野生苋属植物籽实中新型α淀粉酶抑制剂的分离纯化和结构与功能研究

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1、湖南师范大学硕士学位论文野生苋属植物籽实中新型α-淀粉酶抑制剂的分离纯化和结构与功能研究姓名：王琳申请学位级别：硕士专业：生物化学与分子生物学指导教师：王贤纯;梁宋平20050501摘要性，但胰蛋白酶处理导致整个环状分子开环时会使抑制活性完全丧失。WAI一2能在弱酸性条件下，以竞争性抑制作用方式有效抑制美洲蜚蠊消化道Q一淀粉酶的活性。最适抑制pH值为5．5。37。C下与酶预温浴约35rain后显示最大抑制活性。当Ⅱ一淀粉酶用量一定时，a一淀粉酶活性的抑制率在约60％的范围内随抑制剂／酶比例的增大而呈线性增加，超过60％后，抑制率随抑制剂／酶比例的

2、增大而缓慢上升，最终达到最大值(约70％)。WAI一2也具有较强的热稳定性。经80。C水浴120inin后，抑制率约为50％：经100。C水浴10min后，抑制率仍有约30％。WAI-2对入唾液淀粉酶活性无影响。对美洲蜚蠊消化道Ⅱ一淀粉酶的抑制活性大于对黄粉虫消化道a一淀粉酶、猪胰腺Ⅱ一淀粉酶和米曲霉Ⅱ一淀粉酶的抑制活性。WAI一2的抑制活性低于WAI一1和AAI。关键词：蛋白质类抑制剂，抑制活性，结构，功能，苋属植物，a一淀粉酶IVABSTRACrABSTRACTTwoBorelproteinaceousinhibitors，namedWAI一

3、1andWA卜2respectivelY，havebeenfoundinthewildamaranth(Ama?'anthus"panicuMtus)weeds．Inthepresentstudy，WAI—landWAI一2werepurlfiedbyRP-HPLC(reverse—phasehighperformance1iquidchromatography)，andtheirproperties，structuresandbiochemicalactivitieswereinvestigated．WAI—IandWAI一2havemolec

4、ularweightsof986．5Daand1027．5DarespectirelY，determinedbyMALDI—TOF—MS(matrix—assistedlaser—desorption／ionizationtime—of-flightmassspectrometry)．TheyarethesmallestproteinaceousinhibitorsofQ—amylasefoundSOfarWAI—lcouldpotentlyinhibitthed—amylaseofPeriplanetaa破ericanadigestiveduc

5、tinanoncompetirivemannerundermiIdacidconditions，withtheoptimalinhibitoryactivityatpit6．0WAI一1exhibitedthehighestinhibitoryactivityafterpreincubationwiththeenzymeat37。Cforabout30min．WhenafixedamountofU—amylaseused，alongwiththeincreaseoftheinhibitory／enzymeratiotheinhibitionper

6、centagesoftheQ—amylaseactivitywere1inearlyincreaseduptoabout50％，andthenincreasedslowlYuptoamaximumofabout65％．AfterVABSTRACTpreincubationat80℃for120minWAl-1couldinhibit58．7％ofd—amylaseactivity：afterpreincubationat100℃for10min，WA[一lcouldsti1lleadtotheinhibitionrateof23．08％．whic

7、hindicatedthatWAI—lhasstrongthermalstabi1ity．WAI—ididnotinhibitthehumansalivaryQ—amylase．hogpancreatiCⅡ一amylase．bacil1ussubi1isQ—amylaseandaspergillusoryzaen—amylase．TheinhibitoryactivityofWAI—ltoPeriplanetaamericanadigestiveductQ—amylasewashigherthanthattoyel10wmealwormdiges

8、tiveductⅡ一amylase．AminoacidcompositionanalysiS，Edmandegradationandta