Cholesterol enhances surface water diffusion of phospholipid bilayers

Abstract

Elucidating the physical effect of cholesterol (Chol) on biological membranes is necessary towards rationalizing their structural and functional role in cell membranes. One of the debated questions is the role of hydration water in Chol-embedding lipid membranes, for which only little direct experimental data are available. Here, we study the hydration dynamics in a series of Chol-rich and depleted bilayer systems using an approach termed 1 H Overhauser dynamic nuclear polarization (ODNP) NMR relaxometry that enables the sensitive and selective determination of water diffusion within 5-10 Å of a nitroxide-based spin label, positioned off the surface of the polar headgroups or within the nonpolar core of lipid membranes. The Chol-rich membrane systems were prepared from mixtures of Chol, dipalmitoyl phosphatidylcholine and/or dioctadecyl phosphatidylcholine lipid that are known to form liquid-ordered, raft-like, domains. Our data reveal that the translational diffusion of local water on the surface and within the hydrocarbon volume of the bilayer is significantly altered, but in opposite directions: accelerated on the membrane surface and dramatically slowed in the bilayer interior with increasing Chol content. Electron paramagnetic resonance (EPR) lineshape analysis shows looser packing of lipid headgroups and concurrently tighter packing in the bilayer core with increasing Chol content, with the effects peaking at lipid compositions reported to form lipid rafts. The complementary capability of ODNP and EPR to site-specifically probe the hydration dynamics and lipid ordering in lipid membrane systems extends the current understanding of how Chol may regulate biological processes. One possible role of Chol is the facilitation of interactions between biological constituents and the lipid membrane through the weakening or disruption of strong hydrogen-bond networks of the surface hydration layers that otherwise exert stronger repulsive forces, as reflected in faster surface water diffusivity. Another is the concurrent tightening of lipid packing that reduces passive, possibly unwanted, diffusion of ions and water across the bilayer.

Recent Experimental Developments in Studying Passive Membrane Transport of Drug Molecules

“…109,293 In another approach, spin polarization from ODNP-enhanced 1 H actively transferred to 13 C nuclei through scalar (J) coupling 294 successfully monitored the permeability of glycine across negatively charged liposomal bilayers composed of DPPC and DPPG and the capacity of cholesterol to reduce the lipid headgroup packing and increase acyl chains ordering. 294,295 With the long acquisition time required, NMR is not suitable for quantitative kinetic studies. 225 Radiolabeling ( 3 H, 14 C, etc.)…”

“…1 H NMR line-broadening method was first developed by Alger and Prestegard to study acetic acid permeation of LUV membranes . When combined with the dynamic light scattering method, permeability coefficients for seven short-chain monocarboxylic acids across DPPC bilayers were obtained. , In another approach, spin polarization from ODNP-enhanced 1 H actively transferred to 13 C nuclei through scalar ( J ) coupling successfully monitored the permeability of glycine across negatively charged liposomal bilayers composed of DPPC and DPPG and the capacity of cholesterol to reduce the lipid headgroup packing and increase acyl chains ordering. , With the long acquisition time required, NMR is not suitable for quantitative kinetic studies …”

“…Cholesterol content of biological membranes has functional roles in biological events, such as the enzymatic activity of the membrane-bound enzyme, Na + /K + -ATPase, which is enhanced under cholesterol-rich conditions at lipid raft domains. , Using the 1 H Overhauser dynamic nuclear polarization (ODNP) NMR relaxometry approach, Cheng et al reported that at higher cholesterol content, bound water populations significantly increase at the lipid–water interface and hydrophobic core, concurrent with loose packing of lipid headgroups and dramatically increased ordering of the lipid acyl chains. MD simulations have shown that cholesterol concentration up to 20 mol % has an inhibitory effect on the permeation of water across mixed DPPC/cholesterol bilayers.…”

Cholesterol-induced suppression of membrane elastic fluctuations at the atomistic level

“…Various biophysical methods have been used to study lipid-cholesterol interactions, including electron spin resonance (ESR) (Cheng et al, 2014; Delmelle et al, 1980; Hubbell and McConnell, 1971; Lai and Freed, 2014; Manukovsky et al, 2013; Semer and Gelerinter, 1979; Stepien et al, 2015; Vitiello et al, 2015; Williams et al, 2013), Raman (Lippert and Peticolas, 1971; Mendelsohn, 1972; Tantipolphan et al, 2006), Fourier transform infrared (FT-IR) (Umemura et al, 1980), fluorescence spectroscopy (Xu and London, 2000; Yasuda et al, 2015a), atomic force microscopy (AFM), multidimensional NMR spectroscopy (Holland and Alam, 2006; Leftin et al, 2013; Leftin et al, 2014a; Warschawski and Devaux, 2005), solid-state 2 H nuclear magnetic resonance (NMR) (Bartels et al, 2008; Brown, 1990; Bunge et al, 2008; Martinez et al, 2004; Martinez et al, 2002; Matsumori et al, 2012; Stockton et al, 1976; Vogel et al, 2016; Weisz et al, 1992; Yasuda et al, 2015a), and neutron diffraction methods (Armstrong et al, 2014; Toppozini et al, 2014). However, a thorough understanding of the physical basis for these observations in relation to the intricate lipid compositions of many biological membranes to some extent remains an enigma (Feigenson, 2015; McConnell, 2005; Meinhardt et al, 2013; Sodt et al, 2014; Stanich et al, 2013).…”

Is the Membrane Lipid Matrix a Key Target for Action of Pharmacologically Active Plant Saponins?

“…The introduction of CHOL into membrane composition was accompanied by a 25–50% decrease in the dipole-modifying ability of digitonin, tribulosin, dioscin, and escin ( Figure 2 b, Table 1 ). This may be due to the higher rigidity of CHOL-enriched bilayers or a difference in hydration water dynamics in CHOL-free and CHOL-enriched lipid membranes [ 24 ].…”