Abstract
Cytochrome a 1 c 1 was highly purified from Nitrobacter agilis. The cytochrome contained heme a and heme c of equimolar amount, and its reduced form showed absorption peaks at 587, 550, 521, 434 and 416 nm. Molecular weight per heme a of the cytochrome was estimated to be approx. 100,000–130,000 from the amino acid composition. A similar value was obtained by determining the protein content per heme a. The cytochrome molecule was composed of three subunits with molecular weights of 55,000, 29,000 and 19,000, respectively. The 29 kd subunit had heme c.
Hemes a and c of cytochrome a 1 c 1 were reduced on addition of nitrite, and the reduced cytochrome was hardly autoxidizable. Exogenously added horse heart cytochrome c was reduced by nitrite in the presence of cytochrome a 1 c 1; K m values of cytochrome a 1 c 1 for nitrite and N. agilis cytochrome c were 0.5 mM and and 6 μM, respectively. V max was 1.7 mol ferricytochrome c reduced/min·mol of cytochrome a 1 c 1 The pH optimum of the reaction was about 8. The nitrite-cytochrome c reduction catalyzed by cytochrome a 1 c 1 was 61% and 88% inhibited by 44μM azide and cyanide, respectively. In the presence of 4.4 mM nitrate, the reaction was 89% inhibited. The nitrite-cytochrome c reduction catalysed by cytochrome a 1 c 1 was 2.5-fold stimulated by 4.5 mM manganous chloride. An activating factor which was present in the crude enzyme preparation stimulated the reaction by 2.8-fold, and presence of both the factor and manganous ion activated the reaction by 7-fold.
Cytochrome a 1 c 1 showed also cytochrome c-nitrate reductase activity. The pH optimum of the reaction was about 6. The nitrate reductase activity was also stimulated by manganous ions and the activating factor.
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Tanaka, Y., Fukumori, Y. & Yamanaka, T. Purification of cytochrome a 1 c 1 from Nitrobacter agilis and characterization of nitrite oxidation system of the bacterium. Arch. Microbiol. 135, 265–271 (1983). https://doi.org/10.1007/BF00413479
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DOI: https://doi.org/10.1007/BF00413479