These tools will no longer be maintained as of December 31, 2024. Archived website can be found here. PubMed4Hh GitHub repository can be found here. Contact NLM Customer Service if you have questions.


Pubmed for Handhelds

PUBMED FOR HANDHELDS

Search MEDLINE/PubMed

  • Title: Determination of the kinetic and chemical mechanism of malic enzyme using (2R,3R)-erythro-fluoromalate as a slow alternate substrate.
    Author: Urbauer JL, Bradshaw DE, Cleland WW.
    Journal: Biochemistry; 1998 Dec 22; 37(51):18026-31. PubMed ID: 9922171.
    Abstract:
    (2R,3R)-erythro-Fluoromalate, but not the threo isomer, is a slow substrate for chicken liver malic enzyme with either NADP or 3-acetylpyridine-NADP (APADP) as the other substrate. The Km for erythro-fluoromalate is similar to that of malate, but the turnover number with NADP is 3300-fold slower, although 5.5-fold faster with APADP than with NADP. Deuteration of fluoromalate at C-2 gave an isotope effect on V/K of 1.39 with NADP and 3.32 with APADP. With NADP, the 13C isotope effects at C-4 were 1.0490 with unlabeled and 1.0364 with deuterated fluoromalate. With APADP, the corresponding values were 1.0138 and 1.0087. These data show that the mechanism is stepwise with both nucleotide substrates, in contrast to the reaction of malate and APADP, which was postulated to be concerted by Karsten et al. [Karsten, W. E., and Cook, P. F. (1994) Biochemistry 33, 2096-2103], a conclusion recently shown to be correct by Edens et al. [Edens, W. A., Urbauer, J. L., and Cleland, W. W. (1997) Biochemistry 36, 1141-1147]. To explain the effect of deuteration on the 13C isotope effect with APADP, it is necessary to assume a secondary 13C isotope effect at C-4 on the hydride transfer step of approximately 1.0064 (assuming 5.7 as the intrinsic primary deuterium isotope effect and 1.054 as the product of the 13C equilibrium isotope effect on hydride transfer and the intrinsic 13C isotope effect on decarboxylation). The secondary 13C isotope effect on hydride transfer is thought to result from hyperconjugation between the carbonyl group and C-4 of the enzyme-bound fluorooxaloacetate intermediate.
    [Abstract] [Full Text] [Related] [New Search]