MEMBRANE LIPID INFLUENCES ON THE ENERGETICS OF THE METARHODOPSIN I and METARHODOPSIN II CONFORMATIONAL STATES OF RHODOPSIN PROBED BY FLASH PHOTOLYSIS

Abstract– We have investigated the relationship between rhodopsin photochemical function and the retinal rod outer segment (ROS) disk membrane lipid composition using flash photolysis techniques. Bovine rhodopsin was combined with various phospholipids to form recombinant membrane vesicles, in which the lipid acyl chain composition was maintained at that of egg phosphatidylcholine (PC), while the nature of the headgroups was varied. The ratio of metarhodopsin II (MII)/metarhodopsin I (MI) in these recombinants produced by an actinic flash was investigated as a function of pH, and compared with the photochemical activity observed for rhodopsin in native ROS membranes and dimyristoylphosphatidylcholine recombinants. In recombinants made with lipids derived from egg PC, as well as in native ROS membranes, MI and Mil were found to be present in a pH‐dependent, acid‐base equilibrium on the millisecond timescale. The recombinants made with phospholipids containing unsaturated acyl chains were capable of full native‐like Mil production, but each demonstrated a titration curve with a different pK. In addition, some of the recombinants exhibited apparent deviations from the Henderson‐Hasselbalch curve shape. The presence of either phosphatidylethanola‐mine (PE) or phosphatidylserine (PS) headgroups appeared to increase the amount of Mil produced. This may result from alteration of the curvature free energy, in the case of PE, and from the influence of the membrane surface potential in the case of PS. An investigation of the effects of temperature on the MI‐MII transition in native ROS membranes and the recombinants was also carried out. The results suggest that the thermodynamic parameters characterizing the MI and Mil conformational states of rhodopsin are influenced by the membrane bilayer environment, indicating a possible role of lipids in the visual process.