Alkylation, reduction, solubilization and enrichment of binding activity do not impair the ability of insulin receptors to convert from a rapid- into a slow-dissociating state.

Hormone binding promotes conversion of rat hepatic insulin receptors from a rapid-dissociating into a slow-dissociating affinity state. Solubilization into detergent does not impair the ability of receptors to convert into a slow-dissociating state, and this property is retained as receptor-binding activity is enriched 11,000-13,000-fold during purification. Hormone binding also induces two conformational changes (alterations of tryptic lability) in the insulin receptor. The first change is rapid and exposes parts of the receptor to tryptic degradation. The second, slower, change occurs with the same time course, and probably mediates the conversion into the slower-dissociating binding state. Reduction of disulphide bonds with dithiothreitol does not prevent conversion of binding sites into a slower-dissociating state, and reduced receptors retain the ability to undergo conformational changes in response to hormone binding. Alkylation with N-ethylmaleimide also does not affect the insulin-induced conversion into a slow-dissociating state. These observations suggest that the conversion into a slow-dissociating state is an intrinsic property of the insulin receptor. Free thiol groups in the insulin receptor and disulphide bonds between the alpha-subunits are not essential to this process.