Cohen Rachel D, Guseman Alex J, Pielak Gary J
Department of Chemistry, University of North Carolina, Chapel Hill, North Carolina, 27599.
Department of Biochemistry and Biophysics, University of North Carolina, Chapel Hill, North Carolina, 27599.
Protein Sci. 2015 Nov;24(11):1748-55. doi: 10.1002/pro.2765. Epub 2015 Aug 30.
NMR spectroscopy can provide information about proteins in living cells. pH is an important characteristic of the intracellular environment because it modulates key protein properties such as net charge and stability. Here, we show that pH modulates quinary interactions, the weak, ubiquitous interactions between proteins and other cellular macromolecules. We use the K10H variant of the B domain of protein G (GB1, 6.2 kDa) as a pH reporter in Escherichia coli cells. By controlling the intracellular pH, we show that quinary interactions influence the quality of in-cell (15) N-(1) H HSQC NMR spectra. At low pH, the quality is degraded because the increase in attractive interactions between E. coli proteins and GB1 slows GB1 tumbling and broadens its crosspeaks. The results demonstrate the importance of quinary interactions for furthering our understanding of protein chemistry in living cells.
核磁共振光谱能够提供有关活细胞中蛋白质的信息。pH值是细胞内环境的一个重要特征，因为它能调节关键的蛋白质特性，如净电荷和稳定性。在此，我们表明pH值可调节第五级相互作用，即蛋白质与其他细胞大分子之间微弱且普遍存在的相互作用。我们将蛋白G（GB1，6.2 kDa）B结构域的K10H变体用作大肠杆菌细胞中的pH报告分子。通过控制细胞内pH值，我们发现第五级相互作用会影响细胞内（15）N-（1）H HSQC核磁共振光谱的质量。在低pH值下，光谱质量会下降，因为大肠杆菌蛋白质与GB1之间吸引力相互作用的增加会减缓GB1的翻滚并拓宽其交叉峰。这些结果证明了第五级相互作用对于加深我们对活细胞中蛋白质化学的理解的重要性。
