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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: EMDB / ID: EMD-3541 | ||||||||||||
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タイトル | Structure of a spliceosome remodeled for exon ligation | ||||||||||||
![]() | Map of the spliceosomal C-star complex resulting from focussed classification on Prp22 | ||||||||||||
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![]() | pre-mRNA splicing / trans-esterification / lariat intermediate / complex C-star / Splicing | ||||||||||||
機能・相同性 | ![]() U2-type post-spliceosomal complex / spliceosomal complex disassembly / mRNA branch site recognition / U2-type post-mRNA release spliceosomal complex / cellular bud site selection / pre-mRNA 3'-splice site binding / post-mRNA release spliceosomal complex / nuclear mRNA surveillance / generation of catalytic spliceosome for first transesterification step / cis assembly of pre-catalytic spliceosome ...U2-type post-spliceosomal complex / spliceosomal complex disassembly / mRNA branch site recognition / U2-type post-mRNA release spliceosomal complex / cellular bud site selection / pre-mRNA 3'-splice site binding / post-mRNA release spliceosomal complex / nuclear mRNA surveillance / generation of catalytic spliceosome for first transesterification step / cis assembly of pre-catalytic spliceosome / splicing factor binding / spliceosome conformational change to release U4 (or U4atac) and U1 (or U11) / U4/U6 snRNP / 7-methylguanosine cap hypermethylation / pICln-Sm protein complex / U2-type catalytic step 1 spliceosome / pre-mRNA binding / snRNP binding / small nuclear ribonucleoprotein complex / SMN-Sm protein complex / spliceosomal tri-snRNP complex / U2-type spliceosomal complex / commitment complex / mRNA cis splicing, via spliceosome / U2-type prespliceosome assembly / U2-type catalytic step 2 spliceosome / U4 snRNP / U2 snRNP / U1 snRNP / poly(U) RNA binding / U2-type prespliceosome / precatalytic spliceosome / generation of catalytic spliceosome for second transesterification step / Formation of TC-NER Pre-Incision Complex / spliceosomal complex assembly / mRNA 5'-splice site recognition / Gap-filling DNA repair synthesis and ligation in TC-NER / mRNA 3'-splice site recognition / DNA replication origin binding / Dual incision in TC-NER / spliceosomal tri-snRNP complex assembly / Prp19 complex / protein K63-linked ubiquitination / U5 snRNA binding / DNA replication initiation / U5 snRNP / U2 snRNA binding / U6 snRNA binding / pre-mRNA intronic binding / spliceosomal snRNP assembly / U1 snRNA binding / U4/U6 x U5 tri-snRNP complex / positive regulation of cell cycle / catalytic step 2 spliceosome / nuclear periphery / positive regulation of RNA splicing / spliceosomal complex / RING-type E3 ubiquitin transferase / mRNA splicing, via spliceosome / metallopeptidase activity / ubiquitin-protein transferase activity / ubiquitin protein ligase activity / RNA helicase activity / RNA helicase / DNA repair / GTPase activity / mRNA binding / chromatin binding / GTP binding / chromatin / ATP hydrolysis activity / mitochondrion / DNA binding / RNA binding / zinc ion binding / ATP binding / identical protein binding / nucleus / cytosol / cytoplasm 類似検索 - 分子機能 | ||||||||||||
生物種 | ![]() ![]() | ||||||||||||
手法 | 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 4.17 Å | ||||||||||||
![]() | Fica SM / Oubridge C | ||||||||||||
資金援助 | ![]()
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![]() | ![]() タイトル: Cryo-EM structure of the spliceosome immediately after branching. 著者: Wojciech P Galej / Max E Wilkinson / Sebastian M Fica / Chris Oubridge / Andrew J Newman / Kiyoshi Nagai / ![]() 要旨: Precursor mRNA (pre-mRNA) splicing proceeds by two consecutive transesterification reactions via a lariat-intron intermediate. Here we present the 3.8 Å cryo-electron microscopy structure of the ...Precursor mRNA (pre-mRNA) splicing proceeds by two consecutive transesterification reactions via a lariat-intron intermediate. Here we present the 3.8 Å cryo-electron microscopy structure of the spliceosome immediately after lariat formation. The 5'-splice site is cleaved but remains close to the catalytic Mg site in the U2/U6 small nuclear RNA (snRNA) triplex, and the 5'-phosphate of the intron nucleotide G(+1) is linked to the branch adenosine 2'OH. The 5'-exon is held between the Prp8 amino-terminal and linker domains, and base-pairs with U5 snRNA loop 1. Non-Watson-Crick interactions between the branch helix and 5'-splice site dock the branch adenosine into the active site, while intron nucleotides +3 to +6 base-pair with the U6 snRNA ACAGAGA sequence. Isy1 and the step-one factors Yju2 and Cwc25 stabilize docking of the branch helix. The intron downstream of the branch site emerges between the Prp8 reverse transcriptase and linker domains and extends towards the Prp16 helicase, suggesting a plausible mechanism of remodelling before exon ligation. | ||||||||||||
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 | EMマップ: ![]() ![]() ![]() |
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 | ![]() | 248.2 MB | ![]() | |
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ヘッダ (付随情報) | ![]() ![]() | 67.6 KB 67.6 KB | 表示 表示 | ![]() |
FSC (解像度算出) | ![]() ![]() | 14.5 KB 14.5 KB | 表示 表示 | ![]() |
画像 | ![]() ![]() | 63.7 KB 62.6 KB | ||
Filedesc metadata | ![]() | 18.2 KB | ||
その他 | ![]() | 243.9 MB | ||
アーカイブディレクトリ | ![]() ![]() | HTTPS FTP |
-関連構造データ
関連構造データ | ![]() 5mq0MC ![]() 3539C ![]() 3542C ![]() 5mpsC 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 ( |
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類似構造データ | |
電子顕微鏡画像生データ | ![]() Data #1: Unaligned movies of C-complex spliceosome with 3' splice site AG to AC mutation (Dataset 1) [micrographs - multiframe] Data #2: Unaligned movies of C and C*-complex spliceosomes with 3' splice site AG to AdG mutation (Dataset 2) [micrographs - multiframe] Data #3: Unaligned movies of C and C*-complex spliceosomes with 3' splice site AG to AdG mutation (Dataset 3) [micrographs - multiframe] Data #4: Aligned movies of C-complex spliceosomes with cold-sensitive prp16-302 mutation, purified with Cwc25 (Dataset 4) [micrographs - multiframe] Data #5: Unaligned movies of C-complex spliceosomes with cold-sensitive prp16-302 mutation, purified with Cwc25 and incubated with ATP and Mg (Dataset 5) [micrographs - multiframe] Data #6: Unaligned movies of C, C*, and P-complex spliceosomes with dominant-negative Prp22 mutation K512A, purified with Slu7 (Dataset 6) [micrographs - multiframe] Data #7: Unaligned movies of P-complex spliceosomes with dominant-negative Prp22 mutation K512A, treated with anti-3'exon RNaseH oligo, purified in presence of Mg (Dataset 9) [micrographs - single frame] Data #8: Selected C-complex particles after polishing [picked particles - single frame - processed] Data #9: Selected P-complex particles after polishing [picked particles - single frame - processed] Data #10: Various signal subtractions for C- and P-complex spliceosomes [picked particles - single frame - processed]) |
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リンク
EMDBのページ | ![]() ![]() |
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「今月の分子」の関連する項目 |
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マップ
ファイル | ![]() | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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注釈 | Map of the spliceosomal C-star complex resulting from focussed classification on Prp22 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
投影像・断面図 | 画像のコントロール
画像は Spider により作成 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 1.43 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
-追加マップ: Map of the spliceosomal C-star complex resulting from...
ファイル | emd_3541_additional.map | ||||||||||||
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注釈 | Map of the spliceosomal C-star complex resulting from focussed classification on peripheral regions of U2 snRNP | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
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試料の構成要素
+全体 : Saccharomyces cerevisiae spliceosome. Complex C just after Prp16-...
+超分子 #1: Saccharomyces cerevisiae spliceosome. Complex C just after Prp16-...
+分子 #1: Yeast UBC4 gene for ubiquitin-conjugating enzyme
+分子 #2: 5'-EXON OF UBC4 PRE-MRNA
+分子 #3: S.cerevisiae chromosome II reading frame ORF YBR230c
+分子 #4: Saccharomyces cerevisiae strain T.52_2H chromosome XII sequence
+分子 #5: Saccharomyces cerevisiae strain WI_C_MBSP_4 chromosome VII sequence
+分子 #34: 3'-EXON OF UBC4 PRE-MRNA, BOUND BY PRP22 HELICASE
+分子 #6: Pre-mRNA-splicing factor 8
+分子 #7: Pre-mRNA-splicing factor SNU114
+分子 #8: Pre-mRNA-splicing factor CWC22
+分子 #9: Pre-mRNA-splicing factor PRP46
+分子 #10: Pre-mRNA-processing protein 45
+分子 #11: Pre-mRNA-splicing factor BUD31
+分子 #12: Pre-mRNA-splicing factor CWC2
+分子 #13: Pre-mRNA-splicing factor SLT11
+分子 #14: Pre-mRNA-splicing factor CEF1
+分子 #15: Pre-mRNA-splicing factor CWC15
+分子 #16: Pre-mRNA-splicing factor CWC21
+分子 #17: Pre-mRNA-splicing factor CLF1
+分子 #18: Pre-mRNA-splicing factor SYF1,PRE-MRNA-SPLICING FACTOR SYF1
+分子 #19: Pre-mRNA-splicing factor 18
+分子 #20: Pre-mRNA-splicing factor SLU7
+分子 #21: Pre-mRNA-processing factor 17
+分子 #22: UNKNOWN PROTEIN
+分子 #23: Pre-mRNA-splicing factor SYF2
+分子 #24: Small nuclear ribonucleoprotein-associated protein B
+分子 #25: Small nuclear ribonucleoprotein Sm D3
+分子 #26: Small nuclear ribonucleoprotein E
+分子 #27: Small nuclear ribonucleoprotein F
+分子 #28: Small nuclear ribonucleoprotein G
+分子 #29: Small nuclear ribonucleoprotein Sm D1
+分子 #30: Small nuclear ribonucleoprotein Sm D2
+分子 #31: Pre-mRNA-splicing factor ATP-dependent RNA helicase PRP22
+分子 #32: U2 small nuclear ribonucleoprotein A'
+分子 #33: U2 small nuclear ribonucleoprotein B''
+分子 #35: Pre-mRNA-splicing factor SNT309
+分子 #36: Pre-mRNA-processing factor 19
+分子 #37: MAGNESIUM ION
+分子 #38: POTASSIUM ION
+分子 #39: INOSITOL HEXAKISPHOSPHATE
+分子 #40: GUANOSINE-5'-TRIPHOSPHATE
+分子 #41: ZINC ION
-実験情報
-構造解析
手法 | クライオ電子顕微鏡法 |
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![]() | 単粒子再構成法 |
試料の集合状態 | particle |
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試料調製
濃度 | 0.3 mg/mL | ||||||||||||||||||
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緩衝液 | pH: 7.9 構成要素:
詳細: NP-40 is also called IGEPAL CA-630 | ||||||||||||||||||
グリッド | モデル: Quantifoil R1.2/1.3 / 材質: COPPER / メッシュ: 400 / 支持フィルム - 材質: CARBON / 支持フィルム - トポロジー: HOLEY ARRAY / 支持フィルム - Film thickness: 6 / 前処理 - タイプ: GLOW DISCHARGE / 前処理 - 時間: 30 sec. / 前処理 - 雰囲気: AIR / 前処理 - 気圧: 20.0 kPa | ||||||||||||||||||
凍結 | 凍結剤: ETHANE / チャンバー内湿度: 100 % / チャンバー内温度: 277 K / 装置: FEI VITROBOT MARK III 詳細: 3.5 microlitres sample were applied to the grid, left for 25 seconds and then blotted for 3.0-3.5 seconds before plunging.. |
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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 Quantum |
詳細 | GIF Quantum energy filter, 20 eV slit width |
撮影 | フィルム・検出器のモデル: GATAN K2 SUMMIT (4k x 4k) 検出モード: SUPER-RESOLUTION / 実像数: 3596 / 平均露光時間: 0.8 sec. / 平均電子線量: 2.0 e/Å2 詳細: Total dose: 40 electrons/Angstrom^2 over 16 seconds. 20 movie frames collected at 1.25 frames per second. |
電子線 | 加速電圧: 300 kV / 電子線源: ![]() |
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): 4.5 µm / 最小 デフォーカス(公称値): 0.5 µm / 倍率(公称値): 81000 |
試料ステージ | 試料ホルダーモデル: FEI TITAN KRIOS AUTOGRID HOLDER ホルダー冷却材: NITROGEN |
実験機器 | ![]() モデル: Titan Krios / 画像提供: FEI Company |
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画像解析
-原子モデル構築 1
詳細 | Used secondary structure restraints generated in ProSMART and LibG. |
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精密化 | 空間: RECIPROCAL / プロトコル: FLEXIBLE FIT / 温度因子: 330 / 当てはまり具合の基準: Fourier Shell Correlation |
得られたモデル | ![]() PDB-5mq0: |