Leftin Avigdor, Molugu Trivikram R, Job Constantin, Beyer Klaus, Brown Michael F
Department of Chemistry and Biochemistry, University of Arizona, Tucson, Arizona.
Department of Chemistry and Biochemistry, University of Arizona, Tucson, Arizona; Department of Physics, University of Arizona, Tucson, Arizona.
Biophys J. 2014 Nov 18;107(10):2274-86. doi: 10.1016/j.bpj.2014.07.044.
Investigations of lipid membranes using NMR spectroscopy generally require isotopic labeling, often precluding structural studies of complex lipid systems. Solid-state (13)C magic-angle spinning NMR spectroscopy at natural isotopic abundance gives site-specific structural information that can aid in the characterization of complex biomembranes. Using the separated local-field experiment DROSS, we resolved (13)C-(1)H residual dipolar couplings that were interpreted with a statistical mean-torque model. Liquid-disordered and liquid-ordered phases were characterized according to membrane thickness and average cross-sectional area per lipid. Knowledge of such structural parameters is vital for molecular dynamics simulations, and provides information about the balance of forces in membrane lipid bilayers. Experiments were conducted with both phosphatidylcholine (dimyristoylphosphatidylcholine (DMPC) and palmitoyloleoylphosphatidylcholine (POPC)) and egg-yolk sphingomyelin (EYSM) lipids, and allowed us to extract segmental order parameters from the (13)C-(1)H residual dipolar couplings. Order parameters were used to calculate membrane structural quantities, including the area per lipid and bilayer thickness. Relative to POPC, EYSM is more ordered in the ld phase and experiences less structural perturbation upon adding 50% cholesterol to form the lo phase. The loss of configurational entropy is smaller for EYSM than for POPC, thus favoring its interaction with cholesterol in raftlike lipid systems. Our studies show that solid-state (13)C NMR spectroscopy is applicable to investigations of complex lipids and makes it possible to obtain structural parameters for biomembrane systems where isotope labeling may be prohibitive.
使用核磁共振光谱法对脂质膜进行研究通常需要同位素标记，这常常排除了对复杂脂质系统的结构研究。在天然同位素丰度下进行的固态（13）C魔角旋转核磁共振光谱法可提供位点特异性结构信息，有助于对复杂生物膜进行表征。利用分离局部场实验DROSS，我们解析了（13）C-（1）H残余偶极耦合，并通过统计平均扭矩模型对其进行了解释。根据膜厚度和每个脂质的平均横截面积对液相无序和液相有序相进行了表征。了解这些结构参数对于分子动力学模拟至关重要，并提供了有关膜脂质双层中力平衡的信息。我们用磷脂酰胆碱（二肉豆蔻酰磷脂酰胆碱（DMPC）和棕榈酰油酰磷脂酰胆碱（POPC））以及蛋黄鞘磷脂（EYSM）脂质进行了实验，并使我们能够从（13）C-（1）H残余偶极耦合中提取片段序参数。序参数用于计算膜结构量，包括每个脂质的面积和双层厚度。相对于POPC，EYSM在液相无序相（ld相）中更有序，并且在添加50%胆固醇形成液相有序相（lo相）时经历的结构扰动更小。EYSM的构型熵损失比POPC小，因此有利于其在类筏脂质系统中与胆固醇的相互作用。我们的研究表明，固态（13）C核磁共振光谱法适用于复杂脂质的研究，并有可能获得生物膜系统的结构参数，而在这些系统中同位素标记可能是不可行的。
