[English] 日本語
Yorodumi
- PDB-3jbw: Cryo-electron microscopy structure of RAG Paired Complex (with NB... -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: PDB / ID: 3jbw
TitleCryo-electron microscopy structure of RAG Paired Complex (with NBD, no symmetry)
Components
  • (Nicked 23-RSS intermediate ...) x 2
  • (V(D)J recombination-activating protein ...) x 2
  • 12-RSS signal end forward strand
  • 5'-D(P*GP*AP*TP*CP*TP*GP*GP*CP*CP*TP*GP*TP*CP*TP*TP*A)-3'
  • Nicked 12-RSS intermediate reverse strand
KeywordsRECOMBINATION/DNA / RAG1 / RAG2 / V(D)J recombination / Paired complex / Antigen receptor gene recombination / T and B cell development / RECOMBINATION-DNA complex
Function / homology
Function and homology information


somatic diversification of immune receptors via germline recombination within a single locus / hematopoietic or lymphoid organ development / protein-DNA complex assembly / DNA recombinase complex / endodeoxyribonuclease complex / lymphocyte differentiation / immunoglobulin V(D)J recombination / V(D)J recombination / phosphatidylinositol-3,4-bisphosphate binding / phosphatidylinositol-3,5-bisphosphate binding ...somatic diversification of immune receptors via germline recombination within a single locus / hematopoietic or lymphoid organ development / protein-DNA complex assembly / DNA recombinase complex / endodeoxyribonuclease complex / lymphocyte differentiation / immunoglobulin V(D)J recombination / V(D)J recombination / phosphatidylinositol-3,4-bisphosphate binding / phosphatidylinositol-3,5-bisphosphate binding / phosphatidylinositol-3,4,5-trisphosphate binding / T cell differentiation / methylated histone binding / phosphatidylinositol-4,5-bisphosphate binding / phosphatidylinositol binding / B cell differentiation / thymus development / RING-type E3 ubiquitin transferase / ubiquitin-protein transferase activity / ubiquitin protein ligase activity / T cell differentiation in thymus / chromatin organization / histone binding / endonuclease activity / DNA recombination / sequence-specific DNA binding / adaptive immune response / Hydrolases; Acting on ester bonds / chromatin binding / magnesium ion binding / protein homodimerization activity / DNA binding / zinc ion binding / metal ion binding / nucleus
Similarity search - Function
V(D)J recombination-activating protein 1, Zinc finger / RAG nonamer-binding domain / NBD domain profile. / Zinc finger RAG1-type profile. / V(D)J recombination-activating protein 1 / RAG1 importin-binding / RAG1 importin binding / Recombination-activation protein 1 (RAG1), recombinase / Recombination activating protein 2 / RAG2 PHD domain ...V(D)J recombination-activating protein 1, Zinc finger / RAG nonamer-binding domain / NBD domain profile. / Zinc finger RAG1-type profile. / V(D)J recombination-activating protein 1 / RAG1 importin-binding / RAG1 importin binding / Recombination-activation protein 1 (RAG1), recombinase / Recombination activating protein 2 / RAG2 PHD domain / V-D-J recombination activating protein 2 / Recombination activating protein 2, PHD domain / Galactose oxidase/kelch, beta-propeller / Kelch-type beta propeller / Zinc finger, C3HC4 RING-type / Zinc finger, C3HC4 type (RING finger) / Zinc finger, RING-type, conserved site / Zinc finger RING-type signature. / Ring finger / Zinc finger RING-type profile. / Zinc finger, RING-type / Zinc finger, FYVE/PHD-type / Zinc finger, RING/FYVE/PHD-type
Similarity search - Domain/homology
DNA / DNA (> 10) / V(D)J recombination-activating protein 1 / V(D)J recombination-activating protein 2 / V(D)J recombination-activating protein 2
Similarity search - Component
Biological speciesDanio rerio (zebrafish)
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.6 Å
AuthorsRu, H. / Chambers, M.G. / Fu, T.-M. / Tong, A.B. / Liao, M. / Wu, H.
CitationJournal: Cell / Year: 2015
Title: Molecular Mechanism of V(D)J Recombination from Synaptic RAG1-RAG2 Complex Structures.
Authors: Heng Ru / Melissa G Chambers / Tian-Min Fu / Alexander B Tong / Maofu Liao / Hao Wu /
Abstract: Diverse repertoires of antigen-receptor genes that result from combinatorial splicing of coding segments by V(D)J recombination are hallmarks of vertebrate immunity. The (RAG1-RAG2)2 recombinase (RAG) ...Diverse repertoires of antigen-receptor genes that result from combinatorial splicing of coding segments by V(D)J recombination are hallmarks of vertebrate immunity. The (RAG1-RAG2)2 recombinase (RAG) recognizes recombination signal sequences (RSSs) containing a heptamer, a spacer of 12 or 23 base pairs, and a nonamer (12-RSS or 23-RSS) and introduces precise breaks at RSS-coding segment junctions. RAG forms synaptic complexes only with one 12-RSS and one 23-RSS, a dogma known as the 12/23 rule that governs the recombination fidelity. We report cryo-electron microscopy structures of synaptic RAG complexes at up to 3.4 Å resolution, which reveal a closed conformation with base flipping and base-specific recognition of RSSs. Distortion at RSS-coding segment junctions and base flipping in coding segments uncover the two-metal-ion catalytic mechanism. Induced asymmetry involving tilting of the nonamer-binding domain dimer of RAG1 upon binding of HMGB1-bent 12-RSS or 23-RSS underlies the molecular mechanism for the 12/23 rule.
History
DepositionOct 21, 2015Deposition site: RCSB / Processing site: RCSB
Revision 1.0Dec 9, 2015Provider: repository / Type: Initial release
Revision 1.1Jul 18, 2018Group: Data collection / Category: em_software / Item: _em_software.image_processing_id / _em_software.name
Revision 1.2Feb 21, 2024Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_struct_conn_angle / refine / struct_conn / struct_ref_seq_dif / struct_site
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_struct_conn_angle.ptnr1_auth_asym_id / _pdbx_struct_conn_angle.ptnr1_auth_comp_id / _pdbx_struct_conn_angle.ptnr1_auth_seq_id / _pdbx_struct_conn_angle.ptnr1_label_asym_id / _pdbx_struct_conn_angle.ptnr1_label_atom_id / _pdbx_struct_conn_angle.ptnr1_label_comp_id / _pdbx_struct_conn_angle.ptnr1_label_seq_id / _pdbx_struct_conn_angle.ptnr2_auth_asym_id / _pdbx_struct_conn_angle.ptnr2_label_asym_id / _pdbx_struct_conn_angle.ptnr3_auth_asym_id / _pdbx_struct_conn_angle.ptnr3_auth_comp_id / _pdbx_struct_conn_angle.ptnr3_auth_seq_id / _pdbx_struct_conn_angle.ptnr3_label_asym_id / _pdbx_struct_conn_angle.ptnr3_label_atom_id / _pdbx_struct_conn_angle.ptnr3_label_comp_id / _pdbx_struct_conn_angle.ptnr3_label_seq_id / _pdbx_struct_conn_angle.value / _refine.ls_d_res_high / _refine.ls_d_res_low / _struct_conn.pdbx_dist_value / _struct_conn.ptnr1_auth_asym_id / _struct_conn.ptnr1_auth_comp_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr1_label_comp_id / _struct_conn.ptnr1_label_seq_id / _struct_conn.ptnr2_auth_asym_id / _struct_conn.ptnr2_label_asym_id / _struct_ref_seq_dif.details / _struct_site.pdbx_auth_asym_id / _struct_site.pdbx_auth_comp_id / _struct_site.pdbx_auth_seq_id

-
Structure visualization

Movie
  • Deposited structure unit
  • Imaged by Jmol
  • Download
  • Superimposition on EM map
  • EMDB-6489
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

-
Assembly

Deposited unit
A: V(D)J recombination-activating protein 1
B: V(D)J recombination-activating protein 2
C: V(D)J recombination-activating protein 1
D: V(D)J recombination-activating protein 2
E: 12-RSS signal end forward strand
F: Nicked 12-RSS intermediate reverse strand
G: Nicked 23-RSS intermediate reverse strand
H: Nicked 23-RSS intermediate forward strand
I: 5'-D(P*GP*AP*TP*CP*TP*GP*GP*CP*CP*TP*GP*TP*CP*TP*TP*A)-3'
J: 5'-D(P*GP*AP*TP*CP*TP*GP*GP*CP*CP*TP*GP*TP*CP*TP*TP*A)-3'
hetero molecules


Theoretical massNumber of molelcules
Total (without water)362,68812
Polymers362,55710
Non-polymers1312
Water0
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

-
Components

-
V(D)J recombination-activating protein ... , 2 types, 4 molecules ACBD

#1: Protein V(D)J recombination-activating protein 1 / RAG-1 / Endonuclease RAG1 / E3 ubiquitin-protein ligase RAG1


Mass: 87684.125 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Danio rerio (zebrafish) / Strain: AB / Cell: B and T lymphocyte / Gene: rag1 / Organelle: nucleusCell nucleus / Plasmid: pFastBac1 / Production host: Spodoptera frugiperda (fall armyworm)
References: UniProt: O13033, Hydrolases; Acting on ester bonds, Ligases; Forming carbon-nitrogen bonds; Acid-amino-acid ligases (peptide synthases)
#2: Protein V(D)J recombination-activating protein 2 / RAG-2


Mass: 59435.930 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Danio rerio (zebrafish) / Strain: AB / Cell: B and T lymphocyte / Gene: rag2, rag-2 / Organelle: nucleusCell nucleus / Plasmid: pFastBac1 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q1RLW7, UniProt: O13034*PLUS

-
DNA chain , 3 types, 4 molecules EFIJ

#3: DNA chain 12-RSS signal end forward strand


Mass: 10439.762 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Homo sapiens (human)
#4: DNA chain Nicked 12-RSS intermediate reverse strand


Mass: 15439.880 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Homo sapiens (human)
#7: DNA chain 5'-D(P*GP*AP*TP*CP*TP*GP*GP*CP*CP*TP*GP*TP*CP*TP*TP*A)-3'


Mass: 4880.164 Da / Num. of mol.: 2 / Source method: obtained synthetically / Details: Coding end DNA forward strand / Source: (synth.) Homo sapiens (human)

-
Nicked 23-RSS intermediate ... , 2 types, 2 molecules GH

#5: DNA chain Nicked 23-RSS intermediate reverse strand


Mass: 18870.094 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Homo sapiens (human)
#6: DNA chain Nicked 23-RSS intermediate forward strand


Mass: 13806.871 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Homo sapiens (human)

-
Non-polymers , 1 types, 2 molecules

#8: Chemical ChemComp-ZN / ZINC ION


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

-
Experimental details

-
Experiment

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

-
Sample preparation

Component
IDNameTypeDetailsParent-ID
1Paired Complex of RAG1-RAG2COMPLEXDimer of RAG1-RAG2 binds to nicked 12-RSS and 23-RSS intermediates with HMGB10
2Recombination Activating Gene 1Recombination-activating gene1
3Recombination Activating Gene 2Recombination-activating gene1
Molecular weightValue: 0.4 MDa / Experimental value: YES
Buffer solutionName: Polymix buffer / pH: 7.5 / Details: 150 mM NaCl, 20 mM HEPES, 10 mM CaCl2, 1 mM TCEP
SpecimenConc.: 0.4 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: 400 mesh Quantifoil holey carbon grid, glow discharged
VitrificationInstrument: GATAN CRYOPLUNGE 3 / Cryogen name: ETHANE / Temp: 120 K / Humidity: 85 %
Details: Blot for 2.5 seconds before plunging into liquid ethane (GATAN CRYOPLUNGE 3).
Method: Blot for 2.5 seconds before plunging.

-
Electron microscopy imaging

Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company
MicroscopyModel: FEI POLARA 300 / Date: Mar 10, 2015
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 31000 X / Calibrated magnification: 40607 X / Nominal defocus max: 2500 nm / Nominal defocus min: 1500 nm / Cs: 2 mm
Astigmatism: Objective lens astigmatism was corrected at 150,000 times magnification.
Specimen holderSpecimen holder model: GATAN LIQUID NITROGEN / Temperature: 100 K / Temperature (max): 105 K / Temperature (min): 80 K
Image recordingElectron dose: 41 e/Å2 / Film or detector model: GATAN K2 (4k x 4k)
Image scansNum. digital images: 550

-
Processing

EM software
IDNameCategory
1RELION3D reconstruction
2SPIDER3D reconstruction
CTF correctionDetails: Each particle
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionMethod: Projection matching / Resolution: 4.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 14129 / Nominal pixel size: 1.23 Å / Actual pixel size: 1.23 Å
Details: (Single particle details: Image processing was carried out using SAMUEL and Relion.) (Single particle--Applied symmetry: C1)
Symmetry type: POINT
RefinementResolution: 4.6→4.6 Å / SU ML: 1.64 / σ(F): 0 / Phase error: 62.82 / Stereochemistry target values: MLHL
RfactorNum. reflection% reflection
Rfree0.4642 551 0.2 %
Rwork0.4735 276860 -
obs0.4735 277411 99.88 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso max: 587.07 Å2 / Biso mean: 180.3618 Å2 / Biso min: 19.35 Å2
Refinement stepCycle: LAST / Resolution: 4.631→236.16 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms15444 4551 2 0 19997
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.01320880
ELECTRON MICROSCOPYf_angle_d1.69329159
ELECTRON MICROSCOPYf_chiral_restr0.0763192
ELECTRON MICROSCOPYf_plane_restr0.0072994
ELECTRON MICROSCOPYf_dihedral_angle_d25.3318168
LS refinement shell

Refine-ID: ELECTRON MICROSCOPY / Total num. of bins used: 4 / % reflection obs: 100 %

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkNum. reflection all
4.6306-5.09660.59051320.54816903769169
5.0966-5.83420.51811440.52166934269486
5.8342-7.35050.48311080.50896955969667
7.3505-236.46760.34851670.35126892269089
Refinement TLS params.Method: refined / Origin x: 113.7436 Å / Origin y: 115.0465 Å / Origin z: 119.5786 Å
111213212223313233
T0.5058 Å2-0.1803 Å20.0956 Å2-0.4103 Å2-0.1405 Å2--0.19 Å2
L1.7574 °2-0.9085 °20.0842 °2-1.9929 °2-1.0694 °2--1.2073 °2
S0.1105 Å °0.4677 Å °-0.4973 Å °-0.5094 Å °0.9294 Å °-0.6722 Å °0.4074 Å °1.4146 Å °0.6086 Å °
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1ELECTRON MICROSCOPY1allA408 - 1029
2ELECTRON MICROSCOPY1allA1030 - 1101
3ELECTRON MICROSCOPY1allB1 - 351
4ELECTRON MICROSCOPY1allC408 - 1029
5ELECTRON MICROSCOPY1allC1030 - 1101
6ELECTRON MICROSCOPY1allD1 - 351
7ELECTRON MICROSCOPY1allE1 - 34
8ELECTRON MICROSCOPY1allF1 - 50
9ELECTRON MICROSCOPY1allG1 - 61
10ELECTRON MICROSCOPY1allH1 - 45
11ELECTRON MICROSCOPY1allI1 - 16
12ELECTRON MICROSCOPY1allJ1 - 16

+
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