Aldehyde dismutase activity of human liver alcohol dehydrogenase

Svensson, Stefan; Lundsjö, Anders; Cronholm, Tomas; Höög, Jan‐Olov

doi:10.1016/0014-5793(96)00954-4

Cited by 40 publications

(30 citation statements)

References 25 publications

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“…The three major human liver ADHs, class I±III, were expressed as recombinant enzymes and purified to homogeneity as judged from Coomassie blue-stained SDS/polyacrylamide gels, and specific activities were in accordance with earlier results [12]. Acetaldehyde and 5-HIAL were incubated with ADH in the presence of NAD + or NADH while ethanol and 5-HTOL were incubated with ADH in the presence of NAD + .…”

Section: Steady-state Kineticssupporting

confidence: 73%

“…Human class-I g 2 g 2 , class-II and class-III ADH were recombinantly expressed in Escherichia coli using pET expression vectors (Novagen). The different recombinant enzymes were purified to homogeneity in a three-step procedure including ionexchange chromatography and gel-filtration chromatography [12]. Protein concentrations were determined by the BioRad protein assay with BSA as standard complemented with amino acid analysis on a Pharmacia LKB AlphaPlus analyzer.…”

Section: Chemicals and Enzyme Preparationsmentioning

confidence: 99%

“…The higher pH (8.8) was required for a total dissociation of the 5-HIAL±bisulfite complex. Coenzyme concentrations were 2.4 mm NAD + in oxidation assays and 0.2 mm NADH in reduction assays, except for butanal reactions where coenzyme concentrations were 10.8 mm [10,12]. Inhibition of ethanol oxidation was studied with various concentrations of acetate, 5-HIAA and NaCl against various concentrations of ethanol with saturating NAD + (2.4 mm) in 0.1 m potassium phosphate, pH 7.5.…”

Section: Kinetic Analysismentioning

confidence: 99%

“…Butanal activities were assayed by GC/MS as described previously [12]. In experiments with [1-2 H]butanal, the peaks at m/z 56, m/z 57 and m/z 58 were used for quantitation of unlabeled, monodeuterated and dideuterated butanol, respectively.…”

Section: Kinetic Analysismentioning

confidence: 99%

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Activities of human alcohol dehydrogenases in the metabolic pathways of ethanol and serotonin

Svensson¹,

Some²,

Lundsjö³

et al. 1999

European Journal of Biochemistry

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Alcohols and aldehydes in the metabolic pathways of ethanol and serotonin are substrates for alcohol dehydrogenases (ADH) of class I and II. In addition to the reversible alcohol oxidation/aldehyde reduction, these enzymes catalyse aldehyde oxidation. Class-I gg ADH catalyses the dismutation of both acetaldehyde and 5-hydroxyindole-3-acetaldehyde (5-HIAL) into their corresponding alcohols and carboxylic acids. The turnover of acetaldehyde dismutation is high (k cat = 180 min 21 ) but saturation is reached first at high concentrations (K m = 30 mm) while dismutation of 5-HIAL is saturated at lower concentrations and is thereby more efficient (K m = 150 mm; k cat = 40 min 21 ). In a system where NAD + is regenerated, the oxidation of 5-hydroxytryptophol to 5-hydroxyindole-3-acetic acid proceeds with concentration levels of the intermediary 5-HIAL expected for a two-step oxidation. Butanal and 5-HIAL oxidation is also observed for class-I ADH in the presence of NADH. The class-II enzyme is less efficient in aldehyde oxidation, and the ethanol-oxidation activity of this enzyme is competitively inhibited by acetate (K i = 12 mm) and 5-hydroxyindole-3-acetic acid (K i = 2 mm).Reduction of 5-HIAL is efficiently catalysed by class-I gg ADH (k cat = 400 min 21 ; K m = 33 mm) in the presence of NADH. This indicates that the increased 5-hydroxytryptophol/5-hydroxyindole-3-acetic acid ratio observed after ethanol intake may be due to the increased NADH/NAD + ratio on the class-I ADH.Keywords: alcohol dehydrogenase; alcohol metabolism; aldehydes; sequential oxidation; serotonin metabolism.The alcohol dehydrogenase (ADH) family is the major enzyme system for metabolism of ingested ethanol [1,2]. In addition to ethanol, a range of substrates has been identified for these enzymes and a possible mechanism for ethanol-induced metabolic changes may be interference with other activities of ADH (Fig. 1). For example, ethanol affects human noradrenaline, dopamine and serotonin metabolism by increasing the relative formation of the alcohol products, 4-hydroxy-3-methoxyphenylglycol, 3,4-dihydroxyphenylethanol and 5-hydroxytryptophol (5-HTOL), while decreasing the formation of the carboxylic acid products, 4-hydroxy-3-methoxymandelic acid, 3,4-dihydroxyphenylacetic acid and 5-hydroxyindole-3-acetic acid (5-HIAA) [3,4]. Clinical monitoring of recent drinking takes advantage of the elevated 5-HTOL/5-HIAA ratio which can be detected in urine for several hours after ethanol is no longer measurable [5,6]. The ADH activities for noradrenaline and dopamine metabolites have been characterized, and alcohols and aldehydes in these catabolic pathways serve as good substrates for ADH of both class I and class II [7,8]. It has also been shown that the serotonin metabolite 5-HTOL is an ADH substrate [9].In addition to reversible alcohol oxidation/aldehyde reduction, horse class-I and human class-I and class-II ADH catalyse aldehyde oxidation and thereby mimic the activity of aldehyde dehydrogenase (ALDH) [10±12]. Incubation of ADH with aldehyde and N...

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Section: Steady-state Kineticssupporting

confidence: 73%

Section: Chemicals and Enzyme Preparationsmentioning

confidence: 99%

Section: Kinetic Analysismentioning

confidence: 99%

Section: Kinetic Analysismentioning

confidence: 99%

See 2 more Smart Citations

Activities of human alcohol dehydrogenases in the metabolic pathways of ethanol and serotonin

Svensson¹,

Some²,

Lundsjö³

et al. 1999

European Journal of Biochemistry

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“…Firstly, most aldehydes are reactive species that readily react with, and modify, cellular proteins. Most cells contain efficient enzyme systems to remove aldehydes in order to avoid such harmful reactions (Kawamura et al, 2002;Svensson et al, 1996). Thus, it would be necessary for such aldehydes to be generated in close proximity to Kvβ2 in order to be effective.…”

Section: Discussionmentioning

confidence: 99%

Substrate profiling and aldehyde dismutase activity of the Kvβ2 subunit of the mammalian Kv1 potassium channel

Kumari¹,

Ryan²,

Dolly³

et al. 2010

The International Journal of Biochemistry & Cell Biology

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Zn‐Dependent Medium‐Chain Dehydrogenases/Reductases

Meijers

Cedergren-Zeppezauer

2004

Handbook of Metalloproteins

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Medium‐chain dehydrogenases/reductases (MDRs) have about 350 to 390 amino acids and belong to a large superfamily of proteins including the Zn‐dependent alcohol dehydrogenases (ADHs) and several other Zn‐dependent activities. They are dimers or tetramers and utilize either NADH or NADPH as electron carriers referred to as cofactors. The oxidation/reduction process is a 2‐electron/1‐proton (hydride ion, H − ) transfer between substrate and cofactor. The compounds oxidized or reduced by these enzymes vary considerably in structure ranging from small primary and secondary alcohols/aldehydes/ketones (aliphatic and aromatic), sugars (sorbitol/fructose, glucose) to large substances like fatty acids, steroids, and retinoids. ADHs are widely distributed in nature and some MDR members are found in all species. An essential Zn ion at the active site is common to all MDRs but some members have two zinc ions per subunit. The 3D Structures of Zn‐dependent MDRs are highly similar and they essentially have the same polypeptide fold. The low sequence identity found sometimes could not immediately reveal this close structural relationship. Now numerous structures have been examined indicating that the evolutionary strategy for MDRs has been to vary amino acids preferably at the reaction center and further away in the substrate binding area. Thus, the size and shape of the substrate channel is adapted for small or large substrates, hydrophilic or hydrophobic substances, modifications attained via few mutations, or significant deletions/additions of amino acids. A further opportunity to create multiplicity has been to vary the control mechanism for the protein conformation change and the oligomeric state. Some enzymes undergo significant structural changes upon cofactor binding, while others do not. A transition state intermediate of NADH in complex with horse liver alcohol dehydrogenase, resolved to atomic resolution (≈1 Å), has revealed structural modifications of the cofactor associated with the activation process for hydride transfer. This involves an OH − pyridine ring adduct, connected to the zinc center. The distortion of the nicotinamide at the enzyme site, affected by the presence of the negatively charged oxygen ligand, influences three parameters: (i) the puckering of the ring, (ii) a deviation from a standard carbon–carbon double bond length, and (iii) accumulation of negative charge at the C4 atom promoting hydride transfer. The active site Zn‐center of this large enzyme group shows great flexibility with respect to metal–ligand geometry and dynamics. Structural and spectroscopic evidence suggest that Zn‐dependent MDRs should be included in the general family of Zn‐enzymes, which require water and hydroxide ion as reactants during catalysis.

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Aldehyde dismutase activity of human liver alcohol dehydrogenase

Cited by 40 publications

References 25 publications

Activities of human alcohol dehydrogenases in the metabolic pathways of ethanol and serotonin

Activities of human alcohol dehydrogenases in the metabolic pathways of ethanol and serotonin

Substrate profiling and aldehyde dismutase activity of the Kvβ2 subunit of the mammalian Kv1 potassium channel

Zn‐Dependent Medium‐Chain Dehydrogenases/Reductases

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