Deoxylysolecithin and a new biphenyl detergent as solubilizing agents for bovine rhodopsin. Functional test by formation of metarhodopsin II and binding of G-protein

Abstract

The protein-detergent interaction in rhodopsin-detergent micelles has been investigated by using formation of metarhodopsin II (MII) as a monitor. Two detergents of different structural rigidity have been applied. One of them is [3-(lauroyloxy)propyl]phosphorylcholine, which has a high conformational flexibility in its hydrophobic moiety like most of the known detergents for rhodopsin. This deoxylysolecithin was originally designed as a detergent for membrane proteins by Weltzien [Weltzien, H. U. (1979) Biochim. Biophys. Acta 559, 259-287]. The other detergent, which is highly rigid in its hydrophobic part, has been developed for this study. It consists of a biphenyl derivative and a hydrophilic octaethylene oxide group. Both the formation kinetics of MII and the position of its equilibrium with its tautomeric form, metarhodopsin I (MI), strongly differed in the deoxylysolecithin and biphenyl detergent. Deoxylysolecithin caused very fast MII formation and shifted the equilibrium strongly to MII, like other detergents with alkyl chains as the hydrophobic part. In the biphenyl detergent, however, formation of MII was slow and the MI/MII equilibrium similar to that in the native system. For rhodopsin reconstituted in lipid bilayers, normal MII formation requires a well-adjusted fluidity of the hydrocarbon environment of the protein [Baldwin, P. A., & Hubbell, W. L. (1984) Biochemistry 24, 2633-2639], which was explained by an appropriate interfacial pressure at the protein-lipid interface. Extension of this concept would indicate that in the micellar core a degree of fluidity comparable to that of the disk membrane is just achieved with the highly rigid biphenyl structure.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of Digitonin on the Rhodopsin Meta I‐Meta II Equilibrium¶

“…The slowing of the rate of conversion of Meta I 480 into Meta II with increasing digitonin‐to‐phospholipid ratio seen here is in general agreement with previous studies (38,39). Similar slowing was previously associated with another synthetic detergent having a rigid structure similar to the steroid portion of digitonin (40), and addition of cholesterol to reconstituted rhodopsin membranes has been shown to slow approach to the Meta I 480 ‐Meta II equilibrium and to back shift it (41). A similar effect was seen for cholesterol‐depleted and ‐enriched disk membranes (42).…”

Amphipols, Nanodiscs, and Fluorinated Surfactants: Three Nonconventional Approaches to Studying Membrane Proteins in Aqueous Solutions

“…Once the protein has been extracted, however, what the biochemist needs is to keep it soluble and to prevent it from aggregating, conditions that can be provided by surfactants that are not necessarily able to solubilize membranes. This has led to the development of such molecules as tripod amphiphiles (17)(18)(19), surfactants with rigid hydrophobic tails containing cycles (20), peptitergents (21), lipopeptides (22,23), peptergents (24), fluorinated surfactants (FSs) (25,26), or amphipathic polymers called amphipols (APols) (27), most of which are not good membrane solubilizers (for brief overviews, see References 11 and 28). A major part of this review is devoted to the latter two approaches.…”

Evidence for the specific interaction of a lipid molecule with rhodopsin which is altered in the transition to the active state metarhodopsin II1

“…Although illuminated rhodopsin in detergent is a potent activator of transducin, assuming that the detergent concentration is low enough [4, 5], it is known that a more fluid lipid bilayer (e.g. [6–10]) or fluid detergents [11–13]favor MII, and that the charge of the lipid head groups modifies the pH dependence of the MI/MII equilibrium by altering the surface charge of the membrane [9, 14]. The increased fluidity of the environment is thought to enhance MII formation by facilitating a volume increase in MII [11, 15].…”