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- EMDB-38410: Cryo-EM structure of colibactin assembly line polyketide synthase... -

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Basic information

Entry
Database: EMDB / ID: EMD-38410
TitleCryo-EM structure of colibactin assembly line polyketide synthase ClbI KS-AT didomain crosslinked with ClbI ACP
Map data
Sample
  • Complex: ACP-bound ClbI
    • Protein or peptide: Polyketide synthase
    • Protein or peptide: Polyketide synthase
Keywordscolibactin / microbiome / polyketide synthase / colorectal cancer / NRPS-PKS hybrid / BIOSYNTHETIC PROTEIN / TRANSFERASE
Function / homology
Function and homology information


beta-ketoacyl-[acyl-carrier-protein] synthase I / fatty acid synthase activity / 3-oxoacyl-[acyl-carrier-protein] synthase activity / fatty acid biosynthetic process
Similarity search - Function
Polyketide synthase, C-terminal extension / Ketoacyl-synthetase C-terminal extension / Malonyl-CoA ACP transacylase, ACP-binding / : / Acyl transferase / Acyl transferase domain / Acyl transferase domain in polyketide synthase (PKS) enzymes. / Acyl transferase domain superfamily / Acyl transferase/acyl hydrolase/lysophospholipase / Beta-ketoacyl synthase ...Polyketide synthase, C-terminal extension / Ketoacyl-synthetase C-terminal extension / Malonyl-CoA ACP transacylase, ACP-binding / : / Acyl transferase / Acyl transferase domain / Acyl transferase domain in polyketide synthase (PKS) enzymes. / Acyl transferase domain superfamily / Acyl transferase/acyl hydrolase/lysophospholipase / Beta-ketoacyl synthase / Ketosynthase family 3 (KS3) domain profile. / Beta-ketoacyl synthase, N-terminal / Beta-ketoacyl synthase, C-terminal / Polyketide synthase, beta-ketoacyl synthase domain / Beta-ketoacyl synthase, N-terminal domain / Beta-ketoacyl synthase, C-terminal domain / Thiolase-like / Phosphopantetheine attachment site / ACP-like superfamily / Carrier protein (CP) domain profile. / Phosphopantetheine binding ACP domain / Alpha/Beta hydrolase fold
Similarity search - Domain/homology
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.29 Å
AuthorsKim M / Kim J / Kang JY
Funding support Korea, Republic Of, 2 items
OrganizationGrant numberCountry
National Research Foundation (NRF, Korea)NRF- 2019M3E5D6063908 Korea, Republic Of
National Research Foundation (NRF, Korea)NRF -2020R1F1A107746211 Korea, Republic Of
CitationJournal: Structure / Year: 2025
Title: Structural study on human microbiome-derived polyketide synthases that assemble genotoxic colibactin.
Authors: Minjae Kim / Jinwoo Kim / Gyu Sung Lee / Paul Dominic B Olinares / Yougant Airan / Jasmine L Chow / Jongseok Park / Yujin Jeong / Jiho Park / Brian T Chait / Seth B Herzon / Chung Sub Kim / Jin Young Kang /
Abstract: Colibactin, a human microbiome-derived genotoxin, promotes colorectal cancer by damaging the host gut epithelial genomes. While colibactin is synthesized via a hybrid non-ribosomal peptide synthetase ...Colibactin, a human microbiome-derived genotoxin, promotes colorectal cancer by damaging the host gut epithelial genomes. While colibactin is synthesized via a hybrid non-ribosomal peptide synthetase (NRPS)-polyketide synthase (PKS) pathway, known as pks or clb, the structural details of its biosynthetic enzymes remain limited, hindering our understanding of its biosynthesis and clinical application. In this study, we report the cryo-EM structures of two colibactin-producing PKS enzymes, ClbC and ClbI, captured in different reaction states using a substrate-mimic crosslinker. Our structural analysis revealed the binding sites of carrier protein (CP) domains of the ClbC and ClbI on their ketosynthase (KS) domains. Further, we identified a novel NRPS-PKS docking interaction between ClbI and its upstream enzyme, ClbH, mediated by the C-terminal peptide ClbH and the dimeric interface of ClbI, establishing a 1:2 stoichiometry. These findings advance our understanding of colibactin assembly line and provide broader insights into NRPS-PKS natural product biosynthesis mechanisms.
History
DepositionDec 22, 2023-
Header (metadata) releaseMay 21, 2025-
Map releaseMay 21, 2025-
UpdateMay 28, 2025-
Current statusMay 28, 2025Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_38410.png

Downloads & links

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Map

FileDownload / File: emd_38410.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.07 Å/pix.
x 256 pix.
= 273.152 Å		1.07 Å/pix.
x 256 pix.
= 273.152 Å		1.07 Å/pix.
x 256 pix.
= 273.152 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.067 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.023
Minimum - Maximum-0.11626997 - 0.20576091
Average (Standard dev.)0.00014233218 (±0.0043530744)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 273.152 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_38410_msk_1.map
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Half map: #2

Fileemd_38410_half_map_1.map
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Half map: #1

Fileemd_38410_half_map_2.map
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Sample components

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Entire : ACP-bound ClbI

EntireName: ACP-bound ClbI
Components
  • Complex: ACP-bound ClbI
    • Protein or peptide: Polyketide synthase
    • Protein or peptide: Polyketide synthase

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Supramolecule #1: ACP-bound ClbI

SupramoleculeName: ACP-bound ClbI / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 110 KDa

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Macromolecule #1: Polyketide synthase

MacromoleculeName: Polyketide synthase / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO / EC number: beta-ketoacyl-[acyl-carrier-protein] synthase I
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 99.125695 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MAENDFGIAI IGMAGRFPQA DTVQAFWENL LASRECISFY SDEELLAMGI SPEFVQHPDY VKAKGEVADI DKFDAAFFGI APREAELMD PQHRVLLETA WAAFEDAGYV AADYPGDVGI FAGKSMDSYL MLNLMPHFKR VFSSGSLQAA IGNDKDSITT T IAYHLNLR ...String:
MAENDFGIAI IGMAGRFPQA DTVQAFWENL LASRECISFY SDEELLAMGI SPEFVQHPDY VKAKGEVADI DKFDAAFFGI APREAELMD PQHRVLLETA WAAFEDAGYV AADYPGDVGI FAGKSMDSYL MLNLMPHFKR VFSSGSLQAA IGNDKDSITT T IAYHLNLR GPAITVQTSS STSLVAVCVA CQSLLTWQCD MAIAGGVTLG PPAKTGYLSQ EGGITAADGH CRAFSDNSSG FV PGTGAGL VVLKRVDEAL RDGDNIYAVI KGFAVNNDGS EKISYTAPSV DAQARAIAQA QRLAGLTPQD ITYVEAHGTG TRL GDPVEF SALSQAFAGA SQKQYCALGS VKTNIGHLDT AAGVAGLIKT ALAVQQGIIP ATLHFERPNA QIDLTNSPFY INTT CQPWQ PESGIRRAGV TSLGMGGTNA HVVLEQAPAV DLQARAPVPA YSILPFSAKT DSALSSGLAR FADFLQHESL PDRRD LAWT LSQGRKAFAH RAALVTRDLH AAGTLLQQAA TAPFARGVAQ TQLGLGLLFS GQGSQYQRMG HQLYQVWPAY ADAFDR CAT LLEREYQLDI RHELFRAEVS LAQGERLAQT CLTQPLLFSV EYALAQLWLS WGITPTVMIG HSLGEWVAAT LAGVFSL ED ALRLVARRAE LMHQAPSGAM LMVALPEAQI RALITAPLAI AAVNAPDYSV IAGPTSEILA VSQRLTEQNI INKRLHTS H AFHSSMMQDA AQALRQAFEN VRLNPPTLTI ISTVTGAHVS ADTLTTPDYW IEQMLMPVQF SAALQEAQAT FDVDFLEIG PGATLTQLTN GHALGDRLAF SSLPAGARSS DEHKHILDTV AALWVRGHNI DLSAFAGEQP RRVSLPTYAF DKIRYWVDSP EEQRSAVTP VADAGSKLSS GLEVLFQGPS SGHHHHHHHH HH

UniProtKB: Polyketide synthase

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Macromolecule #2: Polyketide synthase

MacromoleculeName: Polyketide synthase / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO / EC number: beta-ketoacyl-[acyl-carrier-protein] synthase I
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 15.369233 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
VIPSEPSVRR QPRPAFSVPY AAPESKTQRG LVAICEALLG IDGLGIDDNF FEAGGHSLML GMLLAQVQER FAVTLSFFDV MEDASVRAL AQLVEQEQQD DGGSALAVLV NDMINEKLSS GLEVLFQGPS SGHHHHHHHH HH

UniProtKB: Polyketide synthase

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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

Concentration1.50 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
200.0 mMNaClsodium chloride
20.0 mMTris-HClTRIS-hydrochloride
10.0 mMMgCl2magnesium chloride
1.0 mMTCEP(tris(2-carboxyethyl)phosphine)
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 3 / Number real images: 15349 / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.8 µm
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 11180913
CTF correctionSoftware - Name: cryoSPARC (ver. 3.0.1) / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: INSILICO MODEL / In silico model: ab-initio reconstruction
Final reconstructionNumber classes used: 2 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.29 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 362401
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.0.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0.1)
Final 3D classificationNumber classes: 3 / Avg.num./class: 166781 / Software - Name: cryoSPARC (ver. 3.0.1)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: AlphaFold / Chain - Initial model type: in silico model
RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-8xjy:
Cryo-EM structure of colibactin assembly line polyketide synthase ClbI KS-AT didomain crosslinked with ClbI ACP

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