Characterization of a Bowman–Birk type trypsin inhibitor purified from seeds of Solanum surattense

Abstract

A Bowman–Birk type trypsin inhibitor protein (SSTI) from seeds of the medicinal plant Solanum surattense was isolated, purified and characterized. SSTI showed a single band on SDS-PAGE corresponding to 11.4 kDa molecular weight. It is a glycoprotein (2.8% glycosylation) that differentially interacted with trypsin and chymotrypsin in a concentration-dependent manner. Its peptide sequence is similar to other Bowman–Birk type protease inhibitors found in Glycine max and Phaseolus acutifolius. The inhibitory activity was stable over a wide range of pH (1–10) and temperatures (10–100° C). Far-UV Circular Dichroism (CD) studies showed that SSTI contains β sheets (~ 23%) and α helix (~ 6%) and demonstrated structural stability at wide pH and high temperature. The kinetic analysis revealed a noncompetitive (mixed) type nature of SSTI and low inhibitor constant (Ki) values (16.6 × 10−8 M) suggested strong inhibitory activity. Isothermal titration calorimetric analysis revealed its high affinity towards trypsin with dissociation constant (Kd) 2.28 µM.

Comparative Binding Affinity Analysis of Soybean Meal Bowman-Birk and Kunitz Trypsin Inhibitors in Interactions with Animal Serine Proteases

“…In the case of soybean BBTI, interactions with chymotrypsin show a K D of ∼3 μM . Likewise, binding of Solanum surattense BBTI with bovine trypsin displays a K D of 2.3 μM . The discrepancy in affinity with our data can be attributed to several factors, including the procedure for obtaining soybean meal and the composition and ratio of the BBTI polymorphic variants.…”