title: The lipoxygenase pathway in zebrafish. Expression and characterization of zebrafish ALOX5 and comparison with its human ortholog.
abstract: The zebrafish (Danio rerio) is frequently employed as model organism to explore vertebrate embryogenesis but little is known about the lipoxygenase pathway in this lower vertebrate. When we searched the zebrafish genome for lipoxygenase genes we detected seven different genes localized on four different chromosomes. Four of these genes (ALOX2, ALOX3a, ALOX3b, ALOX3c) share a high degree of sequence conservation with the human ALOX5 gene, which encodes for the key enzyme of mammalian leukotriene biosynthesis. Expression profiles of the corresponding transcripts indicated that the ALOX2 gene is high level expressed during zebrafish embryogenesis whereas transcripts originating from the other three paralog genes could not be detected. To functionally compare human ALOX5 with the putative zebrafish ortholog (zebrafish ALOX2) we cloned and expressed the zebrafish enzyme in pro- and eukaryotic expression systems and characterized it as arachidonic acid 5S-lipoxygenating enzyme, which also exhibits leukotriene synthase activity. Mutagenesis studies of the triad determinants (Phe359Trp, Ala424Ile, Asn425Met) altered the reaction specificity from 5S- to dominant 15S-lipoxygenation. The finding that zebrafish expresses a functional ALOX5 together with the observation that most other human leukotriene relevant genes have an ortholog in the zebrafish genome suggests the biological relevance of leukotriene signaling in lower vertebrates.
authors.affiliation: Institute of Biochemistry, University Medicine Berlin, Charité, Chariteplatz 1, D-10117 Berlin, Germany.
authors.forename: Susan
authors.initials: S
authors.lastname: Adel
authors.affiliation: Institute of Biochemistry, University Medicine Berlin, Charité, Chariteplatz 1, D-10117 Berlin, Germany.
authors.forename: Dagmar
authors.initials: D
authors.lastname: Heydeck
authors.affiliation: Institute of Biochemistry, University Medicine Berlin, Charité, Chariteplatz 1, D-10117 Berlin, Germany. Electronic address: hartmut.kuehn@charite.de.
authors.forename: Hartmut
authors.initials: H
authors.lastname: Kuhn
authors.affiliation: Institute of Biochemistry, University Medicine Berlin, Charité, Chariteplatz 1, D-10117 Berlin, Germany.
authors.forename: Christoph
authors.initials: C
authors.lastname: Ufer
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001094:Arachidonate 5-Lipoxygenase / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010802:Phylogeny; D016415:Sequence Alignment; D015027:Zebrafish / Q000196:embryology / Q000378:metabolism*; D029961:Zebrafish Proteins / Q000737:chemistry / Q000235:genetics*
chemical_list: D007527:Isoenzymes; D029961:Zebrafish Proteins; D001094:Arachidonate 5-Lipoxygenase; C583212:ALOX5 protein, human; C000608463:alox5a protein, zebrafish
keywords: Eicosanoids; Evolution; Leukotrienes; Oxidative stress; Zebrafish