Chemical and functional analysis of components of adenylyl cyclase from human platelets treated with phorbolesters

FEBS Letters

Palm, Dieter¹  Simmoteit, Robert¹  Mollner, Stefan¹  Pfeuffer, Thomas¹  Schulzki, Horst-Dieter¹ 

[1] Department of Physiological Chemistry, University of Würzburg, Medical School, Würzburg, Germany

关键词: Adenylyl cyclase;    Tetradecanoylphorbol acetate;    Phosphorylation;    Hormonal inhibition;    TPA;    tetradecanoylphorbol acetate;    Gpp(NH)p;    guanosine 5′-(β;    γ-imido)triphosphate;    GTPγS;    guanosine 5′-(γ-thio)-triphosphate;    C;    catalytic component of adenylyl cyclase;    Gs;    guanine nucleotide binding regulatory component responsible for stimulation of adenylyl cyclase;    Gi;    guanine nucleotide binding regulatory component responsible for inhibition of adenylyl cyclase;    SDS;    sodium dodecyl sulfate;    Enzymes: adenylyl cyclase (EC 4.6.1.1);    alkaline phosphatase (EC 3.1.3.1);    creatine kinase (EC 2.7.3.2);   

DOI  :  10.1016/0014-5793(91)80734-K

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Human platelets, prelabeled with [³²P]phospate were treated with tetradecanoylphorbol acetate (TPA) for 5 min at 37°C. Phosphorylation of the components of adenylyl cyclase was determined in membranes using specific antibodies against G-proteins and the catalytic moiety. Less than 0.01 mol of [³²P]phosphate/mol could be detected in immunoprecipitates using antibodies against sequences within the α-subunit of the GTP binding protein G_i. TPA, however, caused the incorporation of 0.67–1.1 mol of [³²P]phosphate per mol of catalyst while 0.13-0.2 mol were found in the absence or TPA. Lack of modification of the α-subunit of G_i was also indicated by the results of reconstitution experiments with purified G_iα from bovine brain: adenylyl cyclase in membranes from untreated platelets was significantly more inhibited by added G_iα, than that from TPA treated cells. While β,γ-subunits were like-wise inhibitory no difference dependent on platelet-pretreatment could be observed.

【 授权许可】

Unknown