Department of Biothermodynamics and Drug Design, Institute of Biotechnology, Life Sciences Center, Vilnius University, Saulėtekio 7, 10257 Vilnius, Lithuania.
Latvian Institute of Organic Synthesis, Aizkraukles 21, 1006 Riga, Latvia.
J Phys Chem B. 2021 Jun 10;125(22):5823-5831. doi: 10.1021/acs.jpcb.1c02917. Epub 2021 May 25.
Proteins undergo changes in their partial volumes in numerous biological processes such as enzymatic catalysis, unfolding-refolding, and ligand binding. The change in the protein volume upon ligand binding-a parameter termed the protein-ligand binding volume-can be extensively studied by high-pressure NMR spectroscopy. In this study, we developed a method to determine the protein-ligand binding volume from a single two-dimensional (2D) H-N heteronuclear single quantum coherence (HSQC) spectrum at different pressures, if the exchange between ligand-free and ligand-bound states of a protein is slow in the NMR time-scale. This approach required a significantly lower amount of protein and NMR time to determine the protein-ligand binding volume of two carbonic anhydrase isozymes upon binding their ligands. The proposed method can be used in other protein-ligand systems and expand the knowledge about protein volume changes upon small-molecule binding.
在许多生物过程中，如酶催化、展开-重折叠和配体结合，蛋白质的部分体积会发生变化。配体结合时蛋白质体积的变化——一个参数称为蛋白质-配体结合体积——可以通过高压 NMR 光谱学进行广泛研究。在这项研究中，如果蛋白质的配体游离态和配体结合态之间的交换在 NMR 时间尺度上较慢，我们开发了一种方法，可以从单个二维（2D）H-N 异核单量子相干（HSQC）光谱在不同压力下确定蛋白质-配体结合体积。该方法需要更少的蛋白质和 NMR 时间来确定两种碳酸酐酶同工酶结合其配体时的蛋白质-配体结合体积。所提出的方法可用于其他蛋白质-配体系统，并扩展关于小分子结合时蛋白质体积变化的知识。
