A steady-state-kinetic model for formaldehyde dehydrogenase from human liver. A mechanism involving NAD+ and the hemimercaptal adduct of glutathione and formaldehyde as substrates and free glutathione as an allosteric activator of the enzyme.
The steady-state kinetics of formaldehyde dehydrogenase from human liver have been explored. Non-linearities were obtained in v-versus-v[S] plots. It was necessary and sufficient to consider two reactants of the equilibrium mixture of formaldehyde, glutathione and their hemimercaptal adduct for a complete description of the kinetics. A random sequential reaction scheme is proposed in which adduct and beta-NAD+ are the substrates. In addition, glutathione can bind to an allosteric regulatory site and only the glutathione-containing enzyme is considered productive. Various alternative reaction models were examined but no simple alterative was superior to the model chosen. The discrimination was largely based on results of non-linear regression analysis. Several S-substituted glutathione derivatives were tested as activators or inhibitors of the enzyme, but all were without effect. Thio-NAD+, nicotinamide--hypoxanthine dinucleotide and 3-acetylpyridine-adenine dinucleotide could substitute for beta-NAD+ as the nucleotide substrate. alpha-NAD+ and ADP-ribose were competitive inhibitors with respect to beta-NAD+ and non-competitive with glutathione and the adduct. When used simultaneously, the inhibitors were linear competitive versus each other, indicating a single nucleotide-binding site or, if more than one, non-co-operative binding sites.
D000445:Aldehyde Oxidoreductases / Q000378:metabolism*; D000494:Allosteric Regulation; D004789:Enzyme Activation / Q000187:drug effects; D005557:Formaldehyde; D005978:Glutathione; D006801:Humans; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008956:Models, Chemical; D009243:NAD; D013379:Substrate Specificity; D013438:Sulfhydryl Compounds
D013438:Sulfhydryl Compounds; D009243:NAD; D005557:Formaldehyde; D000445:Aldehyde Oxidoreductases; D005978:Glutathione