The Cytochrome cbo from the Obligate Methylotroph Methylobacillus flagellatus KT Is a Cytochrome c Oxidase |
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Authors: | Strom E V Dinarieva T Yu Netrusov A I |
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Institution: | (1) Department of Microbiology, Biological Faculty, Moscow State University, Vorob'evy gory, Moscow, 119899, Russia |
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Abstract: | The cbo-type oxidase of Methylobacillus flagellatus KT was purified to homogeneity by preparative native 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 cytochrome cbo with a pH optimum of 8.3. With TMPD as an electron donor for the cbo-type oxidase, the optimal pH (7.0 to 7.6) was determined from the difference between respiration rates in the presence of ascorbate/TMPD and only ascorbate. The kinetic constants determined at pH 7.0 were as follows: oxidation by the enzyme of reduced TMPD was characterized by K
M = 0.86 mM and V
max = 1.1 mol O2/(min mg protein), and oxidation of reduced horse heart cytochrome c was characterized by K
M = 0.09 mM and V
max = 0.9 mol O2/(min mg protein). Cyanide inhibited ascorbate/TMPD–oxidase activity (K
i = 4.5–5.0 M). The soluble cytochrome c
H (12 kDa), partially purified from M. flagellatus KT, was found to serve as a natural electron donor for the cbo-type oxidase. |
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Keywords: | obligate methylotroph Methylobacillus flagellatus KT cytochrome c cytochrome c oxidase cbo-type oxidase |
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