19613.重組日本七鰓鰻rljrgd2與其4種突變體蛋白對lewis細胞生物活性的比較

1、遼寧師范大學(xué)碩士學(xué)位論文重組日本七鰓鰻rLjRGD2與其4種突變體蛋白對Lewis細胞生物活性的比較姓名：劉洋申請學(xué)位級別：碩士專業(yè)：細胞生物學(xué)指導(dǎo)教師：王繼紅201204重組日本七鰓鰻山RGD2與其4種突變體蛋白對Lewis細胞生物活性的比較TheComparisonofTumorCellActivitiesamongTheLj—RGD2ToxinProteinandIts’RGDDeleteMutationsAbstractLjRG

2、D2isahistidinerichpeptidewithtwoRGD(Arg—G1y—Asp)motifsfromthebuccalglandofLampetrajaponica，whichmolecularweightis8kDaBecauseRGDmotifsandhistidinerichstructurearea11helpfultoantitumorandantiangiogenesisfunctionwestudiedth

3、erelationshipbetweenthefunctionandthestructureofLjRGD2WesynthesizedthegeneoftheRGDdeletemutations：themutationwhichkeptthesecondRGDmotifwasnamedLj738，themutationwhichkeptthefirstRGDmotifwasnamedL卜739，themutationwhichwasde

4、letedalltwGDmotifsofLiRGD2wasnamedLj740andthemutationwhichthea112RGDmotifswerereplacedby2KGDwasnamedLj741AfterconstructedabovegenestopET23b，wetransformedtherecombinationplasmidsintoEcoilBL21Furthermore，therecombinantprot

5、einsexpressionwasinducedbyIPTGandpurifiedbyHisaffinitychromatographyTheresultofcellproliferationassaydemonstratedthatrLjRGD2andthefourformsofmutationproteinrLi738，r“739，rLj740andrLj741inhibitedbFGFinducedproliferationofL

6、ewiscellsinadosedependentmannerwithIC50at1129mol／L，1239mol／L，111Bm01／L，118pm01／Land103gm01／L，respectivelyInordertodeterminetheeffectofrLiRGD2andthefourmutationproteinonLewiscellsmigrationtowardbFGFweusedthetranswellconta

7、ininginsertfilter(80gmporesize，polycarbonatefilter)Theresultsshowedthat，allofrLjRGD2，rLj一738，rLj一739，rLj740andrLj一741showedsignificantinhibitiononLewiscellsmigration，theinhibitionrateswere30％，3l％，31％，34％and25％，respective

8、lyIntheinvasionassaytheMatrigelandTranswellwereusedtoimitateenvironmentinvivoTheresultsofinvasionassayrevealedthat，rLjRGD2andthemutationproteinssignificantlyinhibitedbFGFinducedinvasionofLewiscells，buttheinhibitioneffect

9、withouttoomuchdifferencerLjRGD2andthemutationproteinssignificantlybreakdownthecytoskeletonoftheLewiscellsAUofrLiRGD2andthemutationproteinscouldinducetheapoptosisofLewiscellsThepartialsignalpathwayofabovewildtypeLjRGD2and

10、mutationsweredetectedtheresultsshowedthatallofaboveproteinsdecreasedtheexpressionlevelsofFAK，ILKandFAKphosphorylationTakentogether，theseresultssuggestedthat，thefunctionsofLj—RGD2arerelativewithRGDmotifsorhistidinerichstr