title: Insulin-dependent phosphorylation of GTP-binding proteins in phospholipid vesicles.
abstract: The involvement of GTP-binding proteins (G proteins) in insulin action has been investigated in an in vitro system. Insulin receptors that have been purified by wheat germ lectin chromatography and either tyrosine-agarose chromatography, sucrose density centrifugation, or insulin-Sepharose chromatography have been co-inserted into phospholipid vesicles with different purified G proteins. The results of these studies indicate that a specific insulin-promoted phosphorylation of two G proteins, Go and Gi, can occur in these phospholipid vesicles. Bovine retinal transducin is a poor substitute for Go and Gi, being only weakly phosphorylated by the insulin receptor, and bovine brain Gs is not a substrate. The phosphorylation of Gi and Go occurs primarily on the alpha-subunits. Under optimal conditions, about one alpha o- or alpha i-subunit is phosphorylated on a tyrosine residue for every two beta-subunits of the insulin receptor, suggesting a 1:1 interaction between these G proteins and the heterotetrameric (alpha 2 beta 2) insulin receptor molecular. The inactive (GDP-bound) form of the alpha-subunits appears to be the preferred substrate, with the phosphorylation being significantly reduced in alpha o and alpha i upon the binding of guanosine 5'-O-thiotriphosphate (GTP gamma S) and completely eliminated in the pure alpha-GTP gamma S complex of transducin. The Gi and Go proteins also cause an enhancement of the insulin-stimulated receptor autophosphorylation. This enhancement is a reflection of an increased incorporation of the insulin receptor into lipid vesicles which is induced by these G proteins. Taken together these results provide evidence for the interactions of G proteins with the insulin receptor in a lipid milieu.
mesh_terms: D000818:Animals; D001923:Brain Chemistry; D002417:Cattle; D002499:Centrifugation, Density Gradient; D002845:Chromatography; D003902:Detergents; D019204:GTP-Binding Proteins / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism; D006801:Humans; D007328:Insulin / Q000494:pharmacology*; D007700:Kinetics; D008081:Liposomes / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism; D008970:Molecular Weight; D010743:Phospholipids / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D010920:Placenta / Q000032:analysis; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011972:Receptor, Insulin / Q000302:isolation & purification / Q000378:metabolism*; D013873:Thionucleotides / Q000378:metabolism; D015293:Transducin
chemical_list: D003902:Detergents; D007328:Insulin; D008081:Liposomes; D008565:Membrane Proteins; D010743:Phospholipids; D010750:Phosphoproteins; D013873:Thionucleotides; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate; D011505:Protein-Tyrosine Kinases; D011972:Receptor, Insulin; D019204:GTP-Binding Proteins; D015293:Transducin