Chemical modification of human aldehyde dehydrogenase by physiological substrate.
Employing 3,4-dihydroxyphenylacetaldehyde (dopal) as a substrate for human aldehyde dehydrogenase (aldehyde:NAD+ oxidoreductase, EC 1.2.1.3) in anaerobic conditions, inactivation of both cytoplasmic E1 and mitochondrial E2 isozymes during catalysis has been observed. Incorporation of 14C-labelled dopal has been demonstrated by retention of label following denaturation and exhaustive dialysis and by peptide mapping following tryptic digestion. Incorporation of label gave linear plots vs. activity remaining with up to two molecules incorporated per molecule of enzyme and 30% activity remaining. Further incorporation (up to 16 molecules) occurred, but was non-linear when plotted vs. activity remaining. Protection against activity loss during incorporation of the first two molecules was afforded by NAD, NADH, chloral, and by chloral and NAD together, the last being the most effective. Saturation kinetics gave y-axis intercepts, suggesting interaction at a specific point on the enzyme surface. The Ki value from saturation kinetics was the same as that from the slope replot in catalytic reaction. Peptide mapping of tryptic digests showed that a single peptide was labelled, confirming specificity of interaction. Even in the absence of complete inactivation, the results suggest that reaction with the first two molecules occurs at some point on the enzyme surface important for enzyme activity. The possibility of such a reaction occurring in vivo is discussed.
D015102:3,4-Dihydroxyphenylacetic Acid / Q000031:analogs & derivatives / Q000378:metabolism*; D000444:Aldehyde Dehydrogenase / Q000378:metabolism*; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002697:Chloral Hydrate / Q000031:analogs & derivatives / Q000378:metabolism; D002942:Circular Dichroism; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D007700:Kinetics; D009243:NAD / Q000378:metabolism; D010648:Phenylacetates / Q000378:metabolism*
D007527:Isoenzymes; D010648:Phenylacetates; D009243:NAD; D015102:3,4-Dihydroxyphenylacetic Acid; D002697:Chloral Hydrate; C007430:3,4-dihydroxyphenylacetaldehyde; D000444:Aldehyde Dehydrogenase; C021100:trichloroacetaldehyde