ジャーナル: Structure / 年: 2021 タイトル: A two-site flexible clamp mechanism for RET-GDNF-GFRα1 assembly reveals both conformational adaptation and strict geometric spacing. 著者: Sarah E Adams / Andrew G Purkiss / Phillip P Knowles / Andrea Nans / David C Briggs / Annabel Borg / Christopher P Earl / Kerry M Goodman / Agata Nawrotek / Aaron J Borg / Pauline B McIntosh ...著者: Sarah E Adams / Andrew G Purkiss / Phillip P Knowles / Andrea Nans / David C Briggs / Annabel Borg / Christopher P Earl / Kerry M Goodman / Agata Nawrotek / Aaron J Borg / Pauline B McIntosh / Francesca M Houghton / Svend Kjær / Neil Q McDonald / 要旨: RET receptor tyrosine kinase plays vital developmental and neuroprotective roles in metazoans. GDNF family ligands (GFLs) when bound to cognate GFRα co-receptors recognize and activate RET ...RET receptor tyrosine kinase plays vital developmental and neuroprotective roles in metazoans. GDNF family ligands (GFLs) when bound to cognate GFRα co-receptors recognize and activate RET stimulating its cytoplasmic kinase function. The principles for RET ligand-co-receptor recognition are incompletely understood. Here, we report a crystal structure of the cadherin-like module (CLD1-4) from zebrafish RET revealing interdomain flexibility between CLD2 and CLD3. Comparison with a cryo-electron microscopy structure of a ligand-engaged zebrafish RET-GDNF-GFRα1a complex indicates conformational changes within a clade-specific CLD3 loop adjacent to the co-receptor. Our observations indicate that RET is a molecular clamp with a flexible calcium-dependent arm that adapts to different GFRα co-receptors, while its rigid arm recognizes a GFL dimer to align both membrane-proximal cysteine-rich domains. We also visualize linear arrays of RET-GDNF-GFRα1a suggesting that a conserved contact stabilizes higher-order species. Our study reveals that ligand-co-receptor recognition by RET involves both receptor plasticity and strict spacing of receptor dimers by GFL ligands.
凍結剤: ETHANE / チャンバー内湿度: 100 % / チャンバー内温度: 295 K / 装置: FEI VITROBOT MARK II / 詳細: 90 s wait time, 5 s blot time.
詳細
Crosslinked using the GraFIX method with a gradient of 0-0.1% glutaraldehyde, 5-20% sucrose. Further purified by size exclusion chromatography.
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電子顕微鏡法
顕微鏡
FEI TITAN KRIOS
撮影
フィルム・検出器のモデル: GATAN K2 SUMMIT (4k x 4k) 検出モード: COUNTING / 撮影したグリッド数: 2 / 実像数: 12475 / 平均露光時間: 9.0 sec. / 平均電子線量: 48.6 e/Å2 詳細: 6105 movies were collected without stage tilt, 6375 movies were collected with a tilt angle of 30 degrees.