The two α2-Macroglobulin-bound trypsin molecules have different affinities for the basic pancreatic trypsin inhibitor

Abstract

We have investigated the enzymatic properties of α 2-macroglobulin-bound porcine trypsin using a substrate: Z-Gly-Gly-Arg- p-nitroanilide and inhibotors; p-aminobenzamidine and basic pancreatic trypsin inhibitor. The ternary α 2-macroglobulin-(trypsin) 2 complex behaves like a mixture of two enzymes which bind basic pancreatic trypsin inhibitor with widely different ffinities ( K i = 0.11 μM and 23 μM). About one-half of the trypsin molecules of the ternary complex are covalently bound to α 2-macroglobulin. Preparation of the complex in the presence of hydroxylamine prevents covalent bond formation, but the two trypsins of this artificial complex still exhibit large differences in affinity for basic pancreatic trypsin inhibitor. The trypsin molecules of the ternary complex also exhibit small differences in their affinity for Z-Gly-Gly-Arg- p-nitroanilide and p-aminobenzamidine.