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- EMDB-36635: Structure of arginine oxidase from Pseudomonas sp. TRU 7192 -

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Basic information

Entry
Database: EMDB / ID: EMD-36635
TitleStructure of arginine oxidase from Pseudomonas sp. TRU 7192
Map data
Sample
  • Complex: homooctamer of arginine oxidase
    • Protein or peptide: Amine oxidoreductase
  • Ligand: FLAVIN-ADENINE DINUCLEOTIDE
  • Ligand: water
KeywordsOXIDOREDUCTASE
Biological speciesPseudomonas sp. (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.34 Å
AuthorsYamaguchi H / Numoto N / Suzuki H / Nishikawa K / Kamegawa A / Takahashi K / Sugiki M / Fujiyoshi Y
Funding support1 items
OrganizationGrant numberCountry
Not funded
CitationJournal: J Biochem / Year: 2025
Title: Open and closed structures of L-arginine oxidase by cryo-electron microscopy and X-ray crystallography.
Authors: Hiroki Yamaguchi / Kazutoshi Takahashi / Nobutaka Numoto / Hiroshi Suzuki / Moemi Tatsumi / Akiko Kamegawa / Kouki Nishikawa / Yasuhisa Asano / Toshimi Mizukoshi / Hiroshi Miyano / Yoshinori ...Authors: Hiroki Yamaguchi / Kazutoshi Takahashi / Nobutaka Numoto / Hiroshi Suzuki / Moemi Tatsumi / Akiko Kamegawa / Kouki Nishikawa / Yasuhisa Asano / Toshimi Mizukoshi / Hiroshi Miyano / Yoshinori Fujiyoshi / Masayuki Sugiki /
Abstract: L-arginine oxidase (AROD, EC 1.4.3.25) is an oxidoreductase that catalyses the deamination of L-arginine, with flavin adenine dinucleotide (FAD) as a cofactor. Recently identified AROD from ...L-arginine oxidase (AROD, EC 1.4.3.25) is an oxidoreductase that catalyses the deamination of L-arginine, with flavin adenine dinucleotide (FAD) as a cofactor. Recently identified AROD from Pseudomonas sp. TPU 7192 (PT-AROD) demonstrates high selectivity for L-arginine. This enzyme is useful for accurate assays of L-arginine in biological samples. The structural characteristics of the FAD-dependent AROD, however, remain unknown. Here, we report the structure of PT-AROD at a resolution of 2.3 Å by cryo-electron microscopy. PT-AROD adopts an octameric structure with D4 symmetry, which is consistent with its molecular weight in solution, estimated by mass photometry. Comparative analysis of this structure with that determined using X-ray crystallography reveals open and closed forms of the lid-like loop at the entrance to the substrate pocket. Furthermore, mutation of Glu493, located at the substrate binding site, diminishes substrate selectivity, suggesting that this residue contributes significantly to the high selectivity of PT-AROD.
History
DepositionJun 21, 2023-
Header (metadata) releaseJun 26, 2024-
Map releaseJun 26, 2024-
UpdateJan 15, 2025-
Current statusJan 15, 2025Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_36635.png

Downloads & links

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Map

FileDownload / File: emd_36635.map.gz / Format: CCP4 / Size: 36.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

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AxesZ (Sec.)Y (Row.)X (Col.)
0.96 Å/pix.
x 212 pix.
= 203.52 Å		0.96 Å/pix.
x 212 pix.
= 203.52 Å		0.96 Å/pix.
x 212 pix.
= 203.52 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.96 Å
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Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.05
Minimum - Maximum-0.11568865 - 0.24384327
Average (Standard dev.)-0.0004557735 (±0.01527924)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions212212212
Spacing212212212
CellA=B=C: 203.51999 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_36635_msk_1.map
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Half map: #2

Fileemd_36635_half_map_1.map
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Half map: #1

Fileemd_36635_half_map_2.map
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Sample components

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Entire : homooctamer of arginine oxidase

EntireName: homooctamer of arginine oxidase
Components
  • Complex: homooctamer of arginine oxidase
    • Protein or peptide: Amine oxidoreductase
  • Ligand: FLAVIN-ADENINE DINUCLEOTIDE
  • Ligand: water

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Supramolecule #1: homooctamer of arginine oxidase

SupramoleculeName: homooctamer of arginine oxidase / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Pseudomonas sp. (bacteria) / Strain: TPU 7192
Molecular weightTheoretical: 540 KDa

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Macromolecule #1: Amine oxidoreductase

MacromoleculeName: Amine oxidoreductase / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas sp. (bacteria) / Strain: TPU 7192
Molecular weightTheoretical: 67.393164 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MHHHHHHMSQ TQPLDVAIIG GGVSGTYSAW RLQEAQGDHQ RIQLFEYSDR IGGRLFSINL PGLPNVVAEV GGMRWMPATK DNTGGHVMV DKLVGELKLE SKNFPMGSNL PDKDPVGAKD NLFYLRGERF RLRDFTEAPD KIPYKLAWSE RGYGPEDLQV K VMHNIYPG ...String:
MHHHHHHMSQ TQPLDVAIIG GGVSGTYSAW RLQEAQGDHQ RIQLFEYSDR IGGRLFSINL PGLPNVVAEV GGMRWMPATK DNTGGHVMV DKLVGELKLE SKNFPMGSNL PDKDPVGAKD NLFYLRGERF RLRDFTEAPD KIPYKLAWSE RGYGPEDLQV K VMHNIYPG FDKLSLAEQM QVKVFGKEIW RYGFWDLLYR VLSNEGYQFM KDAGGYEANV ANASAVTQLP ATEYSDKTVF LA LKKGFQA LPLTLAKRFA EVPGGLIAGE QRIRMNRRLA SVQFSDDTEY PYRLHFQATR TVDGKTSDVP GAEEIIHARQ VIL ALPRRS LELIQSPLFD DPWLKENIDS VLVQSAFKLF LAYEQPWWRS QGLVAGRSVT DLPIRQCYYM GTECEQDGGE KTLN SLLMA SYNDIGTVPF WKGLEDGAPF EGYQPKSLQG RIDANEVVPK MQYQISEEMV RIAQRQVTSL HDQIELPAPY SAVYH AWDA DPFGGGWHEW KANYRLDLII QRMRHPVQEQ EVYIVGEAYS YGQGWVEGAL TTAESTLQDF FGLPRPAWLP EAYQLL PAP APVDIDNPPA LACTDCKKTL TEVTEFAYTG IKA

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Macromolecule #2: FLAVIN-ADENINE DINUCLEOTIDE

MacromoleculeName: FLAVIN-ADENINE DINUCLEOTIDE / type: ligand / ID: 2 / 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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Macromolecule #3: water

MacromoleculeName: water / type: ligand / ID: 3 / Number of copies: 29 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
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: 8
Component:
ConcentrationFormulaName
20.0 mMHEPES4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid
50.0 mMNaClsodium chloride
GridModel: Quantifoil R2/2 / Material: MOLYBDENUM / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Instrument: LEICA KF80

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Electron microscopy

MicroscopeJEOL CRYO ARM 300
Specialist opticsEnergy filter - Name: In-column Omega Filter / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 5035 / Average exposure time: 8.0 sec. / Average electron dose: 69.6 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 3.4 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Sample stageCooling holder cryogen: NITROGEN

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Image processing

Startup modelType of model: INSILICO MODEL
Final reconstructionApplied symmetry - Point group: D4 (2x4 fold dihedral) / Resolution.type: BY AUTHOR / Resolution: 2.34 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 254111
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: AlphaFold / Chain - Initial model type: in silico model
Output model

PDB-8jt7:
Structure of arginine oxidase from Pseudomonas sp. TRU 7192

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