Isolation of the Insulin Receptor of Liver and Fat-Cell Membranes
- 1 February 1972
- journal article
- Published by Proceedings of the National Academy of Sciences in Proceedings of the National Academy of Sciences of the United States of America
- Vol. 69 (2), 318-322
- https://doi.org/10.1073/pnas.69.2.318
Abstract
Extraction of liver and fat-cell membranes with the nonionic detergent Triton X-100 prevents specific binding of (125)I-labeled insulin to these membranes. This loss of binding to particulate material is quantitatively recovered in a high-speed (300,000 x g, 2 hr) supernatant of the extract. Specific and reversible insulin binding to soluble proteins is readily demonstrable by gel filtration. A simple and sensitive assay for detection of specific macromolecule-insulin complexes has been developed based on the selective precipitation of the complex by polyethylene glycol. Extraction of membrane lipids with organic solvents or by phospholipase digestion does not impair the subsequent extraction of the insulin-binding protein with detergent. Binding of insulin to the soluble protein is a saturable and dissociable process having a dissociation constant of about 100 nM. Derivatives of insulin compete for binding in direct proportion to their biological activity; other peptide hormones are without effect. The quantitative features of the detergent extractions and the specific insulin-binding properties of the material so obtained indicate that the protein solubilized is the biologically significant insulin receptor, whose insulin-binding function is essentially unaltered.Keywords
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