Kinetic behaviour and properties of aldehyde dehydrogenase from rat testis mitochondria. Effect of Mg2+ ions.
1. Mitochondrial aldehyde dehydrogenase is purified to near homogeneity by hydroxylapatite-, affinity- and hydrophobic interaction-chromatography. 2. The enzyme is an oligomeric protein and its molecular weight, as determined by gel-filtration, is 117,000 +/- 5000. 3. Active only in the presence of exogenous sulfhydryl compounds and NAD(+)-dependent, aldehyde dehydrogenase works optimally with linear-chain aliphatic aldehydes and is practically inactive with benzaldehyde. The pH-optimum is at about pH 8.5. 4. Km-Values for aliphatic aldehydes (C2-C6) range between 0.17 and 0.32 microM. The Km for NAD+ increases from 16 microM with acetaldehyde to 71 microM with capronaldehyde. 5. Millimolar concentrations of Mg2+ promote high increases of both V and Km for NAD+. At the same time, saturation curves with C4-C6 aldehydes can be simulated with a substrate inhibition model. 6. Inhibition by NADH is competitive: with capronaldehyde, the inhibition constant for NADH is 52 microM in the absence of Mg2+ and 14 microM in the presence of 4 mM Mg2+; with acetaldehyde, the inhibition constant is about three times higher (36 and 159 microM, respectively).
Bedino
S
S
Dipartimento di Medicina e Oncologia Sperimentale, Università di Torino, Italy.
Testore
G
G
Obert
F
F
D000444:Aldehyde Dehydrogenase / Q000187:drug effects / Q000302:isolation & purification / Q000502:physiology*; D000447:Aldehydes / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D004795:Enzyme Stability; D005260:Female; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology*; D008297:Male; D008928:Mitochondria / Q000201:enzymology*; D008970:Molecular Weight; D009243:NAD / Q000494:pharmacology; D009249:NADP / Q000494:pharmacology; D010084:Oxidation-Reduction; D051381:Rats; D017208:Rats, Wistar; D013696:Temperature; D013737:Testis / Q000201:enzymology* / Q000648:ultrastructure
D000447:Aldehydes; D009243:NAD; D009249:NADP; C010463:n-hexanal; D000444:Aldehyde Dehydrogenase; D008274:Magnesium