Insulin receptor aggregation and autophosphorylation in the presence of cationic polyamino acids.
Aggregation and autophosphorylation of the insulin receptor-protein kinase, from cultured 3T3-L1 adipocytes, were studied in the presence of cationic polyamino acids. Poly-L-lysine and poly-L-arginine produced the following effects with the purified receptor: first, the autophosphorylation rate was increased by polycations. Half-maximal stimulation was proportional to polymer length. The rate enhancement was greater at lower ATP concentrations. Second, near-endpoint (equilibrium) autophosphorylation was greater in the presence of the polycations. Polycations inhibited the reverse reaction: ADP + phosphoreceptor yielding ATP + aporeceptor. Third, the [32P]phosphopeptides generated by trypsin digestion of the 32P-beta-subunit, showed that no new autophosphorylation sites resulted from the presence of polycations. Fourth, the polycations, but not insulin, promoted receptor aggregation, and phosphoreceptor aggregated more readily than aporeceptor. Insulin receptor enriched through the wheat germ agglutinin eluate step was compared with purified receptor. Higher concentrations of poly-L-arginine were required to stimulate autophosphorylation and to promote aggregation. Finally, several polycation-dependent substrates present in the wheat germ agglutinin eluate co-aggregated with the insulin receptor. Polycation-stimulated receptor autophosphorylation is linked to a lower KM,app for ATP, but substrate phosphorylation may require the aggregation.
Kohanski
R A
RA
Department of Biochemistry, Mount Sinai School of Medicine, New York, New York 10029.
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D007700:Kinetics; D046911:Macromolecular Substances; D051379:Mice; D010455:Peptides / Q000494:pharmacology*; D010761:Phosphorus Radioisotopes; D010766:Phosphorylation; D011107:Polylysine / Q000494:pharmacology*; D011972:Receptor, Insulin / Q000187:drug effects / Q000378:metabolism*
D046911:Macromolecular Substances; D010455:Peptides; D010761:Phosphorus Radioisotopes; D011107:Polylysine; C015462:polyarginine; D000255:Adenosine Triphosphate; D011972:Receptor, Insulin