Insulin Binding and Internalization in Hagfish Red Blood Cells

Abstract

Binding of porcine 125I-insulin (0.15 nM) to hagfish red blood cells was time-dependent, reaching equilibrium after 1 hr at 10°. The specific 125I-Insulin bound to receptors from a T1/2 of 60 min in cells suspended in medium alone to 23 min in medium containing 8 μM nonradioactive insulin. Porcine insulin and desoctapeptide insulin competed for specific binding of 125I-insulin in a dose-dependent manner, whereas glucagon and somatostatin did not. For porcine insulin. Scatchard analysis produced a curvilinear plot, suggesting multiple affinity binding sites with high-affinity and low-affinity association constant (Ka) 0.2 × 109M-1 and 0.27 × 107M-1 respectively. A total of 2090 binding sites per hagfish red blood cell was calculated. Sixty-two percent of the bound 125I-insulin was found to be internalized into the hagfish red blood cells. Less degradation of 125I-insulin was observed by Sephadex G-50 chromatography compared to human red blood cells.