Institut für Organische Chemie, Universität Innsbruck, 6020 Innsbruck, Austria.
J Am Chem Soc. 2012 Sep 12;134(36):14800-7. doi: 10.1021/ja303591y. Epub 2012 Aug 28.
We demonstrate that conformational exchange processes in proteins on microsecond-to-millisecond time scales can be detected and quantified by solid-state NMR spectroscopy. We show two independent approaches that measure the effect of conformational exchange on transverse relaxation parameters, namely Carr-Purcell-Meiboom-Gill relaxation-dispersion experiments and measurement of differential multiple-quantum coherence decay. Long coherence lifetimes, as required for these experiments, are achieved by the use of highly deuterated samples and fast magic-angle spinning. The usefulness of the approaches is demonstrated by application to microcrystalline ubiquitin. We detect a conformational exchange process in a region of the protein for which dynamics have also been observed in solution. Interestingly, quantitative analysis of the data reveals that the exchange process is more than 1 order of magnitude slower than in solution, and this points to the impact of the crystalline environment on free energy barriers.
我们证明，通过固态 NMR 光谱学可以检测和量化微秒到毫秒时间尺度上蛋白质中的构象交换过程。我们展示了两种独立的方法，这些方法测量构象交换对横向弛豫参数的影响，即 Carr-Purcell-Meiboom-Gill 弛豫-弥散实验和测量差示多量子相干衰减。通过使用高度氘化的样品和快速魔角旋转，实现了这些实验所需的长相干寿命。这些方法的有用性通过应用于微结晶泛素得到了证明。我们在蛋白质的一个区域中检测到构象交换过程，在该区域中也观察到了溶液中的动力学。有趣的是，对数据的定量分析表明，交换过程比溶液中的速度慢一个数量级以上，这表明结晶环境对自由能势垒有影响。
