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Yorodumi- PDB-8dnl: Acidipropionibacterium acidipropionici encapsulin in an open stat... -
+Open data
-Basic information
Entry | Database: PDB / ID: 8dnl | ||||||
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Title | Acidipropionibacterium acidipropionici encapsulin in an open state at pH 7.5 | ||||||
Components | 29 kDa antigen cfp29 | ||||||
Keywords | VIRUS LIKE PARTICLE / Encapsulin | ||||||
Function / homology | Type 1 encapsulin shell protein / : / Encapsulating protein for peroxidase / encapsulin nanocompartment / 29 kDa antigen cfp29 Function and homology information | ||||||
Biological species | Acidipropionibacterium acidipropionici ATCC 4875 (bacteria) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.32 Å | ||||||
Authors | Jones, J.A. / Andreas, M.P. / Giessen, T.W. | ||||||
Funding support | United States, 1items
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Citation | Journal: Biomacromolecules / Year: 2023 Title: Exploring the Extreme Acid Tolerance of a Dynamic Protein Nanocage. Authors: Jesse A Jones / Michael P Andreas / Tobias W Giessen / Abstract: Encapsulins are microbial protein nanocages capable of efficient self-assembly and cargo enzyme encapsulation. Due to their favorable properties, including high thermostability, protease resistance, ...Encapsulins are microbial protein nanocages capable of efficient self-assembly and cargo enzyme encapsulation. Due to their favorable properties, including high thermostability, protease resistance, and robust heterologous expression, encapsulins have become popular bioengineering tools for applications in medicine, catalysis, and nanotechnology. Resistance against physicochemical extremes like high temperature and low pH is a highly desirable feature for many biotechnological applications. However, no systematic search for acid-stable encapsulins has been carried out, while the influence of pH on encapsulin shells has so far not been thoroughly explored. Here, we report on a newly identified encapsulin nanocage from the acid-tolerant bacterium . Using transmission electron microscopy, dynamic light scattering, and proteolytic assays, we demonstrate its extreme acid tolerance and resilience against proteases. We structurally characterize the novel nanocage using cryo-electron microscopy, revealing a dynamic five-fold pore that displays distinct "closed" and "open" states at neutral pH but only a singular "closed" state under strongly acidic conditions. Further, the "open" state exhibits the largest pore in an encapsulin shell reported to date. Non-native protein encapsulation capabilities are demonstrated, and the influence of external pH on internalized cargo is explored. Our results expand the biotechnological application range of encapsulin nanocages toward potential uses under strongly acidic conditions and highlight pH-responsive encapsulin pore dynamics. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8dnl.cif.gz | 93.8 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8dnl.ent.gz | 72.4 KB | Display | PDB format |
PDBx/mmJSON format | 8dnl.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8dnl_validation.pdf.gz | 1.6 MB | Display | wwPDB validaton report |
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Full document | 8dnl_full_validation.pdf.gz | 1.6 MB | Display | |
Data in XML | 8dnl_validation.xml.gz | 30.2 KB | Display | |
Data in CIF | 8dnl_validation.cif.gz | 40.6 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/dn/8dnl ftp://data.pdbj.org/pub/pdb/validation_reports/dn/8dnl | HTTPS FTP |
-Related structure data
Related structure data | 27573MC 8dn9C 8dnaC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
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Symmetry | Point symmetry: (Schoenflies symbol: I (icosahedral)) |
-Components
#1: Protein | Mass: 28522.725 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Acidipropionibacterium acidipropionici ATCC 4875 (bacteria) Strain: ATCC 4875 / DSM 20272 / JCM 6432 / NBRC 12425 / NCIMB 8070 / 4 Gene: PACID_10050 / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21(DE3) / References: UniProt: K7RV67 |
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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
Component | Name: Acidipropionibacterium acidipropionici encapsulin in an open state at pH 7.5 Type: COMPLEX / Entity ID: all / Source: RECOMBINANT | |||||||||||||||
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Molecular weight | Value: 1.71 MDa / Experimental value: YES | |||||||||||||||
Source (natural) | Organism: Acidipropionibacterium acidipropionici ATCC 4875 (bacteria) | |||||||||||||||
Source (recombinant) | Organism: Escherichia coli BL21(DE3) (bacteria) / Strain: BL21(DE3) | |||||||||||||||
Buffer solution | pH: 7.5 | |||||||||||||||
Buffer component |
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Specimen | Conc.: 3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | |||||||||||||||
Specimen support | Details: The grid was glow discharged for 60 seconds at 5 mA. Grid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R1.2/1.3 | |||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 295 K Details: Additional settings were : Blot force- 20, blot time - 4 seconds, drain time - 0 seconds, wait time - 0 seconds |
-Electron microscopy imaging
Experimental equipment | Model: Talos Arctica / Image courtesy: FEI Company |
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Microscopy | Model: FEI TALOS ARCTICA |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 45000 X / Nominal defocus max: 1800 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm |
Specimen holder | Cryogen: NITROGEN |
Image recording | Average exposure time: 4 sec. / Electron dose: 41 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 975 |
Image scans | Width: 3838 / Height: 3710 / Movie frames/image: 20 |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: I (icosahedral) | ||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.32 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 13581 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | B value: 77.81 / Protocol: AB INITIO MODEL / Space: REAL / Target criteria: Correlation coefficient Details: A starting model was created using RosettaFold. The starting model was placed in the map manually using UCSF Chimera. The model was then further refined using Coot and Phenix Real Space Refine. |