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Yorodumi- PDB-7o24: Structure of the foamy viral protease-reverse transcriptase in co... -
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-Basic information
Entry | Database: PDB / ID: 7o24 | ||||||
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Title | Structure of the foamy viral protease-reverse transcriptase in complex with dsDNA. | ||||||
Components |
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Keywords | VIRAL PROTEIN / reverse trancscriptase / complex with dsDNA | ||||||
Function / homology | Function and homology information DNA integration / virion component / viral penetration into host nucleus / RNA-directed DNA polymerase activity / host cell / RNA-DNA hybrid ribonuclease activity / aspartic-type endopeptidase activity / DNA recombination / nucleic acid binding / host cell cytoplasm ...DNA integration / virion component / viral penetration into host nucleus / RNA-directed DNA polymerase activity / host cell / RNA-DNA hybrid ribonuclease activity / aspartic-type endopeptidase activity / DNA recombination / nucleic acid binding / host cell cytoplasm / DNA-directed DNA polymerase activity / proteolysis / metal ion binding / cytoplasm Similarity search - Function | ||||||
Biological species | White-tufted-ear marmoset simian foamy virus synthetic construct (others) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.8 Å | ||||||
Authors | Nowotny, M. / Czarnocki-Cieciura, M. | ||||||
Funding support | Poland, 1items
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Citation | Journal: J Virol / Year: 2021 Title: Structures of Substrate Complexes of Foamy Viral Protease-Reverse Transcriptase. Authors: Marzena Nowacka / Elżbieta Nowak / Mariusz Czarnocki-Cieciura / Justyna Jackiewicz / Krzysztof Skowronek / Roman H Szczepanowski / Birgitta M Wöhrl / Marcin Nowotny / Abstract: Reverse transcriptases (RTs) use their DNA polymerase and RNase H activities to catalyze the conversion of single-stranded RNA to double-stranded DNA (dsDNA), a crucial process for the replication of ...Reverse transcriptases (RTs) use their DNA polymerase and RNase H activities to catalyze the conversion of single-stranded RNA to double-stranded DNA (dsDNA), a crucial process for the replication of retroviruses. Foamy viruses (FVs) possess a unique RT, which is a fusion with the protease (PR) domain. The mechanism of substrate binding by this enzyme has been unknown. Here, we report a crystal structure of monomeric full-length marmoset FV (MFV) PR-RT in complex with an RNA/DNA hybrid substrate. We also describe a structure of MFV PR-RT with an RNase H deletion in complex with a dsDNA substrate in which the enzyme forms an asymmetric homodimer. Cryo-electron microscopy reconstruction of the full-length MFV PR-RT-dsDNA complex confirmed the dimeric architecture. These findings represent the first structural description of nucleic acid binding by a foamy viral RT and demonstrate its ability to change its oligomeric state depending on the type of bound nucleic acid. Reverse transcriptases (RTs) are intriguing enzymes converting single-stranded RNA to dsDNA. Their activity is essential for retroviruses, which are divided into two subfamilies differing significantly in their life cycles: and . The latter family is much more ancient and comprises five genera. A unique feature of foamy viral RTs is that they contain N-terminal protease (PR) domains, which are not present in orthoretroviral enzymes. So far, no structural information for full-length foamy viral PR-RT interacting with nucleic substrates has been reported. Here, we present crystal and cryo-electron microscopy structures of marmoset foamy virus (MFV) PR-RT. These structures revealed the mode of binding of RNA/DNA and dsDNA substrates. Moreover, unexpectedly, the structures and biochemical data showed that foamy viral PR-RT can adopt both a monomeric configuration, which is observed in our structures in the presence of an RNA/DNA hybrid, and an asymmetric dimer arrangement, which we observed in the presence of dsDNA. | ||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 7o24.cif.gz | 235.5 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7o24.ent.gz | 175.2 KB | Display | PDB format |
PDBx/mmJSON format | 7o24.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7o24_validation.pdf.gz | 1.5 MB | Display | wwPDB validaton report |
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Full document | 7o24_full_validation.pdf.gz | 1.5 MB | Display | |
Data in XML | 7o24_validation.xml.gz | 50.9 KB | Display | |
Data in CIF | 7o24_validation.cif.gz | 74.4 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/o2/7o24 ftp://data.pdbj.org/pub/pdb/validation_reports/o2/7o24 | HTTPS FTP |
-Related structure data
Related structure data | 12698MC 7o0gC 7o0hC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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-Components
#1: Protein | Mass: 85467.289 Da / Num. of mol.: 3 Source method: isolated from a genetically manipulated source Details: foamy virus Source: (gene. exp.) White-tufted-ear marmoset simian foamy virus Gene: pol Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) References: UniProt: D5JWV1, RNA-directed DNA polymerase, DNA-directed DNA polymerase, ribonuclease H #2: DNA chain | | Mass: 7347.770 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) #3: DNA chain | | Mass: 6783.375 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
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Molecular weight | Value: 0.19 MDa / Experimental value: YES | ||||||||||||||||||||||||
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Source (recombinant) |
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Buffer solution | pH: 7 | ||||||||||||||||||||||||
Specimen | Conc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||
Specimen support | Grid material: GOLD / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R2/1 | ||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 277 K |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD |
Image recording | Electron dose: 40 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
-Processing
Software | Name: PHENIX / Version: 1.19_4092: / Classification: refinement | |||||||||||||||||||||||||||||||||||
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 344421 | |||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 4.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 20071 / Symmetry type: POINT | |||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: RIGID BODY FIT | |||||||||||||||||||||||||||||||||||
Refine LS restraints |
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