Spectroscopic studies of specifically deuterium labeled membrane systems. Nuclear magnetic resonance investigation of the effects of cholesterol in model systems

Abstract

Deuterium nuclear magnetic resonance spectra of dimyristoylphosphatidylcholines specifically labeled in positions 2', 3', 4', 6', 8', 10', 12', and 14', of the 2 chain, of an N-deuteriomethylphosphatidylcholine, and of cholesterol-3alpha-d1, have been obtained by the Fourier transform method at 5.46 and 3.52 T on two "home-built" widebore superconducting magnet spectrometers, as a function of temperature and composition. Data on the specifically deuterium-labeled cholesterol molecule (in nonsonicated membrane systems) permits an estimate of the most probable angle of tilt of the sterol in the membrane, and evaluation of the order parameter (Salpha) describing rigid body motions in the bilayer. Segmental order parameters derived from the data presented allow calculation of individual chain segment projections onto the director axis and, consequently, estimation of effective chain length. It is shown that mathematical models which include chain tilt as well as those which neglect this type of rigid body motion give essentially identical results when applied to the dimyristoylphosphatidylcholine and dimyristoylphosphatidylcholine-cholesterol bilayer systems (in excess water, between 23 and 60 degrees C). Results of calculations of chain length and membrane thickness of a dimyristoylphosphatidylcholine-30 mol % cholesterol membrane system at 23 degrees C give excellent agreement when compared with recent high-resolution neutron diffraction data obtained on specifically deuterium labeled lecithin-cholesterol systems. No evidence for formation of lecithin-cholesterol complexes having lifetimes of approximately 30 ms has been found. Below the pure-lipid gel-liquid crystal phase transition temperature Tc but in the presence of cholesterol, we have obtained further evidence for 1-chain/2-chain nonequivalence. At 10 degrees C, the 2' segment of the 2 chain, but not the 2' segment of the 1 chain or the 3', 6', or 12' segments of the 2 chain, is broadened almost beyond detection. These results are in agreement with similar effects reported recently for the dipalmitoylphosphatidylcholine-cholesterol system and may indicate a bent configuration for the 2 chain, in the lecithin-cholesterol system. Further cooling below Tc results in loss of the 1-chain 2'-position signal intensity plus 2-chain 3', 6', and 12' signals simultaneously. The increase in length of the 2 chain of dimyristoylphosphatidylcholine upon addition of 30 mol % cholesterol of 23 degrees C is about 2.3 A. Addition of cholesterol to a choline-labeled lecithin results in complex behavior of the head group deuterium quadrupole splitting as a function of temperature, and cholesterol mole fraction. Above approximately 20 mol % cholesterol, the main effect is a decrease in quadrupole splitting as cholesterol content increases, the opposite effect to that observed with hydrocarbon chains.

Regulation of membrane protein function by lipid bilayer elasticity—a single molecule technology to measure the bilayer properties experienced by an embedded protein

“…Micelle-forming amphiphiles decrease the acyl chain order of lipid bilayers and increase the fluorescent polarization of bilayer-embedded diphenylhexatriene, which has been interpreted to signify that the bilayer fluidity is increased (e.g., [152,153]). Capsaicin has similar effects [154], while cholesterol decreases the bilayer fluidity [155,156]. Nevertheless, the changes in gA channel behaviour cannot be explained in terms of altered fluidity.…”

General features of phospholipid conformation in membranes

“…The method has been applied mainly to bilayers containing the phosphocholine head group, i.e. 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) [3,4], 1 -palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) [ 5,6], DPPC plus cholesterol [7], 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) [8] and egg-yolk lecithin [9]. In addition, a deuterium order profile has been published for Acholeplasma laidlawii membranes biosynthetically enriched with deuterated palmitate [lo].…”

Mechanism of Lipid Bilayer Disruption by the Human Antimicrobial Peptide, LL-37

“…In addition, the interaction of LL-37 with lipids containing choline headgroups is of interest because of the possible role of phosphorylcholine in bacterial resistance to LL-37 (24), although the presence of phosphatidylcholine headgroups on phospholipids does not prevent membrane disruption by LL-37. DMPC has also been widely used in other studies of peptides that interact with lipid bilayers, enabling comparison of the results of this work with studies of other antimicrobial peptides and membrane-perturbing molecules (31)(32)(33)(34)(35)(36)(37)(38)(39), thus allowing insight into the mechanism of membrane disruption and the similarities and differences exhibited by the various cationic, amphipathic, R-helical peptides. It also provides a good baseline for comparison when the bilayer composition is varied.…”