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- EMDB-53820: Pseudomonas putida Pore-Forming Toxin Tke5 in complex with its co... -

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Basic information

Entry
Database: EMDB / ID: EMD-53820
TitlePseudomonas putida Pore-Forming Toxin Tke5 in complex with its cognate Type VI adaptor protein Tap3
Map data
Sample
  • Complex: Pseudomonas putida Pore-Forming Toxin Tke5 in complex with its cognate Type VI adaptor protein Tap3
    • Protein or peptide: Toxin VasX N-terminal region domain-containing protein
    • Protein or peptide: DUF4123 domain-containing protein
KeywordsPseudomonas putida T6SS effector / Pore-Forming Toxin / MIX family domain / adaptor protein / TOXIN
Function / homologyDomain of unknown function DUF4123 / : / Domain of unknown function (DUF4123) / Toxin VasX, N-terminal region / VasX toxin N-terminal region / membrane / DUF4123 domain-containing protein / Toxin VasX N-terminal region domain-containing protein
Function and homology information
Biological speciesPseudomonas putida KT2440 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.77 Å
AuthorsVelazquez C / Zabala-Zearreta M / Altuna-Alvarez J / Albesa-Jove D
Funding support Spain, 1 items
OrganizationGrant numberCountry
Spanish Ministry of Science, Innovation, and UniversitiesPID2021-127816NB-I00 Spain
CitationJournal: EMBO J / Year: 2026
Title: Structural insights into the antibacterial function of the Pseudomonas putida effector Tke5.
Authors: Carmen Velázquez / Maialen Zabala-Zearreta / Carmen Paredes / Cristina Civantos / Jon Altuna-Alvarez / Patricia Bernal / David Albesa-Jové /
Abstract: Pseudomonas putida is a plant-beneficial rhizobacterium that encodes multiple type-VI secretion systems (T6SS) to outcompete phytopathogens in the rhizosphere. Among its antibacterial effectors, Tke5 ...Pseudomonas putida is a plant-beneficial rhizobacterium that encodes multiple type-VI secretion systems (T6SS) to outcompete phytopathogens in the rhizosphere. Among its antibacterial effectors, Tke5 (a member of the BTH_I2691 protein family) is a potent pore-forming toxin that disrupts ion homeostasis without causing considerable membrane damage. Tke5 harbours an N-terminal MIX domain, which is required for T6SS-dependent secretion in other systems. Many MIX domain-containing effectors require T6SS adaptor proteins (Tap) for secretion, but their molecular mechanisms of adaptor-effector binding remain elusive. Here, we report the 2.8 Å cryo-EM structure of the Tap3-Tke5 complex of P. putida strain KT2440, providing structural and functional insights into how effector Tke5 is recruited by its cognate adaptor protein Tap3. Functional dissection shows that the α-helical region of Tke5 is sufficient to kill intoxicated bacteria, while its β-rich region likely contributes to target membrane specificity. These findings delineate a mechanism of BTH_I2691 proteins for Tap recruitment and toxin activity, contributing to our understanding of a widespread yet understudied toxin family.
History
DepositionMay 16, 2025-
Header (metadata) releaseJan 14, 2026-
Map releaseJan 14, 2026-
UpdateJan 21, 2026-
Current statusJan 21, 2026Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_53820.png

Downloads & links

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Map

FileDownload / File: emd_53820.map.gz / Format: CCP4 / Size: 824 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 0.6462 Å
Density
Contour LevelBy AUTHOR: 0.0397
Minimum - Maximum-0.08526547 - 0.17399554
Average (Standard dev.)-0.0001643626 (±0.0021701816)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions600600600
Spacing600600600
CellA=B=C: 387.72 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_53820_msk_1.map
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Additional map: #1

Fileemd_53820_additional_1.map
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Half map: #1

Fileemd_53820_half_map_1.map
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Half map: #2

Fileemd_53820_half_map_2.map
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Sample components

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Entire : Pseudomonas putida Pore-Forming Toxin Tke5 in complex with its co...

EntireName: Pseudomonas putida Pore-Forming Toxin Tke5 in complex with its cognate Type VI adaptor protein Tap3
Components
  • Complex: Pseudomonas putida Pore-Forming Toxin Tke5 in complex with its cognate Type VI adaptor protein Tap3
    • Protein or peptide: Toxin VasX N-terminal region domain-containing protein
    • Protein or peptide: DUF4123 domain-containing protein

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Supramolecule #1: Pseudomonas putida Pore-Forming Toxin Tke5 in complex with its co...

SupramoleculeName: Pseudomonas putida Pore-Forming Toxin Tke5 in complex with its cognate Type VI adaptor protein Tap3
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Pseudomonas putida KT2440 (bacteria)
Molecular weightTheoretical: 145 KDa

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Macromolecule #1: Toxin VasX N-terminal region domain-containing protein

MacromoleculeName: Toxin VasX N-terminal region domain-containing protein
type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas putida KT2440 (bacteria)
Molecular weightTheoretical: 109.903414 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MTNASVSSAT SGPACSARVP ILPVRYAIVP RTADAPACRY ADAGFNLEQG FPTLQHSAYT LRALRPGYVY VFMKGPRGEK LVIHEYVGA GLYKELRYQG LEDYHRRERY LSGQSTGWVW ADTCQDTAEE VWIGYSPHLW TNAITARITG SAALRKRHMR Q LDMAELVS ...String:
MTNASVSSAT SGPACSARVP ILPVRYAIVP RTADAPACRY ADAGFNLEQG FPTLQHSAYT LRALRPGYVY VFMKGPRGEK LVIHEYVGA GLYKELRYQG LEDYHRRERY LSGQSTGWVW ADTCQDTAEE VWIGYSPHLW TNAITARITG SAALRKRHMR Q LDMAELVS GNQAPSTQPH VLPVSALQTW VEDFKPTERR MPLTWSSDPV TETLPIGNLH AIARHYPWTQ PKVPVVVALA DA EGMALDL SLSVSAYQHQ LRDLMPAEQL EHTKPAQPPE QERVPACYRL DAEQLSPQSR DFHHRNLVAM LLNKTLESLY PAD APSPEL AAFRLGTSRR GPALSPAESH FQALTHEDYS PNGARLAQRL DVDKYRRFLA ERDALERRIA ALRNLALQAS NDHD AWLAT AEPQHIDNPY SLAAALACYD RDERTSARGL EISLALLIHP MGQPTPGTED HDRRFKRLEQ WLDQHDSPLY TALAP FNPF KDKADSIGTL FGASDNVIEG LAGRFPAMAD ITDLTAQSVN TVVLKRLRGQ TRWDASHGLR RQVQLAAREA NAEKAL GLL AARYRITEQA ITENPFSQEV QRYLKSGMAQ VEEMKALRIS GSRTVSIELT TTARAKPNFL GLLTSGGGGG LNAGMLW FN ILSLKTAYNS LQKSDAPEYT LGFASSVFGV IGAAAATLVS VRATQKAMML KWISTAPGMA FGNGIINFLT SNLFARAS G YPAIIMGFSS DIFKAIRQWQ NGNLISGLYT AAASATTALG SILVLEGSLA IAGTTVLIPF AGWTAAGIAL IGATLIGGG LFLYSKAHSR LHSPLELWAS RSIFGTREND GEIRDNIKLD FDKKLPPFRH IHEEIKSWQI EFFSPKALLA PYSEQLGLGA LTTHLDINN KWPTPDWATI IHTQVTKQNA TLELTILLPE FVVGQSEWSG SLVSQNLDRQ TTPLQINPDC HITQAGLLLH Y KTDARQIR SATLTINYYA NQGLDDDTPS TTTIHLER

UniProtKB: Toxin VasX N-terminal region domain-containing protein

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Macromolecule #2: DUF4123 domain-containing protein

MacromoleculeName: DUF4123 domain-containing protein / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas putida KT2440 (bacteria)
Molecular weightTheoretical: 35.710516 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MGSSHHHHHH SMTTLSSPPP RSEDFAHNCR FLLLMTSAIE NWAYRPIPQE GELVPAYAPS VLDLINAVSR NTQKAWLWKN SALDDRHEF GPLLVDVADA HELLAHAITT WMPIGGAIAL DAEVGLAALA DHFTSLVQVV LPDHSVATHH VTPDHLAAWL N ALDDDHRK ...String:
MGSSHHHHHH SMTTLSSPPP RSEDFAHNCR FLLLMTSAIE NWAYRPIPQE GELVPAYAPS VLDLINAVSR NTQKAWLWKN SALDDRHEF GPLLVDVADA HELLAHAITT WMPIGGAIAL DAEVGLAALA DHFTSLVQVV LPDHSVATHH VTPDHLAAWL N ALDDDHRK AWLGPVSRLA WRVNWGPAHE WKTHENKPIA ARSKSELPLT LQQLELDRLQ AGMHEHFVLS LAHEVLAMPP HA NHTLADI RLWIETLLPQ LTALNFRSEE VAGQFIRLIA GHMWLMSNDK AGKIYTNLEE SPQARLRELQ ALVESKEPAH D

UniProtKB: DUF4123 domain-containing protein

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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

Concentration2.2 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
150.0 mMNaClSodium chloride
20.0 mMTris-HClTris (hydroxymethyl) aminomethane (THAM) hydrochloride
2.0 mMDTTDithiothreitol
0.05 %w/vCHAPS3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 12 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 90 sec. / Pretreatment - Pressure: 0.041 kPa
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 277.15 K / Instrument: LEICA EM GP

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 48.8 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.8 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.77 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 381732
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE

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