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- PDB-8dq0: Quorum-sensing receptor RhlR bound to PqsE -

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Basic information

Entry
Database: PDB / ID: 8dq0
TitleQuorum-sensing receptor RhlR bound to PqsE
Components
  • 2-aminobenzoylacetyl-CoA thioesterase
  • RhlR protein
KeywordsTRANSCRIPTION / transcriptional regulators / quorum sensing / DNA binding
Function / homology
Function and homology information


2-aminobenzoylacetyl-CoA thioesterase / secondary metabolite biosynthetic process / hydrolase activity / regulation of DNA-templated transcription / DNA binding / metal ion binding
Similarity search - Function
Transcription factor LuxR-like, autoinducer-binding domain / Transcription factor LuxR-like, autoinducer-binding domain superfamily / Autoinducer binding domain / LuxR-type HTH domain signature. / LuxR-type HTH domain profile. / Transcription regulator LuxR, C-terminal / Bacterial regulatory proteins, luxR family / helix_turn_helix, Lux Regulon / Signal transduction response regulator, C-terminal effector / Metallo-beta-lactamase superfamily ...Transcription factor LuxR-like, autoinducer-binding domain / Transcription factor LuxR-like, autoinducer-binding domain superfamily / Autoinducer binding domain / LuxR-type HTH domain signature. / LuxR-type HTH domain profile. / Transcription regulator LuxR, C-terminal / Bacterial regulatory proteins, luxR family / helix_turn_helix, Lux Regulon / Signal transduction response regulator, C-terminal effector / Metallo-beta-lactamase superfamily / Metallo-beta-lactamase superfamily / Metallo-beta-lactamase / Ribonuclease Z/Hydroxyacylglutathione hydrolase-like / Winged helix-like DNA-binding domain superfamily
Similarity search - Domain/homology
Chem-K5G / RhlR protein / 2-aminobenzoylacetyl-CoA thioesterase
Similarity search - Component
Biological speciesPseudomonas aeruginosa (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.74 Å
AuthorsPaczkowski, J.E. / Fromme, J.C. / Feathers, J.R.
Funding support United States, 4items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)1R01GM14436101 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM136258 United States
Cystic Fibrosis FoundationPACZKO21G0 United States
National Science Foundation (NSF, United States)DMR-1719875 United States
CitationJournal: Structure / Year: 2022
Title: Structure of the RhlR-PqsE complex from Pseudomonas aeruginosa reveals mechanistic insights into quorum-sensing gene regulation.
Authors: J Ryan Feathers / Erica K Richael / Kayla A Simanek / J Christopher Fromme / Jon E Paczkowski /
Abstract: Pseudomonas aeruginosa is an opportunistic pathogen that is responsible for thousands of deaths every year in the United States. P. aeruginosa virulence factor production is mediated by quorum ...Pseudomonas aeruginosa is an opportunistic pathogen that is responsible for thousands of deaths every year in the United States. P. aeruginosa virulence factor production is mediated by quorum sensing, a mechanism of bacterial cell-cell communication that relies on the production and detection of signal molecules called autoinducers. In P. aeruginosa, the transcription factor receptor RhlR is activated by a RhlI-synthesized autoinducer. We recently showed that RhlR-dependent transcription is enhanced by a physical interaction with the enzyme PqsE via increased affinity of RhlR for promoter DNA. However, the molecular basis for complex formation and how complex formation enhanced RhlR transcriptional activity remained unclear. Here, we report the structure of ligand-bound RhlR in complex with PqsE. Additionally, we determined the structure of the complex bound with DNA, revealing the mechanism by which RhlR-mediated transcription is enhanced by PqsE, thereby establishing the molecular basis for RhlR-dependent virulence factor production in P. aeruginosa.
History
DepositionJul 18, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Dec 7, 2022Provider: repository / Type: Initial release
Revision 1.1Dec 14, 2022Group: Database references / Category: citation / Item: _citation.journal_volume / _citation.page_first

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
C: RhlR protein
D: RhlR protein
A: 2-aminobenzoylacetyl-CoA thioesterase
B: 2-aminobenzoylacetyl-CoA thioesterase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)124,6406
Polymers123,9244
Non-polymers7172
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein RhlR protein / Transcriptional regulator / Transcriptional regulator RhlR / Transcriptional regulator SdiA / ...Transcriptional regulator / Transcriptional regulator RhlR / Transcriptional regulator SdiA / regulatory protein RhlR


Mass: 27611.596 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pseudomonas aeruginosa (bacteria)
Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
References: UniProt: A9JPX4
#2: Protein 2-aminobenzoylacetyl-CoA thioesterase


Mass: 34350.297 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pseudomonas aeruginosa (bacteria) / Gene: pqsE, PA1000
Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
References: UniProt: P20581, 2-aminobenzoylacetyl-CoA thioesterase
#3: Chemical ChemComp-K5G / 4-(3-bromophenoxy)-N-[(3S)-2-oxothiolan-3-yl]butanamide


Mass: 358.251 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C14H16BrNO3S / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY

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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: Tetrameric complex of RhlR:mBTL bound to PqsE / Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT
Molecular weightValue: .123 MDa / Experimental value: NO
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Source (recombinant)Organism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
Buffer solutionpH: 8
SpecimenConc.: 5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
MicroscopyModel: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 1500 nm / Nominal defocus min: 800 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.18.2_3874: / Classification: refinement
EM software
IDNameVersionCategory
2SerialEMimage acquisition
4GctfCTF correction
10cryoSPARCinitial Euler assignment
11RELION4.0-betafinal Euler assignment
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.74 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 40761 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0058889
ELECTRON MICROSCOPYf_angle_d1.02112057
ELECTRON MICROSCOPYf_dihedral_angle_d7.1931222
ELECTRON MICROSCOPYf_chiral_restr0.1291302
ELECTRON MICROSCOPYf_plane_restr0.0061565

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