Catalysis of serine and tyrosine autophosphorylation by the human insulin receptor

K. Baltensperger, Robert E Lewis, C. W. Woon, P. Vissavajjhala, A. H. Ross, M. P. Czech

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Abstract

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 [γ-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 [γ-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.

Original languageEnglish (US)
Pages (from-to)7885-7889
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume89
Issue number17
DOIs
StatePublished - Jan 1 1992

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Catalysis
Insulin Receptor
Serine
Tyrosine
Phosphorylation
Adenosine Triphosphate
Silver Proteins
Sf9 Cells
Wheat Germ Agglutinins
human INSR protein
Baculoviridae
Cyclic AMP-Dependent Protein Kinases
Lectins
Protein-Tyrosine Kinases
Protein Kinases
Protein Kinase C
Insects
Catalytic Domain
Binding Sites
Insulin

ASJC Scopus subject areas

  • General

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Catalysis of serine and tyrosine autophosphorylation by the human insulin receptor. / Baltensperger, K.; Lewis, Robert E; Woon, C. W.; Vissavajjhala, P.; Ross, A. H.; Czech, M. P.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 89, No. 17, 01.01.1992, p. 7885-7889.

Research output: Contribution to journalArticle

Baltensperger, K. ; Lewis, Robert E ; Woon, C. W. ; Vissavajjhala, P. ; Ross, A. H. ; Czech, M. P. / Catalysis of serine and tyrosine autophosphorylation by the human insulin receptor. In: Proceedings of the National Academy of Sciences of the United States of America. 1992 ; Vol. 89, No. 17. pp. 7885-7889.
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