Optimization of the design and preparation of nanoscale phospholipid bilayers for its application to solution NMR
Optimization of the design and preparation of nanoscale phospholipid bilayers for its application to solution NMR
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DOI：
10.1002/prot.24271
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发表时间：
2013-07-01
影响因子：
2.9
通讯作者：
Vinogradova, Olga
中科院分区：
文献类型：
--
作者：
Puthenveetil, Robbins;Vinogradova, Olga
尽管有艰苦的努力和最近的技术发展，结构的调查，整合膜蛋白仍然是一个挑战。主要的阻遏物包括它们的表达困难、固有的低溶解度以及与现有的膜模拟系统相关的问题。一类相对较新的膜模拟物，纳米盘，正在成为一个有前途的替代品。虽然纳米盘已被证明是成功的几个生物物理应用，他们仍然成为系统的结构确定的首选。因此，我们通过将自组装复合物的直径减小到其潜在的极限，使纳米盘更适合于溶液NMR应用。我们取得了显着的改善，在质量的NMR光谱的整合素IIb的跨膜和胞质结构域纳入这些较小的光盘，使他们容易进行彻底的结构调查。此外，我们还提出了一种快速，高效，单步制备高浓度蛋白质掺入纳米盘的柱上方法。这些光盘已充分其特征在于透射电子显微镜，动态光散射，差示扫描量热法。Proteins 2013; 81：1222-1231. (c)2013 Wiley Periodicals，Inc.
Despite arduous efforts and recent technological developments structural investigation of integral membrane proteins remains a challenge. The primary deterrents include difficulties with their expression, low inherent solubility, and problems associated with existing membrane mimicking systems. A relatively new class of membrane mimetics, nanodiscs, is emerging as a promising alternative. Although nanodiscs have been proven successful for several biophysical applications, they yet remain to become the system of preferred choice for structure determination. We have hereby made nanodiscs more suitable for solution NMR applications by reducing the diameter of the self-assembly complex to its potential limit. We achieved a noticeable improvement in the quality of NMR spectra obtained for the transmembrane and cytoplasmic domains of integrin IIb incorporated into these smaller discs rendering them susceptible for a thorough structural investigation. In addition, we also present an on-column method for a rapid, efficient, single-step preparation of protein incorporated nanodiscs at high concentrations. These discs have been fully characterized by transmission electron microscopy, dynamic light scattering, and differential scanning calorimetry. Proteins 2013; 81:1222-1231. (c) 2013 Wiley Periodicals, Inc.