Dibenzothiophene desulfurizing enzymes from moderately thermophilic bacterium Bacillus subtilis WU-S2B: purification, characterization and overexpression.
The moderately thermophilic bacterium Bacillus subtilis WU-S2B desulfurized dibenzothiophene (DBT) at 50 degrees C through the selective cleavage of carbon-sulfur bonds. In this study, three enzymes involved in the microbial DBT desulfurization were purified and characterized. The first two enzymes, DBT monooxygenase (BdsC) and DBT sulfone monooxygenase (BdsA), were purified from the wild-type strain, and the last one, 2'-hydroxybiphenyl 2-sulfinic acid desulfinase (BdsB), was purified from the recombinant Escherichia coli overexpressing the gene, bdsB, with chaperonin genes, groEL/ES. The genes of BdsC and BdsA were also overexpressed. The molecular weights of BdsC and BdsA were determined to be 200 and 174 kDa, respectively, by gel filtration chromatography, suggesting that both enzymes had four identical subunits. BdsB had a monomeric structure of 40 kDa. The three enzymes were characterized and compared with the corresponding enzymes (DszC, DszA, and DszB) of mesophilic desulfurization bacteria. The specific activities of BdsC, BdsA, and BdsB were 84.2, 855, and 280 units/mg, respectively, and the latter two activities were higher than those of DszA and DszB. The heat stability and optimum temperature of BdsC, BdsA, and BdsB were higher than those of DszC, DszA, and DszB. Other enzymatic properties were investigated in detail.
Ohshiro
Takashi
T
Department of Biotechnology, Tottori University, 4-101 Koyamacho-minami, Tottori 680-8552, Japan.
Ishii
Yoshitaka
Y
Matsubara
Toshiyuki
T
Ueda
Koichi
K
Izumi
Yoshikazu
Y
Kino
Kuniki
K
Kirimura
Kohtaro
K
D001412:Bacillus subtilis / Q000201:enzymology*; D001709:Biotechnology / Q000379:methods*; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D010088:Oxidoreductases / Q000096:biosynthesis* / Q000737:chemistry* / Q000302:isolation & purification; D010105:Oxygenases / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D013379:Substrate Specificity; D013696:Temperature; D013876:Thiophenes / Q000378:metabolism*; D015533:Transcriptional Activation
D013876:Thiophenes; D010088:Oxidoreductases; D010105:Oxygenases; C421210:dibenzothiophene sulfone monooxygenase; C090100:dibenzothiophene monooxygenase; C016366:dibenzothiophene