title: Purification and characterization of NADPH-dependent Cr(VI) reductase from Escherichia coli ATCC 33456.
abstract: A soluble Cr(VI) reductase was purified from the cytoplasm of Escherichia coli ATCC 33456. The molecular mass was estimated to be 84 and 42 kDa by gel filtration and SDS-polyacrylamide gel electrophoresis, respectively, indicating a dimeric structure. The pI was 4.66, and optimal enzyme activity was obtained at pH 6.5 and 37 degrees C. The most stable condition existed at pH 7.0. The purified enzyme used both NADPH and NADH as electron donors for Cr(VI) reduction, while NADPH was the better, conferring 61%; higher activity than NADH. The Km values for NADPH and NADH were determined to be 47.5 and 17.2 micromol, and the Vmax values 322.2 and 130.7 micromol Cr(VI) min(-1)mg(-1) protein, respectively. The activity was strongly inhibited by N-ethylmalemide, Ag2+, Cd2+, Hg2+, and Zn2+. The antibody against the enzyme showed no immunological cross reaction with those of other Cr(VI) reducing strains.
authors.affiliation: Division of Bioscience and Bioinformatics, Myongji University, Yongin, 449-728, Republic of Korea.
authors.forename: Woo-Chul
authors.initials: WC
authors.lastname: Bae
authors.forename: Han-Ki
authors.initials: HK
authors.lastname: Lee
authors.forename: Young-Chool
authors.initials: YC
authors.lastname: Choe
authors.forename: Deok-Jin
authors.initials: DJ
authors.lastname: Jahng
authors.forename: Sang-Hee
authors.initials: SH
authors.lastname: Lee
authors.forename: Sang-Jin
authors.initials: SJ
authors.lastname: Kim
authors.forename: Jung-Hyun
authors.initials: JH
authors.lastname: Lee
authors.forename: Byeong-Chul
authors.initials: BC
authors.lastname: Jeong
mesh_terms: D000595:Amino Acid Sequence; D019281:Dimerization; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004926:Escherichia coli / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D007700:Kinetics; D008670:Metals / Q000494:pharmacology; D008970:Molecular Weight; D009249:NADP / Q000378:metabolism; D010088:Oxidoreductases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D020836:Protein Structure, Quaternary; D012995:Solubility; D013696:Temperature
chemical_list: D004791:Enzyme Inhibitors; D008670:Metals; D009249:NADP; D010088:Oxidoreductases; C075965:chromium(VI) reductase