European Molecular Biology Laboratory, 71 avenue des Martyrs, CS 90181, 38042 Grenoble, France.
School of Biosciences, The University of Sheffield, Firth Court, Western Bank, Sheffield S10 2TN, UK.
Structure. 2024 Oct 3;32(10):1834-1846.e3. doi: 10.1016/j.str.2024.07.007. Epub 2024 Aug 5.
Enzymes facilitating the transfer of phosphate groups constitute the most extensive protein families across all kingdoms of life. They make up approximately 10% of the proteins found in the human genome. Understanding the mechanisms by which enzymes catalyze these reactions is essential in characterizing the processes they regulate. Metal fluorides can be used as multifunctional tools to study these enzymes. These ionic species bear the same charge as phosphate and the transferring phosphoryl group and, in addition, allow the enzyme to be trapped in catalytically important states with spectroscopically sensitive atoms interacting directly with active site residues. The ionic nature of these phosphate surrogates also allows their removal and replacement with other analogs. Here, we describe the best practices to obtain these complexes, their use in NMR, X-ray crystallography, cryo-EM, and SAXS and describe a new metal fluoride, scandium tetrafluoride, which has significant anomalous signal using soft X-rays.
酶促进磷酸基团的转移，构成了所有生命领域中最广泛的蛋白质家族。它们约占人类基因组中发现的蛋白质的 10%。了解酶催化这些反应的机制对于表征它们所调节的过程至关重要。金属氟化物可用作研究这些酶的多功能工具。这些离子物种带有与磷酸相同的电荷和正在转移的磷酸基团，此外，还允许酶被捕获在具有光谱敏感原子与活性位点残基直接相互作用的催化重要状态。这些磷酸替代物的离子性质还允许它们的去除和用其他类似物取代。在这里，我们描述了获得这些配合物的最佳实践，以及它们在 NMR、X 射线晶体学、冷冻电镜和小角 X 射线散射中的应用，并描述了一种新的金属氟化物——四氟化钪，它在软 X 射线下具有显著的异常信号。
