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- PDB-8brj: Escherichia coli anaerobic fatty acid beta oxidation trifunctiona... -

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Basic information

Entry
Database: PDB / ID: 8brj
TitleEscherichia coli anaerobic fatty acid beta oxidation trifunctional enzyme (anEcTFE) trimeric complex
Components
  • 3-ketoacyl-CoA thiolase FadI
  • Fatty acid oxidation complex subunit alpha
KeywordsMEMBRANE PROTEIN / complex / heterotrimer
Function / homology
Function and homology information


fatty acid beta-oxidation, unsaturated, even number / fatty acid beta-oxidation multienzyme complex / 3-hydroxybutyryl-CoA epimerase / 3-hydroxybutyryl-CoA epimerase activity / acetyl-CoA C-acyltransferase / acetyl-CoA C-acyltransferase activity / long-chain (3S)-3-hydroxyacyl-CoA dehydrogenase (NAD+) activity / 3-hydroxyacyl-CoA dehydrogenase / enoyl-CoA hydratase / (3S)-3-hydroxyacyl-CoA dehydrogenase (NAD+) activity ...fatty acid beta-oxidation, unsaturated, even number / fatty acid beta-oxidation multienzyme complex / 3-hydroxybutyryl-CoA epimerase / 3-hydroxybutyryl-CoA epimerase activity / acetyl-CoA C-acyltransferase / acetyl-CoA C-acyltransferase activity / long-chain (3S)-3-hydroxyacyl-CoA dehydrogenase (NAD+) activity / 3-hydroxyacyl-CoA dehydrogenase / enoyl-CoA hydratase / (3S)-3-hydroxyacyl-CoA dehydrogenase (NAD+) activity / acetyl-CoA C-acetyltransferase activity / enoyl-CoA hydratase activity / fatty acid beta-oxidation / NAD+ binding / cytosol / cytoplasm
Similarity search - Function
Acetyl-CoA C-acyltransferase FadI / Fatty oxidation complex, alpha subunit FadJ / : / 3-hydroxyacyl-CoA dehydrogenase, conserved site / 3-hydroxyacyl-CoA dehydrogenase signature. / 3-hydroxyacyl-CoA dehydrogenase, C-terminal / 3-hydroxyacyl-CoA dehydrogenase, NAD binding / 3-hydroxyacyl-CoA dehydrogenase, C-terminal domain / 3-hydroxyacyl-CoA dehydrogenase, NAD binding domain / Thiolase, active site ...Acetyl-CoA C-acyltransferase FadI / Fatty oxidation complex, alpha subunit FadJ / : / 3-hydroxyacyl-CoA dehydrogenase, conserved site / 3-hydroxyacyl-CoA dehydrogenase signature. / 3-hydroxyacyl-CoA dehydrogenase, C-terminal / 3-hydroxyacyl-CoA dehydrogenase, NAD binding / 3-hydroxyacyl-CoA dehydrogenase, C-terminal domain / 3-hydroxyacyl-CoA dehydrogenase, NAD binding domain / Thiolase, active site / Thiolases active site. / Thiolase, conserved site / Thiolases signature 2. / Thiolase, acyl-enzyme intermediate active site / Thiolases acyl-enzyme intermediate signature. / Thiolase, C-terminal / Thiolase, C-terminal domain / Thiolase / Thiolase, N-terminal / Thiolase, N-terminal domain / Enoyl-CoA hydratase/isomerase / Enoyl-CoA hydratase/isomerase / 6-phosphogluconate dehydrogenase-like, C-terminal domain superfamily / ClpP/crotonase-like domain superfamily / Thiolase-like / NAD(P)-binding domain superfamily
Similarity search - Domain/homology
3-ketoacyl-CoA thiolase FadI / Fatty acid oxidation complex subunit alpha
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.08 Å
AuthorsSah-Teli, S.K. / Pinkas, M. / Novacek, J. / Venkatesan, R.
Funding support Czech Republic, Finland, European Union, 3items
OrganizationGrant numberCountry
Ministry of Education, Youth and Sports of the Czech RepublicLM2018127 Czech Republic
Academy of Finland293369, 289024 and 319194 Finland
H2020 Marie Curie Actions of the European Commission871037, 871037European Union
CitationJournal: Structure / Year: 2023
Title: Structural basis for different membrane-binding properties of E. coli anaerobic and human mitochondrial β-oxidation trifunctional enzymes.
Authors: Shiv K Sah-Teli / Matyas Pinkas / Mikko J Hynönen / Sarah J Butcher / Rik K Wierenga / Jiri Novacek / Rajaram Venkatesan /
Abstract: Facultative anaerobic bacteria such as Escherichia coli have two αβ heterotetrameric trifunctional enzymes (TFE), catalyzing the last three steps of the β-oxidation cycle: soluble aerobic TFE ...Facultative anaerobic bacteria such as Escherichia coli have two αβ heterotetrameric trifunctional enzymes (TFE), catalyzing the last three steps of the β-oxidation cycle: soluble aerobic TFE (EcTFE) and membrane-associated anaerobic TFE (anEcTFE), closely related to the human mitochondrial TFE (HsTFE). The cryo-EM structure of anEcTFE and crystal structures of anEcTFE-α show that the overall assembly of anEcTFE and HsTFE is similar. However, their membrane-binding properties differ considerably. The shorter A5-H7 and H8 regions of anEcTFE-α result in weaker α-β as well as α-membrane interactions, respectively. The protruding H-H region of anEcTFE-β is therefore more critical for membrane-association. Mutational studies also show that this region is important for the stability of the anEcTFE-β dimer and anEcTFE heterotetramer. The fatty acyl tail binding tunnel of the anEcTFE-α hydratase domain, as in HsTFE-α, is wider than in EcTFE-α, accommodating longer fatty acyl tails, in good agreement with their respective substrate specificities.
History
DepositionNov 23, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0May 17, 2023Provider: repository / Type: Initial release
Revision 1.1May 31, 2023Group: Database references / Category: citation / citation_author
Item: _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID
Revision 1.2Jul 19, 2023Group: Database references / Category: citation / Item: _citation.journal_volume / _citation.page_first
Revision 1.3Jul 24, 2024Group: Data collection / Category: chem_comp_atom / chem_comp_bond / em_admin / Item: _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: 3-ketoacyl-CoA thiolase FadI
B: 3-ketoacyl-CoA thiolase FadI
C: Fatty acid oxidation complex subunit alpha


Theoretical massNumber of molelcules
Total (without water)173,5733
Polymers173,5733
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area6210 Å2
ΔGint-22 kcal/mol
Surface area62940 Å2
MethodPISA

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Components

#1: Protein 3-ketoacyl-CoA thiolase FadI / ACSs / Acetyl-CoA acyltransferase / Acyl-CoA ligase / Beta-ketothiolase / Fatty acid oxidation ...ACSs / Acetyl-CoA acyltransferase / Acyl-CoA ligase / Beta-ketothiolase / Fatty acid oxidation complex subunit beta


Mass: 48202.070 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Gene: fadI, yfcY, b2342, JW2339
Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
References: UniProt: P76503, acetyl-CoA C-acyltransferase
#2: Protein Fatty acid oxidation complex subunit alpha


Mass: 77169.125 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Gene: fadJ, yfcX, b2341, JW2338
Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
References: UniProt: P77399, enoyl-CoA hydratase, 3-hydroxybutyryl-CoA epimerase, 3-hydroxyacyl-CoA dehydrogenase

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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: anEcTFE trimer / Type: COMPLEX / Details: heterotrimeric complex / Entity ID: all / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Escherichia coli (E. coli)
Source (recombinant)Organism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
Buffer solutionpH: 8
Details: 50 mM Tris, 500 mM NaCl, 5% glycerol, 2.5 mM DTT, 0.0033% amphipol A8-35, pH 8
SpecimenConc.: 0.1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: C-flat-1.2/1.3
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 defocus max: 2500 nm / Nominal defocus min: 1200 nm / Cs: 2.7 mm / C2 aperture diameter: 30 µm / Alignment procedure: ZEMLIN TABLEAU
Image recordingAverage exposure time: 5 sec. / Electron dose: 48 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 QUANTUM (4k x 4k) / Num. of grids imaged: 3 / Num. of real images: 18669
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 10 eV
Image scansSampling size: 5 µm / Width: 3838 / Height: 3710 / Movie frames/image: 40

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Processing

SoftwareName: PHENIX / Version: 1.19.2_4158: / Classification: refinement
EM software
IDNameCategory
1cryoSPARCparticle selection
2SerialEMimage acquisition
4GctfCTF correction
5cryoSPARCCTF correction
8Cootmodel fitting
10cryoSPARCinitial Euler assignment
11cryoSPARCfinal Euler assignment
12RELIONclassification
13cryoSPARCclassification
14cryoSPARC3D reconstruction
15PHENIXmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 4.08 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 37357 / Algorithm: FOURIER SPACE / Symmetry type: POINT
Atomic model buildingSpace: REAL / Target criteria: cross-correlation coefficient
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00311824
ELECTRON MICROSCOPYf_angle_d0.58516057
ELECTRON MICROSCOPYf_dihedral_angle_d5.2641701
ELECTRON MICROSCOPYf_chiral_restr0.0411888
ELECTRON MICROSCOPYf_plane_restr0.0052103

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