Graduate Group in Biochemistry & Molecular Biophysics and Department of Biochemistry & Biophysics, University of Pennsylvania, 422 Curie Boulevard, Philadelphia, Pennsylvania 19104-6059, United States.
J Am Chem Soc. 2011 Aug 17;133(32):12326-9. doi: 10.1021/ja202033k. Epub 2011 Jul 22.
The nature of water's interaction with biomolecules such as proteins has been difficult to examine in detail at atomic resolution. Solution NMR spectroscopy is potentially a powerful method for characterizing both the structural and temporal aspects of protein hydration but has been plagued by artifacts. Encapsulation of the protein of interest within the aqueous core of a reverse micelle particle results in a general slowing of water dynamics, significant reduction in hydrogen exchange chemistry and elimination of contributions from bulk water thereby enabling the use of nuclear Overhauser effects to quantify interactions between the protein surface and hydration water. Here we extend this approach to allow use of dipolar interactions between hydration water and hydrogens bonded to protein carbon atoms. By manipulating the molecular reorientation time of the reverse micelle particle through use of low viscosity liquid propane, the T(1ρ) relaxation time constants of (1)H bonded to (13)C were sufficiently lengthened to allow high quality rotating frame nuclear Overhauser effects to be obtained. These data supplement previous results obtained from dipolar interactions between the protein and hydrogens bonded to nitrogen and in aggregate cover the majority of the molecular surface of the protein. A wide range of hydration dynamics is observed. Clustering of hydration dynamics on the molecular surface is also seen. Regions of long-lived hydration water correspond with regions of the protein that participate in molecular recognition of binding partners suggesting that the contribution of the solvent entropy to the entropy of binding has been maximized through evolution.
水与生物分子（如蛋白质）相互作用的性质一直难以在原子分辨率下进行详细研究。溶液 NMR 光谱学是一种用于描述蛋白质水合作用的结构和时间方面的潜在强大方法，但一直受到伪影的困扰。将感兴趣的蛋白质封装在反胶束颗粒的水核内，会导致水动力学普遍减慢，氢交换化学显著减少，并消除来自体相水的贡献，从而能够利用核 Overhauser 效应来定量蛋白质表面与水合水之间的相互作用。在这里，我们扩展了这种方法，以允许使用水合水与与蛋白质碳原子结合的氢之间的偶极相互作用。通过使用低粘度的液体丙烷来操纵反胶束颗粒的分子重定向时间，可以充分延长与（13）C 结合的（1）H 的 T（1ρ）弛豫时间常数，从而可以获得高质量的旋转框架核 Overhauser 效应。这些数据补充了先前从蛋白质与与氮结合的氢之间的偶极相互作用获得的结果，并且总体上涵盖了蛋白质的大部分分子表面。观察到广泛的水合动力学。在分子表面上也观察到水合动力学的聚类。长寿命水合水的区域与参与结合伴侣分子识别的蛋白质区域相对应，这表明溶剂熵对结合熵的贡献已通过进化最大化。
