Purification and Characterization of a Proteinase Inhibitor from Field Bean, Dolichos lablab perpureus L.

Abstract

A proteinase inhibitor resembling Bowman-Birk family inhibitors has been purified from the seeds of cultivar HA-3 of Dolichos lablab perpureus L. The protein was apparently homogeneous as judged by SDS-PAGE, PAGE, IEF, and immunodiffusion. The inhibitor had 12 mole% 1/2-cystine and a few aromatic amino acids, and lacks tryptophan. Field bean proteinase inhibitor (FBPI) exhibited a pl of 4.3 and an M r of 18,500 Da. CD spectral studies showed random coiled secondary structure. Conformational changes were detected in the FBPI-trypsin/chymotrypsin complexes by difference spectral studies. Apparent K a values of complexes of inhibitor with trypsin and chymotrypsin were 2.1 X 10 7 M -1 and 3.1 X 10 7 M -1 , respectively. The binary and ternary complexes of FBPI with trypsin and chymotrypsin have been isolated indicating 1:1 stoichiometry with independent sites for cognate enzymes. Amino acid modification studies showed lysine and tyrosine at the reactive sites of FBPI for trypsin and chymotrypsin, respectively.

Mechanistic study on the anti‐proinflammatory activity of Kunitz type inhibitor fromCaesalpinia decapetalaseeds

“…Serine PIs have been identified and described from a variety of plant sources, including the BBI and KTI families 19,20 . Based on molecular weight obtained by SDS‐PAGE and N‐terminal sequence of MALDI‐TOF (Figure 2C) gives clear evidence to support purified CdTI belongs to the KTI family and has protease inhibitory activity of 330 TIU/mg and 250 CTIU/mg of protein.…”