Glück Julian M, Wittlich Marc, Feuerstein Sophie, Hoffmann Silke, Willbold Dieter, Koenig Bernd W
Institute of Structural Biology and Biophysics (ISB-3), Research Centre Jülich, D-52425 Jülich, Germany.
J Am Chem Soc. 2009 Sep 2;131(34):12060-1. doi: 10.1021/ja904897p.
We present a two-dimensional solution NMR spectrum of an integral membrane protein (IMP) in a nanodisc. Solution NMR relies on rapid isotropic tumbling of the analyte with correlation times in the nanosecond range. IMPs in a cellular membrane do not satisfy this condition. Previous liquid-state NMR studies on IMPs were conducted in organic solvent or artificial membrane mimicking particles like detergent micelles. Nanodiscs are relatively small (150 kDa), detergent-free model membranes that are suitable for functional reconstitution of IMPs. Nanodiscs allow solubilization of integral membrane proteins in a nearly native lipid bilayer environment. The 70 residue polypeptide CD4mut was incorporated into nanodiscs. CD4mut features one transmembrane helix. The aliphatic (1)H-(13)C HSQC spectrum of nanodiscs with inserted, ((13)C, (15)N)-labeled CD4mut exhibits reasonably dispersed protein and lipid NMR signals. Our results demonstrate that IMPs in nanodiscs are amenable to liquid-state NMR methodology.
我们展示了纳米盘中一种整合膜蛋白（IMP）的二维溶液核磁共振谱。溶液核磁共振依赖于分析物在纳秒范围内的快速各向同性翻滚。细胞膜中的整合膜蛋白不满足此条件。先前对整合膜蛋白的液态核磁共振研究是在有机溶剂或模拟颗粒（如去污剂胶束）的人工膜中进行的。纳米盘相对较小（150 kDa），是不含去污剂的模型膜，适用于整合膜蛋白的功能重构。纳米盘能使整合膜蛋白在几乎天然的脂质双层环境中溶解。70个残基的多肽CD4mut被整合到纳米盘中。CD4mut具有一个跨膜螺旋。插入了（（13）C，（15）N）标记的CD4mut的纳米盘的脂肪族（1）H-（13）C HSQC谱显示出合理分散的蛋白质和脂质核磁共振信号。我们的结果表明，纳米盘中的整合膜蛋白适用于液态核磁共振方法。
