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Cryo-electron microscopy for the study of virus assembly

Abstract

Although viruses are extremely diverse in shape and size, evolution has led to a limited number of viral classes or lineages, which is probably linked to the assembly constraints of a viable capsid. Viral assembly mechanisms are restricted to two general pathways, (i) co-assembly of capsid proteins and single-stranded nucleic acids and (ii) a sequential mechanism in which scaffolding-mediated capsid precursor assembly is followed by genome packaging. Cryo-electron microscopy (cryo-EM) and cryo-electron tomography (cryo-ET), which are revolutionizing structural biology, are central to determining the high-resolution structures of many viral assemblies as well as those of assembly intermediates. This wealth of cryo-EM data has also led to the development and redesign of virus-based platforms for biomedical and biotechnological applications. In this Review, we will discuss recent viral assembly analyses by cryo-EM and cryo-ET showing how natural assembly mechanisms are used to encapsulate heterologous cargos including chemicals, enzymes, and/or nucleic acids for a variety of nanotechnological applications.

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Fig. 1: Overview of cryo-EM and cryo-ET workflow, from data acquisition to 3D model.
Fig. 2: Viral lineages and their representative folds and capsid structures.
Fig. 3: Co-assembly of CPs and viral single-stranded genome.
Fig. 4: Scaffolding protein-assisted capsid assembly.
Fig. 5: Assembly of dsRNA viruses assisted by a permanent scaffold protein shell.
Fig. 6: IBDV capsid assembly model.
Fig. 7: Cryo-ET of pleomorphic viruses and intracellular viral assembly.
Fig. 8: Assembly strategies to develop hybrid P22 and CCMV.

References

  1. 1.

    Harrison, S.C. In Fields Virology, Vol. 1 (eds. Knipe, D.M. et al.) 59–98 (Lippincott Williams & Wilkins, Philadelphia, 2007).

  2. 2.

    Castón, J. R. & Carrascosa, J. L. The basic architecture of viruses. Subcell. Biochem. 68, 53–75 (2013).

    PubMed  Google Scholar 

  3. 3.

    Kondylis, P., Schlicksup, C. J., Zlotnick, A. & Jacobson, S. C. Analytical techniques to characterize the structure, properties, and assembly of virus capsids. Anal. Chem. 91, 622–636 (2019).

    CAS  PubMed  Google Scholar 

  4. 4.

    Schlicksup, C. J. et al. Hepatitis B virus core protein allosteric modulators can distort and disrupt intact capsids. eLife 7, e31473 (2018).

    PubMed  PubMed Central  Google Scholar 

  5. 5.

    Schwarz, B., Uchida, M. & Douglas, T. Biomedical and catalytic opportunities of virus-like particles in nanotechnology. Adv. Virus Res. 97, 1–60 (2017).

    CAS  PubMed  Google Scholar 

  6. 6.

    Merk, A. et al. Breaking cryo-EM resolution barriers to facilitate drug discovery. Cell 165, 1698–1707 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  7. 7.

    Liu, Y., Huynh, D. T. & Yeates, T. O. A. A 3.8 Å resolution cryo-EM structure of a small protein bound to an imaging scaffold. Nat. Commun. 10, 1864 (2019).

    PubMed  PubMed Central  Google Scholar 

  8. 8.

    Lučič, V., Rigort, A. & Baumeister, W. Cryo-electron tomography: the challenge of doing structural biology in situ. J. Cell Biol. 202, 407–419 (2013).

    PubMed  PubMed Central  Google Scholar 

  9. 9.

    Wan, W. & Briggs, J. A. Cryo-electron tomography and subtomogram averaging. Methods Enzymol. 579, 329–367 (2016).

    CAS  PubMed  Google Scholar 

  10. 10.

    Schaffer, M. et al. Optimized cryo-focused ion beam sample preparation aimed at in situ structural studies of membrane proteins. J. Struct. Biol. 197, 73–82 (2017).

    CAS  PubMed  Google Scholar 

  11. 11.

    Schur, F. K. et al. An atomic model of HIV-1 capsid-SP1 reveals structures regulating assembly and maturation. Science 353, 506–508 (2016). This study describes the 3.9-Å resolution structure of the capsid protein and spacer peptide 1 in immature HIV-1 virions, calculated from cryo-ET and subtomogram averaging.

    CAS  PubMed  Google Scholar 

  12. 12.

    Jiang, W. & Tang, L. Atomic cryo-EM structures of viruses. Curr. Opin. Struct. Biol. 46, 122–129 (2017).

    PubMed  PubMed Central  Google Scholar 

  13. 13.

    Kaelber, J. T., Hryc, C. F. & Chiu, W. Electron cryomicroscopy of viruses at near-atomic resolutions. Annu. Rev. Virol. 4, 287–308 (2017).

    CAS  PubMed  Google Scholar 

  14. 14.

    Dubrovsky, A., Sorrentino, S., Harapin, J., Sapra, K. T. & Medalia, O. Developments in cryo-electron tomography for in situ structural analysis. Arch. Biochem. Biophys. 581, 78–85 (2015).

    CAS  PubMed  Google Scholar 

  15. 15.

    Abrescia, N. G., Bamford, D. H., Grimes, J. M. & Stuart, D. I. Structure unifies the viral universe. Annu. Rev. Biochem. 81, 795–822 (2012).

    CAS  PubMed  Google Scholar 

  16. 16.

    Bamford, D. H., Grimes, J. M. & Stuart, D. I. What does structure tell us about virus evolution? Curr. Opin. Struct. Biol. 15, 655–663 (2005).

    CAS  PubMed  Google Scholar 

  17. 17.

    Sinclair, R. M., Ravantti, J. J. & Bamford, D. H. Nucleic and amino acid sequences support structure-based viral classification. J. Virol. 91, e02275–16 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  18. 18.

    Caspar, D. L. D. & Klug, A. Physical principles in the construction of regular viruses. Cold Spring Harb. Symp. Quant. Biol. 27, 1–24 (1962).

    CAS  PubMed  Google Scholar 

  19. 19.

    Bahar, M. W., Graham, S. C., Stuart, D. I. & Grimes, J. M. Insights into the evolution of a complex virus from the crystal structure of vaccinia virus D13. Structure 19, 1011–1020 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  20. 20.

    Nasir, A. & Caetano-Anollés, G. Identification of capsid/coat related protein folds and their utility for virus classification. Front. Microbiol. 8, 380 (2017).

    PubMed  PubMed Central  Google Scholar 

  21. 21.

    Zamora, M. et al. Potyvirus virion structure shows conserved protein fold and RNA binding site in ssRNA viruses. Sci. Adv. 3, eaao2182 (2017).

    PubMed  PubMed Central  Google Scholar 

  22. 22.

    Valle, M. Structural homology between nucleoproteins of ssRNA Viruses. Subcell. Biochem. 88, 129–145 (2018).

    CAS  PubMed  Google Scholar 

  23. 23.

    Laanto, E. et al. Virus found in a boreal lake links ssDNA and dsDNA viruses. Proc. Natl. Acad. Sci. USA 114, 8378–8383 (2017).

    CAS  PubMed  Google Scholar 

  24. 24.

    Feiss, M. & Rao, V. B. The bacteriophage DNA packaging machine. Adv. Exp. Med. Biol. 726, 489–509 (2012).

    CAS  PubMed  Google Scholar 

  25. 25.

    Twarock, R., Bingham, R. J., Dykeman, E. C. & Stockley, P. G. A modelling paradigm for RNA virus assembly. Curr. Opin. Virol. 31, 74–81 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  26. 26.

    Twarock, R. & Stockley, P. G. RNA-mediated virus assembly: mechanisms and consequences for viral evolution and therapy. Annu. Rev. Biophys. 48, 495–514 (2019).

    CAS  PubMed  Google Scholar 

  27. 27.

    Hesketh, E. L. et al. Mechanisms of assembly and genome packaging in an RNA virus revealed by high-resolution cryo-EM. Nat. Commun. 6, 10113 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  28. 28.

    Dai, X. et al. In situ structures of the genome and genome-delivery apparatus in a single-stranded RNA virus. Nature 541, 112–116 (2017). This paper reports the asymmetrical cryo-EM structure of phage MS2 at 3.6-Å resolution. Most of the viral genome, a 3,569-nucleotide ssRNA molecule, is traced, and 16 stem-loops or packaging signals (as dsRNA segments) are identified.

    CAS  PubMed  Google Scholar 

  29. 29.

    Shakeel, S. et al. Genomic RNA folding mediates assembly of human parechovirus. Nat. Commun. 8, 5 (2017).

    PubMed  PubMed Central  Google Scholar 

  30. 30.

    Sarker, S. et al. Structural insights into the assembly and regulation of distinct viral capsid complexes. Nat. Commun. 7, 13014 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  31. 31.

    Patel, N. et al. HBV RNA pre-genome encodes specific motifs that mediate interactions with the viral core protein that promote nucleocapsid assembly. Nat. Microbiol. 2, 17098 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  32. 32.

    Steven, A. C., Heymann, J. B., Cheng, N., Trus, B. L. & Conway, J. F. Virus maturation: dynamics and mechanism of a stabilizing structural transition that leads to infectivity. Curr. Opin. Struct. Biol. 15, 227–236 (2005).

    CAS  PubMed  PubMed Central  Google Scholar 

  33. 33.

    Ignatiou, A. et al. Structural transitions during the scaffolding-driven assembly of a viral capsid. Nat. Commun. 10, 4840 (2019). This is a report of the molecular basis of the sequential structural rearrangement during viral capsid maturation of phage SPP1 (with a linear dsDNA genome). SP release from the procapsid leads to the stable expanded state, independently of DNA packaging.

    PubMed  PubMed Central  Google Scholar 

  34. 34.

    Yu, X., Jih, J., Jiang, J. & Zhou, Z. H. Atomic structure of the human cytomegalovirus capsid with its securing tegument layer of pp150. Science 356, eaam6892 (2017). Using electron-counting cryo-EM, the authors describe the highly pressurized nucleocapsid of human cytomegalovirus. The major CP is folded into seven domains; the floor domain at the shell has an HK97-like fold and is connected to a six-domain protruding tower that interacts with several outer CPs.

    PubMed  PubMed Central  Google Scholar 

  35. 35.

    Bayfield, O. W. et al. Cryo-EM structure and in vitro DNA packaging of a thermophilic virus with supersized T = 7 capsids. Proc. Natl. Acad. Sci. USA 116, 3556–3561 (2019).

    CAS  PubMed  Google Scholar 

  36. 36.

    Putri, R. M. et al. Structural characterization of native and modified encapsulins as nanoplatforms for in vitro catalysis and cellular uptake. ACS Nano 11, 12796–12804 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  37. 37.

    Hendrix, R. W. & Johnson, J. E. Bacteriophage HK97 capsid assembly and maturation. Adv. Exp. Med. Biol. 726, 351–363 (2012).

    CAS  PubMed  Google Scholar 

  38. 38.

    Prevelige, P. E. Jr. & Cortines, J. R. Phage assembly and the special role of the portal protein. Curr. Opin. Virol. 31, 66–73 (2018).

    CAS  PubMed  Google Scholar 

  39. 39.

    Lokareddy, R. K. et al. Portal protein functions akin to a DNA-sensor that couples genome-packaging to icosahedral capsid maturation. Nat. Commun. 8, 14310 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  40. 40.

    Liu, Y. T., Jih, J., Dai, X., Bi, G. Q. & Zhou, Z. H. Cryo-EM structures of herpes simplex virus type 1 portal vertex and packaged genome. Nature 570, 257–261 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  41. 41.

    Hong, C. et al. A structural model of the genome packaging process in a membrane-containing double stranded DNA virus. PLoS Biol. 12, e1002024 (2014).

    PubMed  PubMed Central  Google Scholar 

  42. 42.

    Smith, D. E. et al. The bacteriophage straight phi29 portal motor can package DNA against a large internal force. Nature 413, 748–752 (2001).

    CAS  PubMed  Google Scholar 

  43. 43.

    Chen, D. H. et al. Structural basis for scaffolding-mediated assembly and maturation of a dsDNA virus. Proc. Natl. Acad. Sci. USA 108, 1355–1360 (2011).

    CAS  PubMed  Google Scholar 

  44. 44.

    Guo, F. et al. Capsid expansion mechanism of bacteriophage T7 revealed by multistate atomic models derived from cryo-EM reconstructions. Proc. Natl. Acad. Sci. USA 111, E4606–E4614 (2014).

    CAS  PubMed  Google Scholar 

  45. 45.

    Li, S., Roy, P., Travesset, A. & Zandi, R. Why large icosahedral viruses need scaffolding proteins. Proc. Natl. Acad. Sci. USA 115, 10971–10976 (2018).

    CAS  PubMed  Google Scholar 

  46. 46.

    Borodavka, A., Desselberger, U. & Patton, J. T. Genome packaging in multi-segmented dsRNA viruses: distinct mechanisms with similar outcomes. Curr. Opin. Virol. 33, 106–112 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  47. 47.

    Sung, P. Y. & Roy, P. Sequential packaging of RNA genomic segments during the assembly of bluetongue virus. Nucleic Acids Res. 42, 13824–13838 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  48. 48.

    Lourenco, S. & Roy, P. In vitro reconstitution of bluetongue virus infectious cores. Proc. Natl. Acad. Sci. USA 108, 13746–13751 (2011).

    CAS  PubMed  Google Scholar 

  49. 49.

    Limn, C. K. & Roy, P. Intermolecular interactions in a two-layered viral capsid that requires a complex symmetry mismatch. J. Virol. 77, 11114–11124 (2003).

    CAS  PubMed  PubMed Central  Google Scholar 

  50. 50.

    Nakamichi, Y. et al. An assembly intermediate structure of rice dwarf virus reveals a hierarchical outer capsid shell assembly mechanism. Structure 27, 439–448.e3 (2019). Reoviruses have multilayered capsids. This study describes the structure of an intermediate assembly using phase-plate cryo-EM. The trimers of the secondary T = 13-layer initiate assembly on the three-fold axis of the T = 1 inner core and follows a hierarchical mechanism.

    CAS  PubMed  Google Scholar 

  51. 51.

    Nemecek, D. et al. Subunit folds and maturation pathway of a dsRNA virus capsid. Structure 21, 1374–1383 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  52. 52.

    Ilca, S. L. et al. Multiple liquid crystalline geometries of highly compacted nucleic acid in a dsRNA virus. Nature 570, 252–256 (2019). This paper details the arrangement of dsRNA in the phage ϕ6 in a study using cryo-EM. Most of the ϕ6 genome adopts a single-spooled genome organization similar to that of dsDNA viruses.

    CAS  PubMed  Google Scholar 

  53. 53.

    Mata, C. P. et al. Acquisition of functions on the outer capsid surface during evolution of double-stranded RNA fungal viruses. PLoS Pathog. 13, e1006755 (2017).

    PubMed  PubMed Central  Google Scholar 

  54. 54.

    Luque, D. et al. Cryo-EM near-atomic structure of a dsRNA fungal virus shows ancient structural motifs preserved in the dsRNA viral lineage. Proc. Natl. Acad. Sci. USA 111, 7641–7646 (2014).

    CAS  PubMed  Google Scholar 

  55. 55.

    Mata, C. P. et al. The RNA-binding protein of a double-stranded RNA virus acts like a scaffold protein. J. Virol. 92, e00968–18 (2018). The authors examine the assembly strategy of IBDV by combining cryo-EM, cryo-ET and atomic force microscopy analysis. IBDV is a dsRNA virus with a picornavirus-like fold CP that lacks the T = 1 inner core; instead, the multifunctional SP builds a transient, irregular shell beneath the single T = 13 capsid.

    PubMed  PubMed Central  Google Scholar 

  56. 56.

    Pascual, E. et al. Structural basis for the development of avian virus capsids that display influenza virus proteins and induce protective immunity. J. Virol. 89, 2563–2574 (2015).

    PubMed  Google Scholar 

  57. 57.

    Liu, H. et al. Atomic structure of human adenovirus by cryo-EM reveals interactions among protein networks. Science 329, 1038–1043 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  58. 58.

    Santos-Pérez, I. et al. Structural basis for assembly of vertical single β-barrel viruses. Nat. Commun. 10, 1184 (2019). This analysis reports the cryo-EM structure of two icosahedral archaeal viruses in which the two major CPs are based on a vertical single β-barrel. The capsid assembly relies on membrane proteins that interact with penton proteins and on membrane-proximal proteins that organize major CP heterodimers.

    PubMed  PubMed Central  Google Scholar 

  59. 59.

    De Colibus, L. et al. Assembly of complex viruses exemplified by a halophilic euryarchaeal virus. Nat. Commun. 10, 1456 (2019).

    PubMed  PubMed Central  Google Scholar 

  60. 60.

    Xiao, C. et al. Structural studies of the giant mimivirus. PLoS Biol. 7, e92 (2009).

    PubMed  Google Scholar 

  61. 61.

    Zhang, X. et al. Structure of Sputnik, a virophage, at 3.5-Å resolution. Proc. Natl. Acad. Sci. USA 109, 18431–18436 (2012).

    CAS  PubMed  Google Scholar 

  62. 62.

    Fang, Q. et al. Near-atomic structure of a giant virus. Nat. Commun. 10, 388 (2019).

    PubMed  PubMed Central  Google Scholar 

  63. 63.

    Born, D. et al. Capsid protein structure, self-assembly, and processing reveal morphogenesis of the marine virophage mavirus. Proc. Natl. Acad. Sci. USA 115, 7332–7337 (2018).

    CAS  PubMed  Google Scholar 

  64. 64.

    Condezo, G. N. & San Martín, C. Localization of adenovirus morphogenesis players, together with visualization of assembly intermediates and failed products, favor a model where assembly and packaging occur concurrently at the periphery of the replication center. PLoS Pathog. 13, e1006320 (2017).

    PubMed  PubMed Central  Google Scholar 

  65. 65.

    Zeev-Ben-Mordehai, T. et al. Two distinct trimeric conformations of natively membrane-anchored full-length herpes simplex virus 1 glycoprotein B. Proc. Natl. Acad. Sci. USA 113, 4176–4181 (2016).

    CAS  PubMed  Google Scholar 

  66. 66.

    Arranz, R. et al. The structure of native influenza virion ribonucleoproteins. Science 338, 1634–1637 (2012).

    CAS  PubMed  Google Scholar 

  67. 67.

    Qu, K. et al. Structure and architecture of immature and mature murine leukemia virus capsids. Proc. Natl. Acad. Sci. USA 115, E11751–E11760 (2018).

    CAS  PubMed  Google Scholar 

  68. 68.

    Mangala Prasad, V., Klose, T. & Rossmann, M. G. Assembly, maturation and three-dimensional helical structure of the teratogenic rubella virus. PLoS Pathog. 13, e1006377 (2017).

    PubMed  PubMed Central  Google Scholar 

  69. 69.

    Ke, Z. et al. Promotion of virus assembly and organization by the measles virus matrix protein. Nat. Commun. 9, 1736 (2018).

    PubMed  PubMed Central  Google Scholar 

  70. 70.

    Hochstein, R. et al. Structural studies of Acidianus tailed spindle virus reveal a structural paradigm used in the assembly of spindle-shaped viruses. Proc. Natl. Acad. Sci. USA 115, 2120–2125 (2018).

    CAS  PubMed  Google Scholar 

  71. 71.

    Hartman, R., Munson-McGee, J., Young, M. J. & Lawrence, C. M. Survey of high-resolution archaeal virus structures. Curr. Opin. Virol. 36, 74–83 (2019).

    CAS  PubMed  Google Scholar 

  72. 72.

    Wan, W. et al. Structure and assembly of the Ebola virus nucleocapsid. Nature 551, 394–397 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  73. 73.

    Hampton, C. M. et al. Correlated fluorescence microscopy and cryo-electron tomography of virus-infected or transfected mammalian cells. Nat. Protoc. 12, 150–167 (2017).

    CAS  PubMed  Google Scholar 

  74. 74.

    Chaikeeratisak, V. et al. Assembly of a nucleus-like structure during viral replication in bacteria. Science 355, 194–197 (2017). Based on a combined fluorescence microscopy and cryo-ET study, this paper reports a Pseudomonas phage that assembles a compartment for DNA replication. Before cell lysis to release phages, assembled empty capsids migrate to the compartment surface for DNA packaging.

    CAS  PubMed  PubMed Central  Google Scholar 

  75. 75.

    Dai, W. et al. Visualizing virus assembly intermediates inside marine cyanobacteria. Nature 502, 707–710 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  76. 76.

    Wen, A. M. & Steinmetz, N. F. Design of virus-based nanomaterials for medicine, biotechnology, and energy. Chem. Soc. Rev. 45, 4074–4126 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  77. 77.

    Janitzek, C. M. et al. A proof-of-concept study for the design of a VLP-based combinatorial HPV and placental malaria vaccine. Sci. Rep. 9, 5260 (2019).

    PubMed  PubMed Central  Google Scholar 

  78. 78.

    Hryc, C. F. et al. Accurate model annotation of a near-atomic resolution cryo-EM map. Proc. Natl. Acad. Sci. USA 114, 3103–3108 (2017).

    CAS  PubMed  Google Scholar 

  79. 79.

    Speir, J. A., Munshi, S., Wang, G., Baker, T. S. & Johnson, J. E. Structures of the native and swollen forms of cowpea chlorotic mottle virus determined by X-ray crystallography and cryo-electron microscopy. Structure 3, 63–78 (1995).

    CAS  PubMed  PubMed Central  Google Scholar 

  80. 80.

    Llauró, A. et al. Cargo-shell and cargo-cargo couplings govern the mechanics of artificially loaded virus-derived cages. Nanoscale 8, 9328–9336 (2016).

    PubMed  PubMed Central  Google Scholar 

  81. 81.

    McCoy, K. et al. Cargo retention inside P22 virus-like particles. Biomacromolecules 19, 3738–3746 (2018). Using cryo-EM combined with biophysical analysis, the authors define the factors essential for the ability of P22 VLPs to retain or release various protein cargos. In addition to the relationship between cargo size and capsid pore size, electrostatic interactions are also important.

    CAS  PubMed  Google Scholar 

  82. 82.

    Brasch, M. et al. Assembling enzymatic cascade pathways inside virus-based nanocages using dual-tasking nucleic acid tags. J. Am. Chem. Soc. 139, 1512–1519 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  83. 83.

    Hu, Y., Zandi, R., Anavitarte, A., Knobler, C. M. & Gelbart, W. M. Packaging of a polymer by a viral capsid: the interplay between polymer length and capsid size. Biophys. J. 94, 1428–1436 (2008).

    CAS  PubMed  Google Scholar 

  84. 84.

    Patterson, D. P., Prevelige, P. E. & Douglas, T. Nanoreactors by programmed enzyme encapsulation inside the capsid of the bacteriophage P22. ACS Nano 6, 5000–5009 (2012).

    CAS  PubMed  Google Scholar 

  85. 85.

    Luque, D. et al. Self-assembly and characterization of small and monodisperse dye nanospheres in a protein cage. Chem. Sci. 5, 575–581 (2014).

    CAS  Google Scholar 

  86. 86.

    Bale, J. B. et al. Accurate design of megadalton-scale two-component icosahedral protein complexes. Science 353, 389–394 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  87. 87.

    Terasaka, N., Azuma, Y. & Hilvert, D. Laboratory evolution of virus-like nucleocapsids from nonviral protein cages. Proc. Natl. Acad. Sci. USA 115, 5432–5437 (2018).

    CAS  PubMed  Google Scholar 

  88. 88.

    Butterfield, G. L. et al. Evolution of a designed protein assembly encapsulating its own RNA genome. Nature 552, 415–420 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  89. 89.

    Heddle, J. G., Chakraborti, S. & Iwasaki, K. Natural and artificial protein cages: design, structure and therapeutic applications. Curr. Opin. Struct. Biol. 43, 148–155 (2017).

    CAS  PubMed  Google Scholar 

  90. 90.

    Moser, F. et al. Cryo-SOFI enabling low-dose super-resolution correlative light and electron cryo-microscopy. Proc. Natl. Acad. Sci. USA 116, 4804–4809 (2019).

    CAS  PubMed  Google Scholar 

  91. 91.

    Bertozzi, C. Atoms out of blobs: cryoEM takes the Nobel prize in chemistry. ACS Cent. Sci. 3, 1056 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  92. 92.

    Sirohi, D. et al. The 3.8 Å resolution cryo-EM structure of Zika virus. Science 352, 467–470 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  93. 93.

    Henderson, R. Image contrast in high-resolution electron microscopy of biological macromolecules: TMV in ice. Ultramicroscopy 46, 1–18 (1992).

    CAS  PubMed  Google Scholar 

  94. 94.

    Bammes, B. E., Jakana, J., Schmid, M. F. & Chiu, W. Radiation damage effects at four specimen temperatures from 4 to 100 K. J. Struct. Biol. 169, 331–341 (2010).

    CAS  PubMed  Google Scholar 

  95. 95.

    Brilot, A. F. et al. Beam-induced motion of vitrified specimen on holey carbon film. J. Struct. Biol. 177, 630–637 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  96. 96.

    Scheres, S. H. Beam-induced motion correction for sub-megadalton cryo-EM particles. eLife 3, e03665 (2014).

    PubMed  PubMed Central  Google Scholar 

  97. 97.

    Li, X. et al. Electron counting and beam-induced motion correction enable near-atomic-resolution single-particle cryo-EM. Nat. Methods 10, 584–590 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

We apologize to our colleagues with outstanding contributions who were not mentioned due to space limitations. The authors thank J.L. Carrascosa, C. San Martín and J.M. Rodríguez for critical reading of the manuscript, and C. Mark for editorial assistance. This work was supported by grants from the Spanish Ministry of Economy and Competitivity (BFU2017-88736-R) and the Comunidad Autónoma de Madrid (P2018/NMT-4389) to J.R.C.

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Luque, D., Castón, J.R. Cryo-electron microscopy for the study of virus assembly. Nat Chem Biol 16, 231–239 (2020). https://doi.org/10.1038/s41589-020-0477-1

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