[English] 日本語
Yorodumi
- PDB-6pr5: Cryo-EM structure of HzTransib strand transfer complex (STC) -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: PDB / ID: 6pr5
TitleCryo-EM structure of HzTransib strand transfer complex (STC)
Components
  • DNA (30-MER)
  • DNA (39-MER)
  • DNA (5'-D(*GP*AP*TP*CP*TP*GP*GP*CP*CP*TP*AP*GP*AP*TP*CP*TP*CP*A)-3')
  • DNA (5'-D(P*CP*AP*CP*GP*GP*TP*GP*GP*AP*TP*CP*GP*AP*AP*AP*A)-3')
  • DNA (5'-D(P*TP*TP*TP*TP*CP*GP*AP*TP*C)-3')
  • DNA-mediated transposase
KeywordsRECOMBINATION/DNA / RAG-like transposase / DDE family enzyme / Transib / Strand transfer. / RECOMBINATION / RECOMBINATION-DNA complex
Function / homologyPutative DNA-mediated transposase
Function and homology information
Biological speciesHelicoverpa zea (corn earworm)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsLiu, C. / Yang, Y. / Schatz, D.G.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases1R01AI137079 United States
CitationJournal: Nature / Year: 2019
Title: Structures of a RAG-like transposase during cut-and-paste transposition
Authors: Liu, C. / Yang, Y. / Schatz, D.G.
Validation Report
SummaryFull reportAbout validation report
History
DepositionJul 10, 2019Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 9, 2019Provider: repository / Type: Initial release

-
Structure visualization

Movie
  • Deposited structure unit
  • Imaged by Jmol
  • Download
  • Superimposition on EM map
  • EMDB-20457
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

-
Assembly

Deposited unit
A: DNA-mediated transposase
E: DNA-mediated transposase
B: DNA (5'-D(*GP*AP*TP*CP*TP*GP*GP*CP*CP*TP*AP*GP*AP*TP*CP*TP*CP*A)-3')
C: DNA (5'-D(P*CP*AP*CP*GP*GP*TP*GP*GP*AP*TP*CP*GP*AP*AP*AP*A)-3')
D: DNA (30-MER)
F: DNA (5'-D(P*TP*TP*TP*TP*CP*GP*AP*TP*C)-3')
G: DNA (5'-D(P*CP*AP*CP*GP*GP*TP*GP*GP*AP*TP*CP*GP*AP*AP*AP*A)-3')
H: DNA (39-MER)
hetero molecules


Theoretical massNumber of molelcules
Total (without water)152,66914
Polymers152,4418
Non-polymers2286
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration, The whole assembly was purified by gel filtration. A protein standard containing 6 different protein ranging from 1.7 kDa to 670 kDa was also passed through the same gel filtration column. Comparison of the two chromatography profiles indicate the assembly has an approximate molecular mass of 220-250 kDa, which is about the same as the combined molecular weight of two copies of HzTransib transposase and two TIR substrate DNA.
  • Download structure data
TypeNameSymmetry operationNumber
identity operation1_5551

-
Components

-
Protein/peptide , 1 types, 2 molecules AE

#1: Protein/peptide DNA-mediated transposase


Mass: 56582.734 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Helicoverpa zea (corn earworm) / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: B0F0C5

-
DNA chain , 5 types, 6 molecules BCGDFH

#2: DNA chain DNA (5'-D(*GP*AP*TP*CP*TP*GP*GP*CP*CP*TP*AP*GP*AP*TP*CP*TP*CP*A)-3')


Mass: 5491.566 Da / Num. of mol.: 1 / Details: Target DNA 5' flank / Source method: obtained synthetically / Source: (synth.) Helicoverpa zea (corn earworm)
#3: DNA chain DNA (5'-D(P*CP*AP*CP*GP*GP*TP*GP*GP*AP*TP*CP*GP*AP*AP*AP*A)-3')


Mass: 4956.244 Da / Num. of mol.: 2 / Details: Non-transferred strand of transposon end DNA / Source method: obtained synthetically / Source: (synth.) Helicoverpa zea (corn earworm)
#4: DNA chain DNA (30-MER)


Mass: 9238.951 Da / Num. of mol.: 1 / Details: Strand transfer product forward strand / Source method: obtained synthetically / Source: (synth.) Helicoverpa zea (corn earworm)
#5: DNA chain DNA (5'-D(P*TP*TP*TP*TP*CP*GP*AP*TP*C)-3')


Mass: 2696.783 Da / Num. of mol.: 1 / Details: Target DNA 3' flank / Source method: obtained synthetically / Source: (synth.) Helicoverpa zea (corn earworm)
#6: DNA chain DNA (39-MER)


Mass: 11935.659 Da / Num. of mol.: 1 / Details: Strand transfer product reverse strand / Source method: obtained synthetically / Source: (synth.) Helicoverpa zea (corn earworm)

-
Non-polymers , 2 types, 6 molecules

#7: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: Mg / Magnesium
#8: Chemical ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Zn / Zinc

-
Details

Has ligand of interestN

-
Experimental details

-
Experiment

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

-
Sample preparation

ComponentName: Strand transfer complex of HzTransib with transposon ends covalently linked to target DNA.
Type: COMPLEX / Entity ID: 1, 2, 3, 4, 5, 6 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Helicoverpa zea (corn earworm)
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm) / Cell: Sf9
Buffer solutionpH: 7.5
Details: Solutions were made fresh from concentrated and filtered to avoid microbial contamination.
Buffer component

Buffer-ID: 1

IDConc.NameFormula
120 mMTrisNH2C(CH2OH)3
250 mMpotassium chlorideKCl
310 mMMagnesium chlorideMgCl2
41 mMTris(2-carboxyethyl)phosphineC9H15O6P
SpecimenConc.: 0.3 mg/ml
Details: Recombinantly expressed HzTransib transposase was mixed with chemically synthesized TIR substrate DNA. The complex was further purified on size-exclusion chromatography column. The final complex was monodisperse.
Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 296 K / Details: Blot for 3 seconds before plunging

-
Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS / Details: Preliminary grid screening was performed manually.
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 130000 X / Nominal defocus max: 2400 nm / Nominal defocus min: 1400 nm / Cs: 2.7 mm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 8 sec. / Electron dose: 54.4 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k)
Details: Images were collected in movie-mode at 5 frames per second.
EM imaging opticsEnergyfilter name: GIF Quantum LS / Energyfilter slit width: 20 eV / Phase plate: No phase plate was used
Image scansMovie frames/image: 40 / Used frames/image: 1-40

-
Processing

SoftwareName: PHENIX / Version: 1.15.2_3472: / Classification: refinement
EM software
IDNameVersionCategory
2SerialEMimage acquisition
4RELION3CTF correction
7UCSF Chimera1.13model fitting
9PHENIX1.15model refinement
10RELION3initial Euler assignment
11RELION3final Euler assignment
12RELION3classification
13RELION33D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 43661 / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL / Target criteria: Correlation coefficient
Details: Initial local fitting was done using UCSF Chimera, then manually adjusted and rebuilt in Coot. Final model was refined using Phenix real-space refinement.
Atomic model buildingPDB-ID: 6PQN
Pdb chain-ID: A / Pdb chain residue range: 21-501
Refine LS restraints

Refinement-ID: ELECTRON MICROSCOPY

TypeDev idealNumber
f_bond_d0.00810638
f_angle_d0.72914892
f_dihedral_angle_d18.7126006
f_chiral_restr0.0451690
f_plane_restr0.0221512

+
About Yorodumi

-
News

-
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. (see PDBe EMDB page)
  • 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 at PDBe / 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. See below links for details.
  • 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

+
Jun 16, 2017. Omokage search with filter

Omokage search with filter

  • Result of Omokage search can be filtered by keywords and the database types

Related info.: Omokage search

+
Sep 15, 2016. EM Navigator & Yorodumi renewed

EM Navigator & Yorodumi renewed

  • New versions of EM Navigator and Yorodumi started

Related info.: Changes in new EM Navigator and Yorodumi

+
Aug 31, 2016. New EM Navigator & Yorodumi

New EM Navigator & Yorodumi

  • In 15th Sep 2016, the development versions of EM Navigator and Yorodumi will replace the official versions.
  • Current version will continue as 'legacy version' for some time.

Related info.: Changes in new EM Navigator and Yorodumi / EM Navigator / Yorodumi

Read more

-
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.

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