PROTEIN HYDRATION IN AQUEOUS-SOLUTION
PROTEIN HYDRATION IN AQUEOUS-SOLUTION
复制标题
DOI：
10.1039/fd9929300035
复制
发表时间：
1992-01-01
影响因子：
3.4
通讯作者：
LIEPINSH, E
中科院分区：
文献类型：
--
作者：
WUTHRICH, K;OTTING, G;LIEPINSH, E
利用质子核磁共振研究了与蛋白质碱性胰蛋白酶抑制剂（BPTI）以及水溶液中的九肽催产素结合的单个水化水分子。实验观察结果是蛋白质的单个氨基酸残基的质子与水化水分子的质子之间的核奥弗豪泽效应（NOE）。这些NOE通过实验室坐标系中的二维（2D）和三维（3D）NOE光谱（NOESY）以及旋转坐标系（ROESY）中的相应实验进行记录。研究表明存在两种性质不同的水合位点类型。BPTI分子内部的四个水分子在晶体结构和溶液中处于相同位置。它们与蛋白质质子的NOE具有较大的负交叉弛豫率σ(NOE)的特征，这表明这些水分子在这些水合位点的停留时间长于约10纳秒。使用外位移试剂进行的额外实验确定在4℃时这些停留时间的上限为20毫秒。球状蛋白质BPTI和柔性无序多肽催产素的表面水合是由停留时间在亚纳秒范围内的水分子进行的，这由它们与附近多肽质子的NOE所观察到的小的正σ(NOE)值所证明。所有表面水合位点的停留时间都较短，无论它们在蛋白质单晶中是否被有序的、可通过X射线观察到的水所占据。
Proton nuclear magnetic resonance was used to study individual molecules of hydration water bound to the protein basic pancreatic trypsin inhibitor (BPTI) and to the nonapeptide oxytocin in aqueous solution. The experimental observations are nuclear Overhauser effects (NOE) between protons of individual amino acid residues of the protein and those of hydration water. These NOEs were recorded by two-dimensional (2D) and three dimensional (3D) NOE spectroscopy (NOESY) in the laboratory frame, and by the corresponding experiments in the rotating frame (ROESY). The studies show that there are two qualitatively different types of hydration sites. Four water molecules in the interior of the BPTI molecule are in identical locations in the crystal structure and in solution. Their NOEs with the protein protons are characterized by large negative cross-relaxation rates sigma(NOE), which indicates that the residence times of the water molecules in these hydration sites are longer than ca. 10 ns. Additional experiments with extrinsic shift reagents established an upper limit of 20 ms at 4-degrees-C for these residence times. Surface hydration of both the globular protein BPTI and the flexibly disordered polypeptide oxytocin is by water molecules with residence times in the subnanosecond range, as evidenced by small positive sigma(NOE) values observed for their NOEs with nearby polypeptide protons. Short residence times prevail for all surface hydration sites, independent of whether or not they are occupied by well ordered, X-ray observable water in the protein single crystals.