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- PDB-8tpw: Crosslinked 6-deoxyerythronolide B synthase (DEBS) Module 3 in co... -

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Basic information

Entry
Database: PDB / ID: 8tpw
TitleCrosslinked 6-deoxyerythronolide B synthase (DEBS) Module 3 in complex with antibody fragment 1B2: cis-oriented 1B2 and ACP
Components
  • Antibody Fragment 1B2, Heavy Chain
  • Antibody Fragment 1B2, Light Chain
  • EryAII,EryAII,EryAII,EryAII,6-deoxyerythronolide-B synthase EryA3, modules 5 and 6
KeywordsBIOSYNTHETIC PROTEIN / polyketide synthase / antibody
Function / homology
Function and homology information


6-deoxyerythronolide-B synthase / macrolide biosynthetic process / fatty acid synthase activity / phosphopantetheine binding / 3-oxoacyl-[acyl-carrier-protein] synthase activity / fatty acid biosynthetic process / oxidoreductase activity
Similarity search - Function
Erythronolide synthase, docking / Erythronolide synthase, docking domain superfamily / Erythronolide synthase, docking / : / Polyketide synthase extender module SpnB, Rossmann fold domain / Polyketide synthase, docking domain / Erythronolide synthase docking domain / Zinc-binding dehydrogenase / : / Polyketide synthase dehydratase N-terminal domain ...Erythronolide synthase, docking / Erythronolide synthase, docking domain superfamily / Erythronolide synthase, docking / : / Polyketide synthase extender module SpnB, Rossmann fold domain / Polyketide synthase, docking domain / Erythronolide synthase docking domain / Zinc-binding dehydrogenase / : / Polyketide synthase dehydratase N-terminal domain / Polyketide synthase, dehydratase domain / PKS_DH / PKS_PP_betabranch / : / Polyketide synthase dehydratase domain / : / Polyketide and metazoan fatty acid synthase dehydratase (PKS/mFAS DH) domain profile. / Polyketide synthase, dehydratase domain superfamily / Polyketide synthase, C-terminal extension / Ketoacyl-synthetase C-terminal extension / Polyketide synthase, ketoreductase domain / KR domain / Malonyl-CoA ACP transacylase, ACP-binding / : / Acyl transferase / Acyl transferase domain / Acyl transferase domain in polyketide synthase (PKS) enzymes. / Acyl transferase domain superfamily / Alcohol dehydrogenase, N-terminal / Alcohol dehydrogenase GroES-like domain / Polyketide synthase, enoylreductase domain / Enoylreductase / Acyl transferase/acyl hydrolase/lysophospholipase / Polyketide synthase, phosphopantetheine-binding domain / Phosphopantetheine attachment site / PKS_KR / Beta-ketoacyl synthase / Beta-ketoacyl synthase, active site / Ketosynthase family 3 (KS3) active site signature. / 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 / GroES-like superfamily / Thiolase-like / Phosphopantetheine attachment site / Phosphopantetheine attachment site. / Phosphopantetheine attachment site / ACP-like superfamily / Carrier protein (CP) domain profile. / Phosphopantetheine binding ACP domain / NAD(P)-binding domain superfamily
Similarity search - Domain/homology
6-deoxyerythronolide-B synthase
Similarity search - Component
Biological speciesSaccharopolyspora erythraea (bacteria)
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.46 Å
AuthorsCogan, D.P. / Soohoo, A.M. / Chen, M. / Brodsky, K.L. / Liu, Y. / Khosla, C.
Funding support United States, 4items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM141799 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM136039 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM129541 United States
National Science Foundation (NSF, United States)DGE-1656518 United States
CitationJournal: Nat Chem Biol / Year: 2025
Title: Structural basis for intermodular communication in assembly-line polyketide biosynthesis.
Authors: Dillon P Cogan / Alexander M Soohoo / Muyuan Chen / Yan Liu / Krystal L Brodsky / Chaitan Khosla /
Abstract: Assembly-line polyketide synthases (PKSs) are modular multi-enzyme systems with considerable potential for genetic reprogramming. Understanding how they selectively transport biosynthetic ...Assembly-line polyketide synthases (PKSs) are modular multi-enzyme systems with considerable potential for genetic reprogramming. Understanding how they selectively transport biosynthetic intermediates along a defined sequence of active sites could be harnessed to rationally alter PKS product structures. To investigate functional interactions between PKS catalytic and substrate acyl carrier protein (ACP) domains, we employed a bifunctional reagent to crosslink transient domain-domain interfaces of a prototypical assembly line, the 6-deoxyerythronolide B synthase, and resolved their structures by single-particle cryogenic electron microscopy (cryo-EM). Together with statistical per-particle image analysis of cryo-EM data, we uncovered interactions between ketosynthase (KS) and ACP domains that discriminate between intra-modular and inter-modular communication while reinforcing the relevance of conformational asymmetry during the catalytic cycle. Our findings provide a foundation for the structure-based design of hybrid PKSs comprising biosynthetic modules from different naturally occurring assembly lines.
History
DepositionAug 5, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Aug 7, 2024Provider: repository / Type: Initial release
Revision 1.1Sep 4, 2024Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID / _em_admin.last_update
Revision 1.2Jun 11, 2025Group: Data collection / Database references / Structure summary
Category: citation / citation_author ...citation / citation_author / em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.year / _citation_author.identifier_ORCID / _em_admin.last_update / _pdbx_entry_details.has_protein_modification

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: EryAII,EryAII,EryAII,EryAII,6-deoxyerythronolide-B synthase EryA3, modules 5 and 6
B: EryAII,EryAII,EryAII,EryAII,6-deoxyerythronolide-B synthase EryA3, modules 5 and 6
H: Antibody Fragment 1B2, Heavy Chain
L: Antibody Fragment 1B2, Light Chain
C: EryAII,EryAII,EryAII,EryAII,6-deoxyerythronolide-B synthase EryA3, modules 5 and 6


Theoretical massNumber of molelcules
Total (without water)611,6325
Polymers611,6325
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein EryAII,EryAII,EryAII,EryAII,6-deoxyerythronolide-B synthase EryA3, modules 5 and 6


Mass: 186489.578 Da / Num. of mol.: 3
Fragment: DEBS Module 3,DEBS Module 3,DEBS Module 3,DEBS Module 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharopolyspora erythraea (bacteria) / Gene: eryAII, eryA / Plasmid: pRSG34 / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BAP1
References: UniProt: Q5UNP5, 6-deoxyerythronolide-B synthase
#2: Antibody Antibody Fragment 1B2, Heavy Chain


Mass: 26447.611 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Escherichia coli BL21(DE3) (bacteria)
#3: Antibody Antibody Fragment 1B2, Light Chain


Mass: 25715.832 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Escherichia coli BL21(DE3) (bacteria)
Has ligand of interestY
Has protein modificationY

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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: Crosslinked DEBS Module 3 in complex with Antibody Fragment 1B2
Type: COMPLEX / Entity ID: all / Source: MULTIPLE SOURCES
Molecular weightValue: 0.42 MDa / Experimental value: YES
Source (natural)Organism: Saccharopolyspora erythraea (bacteria)
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria) / Strain: BAP1 / Plasmid: pRSG34
Buffer solutionpH: 7.2
Buffer component
IDConc.NameFormulaBuffer-ID
1100 mMcitric acidC6H8O71
2100 mMsodium chlorideNaCl1
310 mMHEPESC8H18N2O4S1
SpecimenConc.: 8 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: 30 s glow, 10 s hold, 15 mA / Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/1
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277.15 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 81000 X / Nominal defocus max: 2500 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 150 µm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 2.79 sec. / Electron dose: 50 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 10480

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Processing

EM softwareName: PHENIX / Version: 1.21_5207 / Category: model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 573745
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.46 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 59205 / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
Atomic model buildingPDB-ID: 6C9U

6c9u


Accession code: 6C9U / Source name: PDB / Type: experimental model
RefinementCross valid method: NONE

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