Biochemistry and Biophysics Center, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, MD, 20892, USA.
Department of Biophysics, University of Mumbai, Maharashtra, Mumbai, 400098, India.
J Biomol NMR. 2021 Mar;75(2-3):109-118. doi: 10.1007/s10858-021-00359-9. Epub 2021 Feb 24.
Paramagnetic relaxation enhancements (PREs) are routinely used to provide long-range distance restraints for the determination of protein structures, to resolve protein dynamics, ligand-protein binding sites, and lowly populated species, using Nuclear Magnetic Resonance Spectroscopy (NMR). Here, we propose a simultaneous H- N, H-C SESAME based pulse scheme for the rapid acquisition of H-R relaxation rates for the determination of backbone and sidechain PREs of proteins. The H-R rates from the traditional and our approach on Ubiquitin (UBQ) are well correlated (R = 0.99), revealing their potential to be used quantitatively. Comparison of the S57C UBQ calculated and experimental PREs provided backbone and side chain Q factors of 0.23 and 0.24, respectively, well-fitted to the UBQ NMR structure, showing that our approach can be used to acquire accurate PRE rates from the functionally important sites of proteins but in at least half the time as traditional methods.
顺磁弛豫增强（PREs）通常用于通过核磁共振光谱（NMR）提供蛋白质结构测定的远程距离约束，以解决蛋白质动力学、配体-蛋白质结合位点和低丰度物种的问题。在这里，我们提出了一种基于 H- N 和 H-C SESAME 的同时脉冲方案，用于快速获取 H-R 弛豫率，以确定蛋白质的骨架和侧链 PREs。传统方法和我们的方法在泛素（UBQ）上的 H-R 速率相关性很好（R = 0.99），表明它们具有定量应用的潜力。比较 S57C UBQ 计算和实验 PREs 的结果，提供了分别为 0.23 和 0.24 的骨架和侧链 Q 因子，与 UBQ NMR 结构拟合良好，表明我们的方法可用于从蛋白质的功能重要位点获取准确的 PRE 速率，但所需时间至少为传统方法的一半。
