Localization of chromatophore proteins of Rhodobacter sphaeroides. II. Topography of cytochrome c1 and the Rieske iron-sulfur protein as determined by proteolytic digestion of the outer and luminal membrane surfaces.
Proteinase K was used to degrade membrane proteins exposed at the outer (cytoplasmic) and inner (periplasmic) surface of sealed, uniformly oriented chromatophore vesicles of Rhodobacter sphaeroides. Exclusive and controlled digestion of the chromatophore interior was achieved after Ca(2+)-induced fusion with large unilamellar phosphatidylglycerol liposomes containing microencapsulated enzyme. Reaction center subunit H, which served as a marker for the outer surface, was degraded to a slightly smaller product in chromatophores. This protein remained intact after liposome-chromatophore fusion, suggesting that the intermixing of lipid bilayers proceeded without significant leakage of the aqueous vesicle contents. In contrast, while cytochrome c1 was not affected in chromatophores, 70-75% was degraded within 60 min after liposome-chromatophore fusion. These results support an arrangement in which the bulk of this protein, including the mesoheme component and active site residues, faces the periplasmic side of the membrane. Although current functional models for the cytochrome bc1 complex predict that the Rieske iron-sulfur center interacts with cytochrome c1 in the periplasm, the iron-sulfur protein resisted proteolytic attack in the liposome-chromatophore fusion products under conditions that caused extensive degradation of cytochrome c1. Two cleavage products of the iron-sulfur protein were observed after the digestion of chromatophores, suggesting both a heterogeneity in the population of this protein and the exposure of at least part of its molecular mass to the cytoplasm.
Theiler
R
R
Department of Molecular Biology and Biochemistry, Rutgers University, Piscataway, New Jersey 08855-1059.
Niederman
R A
RA
D020130:Bacterial Chromatophores / Q000737:chemistry*; D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D003575:Cytochromes c1 / Q000032:analysis*; D014450:Electron Transport Complex III; D019286:Endopeptidase K; D007506:Iron-Sulfur Proteins / Q000032:analysis*; D008081:Liposomes; D010715:Phosphatidylglycerols / Q000378:metabolism; D012242:Rhodobacter sphaeroides / Q000737:chemistry*; D012697:Serine Endopeptidases
D007506:Iron-Sulfur Proteins; D008081:Liposomes; D010715:Phosphatidylglycerols; C037989:Rieske iron-sulfur protein; D003575:Cytochromes c1; D012697:Serine Endopeptidases; D019286:Endopeptidase K; D014450:Electron Transport Complex III; D002118:Calcium