Methods in molecular biology (Clifton, N.J.) · 2004
Determination of the catalytic activities of mTOR and other members of the phosphoinositide-3-kinase-related kinase family.
СтатьяПоддержка службы здравоохранения СШАНазвание и аннотация: Перевод в очереди
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Chiang Gary G, Abraham Robert T
Аннотация
Members of the phosphoinositide-3-kinase-related kinase (PIKK) family, which includes mTOR, ATM, ATR, and hSMG-1, play important roles in regulating the cellular response to environmental stimuli. Despite the similarity of their catalytic domain to that of phosphoinositide-3-kinase, these extremely large (>250 kDa) polypeptides function as serine/threonine protein kinases. The catalytic activities of these PIKK family members can now be measured in immune-complex kinase assays. This assay involves isolation of the kinase by immunoprecipitation and the in vitro phosphorylation of a specific substrate in the presence of radio-labeled ATP. Here we describe, in detail, the determination of PIKK catalytic activity with a standardized immune-complex kinase assay protocol.
Предметные рубрики MeSH
1-Phosphatidylinositol 4-KinaseAdenosine TriphosphateAntibodies, MonoclonalAntigens, SurfaceAtaxia Telangiectasia Mutated ProteinsCatalysisCell Cycle ProteinsCells, CulturedDNA-Binding ProteinsHumansMembrane ProteinsPhosphorus RadioisotopesPhosphorylationPrecipitin TestsProtein KinasesProtein Serine-Threonine KinasesSubstrate SpecificityTOR Serine-Threonine KinasesTumor Suppressor Proteins