Biochemical and spectroscopic properties of cyanide-insensitive quinol oxidase from Gluconobacter oxydans.
Cyanide-insensitive quinol oxidase (CioAB), a relative of cytochrome bd, has no spectroscopic features of hemes b(595) and d in the wild-type bacteria and is difficult to purify for detailed characterization. Here we studied enzymatic and spectroscopic properties of CioAB from the acetic acid bacterium Gluconobacter oxydans. Gluconobacter oxydans CioAB showed the K(m) value for ubiquinol-1 comparable to that of Escherichia coli cytochrome bd but it was more resistant to KCN and quinone-analogue inhibitors except piericidin A and LL-Z1272gamma. We obtained the spectroscopic evidence for the presence of hemes b(595) and d. Heme b(595) showed the alpha peak at 587 nm in the reduced state and a rhombic high-spin signal at g = 6.3 and 5.5 in the air-oxidized state. Heme d showed the alpha peak at 626 and 644 nm in the reduced and air-oxidized state, respectively, and an axial high-spin signal at g = 6.0 and low-spin signals at g = 2.63, 2.37 and 2.32. We found also a broad low-spin signal at g = 3.2, attributable to heme b(558). Further, we identified the presence of heme D by mass spectrometry. In conclusion, CioAB binds all three ham species present in cytochrome bd quinol oxidase.
Department of Biomedical Chemistry, the University of Tokyo, Bunkyo-ku, Japan. tmogi@m.u-tokyo.ac.jp
Tatsushi
T
Mogi
Yoshitaka
Y
Ano
Tomoko
T
Nakatsuka
Hirohide
H
Toyama
Atsushi
A
Muroi
Hideto
H
Miyoshi
Catharina T
CT
Migita
Hideaki
H
Ui
Kazuro
K
Shiomi
Satoshi
S
Omura
Kiyoshi
K
Kita
Kazunobu
K
Matsushita
D002851:Chromatography, High Pressure Liquid; D003486:Cyanides / Q000494:pharmacology*; D020600:Gluconobacter oxydans / Q000737:chemistry / Q000187:drug effects / Q000201:enzymology; D020128:Inhibitory Concentration 50; D015394:Molecular Structure; D010088:Oxidoreductases / Q000037:antagonists & inhibitors / Q000737:chemistry*; D013057:Spectrum Analysis
D003486:Cyanides; D010088:Oxidoreductases; C020094:duroquinol oxidase