Purification and properties of an unusual membrane-derived cytochrome b-561 from the purple phototrophic bacterium Rhodobacter capsulatus, which is structurally related to the bacteriochlorophyll-binding protein, LHII beta.
An abundant cytochrome b-561 was solubilized from Rhodobacter capsulatus membranes by successive treatments with perchlorate and butanol/water. Neither procedure was effective alone although they could be combined into a single step. Once solubilized, cytochrome b-561 was purified by standard chromatographic procedures used for water-soluble proteins without addition of butanol or detergents. Cytochrome b-561 appears to be highly acidic, it has a size greater than about 1000 kDa as isolated, and the subunit size measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis is less than 8 kDa. The redox potential measured by cyclic voltammetry is -65 mV at pH 7. The N-terminal amino acid sequence is identical to that of the Rb. capsulatus LHII beta light-harvesting bacteriochlorophyll binding protein subunit which has only 48 amino acid residues, and the mass, determined by mass spectroscopy, is identical to that of LHII beta. There is but one heme per two to three peptide chains of 5 kDa, which suggests that the two extraplanar ligands to the heme are on separate subunits. There is strong exciton splitting in the circular dichroism spectrum in the Soret region indicative of heme-heme interaction. The helix content based on far-uv CD is 41%. Together, these properties of cytochrome b-561 are very similar to those of isolated LHII alpha beta bacteriochlorophyll-protein complexes.
Bartsch
R G
RG
Department of Biochemistry, University of Arizona, Tucson 85721.
Caffrey
M S
MS
Van Beeumen
J J
JJ
Salamon
Z
Z
Tollin
G
G
Meyer
T E
TE
Cusanovich
M A
MA
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry; D002942:Circular Dichroism; D003573:Cytochrome b Group / Q000737:chemistry*; D045342:Light-Harvesting Protein Complexes; D008565:Membrane Proteins / Q000737:chemistry*; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D045322:Photosynthetic Reaction Center Complex Proteins / Q000737:chemistry*; D016354:Rhodobacter capsulatus / Q000737:chemistry*; D013057:Spectrum Analysis
D001426:Bacterial Proteins; D003573:Cytochrome b Group; D045342:Light-Harvesting Protein Complexes; D008565:Membrane Proteins; D045322:Photosynthetic Reaction Center Complex Proteins; C081485:light-harvesting protein complex II, Rhodobacter capsulatus; C023646:cytochrome b561