DESIGN OF EFFICIENT HUMAN LEUKOCYTE ELASTASE INHIBITORS; VARIANTS OF HUMAN PANCREATIC SECRETORY TRYPSIN INHIBITOR (hPSTI)

Abstract

A set of hPSTI variants were constructed by total gene synthesis or site-specific mutagenesis, with the aim of producing human leukocyte elastase(HLE)-specific inhibitors and obtaining a better insight into the parameters effecting inhibitor/protease interaction. In the initial planning the structure of related Kazal-type inhibitors were taken into account. For subsequent protein design, models were made of HLE based on the structure of porcine pancreatic elastase, and of hPSTI based on the stucture of porcine PSTI. Models of the hPSTI/HLE and hPSTI/chymotrypsin complexes were generated by CAPD "docking". The modelled complexes could be used to rationalise a posteriori the inhibitory properties of the hPSTI variants, and to postulate the structure of better elastase inhibitors which were duly generated. Excellent specific inhibitors (Ky=1.5x107 4" M for SHEE) were obtained and the contribution of individual amino acid residues Or exchanges to the binding constant were estimated.