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- EMDB-16376: CryoEM structure of a tungsten-containing aldehyde oxidoreductase... -

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Entry
Database: EMDB / ID: EMD-16376
TitleCryoEM structure of a tungsten-containing aldehyde oxidoreductase from Aromatoleum aromaticum
Map data
Sample
  • Complex: Tungsten-containing aldehyde oxidoreductase from Aromatoleum aromaticum
    • Protein or peptide: Aldehyde:ferredoxin oxidoreductase,tungsten-containing
    • Protein or peptide: Iron-sulfur cluster-binding protein potential subunit of aldehyde oxidoreductase
    • Protein or peptide: Similar to ferredoxin:NADH oxidoreductases or NADH oxidases,potential subunit of aldehyde oxidoreductase
  • Ligand: IRON/SULFUR CLUSTER
  • Ligand: tungsten cofactor
  • Ligand: MAGNESIUM ION
  • Ligand: BENZOIC ACID
  • Ligand: FLAVIN-ADENINE DINUCLEOTIDE
KeywordsTungsten-containing enzyme / nanowires. / OXIDOREDUCTASE
Function / homology
Function and homology information


Oxidoreductases; Acting on the aldehyde or oxo group of donors; With an iron-sulfur protein as acceptor / oxidoreductase activity, acting on the aldehyde or oxo group of donors, iron-sulfur protein as acceptor / iron-sulfur cluster binding / 4 iron, 4 sulfur cluster binding / electron transfer activity / oxidoreductase activity / metal ion binding
Similarity search - Function
4Fe-4S binding domain / Aldehyde ferredoxin oxidoreductase, C-terminal / Aldehyde ferredoxin oxidoreductase, N-terminal / Aldehyde ferredoxin oxidoreductase, domain 2 / Aldehyde ferredoxin oxidoreductase, domain 3 / Aldehyde ferredoxin oxidoreductase-like, C-terminal / Aldehyde ferredoxin oxidoreductase, N-terminal domain superfamily / : / Aldehyde ferredoxin oxidoreductase, domains 2 & 3 / Aldehyde ferredoxin oxidoreductase, N-terminal domain ...4Fe-4S binding domain / Aldehyde ferredoxin oxidoreductase, C-terminal / Aldehyde ferredoxin oxidoreductase, N-terminal / Aldehyde ferredoxin oxidoreductase, domain 2 / Aldehyde ferredoxin oxidoreductase, domain 3 / Aldehyde ferredoxin oxidoreductase-like, C-terminal / Aldehyde ferredoxin oxidoreductase, N-terminal domain superfamily / : / Aldehyde ferredoxin oxidoreductase, domains 2 & 3 / Aldehyde ferredoxin oxidoreductase, N-terminal domain / Aldehyde ferredoxin oxidoreductase, N-terminal domain / : / : / 4Fe-4S dicluster domain / FAD/NAD(P)-binding domain / Pyridine nucleotide-disulphide oxidoreductase / 4Fe-4S ferredoxin, iron-sulphur binding, conserved site / 4Fe-4S ferredoxin-type iron-sulfur binding region signature. / 4Fe-4S ferredoxin-type iron-sulfur binding domain profile. / 4Fe-4S ferredoxin-type, iron-sulphur binding domain / FAD/NAD(P)-binding domain superfamily
Similarity search - Domain/homology
Similar to ferredoxin:NADH oxidoreductases or NADH oxidases,potential subunit of aldehyde oxidoreductase / Aldehyde:ferredoxin oxidoreductase,tungsten-containing / Iron-sulfur cluster-binding protein potential subunit of aldehyde oxidoreductase
Similarity search - Component
Biological speciesAromatoleum aromaticum (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.22 Å
AuthorsWiniarska A / Ramirez-Amador F / Hege D / Gemmecker Y / Prinz S / Hochberg G / Heider J / Szaleniec M / Schuller JM
Funding supportEuropean Union, Poland, Germany, 3 items
OrganizationGrant numberCountry
European Union (EU)European Union
Polish National Science Centre Poland
German Research Foundation (DFG) Germany
CitationJournal: Sci Adv / Year: 2023
Title: A bacterial tungsten-containing aldehyde oxidoreductase forms an enzymatic decorated protein nanowire.
Authors: Agnieszka Winiarska / Fidel Ramírez-Amador / Dominik Hege / Yvonne Gemmecker / Simone Prinz / Georg Hochberg / Johann Heider / Maciej Szaleniec / Jan Michael Schuller /
Abstract: Aldehyde oxidoreductases (AORs) are tungsten enzymes catalyzing the oxidation of many different aldehydes to the corresponding carboxylic acids. In contrast to other known AORs, the enzyme from the ...Aldehyde oxidoreductases (AORs) are tungsten enzymes catalyzing the oxidation of many different aldehydes to the corresponding carboxylic acids. In contrast to other known AORs, the enzyme from the denitrifying betaproteobacterium (AOR) consists of three different subunits (AorABC) and uses nicotinamide adenine dinucleotide (NAD) as an electron acceptor. Here, we reveal that the enzyme forms filaments of repeating AorAB protomers that are capped by a single NAD-binding AorC subunit, based on solving its structure via cryo-electron microscopy. The polyferredoxin-like subunit AorA oligomerizes to an electron-conducting nanowire that is decorated with enzymatically active and W-cofactor (W-co) containing AorB subunits. Our structure further reveals the binding mode of the native substrate benzoate in the AorB active site. This, together with quantum mechanics:molecular mechanics (QM:MM)-based modeling for the coordination of the W-co, enables formulation of a hypothetical catalytic mechanism that paves the way to further engineering for applications in synthetic biology and biotechnology.
History
DepositionDec 19, 2022-
Header (metadata) releaseMay 31, 2023-
Map releaseMay 31, 2023-
UpdateJul 24, 2024-
Current statusJul 24, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_16376.png

Downloads & links

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Map

FileDownload / File: emd_16376.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 Å/pix.
x 256 pix.
= 256. Å		1 Å/pix.
x 256 pix.
= 256. Å		1 Å/pix.
x 256 pix.
= 256. Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.55
Minimum - Maximum-3.2287517 - 7.2997375
Average (Standard dev.)0.0029564924 (±0.1502756)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 256.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_16376_msk_1.map
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Histogram (log scale)

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Half map: #2

Fileemd_16376_half_map_1.map
Projections & Slices
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Half map: #1

Fileemd_16376_half_map_2.map
Projections & Slices
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Sample components

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Entire : Tungsten-containing aldehyde oxidoreductase from Aromatoleum arom...

EntireName: Tungsten-containing aldehyde oxidoreductase from Aromatoleum aromaticum
Components
  • Complex: Tungsten-containing aldehyde oxidoreductase from Aromatoleum aromaticum
    • Protein or peptide: Aldehyde:ferredoxin oxidoreductase,tungsten-containing
    • Protein or peptide: Iron-sulfur cluster-binding protein potential subunit of aldehyde oxidoreductase
    • Protein or peptide: Similar to ferredoxin:NADH oxidoreductases or NADH oxidases,potential subunit of aldehyde oxidoreductase
  • Ligand: IRON/SULFUR CLUSTER
  • Ligand: tungsten cofactor
  • Ligand: MAGNESIUM ION
  • Ligand: BENZOIC ACID
  • Ligand: FLAVIN-ADENINE DINUCLEOTIDE

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Supramolecule #1: Tungsten-containing aldehyde oxidoreductase from Aromatoleum arom...

SupramoleculeName: Tungsten-containing aldehyde oxidoreductase from Aromatoleum aromaticum
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3
Source (natural)Organism: Aromatoleum aromaticum (bacteria)
Molecular weightTheoretical: 304 KDa

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Macromolecule #1: Aldehyde:ferredoxin oxidoreductase,tungsten-containing

MacromoleculeName: Aldehyde:ferredoxin oxidoreductase,tungsten-containing
type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
EC number: Oxidoreductases; Acting on the aldehyde or oxo group of donors; With an iron-sulfur protein as acceptor
Source (natural)Organism: Aromatoleum aromaticum (bacteria)
Molecular weightTheoretical: 65.954648 KDa
Recombinant expressionOrganism: Aromatoleum evansii (bacteria)
SequenceString: MGWNRKVLRV NLAEGTCTPE PLNMQWADEY LGSRGLATKY LVSETDPKVD PLSPDNKMIM ATGPLTGTMA STGGRYTVVT KGPLTGAIA CSNSGGFFGA EMKFAGWDMV IFEGRSPTPV YLFIENERAE LRDASYLWGR SCWETEESIR AQHQDPLIRV S SIGRAGEN ...String:
MGWNRKVLRV NLAEGTCTPE PLNMQWADEY LGSRGLATKY LVSETDPKVD PLSPDNKMIM ATGPLTGTMA STGGRYTVVT KGPLTGAIA CSNSGGFFGA EMKFAGWDMV IFEGRSPTPV YLFIENERAE LRDASYLWGR SCWETEESIR AQHQDPLIRV S SIGRAGEN QVMFACIVND LHRAAGRSGV GAVMGSKNLK AVAIRGTKGV SGIRDFPGFV RATSEAKKVL AGNPVTSEGL PK FGTQVLM NVINEMGALP TRNHRDVQFE DASKISAEAM HEKRPSDGKP QLVTNAACFG CTIACGRISA IDKTHFTVKN NPK YWGASG GLEYEAAWAL GAANGVGDLE ALQYANLLCN EQGMDPISFG ATVGAAMELY ETGVLTKERI GLDAPFGSAD ALAK LAEMT ATGEGFGKEI GLGSKRLCEK YGHPELSMSV KGQEFPAYDS RGIQGMGLAY ATSNRGACHL RGYTVASEVL GVPVK TDPH VIEGKAELVK AFQDATAVFD SAGICVFTSF AWTLADVQPQ IAAACDGDWS MDKLATVGER IWNMERQFNN AAGLGA QDD NLPPRLTSEP AKSGPAKGMV NRLAEMLPEY YGVRGWTPEG TPTPETLSRL GLS

UniProtKB: Aldehyde:ferredoxin oxidoreductase,tungsten-containing

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Macromolecule #2: Iron-sulfur cluster-binding protein potential subunit of aldehyde...

MacromoleculeName: Iron-sulfur cluster-binding protein potential subunit of aldehyde oxidoreductase
type: protein_or_peptide / ID: 2
Details: AorA (chain C,F in the model) is fused to a N-terminal Twin-Strep-tag (two Strep-tags linked by a glycin/serin peptide) for purification purposes.
Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Aromatoleum aromaticum (bacteria)
Molecular weightTheoretical: 20.524352 KDa
Recombinant expressionOrganism: Aromatoleum evansii (bacteria)
SequenceString: MASAWSHPQF EKGGGSGGGS GGSAWSHPQF EKSGMWKSLH IDPAKCTGCL QCEMACSYEH TGVINPSKSR IKVFSFEHEG RKVPYTCTQ CTEAWCLHSC PVDAIRLDLT TGAKMVFEDT CVGCKVCTIA CPFGTINYNQ DTGKVQKCDL CEGDPACAKA C PTAAITYI ...String:
MASAWSHPQF EKGGGSGGGS GGSAWSHPQF EKSGMWKSLH IDPAKCTGCL QCEMACSYEH TGVINPSKSR IKVFSFEHEG RKVPYTCTQ CTEAWCLHSC PVDAIRLDLT TGAKMVFEDT CVGCKVCTIA CPFGTINYNQ DTGKVQKCDL CEGDPACAKA C PTAAITYI DADWTGLARM QAWAAKANTP ASAA

UniProtKB: Iron-sulfur cluster-binding protein potential subunit of aldehyde oxidoreductase

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Macromolecule #3: Similar to ferredoxin:NADH oxidoreductases or NADH oxidases,poten...

MacromoleculeName: Similar to ferredoxin:NADH oxidoreductases or NADH oxidases,potential subunit of aldehyde oxidoreductase
type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Aromatoleum aromaticum (bacteria)
Molecular weightTheoretical: 45.798395 KDa
Recombinant expressionOrganism: Aromatoleum evansii (bacteria)
SequenceString: MKHVILGNGP AGVIAAETLR RAAPTDDILL FGSEDAPPYS RMAIPYLLEG NIDESGTWLR KSPGHFDRLR IHEMRGRAVS LDSERRRIL FDDGHFESWD RLLIATGSHP VRPPIPGIDL PEVQTCWTLE DARAIARFAT PGARVLQLGA GFIGCIIMEA L AARGVELT ...String:
MKHVILGNGP AGVIAAETLR RAAPTDDILL FGSEDAPPYS RMAIPYLLEG NIDESGTWLR KSPGHFDRLR IHEMRGRAVS LDSERRRIL FDDGHFESWD RLLIATGSHP VRPPIPGIDL PEVQTCWTLE DARAIARFAT PGARVLQLGA GFIGCIIMEA L AARGVELT VVEMGDRMVP RMMTPTAGGM IRKWVEDQGV RVVTNAGVSR IDCRASNDAP LDVTLSTGEV VVADLVIVAA GV APNIAFL EATPVHVAKG VLVDDRLQTS VPGIFAAGDV AEAPDLFTGA HLVAAIQPNA ADQARVAALN MAGHEARLKG VLA INVLDT LGLISSSFGQ WWGEERERGG AGVEHVDEAA YRYLSLQFKD DVLIGATSIG LTEHVGALRG LIHGRVRLGE WKER LLHSP LQFVDAYIAR SQQPMALVR

UniProtKB: Similar to ferredoxin:NADH oxidoreductases or NADH oxidases,potential subunit of aldehyde oxidoreductase

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Macromolecule #4: IRON/SULFUR CLUSTER

MacromoleculeName: IRON/SULFUR CLUSTER / type: ligand / ID: 4 / Number of copies: 10 / Formula: SF4
Molecular weightTheoretical: 351.64 Da
Chemical component information

ChemComp-FS1:
IRON/SULFUR CLUSTER

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Macromolecule #5: tungsten cofactor

MacromoleculeName: tungsten cofactor / type: ligand / ID: 5 / Number of copies: 2 / Formula: T7R
Molecular weightTheoretical: 1.002511 KDa
Chemical component information

ChemComp-T7R:
tungsten cofactor

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Macromolecule #6: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 6 / Number of copies: 2 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #7: BENZOIC ACID

MacromoleculeName: BENZOIC ACID / type: ligand / ID: 7 / Number of copies: 2 / Formula: BEZ
Molecular weightTheoretical: 122.121 Da
Chemical component information

ChemComp-BEZ:
BENZOIC ACID

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Macromolecule #8: FLAVIN-ADENINE DINUCLEOTIDE

MacromoleculeName: FLAVIN-ADENINE DINUCLEOTIDE / type: ligand / ID: 8 / Number of copies: 1 / Formula: FAD
Molecular weightTheoretical: 785.55 Da
Chemical component information

ChemComp-FAD:
FLAVIN-ADENINE DINUCLEOTIDE / FAD*YM

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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

Concentration0.70 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
45.0 mMHEPESHEPES
50.0 mMNaClsodium chloride
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 200 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 25 sec. / Details: 15 mA
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS GLACIOS
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Number real images: 896 / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: OTHER / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.5 µm

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Image processing #1

Image processing ID1
Particle selectionNumber selected: 549381
Startup modelType of model: INSILICO MODEL / In silico model: Ab initio
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.22 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 79731
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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Image processing #2

Image processing ID2
Particle selectionNumber selected: 549381
Startup modelType of model: INSILICO MODEL / In silico model: Ab initio
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.22 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 79731
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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