Structure-Energy Correlation in Proteins

描述（由申请人提供）：蛋白质对内部瞬时存在的电荷作出反应的能力对其生物学功能至关重要。这是控制能量转换的东西，也是催化的关键。尽管普遍的重要性，这一功能特性的方式，其中电荷被容纳在蛋白质内部，内部基团的电离的结构响应的性质，和内部残基的pKa值的分子决定因素，仍然未知。实验研究提出的特点，在一个前所未有的详细程度，结构和能量的后果内的蛋白质电离过程。葡萄球菌核酸酶的50种变体将用于此目的，这些变体先前在我们的实验室中在25个不同的内部位置用Lys或Glu进行了工程化。X射线晶体学，NMR光谱学和平衡热力学实验将被用来表征静态和动态响应，由内部残留物的电离触发。永久性和诱导偶极子，内部和表面电荷之间的相互作用，水的渗透，和当地展开的作用将被研究。必要时，还将采用计算方法来解决这个问题。例如，在与其他小组的合作中，局部极化率将用量子方法计算，标准分子动力学（MD）将用于研究水渗透和偶极弛豫，复制交换MD方法将用于研究内部基团的电离如何重新排列骨架。从这些研究中获得的实验数据将用于改进和测试最流行的基于结构的pKa计算的计算方法的性能。从实验和计算研究的结合中产生的物理洞察力将进一步理解对所有生命系统都至关重要的关键功能基序。这可能会影响我们对许多疾病分子基础的理解，并将提高我们合理设计药物和蛋白质的能力。

DESCRIPTION (provided by applicant): The ability of proteins to respond to the transient presence of charge in their interior is critical for their biological function. This is what governs energy transduction, and it is also critical in catalysis. Despite the universal importance of this functional property, the manner in which charge is accommodated in the protein interior, the nature of the structural response to the ionization of an internal group, and the molecular determinants of the pKa values of internal residues, remain unknown. Experimental studies are proposed to characterize, at an unprecedented level of detail, the structural and energetic consequences of ionization processes inside proteins. 50 variants of staphylococcal nuclease, engineered previously in our laboratory with Lys or Glu in 25 different internal positions, will be used for this purpose. X-ray crystallography, NMR spectroscopy, and equilibrium thermodynamic experiments will be used to characterize static and dynamic responses, triggered by the ionization of internal residues. The roles of permanent and induced dipoles, of interactions between interior and surface charges, of water penetration, and of local unfolding will be studied. Computational methods will also be brought to bear on the problem, as necessary. For example, in collaboration with other groups, local polarizabilities will be calculated with quantum methods, standard molecular dynamics (MD) will be used to study water penetration and dipolar relaxation, and replica-exchange MD methods will be used to study how ionization of internal groups can rearrange the backbone. The experimental data obtained from these studies will be used to refine and to test the performance of the most popular computational methods for structure-based pKa calculations. The physical insight that will emerge from the combined experimental and computational studies will further understanding of a key functional motif that is central to all living systems. This could impact our understanding of the molecular basis of many diseases, and it will improve our ability to design drugs and proteins rationally.