+データを開く
-基本情報
登録情報 | データベース: EMDB / ID: EMD-21666 | ||||||||||||||||||
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タイトル | Cryo-EM map of the mutant OCCM (ORC-Cdc6-Cdt1-Mcm2-7 with Mcm6 WHD truncation) loaded on DNA at 10.5 A resolution | ||||||||||||||||||
マップデータ | |||||||||||||||||||
試料 |
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キーワード | DNA replication / OCCM-deltaC6 / REPLICATION-DNA complex | ||||||||||||||||||
機能・相同性 | 機能・相同性情報 MCM complex loading / CDC6 association with the ORC:origin complex / Cul8-RING ubiquitin ligase complex / maintenance of rDNA / MCM core complex / Assembly of the pre-replicative complex / Switching of origins to a post-replicative state / MCM complex binding / nuclear DNA replication / Assembly of the ORC complex at the origin of replication ...MCM complex loading / CDC6 association with the ORC:origin complex / Cul8-RING ubiquitin ligase complex / maintenance of rDNA / MCM core complex / Assembly of the pre-replicative complex / Switching of origins to a post-replicative state / MCM complex binding / nuclear DNA replication / Assembly of the ORC complex at the origin of replication / nuclear origin of replication recognition complex / premeiotic DNA replication / pre-replicative complex assembly involved in nuclear cell cycle DNA replication / mitotic DNA replication / Activation of the pre-replicative complex / CMG complex / nucleosome organization / nuclear pre-replicative complex / MCM complex / Activation of ATR in response to replication stress / DNA replication preinitiation complex / cyclin-dependent protein serine/threonine kinase inhibitor activity / mitotic DNA replication checkpoint signaling / replication fork protection complex / double-strand break repair via break-induced replication / mitotic DNA replication initiation / single-stranded DNA helicase activity / silent mating-type cassette heterochromatin formation / regulation of DNA-templated DNA replication initiation / DNA strand elongation involved in DNA replication / CDK-mediated phosphorylation and removal of Cdc6 / DNA unwinding involved in DNA replication / Orc1 removal from chromatin / nuclear replication fork / regulation of DNA replication / DNA replication origin binding / DNA replication initiation / subtelomeric heterochromatin formation / nucleosome binding / DNA helicase activity / helicase activity / transcription elongation by RNA polymerase II / G1/S transition of mitotic cell cycle / heterochromatin formation / chromosome / single-stranded DNA binding / chromosome, telomeric region / DNA helicase / cell division / chromatin binding / GTPase activity / DNA damage response / GTP binding / ATP hydrolysis activity / nucleoplasm / ATP binding / nucleus / metal ion binding / cytoplasm 類似検索 - 分子機能 | ||||||||||||||||||
生物種 | Saccharomyces cerevisiae (パン酵母) | ||||||||||||||||||
手法 | 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 10.0 Å | ||||||||||||||||||
データ登録者 | Yuan Z / Schneider S | ||||||||||||||||||
資金援助 | 米国, 英国, 5件
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引用 | ジャーナル: Proc Natl Acad Sci U S A / 年: 2020 タイトル: Structural mechanism of helicase loading onto replication origin DNA by ORC-Cdc6. 著者: Zuanning Yuan / Sarah Schneider / Thomas Dodd / Alberto Riera / Lin Bai / Chunli Yan / Indiana Magdalou / Ivaylo Ivanov / Bruce Stillman / Huilin Li / Christian Speck / 要旨: DNA replication origins serve as sites of replicative helicase loading. In all eukaryotes, the six-subunit origin recognition complex (Orc1-6; ORC) recognizes the replication origin. During late M- ...DNA replication origins serve as sites of replicative helicase loading. In all eukaryotes, the six-subunit origin recognition complex (Orc1-6; ORC) recognizes the replication origin. During late M-phase of the cell-cycle, Cdc6 binds to ORC and the ORC-Cdc6 complex loads in a multistep reaction and, with the help of Cdt1, the core Mcm2-7 helicase onto DNA. A key intermediate is the ORC-Cdc6-Cdt1-Mcm2-7 (OCCM) complex in which DNA has been already inserted into the central channel of Mcm2-7. Until now, it has been unclear how the origin DNA is guided by ORC-Cdc6 and inserted into the Mcm2-7 hexamer. Here, we truncated the C-terminal winged-helix-domain (WHD) of Mcm6 to slow down the loading reaction, thereby capturing two loading intermediates prior to DNA insertion in budding yeast. In "semi-attached OCCM," the Mcm3 and Mcm7 WHDs latch onto ORC-Cdc6 while the main body of the Mcm2-7 hexamer is not connected. In "pre-insertion OCCM," the main body of Mcm2-7 docks onto ORC-Cdc6, and the origin DNA is bent and positioned adjacent to the open DNA entry gate, poised for insertion, at the Mcm2-Mcm5 interface. We used molecular simulations to reveal the dynamic transition from preloading conformers to the loaded conformers in which the loading of Mcm2-7 on DNA is complete and the DNA entry gate is fully closed. Our work provides multiple molecular insights into a key event of eukaryotic DNA replication. | ||||||||||||||||||
履歴 |
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-構造の表示
ムービー |
ムービービューア |
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構造ビューア | EMマップ: SurfViewMolmilJmol/JSmol |
添付画像 |
-ダウンロードとリンク
-EMDBアーカイブ
マップデータ | emd_21666.map.gz | 54.2 MB | EMDBマップデータ形式 | |
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ヘッダ (付随情報) | emd-21666-v30.xml emd-21666.xml | 37.6 KB 37.6 KB | 表示 表示 | EMDBヘッダ |
画像 | emd_21666.png | 149.6 KB | ||
Filedesc metadata | emd-21666.cif.gz | 13 KB | ||
アーカイブディレクトリ | http://ftp.pdbj.org/pub/emdb/structures/EMD-21666 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-21666 | HTTPS FTP |
-検証レポート
文書・要旨 | emd_21666_validation.pdf.gz | 372.3 KB | 表示 | EMDB検証レポート |
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文書・詳細版 | emd_21666_full_validation.pdf.gz | 371.8 KB | 表示 | |
XML形式データ | emd_21666_validation.xml.gz | 6.2 KB | 表示 | |
CIF形式データ | emd_21666_validation.cif.gz | 7 KB | 表示 | |
アーカイブディレクトリ | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-21666 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-21666 | HTTPS FTP |
-関連構造データ
-リンク
EMDBのページ | EMDB (EBI/PDBe) / EMDataResource |
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「今月の分子」の関連する項目 |
-マップ
ファイル | ダウンロード / ファイル: emd_21666.map.gz / 形式: CCP4 / 大きさ: 64 MB / タイプ: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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ボクセルのサイズ | X=Y=Z: 1.31 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
密度 |
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対称性 | 空間群: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
詳細 | EMDB XML:
CCP4マップ ヘッダ情報:
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-添付データ
-試料の構成要素
+全体 : ORC-Cdc6-Mcm-Cdt1-dsDNA
+超分子 #1: ORC-Cdc6-Mcm-Cdt1-dsDNA
+分子 #1: DNA (34-MER)
+分子 #2: DNA (34-MER)
+分子 #3: Cell division control protein 6
+分子 #4: Origin recognition complex subunit 1
+分子 #5: Origin recognition complex subunit 2
+分子 #6: Origin recognition complex subunit 3
+分子 #7: Origin recognition complex subunit 5
+分子 #8: Origin recognition complex subunit 4
+分子 #9: Origin recognition complex subunit 6
+分子 #10: DNA replication licensing factor MCM2
+分子 #11: DNA replication licensing factor MCM3
+分子 #12: DNA replication licensing factor MCM4
+分子 #13: Minichromosome maintenance protein 5
+分子 #14: Cell division cycle protein CDT1
+分子 #15: DNA replication licensing factor MCM6
+分子 #16: DNA replication licensing factor MCM7
+分子 #17: PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER
+分子 #18: MAGNESIUM ION
-実験情報
-構造解析
手法 | クライオ電子顕微鏡法 |
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解析 | 単粒子再構成法 |
試料の集合状態 | particle |
-試料調製
緩衝液 | pH: 7.5 |
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グリッド | 詳細: unspecified |
凍結 | 凍結剤: ETHANE |
-電子顕微鏡法
顕微鏡 | FEI TITAN KRIOS |
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撮影 | フィルム・検出器のモデル: GATAN K2 SUMMIT (4k x 4k) 平均電子線量: 50.0 e/Å2 |
電子線 | 加速電圧: 300 kV / 電子線源: FIELD EMISSION GUN |
電子光学系 | 照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELD |
実験機器 | モデル: Titan Krios / 画像提供: FEI Company |
-画像解析
初期モデル | モデルのタイプ: PDB ENTRY |
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最終 再構成 | 想定した対称性 - 点群: C1 (非対称) / 解像度のタイプ: BY AUTHOR / 解像度: 10.0 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 使用した粒子像数: 27830 |
初期 角度割当 | タイプ: PROJECTION MATCHING |
最終 角度割当 | タイプ: PROJECTION MATCHING |
-原子モデル構築 1
精密化 | プロトコル: RIGID BODY FIT |
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得られたモデル | PDB-6wgi: |