RPB4-RPB7 complex / nuclear-transcribed mRNA catabolic process, deadenylation-dependent decay / RNA Polymerase I Transcription Initiation / Processing of Capped Intron-Containing Pre-mRNA / RNA Polymerase III Transcription Initiation From Type 2 Promoter / RNA Pol II CTD phosphorylation and interaction with CE / Formation of the Early Elongation Complex / mRNA Capping / RNA polymerase II transcribes snRNA genes / TP53 Regulates Transcription of DNA Repair Genes ...RPB4-RPB7 complex / nuclear-transcribed mRNA catabolic process, deadenylation-dependent decay / RNA Polymerase I Transcription Initiation / Processing of Capped Intron-Containing Pre-mRNA / RNA Polymerase III Transcription Initiation From Type 2 Promoter / RNA Pol II CTD phosphorylation and interaction with CE / Formation of the Early Elongation Complex / mRNA Capping / RNA polymerase II transcribes snRNA genes / TP53 Regulates Transcription of DNA Repair Genes / termination of RNA polymerase II transcription / RNA Polymerase II Promoter Escape / RNA Polymerase II Transcription Pre-Initiation And Promoter Opening / RNA Polymerase II Transcription Initiation / RNA Polymerase II Transcription Initiation And Promoter Clearance / RNA-templated transcription / RNA Polymerase II Pre-transcription Events / termination of RNA polymerase III transcription / Formation of TC-NER Pre-Incision Complex / termination of RNA polymerase I transcription / transcription initiation at RNA polymerase III promoter / RNA Polymerase I Promoter Escape / nucleolar large rRNA transcription by RNA polymerase I / maintenance of transcriptional fidelity during transcription elongation by RNA polymerase II / Gap-filling DNA repair synthesis and ligation in TC-NER / transcription initiation at RNA polymerase I promoter / transcription by RNA polymerase I / Estrogen-dependent gene expression / positive regulation of nuclear-transcribed mRNA poly(A) tail shortening / nuclear-transcribed mRNA catabolic process / transcription by RNA polymerase III / RNA polymerase II activity / Dual incision in TC-NER / transcription elongation by RNA polymerase I / tRNA transcription by RNA polymerase III / RNA polymerase I activity / RNA polymerase I complex / RNA polymerase III complex / positive regulation of translational initiation / transcription-coupled nucleotide-excision repair / RNA polymerase II, core complex / translesion synthesis / translation initiation factor binding / DNA-templated transcription initiation / transcription initiation at RNA polymerase II promoter / transcription elongation by RNA polymerase II / P-body / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / cytoplasmic stress granule / mRNA processing / ribosome biogenesis / single-stranded DNA binding / nucleic acid binding / transcription by RNA polymerase II / single-stranded RNA binding / protein dimerization activity / mRNA binding / nucleotide binding / nucleolus / mitochondrion / DNA binding / zinc ion binding / nucleoplasm / nucleus / metal ion binding / cytoplasm 類似検索 - 分子機能
DNA-directed RNA polymerase II subunit RPB1 / DNA-directed RNA polymerase II subunit RPB2 / DNA-directed RNA polymerase II subunit RPB3 / DNA-directed RNA polymerase II subunit RPB4 / DNA-directed RNA polymerases I, II, and III subunit RPABC1 / DNA-directed RNA polymerases I, II, and III subunit RPABC2 / DNA-directed RNA polymerases I, II, and III subunit RPABC3 / DNA-directed RNA polymerases I, II, and III subunit RPABC5 / DNA-directed RNA polymerase II subunit RPB9 / DNA-directed RNA polymerase II subunit RPB7 / DNA-directed RNA polymerase II subunit RPB11 類似検索 - 構成要素
ジャーナル: PLoS Biol / 年: 2011 タイトル: Molecular architecture of the human Mediator-RNA polymerase II-TFIIF assembly. 著者: Carrie Bernecky / Patricia Grob / Christopher C Ebmeier / Eva Nogales / Dylan J Taatjes / 要旨: The macromolecular assembly required to initiate transcription of protein-coding genes, known as the Pre-Initiation Complex (PIC), consists of multiple protein complexes and is approximately 3.5 MDa ...The macromolecular assembly required to initiate transcription of protein-coding genes, known as the Pre-Initiation Complex (PIC), consists of multiple protein complexes and is approximately 3.5 MDa in size. At the heart of this assembly is the Mediator complex, which helps regulate PIC activity and interacts with the RNA polymerase II (pol II) enzyme. The structure of the human Mediator-pol II interface is not well-characterized, whereas attempts to structurally define the Mediator-pol II interaction in yeast have relied on incomplete assemblies of Mediator and/or pol II and have yielded inconsistent interpretations. We have assembled the complete, 1.9 MDa human Mediator-pol II-TFIIF complex from purified components and have characterized its structural organization using cryo-electron microscopy and single-particle reconstruction techniques. The orientation of pol II within this assembly was determined by crystal structure docking and further validated with projection matching experiments, allowing the structural organization of the entire human PIC to be envisioned. Significantly, pol II orientation within the Mediator-pol II-TFIIF assembly can be reconciled with past studies that determined the location of other PIC components relative to pol II itself. Pol II surfaces required for interacting with TFIIB, TFIIE, and promoter DNA (i.e., the pol II cleft) are exposed within the Mediator-pol II-TFIIF structure; RNA exit is unhindered along the RPB4/7 subunits; upstream and downstream DNA is accessible for binding additional factors; and no major structural re-organization is necessary to accommodate the large, multi-subunit TFIIH or TFIID complexes. The data also reveal how pol II binding excludes Mediator-CDK8 subcomplex interactions and provide a structural basis for Mediator-dependent control of PIC assembly and function. Finally, parallel structural analysis of Mediator-pol II complexes lacking TFIIF reveal that TFIIF plays a key role in stabilizing pol II orientation within the assembly.
ダウンロード / ファイル: emd_5343.map.gz / 形式: CCP4 / 大きさ: 15.5 MB / タイプ: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
注釈
none
ボクセルのサイズ
X=Y=Z: 4.29 Å
密度
表面レベル
登録者による: 0.0284 / ムービー #1: 0.0284
最小 - 最大
-0.00885565 - 0.0734321
平均 (標準偏差)
0.000528866 (±0.00409114)
対称性
空間群: 1
詳細
EMDB XML:
マップ形状
Axis order
X
Y
Z
Origin
0
0
0
サイズ
161
161
161
Spacing
161
161
161
セル
A=B=C: 690.69 Å α=β=γ: 90 °
CCP4マップ ヘッダ情報:
mode
Image stored as Reals
Å/pix. X/Y/Z
4.29
4.29
4.29
M x/y/z
161
161
161
origin x/y/z
0.000
0.000
0.000
length x/y/z
690.690
690.690
690.690
α/β/γ
90.000
90.000
90.000
start NX/NY/NZ
-62
-62
-62
NX/NY/NZ
125
125
125
MAP C/R/S
1
2
3
start NC/NR/NS
0
0
0
NC/NR/NS
161
161
161
D min/max/mean
-0.009
0.073
0.001
-
添付データ
-
試料の構成要素
-
全体 : Assembly of VP16-bound human Mediator, RNA polymerase II, and TFIIF
全体
名称: Assembly of VP16-bound human Mediator, RNA polymerase II, and TFIIF
要素
試料: Assembly of VP16-bound human Mediator, RNA polymerase II, and TFIIF
タンパク質・ペプチド: core Mediator
タンパク質・ペプチド: RNA polymerase II
タンパク質・ペプチド: TFIIF
-
超分子 #1000: Assembly of VP16-bound human Mediator, RNA polymerase II, and TFIIF
超分子
名称: Assembly of VP16-bound human Mediator, RNA polymerase II, and TFIIF タイプ: sample / ID: 1000 / 集合状態: one Mediator complex binds one pol II-TFIIF / Number unique components: 3
pH: 7.9 詳細: 20 mM HEPES, 0.10 mM EDTA, 150 mM KCl, 0.02% NP-40, 35% glycerol
染色
タイプ: NEGATIVE 詳細: grids with adsorbed protein washed 3x with buffer containing 5% trehalose, 20 mM HEPES, 100 mM KCl, and 0.10 mM EDTA, then subjected to cryo-negative staining in a saturated solution (1.2M) ...詳細: grids with adsorbed protein washed 3x with buffer containing 5% trehalose, 20 mM HEPES, 100 mM KCl, and 0.10 mM EDTA, then subjected to cryo-negative staining in a saturated solution (1.2M) of ammonium molybdate (pH 7.5)