Role of the carboxyterminal residue in peptide binding to protein farnesyltransferase and protein geranylgeranyltransferase.

Protein farnesyltransferase and protein geranylgeranyltransferase-I catalyze the prenylation of a cysteinyl group located four residues upstream of the carboxyl terminus. The identity of the carboxyterminal residue plays a significant role in determining the ability of compounds to bind to each enzyme and to serve as substrate. We compared the binding and substrate specificities of peptides with carboxyterminal substitutions to determine which residues promote selectivity and which residues promote recognition by both enzymes. Using tetrapeptide inhibitors with the general structure l-penicillamine-valine-isoleucine-X and substrates with the structure Lys-Lys-Ser-Ser-Cys-Val-Ile-X, we measured their respective Ki, Km, and kcat values for both recombinant rat protein farnesyltransferase and recombinant rat protein geranylgeranyltransferase-I. We studied the roles of carboxyterminal branched residues (leucine, isoleucine, valine, and penicillamine) and linear residues (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 Km values that were 5- to 15-fold greater than the Ki values of the corresponding peptide inhibitors. For protein farnesyltransferase, peptide substrates with carboxyterminal branched residues, proline, or homoserine had Km values that were 7- to 200-fold greater than the Ki values of the corresponding peptide inhibitors. For protein farnesyltransferase the Km and Ki values for peptides ending with linear residues were in general agreement. Our studies indicate that the substrate and inhibitor binding specificities of protein geranylgeranyltransferase was much more restricted than those of protein farnesyltransferase.

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