Abteilung für Molekulare Strukturbiologie, Institut für Mikrobiologie und Genetik Göttinger Zentrum für Molekulare Biowissenschaften Georg-August-Universität Göttingen, Göttingen, Germany
Abteilung für Molekulare Strukturbiologie, Institut für Mikrobiologie und Genetik Göttinger Zentrum für Molekulare Biowissenschaften Georg-August-Universität Göttingen, Göttingen, Germany.
EMBO J. 2014 Nov 3;33(21):2547-63. doi: 10.15252/embj.201488517. Epub 2014 Sep 15.
Translational GTPases are universally conserved GTP hydrolyzing enzymes, critical for fidelity and speed of ribosomal protein biosynthesis. Despite their central roles, the mechanisms of GTP-dependent conformational switching and GTP hydrolysis that govern the function of trGTPases remain poorly understood. Here, we provide biochemical and high-resolution structural evidence that eIF5B and aEF1A/EF-Tu bound to GTP or GTPγS coordinate a monovalent cation (M(+)) in their active site. Our data reveal that M(+) ions form constitutive components of the catalytic machinery in trGTPases acting as structural cofactor to stabilize the GTP-bound "on" state. Additionally, the M(+) ion provides a positive charge into the active site analogous to the arginine-finger in the Ras-RasGAP system indicating a similar role as catalytic element that stabilizes the transition state of the hydrolysis reaction. In sequence and structure, the coordination shell for the M(+) ion is, with exception of eIF2γ, highly conserved among trGTPases from bacteria to human. We therefore propose a universal mechanism of M(+)-dependent conformational switching and GTP hydrolysis among trGTPases with important consequences for the interpretation of available biochemical and structural data.
转译 GTP 酶是普遍保守的 GTP 水解酶，对于核糖体蛋白生物合成的保真度和速度至关重要。尽管它们具有核心作用，但控制 trGTP 酶功能的 GTP 依赖性构象转换和 GTP 水解的机制仍知之甚少。在这里，我们提供了生化和高分辨率结构证据，表明 eIF5B 和 aEF1A/EF-Tu 与 GTP 或 GTPγS 结合，在其活性位点中协调单价阳离子 (M(+))。我们的数据表明，M(+) 离子形成 trGTP 酶催化机制的组成部分，作为结构辅助因子稳定 GTP 结合的“开”状态。此外，M(+) 离子在活性位点提供正电荷，类似于 Ras-RasGAP 系统中的精氨酸指，表明其作为催化元件的类似作用，稳定水解反应的过渡态。在序列和结构上，除了 eIF2γ 之外，M(+) 离子的配位壳在细菌到人类的 trGTP 酶中高度保守。因此，我们提出了一种普遍的 M(+)-依赖性构象转换和 trGTP 酶 GTP 水解机制，这对解释现有生化和结构数据具有重要意义。
