1、華中科技大學(xué)碩士學(xué)位論文煙草黃酮醇合成酶基因的分離及其序列分析姓名：黃偉偉申請學(xué)位級別：碩士專業(yè)：生物化學(xué)與分子生物學(xué)指導(dǎo)教師：何光源20070208華中科技大學(xué)碩士學(xué)位論文AbstractFlavonoidscomprisethemostcommongroupofpolyphenolicplantsecondarymetabolites.Therearediversegroupofplantscontainingflavoneisof

2、lavoneflavonolchalconeetc.Theyplaykeyrolesinbiologicalactivities.Flavonolsareanimptantpartofflavonoidsproducedbyflavonolsynthase.AccdingtotheconservedcDNAsequenceofflavonolsynthase(FLS)genesapairofprimerswasdesigned，flav

3、onolsynthasegenewasclonedfromtobacco(NicotianatabacunL.)leavesusingRTPCRRACEmethods.ThefulllengthcDNAsequencewas1186bpincludedawholeF，whichencoded333residues.TheaminoacididentityamongFLSsfromdifferentspecieswashighrangin

4、gfrom47%to87%.Theenzymebelongstotheclass2oxoglutaratedependentdioxygenases.SomesimilardomainstoIsopenicillinIIsynthase2OGFe(II)oxygenasesuperfamilywerefoundwhensearchingtheproteinconserveddomainsinGenBank.Polypeptidealig

5、nmentsofFLSswith2oxoglutaratedependentdioxygenasesrelated2oxoglutarateindependentenzymesrevealedeightstrictlyconservedaminoacidsinthreeregions.Homologymodelingwasusedtopredictthetertiarystructure.Theresultsofhomologymode

6、lingindicatedthatthemainchainofFLScontained11βstrsofwhichfmedajellyrolldoublestredβhelixtopology.Thejellyrollfmedahyhobiccavityoneendofwhichfmedtheactivesite.Fourstrictlyconservedaminoacids(His219His275Asp221Arg285)werei

7、nthehyhobiccavityothers(Gly65His72Pro205Gly259)inturn.LiketheotherdioxygenasesofthisclassflavonolsynthasecDNAencodedeightstrictlyconservedaminoacidresidueswhichincludedtwohistidines(His219His275)oneacidicaminoacid(Asp221

8、)residuefFe(II)codinationanarginine(Arg285)proposedtobind2oxoglutaratetherestfour(Gly65His72Pro205Gly259)requiredfproperfoldingoftheflavonolsynthasepolypeptide.Keywds:TobaccoFlavonolsynthaseRACEPolypeptidealignmentsHomol