TitleTwo zebrafish alcohol dehydrogenases share common ancestry with mammalian class I, II, IV, and V alcohol dehydrogenase genes but have distinct functional characteristics.
Publication TypeJournal Article
Year of Publication2004
AuthorsReimers, MJ, Hahn, ME, Tanguay, RL
JournalJ Biol Chem
Volume279
Issue37
Pagination38303-12
Date Published2004 Sep 10
ISSN0021-9258
KeywordsAlcohol Dehydrogenase, Amino Acid Sequence, Animals, Binding, Competitive, Cell Line, Chromosome Mapping, Conserved Sequence, DNA, Complementary, Ethanol, Glutathione, Humans, Insecta, Kinetics, Mice, Molecular Sequence Data, Open Reading Frames, Phylogeny, Rats, Recombinant Proteins, RNA, Messenger, Sequence Homology, Amino Acid, Species Specificity, Time Factors, Tissue Distribution, Zebrafish
Abstract

Ethanol is teratogenic to many vertebrates. We are utilizing zebrafish as a model system to determine whether there is an association between ethanol metabolism and ethanol-mediated developmental toxicity. Here we report the isolation and characterization of two cDNAs encoding zebrafish alcohol dehydrogenases (ADHs). Phylogenetic analysis of these zebrafish ADHs indicates that they share a common ancestor with mammalian class I, II, IV, and V ADHs. The genes encoding these zebrafish ADHs have been named Adh8a and Adh8b by the nomenclature committee. Both genes were genetically mapped to chromosome 13. The 1450-bp Adh8a is 82, 73, 72, and 72% similar at the amino acid level to the Baltic cod ADH8 (previously named ADH1), the human ADH1B2, the mouse ADH1, and the rat ADH1, respectively. Also, the 1484-bp Adh8b is 77, 68, 67, and 66% similar at the amino acid level to the Baltic cod ADH8, the human ADH1B2, the mouse ADH1, and the rat ADH1, respectively. ADH8A and ADH8B share 86% amino acid similarity. To characterize the functional properties of ADH8A and ADH8B, recombinant proteins were purified from SF-9 insect cells. Kinetic studies demonstrate that ADH8A metabolizes ethanol, with a V(max) of 13.4 nmol/min/mg protein, whereas ADH8B does not metabolize ethanol. The ADH8A K(m) for ethanol as a substrate is 0.7 mm. 4-Methyl pyrazole, a classical competitive inhibitor of class I ADH, failed to inhibit ADH8A. ADH8B has the capacity to efficiently biotransform longer chain primary alcohols (>/=5 carbons) and S-hydroxymethlyglutathione, whereas ADH8A does not efficiently metabolize these substrates. Finally, mRNA expression studies indicate that both ADH8A and ADH8B mRNA are expressed during early development and in the adult brain, fin, gill, heart, kidney, muscle, and liver. Together these results indicate that class I-like ADH is conserved in zebrafish, albeit with mixed functional properties.

DOI10.1074/jbc.M401165200
Alternate JournalJ. Biol. Chem.
PubMed ID15231826
PubMed Central IDPMC3261772
Grant ListR01 ES006272 / ES / NIEHS NIH HHS / United States
R21 AA012783-04 / AA / NIAAA NIH HHS / United States
ES00210 / ES / NIEHS NIH HHS / United States
P30 ES003850 / ES / NIEHS NIH HHS / United States
R21 AA012783 / AA / NIAAA NIH HHS / United States
ES03850 / ES / NIEHS NIH HHS / United States
R01ES06272 / ES / NIEHS NIH HHS / United States
P30 ES000210-38 / ES / NIEHS NIH HHS / United States
AA12783 / AA / NIAAA NIH HHS / United States
R01 ES006272-13 / ES / NIEHS NIH HHS / United States
P30 ES000210 / ES / NIEHS NIH HHS / United States