LIGAND-INDUCED CONFORMATIONAL CHANGE IN PENICILLIN ACYLASE

The enzyme penicillin acylase (penicillin amidohydrolase EC 3.5.1.11) catalyses the cleavage of the amide bond in the benzylpenicillin (peni cillin G) side-chain to produce phenylacetic acid and 6-aminopenicilla nic acid (6-APA). The enzyme is of great pharmaceutical importance, as the product 6-APA is the starting point for the synthesis of many sem i-synthetic penicillin antibiotics. Studies have shown that the enzyme is specific for hydrolysis of phenylacetamide derivatives, but is mor e tolerant of features in the rest of the substrate. It is this proper ty that has led to many other applications for the enzyme, and greater knowledge of the enzyme's structure and specificity could facilitate engineering of the enzyme, enhancing its potential for chemical and in dustrial applications. An extensive study of the binding of a series o f phenylacetic acid derivatives has been carried out. A measure of the relative degree of inhibition of the enzyme by each of the compounds has been obtained using a competitive inhibition assay, and the struct ures of a number of these complexes have been determined by X-ray crys tallography. The structures reveal a clear rationale for the observed kinetic results, but show also that some of the ligands cause a confor mational change within the binding pocket. This change can generally b e understood in terms of the size and orientation of the ligand within the active site.The results reveal that ligand binding in penicillin acylase is facilitated by certain amino acid residues that can adopt t wo distinct, energetically favourable positions in order to accommodat e a variety of compounds within the active site. The structures of the se complexes provide evidence for conformational changes in the substr ate-binding region that may act as a switch in the mechanism of autoca talytic processing of this enzyme. (C) 1998 Academic Press.