Isolation and properties of a chymotrypsin inhibitor from winged bean seed (Psophocarpus tetragonolobus (L) Dc.).

A new inhibitor of bovine alpha-chymotrypsin has been isolated from winged bean seed. The inhibitor was purified to homogeneity by affinity chromatography on chymotrypsin-Sepharose, following the removal of the trypsin inhibitors on trypsin-Sepharose. The inhibitor has a molecular weight of approx. 21,000 and amino acid analysis showed that it contains four half-cystine residues, lacks methionine, and is rich in aspartic acid, glutamic acid, valine and leucine. The inhibitor does not inhibit bovine trypsin in the standard inhibitor assay and does not bind to trypsin-Sepharose at ph 8.0. Inhibition data show that 1 mol of inhibitor inhibits 2 mol of alpha-chymotrypsin to form a 1:2 complex. The inhibition, however, is characterized by substrate induced dissociation of the complex and complete inhibition, even at high inhibitor concentration, is not attained. The inhibitor-chymotrypsin complex is stable at pH 8.0 and was isolated by gel-filtration on Sephadex G-100. An apparent molecular weight of approx. 70,000 was obtained for the complex, measured by gel filtration and ultracentrifugal analysis, consistent with a 1:2 molar stoichiometry.