Structure of Escherichia coli membranes. Glycerol auxotrophs as a tool for the analysis of the phospholipid head-group region by deuterium magnetic resonance

Abstract

Glycerol selectively deuterated at various positions was synthesized and supplied to the growth medium of E. coli strain T131 GP, which is defective in endogenous glycerol synthesis and glycerol degradation and lacks the ability to synthesize cardiolipin. The procedure enables the stereospecific labeling of the membrane phospholipids (.apprx. 80% phosphatidylethanolamine, .apprx. 20% phosphatidylglycerol). Deuterium magnetic resonance [2H-NMR] spectra were obtained for cell membranes and lipid dispersions either from total lipid extracts or from purified phosphatidylglycerol or phosphatidylethanolamine. When glycerol deuterated at various positions was used, all resonances of the phospholipid glycerol backbone and the terminal glycerol moiety in phosphatidylglycerol could be assigned. The molecular conformation of the glycerol backbone is independent of the phospholipid species investigated and also is not altered by the presence of high amounts of membrane proteins. For the quantitative interpretation of the 2H-NMR splittings, a model is proposed which assumes essentially free rotation around the glycerol c(2).sbd.C(3) bond combined with an asymmetric and restricted jump process around the C(1).sbd.C(2) bond. This model is compatible with known X-ray structures of phospholipid molecules. The 2 deuterons of both the glycerol backbone C(1) and C(3) segments were magnetically inequivalent. Stereoselective monodeuteration eliminated one set of quadrupole splittings in both cases.