Area Per Lipid of Membranes from Natural Abundance Solid-State 13C NMR Spectroscopy

Abstract

The properties of liquid-crystalline membranes vary according to the molecular composition of the lipid bilayer [1]. Structural investigations of lipid membranes using NMR spectroscopy generally require isotopic labeling of the lipids, thereby precluding investigations of complex lipid systems. Combining natural-abundance solid-state 13C magic-angle-spinning (MAS) NMR with a statistical mean torque (MT) model [2] enabled us to obtain the average area per lipid and volumetric bilayer thickness DB for lipid bilayers. The 2D separated local field (SLF) NMR experiment DROSS gives isotropic chemical shifts in the F2 frequency dimension and site-specific dipolar Pake doublets in the F1 dimension [1]. By analyzing the Pake patterns we extracted 13C-1H residual dipolar couplings (RDCs) and calculated segmental |SCH| order parameters. The |SCH| order parameters carry the structural information that aid calculation of and DB using the MT model. Experiments were conducted with DMPC, POPC, and EYSM lipids as single component (liquid-disordered, ld) lipid bilayers as well as with cholesterol. EYSM is more ordered in the ld phase and hence experiences less structural perturbation upon adding cholesterol to form the liquid-ordered (lo) phase relative to POPC. We infer that the cholesterol stiffening effect on lipid bilayers is limited by the maximum volumetric hydrocarbon thickness [2]. Knowledge of structural parameters like and DB is important for molecular dynamics (MD) simulations and provides information about the balance of forces in membrane lipid bilayers [3]. Our studies demonstrate the applicability of solid-state 13C NMR spectroscopy to site-specific structural investigations of complex lipids, where isotope labeling may be prohibitive, allowing structural studies of biologically relevant membrane systems. [1] A. Leftin et al.(2013) JMB425 2973-2987. [2] H. I. Petrache et al.(2000) BJ.79 3172-3192. [3] A. Leftin et al.(2011) BBA1808 818-839.