title: Time-dependent inhibition of protein farnesyltransferase by a benzodiazepine peptide mimetic.
abstract: Protein farnesyltransferase (FTase) and protein geranylgeranyltransferase-I (GGTase-I) catalyze the prenylation of proteins with a carboxy-terminal tetrapeptide sequence called a CaaX box, where C refers to cysteine, "a" refers to an aliphatic residue, and X typically refers to methionine, serine, or glutamine (FTase), or to leucine (GGTase-I). Marsters and co-workers [(1994) Bioorg. Med. Chem. 2, 949--957] developed inhibitors of FTase with cysteine and methionine attached to an inner hydrophobic benzodiazepine scaffold. We found that the most potent of these compounds (BZA-2B) resulted in the time-dependent inhibition of FTase. The K(i) of BZA-2B for FTase, which is the dissociation constant of the initial complex, was 79 +/- 13 nM, and the K(i)*, which is the overall dissociation of inhibitor for all enzyme forms, was 0.91 +/- 0.12 nM. The first-order rate constant for the conversion of the initial complex to the final complex was 1.4 +/- 0.2 min(-1), and that for the reverse process was 0.016 +/- 0.002 min(-1). The latter rate constant corresponds to a half-life of the final complex of 45 min. Our experiments favor the notion that the inhibitor binds to the FTase--farnesyl diphosphate complex which then undergoes an isomerization to form a tighter FTase*--farnesyl diphosphate--BZA2-B complex. Diazepam, a compound with a benzodiazepine nucleus but lacking amino acid extensions, was a weak (K(i) = 870 microM) but not time-dependent inhibitor of FTase. Cys-Val-Phe-Met and Cys-4-aminobenzoyl-Met were instantaneous and not time-dependent inhibitors of FTase. Furthermore, BZA-4B, with a leucine specificity determinant, was a classical competitive inhibitor of GGTase-I and not a time-dependent inhibitor.
authors.affiliation: Department of Biochemistry and Molecular Biology, Louisiana State University Health Sciences Center, New Orleans, Louisiana 70119, USA. biocrr@lsuhsc.edu
authors.forename: R
authors.initials: R
authors.lastname: Roskoski
authors.forename: P A
authors.initials: PA
authors.lastname: Ritchie
mesh_terms: D019883:Alkyl and Aryl Transferases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D000818:Animals; D001569:Benzodiazepines / Q000378:metabolism / Q000494:pharmacology*; D001667:Binding, Competitive; D002478:Cells, Cultured; D004791:Enzyme Inhibitors / Q000378:metabolism / Q000494:pharmacology*; D051231:Farnesyltranstransferase; D007700:Kinetics; D008956:Models, Chemical; D009842:Oligopeptides / Q000378:metabolism / Q000494:pharmacology*; D017368:Protein Prenylation; D051381:Rats; D018411:Spodoptera; D013379:Substrate Specificity; D013997:Time Factors
chemical_list: C437904:BZA-2B; D004791:Enzyme Inhibitors; D009842:Oligopeptides; D001569:Benzodiazepines; C088459:B 581; D019883:Alkyl and Aryl Transferases; C083291:geranylgeranyltransferase type-I; D051231:Farnesyltranstransferase