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- EMDB-40785: MBP tagged mixed chain FAS, fusFAS_deltaACP -

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Basic information

Entry
Database: EMDB / ID: EMD-40785
TitleMBP tagged mixed chain FAS, fusFAS_deltaACP
Map dataMBP tagged mixed chain FAS, fusFAS_deltaACP_corrected
Sample
  • Complex: MBP-tagged mixed chain Fatty acid synthase, fusFAS_deltaACP
KeywordsFatty acid synthase / TRANSFERASE
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.87 Å
AuthorsMazhab-Jafari MT / Khalili Samani E
Funding support Canada, 2 items
OrganizationGrant numberCountry
Canadian Institutes of Health Research (CIHR)419240 Canada
Natural Sciences and Engineering Research Council (NSERC, Canada)RGPIN-2018-06070 Canada
CitationJournal: Commun Biol / Year: 2024
Title: Direct structural analysis of a single acyl carrier protein domain in fatty acid synthase from the fungus Saccharomyces cerevisiae.
Authors: Elnaz Khalili Samani / Amy C Chen / Jennifer W Lou / David L Dai / Alexander F A Keszei / Guihong Tan / Charles Boone / Martin Grininger / Mohammad T Mazhab-Jafari /
Abstract: Acyl carrier protein (ACP) is the work horse of polyketide (PKS) and fatty acid synthases (FAS) and acts as a substrate shuttling domain in these mega enzymes. In fungi, FAS forms a 2.6 MDa symmetric ...Acyl carrier protein (ACP) is the work horse of polyketide (PKS) and fatty acid synthases (FAS) and acts as a substrate shuttling domain in these mega enzymes. In fungi, FAS forms a 2.6 MDa symmetric assembly with six identical copies of FAS1 and FAS2 polypeptides. However, ACP spatial distribution is not restricted by symmetry owing to the long and flexible loops that tether the shuttling domain to its corresponding FAS2 polypeptide. This symmetry breaking has hampered experimental investigation of substrate shuttling route in fungal FAS. Here, we develop a protein engineering and expression method to isolate asymmetric fungal FAS proteins containing odd numbers of ACP domains. Electron cryomicroscopy (cryoEM) observation of the engineered complex reveals a non-uniform distribution of the substrate shuttling domain relative to its corresponding FAS2 polypeptide at 2.9 Å resolution. This work lays the methodological foundation for experimental study of ACP shuttling route in fungi.
History
DepositionMay 15, 2023-
Header (metadata) releaseJan 24, 2024-
Map releaseJan 24, 2024-
UpdateMay 8, 2024-
Current statusMay 8, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

Downloads & links

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Map

FileDownload / File: emd_40785.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMBP tagged mixed chain FAS, fusFAS_deltaACP_corrected
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.03 Å/pix.
x 400 pix.
= 412. Å
1.03 Å/pix.
x 400 pix.
= 412. Å
1.03 Å/pix.
x 400 pix.
= 412. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.03 Å
Density
Contour LevelBy AUTHOR: 0.17
Minimum - Maximum-0.57455236 - 1.4465778
Average (Standard dev.)0.0022933148 (±0.08373164)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 412.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: Half map A

Fileemd_40785_half_map_1.map
AnnotationHalf map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: Half map B

Fileemd_40785_half_map_2.map
AnnotationHalf map B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire : MBP-tagged mixed chain Fatty acid synthase, fusFAS_deltaACP

EntireName: MBP-tagged mixed chain Fatty acid synthase, fusFAS_deltaACP
Components
  • Complex: MBP-tagged mixed chain Fatty acid synthase, fusFAS_deltaACP

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Supramolecule #1: MBP-tagged mixed chain Fatty acid synthase, fusFAS_deltaACP

SupramoleculeName: MBP-tagged mixed chain Fatty acid synthase, fusFAS_deltaACP
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 2.7 MDa

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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 mg/mL
BufferpH: 7.4
GridModel: Homemade / Material: COPPER/RHODIUM / Mesh: 400 / Support film - Material: GOLD / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 25 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Number grids imaged: 1 / Number real images: 15032 / Average electron dose: 36.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.6 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.87 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.0.2) / Number images used: 167630
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4)

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