Department of Theoretical Chemistry, Lund University, PO Box 124, 221 00 Lund, Sweden.
Acta Crystallogr D Struct Biol. 2020 Nov 1;76(Pt 11):1145-1156. doi: 10.1107/S2059798320012917. Epub 2020 Oct 16.
X-ray crystallography is the main source of atomistic information on the structure of proteins. Normal crystal structures are obtained as a compromise between the X-ray scattering data and a set of empirical restraints that ensure chemically reasonable bond lengths and angles. However, such restraints are not always available or accurate for nonstandard parts of the structure, for example substrates, inhibitors and metal sites. The method of quantum refinement, in which these empirical restraints are replaced by quantum-mechanical (QM) calculations, has previously been suggested for small but interesting parts of the protein. Here, this approach is extended to allow for multiple conformations in the QM region by performing separate QM calculations for each conformation. This approach is shown to work properly and leads to improved structures in terms of electron-density maps and real-space difference density Z-scores. It is also shown that the quality of the structures can be gauged using QM strain energies. The approach, called ComQumX-2QM, is applied to the P-cluster in two different crystal structures of the enzyme nitrogenase, i.e. an FeSCys cluster, used for electron transfer. One structure is at a very high resolution (1.0 Å) and shows a mixture of two different oxidation states, the fully reduced P state (Fe, 20%) and the doubly oxidized P state (80%). In the original crystal structure the coordinates differed for only two iron ions, but here it is shown that the two states also show differences in other atoms of up to 0.7 Å. The second structure is at a more modest resolution, 2.1 Å, and was originally suggested to show only the one-electron oxidized state, P. Here, it is shown that it is rather a 50/50% mixture of the P and P states and that many of the Fe-Fe and Fe-S distances in the original structure were quite inaccurate (by up to 0.8 Å). This shows that the new ComQumX-2QM approach can be used to sort out what is actually seen in crystal structures with dual conformations and to give locally improved coordinates.
X 射线晶体学是获取蛋白质结构原子级信息的主要手段。通常情况下，晶体结构是在 X 射线散射数据和一组经验约束之间达成平衡的结果，这些经验约束确保了化学意义上合理的键长和键角。然而，对于结构的非标准部分（例如底物、抑制剂和金属结合位点），这种约束并不总是可用或准确的。量子精修方法曾被提议用于蛋白质的小但有趣的部分，该方法用量子力学（QM）计算取代这些经验约束。在这里，通过为每个构象执行单独的 QM 计算，将这种方法扩展到允许 QM 区域存在多个构象。结果表明，该方法能够正确工作，并在电子密度图和实空间差分密度 Z 分数方面改善结构。还表明，可以使用 QM 应变能来评估结构的质量。该方法称为 ComQumX-2QM，应用于两种不同晶体结构的氮酶 P 簇，即用于电子转移的 FeSCys 簇。一个结构的分辨率非常高（1.0 Å），显示出两种不同氧化态的混合物，完全还原的 P 态（Fe，20%）和双氧化的 P 态（80%）。在原始晶体结构中，只有两个铁离子的坐标不同，但这里表明，这两种状态在其他原子上也存在高达 0.7 Å 的差异。第二个结构的分辨率稍低，为 2.1 Å，最初被认为仅显示单电子氧化态 P。在这里，表明它实际上是 P 和 P 态的 50/50%混合物，并且原始结构中的许多 Fe-Fe 和 Fe-S 距离非常不准确（高达 0.8 Å）。这表明，新的 ComQumX-2QM 方法可用于理清具有双重构象的晶体结构中实际观察到的情况，并给出局部改善的坐标。
