title: Purification, characterization, and gene cloning of thermophilic cytochrome cd1 nitrite reductase from Hydrogenobacter thermophilus TK-6.
abstract: A thermophilic, chemolithoautotrophic hydrogen-oxidizing bacterium, Hydrogenobacter thermophilus TK-6 can grow autotrophically under anaerobic conditions by denitrification. One of the denitrification enzymes, cytochrome cd(1) nitrite reductase, was isolated and its gene was cloned from strain TK-6. The subunit molecular mass of the purified enzyme was 61.5 kDa and the isoelectric point was determined to be 9.3. The optimum temperature and pH for the enzymatic reaction were 70-75 degrees C and 6.5-7.0, respectively. The structural gene for the enzyme, nirS, is probably transcribed as a hexacistronic operon with the following genes encoding a putative diheme cytochrome c and the proteins required for biosynthesis of heme d(1). The NirS sequence was phylogenetically distinct from those of proteobacteria. The consensus -35 and -10 sequences were found in the putative nirS promoter region, but the consensus sequences for the DNR/NnrR-type or the NorR/FhpR-type nitric oxide sensing regulators were not found in this region.
authors.affiliation: Department of Biotechnology, The University of Tokyo, Japan.
authors.forename: Miho
authors.initials: M
authors.lastname: Suzuki
authors.forename: Tadao
authors.initials: T
authors.lastname: Hirai
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Arai
authors.forename: Masaharu
authors.initials: M
authors.lastname: Ishii
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Igarashi
mesh_terms: D000595:Amino Acid Sequence; D001419:Bacteria / Q000201:enzymology*; D003001:Cloning, Molecular; D003580:Cytochromes / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D004789:Enzyme Activation; D004795:Enzyme Stability; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009572:Nitrite Reductases / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D015202:Protein Engineering / Q000379:methods*; D017386:Sequence Homology, Amino Acid; D013696:Temperature
chemical_list: D003580:Cytochromes; D009572:Nitrite Reductases; C028123:nitrite reductase (NO-forming)