5] Phospholipid substitution of the purple membrane

Abstract

This chapter discusses the phospholipid substitution of the purple membrane. Bacteriorhodopsin is an ideal protein for studies on protein–lipid interactions in artificial membranes. It can easily be reconstituted with various phospholipids. A complication is, however, the fact that the purple membrane sheets used for the reconstitution experiments contain not only bacteriorhodopsin but also endogenous lipids. Consequently, these endogenous lipids will be co-reconstituted with bacteriorhodopsin, and under some conditions these lipids may remain specifically associated with the protein. For that reason, it is a prerequisite for studies on the interaction between bacteriorhodopsin and lipids to remove the endogenous purple membrane lipids from the protein before it is reconstituted. This can be achieved in two different ways. First, bacteriorhodopsin can be delipidated with detergents. It has been reported that 99% of the endogenous lipids have been removed by this method. The second way—somewhat less laborious method—is based on exchange of endogenous for exogenous lipids, mediated by the detergent cholate.