15490.日本七鰓鰻組織蛋白酶l的克隆表達及功能分析

1、學位論文獨創(chuàng)性聲明本人承諾：所呈交的學位論文是本人在導師指導下所取得的研究成果。論文中除特別加以標注和致謝的地方外，不包含他人和其他機構已經(jīng)撰寫或發(fā)表過的研究成果，其他同志的研究成果對本人的啟示和所提供的幫助，均已在論文中做了明確的聲明并表示謝意。學位論文作者簽名：盞舀薹孥學位論文版權的使用授權書本學位論文作者完全了解遼寧師范大學有關保留、使用學位論文的規(guī)定，及學校有權保留并向國家有關部門或機構送交復印件或磁盤，允許論文被查閱和借閱。本

2、文授權遼寧師范大學，可以將學位論文的全部或部分內容編入有關數(shù)據(jù)庫并進行檢索，可以采用影印、縮印或掃描等復制手段保存、匯編學位論文，并且本人電子文檔的內容和紙質論文的內容相一致。保密的學位論文在解密后使用本授權書。學位論文作者簽名：盔矗，姿指導教師簽名：簽名日期：zal)年廠月70日日本七鰓鰻組織蛋白酶L的克隆表達及功能分析UOmngExpressionandFunctionA11alysisotcathepsinLgene拉Om／一‘‘

3、一‘●一‘^‘‘一^LampetrajaponicaAbstraotCathepsinL(CTSL)belongstothelysosomalcysteineproteasefamily，whichiscomposedofasignalpeptide，apropeptideandamaturepeptideCTSLplaysimportantrolesinavarietyofbiologicalprocesses，suchasprote

4、olysis，antigenpresentingcellapoptosis，TcellsortingaswellasembryonicdevebpmentInadditiontheabnormalchangeoftheexpressionlevelofCTSLaccountsfortheinductionofavarietyoftumors，cardiovascularandkidneydiseaseInparasiticanimals

5、，CTSLmainlyparticipatedinnutritiomlintake，tissuehvasiortimllauneescapeandtissuedifferentiatiorLInthepresentstudy，thefulllengtheDNA(1457bp)ofJapaneselampreyCTSL田CTSL)wasobtainedthroughgenecbning，5and3’endoftherapidamplifi

6、cationexperiments，whichiscomposedbythe5’UTRnoncodingareaof104bp，3UTRnoncodingregionof333bp，andtheopenreadingframeof1020bp，encoding339aminoacidsBioinformaticsanalysisshowedthattheLj—CTSLaminoacidsequencecontainsasignalpep

7、tide(MetlAlal8)，aprecursordomain(Thrj9Glull7)andamaturedomain(AhllSVal339)Thereisapotentialglycosyhtionsites(ASh226)，sixsubstratebindingsites(I_eul86，Met】87，Ah2“，kf80，GIF83andSer335)，andacatalyticsiteconstitutedbythecyst

8、eine(Cysl42)，histidine(His282)andtheasparagine(Ash306)residuesSequencealignmentshowedthattheaminoacidsequenceoftheLjCTSLhasahighhomologywiththeCTSLsfromtheotherspecies，andthehomologywasupto81。1％withtheCTSL舶mXenopuslaevis

9、Besides，巧CTSLcontainsthehighlyconservedmotifs，ERF／WNINandGNFDPhylogenetictreeanalysisshowedthatthe可一CTSLisclosertOtheCTSLfromtheamphibianXenopuslaevis，inaccordancewiththerolesofcreaturesareevolutedfiomlowtOhighHoweverLjC

10、TSLisalsobcatedonitsownbranch，whichdisplaysitsuniqueevolutionarystatusSemiquantitativeRTPCRshowedthatthe碼CTSLiswidelydistributedinthehmpreyheartoralgland，kidneyintestine，livergillsandgonads，andtheexpressionlevelof巧CTSLwa

11、sthemostintheoralgland，whichsuggeststhe巧CTSLplaysimportantrolesinthenutritionalintakeprocessfortheJapaneselampreysThe巧一CTSLwassubcbnedintothepmkaryoticexpressionvectorpTWIN，andsuccessfullyexpressedinEcoliBL21Thesolublere