2009年7月21日，北京生命科学研究所柴继杰实验室在Nature Structural & Molecular Biology杂志在线发表题为“Structural insights into host GTPase isoform selection by a family of bacterial GEF mimics”的文章。该研究揭示了来自一类细菌毒性因子大家族的Map通过模拟GEF活性实现其功能，并提出了这类家族成员对不同GTPase的选择模型。生化试验证实了该模型的预测：来自Shigella的该家族另外两个成员IpgB1，IpgB2分别对宿主Rac1，RhoA具有GEF活性。
含有WXXXE模块的效应蛋白属于一类大的细菌毒性因子家族，它们通过一种未知的GTPase信号传递机制调细胞骨架动态结构。这些效应蛋白以前被认为是直接模拟激活的GTPases而行使其功能。但我们最近解析的沙门氏菌SifA的晶体结构对这一观点提出了质疑。该结构显示其C端和具有GEF活性的SopE具有相似的结构。基于这些结构信息，我们推测这个家族可能具有GEF活性。通过进一步实验，最终鉴定出来自大肠杆菌的该家族成员Map具有对人GTPase Cdc42的GEF活性。Map–Cdc42复合物结构显示Map模拟哺乳动物Dbl（GEF）的催化机制，但却具有和沙门氏菌GEF SopE几乎一致的三维结构。通过对人和细菌GEF的结构比较分析，发现Map和Cdc42内开关区域的可变区b2-3以一种独特的方式配对。同时，我们提出这类家族成员对不同GTPase的选择模型。通过该模型发现来自志贺氏菌的该家族另外两个成员IpgB1，IpgB2分别对宿主Rac1，RhoA具有GEF活性。这些结果扩大了人们对细菌的GEF模拟以及其对宿主GTPase底物选择机制的认识。
北京生命科学研究所博士后黄志伟和德州西南医学中心的Sarah E Sutton为本文共同第一作者。该文其他作者还有德州西南医学中心的Adam J Wallenfang和Robert C Orchard，以及北京生命科学研究所吴晓静和冯英才技术员。柴继杰博士和德州西南医学中心的Neal M Alto博士为本文共同通讯作者。此项研究为科技部863和北京市科委资助课题，结构及部分生化工作在北京生命科学研究所完成。（生物谷Bioon.com）
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Nature Structural & Molecular Biology 20 July 2009 | doi:10.1038/nsmb.1647
Structural insights into host GTPase isoform selection by a family of bacterial GEF mimics
Zhiwei Huang1,4, Sarah E Sutton2,4, Adam J Wallenfang2, Robert C Orchard2, Xiaojing Wu1, Yingcai Feng1, Jijie Chai1,3 & Neal M Alto2
The Escherichia coli type III effector Map belongs to a large family of bacterial virulence factors that activate host Rho GTPase signaling pathways through an unknown molecular mechanism. Here we report direct evidence that Map functions as a potent and selective guanine-nucleotide exchange factor (GEF) for Cdc42. The 2.3-? structure of the Map–Cdc42 complex revealed that Map mimics the GEF strategy of the mammalian Dbl family but has a three-dimensional architecture that is nearly identical to the bacterial GEF Salmonella spp. SopE. A comparative analysis between human and bacterial GEFs revealed a previously uncharacterized pairing mechanism between Map and the variable 2-3 interswitch region of Cdc42. We propose a GTPase selection model that is experimentally validated by the preferential activation Rac1 and RhoA by the Shigella spp. effectors IpgB1 and IpgB2, respectively. These results significantly expand the repertoire of bacterial GEF mimics and unify a GEF selection mechanism for host GTPase substrates.
1 National Institute of Biological Sciences, Zhong Guan Cun Life Science Park, Beijing, China.
2 Department of Microbiology, University of Texas Southwestern Medical Center, Dallas, Texas, USA.
3 Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing, China.
4 These authors contributed equally to this work.
