21] Removal of detergents from membrane proteins

Abstract

Publisher Summary This chapter describes the different circumstances under which detergent removal may be advisable. It also discusses the detergent properties, which may influence the ease with which detergents may be removed. During the course of the purification of a membrane protein, it may be necessary to remove excess detergent at three distinct points in the protocol. The first arises after initial solubilization of membranes. As this step usually requires detergent excess to give maximal dispersion of membrane proteins and lipids, removal of excess detergent at this stage usually improves the quality of subsequent chromatographic steps. The most efficient processes are those, which involve adsorption of the detergent–membrane protein complex to a chromatographic support, followed by extensive washing with buffer containing the new detergent. Many analytical procedures have specific requirements related to the physical properties of the detergents to be used; thus, preparation of samples may necessitate detergent exchange or removal. Chromatographic techniques for detergent removal or exchange come in two basic varieties. The first procedure involves the adsorption of the membrane detergent–protein complex to the chromatographic medium, followed by thorough washing with buffer including the new detergent. The second procedure involves gel filtration chromatography.