Structure and dynamics of the phosphatidylcholine and the phosphatidylethanolamine head group in L-M fibroblasts as studied by deuterium nuclear magnetic resonance.

Abstract

Mouse fibroblast L‐M cells were grown in tissue culture medium containing selectively deuterated choline or ethanolamine. Both compounds were incorporated into the corresponding phospholipids at levels greater than 50% thus leading to a selective deuteration of these phospholipid head groups. Choline and ethanolamine were labeled at either the alpha‐ or the beta‐carbon atom and well‐resolved deuterium and phosphorus n.m.r. spectra were obtained from intact cells, crude plasma membranes and lipid extracts, leading to the following conclusions. (i) A large fraction, if not all, of the phospholipids in the intact L‐M cell membranes were organized in a liquid crystalline bilayer. (ii) The phosphoethanolamine and the phosphocholine head group conformation were found to be remarkably similar in pure lipid bilayers and in intact L‐M cell membranes with the head group dipoles being oriented parallel to the membrane surface. (iii) The deuterium T1 spin lattice relaxation times fell in the range of 7‐25 ms and were similar in intact L‐M cells and in pure lipid model membranes, suggesting that the two head groups are not involved in strong interactions with membrane proteins. The rotational diffusion rate of the two head groups was reduced by at least a factor of 10 compared to molecules of the same size in aqueous solution. (iv) The phosphocholine head group was sensitive to the size and sign of membrane surface charges as verified in mixing experiments with charged lipids. In L‐M cell membranes the phosphocholine appeared to sense an electrically neutral environment in spite of the fact that L‐M cell membranes contain 10‐20% negatively charged lipids.

Effects of diacylglycerols on conformation of phosphatidylcholine headgroups in phosphatidylcholine/phosphatidylserine bilayers

“…A different picture emerges when considering the headgroup region. The T1z values in the absence of DAGs are similar to those reported by Scherer and Seelig (1987). The presence of all DAGs other than DP caused both a-and ,B-deuteron TI, values to increase nearly twofold (Table 1), reflecting an increased motional freedom of the choline headgroups.…”

“…A change in the average order parameters of the PC headgroup would be expected to shift both a-and ,B-splittings in the same direction and thus cannot account for the counterdirectional change (Scherer and Seelig, 1989). A similar change in PC headgroup quadrupole splittings has been reported by others investigating effects on membrane surface charge density of various charge carriers including metal ions (Brown and Seelig, 1977;Akutsu and Seelig, 1981;Altenbach and Seelig, 1984;Macdonald and Seelig, 1987), hydrophobic ions (Altenbach and Seelig, 1985), the membrane-bound anion SCN- (Macdonald and Seelig, 1988), charged amphiphiles (Seelig et al, 1987; Scherer and ), the zwitterionic lipophilic molecule phloretin (Bechinger and Seelig, 1991), charged phospholipids (Sixl and Watts, 1983;Scherer and Seelig, 1987), and peptides (Dempsey and Watts, 1987;Dempsey et al, 1989; Kuchinka and Roux et al, 1989; Beschiaschvili and Seelig, 1990a,b). These results have been used to develop a molecular voltmeter model (Scherer and Seelig, 1989) in which membrane-bound ions assert a Coulombic force on the P--N+ dipole of the choline moiety, repelling or attracting the positive nitrogen with respect to the membrane surface, which can cause changes in the conformation of the headgroup and alterations of average C-D bond angles with the molecular axis.…”

“…A 2H NMR spectrum of fully hydrated lipid bilayers in the liquid crystalline phase is a superposition of the powder patterns arising from the deuterons at the a-and /3-carbon positions of the choline headgroup. In the DPPC-d4 spectra shown, the outer symmetric pair of peaks corresponds to the deuterons located at the a-carbon of the choline moiety and the inner pair corresponds to the deuterons at the ,8-carbon (Scherer and Seelig, 1987). The peak-to-peak quadrupole splittings (Av) are determined by measuring the distance between symmetric peaks.…”

Calculations of the electrostatic potential adjacent to model phospholipid bilayers

“…Subsequently, we rotated several bonds to obtain a conformation that satisfied the following criteria: 1) the glycerol backbone would be oriented in the same direction as the fatty acid chains, perpendicular to the plane of the bilayer, 2) the polar headgroup would lie approximately parallel to the membrane's surface, and 3) all saturated fatty acid chain bonds would be in the trans conformation. The glycerol backbone and headgroup orientations are based on the available experimental evidence from nuclear magnetic resonance (NMR) (Scherer and Seelig, 1987;Seelig et al, 1987), neutron and x-ray diffraction (Buldt et al, 1979;Pearson and Pascher, 1979;Hauser et al, 1981;Weiner and White, 1992), and Raman scattering (Akutsu and Nagamori, 1991) measurements. For example, the polar headgroup of our model PC has a -15°angle of inclination to the bilayer surface, compared with the estimate of 20°by Weiner and White (1992).…”

Thermodynamics of the Coil ⇆ β-Sheet Transition in a Membrane Environment

“…This is smaller than that observed in the absence of POPG, which is ν Q ∼ 5.9 kHz and can be explained by the influence of the negative POPG charge on the − P-N + dipole. 47 As only a single quadrupole splitting is observed, this requires a homogeneous distribution of anionic POPG molecules in the electrically neutral POPC matrix.…”