Conformational Studies of Membrane Proteins by High-Resolution NMR

Abstract

The lack of conformational information for membrane proteins is a great impediment to understanding the functional mechanisms of biological membranes. The problem of associated lipids has prevented extensive application of X-ray diffraction or high-resolution NMR to conformational characterization of membrane proteins. Difficulties in obtaining crystals suggests that high-resolution NMR might be a more promising approach for such proteins. With currently available methods, high-resolution 1H NMR spectra suitable for detailed conformational analyses are not obtained from proteins bound to native membranes1,2 or from reconstituted systems consisting of proteins bound to phospholipid vesicles.3,4 We have been able to obtain appropriate NMR spectra for membrane proteins and polypeptides incorporated into fully deuterated micelles. Proteins which have been tested include melittin,5 glucagon,6 phospholipase A2, glycophorin A, snake venom cardiotoxin and myelin basic protein (unpublished results). For these proteins, the conformation of the membrane protein is preserved in the micelle and the micellar protein-lipid complex is homogeneous and stoichiometrically well-defined5–7 (unpublished results).