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- EMDB-18499: Structure of the plastid-encoded RNA polymerase complex (PEP) fro... -

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Basic information

Entry
Database: EMDB / ID: EMD-18499
TitleStructure of the plastid-encoded RNA polymerase complex (PEP) from Sinapis alba - Map A
Map dataMap A, sharpened.
Sample
  • Complex: Plastid-encoded DNA-dependent RNA polymerase (PEP)
KeywordsTranscription / Chloroplasts / Gene Expression / RNA / Polymerase
Biological speciesSinapis alba (white mustard)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.43 Å
Authorsdo Prado PFV / Ahrens FM / Pfannschmidt T / Hillen HS
Funding support Germany, 5 items
OrganizationGrant numberCountry
German Research Foundation (DFG)FOR2848 Germany
German Research Foundation (DFG)SFB1190 Germany
German Research Foundation (DFG)SFB1565 Germany
German Research Foundation (DFG)EXC 2067/1 390729940 Germany
German Research Foundation (DFG)PF323-7 Germany
CitationJournal: Mol Cell / Year: 2024
Title: Structure of the multi-subunit chloroplast RNA polymerase.
Authors: Paula F V do Prado / Frederik M Ahrens / Monique Liebers / Noah Ditz / Hans-Peter Braun / Thomas Pfannschmidt / Hauke S Hillen /
Abstract: Chloroplasts contain a dedicated genome that encodes subunits of the photosynthesis machinery. Transcription of photosynthesis genes is predominantly carried out by a plastid-encoded RNA polymerase ...Chloroplasts contain a dedicated genome that encodes subunits of the photosynthesis machinery. Transcription of photosynthesis genes is predominantly carried out by a plastid-encoded RNA polymerase (PEP), a nearly 1 MDa complex composed of core subunits with homology to eubacterial RNA polymerases (RNAPs) and at least 12 additional chloroplast-specific PEP-associated proteins (PAPs). However, the architecture of this complex and the functions of the PAPs remain unknown. Here, we report the cryo-EM structure of a 19-subunit PEP complex from Sinapis alba (white mustard). The structure reveals that the PEP core resembles prokaryotic and nuclear RNAPs but contains chloroplast-specific features that mediate interactions with the PAPs. The PAPs are unrelated to known transcription factors and arrange around the core in a unique fashion. Their structures suggest potential functions during transcription in the chemical environment of chloroplasts. These results reveal structural insights into chloroplast transcription and provide a framework for understanding photosynthesis gene expression.
History
DepositionSep 24, 2023-
Header (metadata) releaseMar 6, 2024-
Map releaseMar 6, 2024-
UpdateMar 20, 2024-
Current statusMar 20, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_18499.png

Downloads & links

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Map

FileDownload / File: emd_18499.map.gz / Format: CCP4 / Size: 325 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMap A, sharpened.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.05 Å/pix.
x 440 pix.
= 462. Å		1.05 Å/pix.
x 440 pix.
= 462. Å		1.05 Å/pix.
x 440 pix.
= 462. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.05 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.216
Minimum - Maximum-0.6956059 - 1.4140964
Average (Standard dev.)0.00044020175 (±0.03412467)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions440440440
Spacing440440440
CellA=B=C: 461.99997 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_18499_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Map A, unsharpened.

Fileemd_18499_additional_1.map
AnnotationMap A, unsharpened.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Map A, half map A.

Fileemd_18499_half_map_1.map
AnnotationMap A, half map A.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Map A, half map B.

Fileemd_18499_half_map_2.map
AnnotationMap A, half map B.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Plastid-encoded DNA-dependent RNA polymerase (PEP)

EntireName: Plastid-encoded DNA-dependent RNA polymerase (PEP)
Components
  • Complex: Plastid-encoded DNA-dependent RNA polymerase (PEP)

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Supramolecule #1: Plastid-encoded DNA-dependent RNA polymerase (PEP)

SupramoleculeName: Plastid-encoded DNA-dependent RNA polymerase (PEP) / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Sinapis alba (white mustard)
Molecular weightTheoretical: 1 MDa

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.6
Component:
ConcentrationNameFormula
50.0 mMTris
0.1 mMEDTA
10.0 mMMagnesium chlorideMgCl2
0.1 mMSodium fluorideNaF
0.1 mMPMSF
1.0 mM2-mercapto ethanol
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average exposure time: 2.7 sec. / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 81000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.43 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 46170
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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