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データを開く
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基本情報
登録情報 | データベース: PDB / ID: 6wgc | ||||||||||||||||||
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タイトル | Atomic model of semi-attached mutant OCCM-DNA complex (ORC-Cdc6-Cdt1-Mcm2-7 with Mcm6 WHD truncation) | ||||||||||||||||||
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![]() | REPLICATION/DNA / DNA replication / Cryo-EM / OCCM-deltaC6 / REPLICATION-DNA complex | ||||||||||||||||||
機能・相同性 | ![]() 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 / 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 / helicase activity / G1/S transition of mitotic cell cycle / chromosome / single-stranded DNA binding / DNA helicase / chromosome, telomeric region / cell division / GTPase activity / chromatin binding / GTP binding / ATP hydrolysis activity / nucleoplasm / ATP binding / nucleus / metal ion binding / cytoplasm 類似検索 - 分子機能 | ||||||||||||||||||
生物種 | ![]() ![]() | ||||||||||||||||||
手法 | 電子顕微鏡法 / 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 4.3 Å | ||||||||||||||||||
![]() | Yuan, Z. / Schneider, S. / Dodd, T. / Riera, A. / Bai, L. / Yan, C. / Magdalou, I. / Ivanov, I. / Stillman, B. / Li, H. / Speck, C. | ||||||||||||||||||
資金援助 | ![]() ![]()
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![]() | ![]() 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 |
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構造ビューア | 分子: ![]() ![]() |
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PDBx/mmCIF形式 | ![]() | 619.4 KB | 表示 | ![]() |
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PDB形式 | ![]() | 459.8 KB | 表示 | ![]() |
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その他 | ![]() |
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アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
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-関連構造データ
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リンク
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集合体
登録構造単位 | ![]()
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要素
-Origin recognition complex subunit ... , 6種, 6分子 ABCEDF
#2: タンパク質 | 分子量: 104433.008 Da / 分子数: 1 / 由来タイプ: 組換発現 由来: (組換発現) ![]() ![]() 遺伝子: ORC1, YML065W / 発現宿主: ![]() ![]() |
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#3: タンパク質 | 分子量: 71342.180 Da / 分子数: 1 / 由来タイプ: 組換発現 由来: (組換発現) ![]() ![]() 遺伝子: ORC2, RRR1, SIR5, YBR060C, YBR0523 / 発現宿主: ![]() ![]() |
#4: タンパク質 | 分子量: 72161.766 Da / 分子数: 1 / 由来タイプ: 組換発現 由来: (組換発現) ![]() ![]() 遺伝子: ORC3, OAF1, OIF1, YLL004W, L1365 / 発現宿主: ![]() ![]() |
#5: タンパク質 | 分子量: 55347.168 Da / 分子数: 1 / 由来タイプ: 組換発現 由来: (組換発現) ![]() ![]() 遺伝子: ORC5, YNL261W, N0834 / 発現宿主: ![]() ![]() |
#6: タンパク質 | 分子量: 60772.152 Da / 分子数: 1 / 由来タイプ: 組換発現 由来: (組換発現) ![]() ![]() 遺伝子: ORC4, YPR162C, P9325.5 / 発現宿主: ![]() ![]() |
#7: タンパク質 | 分子量: 50369.531 Da / 分子数: 1 / 由来タイプ: 組換発現 由来: (組換発現) ![]() ![]() 遺伝子: ORC6, AAP1, YHR118C / 発現宿主: ![]() ![]() |
-DNA replication licensing factor ... , 2種, 2分子 37
#8: タンパク質 | 分子量: 107653.508 Da / 分子数: 1 / 由来タイプ: 組換発現 由来: (組換発現) ![]() ![]() 遺伝子: MCM3, YEL032W, SYGP-ORF23 / 発現宿主: ![]() ![]() |
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#9: タンパク質 | 分子量: 95049.875 Da / 分子数: 1 / 由来タイプ: 組換発現 由来: (組換発現) ![]() ![]() 遺伝子: MCM7, CDC47, YBR202W, YBR1441 / 発現宿主: ![]() ![]() |
-DNA鎖 , 2種, 2分子 GH
#10: DNA鎖 | 分子量: 12577.062 Da / 分子数: 1 / 由来タイプ: 組換発現 由来: (組換発現) ![]() ![]() 発現宿主: ![]() ![]() |
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#11: DNA鎖 | 分子量: 12655.295 Da / 分子数: 1 / 由来タイプ: 組換発現 由来: (組換発現) ![]() ![]() 発現宿主: ![]() ![]() |
-タンパク質 / 非ポリマー , 2種, 5分子 9![](data/chem/img/AGS.gif)
![](data/chem/img/AGS.gif)
#12: 化合物 | ChemComp-AGS / #1: タンパク質 | | 分子量: 58112.367 Da / 分子数: 1 / 由来タイプ: 組換発現 由来: (組換発現) ![]() ![]() 遺伝子: CDC6, YJL194W, J0347 / 発現宿主: ![]() ![]() |
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-詳細
研究の焦点であるリガンドがあるか | Y |
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-実験情報
-実験
実験 | 手法: 電子顕微鏡法 |
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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: ORC-Cdc6-Mcm3-Mcm7-dsDNA / タイプ: COMPLEX / Entity ID: #1-#11 / 由来: RECOMBINANT |
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由来(天然) | 生物種: ![]() ![]() |
由来(組換発現) | 生物種: ![]() ![]() |
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 |
試料支持 | 詳細: unspecified |
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 |
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顕微鏡 | モデル: FEI TITAN KRIOS |
電子銃 | 電子線源: ![]() |
電子レンズ | モード: BRIGHT FIELD |
撮影 | 電子線照射量: 50 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
CTF補正 | タイプ: PHASE FLIPPING AND AMPLITUDE CORRECTION |
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対称性 | 点対称性: C1 (非対称) |
3次元再構成 | 解像度: 4.3 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 86074 / 対称性のタイプ: POINT |
原子モデル構築 | プロトコル: RIGID BODY FIT |