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- PDB-8atv: Cryo-EM structure of yeast mitochondrial RNA polymerase transcrip... -
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Open data
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Basic information
Entry | Database: PDB / ID: 8atv | ||||||
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Title | Cryo-EM structure of yeast mitochondrial RNA polymerase transcription initiation complex with 5-mer RNA, pppGpGpApApA (IC5) | ||||||
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![]() | TRANSCRIPTION / gene transcription / polymerase / RDRP / MTF1 / RPO41 / POLRMT / mtRNAP / DNA / transcription initiation / RNA polymerase / mitochondria | ||||||
Function / homology | ![]() Mitochondrial transcription initiation / mitochondrial DNA-directed RNA polymerase complex / mitochondrial transcription factor activity / transcription initiation at mitochondrial promoter / mitochondrial promoter sequence-specific DNA binding / mitochondrial transcription / mitochondrial genome maintenance / DNA primase activity / DNA replication, synthesis of primer / positive regulation of DNA-templated transcription, elongation ...Mitochondrial transcription initiation / mitochondrial DNA-directed RNA polymerase complex / mitochondrial transcription factor activity / transcription initiation at mitochondrial promoter / mitochondrial promoter sequence-specific DNA binding / mitochondrial transcription / mitochondrial genome maintenance / DNA primase activity / DNA replication, synthesis of primer / positive regulation of DNA-templated transcription, elongation / mitochondrial nucleoid / Transferases; Transferring one-carbon groups; Methyltransferases / methyltransferase activity / mitochondrial intermembrane space / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / methylation / mitochondrial matrix / mitochondrion / DNA binding / RNA binding / cytoplasm Similarity search - Function | ||||||
Biological species | ![]() ![]() | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.39 Å | ||||||
![]() | Goovaerts, Q. / Shen, J. / Patel, S.S. / Das, K. | ||||||
Funding support | ![]()
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![]() | ![]() Title: Structures illustrate step-by-step mitochondrial transcription initiation. Authors: Quinten Goovaerts / Jiayu Shen / Brent De Wijngaert / Urmimala Basu / Smita S Patel / Kalyan Das / ![]() ![]() Abstract: Transcription initiation is a key regulatory step in gene expression during which RNA polymerase (RNAP) initiates RNA synthesis de novo, and the synthesized RNA at a specific length triggers the ...Transcription initiation is a key regulatory step in gene expression during which RNA polymerase (RNAP) initiates RNA synthesis de novo, and the synthesized RNA at a specific length triggers the transition to the elongation phase. Mitochondria recruit a single-subunit RNAP and one or two auxiliary factors to initiate transcription. Previous studies have revealed the molecular architectures of yeast and human mitochondrial RNAP initiation complexes (ICs). Here we provide a comprehensive, stepwise mechanism of transcription initiation by solving high-resolution cryogenic electron microscopy (cryo-EM) structures of yeast mitochondrial RNAP and the transcription factor Mtf1 catalysing two- to eight-nucleotide RNA synthesis at single-nucleotide addition steps. The growing RNA-DNA is accommodated in the polymerase cleft by template scrunching and non-template reorganization, creating stressed intermediates. During early initiation, non-template strand scrunching and unscrunching destabilize the short two- and three-nucleotide RNAs, triggering abortive synthesis. Subsequently, the non-template reorganizes into a base-stacked staircase-like structure supporting processive five- to eight-nucleotide RNA synthesis. The expanded non-template staircase and highly scrunched template in IC8 destabilize the promoter interactions with Mtf1 to facilitate initiation bubble collapse and promoter escape for the transition from initiation to the elongation complex (EC). The series of transcription initiation steps, each guided by the interplay of multiple structural components, reveal a finely tuned mechanism for potential regulatory control. | ||||||
History |
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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PDBx/mmCIF format | ![]() | 275.1 KB | Display | ![]() |
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PDB format | ![]() | 205.5 KB | Display | ![]() |
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-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 15664MC ![]() 8ap1C ![]() 8attC ![]() 8atwC ![]() 8c5sC ![]() 8c5uC ![]() 8q63C M: map data used to model this data C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
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Assembly
Deposited unit | ![]()
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Components
-Protein , 2 types, 2 molecules BA
#1: Protein | Mass: 41151.203 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() References: UniProt: P14908, Transferases; Transferring one-carbon groups; Methyltransferases |
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#2: Protein | Mass: 143282.656 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Details: Residues 101-1351 / Source: (gene. exp.) ![]() ![]() ![]() ![]() |
-DNA chain , 2 types, 2 molecules NT
#3: DNA chain | Mass: 11164.266 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) ![]() ![]() |
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#4: DNA chain | Mass: 10985.101 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) ![]() ![]() |
-RNA chain / Non-polymers , 2 types, 2 molecules C![](data/chem/img/GTP.gif)
![](data/chem/img/GTP.gif)
#5: RNA chain | Mass: 1287.866 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) ![]() ![]() |
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#6: Chemical | ChemComp-GTP / |
-Details
Has ligand of interest | N |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
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Buffer solution | pH: 7 Details: 50 mM Bis-Tris propane; 100 mM NaCl, 5 mM MgCl2, 1 mM EDTA, 2 mM DTT | ||||||||||||||||||||||||||||||||||||||||||
Specimen | Conc.: 0.9 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||||||||||||||||||||
Specimen support | Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3 | ||||||||||||||||||||||||||||||||||||||||||
Vitrification | Instrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 281.15 K / Details: 3 ul sample, back blotting for 12 seconds |
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Electron microscopy imaging
Microscopy | Model: TFS GLACIOS |
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Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 150000 X / Nominal defocus max: 2200 nm / Nominal defocus min: 800 nm / Calibrated defocus min: 550 nm / Calibrated defocus max: 2600 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: OTHER |
Image recording | Average exposure time: 38.48 sec. / Electron dose: 40 e/Å2 / Detector mode: COUNTING / Film or detector model: FEI FALCON III (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 602 |
EM imaging optics | Energyfilter slit width: 20 eV |
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Processing
EM software |
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CTF correction | Type: NONE | ||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 549988 | ||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.39 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 91298 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||
Atomic model building | B value: 140 / Protocol: FLEXIBLE FIT / Space: REAL / Target criteria: Correlation coefficient / Details: Realspace refinement | ||||||||||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 6YMV Accession code: 6YMV / Source name: PDB / Type: experimental model |