Tyrosine phosphorylation of phosphatase inhibitor 2 |
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Authors: | John P. Williams Hanjoong Jo Ruthann E. Hunnicutt David L. Brautigan Jay M. McDonald Dr. |
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Affiliation: | 1. Department of Pathology, University of Alabama at Birmingham, Birmingham, Alabama 35294;2. Division of Biology and Medicine, Brown University, Providence, Rhode Island 02912 |
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Abstract: | Inhibitor 2 is a heat-stable protein that complexes with the catalytic subunit of type-1 protein phosphatase. The reversible phosphorylation of Thr 72 of the inhibitor in this complex has been shown to regulate phosphatase activity. Here we show that inhibitor 2 can also be phosphorylated on tyrosine residues. Inhibitor 2 was 32P-labeled by the insulin receptor kinase in vitro, in the presence of polylysine. Phosphorylation of inhibitor 2 was accompanied by decreased electrophoretic mobility. Dephosphorylation of inhibitor 2 by tyrosine phosphatase 1B, restored normal electrophoretic mobility. Phosphotyrosine in inhibitor 2 was detected by immunoblotting with antiphosphotyrosine antibodies and phosphoamino acid analysis. In addition, following tryptic digestion, one predominant phosphopeptide was recovered at the anode. The ability of inhibitor 2 to inhibit type-1 phosphatase activity was diminished with increasing phosphorylation up to a stoichiometry of 1 mole phosphate incorporated/mole of inhibitor 2, where inhibitory activity was completely lost. These data demonstrate that inhibitor 2 can be phosphorylated on tyrosine residues by the insulin receptor kinase, resulting in a molecule with decreased ability to inhibit type-1 phosphatase activity. |
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Keywords: | insulin dephosporylation type 1 phosphatase tyrosine kinase polylysine insulin receptor |
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