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- PDB-9dze: Computationally Designed Bifaceted Protein Nanomaterial pD5-14 -

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Basic information

Entry
Database: PDB / ID: 9dze
TitleComputationally Designed Bifaceted Protein Nanomaterial pD5-14
Components
  • pD5-14 A component
  • pD5-14 B component
  • pD5-14 C component
  • pD5-14 D component
KeywordsDE NOVO PROTEIN / nanomaterial / 4 component / bifaceted / penton
Biological speciessynthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.3 Å
AuthorsCarr, K.D. / Borst, A.J. / Weidle, C.
Funding support United States, Sweden, 4items
OrganizationGrant numberCountry
Bill & Melinda Gates Foundation United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID) United States
Howard Hughes Medical Institute (HHMI) United States
Swedish Research Council Sweden
CitationJournal: bioRxiv / Year: 2024
Title: Computational design of bifaceted protein nanomaterials.
Authors: Sanela Rankovic / Kenneth D Carr / Justin Decarreau / Rebecca Skotheim / Ryan D Kibler / Sebastian Ols / Sangmin Lee / Jung-Ho Chun / Marti R Tooley / Justas Dauparas / Helen E Eisenach / ...Authors: Sanela Rankovic / Kenneth D Carr / Justin Decarreau / Rebecca Skotheim / Ryan D Kibler / Sebastian Ols / Sangmin Lee / Jung-Ho Chun / Marti R Tooley / Justas Dauparas / Helen E Eisenach / Matthias Glögl / Connor Weidle / Andrew J Borst / David Baker / Neil P King /
Abstract: Recent advances in computational methods have led to considerable progress in the design of self-assembling protein nanoparticles. However, nearly all nanoparticles designed to date exhibit strict ...Recent advances in computational methods have led to considerable progress in the design of self-assembling protein nanoparticles. However, nearly all nanoparticles designed to date exhibit strict point group symmetry, with each subunit occupying an identical, symmetrically related environment. This limits the structural diversity that can be achieved and precludes anisotropic functionalization. Here, we describe a general computational strategy for designing multi-component bifaceted protein nanomaterials with two distinctly addressable sides. The method centers on docking pseudosymmetric heterooligomeric building blocks in architectures with dihedral symmetry and designing an asymmetric protein-protein interface between them. We used this approach to obtain an initial 30-subunit assembly with pseudo-D5 symmetry, and then generated an additional 15 variants in which we controllably altered the size and morphology of the bifaceted nanoparticles by designing extensions to one of the subunits. Functionalization of the two distinct faces of the nanoparticles with protein minibinders enabled specific colocalization of two populations of polystyrene microparticles coated with target protein receptors. The ability to accurately design anisotropic protein nanomaterials with precisely tunable structures and functions could be broadly useful in applications that require colocalizing two or more distinct target moieties.
History
DepositionOct 16, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 30, 2024Provider: repository / Type: Initial release
Revision 1.1Nov 6, 2024Group: Data collection / Structure summary / Category: audit_author / em_admin / em_author_list
Item: _audit_author.name / _em_admin.last_update / _em_author_list.author
Revision 1.2Nov 13, 2024Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.pdbx_database_id_PubMed / _citation.title ..._citation.pdbx_database_id_PubMed / _citation.title / _citation_author.name / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: pD5-14 A component
B: pD5-14 A component
C: pD5-14 A component
D: pD5-14 A component
E: pD5-14 A component
F: pD5-14 A component
G: pD5-14 A component
H: pD5-14 A component
I: pD5-14 A component
J: pD5-14 A component
K: pD5-14 B component
L: pD5-14 B component
M: pD5-14 B component
N: pD5-14 B component
O: pD5-14 B component
P: pD5-14 B component
Q: pD5-14 B component
R: pD5-14 B component
S: pD5-14 B component
T: pD5-14 B component
a: pD5-14 C component
b: pD5-14 C component
c: pD5-14 C component
d: pD5-14 C component
e: pD5-14 C component
f: pD5-14 D component
g: pD5-14 D component
h: pD5-14 D component
i: pD5-14 D component
j: pD5-14 D component


Theoretical massNumber of molelcules
Total (without water)1,257,93130
Polymers1,257,93130
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Protein
pD5-14 A component


Mass: 35302.070 Da / Num. of mol.: 10
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others) / Production host: Escherichia coli (E. coli)
#2: Protein
pD5-14 B component


Mass: 34050.781 Da / Num. of mol.: 10
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others) / Production host: Escherichia coli (E. coli)
#3: Protein
pD5-14 C component


Mass: 56306.781 Da / Num. of mol.: 5
Source method: isolated from a genetically manipulated source
Details: mScarlet fused to the N-terminus of the C component of pD5-14
Source: (gene. exp.) synthetic construct (others) / Production host: Escherichia coli (E. coli)
#4: Protein
pD5-14 D component


Mass: 56573.711 Da / Num. of mol.: 5
Source method: isolated from a genetically manipulated source
Details: mNeonGreen fused to the N-terminus of the D component of pD5-14
Source: (gene. exp.) synthetic construct (others) / Production host: Escherichia coli (E. coli)
Has protein modificationN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Computationally Designed Bifaceted Protein Nanomaterial pD5-14
Type: COMPLEX
Details: Chains were expressed separately in E. coli. Batches of A, B, and C components or A, B, and D components were mixed, subsequently lysed and centrifuged to yield 2 distinct species of cyclic ...Details: Chains were expressed separately in E. coli. Batches of A, B, and C components or A, B, and D components were mixed, subsequently lysed and centrifuged to yield 2 distinct species of cyclic assemblies - (ABC)5 and (ABD)5. Following IMAC and SEC, (ABC)5 and (ABD)5 assemblies were mixed to form pseudo-D5 (ABC)5-(ABD)5 assembly pD5-14.
Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 1.256 MDa / Experimental value: NO
Source (natural)Organism: synthetic construct (others)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 8
SpecimenConc.: 1.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: EMS Lacey Carbon
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 295.15 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 1800 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm
Image recordingElectron dose: 45.21 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of real images: 4871
EM imaging opticsEnergyfilter name: GIF Bioquantum

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Processing

EM software
IDNameVersionCategory
1cryoSPARC4.4.0particle selection
4cryoSPARC4.4.0CTF correction
7UCSF ChimeraXmodel fitting
9PHENIXmodel refinement
10ISOLDEmodel refinement
11Cootmodel refinement
12cryoSPARC4.4.0initial Euler assignment
13cryoSPARC4.4.0final Euler assignment
14cryoSPARC4.4.0classification
15cryoSPARC4.4.03D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 640897
SymmetryPoint symmetry: C5 (5 fold cyclic)
3D reconstructionResolution: 4.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 209004 / Algorithm: FOURIER SPACE / Num. of class averages: 18 / Symmetry type: POINT
Atomic model buildingB value: 239.9 / Protocol: OTHER / Space: REAL / Target criteria: Cross-correlation coefficient
Details: Initial fitting was performed in ChimeraX. Phenix, Namdinator, ISOLDE, and Coot were used for relaxation of the model to better fit the density.

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