Approaches to determining membrane protein structures to high resolution: do selections of subpopulations occur?

Abstract

Three different methods are currently used for the study of high-resolution structures of membrane proteins: X-ray crystallography, electron crystallography, and nuclear magnetic resonance (NMR) spectroscopy. Thus far, all methods combined have yielded a rather modest number of crystal structures that have been solved at the atomic level. It is hypothesized here that different methods may select different populations of proteins on the basis of various properties. Thus, protein stability may be a significant factor in the formation of three-dimensional (3D) crystals from detergent solutions, since exposure of hydrophobic protein zones to water may cause structural perturbation or denaturation in conformationally labile proteins. This is different in the formation of two-dimensional (2D) crystals where a protein remains protected in its native membrane environment. A biological selection mechanism may therefore be operative in that highly ordered lattices may form only if strong protein–protein interactions are relevant in vivo, thereby limiting the number of proteins that are amenable to electron crystallography. Keeping a protein in a bilayer environment throughout 3D crystallization maintains the lateral pressure existing in native membranes. This can be accomplished by using lipidic cubic phases. Alternatively, the hydrophobic interface of a membrane protein may be spared from contact with water by crystallization from organic solvents where the polar caps are protected in reverse micelles by using appropriate detergents. Some of the criteria that are useful in optimizing the various approaches are given. While the usefulness of complementary methods seems obvious, the results presented may be particularly critical in recognizing key problems in other structural approaches.