Optimal Bicelle Size q for Solution NMR Studies of the Protein Transmembrane Partition.
Optimal Bicelle Size q for Solution NMR Studies of the Protein Transmembrane Partition.
复制标题
用于蛋白跨膜分区的溶液NMR研究的最佳bicelle尺寸Q。
DOI：
10.1002/chem.201604206
复制
发表时间：
2017-01-26
期刊：
影响因子：
--
通讯作者：
Chou JJ
中科院分区：
文献类型：
--
作者：
Piai A;Fu Q;Dev J;Chou JJ
各向同性双胞中跨膜蛋白的结构表征已经成为溶液NMR光谱学的越来越受欢迎的应用，因为快速翻滚的双胞是膜状的，但通常可以产生与洗涤剂胶束相当的光谱质量。虽然较大的双胞更接近真正的脂质双层，但仍不清楚双胞需要多大才能准确评估脂质双层中的蛋白质跨膜分配。在这里，我们解决了上述问题，从蛋白质的角度驻留在bicelles，通过系统测量蛋白质的化学位移和跨膜分配在不同的脂质：去污剂的比例（q），从0.3到0.7，使用跨膜结构域的人Fas受体作为模型系统。我们发现，脂质环境的bicelles，所反映的蛋白质化学位移，开始被扰动时，q降低到0.6以下。我们还实施了溶剂顺磁弛豫增强（PRE）的方法，表明在bicelles与q = 0.5和0.7的蛋白质跨膜分配是非常相似的，但在q = 0.3的溶剂PRE的配置文件是显着不同的。我们的数据表明，q值在0.5和0.6之间是高分辨率NMR和接近膜环境之间的良好折衷，并允许准确表征脂质双层中的蛋白质位置。描述了一种稳健的方法来评估最佳的双胞尺寸，以准确地确定寡聚跨膜蛋白结构域在脂质双层中的位置。该方法将溶剂顺磁滴定和顺磁弛豫数据的S形函数拟合相结合。我们的研究结果表明，具有脂质/去污剂比（q）在0.5和0.6之间的bicelles确保了几乎理想的脂质双层环境，同时提供了与高分辨率NMR光谱兼容的有利的弛豫特性。
Structural characterization of transmembrane proteins in isotropic bicelles has become an increasingly popular application of solution NMR spectroscopy, as the fast-tumbling bicelles are membrane-like yet can often yield spectral quality comparable to those of detergent micelles. While larger bicelles are closer to the true lipid bilayer, it remains unclear how large the bicelles need to be to allow accurate assessment of protein transmembrane partition in lipid bilayer. Here, we address the above question from the perspective of protein residing in the bicelles, through systematic measurement of protein chemical shift and transmembrane partition at different lipid:detergent ratios (q), ranging from 0.3 to 0.7, using the transmembrane domain of human Fas receptor as model system. We found that the lipid environment of the bicelles, as reflected by the protein chemical shift, begins to be perturbed when the q is reduced to below 0.6. We also implemented a solvent paramagnetic relaxation enhancement (PRE) approach for bicelles to show that the protein transmembrane partition in bicelles with q = 0.5 and 0.7 are very similar, but at q = 0.3 the solvent PRE profile is significantly different. Our data indicate that q values between 0.5 and 0.6 are good compromise between high resolution NMR and closeness to the membrane environment, and allow accurate characterization of protein position in lipid bilayer. A robust method is described to assess the optimal bicelle size(s) to accurately determine the position of oligomeric transmembrane protein domains in lipid bilayer. This method combines solvent paramagnetic agent titration and paramagnetic relaxation data fitting with sigmoid function. Our results show that bicelles with lipid/detergent ratio (q) between 0.5 and 0.6 ensure an almost-ideal lipid bilayer environment, while at the same time providing favorable relaxation properties that are compatible with high resolution NMR spectroscopy.