Sampling long time scale protein motions: OSRW simulation of active site loop conformational free energies in formyl-CoA:oxalate CoA transferase.
Sampling long time scale protein motions: OSRW simulation of active site loop conformational free energies in formyl-CoA:oxalate CoA transferase.
复制标题
长时间尺度蛋白质运动采样：甲酰辅酶 A：草酸辅酶 A 转移酶活性位点环构象自由能的 OSRW 模拟。
DOI：
10.1021/ja101446u
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发表时间：
2010
影响因子：
15
通讯作者：
Richards,NigelGJ
中科院分区：
文献类型：
--
作者：
Lee,Sangbae;Chen,Mengen;Yang,Wei;Richards,NigelGJ
x射线晶体快照显示，甲酰基辅酶a：草酸辅酶a转移酶（FRC）活性位点的四赖氨酸环的构象变化在酶的催化循环中起重要作用。应用正交空间随机游走（OSRW）模拟得到了该环的“开”和“闭”构象的相对自由能的定量计算估计，以及这些状态在野生型FRC中相互转换的能垒。这些OSRW计算不仅表明这两种构象具有相似的自由能，而且还预测了一个与观察到的酶的周转数一致的势垒。为了量化四甘氨酸环中特定残基的重要性，还对G258A、G259A、G260A和G261A FRC变体进行了OSRW模拟，以检查替换每个甘氨酸残基的能量影响，并将计算的能量与动力学和结构观测相关联。在催化效率（kcat/KM）大幅降低的酶中，OSRW模拟显示采用了额外的低能环构象。在G260A FRC变体的情况下，通过模拟鉴定的新构象与在蛋白质的x射线晶体结构中观察到的构象相似。这些结果进一步证明了OSRW方法在采样构象空间中的能力，从而为功能重要的活性位点环所采用的构象提供了定量的自由能估计。此外，这些模拟模拟了与环构象变化相关的侧链的运动，从而允许通过使用纳秒模拟获得长时间尺度的运动。
X-ray crystallographic snapshots have shown that conformational changes of a tetraglycine loop in the active site of formyl-CoA:oxalate CoA transferase (FRC) play an important role in the catalytic cycle of the enzyme. Orthogonal space random walk (OSRW) simulations have been applied to obtain quantitative computational estimates of the relative free energy of the “open” and “closed” conformations of this loop together with the energetic barrier for interconversion of these states in wild type FRC. These OSRW calculations not only show that the two conformations have similar free energies but also predict a barrier that is consistent with the observed turnover number of the enzyme. In an effort to quantitate the importance of specific residues in the tetraglycine loop, OSRW simulations have also been performed on the G258A, G259A, G260A, and G261A FRC variants both to examine the energetic effects of replacing each glycine residue and to correlate the computed energies with kinetic and structural observations. In enzymes with substantially reduced catalytic efficiency (kcat/KM), the OSRW simulations reveal the adoption of additional low energy loop conformations. In the case of the G260A FRC variant, the new conformation identified by simulation is similar to that observed in the X-ray crystal structure of the protein. These results provide further evidence for the power of the OSRW method in sampling conformational space and, hence, in providing quantitative free energy estimates for the conformations adopted by functionally important active site loops. In addition, these simulations model the motions of side chains that are correlated with changes in loop conformation thereby permitting access to long time-scale motions through the use of nanosecond simulations.