NMR INVESTIGATIONS OF CELL MEMBRANE STRUCTURE

细胞膜结构的核磁共振研究

• 批准号: 2565412
• 负责人: KLAUS GAWRISCH
• 金额: --
• 依托单位: NATIONAL INSTITUTE ON ALCOHOL ABUSE AND ALCOHOLISM
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/2565412
• 关键词: alcoholism /alcohol abuse; cell membrane; ethanol; lipid bilayer membrane; lipid structure; membrane lipids; membrane proteins; membrane structure; nuclear magnetic resonance spectroscopy; physical chemical interaction; protein structure; unsaturated fatty acids

The objectives of this project are to: (1) investigate the interaction of alcohol with proteins and lipids in biological membranes; (2) study structure and dynamics of membranes composed of lipids with polyunsaturated fatty acids such as docosahexaenoic acid (DHA, 22:6n-3); and (3) study lipid-protein interactions related to alcoholism and lipid polyunsaturation. (1) The location of ethanol within a membrane has been investigated directly with two-dimensional NOESY and magic-angle-spinning NMR. From NOESY crosspeak intensities we determined that ethanol resides with highest probability at the lipid-water interface near the lipid glycerol and the upper methylene segments of lipid hydrocarbon chains. Ethanol concentration in the bilayer hydrophobic core is significantly lower. (2) (2)H NMR spectroscopy was used to measure order in PCs with a perdeuterated stearoyl sn-1 chain and four unsaturated sn-2 chains - 18:1n-9, 18:2n-6, 20:4n-6, and 22:6n-3 - combined with PE of identical fatty acid composition and cholesterol. The polyunsaturated bilayers are distinct from the mono- and diunsaturated bilayers in that they had the lowest order at all cholesterol concentrations and were the least ordered by PE. Polyunsaturated membranes show less ethanol-induced chain disorder than mono- and diunsaturated membranes. A modified (17)O NMR method has been developed for measurement of fast water permeation through lipid bilayers. The lower hydrocarbon chain order in polyunsaturated lipids appears to correlate with higher water permeation rates through polyunsaturated membranes. Addition of ethanol reduces water permeation, most likely by restricting entry of water molecules into the membrane. (3) The peptide fragment 828-848 with the sequence RVIEVVQGACRAIRHIPRRIR from the carboxy terminal region of the envelope glycoprotein gp41 of HIV-1 deeply incorporates into negatively charged DMPC bilayers as an amphipathic alpha-helix. Measurements on peptide with specifically deuterated isoleucine amino acids demonstrate that three of the four isoleucine residues are deeply imbedded in the hydrocarbon core of the membrane. The positively charged carboxy terminus of the peptide is located at the bilayer surface and shows a higher degree of flexibility.

该项目的目标是：（1）调查交互 生物膜中的蛋白质和脂质的酒精； （2）研究 由脂质组成的膜的结构和动力学 多不饱和脂肪酸，例如二十六烯酸（DHA，22：6n-3）； （3）研究与酒精中毒和脂质有关的脂质 - 蛋白质相互作用 多不饱和。 （1）乙醇在膜中的位置已在 直接用二维贵族和魔术角旋转进行调查 NMR。 从Noesy Crosspeak强度中，我们确定乙醇属于 在脂质附近的脂质水接口处的概率最高 甘油和脂质烃链的上甲基段。 双层疏水核中的乙醇浓度显着 降低。 （2）（2）H NMR光谱法用于测量具有 一个perderate的硬质Sn-1链和四个不饱和SN-2链 - 18：1n-9，18：2n-6，20：4n-6和22：6n-3-与同一PE结合 脂肪酸成分和胆固醇。 多不饱和双层是 与单一和diunsated双层不同，因为他们有 在所有胆固醇浓度下的最低顺序，是有序最少的 由PE。 多不饱和膜显示乙醇诱导的链较少 疾病比单饱和膜和偶性膜。 修改后的（17）o nmr 已经开发了用于测量快速水渗透的方法 通过脂质双层。 较低的碳氢化合物链顺序 多不饱和脂质似乎与较高的水渗透相关 通过多不饱和膜的速率。 添加乙醇减少 水渗透，最有可能限制水分子的进入 进入膜。 （3）带有序列的肽片段828-848 来自信封的羧基终端区域的RVIEVVQGACRAIRHIPRIR HIV-1的糖蛋白GP41深入纳入负电荷 DMPC双层作为两亲α-螺旋。 对肽的测量 特定的氘化异亮氨酸氨基酸表明， 四个异亮氨酸残基中的三个被深层嵌入 膜的烃核。 带正电荷的羧基 肽的末端位于双层表面，显示 较高的灵活性。