In meso crystallogenesis. Compatibility of the lipid cubic phase with the synthetic digitonin analogue, glyco-diosgenin

Abstract

Digitonin has long been used as a mild detergent for extracting proteins from membranes for structure and function studies. As supplied commercially, digitonin is inhomogeneous and requires lengthy pre-treatment for reliable downstream use. Glyco-diosgenin (GDN) is a recently introduced synthetic surfactant with features that mimic digitonin. It is available in homogeneously pure form. GDN is proving to be a useful detergent, particularly in the area of single-particle cryo-electron microscopic studies of membrane integral proteins. With a view to using it as a detergent for crystallization trials by the in meso or lipid cubic phase method, it was important to establish the carrying capacity of the cubic mesophase for GDN. This was quantified in the current study using small-angle X-ray scattering for mesophase identification and phase microstructure characterization as a function of temperature and GDN concentration. The data show that the lipid cubic phase formed by hydrated monoolein tolerates GDN to concentrations orders of magnitude in excess of those used for membrane protein studies. Thus, having GDN in a typical membrane protein preparation should not deter use of the in meso method for crystallogenesis.