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- PDB-8e6w: Escherichia coli Rho-dependent transcription pre-termination comp... -

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Basic information

Entry
Database: PDB / ID: 8e6w
TitleEscherichia coli Rho-dependent transcription pre-termination complex containing 18 nt long RNA spacer, lambda-tR1 rut RNA, Mg-ADP-BeF3, and NusG; Rho part
Components
  • Transcription termination factor Rho
  • lambda-tR1 rut RNA
KeywordsTRANSCRIPTION/RNA / factor-dependent termination / Rho / transcription termination / transcription elongation complex / helicase / ATPase / TRANSCRIPTION-RNA complex
Function / homology
Function and homology information


ATP-dependent activity, acting on RNA / helicase activity / DNA-templated transcription termination / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / ATP hydrolysis activity / RNA binding / ATP binding / cytosol
Similarity search - Function
Transcription termination factor Rho / Rho termination factor, N-terminal / Rho termination factor, RNA-binding domain / Transcription termination factor Rho, ATP binding domain / Rho termination factor, RNA-binding domain / Rho termination factor, N-terminal domain / Rho RNA-binding domain profile. / Rho termination factor, N-terminal domain / Rho termination factor, N-terminal domain superfamily / Cold shock domain ...Transcription termination factor Rho / Rho termination factor, N-terminal / Rho termination factor, RNA-binding domain / Transcription termination factor Rho, ATP binding domain / Rho termination factor, RNA-binding domain / Rho termination factor, N-terminal domain / Rho RNA-binding domain profile. / Rho termination factor, N-terminal domain / Rho termination factor, N-terminal domain superfamily / Cold shock domain / Cold shock protein domain / ATPase, F1/V1/A1 complex, alpha/beta subunit, nucleotide-binding domain / ATP synthase alpha/beta family, nucleotide-binding domain / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / Nucleic acid-binding, OB-fold / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
ADENOSINE-5'-DIPHOSPHATE / BERYLLIUM TRIFLUORIDE ION / : / RNA / RNA (> 10) / Transcription termination factor Rho
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
synthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.27 Å
AuthorsMolodtsov, V. / Wang, C. / Ebright, R.H.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS) United States
Citation
Journal: Nature / Year: 2023
Title: Structural basis of Rho-dependent transcription termination.
Authors: Vadim Molodtsov / Chengyuan Wang / Emre Firlar / Jason T Kaelber / Richard H Ebright /
Abstract: Rho is a ring-shaped hexameric ATP-dependent molecular motor. Together with the transcription elongation factor NusG, Rho mediates factor-dependent transcription termination and transcription- ...Rho is a ring-shaped hexameric ATP-dependent molecular motor. Together with the transcription elongation factor NusG, Rho mediates factor-dependent transcription termination and transcription-translation-coupling quality control in Escherichia coli. Here we report the preparation of complexes that are functional in factor-dependent transcription termination from Rho, NusG, RNA polymerase (RNAP), and synthetic nucleic acid scaffolds, and we report cryogenic electron microscopy structures of the complexes. The structures show that functional factor-dependent pre-termination complexes contain a closed-ring Rho hexamer; have RNA threaded through the central channel of Rho; have 60 nucleotides of RNA interacting sequence-specifically with the exterior of Rho and 6 nucleotides of RNA interacting sequence-specifically with the central channel of Rho; have Rho oriented relative to RNAP such that ATP-dependent translocation by Rho exerts mechanical force on RNAP; and have NusG bridging Rho and RNAP. The results explain five decades of research on Rho and provide a foundation for understanding Rho's function.
#1: Journal: Biorxiv / Year: 2022
Title: Structural basis of Rho-dependent transcription termination
Authors: Molodtsov, V. / Wang, C. / Firlar, E. / Kaelber, J.T. / Ebright, R.H.
History
DepositionAug 23, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Sep 7, 2022Provider: repository / Type: Initial release
Revision 1.1Jan 25, 2023Group: Database references / Category: citation / citation_author
Revision 1.2Feb 8, 2023Group: Database references / Category: citation / Item: _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.3Feb 22, 2023Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
8: lambda-tR1 rut RNA
e: Transcription termination factor Rho
f: Transcription termination factor Rho
c: Transcription termination factor Rho
b: Transcription termination factor Rho
a: Transcription termination factor Rho
d: Transcription termination factor Rho
hetero molecules


Theoretical massNumber of molelcules
Total (without water)304,58925
Polymers301,4837
Non-polymers3,10518
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: RNA chain lambda-tR1 rut RNA


Mass: 19062.428 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) / References: GenBank: 1227111474
#2: Protein
Transcription termination factor Rho / ATP-dependent helicase Rho


Mass: 47070.168 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Gene: rho, EL79_4648, EL80_4555 / Production host: Escherichia coli (E. coli)
References: UniProt: A0A0A0GPI6, Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement
#3: Chemical
ChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE / Adenosine diphosphate


Mass: 427.201 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C10H15N5O10P2 / Comment: ADP, energy-carrying molecule*YM
#4: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: Mg
#5: Chemical
ChemComp-BEF / BERYLLIUM TRIFLUORIDE ION


Mass: 66.007 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: BeF3
Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Escherichia coli Rho-dependent transcription pre-termination complex containing 18 nt long RNA spacer, lambda-tR1 rut RNA, Mg-ADP-BeF3, and NusG; Rho part
Type: COMPLEX / Entity ID: #1-#2 / Source: MULTIPLE SOURCES
Source (natural)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 7.9
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2500 nm / Nominal defocus min: 1250 nm
Image recordingElectron dose: 35 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 4.27 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 69316 / Symmetry type: POINT

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