Max-Planck-Institut für Kohlenforschung, Kaiser-Wilhelm-Platz 1, D-45470, Mülheim an der Ruhr, Germany.
J Phys Chem B. 2010 Nov 25;114(46):15413-23. doi: 10.1021/jp108095n. Epub 2010 Oct 26.
Quantum refinement is an improvement upon the molecular mechanics (MM)-based crystallographic refinement. In the latter, X-ray data are supplemented with additional chemical information through MM force fields, whereas quantum refinement describes crucial regions of interest in the macromolecule by quantum mechanics (QM) instead of MM. In this paper, we report the implementation of quantum refinement in the ChemShell QM/MM framework and its application in an investigation of the chromophore structure of the red fluorescent protein DsRed.M1. Both mechanical and electrostatic QM/MM embedding schemes are implemented and tested. In the quantum refinement of DsRed.M1, the anionic red acylimine chromophore adopts a nearly orthogonal arrangement (rather than a cis or trans form), and the bond lengths in the acylimine moiety are more consistent with a phenolate (rather than a quinoid) structure. These findings are in contrast to the structure deduced from a standard crystallographic refinement (PDB: 2VAD), but in agreement with our earlier results from a purely theoretical QM/MM study. On the other hand, the quantum refinement of the anionic acylimine form of DsRed.M1 yields a hydrogen bonding network around the chromophore, especially with regard to the arrangement of the water molecules and the Glu148 residue, that is closer to the 2VAD structure than to the previously optimized QM/MM structure. In our earlier study the initial classical molecular dynamics (MD) simulations during QM/MM setup apparently exaggerated the mobility of the water molecules around the chromophore. On the basis of the present results, it seems likely that the Glu148 residue is protonated in the DsRed.M1 protein. The calculation of electronic excitation energies allows for further assessment of the proposed structures, especially in the chromophore region. Using a combination of density functional theory and multireference configuration interaction (DFT/MRCI), we find excellent agreement between experiment and theory only for the structures obtained from quantum refinement and from QM/MM optimization, but not for the 2VAD structure. The present case study on DsRed.M1 thus demonstrates the merits of combining reliable theoretical and experimental information in the determination of protein structures.
量子精修是对基于分子力学（MM）的晶体学精修的改进。在后一种方法中，通过 MM 力场补充 X 射线数据的其他化学信息，而量子精修则通过量子力学（QM）而非 MM 来描述大分子中关键的感兴趣区域。在本文中，我们报告了 ChemShell QM/MM 框架中量子精修的实现及其在研究红色荧光蛋白 DsRed.M1 发色团结构中的应用。我们实现并测试了机械和静电 QM/MM 嵌入方案。在 DsRed.M1 的量子精修中，阴离子红色酰亚胺发色团采用近乎正交的排列（而不是顺式或反式形式），并且酰亚胺部分的键长更符合酚盐（而不是醌型）结构。这些发现与标准晶体学精修（PDB：2VAD）推断的结构形成对比，但与我们之前从纯理论 QM/MM 研究中得出的结果一致。另一方面，DsRed.M1 的阴离子酰亚胺形式的量子精修产生了发色团周围的氢键网络，特别是在水分子和 Glu148 残基的排列方面，与 2VAD 结构比与之前优化的 QM/MM 结构更接近。在我们之前的研究中，QM/MM 设置过程中的初始经典分子动力学（MD）模拟显然夸大了水分子围绕发色团的迁移率。根据目前的结果，似乎很可能在 DsRed.M1 蛋白中 Glu148 残基被质子化。电子激发能的计算允许进一步评估所提出的结构，特别是在发色团区域。使用密度泛函理论和多参考组态相互作用（DFT/MRCI）的组合，我们仅在从量子精修和 QM/MM 优化获得的结构中找到实验和理论之间的极好一致性，而不是在 2VAD 结构中。因此，对 DsRed.M1 的案例研究证明了在确定蛋白质结构时结合可靠的理论和实验信息的优点。
