Moreira Irina S, Fernandes Pedro A, Ramos Maria J
Requimte/Departamento de Química, Faculdade de Ciências da Universidade do Porto, Rua do Campo Alegre 687, 4169-007 Porto- Portugal.
J Phys Chem B. 2007 Mar 15;111(10):2697-706. doi: 10.1021/jp067096p. Epub 2007 Feb 22.
Alanine scanning of protein-protein interfaces has shown that there are some residues in the protein-protein interfaces, responsible for most of the binding free energy, which are called hot spots. Hot spots tend to exist in densely packed central clusters, and a hypothesis has been proposed that considers that inaccessibility to the solvent must be a necessary condition to define a residue as a binding hot spot. This O-ring hypothesis is mainly based on the analysis of the accessible surface area (ASA) of 23 static, crystallographic structures of protein complexes. It is known, however, that protein flexibility allows for temporary exposures of buried interfacial groups, and even though the ASA provides a general trend of the propensity for hydration, protein/solvent-specific interactions or hydrogen bonding cannot be considered here. Therefore, a microscopic level, atomistic picture of hot spot solvation is needed to support the O-ring hypothesis. In this study, we began by applying a computational alanine-scanning mutagenesis technique, which reproduces the experimental results and allows for decomposing the binding free energy difference in its different energetic factors. Subsequently, we calculated the radial distribution function and residence times of the water molecules near the hot/warm spots to study the importance of the water environment around those energetically important amino acid residues. This study shows that within a flexible, dynamic protein framework, the warm/hot spot residues are, indeed, kept sheltered from the bulk solvent during the whole simulation, which allows a better interacting microenvironment.
蛋白质 - 蛋白质界面的丙氨酸扫描表明，在蛋白质 - 蛋白质界面中存在一些对大部分结合自由能起作用的残基，这些残基被称为热点。热点往往存在于紧密堆积的中心簇中，并且有人提出了一种假设，认为溶剂不可及性必定是将一个残基定义为结合热点的必要条件。这种O环假设主要基于对23个蛋白质复合物静态晶体结构的可及表面积（ASA）的分析。然而，众所周知，蛋白质的柔韧性会使埋藏的界面基团出现暂时暴露的情况，而且尽管ASA提供了水合倾向的总体趋势，但这里无法考虑蛋白质/溶剂特异性相互作用或氢键。因此，需要一个微观层面、原子水平的热点溶剂化图景来支持O环假设。在本研究中，我们首先应用了一种计算丙氨酸扫描诱变技术，该技术重现了实验结果，并能将结合自由能差分解为不同的能量因素。随后，我们计算了热点/温点附近水分子的径向分布函数和停留时间，以研究这些能量上重要的氨基酸残基周围水环境的重要性。这项研究表明，在一个灵活、动态的蛋白质框架内，在整个模拟过程中，温点/热点残基确实与大量溶剂隔绝，这使得它们有一个更好的相互作用微环境。
