Influence of the intrinsic membrane protein bacteriorhodopsin on gel-phase domain topology in two-component phase-separated bilayers.
We have investigated the effect of the intrinsic membrane protein bacteriorhodopsin of Halobacterium halobium on the lateral organization of the lipid phase structure in the coexistence region of an equimolar mixture of dimyristoylphos-phatidylcholine and distearoylphosphatidylcholine. The fluorescence recovery after photobleaching (FRAP) technique was used to monitor the diffusion of both a lipid analog (N-(7-nitrobenzoxa-2,3-diazol-4-yl)-dimyristoylphosphatidyle thanolamine, NBD-DMPE) and fluorescein-labeled bacteriorhodopsin (Fl-BR). In the presence of bacteriorhodopsin, the mobile fractions of the two fluorescent probes display a shift of the percolation threshold toward lower temperatures (larger gel-phase fractions), independent of the protein concentration, from 43 degrees C (without bacteriorhodopsin) to 39 degrees C and 41 degrees C for NBD-DMPE and Fl-BR, respectively. Moreover, in the presence of bacteriorhodopsin, the gel-phase domains are much less efficient in restricting the diffusion of both probes than they are in the absence of the protein in the two-phase coexistence region. Bacteriorhodopsin itself, however, obstructs diffusion of NBD-DMPE and Fl-BR to about the same extent in the fluid phase of the two-phase region as it does in the homogeneous fluid phase. These observations suggest that 1) the protein induces the formation of much larger and/or more centrosymmetrical gel-phase domains than those formed in its absence, and 2) bacteriorhodopsin partitions almost equally between the coexisting fluid and gel phases. Although the molecular mechanisms involved are not clear, this phenomenon is fully consistent with the effect of the transmembrane peptide pOmpA of Escherichia coli investigated by electron spin resonance in the same lipid system.
Schram
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Department of Biochemistry, University of Virginia, Charlottesville 22908, USA.
Thompson
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D000345:Affinity Labels; D001436:Bacteriorhodopsins / Q000378:metabolism*; D004058:Diffusion; D004134:Dimyristoylphosphatidylcholine / Q000737:chemistry; D008051:Lipid Bilayers / Q000737:chemistry* / Q000378:metabolism; D010713:Phosphatidylcholines / Q000737:chemistry / Q000378:metabolism; D013696:Temperature
D000345:Affinity Labels; D008051:Lipid Bilayers; D010713:Phosphatidylcholines; D001436:Bacteriorhodopsins; C010942:1,2-distearoyllecithin; D004134:Dimyristoylphosphatidylcholine