Antibodies to the extracellular receptor domain restore the hormone-insensitive kinase and conformation of the mutant insulin receptor valine 382.
A mutation substituting a valine for phenylalanine at residue 382 in the insulin receptor alpha-subunit has been found in two sisters with a genetic form of extreme insulin resistance. This receptor mutation impairs the ability of the hormone to activate autophosphorylation of solubilized receptors and phosphorylation of substrates (Accili, D., Mosthaf, L., Ullrich, A., and Taylor, S. I. (1991) J. Biol. Chem. 266, 434-439). We have previously demonstrated that in native receptors insulin induces a conformational change in the receptor beta-subunit, which is thought to be necessary for receptor activation (Baron, V., Gautier, N., Komoriya, A., Hainaut, P., Scimeca, J. C., Mervic, M., Lavielle, S., Dolais-Kitabgi, J., and Van Obberghen, E. (1990) Biochemistry 29, 4634-4641). Hence, it was thought that a defect in this conformational change might explain the functional defect of the mutant receptor. This appears to be the case, as we demonstrate here that the mutant receptor is locked in its inactive configuration. However, we found two monoclonal antibodies, directed to the extracellular domain, which are capable of restoring the mutant receptor kinase activity. The activation of the mutant receptor was accompanied by restoration of conformational changes in the beta-subunit C terminus. From these data, we draw the two following conclusions. (i) A causal link exists between receptor kinase activation and the occurrence of conformational changes. (ii) Ligands other than insulin, such as antibodies, which perturb the extracellular domain, can function as alternative ways to restore the mutant receptor kinase.
Institut National de la Santé et de la Recherche Médicale, Unité 145, Faculté de Médecine, Nice, France.
C
C
Lebrun
V
V
Baron
P
P
Kaliman
N
N
Gautier
J
J
Dolais-Kitabgi
S
S
Taylor
D
D
Accili
E
E
Van Obberghen
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology*; D006801:Humans; D007328:Insulin / Q000378:metabolism; D007333:Insulin Resistance; D007700:Kinetics; D051379:Mice; D016297:Mutagenesis, Site-Directed; D010455:Peptides / Q000138:chemical synthesis / Q000276:immunology; D010766:Phosphorylation; D011487:Protein Conformation; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D011972:Receptor, Insulin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D014162:Transfection; D014633:Valine
D000911:Antibodies, Monoclonal; D007328:Insulin; D010455:Peptides; D011994:Recombinant Proteins; D011505:Protein-Tyrosine Kinases; D011972:Receptor, Insulin; D014633:Valine