Journal: Nat Commun / Year: 2019 Title: Structural basis for transcription antitermination at bacterial intrinsic terminator. Authors: Linlin You / Jing Shi / Liqiang Shen / Lingting Li / Chengli Fang / Chengzhi Yu / Wenbo Cheng / Yu Feng / Yu Zhang / Abstract: Bacteriophages typically hijack the host bacterial transcriptional machinery to regulate their own gene expression and that of the host bacteria. The structural basis for bacteriophage protein- ...Bacteriophages typically hijack the host bacterial transcriptional machinery to regulate their own gene expression and that of the host bacteria. The structural basis for bacteriophage protein-mediated transcription regulation-in particular transcription antitermination-is largely unknown. Here we report the 3.4 Å and 4.0 Å cryo-EM structures of two bacterial transcription elongation complexes (P7-NusA-TEC and P7-TEC) comprising the bacteriophage protein P7, a master host-transcription regulator encoded by bacteriophage Xp10 of the rice pathogen Xanthomonas oryzae pv. Oryzae (Xoo) and discuss the mechanisms by which P7 modulates the host bacterial RNAP. The structures together with biochemical evidence demonstrate that P7 prevents transcription termination by plugging up the RNAP RNA-exit channel and impeding RNA-hairpin formation at the intrinsic terminator. Moreover, P7 inhibits transcription initiation by restraining RNAP-clamp motions. Our study reveals the structural basis for transcription antitermination by phage proteins and provides insights into bacterial transcription regulation.
History
Deposition
Jan 22, 2019
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Header (metadata) release
Jul 17, 2019
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Map release
Jul 17, 2019
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Update
Mar 27, 2024
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Current status
Mar 27, 2024
Processing site: PDBj / Status: Released
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Structure visualization
Movie
Surface view with section colored by density value
Name: RNA (5'-R(P*AP*GP*CP*GP*GP*AP*GP*AP*GP*GP*UP*A)-3') / type: rna / ID: 7 / Number of copies: 1
Source (natural)
Organism: synthetic construct (others)
Molecular weight
Theoretical: 6.509968 KDa
Sequence
String:
GCAUUCAAAG CGGAGAGGUA
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Macromolecule #9: MAGNESIUM ION
Macromolecule
Name: MAGNESIUM ION / type: ligand / ID: 9 / Number of copies: 1 / Formula: MG
Molecular weight
Theoretical: 24.305 Da
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Macromolecule #10: ZINC ION
Macromolecule
Name: ZINC ION / type: ligand / ID: 10 / Number of copies: 2 / Formula: ZN
Molecular weight
Theoretical: 65.409 Da
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Experimental details
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Structure determination
Method
cryo EM
Processing
single particle reconstruction
Aggregation state
particle
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Sample preparation
Concentration
9.5 mg/mL
Buffer
pH: 7.5 Component:
Concentration
Formula
Name
10.0 mM
Hepes
Hepes
50.0 mM
KCL
Potassium Chloride
5.0 mM
MgCL2
Magnesium chloride
3.0 mM
DTT
DL-Dithiothreitol
Grid
Model: C-flat-1.2/1.3 4C / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 120 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 120 sec. / Pretreatment - Atmosphere: OTHER
Vitrification
Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 283 K / Instrument: FEI VITROBOT MARK IV / Details: blot for 8 seconds before plunging.
Details
This sample was monodisperse
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Electron microscopy
Microscope
FEI TITAN KRIOS
Image recording
Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Number grids imaged: 1 / Number real images: 2271 / Average exposure time: 8.0 sec. / Average electron dose: 1.675 e/Å2
Electron beam
Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
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