STUDIES ON SIMULTANEOUS INHIBITION OF TRYPSIN AND CHYMOTRYPSIN BY HORSEGRAM BOWMAN-BIRK INHIBITOR

Bowman-Birk inhibitors (BBI) isolated from plant seeds are small prote ins active against trypsin and/or chymotrypsin. These inhibitors have been extensively studied in terms of their structure, interactions, fu nction and evolution. Examination of the known three-dimensional struc tures of BBIs revealed similarities and subtle differences. The hydrop hobic core, deduced from surface accessibility and hydrophobicity plot s, corresponding to the two tandem structural domains of the double he aded BBI are related by an almost exact two-fold, in contrast to the r eactive site loops which depart appreciably from the two-fold symmetry . Also, the orientations of inhibitory loops in soybean and peanut inh ibitors were different with respect to the rigid core. Based on the st ructure of Adzuki bean BBI-trypsin complex, models of trypsin and chym otryspin bound to the monomeric soybean BBI (SBI) were constructed. Th ere were minor short contacts between the two enzymes bound to the inh ibitor suggesting near independence of binding. Binding studies reveal ed that the inhibition of one enzyme in the presence of the other is a ssociated with a minor negative cooperativity. In order to assess the functional significance of the reported oligomeric forms of BBI, bindi ng of proteases to the crystallographic and non-crystallographic dimer s as found in the crystal structure of peanut inhibitor were examined. It was found that all the active sites in these oligomers cannot simu ltaneously participate in inhibition.