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首页> 外文期刊>Biochemistry >Lipid-Protein Nanodiscs: Possible Application in High-Resolution NMR Investigations of Membrane Proteins and Membrane-Active Peptides
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Lipid-Protein Nanodiscs: Possible Application in High-Resolution NMR Investigations of Membrane Proteins and Membrane-Active Peptides

机译：脂质蛋白纳米光盘：在膜蛋白和膜活性肽的高分辨率NMR研究中的可能应用

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摘要

High-resolution NMR is shown to be applicable for investigation of membrane proteins and membrane-active peptides embedded into lipid-protein nanodiscs (LPNs). ~(15)N-Labeled K~+-channel from Streptomyces lividans (KcsA) and the antibiotic antiamoebin I from Emericellopsis minima (Aam-I) were embedded in LPNs of different lipid composition. Formation of stable complexes undergoing isotropic motion in solution was confirmed by size-exclusion chromatography and ~(31)P-NMR spectroscopy. The 2D ~1H-~(15)N-correlation spectra were recorded for KcsA in the complex with LPN containing DMPC and for Aam-I in LPNs based on DOPG, DLPC, DMPC, and POPC. The spectra recorded were compared with those in detergent-containing micelles and small bicelles commonly used in high-resolution NMR spectroscopy of membrane proteins. The spectra recorded in LPN environments demonstrated similar signal dispersion but significantly increased ~1H_N line width. The spectra of Aam-I embedded in LPNs containing phosphatidylcholine showed significant selective line broadening, thus suggesting exchange process(es) between several membrane-bound states of the peptide. ~(15)N relaxation rates were measured to obtain the effective rotational correlation time of the Aam-I molecule. The obtained value (~40 nsec at 45°C) is indicative of additional peptide motions within the Aam-I/LPN complex.
机译：高分辨率NMR已显示可用于研究嵌入脂蛋白纳米光盘（LPN）的膜蛋白和膜活性肽。将来自链霉菌（Streptomyces lividans）（KcsA）的〜（15）N标签的K〜+通道和来自最小拟南芥（Aam-1）的抗生素抗变形蛋白I嵌入不同脂质组成的LPN中。通过尺寸排阻色谱法和〜（31）P-NMR光谱确认了在溶液中经历各向同性运动的稳定络合物的形成。基于DOPG，DLPC，DMPC和POPC，记录了LCS含有LPN的配合物中KcsA的2D〜1H-〜（15）N相关光谱。将记录的光谱与通常在膜蛋白的高分辨率NMR光谱中使用的含洗涤剂的胶束和小单体的光谱进行比较。在LPN环境中记录的光谱显示出相似的信号色散，但显着增加了〜1H_N的线宽。嵌入包含磷脂酰胆碱的LPN中的Aam-1光谱显示出显着的选择性谱线加宽，因此表明该肽在几种膜结合状态之间的交换过程。测量〜（15）N的弛豫速率以获得Aam-1分子的有效旋转相关时间。所获得的值（在45°C时约为40纳秒）表明Aam-I / LPN复合物中的其他肽运动。

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