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- EMDB-62861: Cryo-electron microscopic structure of a highly efficient ochrato... -

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Basic information

Entry
Database: EMDB / ID: EMD-62861
TitleCryo-electron microscopic structure of a highly efficient ochratoxin detoxification enzyme LlADH
Map data
Sample
  • Complex: LL-WT
    • Protein or peptide: LlADH-WT
  • Ligand: ZINC ION
Keywordsamide hydrolase / Iron-binding / Ochratoxin A / ZN / HYDROLASE / TOXIN
Biological speciesNovilysobacter luteus (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsDai LH / Xu YH / Hu YM / Niu D / He BY / Huang JP / Xie ZZ / Li H / Guo R-T / Chen C-C
Funding support1 items
OrganizationGrant numberCountry
Other government
CitationJournal: Int J Biol Macromol / Year: 2026
Title: Structure-guided protein engineering and immobilization of an amidohydrolase for efficient ochratoxin A detoxification.
Authors: Yumei Hu / Yuhang Xu / Biyu He / Du Niu / Xuechun Yang / Jiaping Huang / Zhenzhen Xie / Panpan Shen / Xian Li / Maosen Bai / Ziwei Liu / Hao Li / Yunyun Yang / Jian-Wen Huang / Chun-Chi Chen ...Authors: Yumei Hu / Yuhang Xu / Biyu He / Du Niu / Xuechun Yang / Jiaping Huang / Zhenzhen Xie / Panpan Shen / Xian Li / Maosen Bai / Ziwei Liu / Hao Li / Yunyun Yang / Jian-Wen Huang / Chun-Chi Chen / Rey-Ting Guo / Longhai Dai /
Abstract: Ochratoxin A (OTA) is a pervasive and significant mycotoxin that poses serious health risks to humans and animals. The development of efficient biocatalysts for the enzymatic detoxification of OTA is ...Ochratoxin A (OTA) is a pervasive and significant mycotoxin that poses serious health risks to humans and animals. The development of efficient biocatalysts for the enzymatic detoxification of OTA is of great importance. In this study, we enhanced the OTA-degrading activity of three amidohydrolases (ADHs) by up to ninefold. This improvement was achieved by reducing steric hindrance in the binding pocket and fine-tuning the hydrophilic interactions between the enzyme and substrate. The most efficient variant, PwADH/DM, was immobilized onto magnetic FeO nanoparticles functionalized by the co-deposition of dopamine and polyethyleneimine. Under optimal conditions, this immobilization process achieved a high immobilization efficiency (85.4%) and activity recovery (76.9%). The immobilized enzyme exhibited enhanced pH stability and thermostability, along with good storage stability, reusability, and recyclability. More importantly, the immobilized enzyme completely degraded 100 ng/mL OTA in contaminated milk without affecting the milk's properties. These findings expand our understanding of the molecular mechanisms governing substrate binding and catalysis in OTA-degrading ADHs. Furthermore, they provide a blueprint for enzyme-based OTA decontamination during food processing.
History
DepositionDec 24, 2024-
Header (metadata) releaseMay 6, 2026-
Map releaseMay 6, 2026-
UpdateMay 6, 2026-
Current statusMay 6, 2026Processing site: PDBc / Status: Released

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Structure visualization

Supplemental images

emd_62861.png

Downloads & links

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Map

FileDownload / File: emd_62861.map.gz / Format: CCP4 / Size: 178 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.)
0.85 Å/pix.
x 360 pix.
= 306. Å		0.85 Å/pix.
x 360 pix.
= 306. Å		0.85 Å/pix.
x 360 pix.
= 306. Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.85 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.3
Minimum - Maximum-0.94959867 - 1.4531914
Average (Standard dev.)-0.00088817027 (±0.044085532)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 306.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_62861_half_map_1.map
Projections & Slices
AxesZYX

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Histogram

Histogram (log scale)

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Half map: #2

Fileemd_62861_half_map_2.map
Projections & Slices
AxesZYX

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Sample components

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Entire : LL-WT

EntireName: LL-WT
Components
  • Complex: LL-WT
    • Protein or peptide: LlADH-WT
  • Ligand: ZINC ION

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Supramolecule #1: LL-WT

SupramoleculeName: LL-WT / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 / Details: amidohydrolase family protein
Source (natural)Organism: Novilysobacter luteus (bacteria)

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Macromolecule #1: LlADH-WT

MacromoleculeName: LlADH-WT / type: protein_or_peptide / ID: 1 / Number of copies: 8 / Enantiomer: LEVO
Source (natural)Organism: Novilysobacter luteus (bacteria)
Molecular weightTheoretical: 44.553848 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: QSPATAVHCG RLFDSRSGKV LGPHTVLVRD GRIAQVTSGG HADVAGVPSI DLSGHTCTPG WTDLHVHLAS QSSPDSYSEG FRLDEVDYA FRAVGYAKQT LLAGFTSVRD LGGEVAPHLR DAINQGLVAG PRIWAAGKSI ATTGGHADPT NGYNSTLSHL L GPPGPTEG ...String:
QSPATAVHCG RLFDSRSGKV LGPHTVLVRD GRIAQVTSGG HADVAGVPSI DLSGHTCTPG WTDLHVHLAS QSSPDSYSEG FRLDEVDYA FRAVGYAKQT LLAGFTSVRD LGGEVAPHLR DAINQGLVAG PRIWAAGKSI ATTGGHADPT NGYNSTLSHL L GPPGPTEG VINSVDDARQ AVRQRYKEGS DVI(KCX)ITATGG VLSYAKSGDA PQFTVDEVAA IVATAHDYGY RVAAHAHG E EGMRRAVEAG VTSIEHGTYM SDEVMALMKR KGTWYIPTVY AGRFVADKAK VDGYFPEVVR PKAARIGALI QDTAARAYR AGVPMAFGTD MGVGPHGDNA REFIYLVEAG IPAAKALQMA TIDAARVLGI DDQGRIAPGQ RADIVAVPGN PVDDIQAVLA VEFVMKDGV VYRQPGAAVP GETDRAGH

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Macromolecule #2: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 2 / Number of copies: 16 / Formula: ZN
Molecular weightTheoretical: 65.409 Da

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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

Concentration0.8 mg/mL
BufferpH: 8 / Details: 20 mM Tris-HCL,pH 8.0
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI MORGAGNI
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 52.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.4 µm / Nominal defocus min: 1.0 µm

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Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: OTHER
Final reconstructionNumber classes used: 30 / Applied symmetry - Point group: C4 (4 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 125111
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: FLEXIBLE FIT
Output model

PDB-9l6p:
Cryo-electron microscopic structure of a highly efficient ochratoxin detoxification enzyme LlADH

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