The reversible receptor binding of insulin in isolated rat adipocytes measured at 37 degrees C. The binding is not rate limiting for cellular uptake.

The bimolecular binding reaction between mono[TyrA14-125I]iodoinsulin and the insulin receptor was investigated at 37 degrees C in intact isolated rat adipocytes in which membrane traffic was inhibited by 1 mM KCN. This treatment decreased the fraction of cell-associated radioactivity resistant to treatment at pH 3 (usually regarded as internalized ligand) from 70% to 17%. The total amount of tracer being cell-associated at steady state was reduced to about half of the control value partly because of a decreased apparent binding affinity. The t1/2 for the forward reaction was reduced from 414 s in the control cell to 26 s in the KCN treated cell. Likewise, the t1/2 for the dissociation was reduced from 461 s to 67 s. Both rate constants were pH sensitive, the association rate constant being 7-8-fold more than the dissociation rate constant. Since both rate constants for the bimolecular reaction were one order of magnitude greater than those for the uptake and the release of label in the untreated cell, other processes than binding constitute the rate-limiting step(s) in the cellular reaction with insulin.