Department of Chemistry, University of Illinois at Urbana-Champaign, 600 South Mathews Avenue, Urbana, IL 61801, USA.
J Biomol NMR. 2010 Feb;46(2):149-55. doi: 10.1007/s10858-009-9389-9. Epub 2009 Dec 2.
Magic-angle spinning (MAS) solid-state NMR (SSNMR) spectroscopy of uniformly-(13)C,(15)N labeled protein samples provides insight into atomic-resolution chemistry and structure. Data collection efficiency has advanced remarkably in the last decade; however, the study of larger proteins is still challenged by relatively low resolution in comparison to solution NMR. In this study, we present a systematic analysis of SSNMR protein spectra acquired at 11.7, 17.6 and 21.1 Tesla ((1)H frequencies of 500, 750, and 900 MHz). For two protein systems--GB1, a 6 kDa nanocrystalline protein and DsbA, a 21 kDa nanocrystalline protein--line narrowing is demonstrated in all spectral regions with increasing field. Resolution enhancement is greatest in the aliphatic region, including methine, methylene and methyl sites. The resolution for GB1 increases markedly as a function of field, and for DsbA, resolution in the C-C region increases by 42%, according to the number of peaks that can be uniquely picked and integrated in the 900 MHz spectra when compared to the 500 MHz spectra. Additionally, chemical exchange is uniquely observed in the highest field spectra for at least two isoleucine C delta 1 sites in DsbA. These results further illustrate the benefits of high-field MAS SSNMR spectroscopy for protein structural studies.
均匀标记的（13）C，（15）N 蛋白样品的魔角旋转（MAS）固态 NMR（SSNMR）光谱提供了原子分辨率化学和结构的深入了解。在过去的十年中，数据采集效率有了显著提高；然而，与溶液 NMR 相比，较大蛋白质的研究仍然受到相对较低分辨率的挑战。在这项研究中，我们对在 11.7、17.6 和 21.1 特斯拉（（1）H 频率为 500、750 和 900 MHz）下获得的 SSNMR 蛋白光谱进行了系统分析。对于两个蛋白质体系 - GB1，一种 6 kDa 的纳米晶体蛋白和 DsbA，一种 21 kDa 的纳米晶体蛋白 - 在所有光谱区域中都证明了随着场的增加线宽变窄。分辨率增强在脂肪族区域最大，包括亚甲基、次甲基和甲基位点。随着场的增加，GB1 的分辨率显著增加，对于 DsbA，根据在 900 MHz 光谱中可以唯一选择和积分的峰的数量，C-C 区域的分辨率增加了 42%，与 500 MHz 光谱相比。此外，在 DsbA 中，至少有两个异亮氨酸 C delta 1 位点的化学交换在最高场光谱中唯一观察到。这些结果进一步说明了高场 MAS SSNMR 光谱在蛋白质结构研究中的优势。
