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Open data
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Basic information
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Title | Cryo-EM structure of PfAgo-guide DNA-target DNA complex | |||||||||
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![]() | nuclease / GENE REGULATION | |||||||||
Function / homology | ![]() Hydrolases; Acting on ester bonds; Site specific endodeoxyribonucleases: cleavage is not sequence specific (deleted sub-subclass) / clearance of foreign intracellular DNA / DNA endonuclease activity / manganese ion binding / DNA binding / RNA binding Similarity search - Function | |||||||||
Biological species | ![]() ![]() ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.9 Å | |||||||||
![]() | Zhuang L | |||||||||
Funding support | 1 items
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![]() | ![]() Title: Molecular mechanism for target recognition, dimerization, and activation of Pyrococcus furiosus Argonaute. Authors: Longyu Wang / Wanping Chen / Chendi Zhang / Xiaochen Xie / Fuyong Huang / Miaomiao Chen / Wuxiang Mao / Na Yu / Qiang Wei / Lixin Ma / Zhuang Li / ![]() Abstract: The Argonaute nuclease from the thermophilic archaeon Pyrococcus furiosus (PfAgo) contributes to host defense and represents a promising biotechnology tool. Here, we report the structure of a PfAgo- ...The Argonaute nuclease from the thermophilic archaeon Pyrococcus furiosus (PfAgo) contributes to host defense and represents a promising biotechnology tool. Here, we report the structure of a PfAgo-guide DNA-target DNA ternary complex at the cleavage-compatible state. The ternary complex is predominantly dimerized, and the dimerization is solely mediated by PfAgo at PIWI-MID, PIWI-PIWI, and PAZ-N interfaces. Additionally, PfAgo accommodates a short 14-bp guide-target DNA duplex with a wedge-type N domain and specifically recognizes 5'-phosphorylated guide DNA. In contrast, the PfAgo-guide DNA binary complex is monomeric, and the engagement of target DNA with 14-bp complementarity induces sufficient dimerization and activation of PfAgo, accompanied by movement of PAZ and N domains. A closely related Argonaute from Thermococcus thioreducens adopts a similar dimerization configuration with an additional zinc finger formed at the dimerization interface. Dimerization of both Argonautes stabilizes the catalytic loops, highlighting the important role of Argonaute dimerization in the activation and target cleavage. | |||||||||
History |
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Structure visualization
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 28.7 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 16.5 KB 16.5 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 6.6 KB | Display | ![]() |
Images | ![]() | 98.8 KB | ||
Filedesc metadata | ![]() | 6 KB | ||
Others | ![]() ![]() | 28.2 MB 28.3 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 8jpxMC ![]() 8wd8C M: atomic model generated by this map C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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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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Voxel size | X=Y=Z: 0.85 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: #1
File | emd_36489_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #2
File | emd_36489_half_map_2.map | ||||||||||||
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Density Histograms |
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Sample components
-Entire : Cryo-EM structure of a PfAgo-guide DNA-target DNA complex
Entire | Name: Cryo-EM structure of a PfAgo-guide DNA-target DNA complex |
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Components |
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-Supramolecule #1: Cryo-EM structure of a PfAgo-guide DNA-target DNA complex
Supramolecule | Name: Cryo-EM structure of a PfAgo-guide DNA-target DNA complex type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#4 |
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Source (natural) | Organism: ![]() ![]() |
-Macromolecule #1: Protein argonaute
Macromolecule | Name: Protein argonaute / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO EC number: Hydrolases; Acting on ester bonds; Site specific endodeoxyribonucleases: cleavage is not sequence specific (deleted sub-subclass) |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 90.510094 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MKAIVVINLV KINKKIIPDK IYVYRLFNDP EEELQKEGYS IYRLAYENVG IVIDPENLII ATTKELEYEG EFIPEGEISF SELRNDYQS KLVLRLLKEN GIGEYELSKL LRKFRKPKTF GDYKVIPSVE MSVIKHDEDF YLVIHIIHQI QSMKTLWELV N KDPKELEE ...String: MKAIVVINLV KINKKIIPDK IYVYRLFNDP EEELQKEGYS IYRLAYENVG IVIDPENLII ATTKELEYEG EFIPEGEISF SELRNDYQS KLVLRLLKEN GIGEYELSKL LRKFRKPKTF GDYKVIPSVE MSVIKHDEDF YLVIHIIHQI QSMKTLWELV N KDPKELEE FLMTHKENLM LKDIASPLKT VYKPCFEEYT KKPKLDHNQE IVKYWYNYHI ERYWNTPEAK LEFYRKFGQV DL KQPAILA KFASKIKKNK NYKIYLLPQL VVPTYNAEQL ESDVAKEILE YTKLMPEERK ELLENILAEV DSDIIDKSLS EIE VEKIAQ ELENKIRVRD DKGNSVPISQ LNVQKSQLLL WTNYSRKYPV ILPYEVPEKF RKIREIPMFI ILDSGLLADI QNFA TNEFR ELVKSMYYSL AKKYNSLAKK ARSTNEIGLP FLDFRGKEKV ITEDLNSDKG IIEVVEQVSS FMKGKELGLA FIAAR NKLS SEKFEEIKRR LFNLNVISQV VNEDTLKNKR DKYDRNRLDL FVRHNLLFQV LSKLGVKYYV LDYRFNYDYI IGIDVA PMK RSEGYIGGSA VMFDSQGYIR KIVPIKIGEQ RGESVDMNEF FKEMVDKFKE FNIKLDNKKI LLLRDGRITN NEEEGLK YI SEMFDIEVVT MDVIKNHPVR AFANMKMYFN LGGAIYLIPH KLKQAKGTPI PIKLAKKRII KNGKVEKQSI TRQDVLDI F ILTRLNYGSI SADMRLPAPV HYAHKFANAI RNEWKIKEEF LAEGFLYFV UniProtKB: Protein argonaute |
-Macromolecule #2: Guide DNA
Macromolecule | Name: Guide DNA / type: dna / ID: 2 / Number of copies: 2 / Classification: DNA |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 5.337467 KDa |
Sequence | String: (DT)(DG)(DA)(DG)(DG)(DT)(DA)(DG)(DT)(DA) (DG)(DG)(DT)(DT)(DG)(DT)(DA) |
-Macromolecule #3: Target DNA
Macromolecule | Name: Target DNA / type: dna / ID: 3 / Number of copies: 2 / Classification: DNA |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 5.075327 KDa |
Sequence | String: (DA)(DC)(DA)(DA)(DC)(DC)(DT)(DA)(DC)(DT) (DA)(DC)(DC)(DT)(DC)(DA)(DT) |
-Macromolecule #4: Excess DNA
Macromolecule | Name: Excess DNA / type: dna / ID: 4 / Number of copies: 2 / Classification: DNA |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 1.780199 KDa |
Sequence | String: (DT)(DT)(DT)(DT)(DT)(DT) |
-Macromolecule #5: MAGNESIUM ION
Macromolecule | Name: MAGNESIUM ION / type: ligand / ID: 5 / Number of copies: 6 / Formula: MG |
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Molecular weight | Theoretical: 24.305 Da |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Buffer | pH: 7.5 |
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Vitrification | Cryogen name: ETHANE |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 52.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.2 µm / Nominal defocus min: 1.2 µm |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |