caa a22 4500 397557965 CHVBK 20180308164808.0 cr unu---uuuuu 161202e199610 xx s 000 0 eng 10.1093/oxfordjournals.jbchem.a021473 doi (NATIONALLICENCE)oxford-10.1093/oxfordjournals.jbchem.a021473 Gene Cloning, Purification, and Characterization of NfsB, a Minor Oxygen-Insensitive Nitroreductase from Escherichia coli, Similar in Biochemical Properties to FRase I, the Major Flavin Reductase in Vibrio fischeri [Elektronische Daten] [Shuhei Zenno, Hideaki Koike, Masaru Tanokura, Kaoru Saigo] nfsB, encoding a minor oxygen-insensitive nitroreductase, was isolated by PCR using primers corresponding to two amino acid sequences conserved among the major flavin reductase from Vibrio fischeri and classical nitroreductases from Salmonella typhimurium and Enterobacter cloacae. The gene product, NfsB, was purified to homogeneity from extracts of Escherichia coli cells overexpressing it. nfsB was found to be situated at 13 min on the E. coli map. Biochemical analysis Indicated NfsB to be a polypeptide having a calculated molecular weight of 23,904, capable of forming a homodimer and associated tightly with FMN as a prosthetic group. Although it exhibited a lower affinity to the NfsB apoenzyme than FMN, FAD could serve as an effective substitute for FMN. It was also shown that NfsB has a broad electron acceptor specificity and is associated with a low level of the NAD(P)H-flavin oxidoreductase. The NfsB catalysis obeys the ping pong Bi-Bimechanism. The Km value for NADH varied depending on the second substrate used. © 1996 BY THE JOURNAL OF BIOCHEMISTRY Regular Papers nationallicence Escherichia coli nationallicence flavin reductase nationallicence flavoenzyme nationallicence nfsB nationallicence nitroreductase nationallicence Zenno Shuhei Department of Biophysics and Biochemistry, Graduate School of Science 7-3-1 Hongo, Bunkyo-ku, Tokyo 113 aut Koike Hideaki Biotechnology Research Center, The University of Tokyo 7-3-1 Hongo, Bunkyo-ku, Tokyo 113 aut Tanokura Masaru Biotechnology Research Center, The University of Tokyo 7-3-1 Hongo, Bunkyo-ku, Tokyo 113 aut Saigo Kaoru Department of Biophysics and Biochemistry, Graduate School of Science 7-3-1 Hongo, Bunkyo-ku, Tokyo 113 aut The Journal of Biochemistry Oxford University Press 120/4(1996-10), 736-744 0021-924X 120:4<736 1996 120 jbchem https://doi.org/10.1093/oxfordjournals.jbchem.a021473 text/html Onlinezugriff via DOI 1 other jats NATIONALLICENCE 856 40 https://doi.org/10.1093/oxfordjournals.jbchem.a021473 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Zenno Shuhei Department of Biophysics and Biochemistry, Graduate School of Science 7-3-1 Hongo, Bunkyo-ku, Tokyo 113 aut NATIONALLICENCE 700 1- Koike Hideaki Biotechnology Research Center, The University of Tokyo 7-3-1 Hongo, Bunkyo-ku, Tokyo 113 aut NATIONALLICENCE 700 1- Tanokura Masaru Biotechnology Research Center, The University of Tokyo 7-3-1 Hongo, Bunkyo-ku, Tokyo 113 aut NATIONALLICENCE 700 1- Saigo Kaoru Department of Biophysics and Biochemistry, Graduate School of Science 7-3-1 Hongo, Bunkyo-ku, Tokyo 113 aut NATIONALLICENCE 773 0- The Journal of Biochemistry Oxford University Press 120/4(1996-10), 736-744 0021-924X 120:4<736 1996 120 jbchem Metadata rights reserved CC BY-NC-4.0 http://creativecommons.org/licenses/by-nc/4.0 nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-oxford