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- EMDB-33034: Focused map on the aCTD-Zur region of the Streptomyces coelicolor... -

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Basic information

Entry
Database: EMDB / ID: EMD-33034
TitleFocused map on the aCTD-Zur region of the Streptomyces coelicolor RNAP-promoter open complex with one Zur dimer
Map data
Sample
  • Complex: Focused map on the aCTD-Zur region of the Streptomyces coelicolor RNAP-promoter open complex with one Zur dimer
Biological speciesStreptomyces coelicolor A3(2) (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 10.5 Å
AuthorsYang X / Zheng J
Funding support China, 1 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)31770068, 32070040 China
CitationJournal: Nucleic Acids Res / Year: 2022
Title: Structural basis of Streptomyces transcription activation by zinc uptake regulator.
Authors: Xu Yang / Yiqun Wang / Guiyang Liu / Zixin Deng / Shuangjun Lin / Jianting Zheng /
Abstract: Streptomyces coelicolor (Sc) is a model organism of actinobacteria to study morphological differentiation and production of bioactive metabolites. Sc zinc uptake regulator (Zur) affects both ...Streptomyces coelicolor (Sc) is a model organism of actinobacteria to study morphological differentiation and production of bioactive metabolites. Sc zinc uptake regulator (Zur) affects both processes by controlling zinc homeostasis. It activates transcription by binding to palindromic Zur-box sequences upstream of -35 elements. Here we deciphered the molecular mechanism by which ScZur interacts with promoter DNA and Sc RNA polymerase (RNAP) by cryo-EM structures and biochemical assays. The ScZur-DNA structures reveal a sequential and cooperative binding of three ScZur dimers surrounding a Zur-box spaced 8 nt upstream from a -35 element. The ScRNAPσHrdB-Zur-DNA structures define protein-protein and protein-DNA interactions involved in the principal housekeeping σHrdB-dependent transcription initiation from a noncanonical promoter with a -10 element lacking the critical adenine residue at position -11 and a TTGCCC -35 element deviating from the canonical TTGACA motif. ScZur interacts with the C-terminal domain of ScRNAP α subunit (αCTD) in a complex structure trapped in an active conformation. Key ScZur-αCTD interfacial residues accounting for ScZur-dependent transcription activation were confirmed by mutational studies. Together, our structural and biochemical results provide a comprehensive model for transcription activation of Zur family regulators.
History
DepositionMar 9, 2022-
Header (metadata) releaseAug 3, 2022-
Map releaseAug 3, 2022-
UpdateAug 24, 2022-
Current statusAug 24, 2022Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_33034.png

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Map

FileDownload / File: emd_33034.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.05 Å
Density
Contour LevelBy AUTHOR: 0.04
Minimum - Maximum-0.11925851 - 0.3909289
Average (Standard dev.)-0.0011323154 (±0.007329301)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 403.19998 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_33034_msk_1.map
Projections & Slices
AxesZYX

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Histogram

Histogram (log scale)

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Half map: #1

Fileemd_33034_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_33034_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Focused map on the aCTD-Zur region of the Streptomyces coelicolor...

EntireName: Focused map on the aCTD-Zur region of the Streptomyces coelicolor RNAP-promoter open complex with one Zur dimer
Components
  • Complex: Focused map on the aCTD-Zur region of the Streptomyces coelicolor RNAP-promoter open complex with one Zur dimer

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Supramolecule #1: Focused map on the aCTD-Zur region of the Streptomyces coelicolor...

SupramoleculeName: Focused map on the aCTD-Zur region of the Streptomyces coelicolor RNAP-promoter open complex with one Zur dimer
type: complex / Chimera: Yes / ID: 1 / Parent: 0 / Macromolecule list: #1-#8
Source (natural)Organism: Streptomyces coelicolor A3(2) (bacteria)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
Molecular weightExperimental: 560 KDa

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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

Concentration1.0 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
20.0 mMHEPESHEPES
100.0 mMKClKCl
2.0 mMDTTDTT
5.0 mMMgCl2MgCl2
0.02 mMZnSO4ZnSO4
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 289 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Image recordingFilm or detector model: GATAN K2 QUANTUM (4k x 4k) / Average electron dose: 40.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 10.5 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 19427
FSC plot (resolution estimation)

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