蛋白质相互作用VIP免费

Protein-proteinInteraction蛋白质-蛋白质相互作用03/27/2009Protein-ProteinInteractions-ACommonThemeinCellBiology蛋白质-蛋白质相互作用细胞生物学领域的普遍主题*DNA*RNA*ProteinCellassemblyandfunction/细胞组装与功能MessengerHeadquarterExecutorStable,DNAmutationStable/Versatile,rRNA,tRNA,mRNAVersatile,ProteinmodificationProtein-ProteininteractionProtein-OthercomponentinteractionProtein-ProteinInteractionsSingleproteinProteincomplexProteincomplexinteractionProteincomplexnetworkCell,Versatile,Dynamic,Accurateregulation……CellassemblyandfunctionProtein-ProteinInteractionsInteractionDomain-Structuralbasisforproteininteraction相互作用区域是蛋白质相互作用的结构基础*信号蛋白，调节蛋白以及大多数其他的蛋白质都具有包括相互作用区域在内的好几个结构域；*有些多肽仅由相互作用区域组成，从而充当蛋白质复合物的核心。InteractionDomain-Structuralbasisforproteininteraction相互作用区域是蛋白质相互作用的结构基础*SH2domain:phosphotyrosine-containingpeptidesrecognitiondomain。SH2分布最为广泛，识别特定的含有磷酸化Tyr的多肽模块。SH3SH2YkinaseSH3SH2SH3pYEEIpYVNVSrcSH2domain:Grb2SH2domain:InteractiondomainandProteininteractionSignalingtransductionpathwayPhysiologicalfunctionsofproteininteractionsLocalization蛋白质的亚细胞定位*蛋白质的亚细胞定位与其功能密切相关；*涉及：蛋白质-蛋白质相互作用；蛋白质与磷脂的相互作用；*蛋白质的组织特异性与亚细胞定位的研究是蛋白质功能研究的起始步骤Protein-proteininteractioncanbeusedtobuildcomplexbiologicalsystems.蛋白质-蛋白质相互作用是功能复合物形成的基础。（如线粒体内膜呼吸链）PhysiologicalfunctionsofproteininteractionsLocalizationandTraffickingAberrantinteractionscancontributetopathogenesis;错误的相互作用可能导致病理形象的出现。如Alzheimer’sdisease,beta-淀粉样结构的堆积；PhysiologicalfunctionsofproteininteractionsLocalizationandTraffickingInducibleproteininteraction&posttranslationalmodifications可诱导的蛋白相互作用与翻译后修饰Dynamicbehaviorofcellsinresponsetochangingconditionsishighlydependentonposttranslationalmodifications;细胞面对不断改变的环境表现出来的动态行为有赖于蛋白质的翻译后修饰以及它带来的蛋白质相互作用的变化。TechniquesandcomputationElucidationofProtein-ProteinInteractionsBiophysicalTechniquesMassSpectrometrySurfacePlasmonResonanceAtomicForceMicroscopyNMRX-rayCrystallographyHighThroughputTechniquesLibraryScreeningDegeneratePeptideLibrariesSpotBlotsCo-IPsGST-pulldownsYeast2-HybridPhageDisplayStandardTechniquesCo-IPsGST-pulldownsYeast2-HybridPhageDisplayInvivoImagingFRETBRETProteinInteractionNetworksTechniques：Principle,Process,AdvantageandDisadvantageGST-PulldownPrinciple:AffinitypurificationAffinitybetweenInteractionmotif/polypeptideanditsinteractionproteins;Advantage:Directinteraction/InvitroProcess:InteractionmotifGSTImmobilizedonGlutathionebeadsAffinitycolumnMakingAffinitypurificationfromtissueorcelllysateTechniques：免疫沉淀质量作用定律决定IP的成功度Techniques：免疫沉淀（非变性）•免疫沉淀的灵敏度取决于ProteinA(orG)与IgG的Kd、IgG与抗原的Kd值。所以增加抗原或抗体的浓度、使用与抗体直接共价偶联的珠子进行免疫沉淀可以提高灵敏度；ProteinABeadIgGAntigenKdKd**IgGKdAntigenCo-Immunoprecipitation/共免疫沉淀Principle:Affinitypurification/亲和纯化Advantage:Invivo(Physiologicalstatus)/代表生理状态下的相互作用XYXYCellCellLysisandImmunoprecipitationofproteinXZZ细胞裂解与蛋白质X的免疫沉淀Techniques：免疫沉淀（非变性）1.生理活性的保持：a.全程低温操作；b.裂解液中加入...