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ID:36602570
大小:6.17 MB
页数:92页
时间:2019-05-12
《一种番茄褐孢霉产生的高选择性转化人参皂甙Rb1为Rd的β葡萄糖苷酶》由会员上传分享,免费在线阅读,更多相关内容在行业资料-天天文库。
1、东北师范大学博士学位论文一种番茄褐孢霉产生的高选择性转化人参皂甙Rb<,1>为Rd的β-葡萄糖苷酶姓名:赵雪淞申请学位级别:博士专业:生物化学与分子生物学指导教师:周义发20090401Knowledgebase),得到了其授予的序列接受号P85516。G.I的氨基酸序列与D.葡萄糖苷酶家族3的几种真菌p.葡萄糖苷酶有高度的同源性,可能属于此家族的成员之一。4、酶反应动力学实验证实,G.I具有较好的pH稳定性和热稳定性,在pH4.0~l1.0范围内和40℃以下表现出良好的D.葡萄糖苷酶活性,最适反应
2、pH值为pH6.0,最适反应温度为45℃。Cu2+,Zn2+在50mM时对酶活性有较强的抑制作用,相反,Nr、K+、CaZ十、Mn2十和Mgz+对酶活性有轻微的刺激作用。O.25M的SDS没有影响G.I的酶活性,O.25mM的EDTA只是轻微抑制了G.I的活性。G.I的Km值为O.18mM,最大反应初速度Vmax为5.52mM/min。底物专一性分析发现,G.I能高特异性水解人工合成的底物pNPG,还能水解p.葡萄糖苷键连接的二糖如纤维二糖、龙胆二糖和槐二糖,再次表明此酶为一种D.葡萄糖苷酶。G.I
3、对人参皂甙的底物专一性研究表明,此酶对人参皂甙Rbl表现了很强的水解活性,可达到州PG的22.1%,而不水解人参皂甙Rb2、Re和IH,说明此酶对人参皂甙Rbl中C.20位的p(1_6)糖苷键更具有特异性,不水解人参皂甙其它糖苷键,G.I对人参皂甙的这种高选择性水解为Rd的工业制备奠定了基础。关键词:人参皂甙Rbl;人参皂甙Rd;番茄褐孢霉(CladosporiumflulvumCooke,异名FuliafulvaCiferri);p.葡萄糖苷酶;生物转化ⅡAbstractPanaxginsengC
4、.A.MeyerhasbeenusedasamedicineinChinaover2000years.Ginsenosidesarethemajoractivecomponentsofginseng.Morethan40ginsenosideshavebeenisolatedandidentified.IthasbeenfoundthattheminorginsenosidessuchasR93,CKandRdhaveverygoodbioactivities.Itisdifficulttoprep
5、areminorginsenosidesbyextractionfromginsengbecauseoftheirlOWconcentration.Basedoncurrenttechnologies,itiSimpossibletopreparetheminorginsenosidesbychemicalsynthesis.Atpresent,apossiblepathwayforpreparationoftheminorginsenosidesistransformationfromstruct
6、urallyrelatedmajorginsenosides.TheamountofthemajorginsenosideRblishighinginsengandithasthesameaglycone(protopanaxadi01)astheminorginsenosides.RblhasoneormoresugarresiduesattheC·20positionthantheminorginsenosides.Theoretically,theminorginsenosidescanbeo
7、btainedbyhydrolysisofRb;toremoveoneormoreglucoseresidueatpositionC-20.Chemicaltransformationusuallyhaspoorselectivityandgeneratesmoreenvironmentalpollution.Biotransformationisthoughttohavemorepotentialforconversionbecauseofitshi.ghspecificityandenviron
8、mentalcompatibility.SomeeffortshavebeenmadetolookforsuitableenzymesthatcanconvertRblintotheminorwhichresultsinalowyieldofRd.ginsenosides.However,mostlackspecificityThispaperreportsthatanovelginsenosideRbl-hydrolyzing13-o-glucosidase(G—I
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