National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)
R35 GM130234
米国
Human Frontier Science Program (HFSP)
LT000025/18-L1
フランス
引用
ジャーナル: Cell / 年: 2020 タイトル: Asymmetric Molecular Architecture of the Human γ-Tubulin Ring Complex. 著者: Michal Wieczorek / Linas Urnavicius / Shih-Chieh Ti / Kelly R Molloy / Brian T Chait / Tarun M Kapoor / 要旨: The γ-tubulin ring complex (γ-TuRC) is an essential regulator of centrosomal and acentrosomal microtubule formation, yet its structure is not known. Here, we present a cryo-EM reconstruction of the ...The γ-tubulin ring complex (γ-TuRC) is an essential regulator of centrosomal and acentrosomal microtubule formation, yet its structure is not known. Here, we present a cryo-EM reconstruction of the native human γ-TuRC at ∼3.8 Å resolution, revealing an asymmetric, cone-shaped structure. Pseudo-atomic models indicate that GCP4, GCP5, and GCP6 form distinct Y-shaped assemblies that structurally mimic GCP2/GCP3 subcomplexes distal to the γ-TuRC "seam." We also identify an unanticipated structural bridge that includes an actin-like protein and spans the γ-TuRC lumen. Despite its asymmetric architecture, the γ-TuRC arranges γ-tubulins into a helical geometry poised to nucleate microtubules. Diversity in the γ-TuRC subunits introduces large (>100,000 Å) surfaces in the complex that allow for interactions with different regulatory factors. The observed compositional complexity of the γ-TuRC could self-regulate its assembly into a cone-shaped structure to control microtubule formation across diverse contexts, e.g., within biological condensates or alongside existing filaments.
分子量: 200733.641 Da / 分子数: 1 / 由来タイプ: 天然 詳細: Please note that the full sequence for GCP6 (Uniprot accession number Q96RT7) was much longer than our modeled region, and the alignment failed to capture the domains we built, despite all ...詳細: Please note that the full sequence for GCP6 (Uniprot accession number Q96RT7) was much longer than our modeled region, and the alignment failed to capture the domains we built, despite all residues being numbered accordingly in the submitted coordinates. See below for full sequence. 由来: (天然) Homo sapiens (ヒト) / 参照: UniProt: Q96RT7
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実験情報
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実験
実験
手法: 電子顕微鏡法
EM実験
試料の集合状態: PARTICLE / 3次元再構成法: 単粒子再構成法
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試料調製
構成要素
名称: Native human gamma-tubulin ring complex / タイプ: COMPLEX / Entity ID: all / 由来: NATURAL
分子量
実験値: NO
由来(天然)
生物種: Homo sapiens (ヒト)
緩衝液
pH: 7.5
試料
包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES
急速凍結
凍結剤: ETHANE
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電子顕微鏡撮影
実験機器
モデル: Titan Krios / 画像提供: FEI Company
顕微鏡
モデル: FEI TITAN KRIOS
電子銃
電子線源: FIELD EMISSION GUN / 加速電圧: 300 kV / 照射モード: OTHER
電子レンズ
モード: BRIGHT FIELD
撮影
電子線照射量: 45 e/Å2 フィルム・検出器のモデル: GATAN K2 SUMMIT (4k x 4k)
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解析
ソフトウェア
名称: PHENIX / バージョン: 1.16_3549: / 分類: 精密化
CTF補正
タイプ: PHASE FLIPPING AND AMPLITUDE CORRECTION
対称性
点対称性: C1 (非対称)
3次元再構成
解像度: 4.5 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 102613 / 対称性のタイプ: POINT