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- EMDB-19771: Cryo-EM structure of Pseudomonas aeruginosa Recombinase A (RecA) ... -

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Basic information

Entry
Database: EMDB / ID: EMD-19771
TitleCryo-EM structure of Pseudomonas aeruginosa Recombinase A (RecA) in complex with LexAS125A mutant
Map data
Sample
  • Complex: Cryo-EM structure of Pseudomonas aeruginosa Recombinase A (RecA) in complex with LexAS125A mutant
    • Complex: Recombinase A (RecA) and LexA repressor
      • Protein or peptide: LexA repressor
      • Protein or peptide: Protein RecA
    • Complex: ssDNA
      • DNA: DNA (36-MER)
  • Ligand: MAGNESIUM ION
  • Ligand: PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER
Keywordsprotease / antibiotic resistance / SOS response / transcription
Function / homology
Function and homology information


repressor LexA / SOS response / DNA-binding transcription repressor activity / ATP-dependent DNA damage sensor activity / protein-DNA complex / single-stranded DNA binding / DNA recombination / sequence-specific DNA binding / damaged DNA binding / DNA replication ...repressor LexA / SOS response / DNA-binding transcription repressor activity / ATP-dependent DNA damage sensor activity / protein-DNA complex / single-stranded DNA binding / DNA recombination / sequence-specific DNA binding / damaged DNA binding / DNA replication / serine-type endopeptidase activity / DNA repair / negative regulation of DNA-templated transcription / regulation of DNA-templated transcription / ATP hydrolysis activity / proteolysis / ATP binding / cytoplasm
Similarity search - Function
LexA repressor, DNA-binding domain / Transcription regulator LexA / LexA DNA binding domain / Peptidase S24, LexA-like / : / LexA-like / Peptidase S24/S26A/S26B/S26C / Peptidase S24-like / LexA/Signal peptidase-like superfamily / DNA recombination/repair protein RecA, conserved site ...LexA repressor, DNA-binding domain / Transcription regulator LexA / LexA DNA binding domain / Peptidase S24, LexA-like / : / LexA-like / Peptidase S24/S26A/S26B/S26C / Peptidase S24-like / LexA/Signal peptidase-like superfamily / DNA recombination/repair protein RecA, conserved site / DNA recombination and repair protein RecA, C-terminal / : / RecA C-terminal domain / recA signature. / DNA recombination and repair protein RecA / : / recA bacterial DNA recombination protein / DNA recombination and repair protein RecA, monomer-monomer interface / RecA family profile 2. / DNA recombination and repair protein RecA-like, ATP-binding domain / RecA family profile 1. / Winged helix DNA-binding domain superfamily / Winged helix-like DNA-binding domain superfamily / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Protein RecA / LexA repressor
Similarity search - Component
Biological speciesPseudomonas aeruginosa (bacteria) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.43 Å
AuthorsDe Felice S / Vascon F / Huber ST / Catalano C / Jakobi AJ / Cendron L
Funding support Italy, 1 items
OrganizationGrant numberCountry
Italian Ministry of Education Italy
CitationJournal: iScience / Year: 2025
Title: Snapshots of SOS response reveal structural requisites for LexA autoproteolysis.
Authors: Filippo Vascon / Sofia De Felice / Matteo Gasparotto / Stefan T Huber / Claudio Catalano / Monica Chinellato / Riccardo Mezzetti / Alessandro Grinzato / Francesco Filippini / Lorenzo Maso / ...Authors: Filippo Vascon / Sofia De Felice / Matteo Gasparotto / Stefan T Huber / Claudio Catalano / Monica Chinellato / Riccardo Mezzetti / Alessandro Grinzato / Francesco Filippini / Lorenzo Maso / Arjen J Jakobi / Laura Cendron /
Abstract: Antimicrobial resistance poses a severe threat to human health and stands out among the pathogens responsible for this emergency. The SOS response to DNA damage is crucial in bacterial evolution, ...Antimicrobial resistance poses a severe threat to human health and stands out among the pathogens responsible for this emergency. The SOS response to DNA damage is crucial in bacterial evolution, influencing resistance development and adaptability in challenging environments, especially under antibiotic exposure. Recombinase A (RecA) and the transcriptional repressor LexA are the key players that orchestrate this process, determining either the silencing or the active transcription of the genes under their control. By integrating state-of-the-art structural approaches with binding and functional assays, we elucidated the molecular events activating the SOS response in , focusing on the RecA-LexA interaction. Our findings identify the conserved determinants and strength of the interactions that allow RecA to trigger LexA autocleavage and inactivation. These results provide the groundwork for designing novel antimicrobial strategies and exploring the potential translation of -derived approaches, to address the implications of infections.
History
DepositionMar 1, 2024-
Header (metadata) releaseJan 15, 2025-
Map releaseJan 15, 2025-
UpdateMar 12, 2025-
Current statusMar 12, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_19771.png

Downloads & links

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Map

FileDownload / File: emd_19771.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
0.84 Å/pix.
x 384 pix.
= 322.56 Å		0.84 Å/pix.
x 384 pix.
= 322.56 Å		0.84 Å/pix.
x 384 pix.
= 322.56 Å

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.84 Å
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Contour LevelBy AUTHOR: 0.119
Minimum - Maximum-0.026637381 - 0.47557095
Average (Standard dev.)0.004331396 (±0.020588089)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 322.56 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_19771_msk_1.map
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Mask #2

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Mask #3

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Mask #4

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Additional map: #2

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Additional map: #1

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Half map: #2

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Half map: #1

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Sample components

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Entire : Cryo-EM structure of Pseudomonas aeruginosa Recombinase A (RecA) ...

EntireName: Cryo-EM structure of Pseudomonas aeruginosa Recombinase A (RecA) in complex with LexAS125A mutant
Components
  • Complex: Cryo-EM structure of Pseudomonas aeruginosa Recombinase A (RecA) in complex with LexAS125A mutant
    • Complex: Recombinase A (RecA) and LexA repressor
      • Protein or peptide: LexA repressor
      • Protein or peptide: Protein RecA
    • Complex: ssDNA
      • DNA: DNA (36-MER)
  • Ligand: MAGNESIUM ION
  • Ligand: PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER

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Supramolecule #1: Cryo-EM structure of Pseudomonas aeruginosa Recombinase A (RecA) ...

SupramoleculeName: Cryo-EM structure of Pseudomonas aeruginosa Recombinase A (RecA) in complex with LexAS125A mutant
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3

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Supramolecule #2: Recombinase A (RecA) and LexA repressor

SupramoleculeName: Recombinase A (RecA) and LexA repressor / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#2
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)

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Supramolecule #3: ssDNA

SupramoleculeName: ssDNA / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #3
Source (natural)Organism: synthetic construct (others)

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Macromolecule #1: LexA repressor

MacromoleculeName: LexA repressor / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO / EC number: repressor LexA
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Molecular weightTheoretical: 23.407885 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MHHHHHHGQK LTPRQAEILS FIKRCLEDHG FPPTRAEIAQ ELGFKSPNAA EEHLKALARK GAIEMTPGAS RGIRIPGFEP HAANDDEGL PVIGRVAAGA PILAEQNIEE SCRINPAFFN PRADYLLRVR GMAMKDIGIL DGDLLAVHVT REARNGQVVV A RIGEEVTV ...String:
MHHHHHHGQK LTPRQAEILS FIKRCLEDHG FPPTRAEIAQ ELGFKSPNAA EEHLKALARK GAIEMTPGAS RGIRIPGFEP HAANDDEGL PVIGRVAAGA PILAEQNIEE SCRINPAFFN PRADYLLRVR GMAMKDIGIL DGDLLAVHVT REARNGQVVV A RIGEEVTV KRFKREGSKV WLLAENPEFA PIEVDLKEQE LIIEGLSVGV IRR

UniProtKB: LexA repressor

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Macromolecule #2: Protein RecA

MacromoleculeName: Protein RecA / type: protein_or_peptide / ID: 2 / Number of copies: 11 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Molecular weightTheoretical: 38.845184 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MHHHHHHKLE NLYFQGDENK KRALAAALGQ IERQFGKGAV MRMGDHERQA IPAISTGSLG LDIALGIGGL PKGRIVEIYG PESSGKTTL TLSVIAEAQK QGATCAFVDA EHALDPDYAG KLGVNVDDLL VSQPDTGEQA LEITDMLVRS NAVDVIIVDS V AALVPKAE ...String:
MHHHHHHKLE NLYFQGDENK KRALAAALGQ IERQFGKGAV MRMGDHERQA IPAISTGSLG LDIALGIGGL PKGRIVEIYG PESSGKTTL TLSVIAEAQK QGATCAFVDA EHALDPDYAG KLGVNVDDLL VSQPDTGEQA LEITDMLVRS NAVDVIIVDS V AALVPKAE IEGEMGDAHV GLQARLMSQA LRKITGNIKN ANCLVIFINQ IRMKIGVMFG NPETTTGGNA LKFYASVRLD IR RTGAVKE GDEVVGSETR VKVVKNKVSP PFRQAEFQIL YGKGIYRTGE IIDLGVQLGL VEKSGAWYSY QGSKIGQGKA NAA KYLEDN PEIGSVLEKT IRDQLLAKSG PVKADAEEVA DAEAD

UniProtKB: Protein RecA

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Macromolecule #3: DNA (36-MER)

MacromoleculeName: DNA (36-MER) / type: dna / ID: 3 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 10.905983 KDa
SequenceString:
(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)

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Macromolecule #4: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 11 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #5: PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER

MacromoleculeName: PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER / type: ligand / ID: 5 / Number of copies: 11 / Formula: AGS
Molecular weightTheoretical: 523.247 Da
Chemical component information

ChemComp-AGS:
PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER / ATP-gamma-S, energy-carrying molecule analogue*YM

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation statehelical array

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Sample preparation

BufferpH: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 55.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 561719
Startup modelType of model: INSILICO MODEL
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.43 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.2.1) / Number images used: 164165
Initial angle assignmentType: PROJECTION MATCHING
Final angle assignmentType: PROJECTION MATCHING
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementProtocol: RIGID BODY FIT
Output model

PDB-8s7g:
Cryo-EM structure of Pseudomonas aeruginosa Recombinase A (RecA) in complex with LexAS125A mutant

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