SUBSTITUTION OF GLUTAMINE FOR ARGININE-1131 - A NEWLY IDENTIFIED MUTATION IN THE CATALYTIC LOOP OF THE TYROSINE KINASE DOMAIN OF THE HUMAN INSULIN-RECEPTOR

We studied a patient with severe insulin resistance and a remarkable d ecrease in the in vivo autophosphorylation of the insulin receptor. Us ing a polymerase chain reaction-single strand conformation polymorphis m method and direct sequencing, we identified a heterozygous mutation substituting Gln for Arg1131 in the putative ''catalytic loop'' of the tyrosine kinase domain of the insulin receptor gene. The Gln113 mutan t receptor was expressed by transfection in Chinese hamster ovary cell s and compared with cells expressing the wild-type insulin receptor. B oth mutant and wild-type receptors were expressed on the cell surface and displayed similar insulin-binding affinity. The Gln1131 mutation i mpaired the activity of the receptor tyrosine kinase and inhibited the ability of insulin to phosphorylate the endogenous substrate insulin receptor substrate-I. In addition, the Gln1131 mutant receptor exhibit ed diminished tyrosine-phosphorylated phosphatidylinositol 3-kinase an d myelin basic protein kinase activities compared with the wild-type c ells. It also demonstrated a defective mediation of the insulin signal stimulating 2-deoxy-D-glucose transport and thymidine incorporation, resistance to endocytosis, and insulin-induced down-regulation. Unlike a previously described mutation in the putative catalytic loop of the receptor that substituted Glu for Ala1135, the Gln1131 mutation retai ned proteolytic cleavage of the proreceptor into separate subunits. Ou r results demonstrate that a naturally occurring mutation (R1131Q) in the putative catalytic loop of the insulin receptor results in severe impairment of the tyrosine kinase function in our patient. In addition , our results indicate that Arg1131 is important for receptor-mediated insulin action in vivo and suggest that the amino acids constituting the catalytic loop of protein kinases may possess different modes in o rder to retain kinase function.