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- PDB-9kti: CryoEM structure of a 2,3-hydroxycinnamic acid 1,2-dioxygenase Mh... -

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Basic information

Entry
Database: PDB / ID: 9kti
TitleCryoEM structure of a 2,3-hydroxycinnamic acid 1,2-dioxygenase MhpB in substrate bound form
Components2,3-dihydroxyphenylpropionate/2,3-dihydroxicinnamic acid 1,2-dioxygenase
KeywordsBIOSYNTHETIC PROTEIN / 2 / 3-hydroxycinnamic acid 1 / 2-dioxygenase
Function / homology
Function and homology information


phenylpropanoid catabolic process / 3-carboxyethylcatechol 2,3-dioxygenase / 3-carboxyethylcatechol 2,3-dioxygenase activity / 3-(3-hydroxy)phenylpropionate catabolic process / 3-phenylpropionate catabolic process / ferrous iron binding
Similarity search - Function
2,3-dihydroxyphenylpropionate/2,3-dihydroxicinnamic acid 1,2-dioxygenase / Extradiol ring-cleavage dioxygenase, class III enzyme, subunit B / Catalytic LigB subunit of aromatic ring-opening dioxygenase
Similarity search - Domain/homology
: / : / 2,3-dihydroxyphenylpropionate/2,3-dihydroxicinnamic acid 1,2-dioxygenase
Similarity search - Component
Biological speciesEscherichia coli K-12 (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.59 Å
AuthorsDong, X. / Jiang, W.X. / Ma, L.X. / Xing, Q.
Funding support China, 2items
OrganizationGrant numberCountry
National Science Foundation (NSF, China)32371277 China
National Science Foundation (NSF, China)32301028 China
CitationJournal: J Hazard Mater / Year: 2025
Title: Structural and catalytic insights into MhpB: A dioxygenase enzyme for degrading catecholic pollutants.
Authors: Xu Dong / Manli Xu / Miao Wu / Ying Wang / Xiaoqi Cheng / Wenxue Jiang / Dule Zheng / Ahmed Habiba Omar / Yibin Cheng / Aitao Li / Lixin Ma / Qiong Xing /
Abstract: The increasing environmental pollution from persistent aromatic compounds requires effective biodegradation strategies. In this study, we focused on MhpB, an extradiol dioxygenase (EDO) from ...The increasing environmental pollution from persistent aromatic compounds requires effective biodegradation strategies. In this study, we focused on MhpB, an extradiol dioxygenase (EDO) from Escherichia coli. It is known for its role in the degradation of catechols, key intermediates in the degradation of aromatic compounds. We report the high-resolution structure of MhpB determined by cryo-electron microscopy, revealing a decameric conformation with the catalytic chamber at the side. The structure-based analysis allowed us to investigate the substrate-enzyme interaction and the substrate selectivity, which are crucial for its catalytic function. Site-directed mutagenesis was used to modulate the in vitro and in vivo substrate preference of MhpB, enhancing its potential for industrial applications in pollutant degradation. The study provides insight into the mechanism of the enzyme and paves the way for the development of engineered EDOs for environmental remediation of aromatic pollutants.
History
DepositionDec 2, 2024Deposition site: PDBJ / Processing site: PDBC
Revision 1.0Feb 12, 2025Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: 2,3-dihydroxyphenylpropionate/2,3-dihydroxicinnamic acid 1,2-dioxygenase
B: 2,3-dihydroxyphenylpropionate/2,3-dihydroxicinnamic acid 1,2-dioxygenase
C: 2,3-dihydroxyphenylpropionate/2,3-dihydroxicinnamic acid 1,2-dioxygenase
D: 2,3-dihydroxyphenylpropionate/2,3-dihydroxicinnamic acid 1,2-dioxygenase
E: 2,3-dihydroxyphenylpropionate/2,3-dihydroxicinnamic acid 1,2-dioxygenase
F: 2,3-dihydroxyphenylpropionate/2,3-dihydroxicinnamic acid 1,2-dioxygenase
G: 2,3-dihydroxyphenylpropionate/2,3-dihydroxicinnamic acid 1,2-dioxygenase
H: 2,3-dihydroxyphenylpropionate/2,3-dihydroxicinnamic acid 1,2-dioxygenase
I: 2,3-dihydroxyphenylpropionate/2,3-dihydroxicinnamic acid 1,2-dioxygenase
J: 2,3-dihydroxyphenylpropionate/2,3-dihydroxicinnamic acid 1,2-dioxygenase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)344,69030
Polymers342,31010
Non-polymers2,38020
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
2,3-dihydroxyphenylpropionate/2,3-dihydroxicinnamic acid 1,2-dioxygenase / 3-carboxyethylcatechol 2 / 3-dioxygenase


Mass: 34230.961 Da / Num. of mol.: 10
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: mhpB, b0348, JW0339
Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
References: UniProt: P0ABR9, 3-carboxyethylcatechol 2,3-dioxygenase
#2: Chemical
ChemComp-FE2 / FE (II) ION


Mass: 55.845 Da / Num. of mol.: 10 / Source method: obtained synthetically / Formula: Fe
#3: Chemical
ChemComp-A1EG2 / 3-[2,3-bis(oxidanyl)phenyl]propanoic acid


Mass: 182.173 Da / Num. of mol.: 10 / Source method: obtained synthetically / Formula: C9H10O4 / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY
Has protein modificationN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: The decamer complex of MhpB bound with its substrate / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Source (natural)Organism: Escherichia coli K-12 (bacteria)
Source (recombinant)Organism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
MicroscopyModel: FEI TECNAI ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: DIFFRACTION / Nominal defocus max: 2200 nm / Nominal defocus min: 1000 nm
Image recordingElectron dose: 37 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.59 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 182651 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 75.27 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.002524870
ELECTRON MICROSCOPYf_angle_d0.489433820
ELECTRON MICROSCOPYf_chiral_restr0.03843680
ELECTRON MICROSCOPYf_plane_restr0.00434480
ELECTRON MICROSCOPYf_dihedral_angle_d4.57223410

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