title: Infrared and EPR studies on cyanide binding to the heme-copper binuclear center of cytochrome bo-type ubiquinol oxidase from Escherichia coli. Release of a CuB-cyano complex in the partially reduced state.
abstract: Cyanide-binding to the heme-copper binuclear center of bo-type ubiquinol oxidase from Escherichia coli was investigated with Fourier transform-infrared and EPR spectroscopies. Upon treatment of the air-oxidized CN-inhibited enzyme with excess sodium dithionite, a 12C-14N stretching vibration at 2146 cm-1 characteristic of the FeO3+ C=N CuB2+ bridging structure was quickly replaced with another stretching mode at 2034.5 cm-1 derived from the FeO2+ C=N moiety. The presence of ubiquinone-8 or ubiquinone-1 caused a gradual autoreduction of the metal center(s) of the air-oxidized CN-inhibited enzyme and a concomitant appearance of a strong cyanide stretching band at 2169 cm-1. This 2169 cm-1 species could not be retained with a membrane filter (molecular weight cutoff = 10,000) and showed unusual cyanide isotope shifts and a D2O shift. These observations together with metal content analyses indicate that the 2169 cm-1 band is due to a CuB.CN complex released from the enzyme. The same species could be produced by anaerobic partial reduction of the CN-inhibited ubiquinol oxidase and, furthermore, of the CN-inhibited cytochrome c oxidase; but not at all from the fully reduced CN-inhibited enzymes. These findings suggest that there is a common intermediate structure at the binuclear center of heme-copper respiratory enzymes in the partially reduced state from which the CuB center can be easily released upon cyanide-binding.
authors.affiliation: Department of Life Science, Faculty of Science, Himeji Institute of Technology, Kamigoori-cho, Akou-gun, Hyogo 678-12, Japan.
authors.forename: M
authors.initials: M
authors.lastname: Tsubaki
authors.forename: T
authors.initials: T
authors.lastname: Mogi
authors.forename: H
authors.initials: H
authors.lastname: Hori
authors.forename: M
authors.initials: M
authors.lastname: Sato-Watanabe
authors.forename: Y
authors.initials: Y
authors.lastname: Anraku
mesh_terms: D000693:Anaerobiosis; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D003300:Copper / Q000378:metabolism; D003486:Cyanides / Q000378:metabolism* / Q000494:pharmacology; D003573:Cytochrome b Group; D003580:Cytochromes / Q000737:chemistry* / Q000378:metabolism*; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods; D003576:Electron Transport Complex IV / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins; D006418:Heme / Q000378:metabolism; D008929:Mitochondria, Heart / Q000201:enzymology; D016297:Mutagenesis, Site-Directed; D010084:Oxidation-Reduction; D017354:Point Mutation; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017550:Spectroscopy, Fourier Transform Infrared / Q000379:methods
chemical_list: D003486:Cyanides; D003573:Cytochrome b Group; D003580:Cytochromes; D029968:Escherichia coli Proteins; D011994:Recombinant Proteins; D006418:Heme; D003300:Copper; C017168:cytochrome bo, E coli; D003576:Electron Transport Complex IV