P-Rex (PI(3,4,5)P₃-dependent Rac exchanger) guanine nucleotide exchange factors potently activate Rho GTPases. P-Rex guanine nucleotide exchange factors are autoinhibited, synergistically activated by Gβγ and PI(3,4,5)P₃ binding and dysregulated in cancer. Here, we use X-ray crystallography, cryogenic electron microscopy and crosslinking mass spectrometry to determine the structural basis of human P-Rex1 autoinhibition. P-Rex1 has a bipartite structure of N- and C-terminal modules connected by a C-terminal four-helix bundle that binds the N-terminal Pleckstrin homology (PH) domain. In the N-terminal module, the Dbl homology (DH) domain catalytic surface is occluded by the compact arrangement of the DH-PH-DEP1 domains. Structural analysis reveals a remarkable conformational transition to release autoinhibition, requiring a 126° opening of the DH domain hinge helix. The off-axis position of Gβγ and PI(3,4,5)P₃ binding sites further suggests a counter-rotation of the P-Rex1 halves by 90° facilitates PH domain uncoupling from the four-helix bundle, releasing the autoinhibited DH domain to drive Rho GTPase signaling.

P-Rex（PI(3,4,5)P ₃依赖性 Rac 交换剂）鸟嘌呤核苷酸交换因子可有效激活 Rho GTPase。P-Rex 鸟嘌呤核苷酸交换因子是自抑制的，由 Gβγ 和 PI(3,4,5)P _{3协同激活}在癌症中的结合和失调。在这里，我们使用 X 射线晶体学、低温电子显微镜和交联质谱来确定人类 P-Rex1 自身抑制的结构基础。P-Rex1 具有由结合 N 端 Pleckstrin 同源 (PH) 域的 C 端四螺旋束连接的 N 端和 C 端模块的二分结构。在 N 端模块中，Dbl 同源 (DH) 域催化表面被 DH-PH-DEP1 域的紧凑排列封闭。结构分析揭示了释放自抑制的显着构象转变，需要 DH 结构域铰链螺旋打开 126°。_{Gβγ和PI(3,4,5)P 3}的离轴位置结合位点进一步表明 P-Rex1 的一半反向旋转 90° 有助于 PH 结构域从四螺旋束中解偶联，释放自身抑制的 DH 结构域以驱动 Rho GTPase 信号传导。