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- EMDB-15960: Ligand-Free Structure of the decameric sulfofructose transaldolas... -

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Basic information

Entry
Database: EMDB / ID: EMD-15960
TitleLigand-Free Structure of the decameric sulfofructose transaldolase BmSF-TAL
Map data
Sample
  • Cell: Decameric complex of BmSF-TAL in a ligand-free state.
    • Protein or peptide: Transaldolase
  • Ligand: water
Keywordstransaldolase / cryo-EM / decamer / sulfofructose / TRANSFERASE
Function / homology6-deoxy-6-sulfo-D-fructose transaldolase / Transaldolase/Fructose-6-phosphate aldolase, archaeal/bacterial / transaldolase activity / Transaldolase/Fructose-6-phosphate aldolase / Transaldolase/Fructose-6-phosphate aldolase / aldehyde-lyase activity / Aldolase-type TIM barrel / carbohydrate metabolic process / 6-deoxy-6-sulfo-D-fructose transaldolase
Function and homology information
Biological speciesPriestia megaterium DSM 319 (bacteria) / Bacillus aryabhattai (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.6 Å
AuthorsSnow AJD / Sharma M / Blaza J / Davies GJ
Funding support United Kingdom, 3 items
OrganizationGrant numberCountry
Leverhulme TrustRPG-2017-190 United Kingdom
Wellcome Trust206161/Z/17/Z United Kingdom
UK Research and Innovation (UKRI)MR/T040742/1 United Kingdom
CitationJournal: Structure / Year: 2023
Title: Structure and mechanism of sulfofructose transaldolase, a key enzyme in sulfoquinovose metabolism.
Authors: Alexander J D Snow / Mahima Sharma / Palika Abayakoon / Spencer J Williams / James N Blaza / Gideon J Davies /
Abstract: Sulfoquinovose (SQ) is a key component of plant sulfolipids (sulfoquinovosyl diacylglycerols) and a major environmental reservoir of biological sulfur. Breakdown of SQ is achieved by bacteria through ...Sulfoquinovose (SQ) is a key component of plant sulfolipids (sulfoquinovosyl diacylglycerols) and a major environmental reservoir of biological sulfur. Breakdown of SQ is achieved by bacteria through the pathways of sulfoglycolysis. The sulfoglycolytic sulfofructose transaldolase (sulfo-SFT) pathway is used by gut-resident firmicutes and soil saprophytes. After isomerization of SQ to sulfofructose (SF), the namesake enzyme catalyzes the transaldol reaction of SF transferring dihydroxyacetone to 3C/4C acceptors to give sulfolactaldehyde and fructose-6-phosphate or sedoheptulose-7-phosphate. We report the 3D cryo-EM structure of SF transaldolase from Bacillus megaterium in apo and ligand bound forms, revealing a decameric structure formed from two pentameric rings of the protomer. We demonstrate a covalent "Schiff base" intermediate formed by reaction of SF with Lys89 within a conserved Asp-Lys-Glu catalytic triad and defined by an Arg-Trp-Arg sulfonate recognition triad. The structural characterization of the signature enzyme of the sulfo-SFT pathway provides key insights into molecular recognition of the sulfonate group of sulfosugars.
History
DepositionOct 14, 2022-
Header (metadata) releaseJan 18, 2023-
Map releaseJan 18, 2023-
UpdateNov 15, 2023-
Current statusNov 15, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_15960.png

Downloads & links

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Map

FileDownload / File: emd_15960.map.gz / Format: CCP4 / Size: 144.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
0.9 Å/pix.
x 336 pix.
= 302.4 Å		0.9 Å/pix.
x 336 pix.
= 302.4 Å		0.9 Å/pix.
x 336 pix.
= 302.4 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.9 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0304
Minimum - Maximum-0.07312639 - 0.12796505
Average (Standard dev.)0.00012992723 (±0.0030385277)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions336336336
Spacing336336336
CellA=B=C: 302.39996 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_15960_msk_1.map
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Half map: #2

Fileemd_15960_half_map_1.map
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Half map: #1

Fileemd_15960_half_map_2.map
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Sample components

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Entire : Decameric complex of BmSF-TAL in a ligand-free state.

EntireName: Decameric complex of BmSF-TAL in a ligand-free state.
Components
  • Cell: Decameric complex of BmSF-TAL in a ligand-free state.
    • Protein or peptide: Transaldolase
  • Ligand: water

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Supramolecule #1: Decameric complex of BmSF-TAL in a ligand-free state.

SupramoleculeName: Decameric complex of BmSF-TAL in a ligand-free state. / type: cell / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Priestia megaterium DSM 319 (bacteria)

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Macromolecule #1: Transaldolase

MacromoleculeName: Transaldolase / type: protein_or_peptide / ID: 1 / Number of copies: 10 / Enantiomer: LEVO / EC number: transaldolase
Source (natural)Organism: Bacillus aryabhattai (bacteria)
Molecular weightTheoretical: 24.443545 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MKYFLDSAIL EEIRYAYENW AIDGVTTNPR HIMNSGKPFL TVLDEFASEF KGVENFPISV EINPHLDNAK DMVEEGTKIA KLSSNFVIK IPCTEPGLIA AKEFEKQGIS TNVTLVFSPS QALQPARIGA KFVSPFVGWK ENSGDDTTYI QDIVNIYKNY N YNTEIIVA ...String:
MKYFLDSAIL EEIRYAYENW AIDGVTTNPR HIMNSGKPFL TVLDEFASEF KGVENFPISV EINPHLDNAK DMVEEGTKIA KLSSNFVIK IPCTEPGLIA AKEFEKQGIS TNVTLVFSPS QALQPARIGA KFVSPFVGWK ENSGDDTTYI QDIVNIYKNY N YNTEIIVA ALRNGKQIVD AAKAGAHIVT CGFDVYKESF QHAFTDYGLN KFRNAWDNTV

UniProtKB: 6-deoxy-6-sulfo-D-fructose transaldolase

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Macromolecule #2: water

MacromoleculeName: water / type: ligand / ID: 2 / Number of copies: 460 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER / Water

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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

Concentration2.5 mg/mL
BufferpH: 7.4
Component:
ConcentrationFormulaName
50.0 mMC4H11NO3TRIS
200.0 mMNaClSodium chlorideNaClSodium chloride

Details: Buffer was clarified in a 0.22um filter and used for final SEC polishing.
GridModel: Quantifoil / Material: GOLD / Support film - Material: GOLD / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS GLACIOS
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 30.0 µm / Illumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Number grids imaged: 1 / Number real images: 4690 / Average exposure time: 3.68 sec. / Average electron dose: 50.0 e/Å2

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Image processing

Particle selectionNumber selected: 153063
Startup modelType of model: INSILICO MODEL
In silico model: Modified alphafold prediction of the monomer.
Details: AF model had the C-terminus truncated, and was fitted to a map lowpass-filtered to 5 A/px. Model was then copied 9 more times and fitted to density with NCS restraints.
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final 3D classificationNumber classes: 4 / Avg.num./class: 38266
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionApplied symmetry - Point group: D5 (2x5 fold dihedral) / Resolution.type: BY AUTHOR / Resolution: 2.6 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 59063
FSC plot (resolution estimation)

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