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Yorodumi- EMDB-10846: Cryo-EM structure of yeast mitochondrial RNA polymerase transcrip... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-10846 | |||||||||
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Title | Cryo-EM structure of yeast mitochondrial RNA polymerase transcription initiation complex | |||||||||
Map data | ||||||||||
Sample |
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Keywords | gene transcription / polymerase / RDRP / MTF1 / RPO41 / POLRMT / mtRNAP / DNA / transcription initiation / RNA polymerase / mitochondria / TRANSCRIPTION | |||||||||
Function / homology | Function and homology information 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 | Saccharomyces cerevisiae (brewer's yeast) / synthetic construct (others) / Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) / Saccharomyces cerevisiae S288C (yeast) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.71 Å | |||||||||
Authors | Das K | |||||||||
Funding support | Belgium, 1 items
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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 | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_10846.map.gz | 14.5 MB | EMDB map data format | |
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Header (meta data) | emd-10846-v30.xml emd-10846.xml | 28.4 KB 28.4 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_10846_fsc.xml | 9.2 KB | Display | FSC data file |
Images | emd_10846.png | 188.8 KB | ||
Masks | emd_10846_msk_1.map | 15.6 MB | Mask map | |
Filedesc metadata | emd-10846.cif.gz | 8 KB | ||
Others | emd_10846_half_map_1.map.gz emd_10846_half_map_2.map.gz | 49.6 MB 49.6 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-10846 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-10846 | HTTPS FTP |
-Validation report
Summary document | emd_10846_validation.pdf.gz | 1 MB | Display | EMDB validaton report |
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Full document | emd_10846_full_validation.pdf.gz | 1 MB | Display | |
Data in XML | emd_10846_validation.xml.gz | 15.9 KB | Display | |
Data in CIF | emd_10846_validation.cif.gz | 20.5 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-10846 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-10846 | HTTPS FTP |
-Related structure data
Related structure data | 6ymwMC 6ymvC C: citing same article (ref.) M: atomic model generated by this map |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_10846.map.gz / Format: CCP4 / Size: 15.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Voxel size | X=Y=Z: 0.827 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Mask #1
File | emd_10846_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #2
File | emd_10846_half_map_1.map | ||||||||||||
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Density Histograms |
-Half map: #1
File | emd_10846_half_map_2.map | ||||||||||||
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Density Histograms |
-Sample components
+Entire : mitochondria DNA-dependent RNA polymerase pre-initiation complex
+Supramolecule #1: mitochondria DNA-dependent RNA polymerase pre-initiation complex
+Supramolecule #2: mitochondria DNA-dependent RNA polymerase pre-initiation complex
+Supramolecule #3: DNA
+Supramolecule #4: RNA (pppGpG)
+Macromolecule #1: Mitochondrial transcription factor 1
+Macromolecule #2: DNA-directed RNA polymerase, mitochondrial
+Macromolecule #3: Chains: N
+Macromolecule #4: Chains: T
+Macromolecule #5: RNA (pppGpG)
+Macromolecule #6: [[(2~{R},3~{S},4~{R},5~{R})-5-[2,4-bis(oxidanylidene)pyrimidin-1-...
+Macromolecule #7: MAGNESIUM ION
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 1.0 mg/mL |
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Buffer | pH: 7 Details: 50mM Bis-tris propane, 100mM NaCl, 5mM MgCl2, 1mM EDTA, 2mM DTT |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 8 K / Instrument: LEICA EM GP / Details: 5 uL sample; back blotting for 12 -14 second. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Specialist optics | Energy filter - Slit width: 20 eV |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 3500 / Average exposure time: 6.0 sec. / Average electron dose: 61.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 70.0 µm / Calibrated defocus max: 26.0 µm / Calibrated defocus min: 5.5 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 22.0 µm / Nominal defocus min: 7.0 µm / Nominal magnification: 165000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
+Image processing
-Atomic model buiding 1
Initial model |
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Details | Realspace refinement | ||||||
Refinement | Space: REAL / Protocol: FLEXIBLE FIT / Target criteria: Coorelation coefficu=ient | ||||||
Output model | PDB-6ymw: |