+データを開く
-基本情報
登録情報 | データベース: PDB / ID: 8rk2 | |||||||||||||||||||||||||||
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タイトル | Human Replication protein A (RPA; trimeric core) - ssDNA complex | |||||||||||||||||||||||||||
要素 |
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キーワード | DNA BINDING PROTEIN (DNA結合タンパク質) / ssDNA binding protein / DNA damage repair (DNA修復) / Single-strand annealing | |||||||||||||||||||||||||||
機能・相同性 | 機能・相同性情報 protein localization to chromosome / DNA replication factor A complex / chromatin-protein adaptor activity / protein localization to site of double-strand break / regulation of DNA damage checkpoint / single-stranded telomeric DNA binding / Removal of the Flap Intermediate / Mismatch repair (MMR) directed by MSH2:MSH3 (MutSbeta) / Mismatch repair (MMR) directed by MSH2:MSH6 (MutSalpha) / Removal of the Flap Intermediate from the C-strand ...protein localization to chromosome / DNA replication factor A complex / chromatin-protein adaptor activity / protein localization to site of double-strand break / regulation of DNA damage checkpoint / single-stranded telomeric DNA binding / Removal of the Flap Intermediate / Mismatch repair (MMR) directed by MSH2:MSH3 (MutSbeta) / Mismatch repair (MMR) directed by MSH2:MSH6 (MutSalpha) / Removal of the Flap Intermediate from the C-strand / G-rich strand telomeric DNA binding / HDR through Single Strand Annealing (SSA) / Impaired BRCA2 binding to RAD51 / regulation of double-strand break repair via homologous recombination / telomeric DNA binding / site of DNA damage / Presynaptic phase of homologous DNA pairing and strand exchange / telomere maintenance via telomerase / Activation of the pre-replicative complex / PCNA-Dependent Long Patch Base Excision Repair / Regulation of HSF1-mediated heat shock response / HSF1 activation / DNAミスマッチ修復 / Activation of ATR in response to replication stress / SUMOylation of DNA damage response and repair proteins / mitotic G1 DNA damage checkpoint signaling / regulation of mitotic cell cycle / telomere maintenance / Translesion synthesis by REV1 / Translesion synthesis by POLK / Translesion synthesis by POLI / Gap-filling DNA repair synthesis and ligation in GG-NER / meiotic cell cycle / nucleotide-excision repair / Fanconi Anemia Pathway / Recognition of DNA damage by PCNA-containing replication complex / Termination of translesion DNA synthesis / double-strand break repair via homologous recombination / Translesion Synthesis by POLH / 塩基除去修復 / HDR through Homologous Recombination (HRR) / G2/M DNA damage checkpoint / Dual Incision in GG-NER / DNA-templated DNA replication / PML body / 遺伝的組換え / Formation of Incision Complex in GG-NER / Dual incision in TC-NER / Gap-filling DNA repair synthesis and ligation in TC-NER / site of double-strand break / single-stranded DNA binding / regulation of cell population proliferation / Processing of DNA double-strand break ends / protein phosphatase binding / DNA recombination / Regulation of TP53 Activity through Phosphorylation / DNA複製 / damaged DNA binding / chromosome, telomeric region / nuclear body / DNA修復 / DNA damage response / ubiquitin protein ligase binding / クロマチン / enzyme binding / 核質 / metal ion binding / 細胞核 類似検索 - 分子機能 | |||||||||||||||||||||||||||
生物種 | Homo sapiens (ヒト) | |||||||||||||||||||||||||||
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: 3.2 Å | |||||||||||||||||||||||||||
データ登録者 | Liang, C.C. / West, S.C. | |||||||||||||||||||||||||||
資金援助 | 英国, European Union, スイス, 8件
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引用 | ジャーナル: Nature / 年: 2024 タイトル: Mechanism of single-stranded DNA annealing by RAD52-RPA complex. 著者: Chih-Chao Liang / Luke A Greenhough / Laura Masino / Sarah Maslen / Ilirjana Bajrami / Marcel Tuppi / Mark Skehel / Ian A Taylor / Stephen C West / 要旨: RAD52 is important for the repair of DNA double-stranded breaks, mitotic DNA synthesis and alternative telomere length maintenance. Central to these functions, RAD52 promotes the annealing of ...RAD52 is important for the repair of DNA double-stranded breaks, mitotic DNA synthesis and alternative telomere length maintenance. Central to these functions, RAD52 promotes the annealing of complementary single-stranded DNA (ssDNA) and provides an alternative to BRCA2/RAD51-dependent homologous recombination repair. Inactivation of RAD52 in homologous-recombination-deficient BRCA1- or BRCA2-defective cells is synthetically lethal, and aberrant expression of RAD52 is associated with poor cancer prognosis. As a consequence, RAD52 is an attractive therapeutic target against homologous-recombination-deficient breast, ovarian and prostate cancers. Here we describe the structure of RAD52 and define the mechanism of annealing. As reported previously, RAD52 forms undecameric (11-subunit) ring structures, but these rings do not represent the active form of the enzyme. Instead, cryo-electron microscopy and biochemical analyses revealed that ssDNA annealing is driven by RAD52 open rings in association with replication protein-A (RPA). Atomic models of the RAD52-ssDNA complex show that ssDNA sits in a positively charged channel around the ring. Annealing is driven by the RAD52 N-terminal domains, whereas the C-terminal regions modulate the open-ring conformation and RPA interaction. RPA associates with RAD52 at the site of ring opening with critical interactions occurring between the RPA-interacting domain of RAD52 and the winged helix domain of RPA2. Our studies provide structural snapshots throughout the annealing process and define the molecular mechanism of ssDNA annealing by the RAD52-RPA complex. | |||||||||||||||||||||||||||
履歴 |
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-構造の表示
構造ビューア | 分子: MolmilJmol/JSmol |
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-ダウンロードとリンク
-ダウンロード
PDBx/mmCIF形式 | 8rk2.cif.gz | 213.5 KB | 表示 | PDBx/mmCIF形式 |
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PDB形式 | pdb8rk2.ent.gz | 133.9 KB | 表示 | PDB形式 |
PDBx/mmJSON形式 | 8rk2.json.gz | ツリー表示 | PDBx/mmJSON形式 | |
その他 | その他のダウンロード |
-検証レポート
アーカイブディレクトリ | https://data.pdbj.org/pub/pdb/validation_reports/rk/8rk2 ftp://data.pdbj.org/pub/pdb/validation_reports/rk/8rk2 | HTTPS FTP |
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-関連構造データ
-リンク
-集合体
登録構造単位 |
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-要素
#1: タンパク質 | 分子量: 68212.977 Da / 分子数: 1 / 由来タイプ: 組換発現 / 由来: (組換発現) Homo sapiens (ヒト) / 遺伝子: RPA1, REPA1, RPA70 / 細胞株 (発現宿主): Sf9 発現宿主: Spodoptera frugiperda (ツマジロクサヨトウ) 参照: UniProt: P27694 |
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#2: タンパク質 | 分子量: 29276.795 Da / 分子数: 1 / 由来タイプ: 組換発現 / 由来: (組換発現) Homo sapiens (ヒト) / 遺伝子: RPA2, REPA2, RPA32, RPA34 / 細胞株 (発現宿主): Sf9 発現宿主: Spodoptera frugiperda (ツマジロクサヨトウ) 参照: UniProt: P15927 |
#3: タンパク質 | 分子量: 13583.714 Da / 分子数: 1 / 由来タイプ: 組換発現 / 由来: (組換発現) Homo sapiens (ヒト) / 遺伝子: RPA3, REPA3, RPA14 / 細胞株 (発現宿主): Sf9 発現宿主: Spodoptera frugiperda (ツマジロクサヨトウ) 参照: UniProt: P35244 |
-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 |
-試料調製
構成要素 | 名称: Open ring conformation of human RAD52 in complex with ssDNA タイプ: COMPLEX 詳細: Recombinant RAD52 purified from E. coli, and the open ring conformation was separated by cation exchange. The RAD52-ssDNA complex was reconstituted in vitro. Entity ID: all / 由来: RECOMBINANT | ||||||||||||||||||||||||||||||
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分子量 | 実験値: NO | ||||||||||||||||||||||||||||||
由来(天然) | 生物種: Homo sapiens (ヒト) | ||||||||||||||||||||||||||||||
由来(組換発現) | 生物種: Escherichia coli (大腸菌) / 株: BL21 (DE3) | ||||||||||||||||||||||||||||||
緩衝液 | pH: 8 | ||||||||||||||||||||||||||||||
緩衝液成分 |
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試料 | 濃度: 0.15 mg/ml / 包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES | ||||||||||||||||||||||||||||||
試料支持 | グリッドの材料: GOLD / グリッドのサイズ: 200 divisions/in. / グリッドのタイプ: UltrAuFoil R2/2 | ||||||||||||||||||||||||||||||
急速凍結 | 装置: FEI VITROBOT MARK IV / 凍結剤: ETHANE / 湿度: 95 % / 凍結前の試料温度: 277.15 K |
-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: FIELD EMISSION GUN / 加速電圧: 300 kV / 照射モード: SPOT SCAN |
電子レンズ | モード: BRIGHT FIELDBright-field microscopy / 倍率(公称値): 130000 X / 最大 デフォーカス(公称値): 3500 nm / 最小 デフォーカス(公称値): 1500 nm / Cs: 2.7 mm / C2レンズ絞り径: 50 µm / アライメント法: COMA FREE |
試料ホルダ | 凍結剤: NITROGEN 試料ホルダーモデル: FEI TITAN KRIOS AUTOGRID HOLDER |
撮影 | 電子線照射量: 46 e/Å2 / 検出モード: COUNTING フィルム・検出器のモデル: GATAN K2 QUANTUM (4k x 4k) |
-解析
EMソフトウェア |
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CTF補正 | タイプ: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||
粒子像の選択 | 選択した粒子像数: 21697126 | ||||||||||||||||||||||||||||||
3次元再構成 | 解像度: 3.2 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 1316062 / 対称性のタイプ: POINT | ||||||||||||||||||||||||||||||
原子モデル構築 | プロトコル: RIGID BODY FIT / 空間: REAL | ||||||||||||||||||||||||||||||
原子モデル構築 | Source name: AlphaFold / タイプ: in silico model | ||||||||||||||||||||||||||||||
精密化 | 交差検証法: NONE 立体化学のターゲット値: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||||||||
原子変位パラメータ | Biso mean: 138.23 Å2 | ||||||||||||||||||||||||||||||
拘束条件 |
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