School of Pharmacy, Key Laboratory of Molecular Pharmacology and Drug Evaluation (Yantai University), Ministry of Education, Collaborative Innovation Center of Advanced Drug Delivery System and Biotech Drugs in Universities of Shandong, Yantai University, Yantai, China.
State Key Laboratory of Long-Acting and Targeting Drug Delivery Technologies, Shandong Luye Pharmaceutical Co. Ltd., Yantai, China.
J Biomol NMR. 2020 Mar;74(2-3):173-181. doi: 10.1007/s10858-020-00302-4. Epub 2020 Feb 1.
Arginine side chains play critical roles in many protein-ligand interactions and enzyme catalysis. Unambiguous resonance assignment is a prerequisite for the nuclear magnetic resonance (NMR) spectroscopy studies of arginine side chains dynamics and hydrogen exchange properties from which one can expect to elucidate in more detail the roles of arginine residues in protein structure and function. Here we present a new mass spectrometry (MS)-based method for assigning the side-chain resonances of arginine residues in 2D H-N NMR spectra. The method requires no additional isotopic labeling, and relies on knowledge of the amino acid sequence, the modification of the guanidino groups and liquid chromatography-mass spectrometry rather than the protein's structure or properties. Correlating the modification rates can connect cross-peak positions from NMR data with MS data to support resonances assignments. In the present work, we have extended our original application to natural abundance human ubiquitin to provide ε-NH assessments of three arginine for this well-studied protein.
精氨酸侧链在许多蛋白质-配体相互作用和酶催化中起着关键作用。明确的共振分配是核磁共振（NMR）光谱研究精氨酸侧链动力学和氢交换性质的前提条件，从中可以更详细地阐明精氨酸残基在蛋白质结构和功能中的作用。在这里，我们提出了一种新的基于质谱（MS）的方法，用于分配 2D H-NMR 光谱中精氨酸残基的侧链共振。该方法不需要额外的同位素标记，而是依赖于氨基酸序列、胍基基团的修饰和液相色谱-质谱，而不是蛋白质的结构或性质。相关修饰速率可以将 NMR 数据中的交叉峰位置与 MS 数据相关联，以支持共振分配。在本工作中，我们将原始方法扩展到天然丰度的人泛素，为这个研究充分的蛋白质提供三个精氨酸的 ε-NH 评估。
