Kinetics of the biotransformation of maleylacetone and chlorofluoroacetic acid by polymorphic variants of human glutathione transferase zeta (hGSTZ1-1).
Glutathione transferase zeta (GSTZ1-1) catalyzes the cis-trans isomerization of maleylacetoacetate and the biotransformation of a range of alpha-haloacids. The objective of this study was to determine the kinetics of the biotransformation of maleylacetone (MA), an analogue of the natural substrate maleylacetoacetate, and chlorofluoroacetic acid (CFA) by polymorphic variants of recombinant hGSTZ1-1. The k(cat) of the four variants of hGSTZ1-1 with MA as the substrate followed the order: 1c-1c > 1b-1b > 1d-1d > 1a-1a whereas the k(cat) for the biotransformation of CFA followed the order: 1a-1a > 1b-1b approximately 1c-1c approximately 1d-1d. The turnover rates of MA were much higher than those of CFA for each variant and ranged from 22-fold (1a-1a) to 980-fold differences (1c-1c). The catalytic efficiencies of hGSTZ1-1 variants with MA as the substrate were much greater than those with CFA as the substrate, but little difference among the polymorphic variants was observed. MA was a mixed inhibitor of all variants with CFA as substrate: the mean competitive inhibition constant (K(ic)(MA)) for all variants was about 100 microM, and the mean uncompetitive inhibition constant (K(iu)(MA)) was about 201 microM. Hence, MA and alpha-haloacids apparently compete for the same active site on the enzyme. DCA-induced inactivation of the four variants showed that the inactivated enzymes show markedly reduced isomerase activities. The residual activities were different for each variant: 1a-1a (12%) > 1b-1b approximately 1c-1c approximately 1d-1d (<5%). This is the first kinetic analysis of polymorphic variants of hGSTZ1-1, and the similarity of the kinetic constants for hGSTZ1-1 variants with either MA or CFA as substrates indicates that few differences in DCA-induced perturbations of tyrosine metabolism would likely be observed in humans.
Department of Pharmacology and Physiology, University of Rochester Medical Center, 601 Elmwood Avenue, Box 711, Rochester, New York 14642, USA.
Hoffman B M
HB
Lantum
Philip G
PG
Board
M W
MW
Anders
D000085:Acetates / Q000378:metabolism*; D000096:Acetone / Q000031:analogs & derivatives* / Q000378:metabolism*; D001711:Biotransformation; D003999:Dichloroacetic Acid / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005982:Glutathione Transferase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008298:Maleates / Q000378:metabolism*; D020641:Polymorphism, Single Nucleotide; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity; D019745:cis-trans-Isomerases / Q000037:antagonists & inhibitors
D000085:Acetates; D004791:Enzyme Inhibitors; D008298:Maleates; D011994:Recombinant Proteins; C460372:maleylacetone; D000096:Acetone; C120963:chlorofluoroacetic acid; D003999:Dichloroacetic Acid; C407705:GSTZ1 protein, human; D005982:Glutathione Transferase; D019745:cis-trans-Isomerases