Mhashal Anil R, Choudhury Chandan Kumar, Roy Sudip
Physical Chemistry Division, National Chemical Laboratory, Pune, 411008, India.
J Mol Model. 2016 Mar;22(3):54. doi: 10.1007/s00894-016-2922-3. Epub 2016 Feb 10.
Helicases are enzymes that unwind double-stranded DNA (dsDNA) into its single-stranded components. It is important to understand the binding and unbinding of ATP from the active sites of helicases, as this knowledge can be used to elucidate the functionality of helicases during the unwinding of dsDNA. In this work, we investigated the unbinding of ATP and its effect on the active-site residues of the helicase PcrA using molecular dynamic simulations. To mimic the unbinding process of ATP from the active site of the helicase, we simulated the application of an external force that pulls ATP from the active site and computed the free-energy change during this process. We estimated an energy cost of ~85 kJ/mol for the transformation of the helicase from the ATP-bound state (1QHH) to the ATP-free state (1PJR). Unbinding led to conformational changes in the residues of the protein at the active site. Some of the residues at the ATP-binding site were significantly reoriented when the ATP was pulled. We observed a clear competition between reorientation of the residues and energy stabilization by hydrogen bonds between the ATP and active-site residues. We also checked the flexibility of the PcrA protein using a principal component analysis of domain motion. We found that the ATP-free state of the helicase is more flexible than the ATP-bound state.
解旋酶是一种将双链DNA（dsDNA）解旋为单链成分的酶。了解ATP在解旋酶活性位点的结合和解离非常重要，因为这些知识可用于阐明解旋酶在dsDNA解旋过程中的功能。在这项工作中，我们使用分子动力学模拟研究了ATP的解离及其对解旋酶PcrA活性位点残基的影响。为了模拟ATP从解旋酶活性位点的解离过程，我们模拟了一种从活性位点拉拽ATP的外力作用，并计算了此过程中的自由能变化。我们估计解旋酶从ATP结合状态（1QHH）转变为无ATP状态（1PJR）的能量消耗约为85kJ/mol。ATP解离导致活性位点处蛋白质残基的构象变化。当ATP被拉拽时，ATP结合位点的一些残基发生了显著的重新定向。我们观察到残基重新定向与ATP和活性位点残基之间氢键导致的能量稳定之间存在明显竞争。我们还通过对结构域运动进行主成分分析来检查PcrA蛋白的灵活性。我们发现解旋酶的无ATP状态比ATP结合状态更灵活。
