Size dependence of the translational diffusion of large integral membrane proteins in liquid-crystalline phase lipid bilayers. A study using fluorescence recovery after photobleaching

Abstract

The translational diffusion of bovine rhodopsin, the Ca2+-activated ATPase of rabbit muscle sarcoplasmic reticulum and the acetylcholine receptor monomer of Torpedo marmorata was examined at a high dilution (molar ratios of lipid/protein .gtoreq. 3000/l) in liquid-crystalline phase phospholipid bilayer membranes by using the fluorescence recovery after photobleaching technique. These integral membrane proteins having MW of .apprx. 37,000 for rhodopsin, .apprx. 100,000 for the ATPase and .apprx. 250,000 for the acetylcholine receptor were reconstituted into membranes of dimyristoylphosphatidylcholine (rhodopsin and acetylcholine receptor), soybean lipids (acetylcholine receptor) and a total lipid extract of rabbit muscle sarcoplasmic reticulum (ATPase). The translational diffusion coefficients of all the proteins at 310 K were in the range (1-3) .times. 10-8 cm2/s. In consideration of the sizes of the membrane-bound portions of these proteins, this result is in agreement with the weak dependence of the translational diffusion coefficient on diffusing particle size predicted by continuum fluid hydrodynamic models for the diffusion in membranes. Lipid diffusion was also examined in the same lipid bilayers with the fluorescent lipid derivative N-(7-nitro-2,1,3-benzoxadiazol-4-yl)dimyristoylphosphatidylethanolamine. The translational diffusion coefficient for this lipid derivative was in the range (9-14) .times. 10-8 cm2/s at 310 K. In consideration of the dimensions if the lipid molecule, this value for the lipid diffusion coefficient is in agreement with the continuum fluid hydrodynamic model only if a near-complete slip boundary condition is assumed at the bilayer midplane. Alternatively, kinetic diffusion models may have to be invoked to explain the lipid diffusion behavior.