[English] 日本語
Yorodumi
- EMDB-1313: Post-translational cleavage of recombinantly expressed nitrilase ... -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-1313
TitlePost-translational cleavage of recombinantly expressed nitrilase from Rhodococcus rhodochrous J1 yields a stable, active helical form.
Map dataMap of the carboxy-terminal truncated nitrilase helix from Rhodococcus rhodochrous J1
Sample
  • Sample: Carboxy-terminal truncated nitrilase from Rhodococcus rhodochrous J1
  • Protein or peptide: J1 nitrilase 1-327
Function / homologynitrilase activity / Carbon-nitrogen hydrolase
Function and homology information
Biological speciesRhodococcus rhodochrous (bacteria)
Methodhelical reconstruction / negative staining / Resolution: 18.0 Å
AuthorsThuku RN / Weber BW / Varsani A / Sewell BT
CitationJournal: FEBS J / Year: 2007
Title: Post-translational cleavage of recombinantly expressed nitrilase from Rhodococcus rhodochrous J1 yields a stable, active helical form.
Authors: R Ndoria Thuku / Brandon W Weber / Arvind Varsani / B Trevor Sewell /
Abstract: Nitrilases convert nitriles to the corresponding carboxylic acids and ammonia. The nitrilase from Rhodococcus rhodochrous J1 is known to be inactive as a dimer but to become active on oligomerization. ...Nitrilases convert nitriles to the corresponding carboxylic acids and ammonia. The nitrilase from Rhodococcus rhodochrous J1 is known to be inactive as a dimer but to become active on oligomerization. The recombinant enzyme undergoes post-translational cleavage at approximately residue 327, resulting in the formation of active, helical homo-oligomers. Determining the 3D structure of these helices using electron microscopy, followed by fitting the stain envelope with a model based on homology with other members of the nitrilase superfamily, enables the interacting surfaces to be identified. This also suggests that the reason for formation of the helices is related to the removal of steric hindrance arising from the 39 C-terminal amino acids from the wild-type protein. The helical form can be generated by expressing only residues 1-327.
History
DepositionJan 2, 2007-
Header (metadata) releaseJan 2, 2007-
Map releaseJan 2, 2007-
UpdateMay 26, 2011-
Current statusMay 26, 2011Processing site: PDBe / Status: Released

-
Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.03
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.03
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

1313.gif

Downloads & links

-
Map

FileDownload / File: emd_1313.map.gz / Format: CCP4 / Size: 7.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMap of the carboxy-terminal truncated nitrilase helix from Rhodococcus rhodochrous J1
Voxel sizeX=Y=Z: 4 Å
Density
Contour Level1: 0.0406 / Movie #1: 0.03
Minimum - Maximum0.0 - 0.0894763
Average (Standard dev.)0.00160742 (±0.0092578)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-64-64-64
Dimensions128128128
Spacing128128128
CellA=B=C: 512 Å
α=β=γ: 90 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z444
M x/y/z128128128
origin x/y/z0.0000.0000.000
length x/y/z512.000512.000512.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-63-63-63
NX/NY/NZ128128128
MAP C/R/S123
start NC/NR/NS-64-64-64
NC/NR/NS128128128
D min/max/mean0.0000.0890.002

-
Supplemental data

-
Sample components

-
Entire : Carboxy-terminal truncated nitrilase from Rhodococcus rhodochrous J1

EntireName: Carboxy-terminal truncated nitrilase from Rhodococcus rhodochrous J1
Components
  • Sample: Carboxy-terminal truncated nitrilase from Rhodococcus rhodochrous J1
  • Protein or peptide: J1 nitrilase 1-327

-
Supramolecule #1000: Carboxy-terminal truncated nitrilase from Rhodococcus rhodochrous J1

SupramoleculeName: Carboxy-terminal truncated nitrilase from Rhodococcus rhodochrous J1
type: sample / ID: 1000 / Number unique components: 1
Molecular weightExperimental: 36 KDa / Theoretical: 36 KDa / Method: SDS-PAGE MALDI-TOF mass spectrometry

-
Macromolecule #1: J1 nitrilase 1-327

MacromoleculeName: J1 nitrilase 1-327 / type: protein_or_peptide / ID: 1 / Oligomeric state: helix / Recombinant expression: Yes
Source (natural)Organism: Rhodococcus rhodochrous (bacteria) / Strain: J1
Molecular weightExperimental: 36 KDa / Theoretical: 36 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli) / Recombinant plasmid: pET30a
SequenceGO: nitrilase activity / InterPro: Carbon-nitrogen hydrolase

-
Experimental details

-
Structure determination

Methodnegative staining
Processinghelical reconstruction
Aggregation statefilament

-
Sample preparation

Concentration0.5 mg/mL
BufferpH: 7.8 / Details: 100mM KH2PO4, 400mM KCl, 10% (v/v) EtOH
StainingType: NEGATIVE
Details: Sample was applied to glow discharged carbon films which were then washed twice with distilled water and then stained with 2% w/v uranyl acetate
GridDetails: 300 mesh
VitrificationCryogen name: NONE

-
Electron microscopy

MicroscopeJEOL 2000EX
Electron beamAcceleration voltage: 120 kV / Electron source: TUNGSTEN HAIRPIN
Electron opticsCalibrated magnification: 50000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 50000
Sample stageSpecimen holder: standard / Specimen holder model: OTHER
Image recordingCategory: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: OTHER / Digitization - Sampling interval: 10 µm / Number real images: 50 / Average electron dose: 30 e/Å2 / Bits/pixel: 16

-
Image processing

Final reconstructionApplied symmetry - Helical parameters - Δz: 15.8 Å
Applied symmetry - Helical parameters - Δ&Phi: 73.65 °
Applied symmetry - Helical parameters - Axial symmetry: D1 (2x1 fold dihedral)
Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 18.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: SPIDER / Details: D1 symmetry was imposed on map after convergence
DetailsHelices were formed by autolysis of the wild-type protein after storage for one month at 4 degrees C

-
Atomic model buiding 1

SoftwareName: Situs
DetailsThe helical symmetry was applied to a homology model to generate a helix containing 9 dimers. This was fitted to the map using CoLoRes. A two dimensional search of radial distance and azimuthal angle was conducted to find the best fit.
RefinementSpace: REAL / Protocol: RIGID BODY FIT

+
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