Aix Marseille Univ, CNRS, BIP, Bioénergétique et Ingénierie des Protéines, IMM, Marseille, France.
Laboratory of Bioinorganic Chemistry, Department of Pharmacy and Biotechnology, University of Bologna, 40127 Bologna, Italy.
Biomolecules. 2020 Jul 16;10(7):1062. doi: 10.3390/biom10071062.
UreG is a P-loop GTP hydrolase involved in the maturation of nickel-containing urease, an essential enzyme found in plants, fungi, bacteria, and archaea. This protein couples the hydrolysis of GTP to the delivery of Ni(II) into the active site of apo-urease, interacting with other urease chaperones in a multi-protein complex necessary for enzyme activation. Whereas the conformation of () UreG was solved by crystallography when it is in complex with two other chaperones, in solution the protein was found in a disordered and flexible form, defining it as an intrinsically disordered enzyme and indicating that the well-folded structure found in the crystal state does not fully reflect the behavior of the protein in solution. Here, isothermal titration calorimetry and site-directed spin labeling coupled to electron paramagnetic spectroscopy were successfully combined to investigate UreG structural dynamics in solution and the effect of Ni(II) and GTP on protein mobility. The results demonstrate that, although the protein maintains a flexible behavior in the metal and nucleotide bound forms, concomitant addition of Ni(II) and GTP exerts a structural change through the crosstalk of different protein regions.
UreG 是一种 P 环 GTP 水解酶，参与镍结合脲酶的成熟，脲酶是一种存在于植物、真菌、细菌和古菌中的必需酶。该蛋白将 GTP 的水解与 Ni(II) 递送入脱辅基脲酶的活性部位偶联，与多蛋白复合物中的其他脲酶伴侣相互作用，该复合物对于酶的激活是必需的。虽然 () UreG 的构象在与另外两种伴侣蛋白形成复合物时通过晶体学方法得到解决，但在溶液中，该蛋白呈无定形和灵活的形式，将其定义为一种固有无序的酶，并表明在晶体状态下发现的折叠良好的结构不能完全反映蛋白质在溶液中的行为。在这里，使用等温滴定量热法和与电子顺磁共振波谱相结合的定点自旋标记成功地结合起来，研究了溶液中 UreG 的结构动力学以及 Ni(II) 和 GTP 对蛋白质迁移率的影响。结果表明，尽管该蛋白在金属和核苷酸结合形式下保持灵活的行为，但 Ni(II) 和 GTP 的同时添加通过不同蛋白区域的串扰施加结构变化。
