Oxidation in H2O and D2O of 6-ethyl-5H-dibenz(c,e)azepine and 1-methylnicotinamide by aldehyde oxidase from rabbit liver.
We report the solvent hydrogen isotope effects associated with the oxidation of 6-ethyl-5H-dibenz(c,e)azepine (6-ED) and 1-methylnicotinamide (1-MN) catalyzed by aldehyde oxidase from rabbit liver using two assay methods. The first uses 2,6-dichlorophenolindophenol (DCI) as electron acceptor in an indirect assay in which the bleaching of DCI is measured as the substrate is oxidized. The second uses molecular oxygen as electron acceptor in a direct assay in which the oxidation of 1-MN to its pyridones is accompanied by an increase in absorbance at 300 nm and the oxidation of 6-ED to its lactam product is accompanied by a decrease in absorbance at 335 nm. We have found a solvent hydrogen isotope effect close to unity in the turnover number for each substrate and for each assay method. The solvent hydrogen isotope effects on kcat/Km ranged from 0.4 to 1.1. We conclude that changes in bonding of hydrogen in solvent water, including hydrolysis of or general base attack on an enzyme-intermediate complex, do not play a rate-contributing role in the maximal velocity of oxidation of 1-MN and 6-ED catalyzed by aldehyde oxidase from rabbit liver.
D042931:Aldehyde Oxidase; D000445:Aldehyde Oxidoreductases / Q000494:pharmacology*; D000818:Animals; D003903:Deuterium; D017666:Deuterium Oxide; D003984:Dibenzazepines; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008099:Liver / Q000201:enzymology; D009536:Niacinamide / Q000031:analogs & derivatives* / Q000378:metabolism; D010084:Oxidation-Reduction; D011817:Rabbits; D014867:Water
D003984:Dibenzazepines; D014867:Water; D009536:Niacinamide; C046563:6-ethyl-5H-dibenz(c,e)azepine; D003903:Deuterium; D000445:Aldehyde Oxidoreductases; D042931:Aldehyde Oxidase; D017666:Deuterium Oxide; C024058:N(1)-methylnicotinamide