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- EMDB-22272: Structure of RAG1 (R848M/E649V)-RAG2-DNA Strand Transfer Complex ... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-22272 | |||||||||
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Title | Structure of RAG1 (R848M/E649V)-RAG2-DNA Strand Transfer Complex (Dynamic-Form) | |||||||||
![]() | RAG1 (R848M/E649V)-RAG2-DNA Strand Transfer Complex (Dynamic-Form) | |||||||||
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![]() | DNA Transposase / RECOMBINATION | |||||||||
Function / homology | ![]() mature B cell differentiation involved in immune response / DNA recombinase complex / B cell homeostatic proliferation / DN2 thymocyte differentiation / negative regulation of T cell differentiation in thymus / endodeoxyribonuclease complex / pre-B cell allelic exclusion / positive regulation of organ growth / regulation of behavioral fear response / V(D)J recombination ...mature B cell differentiation involved in immune response / DNA recombinase complex / B cell homeostatic proliferation / DN2 thymocyte differentiation / negative regulation of T cell differentiation in thymus / endodeoxyribonuclease complex / pre-B cell allelic exclusion / positive regulation of organ growth / regulation of behavioral fear response / V(D)J recombination / negative regulation of T cell apoptotic process / phosphatidylinositol-3,4-bisphosphate binding / negative regulation of thymocyte apoptotic process / phosphatidylinositol-3,5-bisphosphate binding / positive regulation of T cell differentiation / regulation of T cell differentiation / organ growth / T cell lineage commitment / B cell lineage commitment / T cell homeostasis / phosphatidylinositol-3,4,5-trisphosphate binding / T cell differentiation / protein autoubiquitination / phosphatidylinositol-4,5-bisphosphate binding / methylated histone binding / phosphatidylinositol binding / B cell differentiation / thymus development / visual learning / RING-type E3 ubiquitin transferase / ubiquitin-protein transferase activity / ubiquitin protein ligase activity / chromatin organization / histone binding / T cell differentiation in thymus / endonuclease activity / DNA recombination / adaptive immune response / sequence-specific DNA binding / Hydrolases; Acting on ester bonds / defense response to bacterium / chromatin binding / protein homodimerization activity / zinc ion binding / nucleoplasm / identical protein binding / nucleus / metal ion binding Similarity search - Function | |||||||||
Biological species | ![]() ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.7 Å | |||||||||
![]() | Zhang Y / Corbett E | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Structural basis for the activation and suppression of transposition during evolution of the RAG recombinase. Authors: Yuhang Zhang / Elizabeth Corbett / Shenping Wu / David G Schatz / ![]() Abstract: Jawed vertebrate adaptive immunity relies on the RAG1/RAG2 (RAG) recombinase, a domesticated transposase, for assembly of antigen receptor genes. Using an integration-activated form of RAG1 with ...Jawed vertebrate adaptive immunity relies on the RAG1/RAG2 (RAG) recombinase, a domesticated transposase, for assembly of antigen receptor genes. Using an integration-activated form of RAG1 with methionine at residue 848 and cryo-electron microscopy, we determined structures that capture RAG engaged with transposon ends and U-shaped target DNA prior to integration (the target capture complex) and two forms of the RAG strand transfer complex that differ based on whether target site DNA is annealed or dynamic. Target site DNA base unstacking, flipping, and melting by RAG1 methionine 848 explain how this residue activates transposition, how RAG can stabilize sharp bends in target DNA, and why replacement of residue 848 by arginine during RAG domestication led to suppression of transposition activity. RAG2 extends a jawed vertebrate-specific loop to interact with target site DNA, and functional assays demonstrate that this loop represents another evolutionary adaptation acquired during RAG domestication to inhibit transposition. Our findings identify mechanistic principles of the final step in cut-and-paste transposition and the molecular and structural logic underlying the transformation of RAG from transposase to recombinase. | |||||||||
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 | ![]() | 51.9 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 21.4 KB 21.4 KB | Display Display | ![]() |
Images | ![]() | 171.6 KB | ||
Filedesc metadata | ![]() | 7.1 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 6xnxMC ![]() 6xnyC ![]() 6xnzC 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 | RAG1 (R848M/E649V)-RAG2-DNA Strand Transfer Complex (Dynamic-Form) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.05 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 : RAG recombinase strand transfer complex
+Supramolecule #1: RAG recombinase strand transfer complex
+Supramolecule #2: V(D)J recombination-activating protein 1, V(D)J recombination-act...
+Supramolecule #3: DNA
+Macromolecule #1: V(D)J recombination-activating protein 1
+Macromolecule #2: V(D)J recombination-activating protein 2
+Macromolecule #3: 12RSS integration strand DNA (55-MER)
+Macromolecule #4: 23RSS integration strand DNA (66-MER)
+Macromolecule #5: Flanking DNA top strand DNA
+Macromolecule #6: 23RSS signal top strand DNA (45-MER)
+Macromolecule #7: 12RSS signal top strand DNA (34-MER)
+Macromolecule #8: MAGNESIUM ION
+Macromolecule #9: ZINC ION
-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 Details: 20 mM HEPES pH7.6, 0.5 mM TCEP, 5 mM MgCl2 and 150 mM KCl |
Grid | Model: C-flat-2/1 / Material: COPPER / Mesh: 400 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 283 K / Instrument: FEI VITROBOT MARK IV |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Number real images: 3527 / Average exposure time: 11.0 sec. / Average electron dose: 72.8 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | C2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.3000000000000003 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 130000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |