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- EMDB-7046: BbRAGL-3'TIR synaptic complex with nicked DNA refined with C2 symmetry -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-7046 | |||||||||
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Title | BbRAGL-3'TIR synaptic complex with nicked DNA refined with C2 symmetry | |||||||||
![]() | BbRAGL-31TIR synaptic complex with nicked DNA refined with C2 symmetry | |||||||||
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![]() | DNA transposase / DNA cut and paste transposition / DDE family RNase H fold DNA transposase / RECOMBINATION | |||||||||
Function / homology | ![]() : / V(D)J recombination / B cell differentiation / RING-type E3 ubiquitin transferase / ubiquitin protein ligase activity / histone binding / T cell differentiation in thymus / endonuclease activity / sequence-specific DNA binding / Hydrolases; Acting on ester bonds ...: / V(D)J recombination / B cell differentiation / RING-type E3 ubiquitin transferase / ubiquitin protein ligase activity / histone binding / T cell differentiation in thymus / endonuclease activity / sequence-specific DNA binding / Hydrolases; Acting on ester bonds / protein homodimerization activity / zinc ion binding / nucleus Similarity search - Function | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.3 Å | |||||||||
![]() | Zhang Y / Cheng TC | |||||||||
Funding support | ![]() ![]()
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![]() | ![]() Title: Transposon molecular domestication and the evolution of the RAG recombinase. Authors: Yuhang Zhang / Tat Cheung Cheng / Guangrui Huang / Qingyi Lu / Marius D Surleac / Jeffrey D Mandell / Pierre Pontarotti / Andrei J Petrescu / Anlong Xu / Yong Xiong / David G Schatz / ![]() ![]() ![]() ![]() Abstract: Domestication of a transposon (a DNA sequence that can change its position in a genome) to give rise to the RAG1-RAG2 recombinase (RAG) and V(D)J recombination, which produces the diverse repertoire ...Domestication of a transposon (a DNA sequence that can change its position in a genome) to give rise to the RAG1-RAG2 recombinase (RAG) and V(D)J recombination, which produces the diverse repertoire of antibodies and T cell receptors, was a pivotal event in the evolution of the adaptive immune system of jawed vertebrates. The evolutionary adaptations that transformed the ancestral RAG transposase into a RAG recombinase with appropriately regulated DNA cleavage and transposition activities are not understood. Here, beginning with cryo-electron microscopy structures of the amphioxus ProtoRAG transposase (an evolutionary relative of RAG), we identify amino acid residues and domains the acquisition or loss of which underpins the propensity of RAG for coupled cleavage, its preference for asymmetric DNA substrates and its inability to perform transposition in cells. In particular, we identify two adaptations specific to jawed-vertebrates-arginine 848 in RAG1 and an acidic region in RAG2-that together suppress RAG-mediated transposition more than 1,000-fold. Our findings reveal a two-tiered mechanism for the suppression of RAG-mediated transposition, illuminate the evolution of V(D)J recombination and provide insight into the principles that govern the molecular domestication of transposons. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 20.3 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 19.3 KB 19.3 KB | Display Display | ![]() |
Images | ![]() | 81.9 KB | ||
Filedesc metadata | ![]() | 6.9 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 6b40MC ![]() 7043C ![]() 7044C ![]() 7045C M: atomic model generated by this map C: citing same article ( |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | BbRAGL-31TIR synaptic complex with nicked DNA refined with C2 symmetry | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.35 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : BbRAGL-31TIR synaptic complex with nicked DNA
Entire | Name: BbRAGL-31TIR synaptic complex with nicked DNA |
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Components |
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-Supramolecule #1: BbRAGL-31TIR synaptic complex with nicked DNA
Supramolecule | Name: BbRAGL-31TIR synaptic complex with nicked DNA / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#5 |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 350 KDa |
-Macromolecule #1: RAG1L,RAG1L
Macromolecule | Name: RAG1L,RAG1L / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 74.440133 KDa |
Recombinant expression | Organism: ![]() |
Sequence | String: ESLQRKRLCK ARLYDVTRHH VKHKRLKPLI EHIDEYCNEK DENKGDVLFF LLRSHLYDTG NRSMAEQIDT LWHGESLSCM TPEECLGMR LDMLMTKNQY SKEYNILKER GFSTLCPPKQ LDAIEKTLMP GTARYSIEGM DYSEHYFHSP VKMTGDLEVH S GETLEPDS ...String: ESLQRKRLCK ARLYDVTRHH VKHKRLKPLI EHIDEYCNEK DENKGDVLFF LLRSHLYDTG NRSMAEQIDT LWHGESLSCM TPEECLGMR LDMLMTKNQY SKEYNILKER GFSTLCPPKQ LDAIEKTLMP GTARYSIEGM DYSEHYFHSP VKMTGDLEVH S GETLEPDS VTMDFHEYVP DFPCPNTKGV RFPYAHAVAK TLEELEDEIV NGLKKLGRDP NDPTLVIHTI CKDGADGMGD VS VHKEKSD HLLPDKALRF SFCVLRCSVM HKDTEVTIYE DPNPNSVRSN RPVLECIGDE NDDGTVAVCV GPIECQRLLM KDK IMRVHM SDGTQRAHYL TFFNSMVDEK WDRAHGGLAG AGSKYLCTLC EAVRDEALEK AGSYKITRTL KKIEVTASKM KYES QEKDT FGVKGYPLLT TEPWERGIDA THTDINMGNY FKSLIVREMA QVHSWAKTAN VKKQIVDAES KLDKHLKESL GLNPT LMMA GNYARELFKA EHADKLVALV DKPDRKSALV EVLAKFRQLR KVYRANWPLN DMSDEVRQYK AKAVEMANDL KTHFPY APC TNYLHKVIEH VQELIEHPSG VGSVGALSSE GNEAGNKLFR QLRLGHARKG NTYNGLRDVL CTHWLYTSKT LRDKAA (UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK) (UNK)(UNK)(UNK) UniProtKB: V(D)J recombination-activating protein 1 |
-Macromolecule #5: RAG2L
Macromolecule | Name: RAG2L / type: protein_or_peptide / ID: 5 / Number of copies: 2 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 39.509254 KDa |
Recombinant expression | Organism: ![]() |
Sequence | String: MSSGPIFSSL VTDSSRSSRK KSDSALGTEF TVVCDRVPLR DPSPTVQRFL CFVFDNGSGA MSTLPIDVGG EGISLSDMKE VKAMPHIAS GCTAVWGPPL PPKPGKDTKQ VILWGGLDKR RWCCSNDLTQ VDITITPKTT TAKVSILPAD KQDGVPSPRT G HTLVAISS ...String: MSSGPIFSSL VTDSSRSSRK KSDSALGTEF TVVCDRVPLR DPSPTVQRFL CFVFDNGSGA MSTLPIDVGG EGISLSDMKE VKAMPHIAS GCTAVWGPPL PPKPGKDTKQ VILWGGLDKR RWCCSNDLTQ VDITITPKTT TAKVSILPAD KQDGVPSPRT G HTLVAISS LQAILFGGLE LASRHARLGT CAQSCKDGYF YLLDMTTLRW NKLPLPPLVP RAYHSSTWVP ASSTMVIVGG IT YSGHCPS ERLSVSDVVC LKISDTSQYT LTEIHMEGVR DSYVSSSSAS ALCDDRFVLY GGYHHDKSGL HPPEPSRDLY VMN LQTKKA VVHHAPTRMA SAGHTCLRLI DNSVVMIGGT CKSVNCCTNL UniProtKB: RAG2L |
-Macromolecule #2: 31TIR intact strand
Macromolecule | Name: 31TIR intact strand / type: dna / ID: 2 / Number of copies: 2 / Classification: DNA |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 19.218312 KDa |
Sequence | String: (DC)(DA)(DA)(DG)(DA)(DT)(DG)(DG)(DC)(DG) (DA)(DC)(DC)(DA)(DG)(DA)(DC)(DA)(DC)(DT) (DG)(DC)(DT)(DG)(DG)(DG)(DT)(DA)(DT) (DA)(DG)(DC)(DG)(DT)(DA)(DA)(DG)(DT)(DA) (DT) (DC)(DA)(DT)(DA)(DG)(DT) ...String: (DC)(DA)(DA)(DG)(DA)(DT)(DG)(DG)(DC)(DG) (DA)(DC)(DC)(DA)(DG)(DA)(DC)(DA)(DC)(DT) (DG)(DC)(DT)(DG)(DG)(DG)(DT)(DA)(DT) (DA)(DG)(DC)(DG)(DT)(DA)(DA)(DG)(DT)(DA) (DT) (DC)(DA)(DT)(DA)(DG)(DT)(DG)(DC) (DA)(DG)(DC)(DG)(DC)(DG)(DC)(DT)(DG)(DC) (DC)(DA) (DA)(DG) |
-Macromolecule #3: 31TIR pre-nicked strand of signal DNA
Macromolecule | Name: 31TIR pre-nicked strand of signal DNA / type: dna / ID: 3 / Number of copies: 2 / Classification: DNA |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 14.357206 KDa |
Sequence | String: (DC)(DA)(DC)(DT)(DA)(DT)(DG)(DA)(DT)(DA) (DC)(DT)(DT)(DA)(DC)(DG)(DC)(DT)(DA)(DT) (DA)(DC)(DC)(DC)(DA)(DG)(DC)(DA)(DG) (DT)(DG)(DT)(DC)(DT)(DG)(DG)(DT)(DC)(DG) (DC) (DC)(DA)(DT)(DC)(DT)(DT)(DG) |
-Macromolecule #4: 31TIR pre-nicked strand of flanking DNA
Macromolecule | Name: 31TIR pre-nicked strand of flanking DNA / type: dna / ID: 4 / Number of copies: 2 / Classification: DNA |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 4.601971 KDa |
Sequence | String: (DC)(DT)(DT)(DG)(DG)(DC)(DA)(DG)(DC)(DG) (DC)(DG)(DC)(DT)(DG) |
-Macromolecule #6: CALCIUM ION
Macromolecule | Name: CALCIUM ION / type: ligand / ID: 6 / Number of copies: 2 / Formula: CA |
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Molecular weight | Theoretical: 40.078 Da |
-Macromolecule #7: ZINC ION
Macromolecule | Name: ZINC ION / type: ligand / ID: 7 / Number of copies: 2 / Formula: ZN |
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Molecular weight | Theoretical: 65.409 Da |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.4 mg/mL |
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Buffer | pH: 7.6 |
Grid | Model: C-flat-1.2/1.3 4C / Material: COPPER / Mesh: 400 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 45 sec. / Pretreatment - Atmosphere: AIR |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 283 K / Instrument: FEI VITROBOT MARK III |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Specialist optics | Energy filter - Name: GIF Quantum LS / Energy filter - Lower energy threshold: 0 eV / Energy filter - Upper energy threshold: 20 eV |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Dimensions - Width: 7676 pixel / Digitization - Dimensions - Height: 7420 pixel / Digitization - Frames/image: 3-40 / Number grids imaged: 1 / Number real images: 4429 / Average exposure time: 10.0 sec. / Average electron dose: 54.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 0.01 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.2 µm / Nominal magnification: 81000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Image processing
-Atomic model buiding 1
Refinement | Space: REAL / Protocol: FLEXIBLE FIT |
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Output model | ![]() PDB-6b40: |