ICAR Research Data Repository for Knowledge Management

The major proteinase inhibitor in horse gram (Dolichos biflorus) is a low molecular weight ~8500 Bowman–Birk inhibitor BBI, HGI-III, that inhibits both trypsin and chymotrypsin simultaneously. Analysis of the reactivity of the polyclonal antibodies raised against native HGI-III, with tryptic, lysylendoproteinase-C and CNBr peptides, in dot-blot assays, revealed the presence of three sequential epitopes (Asp¹–Lys¹⁴ (I), Leu³⁷–Lys⁶³ II and Asp⁵⁴–Lys⁷¹ (III)). Of
these, epitope II and III occur consecutively in the sequence of HGI-III. The reactive site peptide bonds were identified by cleavage with catalytic quantities of either trypsin or chymotrypsin at acidic pH. The reactive site peptide bond for trypsin was found to be Lys²⁴–Ser²⁵, whereas for chymotrypsin it was Phe⁵¹–Ser⁵². The highly conserved reactive site loop residues of the Bowman–Birk inhibitors are also conserved in HGI-III. The less immunogenic peptide sequence,Leu³⁷–Lys⁶³, also contains the chymotrypsin reactive site. The recognition of this polypeptide by the immune system provides for a new strategy in the design of ideal, smaller proteinase inhibitors as cancer preventive agents.