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基本情報
登録情報 | データベース: PDB / ID: 5flm | |||||||||
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タイトル | Structure of transcribing mammalian RNA polymerase II | |||||||||
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![]() | TRANSCRIPTION / ELONGATION | |||||||||
機能・相同性 | ![]() Formation of RNA Pol II elongation complex / Formation of the Early Elongation Complex / Transcriptional regulation by small RNAs / TP53 Regulates Transcription of DNA Repair Genes / FGFR2 alternative splicing / RNA polymerase II transcribes snRNA genes / mRNA Capping / mRNA Splicing - Minor Pathway / RNA Polymerase I Transcription Initiation / RNA Polymerase I Promoter Escape ...Formation of RNA Pol II elongation complex / Formation of the Early Elongation Complex / Transcriptional regulation by small RNAs / TP53 Regulates Transcription of DNA Repair Genes / FGFR2 alternative splicing / RNA polymerase II transcribes snRNA genes / mRNA Capping / mRNA Splicing - Minor Pathway / RNA Polymerase I Transcription Initiation / RNA Polymerase I Promoter Escape / RNA Polymerase II Promoter Escape / RNA Polymerase II Transcription Pre-Initiation And Promoter Opening / RNA Polymerase I Transcription Termination / RNA Polymerase II Transcription Initiation / RNA Polymerase II Transcription Elongation / RNA Polymerase II Transcription Initiation And Promoter Clearance / RNA Polymerase III Transcription Initiation From Type 1 Promoter / RNA Polymerase III Transcription Initiation From Type 2 Promoter / RNA Polymerase III Transcription Initiation From Type 3 Promoter / RNA Pol II CTD phosphorylation and interaction with CE / Estrogen-dependent gene expression / RNA Polymerase II Pre-transcription Events / Processing of Capped Intron-Containing Pre-mRNA / B-WICH complex positively regulates rRNA expression / mRNA Splicing - Major Pathway / Formation of TC-NER Pre-Incision Complex / Dual incision in TC-NER / Gap-filling DNA repair synthesis and ligation in TC-NER / DNA/RNA hybrid binding / maintenance of transcriptional fidelity during transcription elongation by RNA polymerase II / transcription by RNA polymerase I / positive regulation of nuclear-transcribed mRNA poly(A) tail shortening / nuclear-transcribed mRNA catabolic process / transcription by RNA polymerase III / RNA polymerase II activity / transcription elongation by RNA polymerase I / tRNA transcription by RNA polymerase III / RNA polymerase I complex / RNA polymerase III complex / positive regulation of translational initiation / transcription-coupled nucleotide-excision repair / RNA polymerase II, core complex / core promoter sequence-specific DNA binding / translation initiation factor binding / transcription initiation at RNA polymerase II promoter / protein-DNA complex / P-body / euchromatin / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / single-stranded DNA binding / 加水分解酵素; エステル加水分解酵素; 5'-リン酸モノエステル産生エキソリボヌクレアーゼ / transcription by RNA polymerase II / nucleic acid binding / chromosome, telomeric region / single-stranded RNA binding / protein dimerization activity / hydrolase activity / RNA-directed RNA polymerase / RNA-dependent RNA polymerase activity / nucleotide binding / chromatin binding / nucleolus / DNA binding / zinc ion binding / nucleus / metal ion binding / cytosol 類似検索 - 分子機能 | |||||||||
生物種 | ![]() ![]() SYNTHETIC CONSTRUCT (人工物) | |||||||||
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: 3.4 Å | |||||||||
![]() | Bernecky, C. / Herzog, F. / Baumeister, W. / Plitzko, J.M. / Cramer, P. | |||||||||
![]() | ![]() タイトル: Structure of transcribing mammalian RNA polymerase II. 著者: Carrie Bernecky / Franz Herzog / Wolfgang Baumeister / Jürgen M Plitzko / Patrick Cramer / ![]() 要旨: RNA polymerase (Pol) II produces messenger RNA during transcription of protein-coding genes in all eukaryotic cells. The Pol II structure is known at high resolution from X-ray crystallography for ...RNA polymerase (Pol) II produces messenger RNA during transcription of protein-coding genes in all eukaryotic cells. The Pol II structure is known at high resolution from X-ray crystallography for two yeast species. Structural studies of mammalian Pol II, however, remain limited to low-resolution electron microscopy analysis of human Pol II and its complexes with various proteins. Here we report the 3.4 Å resolution cryo-electron microscopy structure of mammalian Pol II in the form of a transcribing complex comprising DNA template and RNA transcript. We use bovine Pol II, which is identical to the human enzyme except for seven amino-acid residues. The obtained atomic model closely resembles its yeast counterpart, but also reveals unknown features. Binding of nucleic acids to the polymerase involves 'induced fit' of the mobile Pol II clamp and active centre region. DNA downstream of the transcription bubble contacts a conserved 'TPSA motif' in the jaw domain of the Pol II subunit RPB5, an interaction that is apparently already established during transcription initiation. Upstream DNA emanates from the active centre cleft at an angle of approximately 105° with respect to downstream DNA. This position of upstream DNA allows for binding of the general transcription elongation factor DSIF (SPT4-SPT5) that we localize over the active centre cleft in a conserved position on the clamp domain of Pol II. Our results define the structure of mammalian Pol II in its functional state, indicate that previous crystallographic analysis of yeast Pol II is relevant for understanding gene transcription in all eukaryotes, and provide a starting point for a mechanistic analysis of human transcription. | |||||||||
履歴 |
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Remark 700 | SHEET DETERMINATION METHOD: DSSP THE SHEETS PRESENTED AS "AG" IN EACH CHAIN ON SHEET RECORDS BELOW ... SHEET DETERMINATION METHOD: DSSP THE SHEETS PRESENTED AS "AG" IN EACH CHAIN ON SHEET RECORDS BELOW IS ACTUALLY AN 6-STRANDED BARREL THIS IS REPRESENTED BY A 7-STRANDED SHEET IN WHICH THE FIRST AND LAST STRANDS ARE IDENTICAL. THE SHEETS PRESENTED AS "AJ" IN EACH CHAIN ON SHEET RECORDS BELOW IS ACTUALLY AN 9-STRANDED BARREL THIS IS REPRESENTED BY A 10-STRANDED SHEET IN WHICH THE FIRST AND LAST STRANDS ARE IDENTICAL. THE SHEETS PRESENTED AS "BO" IN EACH CHAIN ON SHEET RECORDS BELOW IS ACTUALLY AN -3-STRANDED BARREL THIS IS REPRESENTED BY A -2-STRANDED SHEET IN WHICH THE FIRST AND LAST STRANDS ARE IDENTICAL. THE SHEETS PRESENTED AS "GA" IN EACH CHAIN ON SHEET RECORDS BELOW IS ACTUALLY AN -1-STRANDED BARREL THIS IS REPRESENTED BY A 0-STRANDED SHEET IN WHICH THE FIRST AND LAST STRANDS ARE IDENTICAL. |
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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 | ![]() | 762.3 KB | 表示 | ![]() |
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PDB形式 | ![]() | 609 KB | 表示 | ![]() |
PDBx/mmJSON形式 | ![]() | ツリー表示 | ![]() | |
その他 | ![]() |
-検証レポート
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
-DNA-DIRECTED RNA POLYMERASE II SUBUNIT ... , 6種, 6分子 BCDGIK
#2: タンパク質 | 分子量: 134041.422 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() ![]() |
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#3: タンパク質 | 分子量: 31466.098 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() ![]() |
#4: タンパク質 | 分子量: 16347.255 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() ![]() |
#7: タンパク質 | 分子量: 19314.283 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() ![]() |
#9: タンパク質 | 分子量: 14507.205 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() ![]() |
#11: タンパク質 | 分子量: 13310.284 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() ![]() |
-DNA-DIRECTED RNA POLYMERASES I, II, AND III SUBUNIT ... , 5種, 5分子 EFHJL
#5: タンパク質 | 分子量: 24514.219 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() ![]() |
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#6: タンパク質 | 分子量: 14491.026 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() ![]() |
#8: タンパク質 | 分子量: 17162.273 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() ![]() |
#10: タンパク質 | 分子量: 7655.123 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() ![]() |
#12: タンパク質 | 分子量: 7018.244 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() ![]() |
-DNA, DNA-RNA ELONGATION ... , 2種, 2分子 NT
#13: DNA鎖 | 分子量: 12085.806 Da / 分子数: 1 / 由来タイプ: 合成 / 由来: (合成) SYNTHETIC CONSTRUCT (人工物) |
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#15: DNA鎖 | 分子量: 11942.696 Da / 分子数: 1 / 由来タイプ: 合成 / 由来: (合成) SYNTHETIC CONSTRUCT (人工物) |
-タンパク質 / RNA鎖 , 2種, 2分子 AP
#14: RNA鎖 | 分子量: 6414.902 Da / 分子数: 1 / 由来タイプ: 合成 / 由来: (合成) SYNTHETIC CONSTRUCT (人工物) |
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#1: タンパク質 | 分子量: 217436.062 Da / 分子数: 1 / 由来タイプ: 天然 / 由来: (天然) ![]() ![]() |
-非ポリマー , 2種, 9分子 ![](data/chem/img/ZN.gif)
![](data/chem/img/MG.gif)
![](data/chem/img/MG.gif)
#16: 化合物 | ChemComp-ZN / #17: 化合物 | ChemComp-MG / | |
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-詳細
配列の詳細 | SYNTHETIC CONSTRUCT, CHAINS N, P, T GENBANK ACCESSION FOR CHAIN A GI|329663165 GENBANK ACCESSION ...SYNTHETIC CONSTRUCT, CHAINS N, P, T GENBANK ACCESSION FOR CHAIN A GI|329663165 GENBANK ACCESSION FOR CHAIN F GI|269315856 |
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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: BOVINE POL II ELONGATION COMPLEX / タイプ: COMPLEX |
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緩衝液 | 名称: 150 MM NACL, 5 MM HEPES, 0.01 MM ZNCL2, 10 MM DTT / pH: 7.25 / 詳細: 150 MM NACL, 5 MM HEPES, 0.01 MM ZNCL2, 10 MM DTT |
試料 | 濃度: 0.3 mg/ml / 包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES |
試料支持 | 詳細: HOLEY CARBON |
急速凍結 | 装置: FEI VITROBOT MARK IV / 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 / 日付: 2014年12月1日 |
電子銃 | 電子線源: ![]() |
電子レンズ | モード: BRIGHT FIELD / 倍率(公称値): 37000 X / 倍率(補正後): 37037 X / 最大 デフォーカス(公称値): 3100 nm / 最小 デフォーカス(公称値): 600 nm / Cs: 2 mm |
撮影 | 電子線照射量: 43 e/Å2 フィルム・検出器のモデル: GATAN K2 SUMMIT (4k x 4k) |
画像スキャン | デジタル画像の数: 1172 |
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解析
EMソフトウェア | 名称: RELION / バージョン: 1.3 / カテゴリ: 3次元再構成 | ||||||||||||
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CTF補正 | 詳細: INDIVIDUAL PARTICLES | ||||||||||||
対称性 | 点対称性: C1 (非対称) | ||||||||||||
3次元再構成 | 解像度: 3.4 Å / 粒子像の数: 264134 / ピクセルサイズ(実測値): 1.35 Å 詳細: THE FOLLOWING REGIONS WERE MODELED INTO THE UNSHARPENED EC1 MAP (EMDB-3218) DUE TO WEAKER DENSITY. RPB1 433-437, RPB1 1103-1108 (POLY-ALANINE), RPB1 1198-1205, RPB1 1262-1267, RPB1 1278-1281, ...詳細: THE FOLLOWING REGIONS WERE MODELED INTO THE UNSHARPENED EC1 MAP (EMDB-3218) DUE TO WEAKER DENSITY. RPB1 433-437, RPB1 1103-1108 (POLY-ALANINE), RPB1 1198-1205, RPB1 1262-1267, RPB1 1278-1281, RPB2 242-252, RPB2 326-331, RPB2 457-461 SUBMISSION BASED ON EXPERIMENTAL DATA FROM EMDB EMD-3218. (DEPOSITION ID: 13928). 対称性のタイプ: POINT | ||||||||||||
原子モデル構築 | B value: 137 / プロトコル: OTHER / 空間: REAL / 詳細: REFINEMENT PROTOCOL--CRYO-EM | ||||||||||||
精密化 | 最高解像度: 3.4 Å | ||||||||||||
精密化ステップ | サイクル: LAST / 最高解像度: 3.4 Å
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