Subtilisin inhibitor from seeds of broad bean (vicia FABA); purification, amino acid sequence and specificity of inhibition

Abstract

A potent inhibitor of microbial serine proteases, including subtilisin, has been purified 1100-fold from seeds of broad bean (Vicia faba, cv. Kleine Thuringer). Chymotrypsin and trypsin were not inhibited, but a weak “temporary” inhibition of pancreas elastase was observed. The preparation of pure inhibitor contained one major molecular form with a blocked N-terminal (MW ∼ 10,000, pl ∼ 4.8) and a minor form (< 10%) with slightly lower MW and pI ∼ 5.0. About 80% of the amino acid sequence of the major form was determined by automatic Edman degradation of a cyanogen bromide fragment and tryptic peptides. The inhibitor is homologous with barley, potato and leech inhibitors of the “potato inhibitor I family” (36–56% of the amino acid residues in identical positions). Cleavage studies suggest one enzyme inhibitory site at an Ala-Asp bond.