[English] 日本語
Yorodumi- PDB-7xm1: Cryo-EM structure of mTIP60-Ba (metal-ion induced TIP60 (K67E) co... -
+Open data
-Basic information
Entry | Database: PDB / ID: 7xm1 | |||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Title | Cryo-EM structure of mTIP60-Ba (metal-ion induced TIP60 (K67E) complex with barium ions | |||||||||||||||
Components | TIP60 K67E mutant | |||||||||||||||
Keywords | DE NOVO PROTEIN / Artificial designed protein complex / Protein cage / Protein nanoparticle / Metal-induced protein assembly / Protein metal complex | |||||||||||||||
Function / homology | : Function and homology information | |||||||||||||||
Biological species | synthetic construct (others) | |||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.96 Å | |||||||||||||||
Authors | Ohara, N. / Kawakami, N. / Arai, R. / Adachi, N. / Moriya, T. / Kawasaki, M. / Miyamoto, K. | |||||||||||||||
Funding support | Japan, 4items
| |||||||||||||||
Citation | Journal: J Am Chem Soc / Year: 2023 Title: Reversible Assembly of an Artificial Protein Nanocage Using Alkaline Earth Metal Ions. Authors: Naoya Ohara / Norifumi Kawakami / Ryoichi Arai / Naruhiko Adachi / Toshio Moriya / Masato Kawasaki / Kenji Miyamoto / Abstract: Protein nanocages are of increasing interest for use as drug capsules, but the encapsulation and release of drug molecules at appropriate times require the reversible association and dissociation of ...Protein nanocages are of increasing interest for use as drug capsules, but the encapsulation and release of drug molecules at appropriate times require the reversible association and dissociation of the nanocages. One promising approach to addressing this challenge is the design of metal-dependent associating proteins. Such designed proteins typically have Cys or His residues at the protein surface for connecting the associating proteins through metal-ion coordination. However, Cys and His residues favor interactions with soft and borderline metal ions, such as Au and Zn, classified by the hard and soft acids and bases concept, restricting the types of metal ions available to drive association. Here, we show the alkaline earth (AE) metal-dependent association of the recently designed artificial protein nanocage TIP60, which is composed of 60-mer fusion proteins. The introduction of a Glu (hard base) mutation to the fusion protein (K67E mutant) prevented the formation of the 60-mer but formed the expected cage structure in the presence of Ca, Sr, or Ba ions (hard acids). Cryogenic electron microscopy (cryo-EM) analysis indicated a Ba ion at the interface of the subunits. Furthermore, we demonstrated the encapsulation and release of single-stranded DNA molecules using this system. Our results provide insights into the design of AE metal-dependent association and dissociation mechanisms for proteins. #1: Journal: Angew Chem Int Ed Engl / Year: 2018 Title: Design of Hollow Protein Nanoparticles with Modifiable Interior and Exterior Surfaces. Authors: Norifumi Kawakami / Hiroki Kondo / Yuki Matsuzawa / Kaoru Hayasaka / Erika Nasu / Kenji Sasahara / Ryoichi Arai / Kenji Miyamoto / Abstract: Protein-based nanoparticles hold promise for a broad range of applications. Here, we report the production of a uniform anionic hollow protein nanoparticle, designated TIP60, which spontaneously ...Protein-based nanoparticles hold promise for a broad range of applications. Here, we report the production of a uniform anionic hollow protein nanoparticle, designated TIP60, which spontaneously assembles from a designed fusion protein subunit based on the geometric features of polyhedra. We show that TIP60 tolerates mutation and both its interior and exterior surfaces can be chemically modified. Moreover, TIP60 forms larger structures upon the addition of a cationic protein. Therefore, TIP60 can be used as a modifiable nano-building block for further molecular assembly. #2: Journal: Chem Commun (Camb) / Year: 2021 Title: Icosahedral 60-meric porous structure of designed supramolecular protein nanoparticle TIP60. Authors: Junya Obata / Norifumi Kawakami / Akihisa Tsutsumi / Erika Nasu / Kenji Miyamoto / Masahide Kikkawa / Ryoichi Arai / Abstract: Supramolecular protein nanoparticles and nanocages have potential in a broad range of applications. Recently, we developed a uniform supramolecular protein nanoparticle, TIP60, symmmetrically self- ...Supramolecular protein nanoparticles and nanocages have potential in a broad range of applications. Recently, we developed a uniform supramolecular protein nanoparticle, TIP60, symmmetrically self-assembled from fusion proteins of a pentameric Sm-like protein and a dimeric MyoX-coil domain. Herein, we report the icosahedral 60-meric structure of TIP60 solved using single-particle cryo-electron microscopy. Interestingly, the structure revealed 20 regular-triangle-like pores on the surface. TIP60 and its mutants have many modifiable sites on their exterior and interior surfaces. The TIP60 architecture will be useful in the development of biomedical and biochemical nanoparticles/nanocages for future applications. | |||||||||||||||
History |
|
-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
---|
-Downloads & links
-Download
PDBx/mmCIF format | 7xm1.cif.gz | 1.3 MB | Display | PDBx/mmCIF format |
---|---|---|---|---|
PDB format | pdb7xm1.ent.gz | Display | PDB format | |
PDBx/mmJSON format | 7xm1.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7xm1_validation.pdf.gz | 1.8 MB | Display | wwPDB validaton report |
---|---|---|---|---|
Full document | 7xm1_full_validation.pdf.gz | 1.8 MB | Display | |
Data in XML | 7xm1_validation.xml.gz | 132 KB | Display | |
Data in CIF | 7xm1_validation.cif.gz | 224.6 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/xm/7xm1 ftp://data.pdbj.org/pub/pdb/validation_reports/xm/7xm1 | HTTPS FTP |
-Related structure data
Related structure data | 33289MC M: map data used to model this data C: citing same article (ref.) |
---|---|
Similar structure data | Similarity search - Function & homologyF&H Search |
Other databases |
-Links
-Assembly
Deposited unit |
|
---|---|
1 |
|
-Components
#1: Protein | Mass: 17794.160 Da / Num. of mol.: 60 Source method: isolated from a genetically manipulated source Source: (gene. exp.) synthetic construct (others) / Gene: TIP60 / Variant: K67E / Plasmid: pETDuet-1 / Production host: Escherichia coli BL21(DE3) (bacteria) #2: Chemical | ChemComp-BA / Has ligand of interest | Y | |
---|
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
---|---|
EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: mTIP60-Ba / Type: COMPLEX Details: metal-ion induced TIP60 (truncated icosahedral protein composed of 60-mer fusion proteins) complexed with barium ions Entity ID: #1 / Source: RECOMBINANT | ||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Molecular weight | Value: 1.074 MDa / Experimental value: YES | ||||||||||||||||||||
Source (natural) | Organism: synthetic construct (others) | ||||||||||||||||||||
Source (recombinant) | Organism: Escherichia coli BL21(DE3) (bacteria) / Plasmid: pETDuet-1 | ||||||||||||||||||||
Buffer solution | pH: 8 | ||||||||||||||||||||
Buffer component |
| ||||||||||||||||||||
Specimen | Conc.: 5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||
Specimen support | Details: The grid was washed by acetone prior to use. / Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3 | ||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 291 K / Details: Blotting time was 15 seconds (blot force 5) |
-Electron microscopy imaging
Experimental equipment | Model: Talos Arctica / Image courtesy: FEI Company |
---|---|
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: 120000 X / Nominal defocus max: 2500 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm |
Specimen holder | Cryogen: NITROGEN |
Image recording | Average exposure time: 66.42 sec. / Electron dose: 50 e/Å2 / Detector mode: COUNTING / Film or detector model: FEI FALCON III (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 2206 |
Image scans | Width: 4096 / Height: 4096 |
-Processing
Software |
| ||||||||||||||||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
EM software |
| ||||||||||||||||||||||||||||||||||||||||||||
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 113155 | ||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: I (icosahedral) | ||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.96 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 61251 / Algorithm: FOURIER SPACE / Num. of class averages: 1 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | B value: 22.77 / Protocol: OTHER / Space: REAL / Target criteria: Correlation coefficient | ||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 7EQ9 Pdb chain-ID: A / Accession code: 7EQ9 / Pdb chain residue range: 2-136 / Source name: PDB / Type: experimental model | ||||||||||||||||||||||||||||||||||||||||||||
Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 22.84 Å2 | ||||||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
|