Mutagenesis of lysine 460 in the human insulin receptor: Effects upon receptor recycling and cooperative interactions among binding sites

Hiroko Kadowaki, Takashi Kadowaki, Alessandro Cama, Bernice Marcus-Samuels, Adela Rovira, Charles L Bevins, Simeon I. Taylor

Research output: Contribution to journalArticle

84 Scopus citations

Abstract

Mutations in the insulin receptor gene can cause insulin resistance. Previously, we have identified a mutation substituting glutamic acid for lysine at position 460 in the α-subunit of the insulin receptor in a patient with a genetic form of insulin resistance. In the present work, we have investigated the effect upon receptor function of amino acid substitutions at position 460. Decreasing the pH from 8.0 to 5.5 caused a progressive acceleration of the dissociation of 125I-insulin from the wild-type insulin receptor. Substitution of acidic amino acids (Glu or Asp) for Lys460 decreased the ability of acid pH to accelerate dissociation of 125I-insulin. In contrast, substitution of Arg or neutral amino acids (Val, Met, Thr, or Gln) had no effect upon the sensitivity to acid pH. Correlated with decreased sensitivity to acid pH, substitution of Glu or Asp at position 460 retarded the dissociation of 125I-insulin from intracellular receptors subsequent to receptor-mediated endocytosis. Furthermore, retardation of dissociation of 125I-insulin from the internalized receptor was associated with a decreased half-life of the receptor. In summary, the Glu460 mutation appears to cause insulin resistance by accelerating receptor degradation and, thereby, decreasing the number of insulin receptors on the cell surface. Additional studies suggested that Lys460 may provide the amino groups whereby disuccinimidyl suberate cross-links the two α-subunits to each other. Consistent with the hypothesis that Lys460 is located at the interface between adjacent α-subunits, substitutions at position 460 impair cooperative interactions among insulin binding sites. The Glu460 mutation decreases positively cooperative binding interactions; the Arg460 mutation impairs negative cooperativity. Mutations at position 460 in the α-subunit did not decrease the ability of insulin to stimulate receptor tyrosine kinase.

Original languageEnglish (US)
Pages (from-to)21285-21296
Number of pages12
JournalJournal of Biological Chemistry
Volume265
Issue number34
StatePublished - Dec 5 1990
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry

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