SYNTHESIS OF A MIXTURE OF CYCLIC-PEPTIDES BASED ON THE BOWMAN-BIRK REACTIVE-SITE LOOP TO SCREEN FOR SERINE-PROTEASE INHIBITORS

A peptide mixture containing 21 peptide sequences has been constructed to test the Bowman-Birk inhibitor reactive-site loop motif as the bas is of inhibition for a range of serine proteases. The 21 peptides are all based on an 11 amino acid sequence designed from a Bowman-Birk lik e inhibitor reactive-site loop. Variation has been introduced at the P 1 site of the loop, which has been randomised to include all the natur al L-amino acids (except for cysteine), plus the non-natural L-amino a cids ornithine and norleucine. The mixture of peptides was screened fo r specific binding to immobilised porcine pancreatic elastase, subtili sin BPN', alpha-chymotrypsin, trypsin, anhydro-alpha-chymotrypsin and anhydrotrypsin. Five peptides from the mixture bind to alpha-chymotryp sin, two of which also bind to anhydro-alpha-chymotrypsin, and two pep tides bind trypsin, neither of which binds to anhydro-trypsin. The com petitive inhibition constants (K-i) and the rates of proteolytic hydro lysis of the individual peptides with their respective enzymes were de termined. The rates of hydrolysis were found to vary widely and show l ittle correlation with the K-i values. In the case of the alpha-chymot rypsin inhibitors, the peptides with the lowest K-i (0.1-0.05 mM) were the only peptides that bound to anhydro-alpha-chymotrypsin. However, no peptides bound to anhydrotrypsin, suggesting a fundamental differen ce in the way that alpha-chymotrypsin and trypsin are inhibited by the se cyclic peptides. (C) Munksgaard 1995.