National Health and Medical Research Council (NHMRC, Australia)
APP1128120
オーストラリア
National Health and Medical Research Council (NHMRC, Australia)
APP1146578
オーストラリア
引用
ジャーナル: Nat Struct Mol Biol / 年: 2022 タイトル: Structure of the metastatic factor P-Rex1 reveals a two-layered autoinhibitory mechanism. 著者: Yong-Gang Chang / Christopher J Lupton / Charles Bayly-Jones / Alastair C Keen / Laura D'Andrea / Christina M Lucato / Joel R Steele / Hari Venugopal / Ralf B Schittenhelm / James C Whisstock ...著者: Yong-Gang Chang / Christopher J Lupton / Charles Bayly-Jones / Alastair C Keen / Laura D'Andrea / Christina M Lucato / Joel R Steele / Hari Venugopal / Ralf B Schittenhelm / James C Whisstock / Michelle L Halls / Andrew M Ellisdon / 要旨: P-Rex (PI(3,4,5)P-dependent Rac exchanger) guanine nucleotide exchange factors potently activate Rho GTPases. P-Rex guanine nucleotide exchange factors are autoinhibited, synergistically activated by ...P-Rex (PI(3,4,5)P-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)P 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)P 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.
T4-Lysozyme is spliced/inserted into the P-Rex1 structure in a loop region to enable ...T4-Lysozyme is spliced/inserted into the P-Rex1 structure in a loop region to enable crystallisation. However, the T4L could not be built into the PDB model as it was too flexible. Density was present but too poor to build/model the T4L.
-
実験情報
-
実験
実験
手法: X線回折 / 使用した結晶の数: 1
-
試料調製
結晶
マシュー密度: 3.86 Å3/Da / 溶媒含有率: 68.1 %
結晶化
温度: 293 K / 手法: 蒸気拡散法, ハンギングドロップ法 / 詳細: 1.8 M (NH4)2SO4, 0.05 MES pH 6.0
-
データ収集
回折
平均測定温度: 100 K / Serial crystal experiment: N
放射光源
由来: シンクロトロン / サイト: Australian Synchrotron / ビームライン: MX2 / 波長: 0.95373 Å