ROLE OF THE CARBOXYTERMINAL RESIDUE IN PEPTIDE BINDING TO PROTEIN FARNESYLTRANSFERASE AND PROTEIN GERANYLGERANYLTRANSFERASE

Protein farnesyltransferase and protein geranylgeranyltransferase-I ca talyze the prenylation of a cysteinyl group located four residues upst ream of the carboxyl terminus. The identity of the carboxyterminal res idue plays a significant role in determining the ability of compounds to bind to each enzyme and to serve as substrate. We compared the bind ing and substrate specificities of peptides with carboxyterminal subst itutions to determine which residues promote selectivity and which res idues promote recognition by both enzymes. Using tetrapeptide inhibito rs with the general structure L-penicillamine-valine-isoleucine-X and substrates with the structure Lys-Lys-Ser-Ser-Cys-Val-Ile-X, we measur ed their respective K-i, K-m, and k(cat) values for both recombinant r at protein farnesyltransferase and recombinant rat protein geranylgera nyltransferase-I. We studied the roles of carboxyterminal branched res idues (leucine, isoleucine, valine, and penicillamine) and linear resi dues (methionine, cysteine, homocysteine, alanine, aminobutyrate, and aminohexanoate) in promoting interaction with the enzymes. For protein geranylgeranyltransferase-I, peptide substrates with carboxyterminal branched or linear residues had K-m values that were 5- to 15-fold gre ater than the K-i values of the corresponding peptide inhibitors. For protein farnesyltransferase, peptide substrates with carboxyterminal b ranched residues, proline, or homoserine had K-m values that were 7- t o 200-fold greater than the K-i values of the corresponding peptide in hibitors. For protein farnesyltransferase the K-m and K-i values for p eptides ending with linear residues were in general agreement, Our stu dies indicate that the substrate and inhibitor binding specificities o f protein geranylgeranyltransferase was much more restricted than thos e of protein farnesyltransferase. (C) 1998 Academic Press.