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- EMDB-33514: Cryo-EM structure of secondary alcohol dehydrogenases TbSADH afte... -

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Basic information

Entry
Database: EMDB / ID: EMD-33514
TitleCryo-EM structure of secondary alcohol dehydrogenases TbSADH after carrier-free immobilization based on weak intermolecular interactions
Map data
Sample
  • Complex: enzyme assembled gel
    • Protein or peptide: NADP-dependent isopropanol dehydrogenase
  • Ligand: ZINC ION
  • Ligand: MAGNESIUM ION
  • Ligand: water
KeywordsCoordination complex / Activity / Stability / Enzyme Immobilization / OXIDOREDUCTASE
Function / homology
Function and homology information


isopropanol dehydrogenase (NADP+) / isopropanol dehydrogenase (NADP+) activity / zinc ion binding
Similarity search - Function
Alcohol dehydrogenase, zinc-type, conserved site / Zinc-containing alcohol dehydrogenases signature. / Alcohol dehydrogenase-like, C-terminal / Zinc-binding dehydrogenase / Alcohol dehydrogenase, N-terminal / Alcohol dehydrogenase GroES-like domain / Polyketide synthase, enoylreductase domain / Enoylreductase / GroES-like superfamily / NAD(P)-binding domain superfamily
Similarity search - Domain/homology
NADP-dependent isopropanol dehydrogenase
Similarity search - Component
Biological speciesThermoanaerobacter brockii (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.12 Å
AuthorsChen Q / Li X / Yang F / Qu G / Sun ZT / Wang YJ
Funding support China, 2 items
OrganizationGrant numberCountry
Ministry of Science and Technology (MoST, China)2019YFA0905100 China
National Natural Science Foundation of China (NSFC)21878169 and 21991102 China
CitationJournal: Nat Commun / Year: 2023
Title: Active and stable alcohol dehydrogenase-assembled hydrogels via synergistic bridging of triazoles and metal ions.
Authors: Qiang Chen / Ge Qu / Xu Li / Mingjian Feng / Fan Yang / Yanjie Li / Jincheng Li / Feifei Tong / Shiyi Song / Yujun Wang / Zhoutong Sun / Guangsheng Luo /
Abstract: Biocatalysis is increasingly replacing traditional methods of manufacturing fine chemicals due to its green, mild, and highly selective nature, but biocatalysts, such as enzymes, are generally ...Biocatalysis is increasingly replacing traditional methods of manufacturing fine chemicals due to its green, mild, and highly selective nature, but biocatalysts, such as enzymes, are generally costly, fragile, and difficult to recycle. Immobilization provides protection for the enzyme and enables its convenient reuse, which makes immobilized enzymes promising heterogeneous biocatalysts; however, their industrial applications are limited by the low specific activity and poor stability. Herein, we report a feasible strategy utilizing the synergistic bridging of triazoles and metal ions to induce the formation of porous enzyme-assembled hydrogels with increased activity. The catalytic efficiency of the prepared enzyme-assembled hydrogels toward acetophenone reduction is 6.3 times higher than that of the free enzyme, and the reusability is confirmed by the high residual catalytic activity after 12 cycles of use. A near-atomic resolution (2.1 Å) structure of the hydrogel enzyme is successfully analyzed via cryogenic electron microscopy, which indicates a structure-property relationship for the enhanced performance. In addition, the possible mechanism of gel formation is elucidated, revealing the indispensability of triazoles and metal ions, which guides the use of two other enzymes to prepare enzyme-assembled hydrogels capable of good reusability. The described strategy can pave the way for the development of practical catalytic biomaterials and immobilized biocatalysts.
History
DepositionJun 1, 2022-
Header (metadata) releaseJun 7, 2023-
Map releaseJun 7, 2023-
UpdateMay 8, 2024-
Current statusMay 8, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_33514.png

Downloads & links

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Map

FileDownload / File: emd_33514.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.84 Å/pix.
x 300 pix.
= 251.22 Å		0.84 Å/pix.
x 300 pix.
= 251.22 Å		0.84 Å/pix.
x 300 pix.
= 251.22 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.8374 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.71
Minimum - Maximum-2.0341616 - 3.8801882
Average (Standard dev.)0.0005439131 (±0.10983915)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 251.22 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_33514_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_33514_half_map_2.map
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : enzyme assembled gel

EntireName: enzyme assembled gel
Components
  • Complex: enzyme assembled gel
    • Protein or peptide: NADP-dependent isopropanol dehydrogenase
  • Ligand: ZINC ION
  • Ligand: MAGNESIUM ION
  • Ligand: water

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Supramolecule #1: enzyme assembled gel

SupramoleculeName: enzyme assembled gel / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Thermoanaerobacter brockii (bacteria)

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Macromolecule #1: NADP-dependent isopropanol dehydrogenase

MacromoleculeName: NADP-dependent isopropanol dehydrogenase / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO / EC number: isopropanol dehydrogenase (NADP+)
Source (natural)Organism: Thermoanaerobacter brockii (bacteria)
Molecular weightTheoretical: 38.524695 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MHHHHHHKGF AMLSIGKVGW IEKEKPAPGP FDAIVRPLAV APCTSDIHTV FEGAIGERHN MILGHEAVGE VVEVGSEVKD FKPGDRVVV PANTPDWRTS EVQRGYHQHS GGMLAGWKFS NVKDGVFGEF FHVNDADMNL AHLPKEIPLE AAVMIPDMMT T GFHGAELA ...String:
MHHHHHHKGF AMLSIGKVGW IEKEKPAPGP FDAIVRPLAV APCTSDIHTV FEGAIGERHN MILGHEAVGE VVEVGSEVKD FKPGDRVVV PANTPDWRTS EVQRGYHQHS GGMLAGWKFS NVKDGVFGEF FHVNDADMNL AHLPKEIPLE AAVMIPDMMT T GFHGAELA DIELGATVAV LGIGPVGLMA VAGAKLRGAG RIIAVGSRPV CVDAAKYYGA TDIVNYKDGP IESQIMNLTE GK GVDAAII AGGNADIMAT AVKIVKPGGT IANVNYFGEG EVLPVPRLEW GCGMAHKTIK GGLCPGGRLR MERLIDLVFY KRV DPSKLV THVFRGFDNI EKAFMLMKDK PKDLIKPVVI LA

UniProtKB: NADP-dependent isopropanol dehydrogenase

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Macromolecule #2: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 2 / Number of copies: 4 / Formula: ZN
Molecular weightTheoretical: 65.409 Da

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Macromolecule #3: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 3 / Number of copies: 8 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #4: water

MacromoleculeName: water / type: ligand / ID: 4 / Number of copies: 574 / 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

BufferpH: 7.4
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.6 µm / Nominal defocus min: 0.7000000000000001 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Final reconstructionApplied symmetry - Point group: D2 (2x2 fold dihedral) / Resolution.type: BY AUTHOR / Resolution: 2.12 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 1254312
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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