Localization of chromatophore proteins of Rhodobacter sphaeroides. I. Rapid Ca(2+)-induced fusion of chromatophores with phosphatidylglycerol liposomes for proteinase delivery to the luminal membrane surface.
A protease delivery system was developed for the exclusive and controlled digestion of proteins exposed at the morphological inside (periplasmic surface) of Rhodobacter sphaeroides chromatophores. In this procedure, proteinase K is encapsulated within large unilamellar liposomes which are fused to the chromatophores in the presence of Ca2+ ions. The liposomes were prepared by a detergent dialysis procedure from native phosphatidylglycerol and found to undergo rapid bilayer fusion with purified chromatophore preparations above a threshold concentration of 12.5 mM CaCl2. The fusion process was complete within 10 min at 35 mM Ca2+ with about 80% of the pigment located in the fusion products. Electron micrographs of freeze-fracture replicas confirmed the intermixing of the lipid bilayers and the unilamellar structure of the fused membrane vesicles. The procedure did not affect the labile B800 chromophore of the B800-850 antenna complex, but reduced slightly the absorption due to the B875 core antenna. Emission from both light-harvesting complexes was increased in the fused membranes, suggesting a partial dissociation of photosynthetic units in the expanded bilayer. The results, together with those presented in the following paper (Theiler, R., and Niederman, R. A. (1991) J. Biol. Chem. 266, 23163-23168), demonstrate that this new method fulfills the stringent requirements for a successful delivery of macromolecules to the chromatophore interior.
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* / Q000648:ultrastructure; D001426:Bacterial Proteins / Q000032:analysis* / Q000378:metabolism; D001431:Bacteriological Techniques; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D002855:Chromatography, Thin Layer; D010450:Endopeptidases / Q000378:metabolism; D005614:Freeze Fracturing; D007700:Kinetics; D008081:Liposomes; D008565:Membrane Proteins / Q000032:analysis* / Q000378:metabolism; D008854:Microscopy, Electron; D010715:Phosphatidylglycerols / Q000378:metabolism*; D012242:Rhodobacter sphaeroides / Q000737:chemistry* / Q000378:metabolism
D001426:Bacterial Proteins; D008081:Liposomes; D008565:Membrane Proteins; D010715:Phosphatidylglycerols; D010450:Endopeptidases; D002118:Calcium