Protein farnesyltransferase: kinetics of farnesyl pyrophosphate binding and product release.
Journal Article (Journal Article)
Protein farnesyltransferase (FTase) catalyzes the prenylation of Ras and several other key proteins involved in cell regulation. The mechanism of the FTase reaction was elucidated by pre-steady-state and steady-state kinetic analysis. FTase catalyzed the farnesylation of biotinylated peptide substrate (BiopepSH) by farnesyl pyrophosphate (FPP) to an S-farnesylated peptide (BiopepS-C15). The steady-state kinetic mechanism was ordered. FTase bound FPP in a two-step process with an effective dissociation rate constant of 0.013 s-1 and an overall Kd of 2.8 nM. BiopepSH reacted with FTase.FPP irreversibly, with a second-order rate constant of 2.2 x 10(5) M-1 s-1, to form FTase.BiopepS-C15. Because most of the FPP in FTase.FPP was trapped as FTase.BiopepS-C15 at high concentrations of BiopepSH, FPP dissociated slowly from the ternary complex relative to catalysis, so that the commitment to catalysis was high. The maximal rate constant for formation of FTase.BiopepS-C15 (enzyme-bound product) is much larger than kcat (0.06 s-1), indicating that product release is the rate-determining step in the reaction mechanism.
Full Text
Duke Authors
Cited Authors
- Furfine, ES; Leban, JJ; Landavazo, A; Moomaw, JF; Casey, PJ
Published Date
- May 23, 1995
Published In
Volume / Issue
- 34 / 20
Start / End Page
- 6857 - 6862
PubMed ID
- 7756316
International Standard Serial Number (ISSN)
- 0006-2960
Digital Object Identifier (DOI)
- 10.1021/bi00020a032
Language
- eng
Conference Location
- United States