Spatial organization of bacteriorhodopsin in model membranes. Light-induced mobility changes.
Bacteriorhodopsin is a proton-transporting membrane protein in Halophilic archaea, and it is considered a prototype of membrane transporters and a model for G-protein-coupled receptors. Oligomerization of the protein has been reported, but it is unknown whether this feature is correlated with, for instance, light activation. Here, we have addressed this issue by reconstituting bacteriorhodopsin into giant unilamellar vesicles. The dynamics of the fully active protein was investigated using fluorescence correlation spectroscopy and freeze fracture electron microscopy. At low protein-to-lipid ratios (<1:10 w/w), a decrease in mobility was observed upon protein photoactivation. This process occurred on a second time scale and was fully reversible, i.e. when the dark-adapted state was reestablished the lateral diffusion rate of the protein was returned to that prior to activation. A similar decrease in lateral mobility as observed upon photoactivation was obtained when bacteriorhodopsin was reconstituted at high protein-to-lipid ratios (>1:10 w/w). We interpret the shifts in mobility during light adaptation as being caused by transient photoinduced oligomerization of bacteriorhodopsin. These observations are fully supported by freeze-fracture electron microscopy, and the size of the clusters during photoactivation was estimated to consist of two or three trimers.
Kahya
Nicoletta
N
Ultrafast Laser and Spectroscopy Laboratory, Optical Sciences, Materials Science Centre, University of Groningen, Nijenborgh 4, 9747 AG Groningen, The Netherlands. N.Kahya@chem.rug.nl
Wiersma
Douwe A
DA
Poolman
Bert
B
Hoekstra
Dick
D
D001436:Bacteriorhodopsins / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D001692:Biological Transport; D002462:Cell Membrane / Q000378:metabolism*; D004058:Diffusion; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D004578:Electron Spin Resonance Spectroscopy; D005614:Freeze Fracturing; D007700:Kinetics; D008027:Light; D008081:Liposomes / Q000378:metabolism; D008854:Microscopy, Electron; D015233:Models, Statistical; D010455:Peptides / Q000737:chemistry; D011522:Protons; D013050:Spectrometry, Fluorescence
D008081:Liposomes; D010455:Peptides; D011522:Protons; D001436:Bacteriorhodopsins