘌呤检测。对探针处理10分钟后，检测420nm激发528nm发射处荧光强度和485nm激发528nm发射处荧光强度比值的变化。探针滴定结果如图2所示，结果表明不同突变体对次黄嘌呤亲和力不同。
对表5和表6中的次黄嘌呤探针进行特异性检测，分别与次黄嘌呤结构类似物，黄嘌呤，腺嘌呤，鸟嘌呤以及尿酸进行反应性检测，结果表明具有良好的特异性，如图3所示。
实施例8：光学探针的亚细胞器定位和光学探针在亚细胞器内的性能
本实施例中，使用不同的定位信号肽与光学探针PurR-W147R/K151V/A152L/E149R/A150I cpYFP-Y1L融合，将光学探针定位到不同的细胞器中。用融合不同定位信号肽的光学探针质粒转染HEK293细胞36小时后，使用PBS冲洗，置于HBSS溶液中使用倒置荧光显微镜进行FITC通道下进行荧光检测。结果如图4所示。次黄嘌呤光学探针通过与不同的特异定位信号肽融合能够定位到包括细胞浆、线粒体、细胞核、核排阻、细胞外膜、内质网中。不同的亚细胞结构中都显示有荧光，并且荧光的分布和强度各不相同。
用胞浆表达的光学探针质粒转染HEK293细胞36小时后，使用PBS冲洗，置于HBSS溶液中，检测30min时间段内420nm激发528nm发射处荧光强度和485nm激发528nm发射处荧光强度比值的变化。结果如图5所示。添加1mM次黄嘌呤，继续检测30分钟。添加次黄嘌呤的样品的485/420逐渐增加，最高可以达到初始值的28倍，而不添加次黄嘌呤的对照组的485/420基本保持不变。
实施例9：在活细胞中基于光学探针进行高通量化合物筛选
本实施例中，我们使用胞浆表达PurR-W147R/K151V/A152L/E149R/A150I cpYFP-Y1L的HEK293细胞进行高通量化合物筛选。
经转染的HEK293细胞使用PBS冲洗，置于HBSS溶液中(无次黄嘌呤)处理1小时，然后使用10μM的化合物处理1小时。各样品中分别滴加次黄嘌呤。使用酶标仪记录420nm激发528nm发射处荧光强度和485nm激发528nm发射处荧光强度比值变化。以未用任何化合物处理的样品作为对照进行标准化。结果如图6所示。在使用的2000种化合物中，绝大部分的化合物对次黄嘌呤进入细胞影响极小。有11种化合物能够提高细胞对次黄嘌呤的摄取能力，另外有8种化合物能够明显降低细胞对次黄嘌呤的摄取。
实施例10:光学探针定量检测血液中的次黄嘌呤
在本实施中，使用纯化的PurR-W147R/K151V/A152L/E149S/A150V对小鼠和人的血液上清中的次黄嘌呤进行分析。
将PurR-W147R/K151V/A152L/E149S/A150V与稀释的血液上清混合处理10分钟后，使用酶标仪检测420nm激发528nm发射处荧光强度和485nm激发528nm发射处荧光强度比值。结果如图7所示，小鼠血液中的次黄嘌呤含量在50μM左右，人血液中的次黄嘌呤含量在6μM左右。
由以上实施例可知，本发明提供的次黄嘌呤光学探针，蛋白分子量相对较小且易于成熟，荧光动态变化大，特异性好，并且能够通过基因操作的方法在细胞中表达，可在细胞内外实时定位、定量检测次黄嘌呤；并且能够进行高通量的化合物筛选。
其它实施方式
本说明书描述了许多实施方式。然而应理解，本领域技术人员通过阅读本说明书获知的不背离本发明的构思和范围的各种改进，也应包括在所附权利要求书的范围内。
SEQUENCE LISTING
<110> 华东理工大学
<120> 一种次黄嘌呤光学探针及其制备方法和应用
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Met Ala Thr Ile Lys Asp Val Ala Lys Arg Ala Asn Val Ser Thr Thr
1 5 10 15
Thr Val Ser His Val Ile Asn Lys Thr Arg Phe Val Ala Glu Glu Thr
20 25 30
Arg Asn Ala Val Trp Ala Ala Ile Lys Glu Leu His Tyr Ser Pro Ser
35 40 45
Ala Val Ala Arg Ser Leu Lys Val Asn His Thr Lys Ser Ile Gly Leu
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Leu Ala Thr Ser Ser Glu Ala Ala Tyr Phe Ala Glu Ile Ile Glu Ala
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Val Glu Lys Asn Cys Phe Gln Lys Gly Tyr Thr Leu Ile Leu Gly Asn
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Ala Trp Asn Asn Leu Glu Lys Gln Arg Ala Tyr Leu Ser Met Met Ala
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Gln Lys Arg Val Asp Gly Leu Leu Val Met Cys Ser Glu Tyr Pro Glu
115 120 125
Pro Leu Leu Ala Met Leu Glu Glu Tyr Arg His Ile Pro Met Val Val
130 135 140
Met Asp Trp Gly Glu Ala Lys Ala Asp Phe Thr Asp Ala Val Ile Asp
145 150 155 160
Asn Ala Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg
165 170 175
Gly His Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr
180 185 190
Gly Ala Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met
195 200 205
Ile Lys Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu
210 215 220
Ser Gly Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro
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Thr Ala Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys
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Ala Ala Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile
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Gly Tyr Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr
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Thr Ile His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met
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Leu Leu Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu
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Val His Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe
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Arg Asp Tyr Arg Arg
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Tyr Asn Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly
1 5 10 15
Ile Lys Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val
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Gln Leu Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro
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Val Leu Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser
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Lys Asp Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val
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Thr Ala Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp
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Gly Gly Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly
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Val Val Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys
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Phe Ser Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu
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Thr Leu Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro
145 150 155 160
Thr Leu Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr
165 170 175
Pro Asp His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu
180 185 190
Gly Tyr Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr
195 200 205
Lys Thr Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg
210 215 220
Ile Glu Leu Lys Gly Ile Asp Phe Lys Glu Asp Gly Asn Ile Leu Gly
225 230 235 240
His Lys Leu Glu Tyr Asn
245
<210> 3
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<212> PRT
<213> Artificial Sequence
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Val Ser Glu Arg Met Tyr Pro Glu Asp Gly Val Leu Lys Ser Glu Ile
1 5 10 15
Lys Lys Gly Leu Arg Leu Lys Asp Gly Gly His Tyr Ala Ala Glu Val
20 25 30
Lys Thr Thr Tyr Lys Ala Lys Lys Pro Val Gln Leu Pro Gly Ala Tyr
35 40 45
Ile Val Asp Ile Lys Leu Asp Ile Val Ser His Asn Glu Asp Tyr Thr
50 55 60
Ile Val Glu Gln Cys Glu Arg Ala Glu Gly Arg His Pro Thr Gly Gly
65 70 75 80
Arg Asp Glu Leu Tyr Lys Gly Gly Thr Gly Gly Ser Leu Val Ser Lys
85 90 95
Gly Glu Glu Asp Asn Met Ala Ile Ile Lys Glu Phe Met Arg Phe Lys
100 105 110
Val His Met Glu Gly Ser Val Asn Gly His Glu Phe Glu Ile Glu Gly
115 120 125
Glu Gly Glu Gly Arg Pro Tyr Glu Ala Phe Gln Thr Ala Lys Leu Lys
130 135 140
Val Thr Lys Gly Gly Pro Leu Pro Phe Ala Trp Asp Ile Leu Ser Pro
145 150 155 160
Gln Phe Thr Tyr Gly Ser Lys Ala Tyr Ile Lys His Pro Ala Asp Ile
165 170 175
Pro Asp Tyr Phe Lys Leu Ser Phe Pro Glu Gly Phe Arg Trp Glu Arg
180 185 190
Val Met Asn Phe Glu Asp Gly Gly Ile Ile His Val Asn Gln Asp Ser
195 200 205
Ser Leu Gln Asp Gly Val Phe Ile Tyr Lys Val Lys Leu Arg Gly Thr
210 215 220
Asn Phe Pro Pro Asp Gly Pro Val Met Gln Lys Lys Thr Met Gly Trp
225 230 235 240
Glu Ala
<210> 4
<211> 250
<212> PRT
<213> Artificial Sequence
<220>
<223> cpmKate
<400> 4
Met Gly Gly Arg Ser Lys Lys Pro Ala Lys Asn Leu Lys Met Pro Gly
1 5 10 15
Val Tyr Tyr Val Asp Arg Arg Leu Glu Arg Ile Lys Glu Ala Asp Lys
20 25 30
Glu Thr Tyr Val Glu Gln His Glu Val Ala Val Ala Arg Tyr Cys Asp
35 40 45
Leu Pro Ser Lys Leu Gly His Lys Leu Asn Gly Gly Thr Gly Gly Ser
50 55 60
Met Val Ser Lys Gly Glu Glu Leu Ile Lys Glu Asn Met His Met Lys
65 70 75 80
Leu Tyr Met Glu Gly Thr Val Asn Asn His His Phe Lys Cys Thr Ser
85 90 95
Glu Gly Glu Gly Lys Pro Tyr Glu Gly Thr Gln Thr Met Arg Ile Lys
100 105 110
Val Val Glu Gly Gly Pro Leu Pro Phe Ala Phe Asp Ile Leu Ala Thr
115 120 125
Ser Phe Met Tyr Gly Ser Lys Thr Phe Ile Asn His Thr Gln Gly Ile
130 135 140
Pro Asp Phe Phe Lys Gln Ser Phe Pro Glu Gly Phe Thr Trp Glu Arg
145 150 155 160
Val Thr Thr Tyr Glu Asp Gly Gly Val Leu Thr Ala Thr Gln Asp Thr
165 170 175
Ser Leu Gln Asp Gly Cys Leu Ile Tyr Asn Val Lys Ile Arg Gly Val
180 185 190
Asn Phe Pro Ser Asn Gly Pro Val Met Gln Lys Lys Thr Leu Gly Trp
195 200 205
Glu Ala Ser Thr Glu Met Leu Tyr Pro Ala Asp Gly Gly Leu Glu Gly
210 215 220
Arg Ser Asp Met Ala Leu Lys Leu Val Gly Gly Gly His Leu Ile Cys
225 230 235 240
Asn Leu Lys Thr Thr Tyr Arg Ser Lys Lys
245 250
<210> 5
<211> 236
<212> PRT
<213> Artificial Sequence
<220>
<223> mCherry
<400> 5
Met Val Ser Lys Gly Glu Glu Asp Asn Met Ala Ile Ile Lys Glu Phe
1 5 10 15
Met Arg Phe Lys Val His Met Glu Gly Ser Val Asn Gly His Glu Phe
20 25 30
Glu Ile Glu Gly Glu Gly Glu Gly Arg Pro Tyr Glu Gly Thr Gln Thr
35 40 45
Ala Lys Leu Lys Val Thr Lys Gly Gly Pro Leu Pro Phe Ala Trp Asp
50 55 60
Ile Leu Ser Pro Gln Phe Met Tyr Gly Ser Lys Ala Tyr Val Lys His
65 70 75 80
Pro Ala Asp Ile Pro Asp Tyr Leu Lys Leu Ser Phe Pro Glu Gly Phe
85 90 95
Lys Trp Glu Arg Val Met Asn Phe Glu Asp Gly Gly Val Val Thr Val
100 105 110
Thr Gln Asp Ser Ser Leu Gln Asp Gly Glu Phe Ile Tyr Lys Val Lys
115 120 125
Leu Arg Gly Thr Asn Phe Pro Ser Asp Gly Pro Val Met Gln Lys Lys
130 135 140
Thr Met Gly Trp Glu Ala Ser Ser Glu Arg Met Tyr Pro Glu Asp Gly
145 150 155 160
Ala Leu Lys Gly Glu Ile Lys Gln Arg Leu Lys Leu Lys Asp Gly Gly
165 170 175
His Tyr Asp Ala Glu Val Lys Thr Thr Tyr Lys Ala Lys Lys Pro Val
180 185 190
Gln Leu Pro Gly Ala Tyr Asn Val Asn Ile Lys Leu Asp Ile Thr Ser
195 200 205
His Asn Glu Asp Tyr Thr Ile Val Glu Gln Tyr Glu Arg Ala Glu Gly
210 215 220
Arg His Ser Thr Gly Gly Met Asp Glu Leu Tyr Lys
225 230 235
<210> 6
<211> 241
<212> PRT
<213> Artificial Sequence
<220>
<223> cpGFP
<400> 6
Asn Val Tyr Ile Lys Ala Asp Lys Gln Lys Asn Gly Ile Lys Ala Asn
1 5 10 15
Phe Lys Ile Arg His Asn Ile Glu Asp Gly Gly Val Gln Leu Ala Tyr
20 25 30
His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu Leu Pro
35 40 45
Asp Asn His Tyr Leu Ser Val Gln Ser Ile Leu Ser Lys Asp Pro Asn
50 55 60
Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala Ala Gly
65 70 75 80
Ile Thr Leu Gly Met Asp Glu Leu Tyr Lys Gly Gly Thr Gly Gly Ser
85 90 95
Met Val Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val Pro Ile Gln
100 105 110
Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser Val Ser Gly
115 120 125
Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu Lys Phe Ile
130 135 140
Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu Val Thr Thr
145 150 155 160
Leu Thr Tyr Gly Val Gln Cys Phe Ser Arg Tyr Pro Asp His Met Lys
165 170 175
Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr Ile Gln Glu
180 185 190
Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr Arg Ala Glu
195 200 205
Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu Leu Lys Gly
210 215 220
Ile Asp Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys Leu Glu Tyr
225 230 235 240
Asn
<210> 7
<211> 243
<212> PRT
<213> Artificial Sequence
<220>
<223> cpBFP
<400> 7
Asn Val Tyr Ile Lys Ala Asp Lys Gln Lys Asn Gly Ile Lys Ala Asn
1 5 10 15
Phe Lys Ile Arg His Asn Ile Glu Gly Gly Gly Val Gln Leu Ala Tyr
20 25 30
His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu Leu Pro
35 40 45
Asp Asn His Tyr Leu Ser Val Gln Ser Ile Leu Ser Lys Asp Pro Asn
50 55 60
Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala Ala Gly
65 70 75 80
Ile Thr Leu Gly Met Asp Glu Leu Tyr Lys Gly Gly Thr Gly Gly Ser
85 90 95
Glu Ser Met Val Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val Pro
100 105 110
Ile Gln Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser Val
115 120 125
Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu Lys
130 135 140
Phe Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu Val
145 150 155 160
Thr Thr Leu Ser His Gly Val Gln Cys Phe Ser Arg Tyr Pro Asp His
165 170 175
Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Gly Gly Tyr Ile
180 185 190
Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr Arg
195 200 205
Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu Leu
210 215 220
Lys Gly Ile Asp Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys Leu
225 230 235 240
Glu Tyr Asn
<210> 8
<211> 233
<212> PRT
<213> Artificial Sequence
<220>
<223> mKate
<400> 8
Met Ser Glu Leu Ile Thr Glu Asn Met His Met Lys Leu Tyr Met Glu
1 5 10 15
Gly Thr Val Asn Asn His His Phe Lys Cys Thr Ser Glu Gly Glu Gly
20 25 30
Lys Pro Tyr Glu Gly Thr Gln Thr Met Arg Ile Lys Val Val Glu Gly
35 40 45
Gly Pro Leu Pro Phe Ala Phe Asp Ile Leu Ala Thr Ser Phe Met Tyr
50 55 60
Gly Ser Lys Thr Phe Ile Asn His Thr Gln Gly Ile Pro Asp Phe Phe
65 70 75 80
Lys Gln Ser Phe Pro Glu Gly Phe Thr Trp Glu Arg Val Thr Thr Tyr
85 90 95
Glu Asp Gly Gly Val Leu Thr Ala Thr Gln Asp Thr Ser Leu Gln Asp
100 105 110
Gly Cys Leu Ile Tyr Asn Val Lys Ile Arg Gly Val Asn Phe Pro Ser
115 120 125
Asn Gly Pro Val Met Gln Lys Lys Thr Leu Gly Trp Glu Ala Ser Thr
130 135 140
Glu Met Leu Tyr Pro Ala Asp Gly Gly Leu Glu Gly Arg Ala Asp Met
145 150 155 160
Ala Leu Lys Leu Val Gly Gly Gly His Leu Ile Cys Asn Leu Lys Thr
165 170 175
Thr Tyr Arg Ser Lys Lys Pro Ala Lys Asn Leu Lys Met Pro Gly Val
180 185 190
Tyr Tyr Val Asp Arg Arg Leu Glu Arg Ile Lys Glu Ala Asp Lys Glu
195 200 205
Thr Tyr Val Glu Gln His Glu Val Ala Val Ala Arg Tyr Cys Asp Leu
210 215 220
Pro Ser Lys Leu Gly His Lys Leu Asn
225 230
<210> 9
<211> 242
<212> PRT
<213> Artificial Sequence
<220>
<223> cpmApple
<400> 9
Val Ser Glu Arg Met Tyr Pro Glu Asp Gly Ala Leu Lys Ser Glu Ile
1 5 10 15
Lys Lys Gly Leu Arg Leu Lys Asp Gly Gly His Tyr Ala Ala Glu Val
20 25 30
Lys Thr Thr Tyr Lys Ala Lys Lys Pro Val Gln Leu Pro Gly Ala Tyr
35 40 45
Ile Val Asp Ile Lys Leu Asp Ile Val Ser His Asn Glu Asp Tyr Thr
50 55 60
Ile Val Glu Gln Cys Glu Arg Ala Glu Gly Arg His Ser Thr Gly Gly
65 70 75 80
Met Asp Glu Leu Tyr Lys Gly Gly Thr Gly Gly Ser Leu Val Ser Lys
85 90 95
Gly Glu Glu Asp Asn Met Ala Ile Ile Lys Glu Phe Met Arg Phe Lys
100 105 110
Val His Met Glu Gly Ser Val Asn Gly His Glu Phe Glu Ile Glu Gly
115 120 125
Glu Gly Glu Gly Arg Pro Tyr Glu Ala Phe Gln Thr Ala Lys Leu Lys
130 135 140
Val Thr Lys Gly Gly Pro Leu Pro Phe Ala Trp Asp Ile Leu Ser Pro
145 150 155 160
Gln Phe Met Tyr Gly Ser Lys Ala Tyr Ile Lys His Pro Ala Asp Ile
165 170 175
Pro Asp Tyr Phe Lys Leu Ser Phe Pro Glu Gly Phe Arg Trp Glu Arg
180 185 190
Val Met Asn Phe Glu Asp Gly Gly Ile Ile His Val Asn Gln Asp Ser
195 200 205
Ser Leu Gln Asp Gly Val Phe Ile Tyr Lys Val Lys Leu Arg Gly Thr
210 215 220
Asn Phe Pro Pro Asp Gly Pro Val Met Gln Lys Lys Thr Met Gly Trp
225 230 235 240
Glu Ala
<210> 10
<211> 530
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针
<400> 10
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Tyr Asn Ser Asp Asn
85 90 95
Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys Ala Asn Phe
100 105 110
Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu Ala Asp His
115 120 125
Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu Leu Pro Asp
130 135 140
Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp Pro Asn Glu
145 150 155 160
Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala Ala Gly Ile
165 170 175
Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly Ser Gly Gly
180 185 190
Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val Pro Ile Leu
195 200 205
Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser Val Ser Gly
210 215 220
Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu Lys Leu Ile
225 230 235 240
Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu Val Thr Thr
245 250 255
Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp His Met Lys
260 265 270
Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr Val Gln Glu
275 280 285
Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr Arg Ala Glu
290 295 300
Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu Leu Lys Gly
305 310 315 320
Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys Leu Glu Tyr
325 330 335
Asn Glu Ala Lys Ala Asp Phe Thr Asp Ala Val Ile Asp Asn Ala Phe
340 345 350
Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His Arg
355 360 365
Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala Gly
370 375 380
Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys Val
385 390 395 400
Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly Tyr
405 410 415
Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala Val
420 425 430
Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala Asp
435 440 445
Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr Asp
450 455 460
Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile His
465 470 475 480
Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu Asp
485 490 495
Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His Pro
500 505 510
Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp Tyr
515 520 525
Arg Arg
530
<210> 11
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针
<400> 11
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Tyr Asn Ser
85 90 95
Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys Ala
100 105 110
Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu Ala
115 120 125
Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu Leu
130 135 140
Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp Pro
145 150 155 160
Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala Ala
165 170 175
Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly Ser
180 185 190
Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val Pro
195 200 205
Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser Val
210 215 220
Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu Lys
225 230 235 240
Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu Val
245 250 255
Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp His
260 265 270
Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr Val
275 280 285
Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr Arg
290 295 300
Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu Leu
305 310 315 320
Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys Leu
325 330 335
Glu Tyr Asn Ala Lys Ala Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 12
<211> 530
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针
<400> 12
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Tyr Asn Ser
85 90 95
Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys Ala
100 105 110
Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu Ala
115 120 125
Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu Leu
130 135 140
Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp Pro
145 150 155 160
Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala Ala
165 170 175
Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly Ser
180 185 190
Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val Pro
195 200 205
Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser Val
210 215 220
Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu Lys
225 230 235 240
Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu Val
245 250 255
Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp His
260 265 270
Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr Val
275 280 285
Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr Arg
290 295 300
Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu Leu
305 310 315 320
Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys Leu
325 330 335
Glu Tyr Asn Lys Ala Asp Phe Thr Asp Ala Val Ile Asp Asn Ala Phe
340 345 350
Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His Arg
355 360 365
Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala Gly
370 375 380
Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys Val
385 390 395 400
Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly Tyr
405 410 415
Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala Val
420 425 430
Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala Asp
435 440 445
Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr Asp
450 455 460
Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile His
465 470 475 480
Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu Asp
485 490 495
Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His Pro
500 505 510
Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp Tyr
515 520 525
Arg Arg
530
<210> 13
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针
<400> 13
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Lys Ala Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 14
<211> 529
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针
<400> 14
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Lys Ala
85 90 95
Asp Phe Thr Asp Ala Val Ile Asp Asn Ala Phe Glu Gly Gly Tyr Met
100 105 110
Ala Gly Arg Tyr Leu Ile Glu Arg Gly His Arg Glu Ile Gly Val Ile
115 120 125
Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala Gly Arg Leu Ala Gly Phe
130 135 140
Met Lys Ala Met Glu Glu Ala Met Ile Lys Val Pro Glu Ser Trp Ile
145 150 155 160
Val Gln Gly Asp Phe Glu Pro Glu Ser Gly Tyr Arg Ala Met Gln Gln
165 170 175
Ile Leu Ser Gln Pro His Arg Pro Thr Ala Val Phe Cys Gly Gly Asp
180 185 190
Ile Met Ala Met Gly Ala Leu Cys Ala Ala Asp Glu Met Gly Leu Arg
195 200 205
Val Pro Gln Asp Val Ser Leu Ile Gly Tyr Asp Asn Val Arg Asn Ala
210 215 220
Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile His Gln Pro Lys Asp Ser
225 230 235 240
Leu Gly Glu Thr Ala Phe Asn Met Leu Leu Asp Arg Ile Val Asn Lys
245 250 255
Tyr Asn Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly
260 265 270
Ile Lys Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val
275 280 285
Gln Leu Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro
290 295 300
Val Leu Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser
305 310 315 320
Lys Asp Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val
325 330 335
Thr Ala Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp
340 345 350
Gly Gly Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly
355 360 365
Val Val Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys
370 375 380
Phe Ser Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu
385 390 395 400
Thr Leu Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro
405 410 415
Thr Leu Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr
420 425 430
Pro Asp His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu
435 440 445
Gly Tyr Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr
450 455 460
Lys Thr Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg
465 470 475 480
Ile Glu Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly
485 490 495
His Lys Leu Glu Tyr Asn Glu Pro Gln Ser Ile Glu Val His Pro Arg
500 505 510
Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp Tyr Arg
515 520 525
Arg
<210> 15
<211> 530
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针
<400> 15
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Lys Ala
85 90 95
Asp Phe Thr Asp Ala Val Ile Asp Asn Ala Phe Glu Gly Gly Tyr Met
100 105 110
Ala Gly Arg Tyr Leu Ile Glu Arg Gly His Arg Glu Ile Gly Val Ile
115 120 125
Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala Gly Arg Leu Ala Gly Phe
130 135 140
Met Lys Ala Met Glu Glu Ala Met Ile Lys Val Pro Glu Ser Trp Ile
145 150 155 160
Val Gln Gly Asp Phe Glu Pro Glu Ser Gly Tyr Arg Ala Met Gln Gln
165 170 175
Ile Leu Ser Gln Pro His Arg Pro Thr Ala Val Phe Cys Gly Gly Asp
180 185 190
Ile Met Ala Met Gly Ala Leu Cys Ala Ala Asp Glu Met Gly Leu Arg
195 200 205
Val Pro Gln Asp Val Ser Leu Ile Gly Tyr Asp Asn Val Arg Asn Ala
210 215 220
Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile His Gln Pro Lys Asp Ser
225 230 235 240
Leu Gly Glu Thr Ala Phe Asn Met Leu Leu Asp Arg Ile Val Asn Lys
245 250 255
Arg Tyr Asn Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn
260 265 270
Gly Ile Lys Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser
275 280 285
Val Gln Leu Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly
290 295 300
Pro Val Leu Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu
305 310 315 320
Ser Lys Asp Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe
325 330 335
Val Thr Ala Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val
340 345 350
Asp Gly Gly Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr
355 360 365
Gly Val Val Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His
370 375 380
Lys Phe Ser Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys
385 390 395 400
Leu Thr Leu Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp
405 410 415
Pro Thr Leu Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg
420 425 430
Tyr Pro Asp His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro
435 440 445
Glu Gly Tyr Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn
450 455 460
Tyr Lys Thr Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn
465 470 475 480
Arg Ile Glu Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu
485 490 495
Gly His Lys Leu Glu Tyr Asn Glu Pro Gln Ser Ile Glu Val His Pro
500 505 510
Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp Tyr
515 520 525
Arg Arg
530
<210> 16
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针
<400> 16
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Lys Ala
85 90 95
Asp Phe Thr Asp Ala Val Ile Asp Asn Ala Phe Glu Gly Gly Tyr Met
100 105 110
Ala Gly Arg Tyr Leu Ile Glu Arg Gly His Arg Glu Ile Gly Val Ile
115 120 125
Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala Gly Arg Leu Ala Gly Phe
130 135 140
Met Lys Ala Met Glu Glu Ala Met Ile Lys Val Pro Glu Ser Trp Ile
145 150 155 160
Val Gln Gly Asp Phe Glu Pro Glu Ser Gly Tyr Arg Ala Met Gln Gln
165 170 175
Ile Leu Ser Gln Pro His Arg Pro Thr Ala Val Phe Cys Gly Gly Asp
180 185 190
Ile Met Ala Met Gly Ala Leu Cys Ala Ala Asp Glu Met Gly Leu Arg
195 200 205
Val Pro Gln Asp Val Ser Leu Ile Gly Tyr Asp Asn Val Arg Asn Ala
210 215 220
Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile His Gln Pro Lys Asp Ser
225 230 235 240
Leu Gly Glu Thr Ala Phe Asn Met Leu Leu Asp Arg Ile Val Asn Lys
245 250 255
Arg Glu Tyr Asn Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys
260 265 270
Asn Gly Ile Lys Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly
275 280 285
Ser Val Gln Leu Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp
290 295 300
Gly Pro Val Leu Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val
305 310 315 320
Leu Ser Lys Asp Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu
325 330 335
Phe Val Thr Ala Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn
340 345 350
Val Asp Gly Gly Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe
355 360 365
Thr Gly Val Val Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly
370 375 380
His Lys Phe Ser Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly
385 390 395 400
Lys Leu Thr Leu Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro
405 410 415
Trp Pro Thr Leu Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala
420 425 430
Arg Tyr Pro Asp His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met
435 440 445
Pro Glu Gly Tyr Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly
450 455 460
Asn Tyr Lys Thr Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val
465 470 475 480
Asn Arg Ile Glu Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile
485 490 495
Leu Gly His Lys Leu Glu Tyr Asn Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 17
<211> 530
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 17
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Tyr Asn Ser Asp Asn
85 90 95
Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys Ala Asn Phe
100 105 110
Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu Ala Asp His
115 120 125
Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu Leu Pro Asp
130 135 140
Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp Pro Asn Glu
145 150 155 160
Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala Ala Gly Ile
165 170 175
Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly Ser Gly Gly
180 185 190
Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val Pro Ile Leu
195 200 205
Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser Val Ser Gly
210 215 220
Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu Lys Leu Ile
225 230 235 240
Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu Val Thr Thr
245 250 255
Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp His Met Lys
260 265 270
Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr Val Gln Glu
275 280 285
Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr Arg Ala Glu
290 295 300
Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu Leu Lys Gly
305 310 315 320
Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys Leu Glu Tyr
325 330 335
Asn Leu Phe Lys Ala Asp Phe Thr Asp Ala Val Ile Asp Asn Ala Phe
340 345 350
Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His Arg
355 360 365
Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala Gly
370 375 380
Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys Val
385 390 395 400
Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly Tyr
405 410 415
Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala Val
420 425 430
Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala Asp
435 440 445
Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr Asp
450 455 460
Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile His
465 470 475 480
Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu Asp
485 490 495
Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His Pro
500 505 510
Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp Tyr
515 520 525
Arg Arg
530
<210> 18
<211> 530
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 18
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Tyr Asn Ser Asp Asn
85 90 95
Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys Ala Asn Phe
100 105 110
Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu Ala Asp His
115 120 125
Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu Leu Pro Asp
130 135 140
Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp Pro Asn Glu
145 150 155 160
Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala Ala Gly Ile
165 170 175
Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly Ser Gly Gly
180 185 190
Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val Pro Ile Leu
195 200 205
Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser Val Ser Gly
210 215 220
Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu Lys Leu Ile
225 230 235 240
Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu Val Thr Thr
245 250 255
Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp His Met Lys
260 265 270
Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr Val Gln Glu
275 280 285
Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr Arg Ala Glu
290 295 300
Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu Leu Lys Gly
305 310 315 320
Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys Leu Glu Tyr
325 330 335
Asn Ile Lys Lys Ala Asp Phe Thr Asp Ala Val Ile Asp Asn Ala Phe
340 345 350
Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His Arg
355 360 365
Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala Gly
370 375 380
Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys Val
385 390 395 400
Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly Tyr
405 410 415
Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala Val
420 425 430
Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala Asp
435 440 445
Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr Asp
450 455 460
Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile His
465 470 475 480
Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu Asp
485 490 495
Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His Pro
500 505 510
Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp Tyr
515 520 525
Arg Arg
530
<210> 19
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 19
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Tyr Asn Ser
85 90 95
Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys Ala
100 105 110
Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu Ala
115 120 125
Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu Leu
130 135 140
Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp Pro
145 150 155 160
Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala Ala
165 170 175
Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly Ser
180 185 190
Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val Pro
195 200 205
Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser Val
210 215 220
Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu Lys
225 230 235 240
Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu Val
245 250 255
Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp His
260 265 270
Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr Val
275 280 285
Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr Arg
290 295 300
Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu Leu
305 310 315 320
Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys Leu
325 330 335
Glu Tyr Asn Met Cys Ala Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 20
<211> 530
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 20
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Pro Gly Tyr Asn Ser
85 90 95
Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys Ala
100 105 110
Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu Ala
115 120 125
Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu Leu
130 135 140
Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp Pro
145 150 155 160
Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala Ala
165 170 175
Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly Ser
180 185 190
Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val Pro
195 200 205
Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser Val
210 215 220
Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu Lys
225 230 235 240
Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu Val
245 250 255
Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp His
260 265 270
Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr Val
275 280 285
Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr Arg
290 295 300
Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu Leu
305 310 315 320
Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys Leu
325 330 335
Glu Tyr Asn Lys Ala Asp Phe Thr Asp Ala Val Ile Asp Asn Ala Phe
340 345 350
Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His Arg
355 360 365
Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala Gly
370 375 380
Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys Val
385 390 395 400
Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly Tyr
405 410 415
Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala Val
420 425 430
Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala Asp
435 440 445
Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr Asp
450 455 460
Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile His
465 470 475 480
Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu Asp
485 490 495
Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His Pro
500 505 510
Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp Tyr
515 520 525
Arg Arg
530
<210> 21
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 21
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Leu Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 22
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 22
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Cys Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 23
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 23
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Gln Phe Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 24
<211> 529
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 24
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Lys Ala
85 90 95
Asp Phe Thr Asp Ala Val Ile Asp Asn Ala Phe Glu Gly Gly Tyr Met
100 105 110
Ala Gly Arg Tyr Leu Ile Glu Arg Gly His Arg Glu Ile Gly Val Ile
115 120 125
Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala Gly Arg Leu Ala Gly Phe
130 135 140
Met Lys Ala Met Glu Glu Ala Met Ile Lys Val Pro Glu Ser Trp Ile
145 150 155 160
Val Gln Gly Asp Phe Glu Pro Glu Ser Gly Tyr Arg Ala Met Gln Gln
165 170 175
Ile Leu Ser Gln Pro His Arg Pro Thr Ala Val Phe Cys Gly Gly Asp
180 185 190
Ile Met Ala Met Gly Ala Leu Cys Ala Ala Asp Glu Met Gly Leu Arg
195 200 205
Val Pro Gln Asp Val Ser Leu Ile Gly Tyr Asp Asn Val Arg Asn Ala
210 215 220
Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile His Gln Pro Lys Asp Ser
225 230 235 240
Leu Gly Glu Thr Ala Phe Asn Met Leu Leu Asp Arg Ile Val Lys Ala
245 250 255
Tyr Asn Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly
260 265 270
Ile Lys Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val
275 280 285
Gln Leu Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro
290 295 300
Val Leu Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser
305 310 315 320
Lys Asp Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val
325 330 335
Thr Ala Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp
340 345 350
Gly Gly Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly
355 360 365
Val Val Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys
370 375 380
Phe Ser Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu
385 390 395 400
Thr Leu Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro
405 410 415
Thr Leu Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr
420 425 430
Pro Asp His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu
435 440 445
Gly Tyr Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr
450 455 460
Lys Thr Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg
465 470 475 480
Ile Glu Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly
485 490 495
His Lys Leu Glu Tyr Asn Glu Pro Gln Ser Ile Glu Val His Pro Arg
500 505 510
Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp Tyr Arg
515 520 525
Arg
<210> 25
<211> 529
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 25
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Lys Ala
85 90 95
Asp Phe Thr Asp Ala Val Ile Asp Asn Ala Phe Glu Gly Gly Tyr Met
100 105 110
Ala Gly Arg Tyr Leu Ile Glu Arg Gly His Arg Glu Ile Gly Val Ile
115 120 125
Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala Gly Arg Leu Ala Gly Phe
130 135 140
Met Lys Ala Met Glu Glu Ala Met Ile Lys Val Pro Glu Ser Trp Ile
145 150 155 160
Val Gln Gly Asp Phe Glu Pro Glu Ser Gly Tyr Arg Ala Met Gln Gln
165 170 175
Ile Leu Ser Gln Pro His Arg Pro Thr Ala Val Phe Cys Gly Gly Asp
180 185 190
Ile Met Ala Met Gly Ala Leu Cys Ala Ala Asp Glu Met Gly Leu Arg
195 200 205
Val Pro Gln Asp Val Ser Leu Ile Gly Tyr Asp Asn Val Arg Asn Ala
210 215 220
Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile His Gln Pro Lys Asp Ser
225 230 235 240
Leu Gly Glu Thr Ala Phe Asn Met Leu Leu Asp Arg Ile Val Lys Gly
245 250 255
Tyr Asn Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly
260 265 270
Ile Lys Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val
275 280 285
Gln Leu Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro
290 295 300
Val Leu Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser
305 310 315 320
Lys Asp Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val
325 330 335
Thr Ala Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp
340 345 350
Gly Gly Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly
355 360 365
Val Val Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys
370 375 380
Phe Ser Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu
385 390 395 400
Thr Leu Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro
405 410 415
Thr Leu Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr
420 425 430
Pro Asp His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu
435 440 445
Gly Tyr Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr
450 455 460
Lys Thr Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg
465 470 475 480
Ile Glu Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly
485 490 495
His Lys Leu Glu Tyr Asn Glu Pro Gln Ser Ile Glu Val His Pro Arg
500 505 510
Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp Tyr Arg
515 520 525
Arg
<210> 26
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 26
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Lys Ala
85 90 95
Asp Phe Thr Asp Ala Val Ile Asp Asn Ala Phe Glu Gly Gly Tyr Met
100 105 110
Ala Gly Arg Tyr Leu Ile Glu Arg Gly His Arg Glu Ile Gly Val Ile
115 120 125
Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala Gly Arg Leu Ala Gly Phe
130 135 140
Met Lys Ala Met Glu Glu Ala Met Ile Lys Val Pro Glu Ser Trp Ile
145 150 155 160
Val Gln Gly Asp Phe Glu Pro Glu Ser Gly Tyr Arg Ala Met Gln Gln
165 170 175
Ile Leu Ser Gln Pro His Arg Pro Thr Ala Val Phe Cys Gly Gly Asp
180 185 190
Ile Met Ala Met Gly Ala Leu Cys Ala Ala Asp Glu Met Gly Leu Arg
195 200 205
Val Pro Gln Asp Val Ser Leu Ile Gly Tyr Asp Asn Val Arg Asn Ala
210 215 220
Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile His Gln Pro Lys Asp Ser
225 230 235 240
Leu Gly Glu Thr Ala Phe Asn Met Leu Leu Asp Arg Ile Val Asn Lys
245 250 255
Arg Glu Tyr Asn Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys
260 265 270
Asn Gly Ile Lys Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly
275 280 285
Ser Val Gln Leu Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp
290 295 300
Gly Pro Val Leu Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val
305 310 315 320
Leu Ser Lys Asp Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu
325 330 335
Phe Val Thr Ala Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn
340 345 350
Val Asp Gly Gly Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe
355 360 365
Thr Gly Val Val Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly
370 375 380
His Lys Phe Ser Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly
385 390 395 400
Lys Leu Thr Leu Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro
405 410 415
Trp Pro Thr Leu Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala
420 425 430
Arg Tyr Pro Asp His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met
435 440 445
Pro Glu Gly Tyr Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly
450 455 460
Asn Tyr Lys Thr Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val
465 470 475 480
Asn Arg Ile Glu Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile
485 490 495
Leu Gly His Lys Leu Glu Tyr Asn Val Gln Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 27
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 27
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Arg Pro Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 28
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 28
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Arg Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 29
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 29
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Ala Gly Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 30
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 30
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Leu Arg Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 31
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 31
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Phe Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 32
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 32
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Tyr Ile Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 33
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 33
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 34
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 34
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Asn Met Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 35
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 35
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Leu Ala Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 36
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 36
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Trp Gly Glu Ala Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Ile Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 37
<211> 530
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 37
His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala Tyr
1 5 10 15
Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys Gly
20 25 30
Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln Arg
35 40 45
Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu Val
50 55 60
Met Cys Ser Glu Tyr Pro Glu Gln Met Leu Ala Met Leu Glu Glu Tyr
65 70 75 80
Arg His Ile Pro Met Val Val Met Asp Arg Gly Ser Ile Cys Asn Ser
85 90 95
Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys Ala
100 105 110
Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu Ala
115 120 125
Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu Leu
130 135 140
Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp Pro
145 150 155 160
Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala Ala
165 170 175
Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly Ser
180 185 190
Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val Pro
195 200 205
Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser Val
210 215 220
Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu Lys
225 230 235 240
Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu Val
245 250 255
Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp His
260 265 270
Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr Val
275 280 285
Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr Arg
290 295 300
Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu Leu
305 310 315 320
Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys Leu
325 330 335
Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala Phe
340 345 350
Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His Arg
355 360 365
Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala Gly
370 375 380
Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys Val
385 390 395 400
Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly Tyr
405 410 415
Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala Val
420 425 430
Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala Asp
435 440 445
Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr Asp
450 455 460
Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile His
465 470 475 480
Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu Asp
485 490 495
Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His Pro
500 505 510
Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp Tyr
515 520 525
Arg Arg
530
<210> 38
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 38
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Thr Val Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 39
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 39
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Gly Val Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 40
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 40
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Gly Leu Met Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 41
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 41
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Cys Leu Val Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 42
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 42
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Tyr Ile Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 43
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 43
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Gly Leu Val Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 44
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 44
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Gly Ile Gln Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 45
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 45
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Gly Val Arg Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 46
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 46
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Cys Ile Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 47
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 47
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Ser Leu Arg Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 48
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 48
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Val Ile Arg Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 49
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 49
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Gly Ile Met Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 50
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 50
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Gly Cys Val Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 51
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 51
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Ser Ile Trp Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 52
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 52
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Ser Val Tyr Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 53
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 53
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Gly Ile Cys Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 54
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 54
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Gly Arg Val Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 55
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 55
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Gly Phe Leu Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 56
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 56
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Ser Val Arg Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 57
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 57
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Thr Ile Val Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 58
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 58
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Gly Phe Arg Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 59
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 59
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Gly Phe Val Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 60
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 60
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Ile Ile Val Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 61
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 61
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Gly Leu Arg Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 62
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 62
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Ser Val Trp Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 63
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 63
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly His Phe Val Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 64
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 64
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Ser Phe Val Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 65
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 65
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Gly Ser Val Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 66
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 66
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Ser Ile Val Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 67
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 67
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Asp Phe Val Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 68
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 68
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Ser Ile Leu Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 69
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 69
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Ser Leu Leu Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 70
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 70
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Arg Ile Leu Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 71
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 71
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Gly Phe His Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 72
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 72
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Gly Trp Val Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 73
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 73
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Ser Val Met Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 74
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 74
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Ser Leu Val Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 75
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 75
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Arg Phe Leu Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530
<210> 76
<211> 531
<212> PRT
<213> Artificial Sequence
<220>
<223> 光学探针变体
<400> 76
Met His Thr Lys Ser Ile Gly Leu Leu Ala Thr Ser Ser Glu Ala Ala
1 5 10 15
Tyr Phe Ala Glu Ile Ile Glu Ala Val Glu Lys Asn Cys Phe Gln Lys
20 25 30
Gly Tyr Thr Leu Ile Leu Gly Asn Ala Trp Asn Asn Leu Glu Lys Gln
35 40 45
Arg Ala Tyr Leu Ser Met Met Ala Gln Lys Arg Val Asp Gly Leu Leu
50 55 60
Val Met Cys Ser Glu Tyr Pro Glu Pro Leu Leu Ala Met Leu Glu Glu
65 70 75 80
Tyr Arg His Ile Pro Met Val Val Met Asp Arg Gly Cys Phe Val Asn
85 90 95
Ser Asp Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn Gly Ile Lys
100 105 110
Ala Asn Phe Lys Ile Arg His Asn Val Glu Asp Gly Ser Val Gln Leu
115 120 125
Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly Asp Gly Pro Val Leu
130 135 140
Leu Pro Asp Asn His Tyr Leu Ser Phe Gln Ser Val Leu Ser Lys Asp
145 150 155 160
Pro Asn Glu Lys Arg Asp His Met Val Leu Leu Glu Phe Val Thr Ala
165 170 175
Ala Gly Ile Thr Leu Gly Met Asp Glu Leu Tyr Asn Val Asp Gly Gly
180 185 190
Ser Gly Gly Thr Gly Ser Lys Gly Glu Glu Leu Phe Thr Gly Val Val
195 200 205
Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly His Lys Phe Ser
210 215 220
Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr Gly Lys Leu Thr Leu
225 230 235 240
Lys Leu Ile Cys Thr Thr Gly Lys Leu Pro Val Pro Trp Pro Thr Leu
245 250 255
Val Thr Thr Leu Gly Tyr Gly Leu Lys Cys Phe Ala Arg Tyr Pro Asp
260 265 270
His Met Lys Gln His Asp Phe Phe Lys Ser Ala Met Pro Glu Gly Tyr
275 280 285
Val Gln Glu Arg Thr Ile Phe Phe Lys Asp Asp Gly Asn Tyr Lys Thr
290 295 300
Arg Ala Glu Val Lys Phe Glu Gly Asp Thr Leu Val Asn Arg Ile Glu
305 310 315 320
Leu Lys Gly Ile Gly Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys
325 330 335
Leu Glu Tyr Asn Val Leu Asp Phe Thr Asp Ala Val Ile Asp Asn Ala
340 345 350
Phe Glu Gly Gly Tyr Met Ala Gly Arg Tyr Leu Ile Glu Arg Gly His
355 360 365
Arg Glu Ile Gly Val Ile Pro Gly Pro Leu Glu Arg Asn Thr Gly Ala
370 375 380
Gly Arg Leu Ala Gly Phe Met Lys Ala Met Glu Glu Ala Met Ile Lys
385 390 395 400
Val Pro Glu Ser Trp Ile Val Gln Gly Asp Phe Glu Pro Glu Ser Gly
405 410 415
Tyr Arg Ala Met Gln Gln Ile Leu Ser Gln Pro His Arg Pro Thr Ala
420 425 430
Val Phe Cys Gly Gly Asp Ile Met Ala Met Gly Ala Leu Cys Ala Ala
435 440 445
Asp Glu Met Gly Leu Arg Val Pro Gln Asp Val Ser Leu Ile Gly Tyr
450 455 460
Asp Asn Val Arg Asn Ala Arg Tyr Phe Thr Pro Ala Leu Thr Thr Ile
465 470 475 480
His Gln Pro Lys Asp Ser Leu Gly Glu Thr Ala Phe Asn Met Leu Leu
485 490 495
Asp Arg Ile Val Asn Lys Arg Glu Glu Pro Gln Ser Ile Glu Val His
500 505 510
Pro Arg Leu Ile Glu Arg Arg Ser Val Ala Asp Gly Pro Phe Arg Asp
515 520 525
Tyr Arg Arg
530

Claims (10)

1.一种次黄嘌呤结合蛋白变体，其：
(1)具有SEQ ID NO:1所示的序列并且在选自以下的1个、2个或3个、4个、5个、6个、7个或更多个位点具有突变：P129、L130、D146、W147、G148、E149、A150、K151、A152、N311、K312、R313、E314、E315、P316，所述氨基酸突变包括氨基酸的修饰、取代或缺失，
(2)是(1)的具有第58-341位氨基酸的截短变体，或
(3)是与(1)或(2)的序列具有至少70％序列相同性并具有(1)所述突变并保留对次黄嘌呤敏感性的序列，
优选地，所述突变的位点选自以下的1个、2个或3个、4个、5个、6个或7个：P129、L130、W147、G148、E149、A150、K151、A152、N311、K312、E315、P316，
更优选地，所述突变包括选自以下任一组的位点处的突变：E149和A150，A150和K151，G148和E149，K151和A152，N311和K312，E315和P316，
更优选地，所述突变包括K151和A152位点处的突变，任选还包括选自P129、L130、W147、E149和A150中的1个、2个、3个、4个或5个位点处的突变，
更优选地，所述突变包括K151和A152位点处的突变，并且还包括W147、E149和A150位点处或P129、L130、W147、E149和A150位点处的突变。
2.一种光学探针，包含次黄嘌呤敏感多肽和光学活性多肽，其中光学活性多肽位于次黄嘌呤敏感多肽的序列内，
所述次黄嘌呤敏感多肽具有：
(1)SEQ ID NO:1所示的序列或其具有第58-341位氨基酸的截短变体，或与它们有至少70％序列相同性并保留对次黄嘌呤敏感性的序列，
(2)权利要求1所述的次黄嘌呤结合蛋白变体的序列，或
(3)与(2)所述序列具有至少70％序列相同性并具有(2)所述突变并保留对次黄嘌呤敏感性的序列，
优选地，光学活性多肽位于次黄嘌呤敏感多肽的残基66-72，97-101，123-128，145-149，147-156，160-163，187-191，210-215，220-223，237-239，246-248，275-279，291-294，312-317或321-324中，
更优选地，光学活性多肽位于次黄嘌呤敏感多肽的选自下述位点中的任一个或多个：66/67、66/68、66/69,66/70、66/71、66/72、67/68、67/69、67/70、67/71、67/72、68/69、68/70、68/71、68/72、69/70、69/71、69/72、70/71、70/72、71/72、97/98、97/99、97/100、97/101、98/99、98/100、98/101、99/100、99/101、100/101、123/124、123/125、123/126、123/127、123/128、124/125、124/126、124/127、124/128、125/126、125/127、125/128、126/127、126/128、127/128、145/146、145/147、145/148、145/149、146/147、146/148、146/149、147/148、147/149、147/150、147/151、147/152、147/153、147/154、147/155、147/156、148/149、148/150、148/151、148/152、148/153、148/154、148/155、148/156、149/150、149/151、149/152、149/153、149/154、149/155、149/156、150/151、150/152、150/153、150/154、150/155、150/156、151/152、151/153、151/154、151/155、151/156、152/153、152/154、152/155、152/156、153/154、153/155、153/156、154/155、154/156、155/156、160/161、160/162,1 60/163、161/162、161/163、162/163、187/188、187/189、187/190、187/191、188/189、188/190、188/191、189/190、189/191、190/191、210/211、210/212、210/213、210/214、210/215、211/212、211/213、211/214、211/215、212/213、212/214、212/215、213/214、213/215、214/215、220/221、220/222、220/223、221/222、221/223、222/223、237/238、237/239、238/239、246/247、246/248、247/248、275/276、275/277、275/278、275/279、276/277、276/278、276/279、277/278、277/279、278/279、291/292、291/293、291/294、292/293、292/294、293/294、312/313、312/314、312/315、312/316、312/317、313/314,313/315、313/316、313/317、314/315、314/316、314/317、315/316、315/317、316/317、321/322、321/323、321/324、322/323、322/324、323/324。
3.如权利要求2所述的光学探针，其特征在于，所述光学活性多肽是荧光蛋白或其功能变体，
优选地，荧光蛋白选自黄色荧光蛋白、橘黄色荧光蛋白、红色荧光蛋白、绿色荧光蛋白、蓝色荧光蛋白、苹果红荧光蛋白，
更优选地，
荧光蛋白具有SEQ ID NO:2-9中任一所示的序列，和/或
荧光蛋白的功能变体具有SEQ ID NO:2所示的序列并在Y1位点具有突变，和/或
荧光蛋白的功能变体的突变为Y1C、Y1M、Y1Q、Y1V、Y1R、Y1W、Y1L或Y1H。
4.如权利要求2或3所述的光学探针，其特征在于，
次黄嘌呤敏感多肽是SEQ ID NO:1的具有第58-341位氨基酸的截短变体，并且光学活性多肽位于次黄嘌呤敏感多肽的选自下述位点中的任一个或多个：147/149、149/150、149/151、150/151、312/315、313/315、314/315；优选地，光学探针具有SEQ ID NO:10-16中任一项所示序列，或者
次黄嘌呤敏感多肽具有权利要求1所述的次黄嘌呤结合蛋白变体的序列，并且光学活性多肽位于次黄嘌呤敏感多肽的选自下述位点中的任一个或多个：147/149、149/150、149/151、150/151、312/315、314/315；优选地，光学活性多肽位于次黄嘌呤敏感多肽的150/151位点；更优选地，光学探针包含氨基酸序列SEQ ID NO:17-76中任一项所示的序列。
5.一种核酸分子，其包含：
(a)权利要求2-4中任一项所述的光学探针的编码序列，
(b)(a)的互补序列，或
(c)(a)或(c)的片段，
优选地，所述片段是引物。
6.包含权利要求5所述的核酸分子的核酸构建物，
优选地，所述核酸构建物是克隆载体、表达载体或重组载体。
7.一种宿主细胞，所述宿主细胞：
(1)表达权利要求2-4中任一项所述的光学探针；
(2)包含权利要求5所述的核酸分子；或
(3)包含权利要求6所述的核酸构建物。
8.一种检测试剂盒，其包含：
(1)权利要求2-4中任一项所述的光学探针，
(2)权利要求5所述的核酸序列，
(3)权利要求6所述的核酸构建物，或
(4)权利要求7所述的细胞，
所述检测试剂盒任选还包含利用光学探针检测次黄嘌呤所需的其他试剂，
优选地，所述检测试剂盒还包含选自以下的一种或多种试剂：缓冲液、培养基、次黄嘌呤标准品。
9.制备权利要求2-4中任一项所述的光学探针的方法，包括：提供权利要求7所述的宿主细胞，在所述光学探针表达的条件下培养所述宿主细胞，和分离光学探针。
10.权利要求2-4中任一项所述的光学探针、权利要求5所述的核酸序列、权利要求6所述的核酸构建物或权利要求7所述的宿主细胞在检测样品中的次黄嘌呤、筛选化合物、次黄嘌呤细胞内/外定位中的应用，
优选地，
检测样品中的次黄嘌呤包括步骤：使所述光学探针或宿主细胞与样品接触，检测光学活性多肽的光学变化，和根据光学活性多肽的光学变化检测样品中的次黄嘌呤，
所述筛选化合物包括步骤：在含次黄嘌呤的体系中使所述光学探针或宿主细胞与候选化合物接触，检测光学活性多肽的光学变化，和根据光学活性多肽的光学变化筛选化合物；优选地，所述筛选化合物包括步骤：在含次黄嘌呤的体系中使所述宿主细胞与候选化合物接触，并且光学活性多肽的光学变化指示所述候选化合物是否调节细胞对次黄嘌呤的摄取，
所述次黄嘌呤细胞内/外定位包括步骤：将含次黄嘌呤的体系与所述光学探针或所述宿主细胞接触，和检测光学活性多肽的光学变化，
更优选地，所述体系是溶液体系、细胞体系或亚细胞体系。
CN202210481174.9A 2022-05-05 2022-05-05 一种次黄嘌呤光学探针及其制备方法和应用 Pending CN117050150A (zh)

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