[English] 日本語
Yorodumi
- EMDB-39101: Cryo-EM structure and rational engineering of a novel efficient o... -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-39101
TitleCryo-EM structure and rational engineering of a novel efficient ochratoxin A-detoxifying amidohydrolase
Map data
Sample
  • Complex: PwADH-G88D-D344N
    • Protein or peptide: Imidazolonepropionase
  • Ligand: ZINC ION
  • Ligand: (2~{S})-2-[[(3~{R})-5-chloranyl-3-methyl-8-oxidanyl-1-oxidanylidene-3,4-dihydroisochromen-7-yl]carbonylamino]-3-phenyl-propanoic acid
Keywordsamide hydrolase / Iron-binding / Ochratoxin A / ZN / HYDROLASE
Function / homology: / hydrolase activity, acting on carbon-nitrogen (but not peptide) bonds / Amidohydrolase family / Metal-dependent hydrolase, composite domain superfamily / Amidohydrolase-related / Metal-dependent hydrolase / Imidazolonepropionase
Function and homology information
Biological speciesPseudoxanthomonas wuyuanensis (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.37 Å
AuthorsDai LH / Xu YH / Hu YM / Niu D / Yang XC / Shen PP / Li X / Xie ZZ / Li H / Guo R-T / Chen C-C
Funding support1 items
OrganizationGrant numberCountry
Other government
CitationJournal: Int J Biol Macromol / Year: 2024
Title: Functional characterization and structural basis of an efficient ochratoxin A-degrading amidohydrolase.
Authors: Yumei Hu / Longhai Dai / Yuhang Xu / Du Niu / Xuechun Yang / Zhenzhen Xie / Panpan Shen / Xian Li / Hao Li / Lilan Zhang / Jian Min / Rey-Ting Guo / Chun-Chi Chen /
Abstract: Ochratoxin A (OTA) contamination in various agro-products poses a serious threat to the global food safety and human health, leading to enormous economic losses. Enzyme-mediated OTA degradation is an ...Ochratoxin A (OTA) contamination in various agro-products poses a serious threat to the global food safety and human health, leading to enormous economic losses. Enzyme-mediated OTA degradation is an appealing strategy, and the search for more efficient enzymes is a prerequisite for achieving this goal. Here, a novel amidohydrolase, termed PwADH, was demonstrated to exhibit 7.3-fold higher activity than that of the most efficient OTA-degrading ADH3 previously reported. Cryo-electron microscopy structure analysis indicated that additional hydrogen-bond interactions among OTA and the adjacent residue H163, the more compact substrate-binding pocket, and the wider entry to the substrate-access cavity might account for the more efficient OTA-degrading activity of PwADH compared with that of ADH3. We conducted a structure-guided rational design of PwADH and obtained an upgraded variant, G88D, whose OTA-degrading activity was elevated by 1.2-fold. In addition, PwADH and the upgraded G88D were successfully expressed in the industrial yeast Pichia pastoris, and their catalytic activities were compared to those of their counterparts produced in E. coli, revealing the feasibility of producing PwADH and its variants in industrial yeast strains. These results illustrate the structural basis of a novel, efficient OTA-degrading amidohydrolase and will be beneficial for the development of high-efficiency OTA-degrading approaches.
History
DepositionFeb 9, 2024-
Header (metadata) releaseDec 18, 2024-
Map releaseDec 18, 2024-
UpdateDec 18, 2024-
Current statusDec 18, 2024Processing site: PDBj / Status: Released

-
Structure visualization

Supplemental images

emd_39101.png

Downloads & links

-
Map

FileDownload / File: emd_39101.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.85 Å/pix.
x 320 pix.
= 272. Å		0.85 Å/pix.
x 320 pix.
= 272. Å		0.85 Å/pix.
x 320 pix.
= 272. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.85 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.25
Minimum - Maximum-0.9102252 - 1.511009
Average (Standard dev.)0.0004899478 (±0.06416916)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 272.0 Å
α=β=γ: 90.0 °

-
Supplemental data

-
Half map: #2

Fileemd_39101_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Half map: #1

Fileemd_39101_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Sample components

-
Entire : PwADH-G88D-D344N

EntireName: PwADH-G88D-D344N
Components
  • Complex: PwADH-G88D-D344N
    • Protein or peptide: Imidazolonepropionase
  • Ligand: ZINC ION
  • Ligand: (2~{S})-2-[[(3~{R})-5-chloranyl-3-methyl-8-oxidanyl-1-oxidanylidene-3,4-dihydroisochromen-7-yl]carbonylamino]-3-phenyl-propanoic acid

-
Supramolecule #1: PwADH-G88D-D344N

SupramoleculeName: PwADH-G88D-D344N / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: amidohydrolase family protein [Pseudoxanthomonas wuyuanensis]
Source (natural)Organism: Pseudoxanthomonas wuyuanensis (bacteria)

-
Macromolecule #1: Imidazolonepropionase

MacromoleculeName: Imidazolonepropionase / type: protein_or_peptide / ID: 1 / Number of copies: 8 / Enantiomer: LEVO
Source (natural)Organism: Pseudoxanthomonas wuyuanensis (bacteria)
Molecular weightTheoretical: 43.475809 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: EPVALHCGKL FDARSGRVLG PHTVVVRDGR IDQLISGGHA DVVGLAAVDL RNRTCLPGWT DLHVHLGDES SPQSYSEGFR LDPIDFAYR SVGYAERTLL AGFTSVRDLG GEVSPHLRDA VNQGLVKGPR IFAAGKSIAT TGGHADPTNG WNDQLSHLIG P PGPTEGVV ...String:
EPVALHCGKL FDARSGRVLG PHTVVVRDGR IDQLISGGHA DVVGLAAVDL RNRTCLPGWT DLHVHLGDES SPQSYSEGFR LDPIDFAYR SVGYAERTLL AGFTSVRDLG GEVSPHLRDA VNQGLVKGPR IFAAGKSIAT TGGHADPTNG WNDQLSHLIG P PGPTEGVV NSVDEARQAV RQRYKDGSDV I(KCX)ITATGGVL SYAKSGDAPQ FTVDEVKAIV DTANDYGYKV AAHAHGTE G MKRAILGGVT SIEHGTYMTD EVMRLMKQHG TWYVPTISAG RFVAEKAKID GYFPEVVRPK AARIGAQIQD TAAKAYRNG VKIAFGTNMG VGPHGDNARE FIYMVEAGIP AATALQSATV LAAEVLGVDD QGAIETGKRA DIIAMPGDPV ADINAVLNVD FVMKDGEIF RQP

UniProtKB: Imidazolonepropionase

-
Macromolecule #2: ZINC ION

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

-
Macromolecule #3: (2~{S})-2-[[(3~{R})-5-chloranyl-3-methyl-8-oxidanyl-1-oxidanylide...

MacromoleculeName: (2~{S})-2-[[(3~{R})-5-chloranyl-3-methyl-8-oxidanyl-1-oxidanylidene-3,4-dihydroisochromen-7-yl]carbonylamino]-3-phenyl-propanoic acid
type: ligand / ID: 3 / Number of copies: 8 / Formula: 97U
Molecular weightTheoretical: 403.813 Da
Chemical component information

ChemComp-97U:
(2~{S})-2-[[(3~{R})-5-chloranyl-3-methyl-8-oxidanyl-1-oxidanylidene-3,4-dihydroisochromen-7-yl]carbonylamino]-3-phenyl-propanoic acid / toxin*YM

-
Experimental details

-
Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

-
Sample preparation

Concentration1.5 mg/mL
BufferpH: 8 / Details: 20 mM Tris-HCL,pH 8.0
VitrificationCryogen name: ETHANE

-
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

-
Image processing

Startup modelType of model: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.37 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 118429
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

-
Atomic model buiding 1

RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-8yak:
Cryo-EM structure and rational engineering of a novel efficient ochratoxin A-detoxifying amidohydrolase

+
About Yorodumi

-
News

-
Feb 9, 2022. New format data for meta-information of EMDB entries

New format data for meta-information of EMDB entries

  • Version 3 of the EMDB header file is now the official format.
  • The previous official version 1.9 will be removed from the archive.

Related info.:EMDB header

External links:wwPDB to switch to version 3 of the EMDB data model

-
Aug 12, 2020. Covid-19 info

Covid-19 info

URL: https://pdbj.org/emnavi/covid19.php

New page: Covid-19 featured information page in EM Navigator.

Related info.:Covid-19 info / Mar 5, 2020. Novel coronavirus structure data

+
Mar 5, 2020. Novel coronavirus structure data

Novel coronavirus structure data

Related info.:Yorodumi Speices / Aug 12, 2020. Covid-19 info

External links:COVID-19 featured content - PDBj / Molecule of the Month (242)：Coronavirus Proteases

+
Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)

EMDB accession codes are about to change! (news from PDBe EMDB page)

  • The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force.
  • The EM Navigator/Yorodumi systems omit the EMD- prefix.

Related info.:Q: What is EMD? / ID/Accession-code notation in Yorodumi/EM Navigator

External links:EMDB Accession Codes are Changing Soon! / Contact to PDBj

+
Jul 12, 2017. Major update of PDB

Major update of PDB

  • wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary.
  • This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated.
  • In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software).
  • Now, EM Navigator and Yorodumi are based on the updated data.

External links:wwPDB Remediation / Enriched Model Files Conforming to OneDep Data Standards Now Available in the PDB FTP Archive

-
Yorodumi

Thousand views of thousand structures

  • Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
  • This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.
  • The word "yorodu" (or yorozu) is an old Japanese word meaning "ten thousand". "mi" (miru) is to see.

Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

Read more