title: Acetylacetone-cleaving enzyme Dke1: a novel C-C-bond-cleaving enzyme from Acinetobacter johnsonii.
abstract: The toxicity of acetylacetone has been demonstrated in various studies. Little is known, however, about metabolic pathways for its detoxification or mineralization. Data presented here describe for the first time the microbial degradation of acetylacetone and the characterization of a novel enzyme that initiates the metabolic pathway. From an Acinetobacter johnsonii strain that grew with acetylacetone as the sole carbon source, an inducible acetylacetone-cleaving enzyme was purified to homogeneity. The corresponding gene, coding for a 153 amino acid sequence that does not show any significant relationship to other known protein sequences, was cloned and overexpressed in Escherichia coli and gave high yields of active enzyme. The enzyme cleaves acetylacetone to equimolar amounts of methylglyoxal and acetate, consuming one equivalent of molecular oxygen. No exogenous cofactor is required, but Fe(2+) is bound to the active protein and essential for its catalytic activity. The enzyme has a high affinity for acetylacetone with a K (m) of 9.1 microM and a k(cat) of 8.5 s(-1). A metabolic pathway for acetylacetone degradation and the putative relationship of this novel enzyme to previously described dioxygenases are discussed.
authors.affiliation: Institute of Biotechnology, Graz University of Technology, Austria. grit.straganz@TUGraz.at
authors.forename: Grit D
authors.initials: GD
authors.lastname: Straganz
authors.forename: Anton
authors.initials: A
authors.lastname: Glieder
authors.forename: Lothar
authors.initials: L
authors.lastname: Brecker
authors.forename: Douglas W
authors.initials: DW
authors.lastname: Ribbons
authors.forename: Walter
authors.initials: W
authors.lastname: Steiner
mesh_terms: D000085:Acetates / Q000378:metabolism; D000150:Acinetobacter / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D001483:Base Sequence; D003001:Cloning, Molecular; D007501:Iron / Q000378:metabolism; D008969:Molecular Sequence Data; D010100:Oxygen / Q000378:metabolism; D010105:Oxygenases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D010422:Pentanones / Q000378:metabolism; D011765:Pyruvaldehyde / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D000085:Acetates; D001426:Bacterial Proteins; D010422:Pentanones; C008790:acetylacetone; D011765:Pyruvaldehyde; D007501:Iron; D010105:Oxygenases; D010100:Oxygen