Department of Biochemistry
Chromatography, Gel; Chromatography, High Pressure Liquid; Detergents; Electrophoresis, Polyacrylamide Gel; Insulin; Liposomes; Octoxynol; Peptide Mapping; Phospholipids; Phosphorylation; Polyethylene Glycols; Protein-Tyrosine Kinases; Receptor, Insulin
Endocrinology, Diabetes, and Metabolism | Medical Biochemistry
Insulin receptor kinase, affinity-purified by adsorption and elution from immobilized insulin, is stimulated 2-3-fold by insulin in detergent solution. Reconstitution of the receptor kinase into leaky vesicles containing phosphatidylcholine and phosphatidylethanolamine (1:1, w/w) by detergent removal on Sephadex G-50 results in the complete loss of receptor kinase sensitivity to activation by insulin. Insulin receptors in these vesicles also exhibit an increase in their apparent affinity for 125I-insulin (Kd = 0.12 nM versus 0.76 nM). Inclusion of 8.3-16.7% phosphatidylserine into the reconstituted vesicles restores 40-50% of the insulin-sensitivity to the receptor kinase. An elevated apparent affinity for 125I-insulin of insulin receptors in vesicles containing phosphatidylcholine and phosphatidylethanolamine is also restored to the value observed in detergent solution by the inclusion of phosphatidylserine in the reconstituted system. The effect of phosphatidylserine on insulin receptor kinase appears specific, because cholesterol, phosphatidylinositol and phosphatidic acid are all unable to restore insulin-sensitivity to the receptor kinase. Autophosphorylation sites on the insulin receptor as analysed by h.p.l.c. of tryptic 32P-labelled receptor phosphopeptides are not different for insulin receptors autophosphorylated in detergent solution or for the reconstituted vesicles in the presence or absence of phosphatidylserine. These data indicate that the phospholipid environment of insulin receptors can modulate its binding and kinase activity, and phosphatidylserine acts to restore insulin-sensitivity to the receptor kinase incorporated into phosphatidylcholine/phosphatidylethanolamine vesicles.
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Citation: Biochem J. 1987 Dec 15;248(3):829-36.
The Biochemical journal
Lewis, Robert E. and Czech, Michael P., "Phospholipid environment alters hormone-sensitivity of the purified insulin receptor kinase" (1987). Open Access Articles. 204.