title: Steady-state kinetic analysis of aldehyde dehydrogenase from human erythrocytes.
abstract: The steady-state kinetics of purified cytoplasmic aldehyde dehydrogenase (EC 1.2.1.3) from human erythrocytes have been studied at 37 degrees C. Previous studies of the enzyme from several mammalian sources, which used a lower assay temperature, have been difficult to interpret because of the substrate activation by acetaldehyde which led to complex kinetic behaviour. At 37 degrees C the initial-rate data do not depart significantly from Michaelis-Menten kinetics. Studies of the variation of initial rates as a function of the concentrations of both substrates and studies of the inhibition by NADH were consistent with a sequential mechanism being followed. High-substrate inhibition by acetaldehyde was competitive with respect to NAD+. The enzyme was not inhibited by the product acetate and thus the results of these studies, although consistent with an ordered mechanism in which NAD+ was the first substrate to bind, were inconclusive. That such a mechanism was followed was confirmed by determination of the initial-rate behaviour in the presence of acetaldehyde and glycolaldehyde as alternative substrates. When the reciprocal of the initial rate of NADH formation was plotted against the acetaldehyde concentration at a series of fixed ratios between that substrate and glycolaldehyde, a linear 'mixed inhibition' pattern was obtained, confirming the mechanism to be ordered with NAD+ being the leading substrate and with kinetically significant ternary complex-formation.
authors.affiliation: Department of Biochemistry, Trinity College, Dublin, Ireland.
authors.forename: G T
authors.initials: GT
authors.lastname: Henehan
authors.forename: K F
authors.initials: KF
authors.lastname: Tipton
mesh_terms: D000079:Acetaldehyde / Q000378:metabolism; D000444:Aldehyde Dehydrogenase / Q000037:antagonists & inhibitors / Q000097:blood*; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans; D007700:Kinetics; D009243:NAD / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D009243:NAD; D000444:Aldehyde Dehydrogenase; D000079:Acetaldehyde