title: Gentisate 1,2-dioxygenase from Xanthobacter polyaromaticivorans 127W.
abstract: A putative gentisate 1,2-dioxygenase was encoded in the dibenzothiophene degradation gene cluster (dbd) from Xanthobacter polyaromaticivorans 127W. The deduced amino acid sequence showed high sequence similarity with gentisate dioxygenases from Pseudomonas alcaligenes (AAD49427, 65% identical), Bradyrhizobium japonicum (NP_766750, 64%), and P. aeruginosa (ZP_00135722, 54%), and moderate similarity with 1-hydroxy-2-naphthoate dioxygenase from Nocardioides sp. KP7 (BAA31235, 33%) and salicylate dioxygenase from Pseudaminobacter salicylatoxidans (AAQ91293, 33%). The enzyme, GDOxp, was heterologously produced in Escherichia coli and purified to homogeneity. GDOxp formed a tetramer and exhibited high dioxygenase activity against 1,4-dihydroxy 2-naphthoate as well as gentisate, suggesting unusually broad substrate specificity. GDOxp easily released ferrous ion under unfavorable temperature and pH conditions to become an inactive monomer protein. An inactive monomer protein can reconstitute a tetramer structure and restore enzyme activity in a cooperative manner upon the addition of ferrous ion. Chymotryptic digestion and protein truncation experiments suggested that the N-terminal region is important for the tetramerization of GDOxp.
authors.affiliation: Department of Material and Life Science, Graduate School of Engineering, Osaka University, Japan.
authors.forename: Shin-ichi
authors.initials: S
authors.lastname: Hirano
authors.forename: Masaaki
authors.initials: M
authors.lastname: Morikawa
authors.forename: Kazufumi
authors.initials: K
authors.lastname: Takano
authors.forename: Tadayuki
authors.initials: T
authors.lastname: Imanaka
authors.forename: Shigenori
authors.initials: S
authors.lastname: Kanaya
mesh_terms: D000595:Amino Acid Sequence; D049308:Dioxygenases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D011489:Protein Denaturation; D016415:Sequence Alignment; D013379:Substrate Specificity; D013696:Temperature; D020584:Xanthobacter / Q000201:enzymology* / Q000235:genetics
chemical_list: D049308:Dioxygenases; C026247:gentisate 1,2-dioxygenase