title: Molecular and catalytic properties of 2,4'-dihydroxyacetophenone dioxygenase from Burkholderia sp. AZ11.
abstract: The gene dad encoding 2,4'-dihydroxyacetophenone (DHAP) dioxygenase was cloned from Burkholderia sp. AZ11. The initiation codon GTG was converted to ATG for high-level expression of the enzyme in Escherichia coli. The enzyme was moderately thermostable, and the recombinant enzyme was briefly purified. The enzyme (M(r)=90 kDa) was a homotetramer with a subunit M(r) of 23 kDa. It contained 1.69 mol of non-heme iron, and had a dark gray color. On anaerobic incubation of it with DHAP, the absorption at around 400 nm increased due to the formation of an enzyme-DHAP complex. Multiple sequence alignment suggested that His77, His79, His115, and Glu96 in the cupin fold were possible metal ligands. The apparent K(m) for DHAP and the apparent V(max) were estimated to be 1.60 µM and 6.28 µmol/min/mg respectively. 2-Hydroxyacetophenone was a poor substrate. CuCl(2) and HgCl(2) strongly inhibited the enzyme, while FeSO(4) weakly activated it.
authors.affiliation: Department of Biomolecular Science, Faculty of Engineering, Gifu University, Gifu, Japan.
authors.forename: Mayu
authors.initials: M
authors.lastname: Enya
authors.forename: Keiko
authors.initials: K
authors.lastname: Aoyagi
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Hishikawa
authors.forename: Azusa
authors.initials: A
authors.lastname: Yoshimura
authors.forename: Koichi
authors.initials: K
authors.lastname: Mitsukura
authors.forename: Kiyofumi
authors.initials: K
authors.lastname: Maruyama
mesh_terms: D000042:Absorption; D000595:Amino Acid Sequence; D001559:Benzhydryl Compounds; D055162:Biocatalysis; D019117:Burkholderia / Q000201:enzymology* / Q000235:genetics / Q000302:isolation & purification; D003001:Cloning, Molecular; D049308:Dioxygenases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004926:Escherichia coli / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D010636:Phenols / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature; D000098826:Bisphenol A Compounds
chemical_list: D001559:Benzhydryl Compounds; D049308:Dioxygenases; D004791:Enzyme Inhibitors; D010636:Phenols; C403717:2,4'-dihydroxyacetophenone dioxygenase; C006780:bisphenol A; D000098826:Bisphenol A Compounds