Purification and characterization of two membrane-bound c-type cytochromes from a facultative alkalophilic Bacillus.
The membrane fraction of the facultative alkalophilic bacterium, Bacillus YN-2000, was found to contain considerably larger amounts of two c-type cytochromes, cytochromes c-553 and c-552, when the bacterium was grown at pH 10 than when it was grown at lower pHs (pH 7-9). In particular, cytochrome c-553 was present in a much higher amount in the cells grown at pH 10 than in those grown at pH 8. Cytochromes c-553 and c-552, which are membrane-bound proteins, were purified to electrophoretically homogeneous states from Bacillus YN-2000. Cytochrome c-553 showed absorption peaks at 553, 524, and 417 nm in the reduced form, and a peak at 411 nm in the oxidized form. Its molecular weight was estimated to be 10,500 from the results of SDS-polyacrylamide gel electrophoresis. However, its molecular weight was estimated to be 127,000 by gel filtration. Therefore, it seemed to occur as an oligomer in solution. The isoelectric point of cytochrome c-553 was determined to be 3.9. Its midpoint redox potential was found to be +87 mV in the pH region from 6 to 8. Cytochrome c-553 reacted with cytochrome c oxidase of the bacterium and the reaction was greatly accelerated in the presence of poly-L-lysine. Cytochrome c-552 showed absorption peaks at 552, 521, and 416 nm in the reduced form, and a peak at 408 nm in the oxidized form. The cytochrome molecule seemed to be composed of six different subunits, with molecular masses of 40, 32, 19, 17, 14, and 12 kDa, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)
Yumoto
I
I
Department of Life Science, Faculty of Bioscience and Biotechnology, Tokyo Institute of Technology, Kanagawa.
Fukumori
Y
Y
Yamanaka
T
T
D000596:Amino Acids / Q000032:analysis; D001407:Bacillus / Q000201:enzymology*; D002462:Cell Membrane / Q000201:enzymology; D002853:Chromatography, Liquid; D003574:Cytochrome c Group / Q000302:isolation & purification*; D003576:Electron Transport Complex IV / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D010084:Oxidation-Reduction; D011199:Potentiometry; D013057:Spectrum Analysis
D000596:Amino Acids; D003574:Cytochrome c Group; D003576:Electron Transport Complex IV