CHARACTERIZATION OF NEPRILYSIN (EC 3.4.24.11) S-2' SUBSITE

Neprilysin is a neutral peptidase that cleaves small peptide substrate s on the amino-side of hydrophobic amino acid residues. In the present study, we have used inhibition of nonmutated and mutated enzymes with dipeptide inhibitors and hydrolysis of the substrate [Leu(5), Arg(6)] enkephalin in order to evaluate the contribution of the S-2' subsite t o substrate and inhibitor binding. Our results suggest that (1) Arg-10 2 and Asn-542 provide major contributions to the interaction of the en zyme with the P-2' residue of the substrate, (2) the S-2' subsite is v ast and can accommodate bulky side chains, and (3) Arg-102 restricts a ccess to the S-2' subsite to some side chains such as arginine. (C) 19 97 Federation of European Biochemical Societies.