NADP⁺‐dependent aldehyde dehydrogenase from ‘Acetobacter rancens’ CCM 1774: Purification and properties

Abstract

NADP+-dependent aldehyde dehydrogenase from the cytosolic fraction of the alkane-degradating 'Acelobacler rancens' CCM 1774 was purified 112-fold (specific activity of 112 nkat mg-1 protein). After polyacrylamide gel electrophoresis of the purified enzyme only one band was visible. The relative molecular weight was estimated to be 82,000 by both gel filtration and disc gel electrophoresis. The substrate specifity of the purified enzj me within the straight chain aliphatic aldehyde series is confined to acetaldehyde and propionaldehyde. Butyraldehyde and formaldehyde are considerably less converted. NADP+ alone served as electron acceptor. Mg*+ stimulated the reaction velocity in a strong manner by 'non-competitive' activation. The estimation of kinetic ammeters and inhibition experiments of the irreversible oxidation of ethanal indicate a random k e t i c mechanism a t optimal aldehyde concentrations and saturating NADP+ concentrations a t optimal pH 8.5 which, however, trends to bgcome ordered with decreasing pK values. Properties described exclude the participation of the enzyme in degradation of n-alkanes. One physiological function of this constitutive aldehyde dehydrogenase may be the intracellular detoxification of short chain aldehydes by conversion to corresponding fatty acids.Cytoplasmic aldehyde dehydrogenases of acetic acid bacteria have been niainly studied underthe view of acetic acid production (NAKAYAMA 1960, ADACHI et aZ. 1980, MURAOKA et al. 1980). However, the participation of such enzymes in this process of coinmercial interest has been excluded by AMEYAMA and ADACHI (1982) who have shown that during vinegar production the conversion of ethanol to acetic acid via intermediary aldehyde is mediated by membrane-bound enzymes. On the other hand, various cytoplasmic aldehyde dehydrogenases with partly different substrate and coenzyme specifities have been detected (MURAOKA et d. 1980 , MIDDELHOVEN et aZ. 1983. The existence of a t least two constitutive pyridine nucleotide-dependent aldehyde dehydrogenases is known for the alkane-degrading Pseudomonas aeruginosa ( GUERRILLOT and VANDECASTEELE 1977, JACOBY 1963). However, a complete understanding of the physiological importance has not been achieved yet. All reports on bacterial aldehyde dehydrogenases lack more coniplex kinetic investigations.I n a previous paper (HOMMEL and KLEBER 1984a) we have demonstrated that 'Acetobacter rancend CCM 1774 can both grow on and oxidize long chain n-alkanes via a monoterminal pathway. Both activities, a membrane-bound one and a cytoplasmic aldehyde dehydrogenase have been detected after growth on n-hexadecane and glucose, respectively, from which the former enzyme should be involved in this metabolic sequence ( HOWEL and KLEBER 1984 b). In order t o gain more information concerning the function of the cytoplasmic enzyme in this alkane assimilating strain we purified the enzyme and studied its properties. Materials and methodsOrganism and growth conditions: 'Acetobacter r a m m ' CCM 1774 was obtained from the Cze...

The Key to Acetate: Metabolic Fluxes of Acetic Acid Bacteria under Cocoa Pulp Fermentation-Simulating Conditions

“…Additionally, A. pasteurianus is able to produce NAD(P)H ϩ H ϩ via soluble alcohol and aldehyde dehydrogenases (21,51). Among species of Acetobacter, only a soluble aldehyde dehydrogenase of Acetobacter rancens (a later synonym of A. pasteurianus), which requires NADP ϩ , has been characterized (70). Furthermore, an NAD ϩ -dependent soluble alcohol dehydrogenase was identified in A. pasteurianus (51).…”

Properties of the quinoprotein aldehyde dehydrogenase from 'Acetobacter rancens'

“…rancens' possesses two constitutive aldehyde dehydrogenases : a cytoplasmic N ADP+-dependent enzyme that is highly specific towards short-chain aldehydes (Hommel et al, 1988) and a membrane-bound dye-linked enzyme. Their relative specific activity is influenced by the carbon source, e.g.…”