Catalysis of serine and tyrosine autophosphorylation by the human insulin receptor
Program in Molecular Medicine; Department of Biochemistry and Molecular Pharmacology
Amino Acid Sequence; Humans; Molecular Sequence Data; Phosphorylation; Phosphoserine; Phosphotyrosine; Protein-Tyrosine Kinases; Receptor, Insulin; Recombinant Proteins; Time Factors; Tyrosine
Life Sciences | Medicine and Health Sciences
The protein kinase activity of human insulin receptors purified from Sf9 insect cells after infection with a recombinant baculovirus was evaluated. The following experimental observations led to the unexpected conclusion that this receptor protein catalyzes both serine and tyrosine autophosphorylation at significant stoichiometries. (i) Phosphorylation of lectin-purified insulin receptors with [gamma-32P]ATP resulted in rapid receptor tyrosine phosphorylation (7 mol of P per high-affinity binding site) and the delayed onset of insulin-stimulated receptor serine phosphorylation (about 7% of total phosphorylation). The tyrosine kinase inhibitor (hydroxy-2-naphthalenylmethyl)phosphonic acid (HNMPA), which has no effect on protein kinase C or cyclic AMP-dependent protein kinase activities, inhibited both the receptor serine and tyrosine phosphorylation. (ii) Phosphorylation of a synthetic peptide substrate composed of insulin receptor residues 1290-1319 on serines-1305/1306 by partially purified insulin receptors was also inhibited by HNMPA. (iii) Insulin receptors sequentially affinity-purified on immobilized wheat germ agglutinin and immobilized insulin showed no apparent contaminant proteins on silver-stained SDS/polyacrylamide gels yet catalyzed autophosphorylation on receptor serine and tyrosine residues when incubated with [gamma-32P]ATP. These results suggest that the catalytic site of the insulin receptor tyrosine kinase also recognizes receptor serine residues as substrates for the phosphotransfer reaction. Furthermore, insulin-stimulated receptor serine phosphorylation in intact cells may occur in part by an autophosphorylation mechanism subsequent to tyrosine phosphorylation of the insulin receptor.
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Citation: Proc Natl Acad Sci U S A. 1992 Sep 1;89(17):7885-9.
Proceedings of the National Academy of Sciences of the United States of America
Baltensperger, Kurt; Lewis, Robert E.; Woon, Chee-Wai; Vissavajjhala, Prabhakar; Ross, Alonzo H.; and Czech, Michael P., "Catalysis of serine and tyrosine autophosphorylation by the human insulin receptor" (1992). Open Access Articles. 1821.