The role of the conserved switch II glutamate in guanine nucleotide exchange factor-mediated nucleotide exchange of GTP-binding proteins
The role of the conserved switch II glutamate in guanine nucleotide exchange factor-mediated nucleotide exchange of GTP-binding proteins
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DOI：
10.1016/j.jmb.2008.03.011
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发表时间：
2008-05-23
影响因子：
5.6
通讯作者：
Wittinghofer, Alfred
中科院分区：
文献类型：
--
作者：
Gasper, Raphael;Thomas, Christoph;Wittinghofer, Alfred
鸟嘌呤核苷酸交换因子（GEF）通过催化GDP与GTP之间的缓慢交换来调节小G蛋白的活性。该机制涉及三聚体G蛋白-核苷酸-GEF复合物的形成，随后释放核苷酸以形成稳定的二元G蛋白-GEF复合物。许多G蛋白-GEF复合物的结构研究表明，在核苷酸结合位点诱导的大的结构变化。再加上最近的三聚体复合物的结构，这些研究不仅提出了一些共同的原则，但在GEF介导的交换反应的细节也有很大的差异。几种结构表明，开关II中的谷氨酸残基，这是DxxGQE基序的一部分，在Ras样G蛋白中高度保守，可能在GEF介导的核苷酸交换反应中具有决定性的机械作用。在这里，我们表明，开关II谷氨酸突变为丙氨酸严重损害GEF催化的核苷酸交换在大多数，但不是所有的Ras家族G蛋白，解释其高序列保守性。该残基决定了GEF对核苷酸负载G蛋白的初始方法，并且不会明显影响二元无核苷酸复合物的形成。因此，它的主要作用似乎是从与核苷酸的相互作用中去除P-环赖氨酸。(C)2008爱思唯尔有限公司保留所有权利。
Guanine nucleotide exchange factors (GEFs) regulate the activity of small G proteins by catalysing the intrinsically slow exchange of GDP for GTP. The mechanism involves the formation of trimeric G protein-nucleotide-GEF complexes, followed by the release of nucleotide to form stable binary G protein-GEF complexes. A number of structural studies of G protein-GEF complexes have shown large structural changes induced in the nucleotide binding site. Together with a recent structure of a trimeric complex, these studies have suggested not only some common principles but also large differences in detail in the GEF-mediated exchange reaction. Several structures suggested that a glutamic acid residue in switch II, which is part of the DxxGQE motif and highly conserved in Ras-like G proteins, might have a decisive mechanistic role in GEF-mediated nucleotide exchange reactions. Here we show that mutation of the switch II glutamate to Ala severely impairs GEF-catalysed nucleotide exchange in most, but not all, Ras family G proteins, explaining its high sequence conservation. The residue determines the initial approach of GEF to the nucleotide-loaded G protein and does not appreciably affect the formation of a binary nucleotide-free complex. Its major effect thus appears to be the removal of the P-loop lysine from its interaction with the nucleotide. (C) 2008 Elsevier Ltd. All rights reserved.