Department of Chemistry, University of Illinois at Urbana-Champaign , 600 South Mathews Avenue, Urbana, Illinois 61801, United States.
Acc Chem Res. 2013 Sep 17;46(9):2080-8. doi: 10.1021/ar4000168. Epub 2013 May 10.
Solid-state NMR (SSNMR) spectroscopy has become an important technique for studying the biophysics and structure biology of proteins. This technique is especially useful for insoluble membrane proteins and amyloid fibrils, which are essential for biological functions and are associated with human diseases. In the past few years, as major contributors to the rapidly advancing discipline of biological SSNMR, we have developed a family of methods for high-resolution structure determination of microcrystalline, fibrous, and membrane proteins. Key developments include order-of-magnitude improvements in sensitivity, resolution, instrument stability, and sample longevity under data collection conditions. These technical advances now enable us to apply new types of 3D and 4D experiments to collect atomic-resolution structural restraints in a site-resolved manner, such as vector angles, chemical shift tensors, and internuclear distances, throughout large proteins. In this Account, we present the technological advances in SSNMR approaches towards protein structure determination. We also describe the application of those methods for large membrane proteins and amyloid fibrils. Particularly, the SSNMR measurements of an integral membrane protein DsbB support the formation of a charge-transfer complex between DsbB and ubiquinone during the disulfide bond transfer pathways. The high-resolution structure of the DsbA-DsbB complex demonstrates that the joint calculation of X-ray and SSNMR restraints for membrane proteins with low-resolution crystal structure is generally applicable. The SSNMR investigations of α-synuclein fibrils from both wild type and familial mutants reveal that the structured regions of α-synuclein fibrils include the early-onset Parkinson's disease mutation sites. These results pave the way to understanding the mechanism of fibrillation in Parkinson's disease.
固态核磁共振（SSNMR）光谱学已成为研究蛋白质生物物理学和结构生物学的重要技术。该技术对于不溶性膜蛋白和淀粉样纤维特别有用，因为它们对于生物功能至关重要，并且与人类疾病有关。在过去的几年中，作为生物 SSNMR 快速发展领域的主要贡献者，我们开发了一系列用于微晶体、纤维和膜蛋白高分辨率结构测定的方法。关键的发展包括在数据收集条件下灵敏度、分辨率、仪器稳定性和样品寿命的数量级提高。这些技术进步现在使我们能够应用新型的 3D 和 4D 实验以站点分辨的方式收集原子分辨率的结构约束，例如矢量角度、化学位移张量和核间距，贯穿整个大蛋白。在本报告中，我们介绍了 SSNMR 方法在蛋白质结构测定方面的技术进步。我们还描述了这些方法在大型膜蛋白和淀粉样纤维中的应用。特别是，对整合膜蛋白 DsbB 的 SSNMR 测量支持了 DsbB 和泛醌在二硫键转移途径中形成电荷转移复合物的形成。DsbA-DsbB 复合物的高分辨率结构表明，对于具有低分辨率晶体结构的膜蛋白，联合计算 X 射线和 SSNMR 约束通常是适用的。对来自野生型和家族突变体的α-突触核蛋白纤维的 SSNMR 研究表明，α-突触核蛋白纤维的结构区域包括早发性帕金森病突变位点。这些结果为理解帕金森病中的纤维形成机制铺平了道路。
