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- EMDB-27646: Quorum-sensing receptor RhlR bound to PqsE -

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Basic information

Entry
Database: EMDB / ID: EMD-27646
TitleQuorum-sensing receptor RhlR bound to PqsE
Map dataDensity modified full map
Sample
  • Complex: Tetrameric complex of RhlR:mBTL bound to PqsE
    • Protein or peptide: RhlR protein
    • Protein or peptide: 2-aminobenzoylacetyl-CoA thioesterase
    • DNA: DNA (5'-D(*AP*CP*CP*TP*GP*CP*CP*AP*GP*AP*CP*TP*GP*CP*AP*CP*AP*G)-3')
    • DNA: DNA (5'-D(*CP*TP*GP*TP*GP*CP*AP*GP*TP*CP*TP*GP*GP*CP*AP*GP*GP*T)-3')
  • Ligand: 4-(3-bromophenoxy)-N-[(3S)-2-oxothiolan-3-yl]butanamide
Function / homology
Function and homology information


2-aminobenzoylacetyl-CoA thioesterase / secondary metabolite biosynthetic process / beta-lactam antibiotic catabolic process / beta-lactamase 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
RhlR protein / 2-aminobenzoylacetyl-CoA thioesterase
Similarity search - Component
Biological speciesPseudomonas aeruginosa (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.1 Å
AuthorsPaczkowski JE / Fromme JC / Feathers JR
Funding support United States, 4 items
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, 2022-
Header (metadata) releaseDec 7, 2022-
Map releaseDec 7, 2022-
UpdateDec 14, 2022-
Current statusDec 14, 2022Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

Downloads & links

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Map

FileDownload / File: emd_27646.map.gz / Format: CCP4 / Size: 34.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationDensity modified full map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesX (Sec.)Y (Row.)Z (Col.)
1.29 Å/pix.
x 208 pix.
= 268.32 Å
1.29 Å/pix.
x 208 pix.
= 268.32 Å
1.29 Å/pix.
x 208 pix.
= 268.32 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.29 Å
Density
Contour LevelBy AUTHOR: 0.115
Minimum - Maximum-1.6226361 - 2.642822
Average (Standard dev.)8.3034865e-06 (±0.05218911)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions208208208
Spacing208208208
CellA=B=C: 268.32 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Sharpened post process map

Fileemd_27646_additional_1.map
AnnotationSharpened post process map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: Density modified half map 1

Fileemd_27646_additional_2.map
AnnotationDensity modified half map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: Density modified half map 2

Fileemd_27646_additional_3.map
AnnotationDensity modified half map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: Refinement full map

Fileemd_27646_additional_4.map
AnnotationRefinement full map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: Refinement half map 2

Fileemd_27646_half_map_1.map
AnnotationRefinement half map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: Refinement half map 1

Fileemd_27646_half_map_2.map
AnnotationRefinement half map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire : Tetrameric complex of RhlR:mBTL bound to PqsE

EntireName: Tetrameric complex of RhlR:mBTL bound to PqsE
Components
  • Complex: Tetrameric complex of RhlR:mBTL bound to PqsE
    • Protein or peptide: RhlR protein
    • Protein or peptide: 2-aminobenzoylacetyl-CoA thioesterase
    • DNA: DNA (5'-D(*AP*CP*CP*TP*GP*CP*CP*AP*GP*AP*CP*TP*GP*CP*AP*CP*AP*G)-3')
    • DNA: DNA (5'-D(*CP*TP*GP*TP*GP*CP*AP*GP*TP*CP*TP*GP*GP*CP*AP*GP*GP*T)-3')
  • Ligand: 4-(3-bromophenoxy)-N-[(3S)-2-oxothiolan-3-yl]butanamide

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Supramolecule #1: Tetrameric complex of RhlR:mBTL bound to PqsE

SupramoleculeName: Tetrameric complex of RhlR:mBTL bound to PqsE / type: complex / Chimera: Yes / ID: 1 / Parent: 0 / Macromolecule list: #1-#4
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Molecular weightTheoretical: 123 KDa

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Macromolecule #1: RhlR protein

MacromoleculeName: RhlR protein / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Molecular weightTheoretical: 27.611596 KDa
Recombinant expressionOrganism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
SequenceString: MRNDGGFLLW WDGLRSEMQP IHDSQGVFAV LEKEVRRLGF DYYAYGVRHT IPFTRPKTEV HGTYPKAWLE RYQMQNYGAV DPAILNGLR SSEMVVWSDS LFDQSRMLWN EARDWGLCVG ATLPIRAPNN LLSVLSVARD QQNISSFERE EIRLRLRCMI E LLTQKLTD ...String:
MRNDGGFLLW WDGLRSEMQP IHDSQGVFAV LEKEVRRLGF DYYAYGVRHT IPFTRPKTEV HGTYPKAWLE RYQMQNYGAV DPAILNGLR SSEMVVWSDS LFDQSRMLWN EARDWGLCVG ATLPIRAPNN LLSVLSVARD QQNISSFERE EIRLRLRCMI E LLTQKLTD LEHPMLMSNP VCLSHREREI LQWTADGKSS GEIAIILSIS ESTVNFHHKN IQKKFDAPNK TLAAAYAAAL GL I

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Macromolecule #2: 2-aminobenzoylacetyl-CoA thioesterase

MacromoleculeName: 2-aminobenzoylacetyl-CoA thioesterase / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Molecular weightTheoretical: 34.350297 KDa
Recombinant expressionOrganism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
SequenceString: MLRLSAPGQL DDDLCLLGDV QVPVFLLRLG EASWALVEGG ISRDAELVWA DLCRWVADPS QVHYWLITHK HYDHCGLLPY LCPRLPNVQ VLASERTCQA WKSESAVRVV ERLNRQLLRA EQRLPEACAW DALPVRAVAD GEWLELGPRH RLQVIEAHGH S DDHVVFYD ...String:
MLRLSAPGQL DDDLCLLGDV QVPVFLLRLG EASWALVEGG ISRDAELVWA DLCRWVADPS QVHYWLITHK HYDHCGLLPY LCPRLPNVQ VLASERTCQA WKSESAVRVV ERLNRQLLRA EQRLPEACAW DALPVRAVAD GEWLELGPRH RLQVIEAHGH S DDHVVFYD VRRRRLFCGD ALGEFDEAEG VWRPLVFDDM EAYLESLERL QRLPTLLQLI PGHGGLLRGR LAADGAESAY TE CLRLCRR LLWRQSMGES LDELSEELHR AWGGQSVDFL PGELHLGSMR RMLEILSRQA LPLD

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Macromolecule #3: DNA (5'-D(*AP*CP*CP*TP*GP*CP*CP*AP*GP*AP*CP*TP*GP*CP*AP*CP*AP*G)-3')

MacromoleculeName: DNA (5'-D(*AP*CP*CP*TP*GP*CP*CP*AP*GP*AP*CP*TP*GP*CP*AP*CP*AP*G)-3')
type: dna / ID: 3 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Molecular weightTheoretical: 5.470558 KDa
SequenceString:
(DA)(DC)(DC)(DT)(DG)(DC)(DC)(DA)(DG)(DA) (DC)(DT)(DG)(DC)(DA)(DC)(DA)(DG)

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Macromolecule #4: DNA (5'-D(*CP*TP*GP*TP*GP*CP*AP*GP*TP*CP*TP*GP*GP*CP*AP*GP*GP*T)-3')

MacromoleculeName: DNA (5'-D(*CP*TP*GP*TP*GP*CP*AP*GP*TP*CP*TP*GP*GP*CP*AP*GP*GP*T)-3')
type: dna / ID: 4 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Molecular weightTheoretical: 5.563588 KDa
SequenceString:
(DC)(DT)(DG)(DT)(DG)(DC)(DA)(DG)(DT)(DC) (DT)(DG)(DG)(DC)(DA)(DG)(DG)(DT)

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Macromolecule #5: 4-(3-bromophenoxy)-N-[(3S)-2-oxothiolan-3-yl]butanamide

MacromoleculeName: 4-(3-bromophenoxy)-N-[(3S)-2-oxothiolan-3-yl]butanamide
type: ligand / ID: 5 / Number of copies: 2 / Formula: K5G
Molecular weightTheoretical: 358.251 Da
Chemical component information

ChemComp-K5G:
4-(3-bromophenoxy)-N-[(3S)-2-oxothiolan-3-yl]butanamide

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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

Concentration5.0 mg/mL
BufferpH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 1.5 µm / Nominal defocus min: 0.8 µm
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Startup modelType of model: EMDB MAP
EMDB ID:
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 4.0-beta)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 4.1 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 84842
FSC plot (resolution estimation)

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