Kinetic characterization of glutathione reductase from the malarial parasite Plasmodium falciparum. Comparison with the human enzyme.
The homodimeric flavoenzyme glutathione reductase (GR) maintains high intracellular concentrations of the antioxidant glutathione (GSSG + NADPH + H(+) <--> 2 GSH + NADP(+)). Due to its central function in cellular redox metabolism, inhibition of GR from the malarial parasite Plasmodium falciparum represents an important approach to antimalarial drug development; therefore, the catalytic mechanism of GR from P. falciparum has been analyzed and compared with the human host enzyme. The reductive half-reaction is similar to the analogous reaction with GR from other species. The oxidative half-reaction is biphasic, reflecting formation and breakdown of a mixed disulfide between the interchange thiol and GSH. The equilibrium between the E(ox)-EH(2) and GSSG-GSH couples has been modeled showing that the Michaelis complex, mixed disulfide-GSH, is the predominant enzyme form as the oxidative half-reaction progresses; rate constants used in modeling allow calculation of an K(eq) from the Haldane relationship, 0.075, very similar to the K(eq) of the same reaction for the yeast enzyme (0.085) (Arscott, L. D., Veine, D. M., and Williams, C. H., Jr. (2000) Biochemistry 39, 4711-4721). Enzyme-monitored turnover indicates that E(FADH(-))(S-S). NADP(+) and E(FAD)(SH)(2).NADPH are dominant enzyme species in turnover. Since the individual forms of the enzyme differ in their susceptibility to inhibitors, the prevailing states of GR in the cell are of practical relevance.
Bohme
C C
CC
Department of Veterans Affairs Medical Center, Ann Arbor, Michigan 48105, Center of Biochemistry, Heidelberg University, Im Neuenheimer Feld 328, D-69120 Heidelberg, Germany.
Arscott
L D
LD
Becker
K
K
Schirmer
R H
RH
Williams
C H
CH
D000818:Animals; D005980:Glutathione Reductase / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D010963:Plasmodium falciparum / Q000201:enzymology*; D011994:Recombinant Proteins / Q000378:metabolism; D013057:Spectrum Analysis
D011994:Recombinant Proteins; D005980:Glutathione Reductase