+データを開く
-基本情報
登録情報 | データベース: EMDB / ID: EMD-13211 | |||||||||||||||
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タイトル | Structure of a DNA-loaded MCM double hexamer engaged with the Dbf4-dependent kinase | |||||||||||||||
マップデータ | Auto-refined structure of the DNA-bound MCM double hexamer engaged with DDK | |||||||||||||||
試料 |
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機能・相同性 | 機能・相同性情報 positive regulation of spindle attachment to meiosis I kinetochore / positive regulation of meiotic DNA double-strand break formation involved in reciprocal meiotic recombination / positive regulation of DNA replication initiation / positive regulation of kinetochore assembly / positive regulation of meiotic DNA double-strand break formation / negative regulation of exit from mitosis / Dbf4-dependent protein kinase complex / positive regulation of meiosis I / regulation of cell cycle phase transition / positive regulation of nuclear cell cycle DNA replication ...positive regulation of spindle attachment to meiosis I kinetochore / positive regulation of meiotic DNA double-strand break formation involved in reciprocal meiotic recombination / positive regulation of DNA replication initiation / positive regulation of kinetochore assembly / positive regulation of meiotic DNA double-strand break formation / negative regulation of exit from mitosis / Dbf4-dependent protein kinase complex / positive regulation of meiosis I / regulation of cell cycle phase transition / positive regulation of nuclear cell cycle DNA replication / mitotic DNA damage checkpoint signaling / MCM core complex / Assembly of the pre-replicative complex / Switching of origins to a post-replicative state / MCM complex binding / nuclear DNA replication / mitotic DNA replication preinitiation complex assembly / 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 / single-stranded 3'-5' DNA helicase activity / nuclear pre-replicative complex / MCM complex / Activation of ATR in response to replication stress / DNA replication preinitiation complex / protein-containing complex localization / replication fork protection complex / mitotic DNA replication checkpoint signaling / mitotic DNA replication initiation / double-strand break repair via break-induced replication / silent mating-type cassette heterochromatin formation / regulation of DNA-templated DNA replication initiation / single-stranded DNA helicase activity / DNA strand elongation involved in DNA replication / DNA unwinding involved in DNA replication / nuclear replication fork / 3'-5' DNA helicase activity / DNA replication origin binding / chromosome, centromeric region / subtelomeric heterochromatin formation / DNA replication initiation / heterochromatin formation / protein serine/threonine kinase activator activity / helicase activity / chromosome segregation / single-stranded DNA binding / DNA helicase / chromosome, telomeric region / non-specific serine/threonine protein kinase / positive regulation of protein phosphorylation / phosphorylation / cell division / protein serine kinase activity / protein serine/threonine kinase activity / centrosome / DNA damage response / chromatin binding / chromatin / signal transduction / ATP hydrolysis activity / zinc ion binding / nucleoplasm / ATP binding / identical protein binding / nucleus / metal ion binding / cytoplasm 類似検索 - 分子機能 | |||||||||||||||
生物種 | Saccharomyces cerevisiae S288C (パン酵母) / Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (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: 3.3 Å | |||||||||||||||
データ登録者 | Greiwe JF / Locke J / Nans A / Costa A | |||||||||||||||
資金援助 | European Union, 4件
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引用 | ジャーナル: Nat Struct Mol Biol / 年: 2022 タイトル: Structural mechanism for the selective phosphorylation of DNA-loaded MCM double hexamers by the Dbf4-dependent kinase. 著者: Julia F Greiwe / Thomas C R Miller / Julia Locke / Fabrizio Martino / Steven Howell / Anne Schreiber / Andrea Nans / John F X Diffley / Alessandro Costa / 要旨: Loading of the eukaryotic replicative helicase onto replication origins involves two MCM hexamers forming a double hexamer (DH) around duplex DNA. During S phase, helicase activation requires MCM ...Loading of the eukaryotic replicative helicase onto replication origins involves two MCM hexamers forming a double hexamer (DH) around duplex DNA. During S phase, helicase activation requires MCM phosphorylation by Dbf4-dependent kinase (DDK), comprising Cdc7 and Dbf4. DDK selectively phosphorylates loaded DHs, but how such fidelity is achieved is unknown. Here, we determine the cryogenic electron microscopy structure of Saccharomyces cerevisiae DDK in the act of phosphorylating a DH. DDK docks onto one MCM ring and phosphorylates the opposed ring. Truncation of the Dbf4 docking domain abrogates DH phosphorylation, yet Cdc7 kinase activity is unaffected. Late origin firing is blocked in response to DNA damage via Dbf4 phosphorylation by the Rad53 checkpoint kinase. DDK phosphorylation by Rad53 impairs DH phosphorylation by blockage of DDK binding to DHs, and also interferes with the Cdc7 active site. Our results explain the structural basis and regulation of the selective phosphorylation of DNA-loaded MCM DHs, which supports bidirectional 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_13211.map.gz | 338 MB | EMDBマップデータ形式 | |
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ヘッダ (付随情報) | emd-13211-v30.xml emd-13211.xml | 39.8 KB 39.8 KB | 表示 表示 | EMDBヘッダ |
FSC (解像度算出) | emd_13211_fsc.xml | 16.9 KB | 表示 | FSCデータファイル |
画像 | emd_13211.png | 148.7 KB | ||
マスクデータ | emd_13211_msk_1.map | 421.9 MB | マスクマップ | |
その他 | emd_13211_additional_1.map.gz emd_13211_additional_2.map.gz emd_13211_half_map_1.map.gz emd_13211_half_map_2.map.gz | 28.9 MB 3.1 GB 338.6 MB 339.7 MB | ||
アーカイブディレクトリ | http://ftp.pdbj.org/pub/emdb/structures/EMD-13211 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-13211 | HTTPS FTP |
-検証レポート
文書・要旨 | emd_13211_validation.pdf.gz | 535.7 KB | 表示 | EMDB検証レポート |
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文書・詳細版 | emd_13211_full_validation.pdf.gz | 535.3 KB | 表示 | |
XML形式データ | emd_13211_validation.xml.gz | 24.9 KB | 表示 | |
CIF形式データ | emd_13211_validation.cif.gz | 32.8 KB | 表示 | |
アーカイブディレクトリ | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-13211 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-13211 | 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_13211.map.gz / 形式: CCP4 / 大きさ: 421.9 MB / タイプ: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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注釈 | Auto-refined structure of the DNA-bound MCM double hexamer engaged with DDK | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 1.08 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
-マスク #1
ファイル | emd_13211_msk_1.map | ||||||||||||
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投影像・断面図 |
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密度ヒストグラム |
-追加マップ: Sharpened map
ファイル | emd_13211_additional_1.map | ||||||||||||
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注釈 | Sharpened map | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-追加マップ: LAFTER filtered map, which was used for docking...
ファイル | emd_13211_additional_2.map | ||||||||||||
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注釈 | LAFTER filtered map, which was used for docking of the Dbf4 BRCT domain. | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-ハーフマップ: #1
ファイル | emd_13211_half_map_1.map | ||||||||||||
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投影像・断面図 |
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密度ヒストグラム |
-ハーフマップ: #2
ファイル | emd_13211_half_map_2.map | ||||||||||||
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投影像・断面図 |
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密度ヒストグラム |
-試料の構成要素
+全体 : DNA-loaded MCM double hexamer engaged with the dimeric Dbf4-depen...
+超分子 #1: DNA-loaded MCM double hexamer engaged with the dimeric Dbf4-depen...
+分子 #1: DNA replication licensing factor MCM2
+分子 #2: DNA replication licensing factor MCM3
+分子 #3: DNA replication licensing factor MCM4
+分子 #4: Minichromosome maintenance protein 5
+分子 #5: DNA replication licensing factor MCM6
+分子 #6: DNA replication licensing factor MCM7
+分子 #7: Cell division control protein 7
+分子 #8: DDK kinase regulatory subunit DBF4
+分子 #9: DNA (53-MER)
+分子 #10: DNA (53-MER)
+分子 #11: ADENOSINE-5'-TRIPHOSPHATE
+分子 #12: MAGNESIUM ION
+分子 #13: ZINC ION
+分子 #14: ADENOSINE-5'-DIPHOSPHATE
-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: 7.6 |
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凍結 | 凍結剤: ETHANE / チャンバー内湿度: 90 % / チャンバー内温度: 295 K / 装置: FEI VITROBOT MARK IV / 詳細: blotted for 3 seconds before plunging. |
詳細 | The entire MCM loading and phosphorylation reaction was applied to the EM grid. |
-電子顕微鏡法
顕微鏡 | FEI TITAN KRIOS |
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特殊光学系 | エネルギーフィルター - 名称: GIF Bioquantum / エネルギーフィルター - スリット幅: 20 eV |
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: 1 / 実像数: 18135 / 平均露光時間: 9.0 sec. / 平均電子線量: 51.3 e/Å2 |
電子線 | 加速電圧: 300 kV / 電子線源: FIELD EMISSION GUN |
電子光学系 | C2レンズ絞り径: 50.0 µm / 照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELD / 最大 デフォーカス(公称値): 4.1 µm / 最小 デフォーカス(公称値): 2.0 µm / 倍率(公称値): 130000 |
試料ステージ | 試料ホルダーモデル: FEI TITAN KRIOS AUTOGRID HOLDER |
実験機器 | モデル: Titan Krios / 画像提供: FEI Company |