+データを開く
-基本情報
登録情報 | データベース: EMDB / ID: EMD-42386 | |||||||||
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タイトル | Cryo-EM map of the human CTF18-RFC-PCNA-DNA ternary complex in the five-subunit binding state (state 4) | |||||||||
マップデータ | EM sharpen map | |||||||||
試料 |
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キーワード | DNA clamp loader complex / REPLICATION | |||||||||
機能・相同性 | 機能・相同性情報 positive regulation of deoxyribonuclease activity / dinucleotide insertion or deletion binding / PCNA-p21 complex / response to organophosphorus / Ctf18 RFC-like complex / mitotic telomere maintenance via semi-conservative replication / Rad17 RFC-like complex / DNA replication factor C complex / Elg1 RFC-like complex / purine-specific mismatch base pair DNA N-glycosylase activity ...positive regulation of deoxyribonuclease activity / dinucleotide insertion or deletion binding / PCNA-p21 complex / response to organophosphorus / Ctf18 RFC-like complex / mitotic telomere maintenance via semi-conservative replication / Rad17 RFC-like complex / DNA replication factor C complex / Elg1 RFC-like complex / purine-specific mismatch base pair DNA N-glycosylase activity / nuclear lamina / MutLalpha complex binding / positive regulation of DNA-directed DNA polymerase activity / Polymerase switching / Telomere C-strand (Lagging Strand) Synthesis / Processive synthesis on the lagging strand / DNA clamp loader activity / PCNA complex / Removal of the Flap Intermediate / Processive synthesis on the C-strand of the telomere / Mismatch repair (MMR) directed by MSH2:MSH3 (MutSbeta) / Polymerase switching on the C-strand of the telomere / Mismatch repair (MMR) directed by MSH2:MSH6 (MutSalpha) / Transcription of E2F targets under negative control by DREAM complex / Removal of the Flap Intermediate from the C-strand / replisome / DNA duplex unwinding / single-stranded DNA helicase activity / response to L-glutamate / HDR through Single Strand Annealing (SSA) / Impaired BRCA2 binding to RAD51 / DNA strand elongation involved in DNA replication / histone acetyltransferase binding / DNA synthesis involved in DNA repair / DNA polymerase processivity factor activity / G1/S-Specific Transcription / leading strand elongation / response to dexamethasone / replication fork processing / nuclear replication fork / Presynaptic phase of homologous DNA pairing and strand exchange / SUMOylation of DNA replication proteins / estrous cycle / PCNA-Dependent Long Patch Base Excision Repair / ATP-dependent activity, acting on DNA / mismatch repair / Activation of ATR in response to replication stress / translesion synthesis / cyclin-dependent protein kinase holoenzyme complex / response to cadmium ion / DNA polymerase binding / epithelial cell differentiation / positive regulation of DNA repair / Translesion synthesis by REV1 / Translesion synthesis by POLK / Translesion synthesis by POLI / Gap-filling DNA repair synthesis and ligation in GG-NER / base-excision repair, gap-filling / TP53 Regulates Transcription of Genes Involved in G2 Cell Cycle Arrest / positive regulation of DNA replication / replication fork / male germ cell nucleus / liver regeneration / nuclear estrogen receptor binding / Recognition of DNA damage by PCNA-containing replication complex / Termination of translesion DNA synthesis / Translesion Synthesis by POLH / HDR through Homologous Recombination (HRR) / G2/M DNA damage checkpoint / Dual Incision in GG-NER / DNA-templated DNA replication / receptor tyrosine kinase binding / cellular response to hydrogen peroxide / Dual incision in TC-NER / Gap-filling DNA repair synthesis and ligation in TC-NER / cellular response to UV / cellular response to xenobiotic stimulus / E3 ubiquitin ligases ubiquitinate target proteins / response to estradiol / heart development / Processing of DNA double-strand break ends / DNA replication / Regulation of TP53 Activity through Phosphorylation / chromosome, telomeric region / damaged DNA binding / nuclear body / cell cycle / DNA repair / centrosome / chromatin binding / protein-containing complex binding / chromatin / enzyme binding / negative regulation of transcription by RNA polymerase II / ATP hydrolysis activity / DNA binding / extracellular exosome / nucleoplasm / ATP binding / identical protein binding 類似検索 - 分子機能 | |||||||||
生物種 | Homo sapiens (ヒト) / synthetic construct (人工物) | |||||||||
手法 | 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 2.93 Å | |||||||||
データ登録者 | Wang F / He Q / Li H | |||||||||
資金援助 | 米国, 1件
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引用 | ジャーナル: Proc Natl Acad Sci U S A / 年: 2024 タイトル: Cryo-EM reveals a nearly complete PCNA loading process and unique features of the human alternative clamp loader CTF18-RFC. 著者: Qing He / Feng Wang / Michael E O'Donnell / Huilin Li / 要旨: The DNA sliding clamp PCNA is a multipurpose platform for DNA polymerases and many other proteins involved in DNA metabolism. The topologically closed PCNA ring needs to be cracked open and loaded ...The DNA sliding clamp PCNA is a multipurpose platform for DNA polymerases and many other proteins involved in DNA metabolism. The topologically closed PCNA ring needs to be cracked open and loaded onto DNA by a clamp loader, e.g., the well-studied pentameric ATPase complex RFC (RFC1-5). The CTF18-RFC complex is an alternative clamp loader found recently to bind the leading strand DNA polymerase ε and load PCNA onto leading strand DNA, but its structure and the loading mechanism have been unknown. By cryo-EM analysis of in vitro assembled human CTF18-RFC-DNA-PCNA complex, we have captured seven loading intermediates, revealing a detailed PCNA loading mechanism onto a 3'-ss/dsDNA junction by CTF18-RFC. Interestingly, the alternative loader has evolved a highly mobile CTF18 AAA+ module likely to lower the loading activity, perhaps to avoid competition with the RFC and to limit its role to leading strand clamp loading. To compensate for the lost stability due to the mobile AAA+ module, CTF18 has evolved a unique β-hairpin motif that reaches across RFC2 to interact with RFC5, thereby stabilizing the pentameric complex. Further, we found that CTF18 also contains a separation pin to locally melt DNA from the 3'-end of the primer; this ensures its ability to load PCNA to any 3'-ss/dsDNA junction, facilitated by the binding energy of the E-plug to the major groove. Our study reveals unique structural features of the human CTF18-RFC and contributes to a broader understanding of PCNA loading by the alternative clamp loaders. | |||||||||
履歴 |
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-構造の表示
添付画像 |
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-ダウンロードとリンク
-EMDBアーカイブ
マップデータ | emd_42386.map.gz | 118 MB | EMDBマップデータ形式 | |
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ヘッダ (付随情報) | emd-42386-v30.xml emd-42386.xml | 28.7 KB 28.7 KB | 表示 表示 | EMDBヘッダ |
FSC (解像度算出) | emd_42386_fsc.xml | 10.6 KB | 表示 | FSCデータファイル |
画像 | emd_42386.png | 75.8 KB | ||
Filedesc metadata | emd-42386.cif.gz | 8.1 KB | ||
その他 | emd_42386_additional_1.map.gz emd_42386_additional_2.map.gz emd_42386_half_map_1.map.gz emd_42386_half_map_2.map.gz | 62.6 MB 110.9 MB 115.9 MB 115.9 MB | ||
アーカイブディレクトリ | http://ftp.pdbj.org/pub/emdb/structures/EMD-42386 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-42386 | HTTPS FTP |
-検証レポート
文書・要旨 | emd_42386_validation.pdf.gz | 1015 KB | 表示 | EMDB検証レポート |
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文書・詳細版 | emd_42386_full_validation.pdf.gz | 1014.5 KB | 表示 | |
XML形式データ | emd_42386_validation.xml.gz | 19 KB | 表示 | |
CIF形式データ | emd_42386_validation.cif.gz | 24.5 KB | 表示 | |
アーカイブディレクトリ | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-42386 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-42386 | HTTPS FTP |
-関連構造データ
関連構造データ | 8umwMC 8umtC 8umuC 8umvC 8umyC 8un0C 8unjC M: このマップから作成された原子モデル C: 同じ文献を引用 (文献) |
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類似構造データ | 類似検索 - 機能・相同性F&H 検索 |
-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_42386.map.gz / 形式: CCP4 / 大きさ: 125 MB / タイプ: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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注釈 | EM sharpen map | ||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 0.828 Å | ||||||||||||||||||||
密度 |
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対称性 | 空間群: 1 | ||||||||||||||||||||
詳細 | EMDB XML:
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-添付データ
-追加マップ: Original unsharpen EM map
ファイル | emd_42386_additional_1.map | ||||||||||||
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注釈 | Original unsharpen EM map | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-追加マップ: EM sharpen map by DeepEM hancer
ファイル | emd_42386_additional_2.map | ||||||||||||
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注釈 | EM sharpen map by DeepEM hancer | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-ハーフマップ: EM half map B
ファイル | emd_42386_half_map_1.map | ||||||||||||
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注釈 | EM half map B | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-ハーフマップ: EM half map A
ファイル | emd_42386_half_map_2.map | ||||||||||||
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注釈 | EM half map A | ||||||||||||
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 : the human clamp-clamp loader complex PCNA-CTF18
+超分子 #1: the human clamp-clamp loader complex PCNA-CTF18
+分子 #1: Chromosome transmission fidelity protein 18 homolog
+分子 #2: Replication factor C subunit 2
+分子 #3: Replication factor C subunit 5
+分子 #4: Replication factor C subunit 4
+分子 #5: Replication factor C subunit 3
+分子 #6: Proliferating cell nuclear antigen
+分子 #7: DNA (40-MER)
+分子 #8: DNA (20-MER)
+分子 #9: MAGNESIUM ION
+分子 #10: PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER
+分子 #11: 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
濃度 | 0.3 mg/mL |
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緩衝液 | pH: 7.5 |
凍結 | 凍結剤: ETHANE / チャンバー内湿度: 100 % / チャンバー内温度: 280 K / 装置: FEI VITROBOT MARK IV |
-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 K3 (6k x 4k) / 平均電子線量: 60.0 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 最大 デフォーカス(公称値): 1.9000000000000001 µm 最小 デフォーカス(公称値): 1.2 µm |
実験機器 | モデル: Titan Krios / 画像提供: FEI Company |
+画像解析
-原子モデル構築 1
精密化 | 空間: REAL / プロトコル: RIGID BODY FIT / 温度因子: 123 |
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得られたモデル | PDB-8umw: |