Binding and degradation of insulin by human peripheral granulocytes. Demonstration of specific receptors with high affinity.

摘要: Abstract The interaction of insulin with human circulating granulocytes was studied the use 125I-insulin. Human granulocytes, isolated from blood by Boyum technique, showed high insulin-degrading activity in vitro which almost obscured presence specific, affinity binding sites. Degradation, measured trichloroacetic acid precipitation and to well characterized receptors on cultured lymphocytes (IM-9 line), due extracellular as cell-bound enzymes. Degradation enhanced Ca2+ thiols inhibited various protease inhibitors sulfhydryl-blocking reagents. Phenylmethylsulfonyl fluoride (5 X 10(-4) M), a serine inhibitor, most potent 125I-insulin degradation 80 90%. Tert-butyl hydroperoxide (2 10(-3) glutathione-oxidizing reagent, 35 50%, possibly an effect glutathione-insulin transhydrogenase. Neither affected cell viability. In degradation, sites for were detected, multiple criteria true receptors. Binding these rapid, saturable, reversible about 1000 sites/cell. Hill coefficient 0.7, Scatchard plot B/F versus B curvilinear, site-site interactions negative cooperative type; latter demonstrated directly kinetic studies. As shown previously all other receptors, highly pH-dependent, analogues had affinities that closely correlated their biological potencies.