Serine protease inhibition by insect peptides containing a cysteine knot and a triple-stranded β-sheet

Abstract

Three insect peptides showing high sequence similarity and belonging to the same structural family incorporating a cysteine knot and a short three- stranded antiparallel β-sheet were studied. Their inhibitory effect on two serine proteases (bovine α-chymotrypsin and human leukocyte elastase) is reported. One of them, PMP-C, is a strong α-chymotrypsin inhibitor (K(i) = 0.2 nM) and interacts with leukocyte elastase with a K(i) of 0.12 μM. The other two peptides, PMP-D2 and HI, interact only weakly with α-chymotrypsin and do not inhibit leukocyte elastase. Synthetic variants of these peptides were prepared by solid-phase synthesis, and their action toward serine proteases was evaluated. This enabled us to locate the P1 residues within the reactive sites (Leu-30 for PMP-C and Arg-29 for PMP-D2 and HI), and, interestingly, variants of PMP-D2 and HI were converted into powerful inhibitors of both α-chymotrypsin and leukocyte elastase, the most potent elastase inhibitor obtained in this study having a K(i) of 3 nM.