Phosphorylation of exogenous substrates by the insulin receptor-associated protein kinase.
A glycoprotein-enriched fraction derived from 3T3-L1 adipocyte membranes by Triton X-100 extraction and chromatography on wheat germ agglutinin-agarose contains an insulin-activable protein kinase that catalyzes the phosphorylation of tyrosine residues in proteins (Petruzzelli, L. M., Ganguly, S., Smith, C.J., Cobb, M. H., Rubin, C.S., and Rosen, O. M. (1982) Proc. Natl. Acad. Sci. U. S. A. 79, 6792-6796). The best peptide substrates for the enzyme are angiotensin II, and a synthetic peptide related to the amino acid sequence surrounding the site of tyrosine phosphorylation in the transforming protein kinase of Rous sarcoma virus. Kinetic analysis of the phosphorylation reaction with angiotensin II showed that insulin decreased the Km from 5.0 to 2.6 mM, and increased the Vmax from 0.6 to 2.2 pmol/min/10 fmol of insulin binding capacity. For the src-related peptide, the addition of insulin decreased the Km from 1.6 to 1.2 mM and increased the Vmax from 0.17 to 2.2 pmol/min/10 fmol of insulin binding capacity. Angiotensin III inhibitor, proctolin, beta-lipotropin (61-65) and Tyr-Arg were poorer substrates for the protein kinase. Protein substrates for the insulin-stimulated protein kinase include anti-src IgG, tubulin, casein, and histone H2b. The data suggest that the substrate specificity of the insulin-activable protein kinase is similar to that reported for the src and epidermal growth factor receptor protein kinases.
D000273:Adipose Tissue / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D007700:Kinetics; D051379:Mice; D015688:Oncogene Protein pp60(v-src); D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D011972:Receptor, Insulin / Q000378:metabolism*; D013379:Substrate Specificity; D014764:Viral Proteins / Q000378:metabolism
D014764:Viral Proteins; D011494:Protein Kinases; D011972:Receptor, Insulin; D015688:Oncogene Protein pp60(v-src)