[The cytochrome cbo from the obligate methylotroph Methylobacillus flagellatus KT is a cytochrome-c oxidase].
The oxidase cho of Methylobacillus flagellatus KT was purified to homogeneity by nondenaturing gel electrophoresis, and the kinetic properties and substrate specificity of the enzyme were studied. Ascorbate and ascorbate/N,N,N',N'-tetramethyl-p-phenylenediamine (TMPD) were oxidized by cbo with a pH optimum of 8.3. When TMPD served as electron donor for the oxidase cho, the optimal pH (7.0 to 7.6) was determined from the difference between respiration rates in the presence of ascorbate/TMPD and of only ascorbate. The kinetic constants, determined at pH 7.0, were as follows: oxidation by the enzyme of reduced TMPD at pH 7.0 was characterized by KM = 0.86 mM and Vmax = 1.1 mumol O2/(min mg protein), and oxidation of reduced cytochrome c from horse heart was characterized by KM = 0.09 mM and Vmax = 0.9 mumol O2/(min mg protein) Cyanide inhibited ascorbate/TMPD oxidase activity (Ki = 4.5-5.0 microM). The soluble cytochrome cH (12 kDa) partially purified from M. flagellatus KT was found to serve as the natural electron donor for the oxidase cbo.
Strom
E V
EV
Department of Microbiology, Biological Faculty, Moscow State University, Vorob'evy gory, Moscow, 119899 Russia.
Dinarieva
T Iu
TIu
Hetrusov
A I
AI
D000818:Animals; D001205:Ascorbic Acid / Q000737:chemistry / Q000378:metabolism; D003486:Cyanides / Q000494:pharmacology; D003580:Cytochromes / Q000302:isolation & purification / Q000378:metabolism*; D003576:Electron Transport Complex IV / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D006736:Horses; D006863:Hydrogen-Ion Concentration; D020631:Methylobacillus / Q000201:enzymology*; D010084:Oxidation-Reduction; D013379:Substrate Specificity; D013772:Tetramethylphenylenediamine / Q000737:chemistry / Q000378:metabolism
D003486:Cyanides; D003580:Cytochromes; C038862:cytochrome o oxidase; D003576:Electron Transport Complex IV; D013772:Tetramethylphenylenediamine; D001205:Ascorbic Acid