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- PDB-8uyo: Structure of a recombinant human PNMA2 capsid -

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Basic information

Entry
Database: PDB / ID: 8uyo
TitleStructure of a recombinant human PNMA2 capsid
ComponentsParaneoplastic antigen Ma2
KeywordsVIRUS LIKE PARTICLE / capsid / endogenous retrovirus-like particle / paraneoplasm / antigen
Function / homologyParaneoplastic antigen Ma / : / : / PNMA / PNMA N-terminal RRM-like domain / positive regulation of apoptotic process / nucleolus / Paraneoplastic antigen Ma2
Function and homology information
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsWilkinson, M.E. / Madigan, V. / Zhang, Y. / Zhang, F.
Funding support United States, 2items
OrganizationGrant numberCountry
Howard Hughes Medical Institute (HHMI) United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID) United States
CitationJournal: Proc Natl Acad Sci U S A / Year: 2024
Title: Human paraneoplastic antigen Ma2 (PNMA2) forms icosahedral capsids that can be engineered for mRNA delivery.
Authors: Victoria Madigan / Yugang Zhang / Rumya Raghavan / Max E Wilkinson / Guilhem Faure / Elena Puccio / Michael Segel / Blake Lash / Rhiannon K Macrae / Feng Zhang /
Abstract: A number of endogenous genes in the human genome encode retroviral -like proteins, which were domesticated from ancient retroelements. The paraneoplastic Ma antigen (PNMA) family members encode a - ...A number of endogenous genes in the human genome encode retroviral -like proteins, which were domesticated from ancient retroelements. The paraneoplastic Ma antigen (PNMA) family members encode a -like capsid domain, but their ability to assemble as capsids and traffic between cells remains mostly uncharacterized. Here, we systematically investigate human PNMA proteins and find that a number of PNMAs are secreted by human cells. We determine that PNMA2 forms icosahedral capsids efficiently but does not naturally encapsidate nucleic acids. We resolve the cryoelectron microscopy (cryo-EM) structure of PNMA2 and leverage the structure to design engineered PNMA2 (ePNMA2) particles with RNA packaging abilities. Recombinantly purified ePNMA2 proteins package mRNA molecules into icosahedral capsids and can function as delivery vehicles in mammalian cell lines, demonstrating the potential for engineered endogenous capsids as a nucleic acid therapy delivery modality.
History
DepositionNov 13, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Mar 20, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
1: Paraneoplastic antigen Ma2


Theoretical massNumber of molelcules
Total (without water)41,5571
Polymers41,5571
Non-polymers00
Water00
1
1: Paraneoplastic antigen Ma2
x 60


Theoretical massNumber of molelcules
Total (without water)2,493,43760
Polymers2,493,43760
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation59
2


  • Idetical with deposited unit
  • icosahedral asymmetric unit
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
3
1: Paraneoplastic antigen Ma2
x 5


  • icosahedral pentamer
  • 208 kDa, 5 polymers
Theoretical massNumber of molelcules
Total (without water)207,7865
Polymers207,7865
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation4
4
1: Paraneoplastic antigen Ma2
x 6


  • icosahedral 23 hexamer
  • 249 kDa, 6 polymers
Theoretical massNumber of molelcules
Total (without water)249,3446
Polymers249,3446
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation5
5


  • Idetical with deposited unit in distinct coordinate
  • icosahedral asymmetric unit, std point frame
TypeNameSymmetry operationNumber
transform to point frame1
SymmetryPoint symmetry: (Schoenflies symbol: I (icosahedral))

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Components

#1: Protein Paraneoplastic antigen Ma2 / 40 kDa neuronal protein / Onconeuronal antigen Ma2 / Paraneoplastic neuronal antigen MM2


Mass: 41557.289 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PNMA2, KIAA0883, MA2 / Production host: Escherichia coli (E. coli) / Strain (production host): Rosetta 2 (DE3) pLysS / References: UniProt: Q9UL42

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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: PNMA2 icosahedral capsid / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 2.49 MDa / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 7.4 / Details: phosphate buffered saline, pH 7.4
SpecimenConc.: 1.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 130000 X / Nominal defocus max: 2600 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 0.6 sec. / Electron dose: 30.68 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 17600

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Processing

EM software
IDNameVersionCategory
1Topazparticle selection
2EPUimage acquisition
7Cootmodel fitting
8ISOLDEmodel fitting
11RELION4.1final Euler assignment
13RELION4.13D reconstruction
14PHENIXmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 722571
SymmetryPoint symmetry: I (icosahedral)
3D reconstructionResolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 88320 / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL
Atomic model buildingAccession code: AF-Q9UL42-F1 / Source name: AlphaFold / Type: in silico model

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