title: Glutathione reductase from Saccharomyces cerevisiae undergoes redox interconversion in situ and in vivo.
abstract: Redox interconversion of glutathione reductase was studied in situ with S. cerevisiae. The enzyme was more sensitive to redox inactivation in 24 hour-starved cells than in freshly-grown ones. While 5 microM NADPH or 100 microM NADH caused 50% inactivation in normal cells in 30 min, 0.75 microM NADPH or 50 microM NADH promoted a similar effect in starved cells. GSSG reactivated the enzyme previously inactivated by NADPH, ascertaining that the enzyme was subjected to redox interconversion. Low EDTA concentrations fully protected the enzyme from NADPH inactivation, thus confirming the participation of metals in such a process. Extensive inactivation was obtained in permeabilized cells incubated with glucose-6-phosphate or 6-phosphogluconate, in agreement with the very high specific activities of the corresponding dehydrogenases. Some inactivation was also observed with malate, L-lactate, gluconate or isocitrate in the presence of low NADP+ concentrations. The inactivation of yeast glutathione reductase has also been studied in vivo. The activity decreased to 75% after 2 hours of growth with glucono-delta-lactone as carbon source, while NADPH rose to 144% and NADPH+ fell to 86% of their initial values. Greater changes were observed in the presence of 1.5 microM rotenone: enzymatic activity descended to 23% of the control value, while the NADH/NAD+ and NADPH/NADP+ ratios rose to 171% and 262% of their initial values, respectively. Such results indicate that the lowered redox potential of the pyridine nucleotide pool existing when glucono-delta-lactone is oxidized promotes in vivo inactivation of glutathione reductase.
authors.affiliation: Departamento de Bioquímica y Biología Molecular, Unidad de Veterinaria, Universidad de Córdoba, Spain.
authors.forename: J
authors.initials: J
authors.lastname: Peinado
authors.forename: J
authors.initials: J
authors.lastname: Florindo
authors.forename: J
authors.initials: J
authors.lastname: López-Barea
mesh_terms: D002463:Cell Membrane Permeability; D005980:Glutathione Reductase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D007700:Kinetics; D010084:Oxidation-Reduction; D011725:Pyridines / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology*
chemical_list: D011725:Pyridines; D005980:Glutathione Reductase