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Yorodumi- PDB-6ymw: Cryo-EM structure of yeast mitochondrial RNA polymerase transcrip... -
+Open data
-Basic information
Entry | Database: PDB / ID: 6ymw | ||||||
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Title | Cryo-EM structure of yeast mitochondrial RNA polymerase transcription initiation complex | ||||||
Components |
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Keywords | TRANSCRIPTION / gene transcription / polymerase / RDRP / MTF1 / RPO41 / POLRMT / mtRNAP / DNA / transcription initiation / RNA polymerase / mitochondria | ||||||
Function / homology | Function and homology information Mitochondrial transcription initiation / mitochondrial DNA-directed RNA polymerase complex / mitochondrial promoter sequence-specific DNA binding / mitochondrial transcription factor activity / transcription initiation at mitochondrial promoter / 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 promoter sequence-specific DNA binding / mitochondrial transcription factor activity / transcription initiation at mitochondrial promoter / 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 | Saccharomyces cerevisiae (brewer's yeast) Saccharomyces cerevisiae S288C (yeast) synthetic construct (others) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.71 Å | ||||||
Authors | Das, K. / Patel, S.S. | ||||||
Funding support | Belgium, 1items
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Citation | Journal: Mol Cell / Year: 2021 Title: Cryo-EM Structures Reveal Transcription Initiation Steps by Yeast Mitochondrial RNA Polymerase. Authors: Brent De Wijngaert / Shemaila Sultana / Anupam Singh / Chhaya Dharia / Hans Vanbuel / Jiayu Shen / Daniel Vasilchuk / Sergio E Martinez / Eaazhisai Kandiah / Smita S Patel / Kalyan Das / Abstract: Mitochondrial RNA polymerase (mtRNAP) is crucial in cellular energy production, yet understanding of mitochondrial DNA transcription initiation lags that of bacterial and nuclear DNA transcription. ...Mitochondrial RNA polymerase (mtRNAP) is crucial in cellular energy production, yet understanding of mitochondrial DNA transcription initiation lags that of bacterial and nuclear DNA transcription. We report structures of two transcription initiation intermediate states of yeast mtRNAP that explain promoter melting, template alignment, DNA scrunching, abortive synthesis, and transition into elongation. In the partially melted initiation complex (PmIC), transcription factor MTF1 makes base-specific interactions with flipped non-template (NT) nucleotides "AAGT" at -4 to -1 positions of the DNA promoter. In the initiation complex (IC), the template in the expanded 7-mer bubble positions the RNA and NTP analog UTPαS, while NT scrunches into an NT loop. The scrunched NT loop is stabilized by the centrally positioned MTF1 C-tail. The IC and PmIC states coexist in solution, revealing a dynamic equilibrium between two functional states. Frequent scrunching/unscruching transitions and the imminent steric clashes of the inflating NT loop and growing RNA:DNA with the C-tail explain abortive synthesis and transition into elongation. | ||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 6ymw.cif.gz | 283 KB | Display | PDBx/mmCIF format |
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PDB format | pdb6ymw.ent.gz | 209.1 KB | Display | PDB format |
PDBx/mmJSON format | 6ymw.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 6ymw_validation.pdf.gz | 1.5 MB | Display | wwPDB validaton report |
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Full document | 6ymw_full_validation.pdf.gz | 1.6 MB | Display | |
Data in XML | 6ymw_validation.xml.gz | 51.8 KB | Display | |
Data in CIF | 6ymw_validation.cif.gz | 78.3 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/ym/6ymw ftp://data.pdbj.org/pub/pdb/validation_reports/ym/6ymw | HTTPS FTP |
-Related structure data
Related structure data | 10846MC 6ymvC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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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.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / Gene: MTF1, YMR228W, YM9959.10 / Production host: Escherichia coli (E. coli) 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 Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / Gene: RPO41, YFL036W / Production host: Escherichia coli (E. coli) / References: UniProt: P13433, DNA-directed RNA polymerase |
-DNA chain , 2 types, 2 molecules NT
#3: DNA chain | Mass: 10248.671 Da / Num. of mol.: 1 / Source method: obtained synthetically / Details: 15S mitochondria / Source: (synth.) Saccharomyces cerevisiae S288C (yeast) |
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#4: DNA chain | Mass: 10047.511 Da / Num. of mol.: 1 / Source method: obtained synthetically / Details: 15S mitochondria / Source: (synth.) Saccharomyces cerevisiae S288C (yeast) |
-RNA chain , 1 types, 1 molecules C
#5: RNA chain | Mass: 805.413 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) |
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-Non-polymers , 2 types, 2 molecules
#6: Chemical | ChemComp-P5E / [[( |
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#7: Chemical | ChemComp-MG / |
-Details
Has ligand of interest | Y |
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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 |
-Sample preparation
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Molecular weight | Value: 0.202 MDa / Experimental value: NO | ||||||||||||||||||||||||||||||
Source (natural) |
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Source (recombinant) |
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Buffer solution | pH: 7 Details: 50mM Bis-tris propane, 100mM NaCl, 5mM MgCl2, 1mM EDTA, 2mM DTT | ||||||||||||||||||||||||||||||
Specimen | Conc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||||||||
Vitrification | Instrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 8 K / Details: 5 uL sample; back blotting for 12 -14 second |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 165000 X / Nominal defocus max: 22000 nm / Nominal defocus min: 7000 nm / Calibrated defocus min: 5500 nm / Calibrated defocus max: 26000 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 6 sec. / Electron dose: 61 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 3500 |
EM imaging optics | Energyfilter slit width: 20 eV |
Image scans | Movie frames/image: 50 |
-Processing
EM software |
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CTF correction | Type: NONE | ||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.71 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 62807 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: FLEXIBLE FIT / Space: REAL / Target criteria: Coorelation coefficu=ient / Details: Realspace refinement | ||||||||||||||||||||||||||||||||||||
Atomic model building | 3D fitting-ID: 1 / Source name: PDB / Type: experimental model
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Refinement | Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 55.32 Å2 | ||||||||||||||||||||||||||||||||||||
Refine LS restraints |
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