ACETALDEHYDE OXIDATION CATALYZED BY ALDEHYDE DEHYDROGENASE (CYANAMIDE, ALCOHOL, ETHANOL)
Abstract
Two factors can affect the rate of acetaldehyde oxidation by aldehyde dehydrogenase (ALDH) in rat liver: the rate of regeneration of NAD by the electron transport system and the level of ALDH activity in the cell. A respiration uncoupler did not affect the rate of acetaldehyde metabolism by liver slices, indicating that NADH re-oxidation was not rate-limiting. The possibility of ALDH activity being in excess was examined by treating rat liver slices and mitochondria with an irreversible inhibitor of ALDH, cyanamide, and then measuring the effect of this treatment on the rate of acetaldehyde metabolism. In as much as relatively little was known about the mechanism by which cyanamide inactivates the enzyme, the interaction of inhibitor and ALDH was partially characterized. Cyanamide at low concentrations (5 uM) required enzymatic conversion, probably by catalase, to a reactive derivative in order to inhibit ALDH activity. Data from protection experiments was consistent with cyanamide binding to the same site on ALDH as does the aldehyde substrate. Since affinity-purified ALDH, free of measurable catalase activity, was inactivated by high concentrations (185 uM) of cyanamide, an alternate pathway for ALDH inactivation may exist. A small number of rats were encountered whose mitochondrial low-Km ALDH was not inactivated by low concentrations of cyanamide. This resistance to cyanamide inhibition was apparently due to the presence of different ALDH isozymes. Inhibition of low-K(,m) ALDH activity in liver slices or liver mitochondria by cyanamide caused a decrease in the rate of acetaldehyde metabolism, indicating that no excess of low-K(,m) ALDH existed. It was also established that low-K(,m) ALDH activity was rate-limiting for acetaldehyde oxidation during concomitant ethanol oxidation by the liver slice. Approximately 40% of the metabolism of 200 uM acetaldehyde did not involve low K(,m) ALDH was capable of catalyzing almost half of this residual acetaldehyde metabolism. Formation of acetaldehyde-protein adducts could account for some of the acetaldehyde disappearance, and the present data is consistent with acetaldehyde primarily binding to cytosolic constituents.
Degree
Ph.D.
Subject Area
Biochemistry
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