Yang Jing, Zhang Ziguo, Roe S Mark, Marshall Christopher J, Barford David
Section of Structural Biology, Institute of Cancer Research, Chester Beatty Laboratories, 237 Fulham Road, London SW3 6JB, UK.
Science. 2009 Sep 11;325(5946):1398-402. doi: 10.1126/science.1174468.
Activation of Rho guanosine triphosphatases (GTPases) to the guanine triphosphate (GTP)-bound state is a critical event in their regulation of the cytoskeleton and cell signaling. Members of the DOCK family of guanine nucleotide exchange factors (GEFs) are important activators of Rho GTPases, but the mechanism of activation by their catalytic DHR2 domain is unknown. Through structural analysis of DOCK9-Cdc42 complexes, we identify a nucleotide sensor within the alpha10 helix of the DHR2 domain that contributes to release of guanine diphosphate (GDP) and then to discharge of the activated GTP-bound Cdc42. Magnesium exclusion, a critical factor in promoting GDP release, is mediated by a conserved valine residue within this sensor, whereas binding of GTP-Mg2+ to the nucleotide-free complex results in magnesium-inducing displacement of the sensor to stimulate discharge of Cdc42-GTP. These studies identify an unusual mechanism of GDP release and define the complete GEF catalytic cycle from GDP dissociation followed by GTP binding and discharge of the activated GTPase.
Rho鸟苷三磷酸酶（GTP酶）激活至结合鸟嘌呤三磷酸（GTP）的状态是其调控细胞骨架和细胞信号传导中的关键事件。鸟嘌呤核苷酸交换因子（GEF）的DOCK家族成员是Rho GTP酶的重要激活剂，但其催化性DHR2结构域的激活机制尚不清楚。通过对DOCK9-Cdc42复合物的结构分析，我们在DHR2结构域的α10螺旋内鉴定出一个核苷酸传感器，它有助于鸟嘌呤二磷酸（GDP）的释放，进而促使结合GTP的活化Cdc42的释放。促进GDP释放的关键因素——镁离子排除，由该传感器内一个保守的缬氨酸残基介导，而GTP-Mg2+与无核苷酸复合物的结合导致该传感器发生镁离子诱导的位移，从而刺激Cdc42-GTP的释放。这些研究确定了一种不同寻常的GDP释放机制，并定义了从GDP解离，随后进行GTP结合以及活化GTP酶释放的完整GEF催化循环。
