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- EMDB-33289: Cryo-EM structure of mTIP60-Ba (metal-ion induced TIP60 (K67E) co... -

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Basic information

Entry
Database: EMDB / ID: EMD-33289
TitleCryo-EM structure of mTIP60-Ba (metal-ion induced TIP60 (K67E) complex with barium ions
Map dataSharpened map by post-processing
Sample
  • Complex: mTIP60-Ba
    • Protein or peptide: TIP60 K67E mutant
  • Ligand: BARIUM ION
KeywordsArtificial designed protein complex / Protein cage / Protein nanoparticle / Metal-induced protein assembly / Protein metal complex / DE NOVO PROTEIN
Biological speciessynthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.96 Å
AuthorsOhara N / Kawakami N / Arai R / Adachi N / Moriya T / Kawasaki M / Miyamoto K
Funding support Japan, 4 items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS)JP19H02522 Japan
Japan Society for the Promotion of Science (JSPS)JP18K05324 Japan
Japan Society for the Promotion of Science (JSPS)JP17KK0104 Japan
Japan Agency for Medical Research and Development (AMED)JP21am0101071 Japan
CitationJournal: 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
DepositionApr 24, 2022-
Header (metadata) releaseJan 4, 2023-
Map releaseJan 4, 2023-
UpdateNov 29, 2023-
Current statusNov 29, 2023Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_33289.png

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Map

FileDownload / File: emd_33289.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSharpened map by post-processing
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
0.88 Å/pix.
x 360 pix.
= 316.8 Å		0.88 Å/pix.
x 360 pix.
= 316.8 Å		0.88 Å/pix.
x 360 pix.
= 316.8 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.88 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.023
Minimum - Maximum-0.050495904 - 0.0973035
Average (Standard dev.)0.00008180819 (±0.005705675)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 316.80002 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_33289_msk_1.map
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Additional map: Raw full map

Fileemd_33289_additional_1.map
AnnotationRaw full map
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Half map: Half map 1

Fileemd_33289_half_map_1.map
AnnotationHalf map 1
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Histogram

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Half map: Half map 2

Fileemd_33289_half_map_2.map
AnnotationHalf map 2
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Sample components

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Entire : mTIP60-Ba

EntireName: mTIP60-Ba
Components
  • Complex: mTIP60-Ba
    • Protein or peptide: TIP60 K67E mutant
  • Ligand: BARIUM ION

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Supramolecule #1: mTIP60-Ba

SupramoleculeName: mTIP60-Ba / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: metal-ion induced TIP60 (truncated icosahedral protein composed of 60-mer fusion proteins) complexed with barium ions
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 1.074 MDa

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Macromolecule #1: TIP60 K67E mutant

MacromoleculeName: TIP60 K67E mutant / type: protein_or_peptide / ID: 1 / Number of copies: 60 / Enantiomer: LEVO
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 17.79416 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
MGSSHHHHHH SQDPKNIKIM RLVTGEDIIG NISESQGLIT IKKAFVIIPM QATPGKPVQL VLSPWQPYTD DKEIVIDDSE VITITSPKD DIIKSYESHT RVLENKQVEE ILRLEKEIED LQRMKEQQEL SLTEASLQKL QERRDQELRR LEEE

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Macromolecule #2: BARIUM ION

MacromoleculeName: BARIUM ION / type: ligand / ID: 2 / Number of copies: 60 / Formula: BA
Molecular weightTheoretical: 137.327 Da

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration5 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
25.0 mMHEPES2-[4-(2-hydroxyethyl)-1-piperazinyl]ethanesulfonic acid
100.0 mMNaClsodium chloride
0.3 mMBaCl2barium chloride
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Pretreatment - Atmosphere: AIR / Details: The grid was washed by acetone prior to use.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 291 K / Instrument: FEI VITROBOT MARK IV / Details: Blotting time was 15 seconds (blot force 5).

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Number grids imaged: 1 / Number real images: 2206 / Average exposure time: 66.42 sec. / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 120000
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 113155
Startup modelType of model: OTHER
Details: An ab initio model was generated using RELION3's own implementation of Stochastic Gradient Descent (SGD) algorithm and low-pass filtered to 60 A for use as an initial model for Refine3D (C1).
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: I (icosahedral) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.96 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 61251
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final 3D classificationNumber classes: 4 / Avg.num./class: 16165 / Software - Name: RELION (ver. 3.1)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

7eq9


Chain - Chain ID: A / Chain - Residue range: 2-136 / Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: OTHER / Overall B value: 22.77 / Target criteria: Correlation coefficient
Output model

PDB-7xm1:
Cryo-EM structure of mTIP60-Ba (metal-ion induced TIP60 (K67E) complex with barium ions

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