Insulin-stimulated tyrosine phosphorylation of the insulin receptor in detergent extracts of human placental membranes. Comparison to epidermal growth factor-stimulated phosphorylation

• Published in: The Journal of biological chemistry Vol. 257; no. 24; pp. 15162 - 15166
• Main Authors: Avruch, J, Nemenoff, R A, Blackshear, P J, Pierce, M W, Osathanondh, R
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 25-12-1982; American Society for Biochemistry and Molecular Biology
• Subjects: Adenosine Triphosphate - metabolism; Cations, Divalent; Cell Membrane - metabolism; Epidermal Growth Factor - pharmacology; Female; Humans; Insulin - pharmacology; Kinetics; Phosphorylation; Placenta - metabolism; Pregnancy; Receptor, Insulin - metabolism; Tyrosine
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(18)33407-0

Addition of insulin to Triton-solubilized extracts of human placental membranes selectively stimulates the incorporation of 32P from [gamma-32P]ATP into an endogenous 95,000-dalton protein, which is identified as a component of the insulin receptor by immunoprecipitation. The insulin-stimulated increment in 32P is recovered largely in [32P]tyrosine after acid hydrolysis. E Epidermal growth factor (EGF) stimulates the phosphorylation of a 150,000-dalton protein in these detergent extracts. This reaction differs in several respects from the insulin-stimulated phosphorylation of the 95,000-dalton protein. Insulin-stimulated phosphorylation exhibits an absolute requirement for Mn2+ as the sole divalent cation, whereas EGF-stimulated phosphorylation is supported by Mg2+ and Co2+ as well as Mn2+. In the presence of Mn2+, insulin-stimulated phosphorylation is not detected at less than 50 microM ATP, whereas EGF-stimulated phosphorylation is well expressed at 5 microM ATP. Thus, in detergent-solubilized membrane extracts, insulin stimulates the phosphorylation of its own receptor on tyrosine residues. This reaction has enzymatic properties distinct from those of the EGF-stimulated phosphorylation in these same extracts. The role of this insulin-stimulated phosphorylation reaction in the initiation of insulin's many biologic actions merits further study.