Laboratory of Physical Chemistry, Swiss Federal Institute of Technology, ETH-Hönggerberg, Zürich, Switzerland.
PLoS One. 2013;8(1):e54378. doi: 10.1371/journal.pone.0054378. Epub 2013 Jan 22.
Because membrane proteins need to be extracted from their natural environment and reconstituted in artificial milieus for the 3D structure determination by X-ray crystallography or NMR, the search for membrane mimetic that conserve the native structure and functional activities remains challenging. We demonstrate here a detergent/nanodisc screening study by NMR of the bacterial α-helical membrane protein YgaP containing a cytoplasmic rhodanese domain. The analysis of 2D [(15)N,(1)H]-TROSY spectra shows that only a careful usage of low amounts of mixed detergents did not perturb the cytoplasmic domain while solubilizing in parallel the transmembrane segments with good spectral quality. In contrast, the incorporation of YgaP into nanodiscs appeared to be straightforward and yielded a surprisingly high quality [(15)N,(1)H]-TROSY spectrum opening an avenue for the structural studies of a helical membrane protein in a bilayer system by solution state NMR.
由于膜蛋白需要从其自然环境中提取出来，并在人工环境中重新构建，以便通过 X 射线晶体学或 NMR 确定其 3D 结构，因此寻找能够保持天然结构和功能活性的膜模拟物仍然具有挑战性。我们在这里通过 NMR 对含有细胞质硫氧还蛋白结构域的细菌α-螺旋膜蛋白 YgaP 进行了去污剂/纳米盘筛选研究。二维 [(15)N,(1)H]-TROSY 谱的分析表明，只有小心使用低浓度的混合去污剂，才能在平行溶解跨膜片段的同时，不干扰细胞质结构域，并且获得具有良好谱质量的跨膜片段。相比之下，将 YgaP 掺入纳米盘中似乎很简单，并且出乎意料地获得了高质量的 [(15)N,(1)H]-TROSY 谱，为通过溶液状态 NMR 在双层系统中研究螺旋膜蛋白的结构开辟了一条途径。
