MODULATION OF METARHODOPSIN FORMATION BY CHOLESTEROL-INDUCED ORDERING OF BILAYER LIPIDS

The effect of lipid ordering on the kinetics and extent of metarhodopsin II (meta II) formation was evaluated in bovine rhodopsin which had been reconstituted into phosphatidylcholine vesicles containing 0, 15, and 30 mol % cholesterol. The rate of establishment of the dynamic equilibrium between metarhodopsin I (meta I) and the two kinetically distinguished forms of meta II in the branched meta II model [meta IIfastand meta IIslow; Straume, M., Mitchell, D. C., Miller, J. L., & Litman, B. J. (1990) Biochemistry (preceding paper in this issue)] is derived from kinetic measurements of rhodopsin photolysis in these vesicle systems at several temperatures. Values of the meta I ↔ meta IItotalequilibrium constant, are calculated from the derived model-dependent rate constants, and are shown to be equivalent to those derived from rapidly acquired absorbance spectra. The presence of 30 mol % cholesterol reduces Keqby approximately 50% between 10 and 37 °C. Analysis of the model-dependent parameters in terms of ΔH and ΔS reveals that cholesterol raises the free energy of meta IIslow, relative to meta I, by increasing ΔH whereas it raises the relative free energy of meta IIfastby making ΔS of meta IIfastrelative to meta I less positive. The reduction in Keqby both temperature and cholesterol is found to be directly correlated with a parameter that reflects the free volume available for molecular motion in the hydrophobic core of the bilayer [Straume, M., & Litman, B.J. (1988) Biochemistry 27, 7723-7733]. This correlation is interpreted as evidence that meta II formation is modulated by the packing properties of the surrounding lipid bilayer rather than by any specific rhodopsin-lipid interactions. Such modification of the functional competence of an integral membrane protein by variation of the lipid bilayer composition provides a potential control mechanism in the essentially isothermal environment of living cells. The implications of the results for the nature of the meta I to meta II transition and for the role of the lipid bilayer in visual signal transduction are discussed. © 1990, American Chemical Society. All rights reserved.