Scrima Andrea, Wittinghofer Alfred
Max-Planck-Institut für Molekulare Physiologie, Dortmund, Germany.
EMBO J. 2006 Jun 21;25(12):2940-51. doi: 10.1038/sj.emboj.7601171. Epub 2006 Jun 8.
MnmE, a Guanine nucleotide-binding protein conserved between bacteria and man, is involved in the modification of tRNAs. Here we provide biochemical and X-ray structural evidence for a new GTP-hydrolysis mechanism, where the G-domains of MnmE dimerise in a potassium-dependent manner and induce GTP hydrolysis. The structure in the presence of GDP-AlFx and potassium shows how juxtaposition of the subunits induces a conformational change around the nucleotide which reorients the catalytic machinery. A critical glutamate is positioned such as to stabilise or activate the attacking water. Potassium provides a positive charge into the catalytic site in a position analogous to the arginine finger in the Ras-RasGAP system. Mutational studies show that potassium-dependent dimerisation and GTP hydrolysis can be uncoupled and that interaction between the G-domains is a prerequisite for subsequent phosphoryl transfer. We propose a model for the juxtaposition of G-domains in the full-length protein and how it induces conformational changes in the putative tRNA-modification centre.
MnmE是一种在细菌和人类之间保守的鸟嘌呤核苷酸结合蛋白，参与tRNA的修饰。在此，我们提供了一种新的GTP水解机制的生化和X射线结构证据，即MnmE的G结构域以钾离子依赖的方式二聚化并诱导GTP水解。在GDP-AlFx和钾离子存在下的结构显示了亚基的并列如何诱导核苷酸周围的构象变化，从而重新定位催化机制。一个关键的谷氨酸定位在稳定或激活进攻水的位置。钾离子在类似于Ras-RasGAP系统中精氨酸指的位置向催化位点提供正电荷。突变研究表明，钾离子依赖的二聚化和GTP水解可以解偶联，并且G结构域之间的相互作用是随后磷酸转移的先决条件。我们提出了一个全长蛋白中G结构域并列的模型，以及它如何在假定的tRNA修饰中心诱导构象变化。
