Purification and characterization of chicken liver alcohol dehydrogenase

Bahr‐Lindström, Hedvig von; Andersson, Lars I.; Mosbach, Klaus; Jörnvall, Hans

doi:10.1016/0014-5793(78)80239-7

Cited by 14 publications

(9 citation statements)

References 17 publications

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“…The mutant YADH represents the S-AA-5 form (11). LADH represents all characterized parent forms of liver alcohol dehydrogenase-i.e., the E-and S-isozyme chains of the horse liver enzyme, the 13 and y isozyme chains of the human liver enzyme, and the subunits of the rat and chicken liver enzymes (1, 17,38). A direct correlation between the histidine/arginine exchange and the binding of NAD' is obvious from kinetic properties of the mutant forms and the parent LADH and YADH forms, respectively (cf.…”

Section: Resultsmentioning

confidence: 99%

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Human liver alcohol dehydrogenase: amino acid substitution in the beta 2 beta 2 Oriental isozyme explains functional properties, establishes an active site structure, and parallels mutational exchanges in the yeast enzyme.

Jörnvall¹,

Hempel²,

Vallée³

et al. 1984

Proc. Natl. Acad. Sci. U.S.A.

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The homodimeric Oriental 132132 isozyme of human liver alcohol dehydrogenase, corresponding to an allelic variant at the ADH2 gene locus, was studied in order to define the amino acid exchange in relation to the .813f3 isozyme, the predominant allelic form among Caucasians. Sequence analysis reveals that the amino acid substitution occurs at position 7 of the largest CNBr fragment, corresponding to position 47 of the whole protein chain. Here, the P2 form has a histidine residue, while, in common with other characterized mammalian liver alcohol dehydrogenases, the 81, form has an arginine residue. This exchange does not affect the adjacent cysteine-46 residue, which is a protein ligand to the active-site zinc atom, thus clarifying previously inconsistent results. The histidine/arginine-47 mutational replacement corresponds to a position that binds the pyrophosphate group of the coenzyme NAD(H); this explains the functional differences between the P13f1 and 82182 isozymes, including both a lower pH optimum and higher turnover number of 382132, which is likely to be the mutant form. The exchange demonstrates the existence of parallel but separate mutations in the evolution of alcohol dehydrogenases because these mammalian enzymes differ at exactly the same position by the same type of substitution as is found between a mutant and the wild-type constitutive forms of the corresponding yeast enzyme.

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Section: Resultsmentioning

confidence: 99%

“…LA; the composition, in Table 1; and the amino acid sequence data, in Table 2. The fragment corresponds to positions (horse, rat, and chicken) that have been investigated (1,38). This exchange is adjacent to the zinc-liganding cysteine-46 (39), which is unaltered.…”

Section: Resultsmentioning

confidence: 99%

Human liver alcohol dehydrogenase: amino acid substitution in the beta 2 beta 2 Oriental isozyme explains functional properties, establishes an active site structure, and parallels mutational exchanges in the yeast enzyme.

Jörnvall¹,

Hempel²,

Vallée³

et al. 1984

Proc. Natl. Acad. Sci. U.S.A.

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“…This is further strengthened when the structure around the reactive cysteine residue in sorbitol dehydrogenase is compared to the structure around the active-site cysteine residue (Cys-46) in horse liver alcohol dehydrogenase and its counterpart in various other alcohol dehydrogenases [14,28,42]. These regions are extensively homologous as shown in Fig.…”

Section: Resemblance To Alcohol Dehydrogenasesmentioning

confidence: 95%

Properties of Sorbitol Dehydrogenase and Characterization of a Reactive Cysteine Residue Reveal Unexpected Similarities to Alcohol Dehydrogenases

et al. 1981

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Sorbitol dehydrogenase was characterized as a homogeneous protein on affinity chromatography and ion‐exchange chromatography. Tests of stability, sensitivity to inhibitors, and protection by coenzyme suggest that the enzyme has essential cysteine, metal, and probably histidine. The native enzyme has a molecular weight around 140000 and a subunit around 35000–40000, suggesting a tetrameric quaternary structure. Subunits are highly similar if not identical as judged by characterization of one unique 45‐residue sequence containing a single reactive cysteine residue. Properties resemble those of mammalian and yeast alcohol dehydrogenases, and the sequence determined for the region around the reactive cysteine residue is homologous to that around one of the zinc‐liganding cysteine residues at the active site of horse liver alcohol dehydrogenase. Sorbitol dehydrogenase thus reveals an unexpected relationship to alcohol dehydrogenases, from which ancestral connections and functional mechanisms in this group of enzymes may be further elucidated.

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“…Apart from the structures included, extensive data exist for alcohol dehydrogenase from chicken liver [30], and Bacillus stearothermophilus [31], as well as from the S-type of protein chain of horse liver alcohol dehydrogenase, class I [18]. These structures are excluded from the alignments given because none covers all positions in complete detail.…”

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confidence: 99%

Characteristics of alcohol/polyol dehydrogenases

Jörnvall

Persson

Jeffery

1987

European Journal of Biochemistry

282

173

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Sixteen characterized alcohol dehydrogenases and one sorbitol dehydrogenase have been aligned. The proteins represent two formally different enzyme activities (EC 1.1.1.1 and EC 1.1.1.14), three different types of molecule (dimeric alcohol dehydrogenase, tetrameric alcohol dehydrogenase, tetrameric sorbitol dehydrogenase), metalloproteins with different zinc contents (1 or 2 atoms per subunit), and polypeptide chains from different kingdoms and orders (mammals, higher plants, fungus, yeasts). Present comparisons utilizing all 17 forms reveal extensive variations in alcohol dehydrogenase, but with evolutionary changes that are of the same order in different branches and at different times. They emphasize the general importance of particular residues, suggesting related overall functional constraints in the molecules. The comparisons also define a few coincidences between intron positions in the genes and gap positions in the gene products. Only 22 residues are strictly conserved; half of these are Gly, and most of the remaining ones are Pro or acidic residues. No basic residues, no straight‐chain hydrophobic residues, no aromatic residues, and essentially no branched‐chain or polar neutral residues are invariable. Tentative consensus sequences were calculated, defining 13 additional residues likely to be typical of but not invariant among the alcohol dehydrogenases. These show a predominance of Val, charged residues, and Gly. Combined, the comparisons, which are particularly relevant to the data base for protein engineering, illustrate the requirements for functionally important binding interactions, and the extent of space restrictions in proteins with related overall conformations and functions.

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Purification and characterization of chicken liver alcohol dehydrogenase

Cited by 14 publications

References 17 publications

Human liver alcohol dehydrogenase: amino acid substitution in the beta 2 beta 2 Oriental isozyme explains functional properties, establishes an active site structure, and parallels mutational exchanges in the yeast enzyme.

Human liver alcohol dehydrogenase: amino acid substitution in the beta 2 beta 2 Oriental isozyme explains functional properties, establishes an active site structure, and parallels mutational exchanges in the yeast enzyme.

Properties of Sorbitol Dehydrogenase and Characterization of a Reactive Cysteine Residue Reveal Unexpected Similarities to Alcohol Dehydrogenases

Characteristics of alcohol/polyol dehydrogenases

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