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Predicate | Object |
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rdf:type | |
lifeskim:mentions | |
pubmed:issue |
8
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pubmed:dateCreated |
1995-3-31
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pubmed:databankReference | |
pubmed:abstractText |
A full-length 1966-base pair clone of the human class IV alcohol dehydrogenase (sigma-ADH) was isolated from a human stomach cDNA library. The 373-amino acid sigma-ADH encoded by this cDNA was expressed in Escherichia coli. The specific activity of the recombinant enzyme for ethanol oxidation at pH 7.5 and 25 degrees C, calculated from active-site titration of NADH binding, was 92 +/- 9 units/mg. Kinetic analysis of the catalytic efficiency (kcat/KM) of recombinant sigma-ADH for oxidation of primary alcohols indicated broad substrate specificity. Recombinant human sigma-ADH exhibited high catalytic efficiency for oxidation of all-trans-retinol to all-trans-retinal. This pathway is important in the synthesis of the transcriptional regulator all-trans-retinoic acid. Secondary alcohols and 3 beta-hydroxysteroids were inactive with sigma-ADH or were oxidized with very low efficiency. The KM of sigma-ADH for ethanol was 25 mM, and the KM for primary straight chain alcohols decreased substantially as chain length increased. There are important amino acid differences in the alcohol-binding site between the human class IV (sigma) and human class I (beta) alcohol dehydrogenases that appear to explain the high catalytic efficiency for all-trans-retinol, the high kcat for ethanol, and the low catalytic efficiency for secondary alcohols of sigma-ADH relative to beta 1-ADH. For example, modeling the binding of all-trans-retinol in the human beta 1-ADH structure suggested that coordination of retinol to the active-site zinc is hindered by a loop from residues 114 to 120 that is at the entrance to the alcohol-binding site. The deletion of Gly-117 in human sigma-ADH and a substitution of Leu for the bulky Tyr-110 appear to facilitate retinol access to the active-site zinc.
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pubmed:grant | |
pubmed:language |
eng
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pubmed:journal | |
pubmed:citationSubset |
IM
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pubmed:chemical | |
pubmed:status |
MEDLINE
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pubmed:month |
Feb
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pubmed:issn |
0021-9258
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pubmed:author | |
pubmed:issnType |
Print
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pubmed:day |
24
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pubmed:volume |
270
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pubmed:owner |
NLM
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pubmed:authorsComplete |
Y
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pubmed:pagination |
3625-30
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pubmed:dateRevised |
2007-11-14
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pubmed:meshHeading |
pubmed-meshheading:7876099-Alcohol Dehydrogenase,
pubmed-meshheading:7876099-Animals,
pubmed-meshheading:7876099-Base Sequence,
pubmed-meshheading:7876099-DNA, Complementary,
pubmed-meshheading:7876099-Humans,
pubmed-meshheading:7876099-Isoenzymes,
pubmed-meshheading:7876099-Kinetics,
pubmed-meshheading:7876099-Liver,
pubmed-meshheading:7876099-Molecular Sequence Data,
pubmed-meshheading:7876099-Recombinant Proteins,
pubmed-meshheading:7876099-Sequence Homology, Amino Acid,
pubmed-meshheading:7876099-Stomach,
pubmed-meshheading:7876099-Substrate Specificity
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pubmed:year |
1995
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pubmed:articleTitle |
Expression and kinetic characterization of recombinant human stomach alcohol dehydrogenase. Active-site amino acid sequence explains substrate specificity compared with liver isozymes.
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pubmed:affiliation |
Dept. of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis 46202-5122.
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pubmed:publicationType |
Journal Article,
Research Support, U.S. Gov't, P.H.S.
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