Puglisi Rita, Brylski Oliver, Alfano Caterina, Martin Stephen R, Pastore Annalisa, Temussi Piero A
UK-DRI at the Wohl Institute of King's College London, 5 Cutcombe Road, SE59RT London, UK.
Institute of Physical and Theoretical Chemistry, Technische Universität Braunschweig, Braunschweig, Germany.
Commun Chem. 2020 Aug 7;3:100. doi: 10.1038/s42004-020-00358-1.
A topic that has attracted considerable interest in recent years is the possibility to perform thermodynamic studies of proteins directly in-cell or in complex environments which mimic the cellular interior. Nuclear magnetic resonance (NMR) could be an attractive technique for these studies but its applicability has so far been limited by technical issues. Here, we demonstrate that 2D NMR methods can be successfully applied to measure thermodynamic parameters provided that a suitable choice of the residues used for the calculation is made. We propose a new parameter, named RAD, which reflects the level of protection of a specific amide proton in the protein core and can guide through the selection of the resonances. We also suggest a way to calibrate the volumes to become independent of technical limitations. The methodology we propose leads to stability curves comparable to that calculated from CD data and provides a new tool for thermodynamic measurements in complex environments.
近年来引起广泛关注的一个话题是能否直接在细胞内或模拟细胞内部的复杂环境中对蛋白质进行热力学研究。核磁共振（NMR）可能是用于这些研究的一种有吸引力的技术，但迄今为止其适用性受到技术问题的限制。在这里，我们证明，只要对用于计算的残基做出合适选择，二维NMR方法就可以成功应用于测量热力学参数。我们提出了一个名为RAD的新参数，它反映了蛋白质核心中特定酰胺质子的保护水平，并可以指导共振的选择。我们还提出了一种校准体积的方法，使其不受技术限制的影响。我们提出的方法产生的稳定性曲线与从圆二色性（CD）数据计算得到的曲线相当，并为在复杂环境中的热力学测量提供了一种新工具。
