Isolation and purification of membrane-bound cytochrome b-560 from photosynthetic bacterium Chromatium vinosum

Abstract

A membrane-bound cytochrome of the b-type (cytochrome b-560) was success-fully purified from chromatophores of the photosynthetic purple sulfur bacterium Chromatium vinosum by treatment with sodium cholate, sodium deoxycholate, sodium thiocyanate, and bacterial alkaline protease (EC 3·4·21·14) followed by gel filtration.The purified cytochrome b-560 showed the absorption maxima at 279, 412.5 and 533 nm in the oxidized form, and 427, 530 and 560 nm in the reduced form. Reduced-minus-oxidized difference millimolar absorption coefficient was 14.0 for a wavelength pair, 560 minus 540 nm.Isolated cytochrome b-560 was electrophoretically homogeneous, and its minimal molecular weight was estimated to the 13,000 by SDS polyacrylamide gel electrophoresis.The midpoint potential at pH 8.0 was -110mV, and was not dependent on the ambient pH in the pH range of 6.8 to 8.8.

GATA‐1 isoforms differently contribute to the production and compartmentation of reactive oxygen species in the myeloid leukemia cell line K562

“…Notably, we observed that spectral features at 560 nm and 412.5 nm could resemble the absorption spectra of the reduced and oxidized form of cytochrome b-560, that, as previously reported, shows absorption maxima at 279, 412.5, and 533 nm in the oxidized form, and at 427, 530, and 560 nm in the reduced form (Doi, Takamiya & Nishimura, 1982). Therefore, the combined absorbance spectra variations at both 412.5 and 560 nm prompted thus us to hypothesize that the expression of specific GATA-1 isoforms could be associated with different redox states of this cytochrome, a component of complex II of the mitochondrial respiratory chain that is responsible for transferring electrons from succinate to ubiquinone (Yu, Xu, Haley & Yu, 1987).…”

Isolation and properties of membrane-bound cytochrome c-552 from photosynthetic bacterium Chromatium vinosum

“…A: chromatophores. B: 0-10% Ammonium sulfate fraction prepared as described earlier [4 ]. C: Fraction A.…”

“…This difference may be due to some modification of the cytochrome molecule caused by isolation and purification or to change of microenvironment surrounding the molecule. The lowering of the midpoint potential was found in cytochrome b isolated from mammalian mitochondria [8] and cytochrome b-560 isolated from Chromatium vinosum [4].…”

“…The highpotential cytochrome c-555 was not isolated, which suggests that this cytochrome is more labile than cytochrome c-552 under the isolation conditions employed in this study. The separation of cytochrome c-552 from other intrinsic proteins could be carried out at present only after the treatment with a proteolytie enzyme, Nagarse [4], although a partial digestion of native cytochromes might occur (see below).…”

“…Recently, we have solubilized and purified a membrane-bound cytochrome b-560 from Chromatium vinosum chromatophores by the treatment with detergents and a proteolytic enzyme [4]. In this study, we successfully purified the membrane-bound cytochrome, cytochrome c-552, from Fraction A of Chromatium vinosum, by a similar treatment and investigated some of its properties.…”