Encapsulation of Nanoparticles in Virus Protein Shells

  • Irina B. Tsvetkova
  • Bogdan G. DragneaEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1252)


The self-assembly of virus-like particles may lead to materials which combine the unique characteristics of viruses, such as precise size control and responsivity to environmental cues, with the properties of abiotic cargo. For a few different viruses, shell proteins are amenable to the in vitro encapsulation of non-genomic cargo in a regular protein cage. In this chapter we describe protocols of high-efficiency in vitro self-assembly around functionalized gold nanoparticles for three examples of icosahedral and non-icosahedral viral protein cages derived from a plant virus, an animal virus, and a human retrovirus. These protocols can be readily adapted with small modifications to work for a broad variety of inorganic and organic nanoparticles.

Key words

Virus-like particle Gold nanoparticle Templated self-assembly Protein cage 



This work was supported by the US Department of Energy, Office of Science, Basic Energy Sciences, under Award # DE-SC0010507.


  1. 1.
    Douglas T, Young M (2006) Viruses: making friends with old foes. Science 312:873–875PubMedCrossRefGoogle Scholar
  2. 2.
    Steinmetz NF, Manchester M (2011) Viral nanoparticles: tools for materials science and biomedicine. Pan Stanford Publishing, SingaporeGoogle Scholar
  3. 3.
    Plummer EM, Manchester M (2011) Viral nanoparticles and virus-like particles: platforms for contemporary vaccine design. WIREs Nanomed Nanobiotechnol 3:174–196CrossRefGoogle Scholar
  4. 4.
    Lee LA, Wang Q (2006) Adaptations of nanoscale viruses and other protein cages for medical applications. Nanomedicine 2:137–149PubMedCrossRefGoogle Scholar
  5. 5.
    Fiedler JD, Brown SD, Lau JL et al (2010) RNA-directed packaging of enzymes within virus-like particles. Angew Chem Int Ed 49:9648–9651CrossRefGoogle Scholar
  6. 6.
    Comellas-Aragones M, Engelkamp H, Claessen VI et al (2007) A virus-based single-enzyme nanoreactor. Nat Nanotechnol 2:635–639PubMedCrossRefGoogle Scholar
  7. 7.
    Patterson DP, Schwarz B, El-Boubbou K et al (2012) Virus-like particle nanoreactors: programmed encapsulation of the thermostable CelB glycosidase inside the P22 capsid. Soft Matter 8:10158–10166CrossRefGoogle Scholar
  8. 8.
    Stephanopoulos N, Carrico ZM, Francis MB (2009) Nanoscale integration of sensitizing chromophores and porphyrins with bacteriophage MS2. Angew Chem Int Ed 48:9498–9502CrossRefGoogle Scholar
  9. 9.
    Nam YS, Magyar AP, Lee D et al (2010) Biologically templated photocatalytic nanostructures for sustained light-driven water oxidation. Nat Nanotechnol 5:340–344PubMedCrossRefGoogle Scholar
  10. 10.
    Nam KT, Kim D-W, Yoo PJ et al (2006) Virus-enabled synthesis and assembly of nanowires for lithium ion battery electrodes. Science 312:885–888PubMedCrossRefGoogle Scholar
  11. 11.
    Niu Z, Liu J, Lee LA et al (2007) Biological templated synthesis of water-soluble conductive polymeric nanowires. Nano Lett 7:3729–3733PubMedCrossRefGoogle Scholar
  12. 12.
    DuFort CC, Dragnea B (2010) Bio-enabled synthesis of metamaterials. Annu Rev Phys Chem 61:323–344PubMedCrossRefGoogle Scholar
  13. 13.
    Kostiainen MA, Hiekkataipale P, Laiho A et al (2013) Electrostatic assembly of binary nanoparticle superlattices using protein cages. Nat Nanotechnol 8:52PubMedCrossRefGoogle Scholar
  14. 14.
    Carette N, Engelkamp H, Akpa E et al (2007) A virus-based biocatalyst. Nat Nanotechnol 2:226–229PubMedCrossRefGoogle Scholar
  15. 15.
    Douglas T, Young M (1998) Host-guest encapsulation of materials by assembled virus protein cages. Nature 393:152–155CrossRefGoogle Scholar
  16. 16.
    Jung B, Rao ALN, Anvari B (2011) Optical nano-constructs composed of genome-depleted brome mosaic virus doped with a near infrared chromophore for potential biomedical applications. ACS Nano 5:1243–1252PubMedCrossRefGoogle Scholar
  17. 17.
    Tsvetkova I, Chen C, Rana S et al (2012) Pathway switching in templated virus-like particle assembly. Soft Matter 8:4571–4577CrossRefGoogle Scholar
  18. 18.
    Cadena-Nava RD, Hu YF, Garmann RF et al (2011) Exploiting fluorescent polymers to probe the self-assembly of virus-like particles. J Phys Chem B 115:2386–2391PubMedCrossRefGoogle Scholar
  19. 19.
    Hu Y, Zandi R, Anavitarte A et al (2008) Packaging of a polymer by a viral capsid: the interplay between polymer length and capsid size. Biophys J 94:1428–1436PubMedCrossRefPubMedCentralGoogle Scholar
  20. 20.
    Porterfield JZ, Dhason MS, Loeb DD et al (2010) Full-length hepatitis B virus core protein packages viral and heterologous RNA with similarly high levels of cooperativity. J Virol 84:7174–7184PubMedCrossRefPubMedCentralGoogle Scholar
  21. 21.
    Daniel M-C, Tsvetkova IB, Quinkert ZT et al (2010) Role of surface charge density in nanoparticle-templated assembly of bromovirus protein cages. ACS Nano 4:3853–3860PubMedCrossRefPubMedCentralGoogle Scholar
  22. 22.
    Mout R, Moyano DF, Rana S et al (2012) Surface functionalization of nanoparticles for nanomedicine. Chem Soc Rev 41:2539–2544PubMedCrossRefPubMedCentralGoogle Scholar
  23. 23.
    Thomas M, Klibanov AM (2003) Conjugation to gold nanoparticles enhances polyethylenimine’s transfer of plasmid DNA into mammalian cells. Proc Natl Acad Sci 100:9138–9143PubMedCrossRefPubMedCentralGoogle Scholar
  24. 24.
    Ghosh P, Han G, De M et al (2008) Gold nanoparticles in delivery applications. Adv Drug Deliv Rev 60:1307–1315PubMedCrossRefGoogle Scholar
  25. 25.
    Cognet L, Tardin C, Boyer D et al (2003) Single metallic nanoparticle imaging for protein detection in cells. Proc Natl Acad Sci 100:11350–11355PubMedCrossRefPubMedCentralGoogle Scholar
  26. 26.
    Berciaud S, Cognet L, Blab GA et al (2004) Photothermal heterodyne imaging of individual nonfluorescent nanoclusters and nanocrystals. Phys Rev Lett 93:257402PubMedCrossRefGoogle Scholar
  27. 27.
    Haiss W, Thanh NTK, Aveyard J et al (2007) Determination of size and concentration of gold nanoparticles from UV−Vis spectra. Anal Chem 79:4215–4221PubMedCrossRefGoogle Scholar
  28. 28.
    Boyer D, Tamarat P, Maali A et al (2002) Photothermal imaging of nanometer-sized metal particles among scatterers. Science 297:1160–1163PubMedCrossRefGoogle Scholar
  29. 29.
    Capehart SL, Coyle MP, Glasgow JE et al (2013) Controlled integration of gold nanoparticles and organic fluorophores using synthetically modified MS2 viral capsids. J Am Chem Soc 135:3011–3016PubMedCrossRefPubMedCentralGoogle Scholar
  30. 30.
    Blum AS, Soto CM, Wilson CD et al (2004) Cowpea mosaic virus as a scaffold for 3-D patterning of gold nanoparticles. Nano Lett 4:867–870CrossRefGoogle Scholar
  31. 31.
    Slocik JM, Naik RR, Stone MO et al (2005) Viral templates for gold nanoparticle synthesis. J Mater Chem 15:749–753CrossRefGoogle Scholar
  32. 32.
    Radloff C, Vaia RA, Brunton J et al (2005) Metal nanoshell assembly on a virus bioscaffold. Nano Lett 5:1187–1191PubMedCrossRefGoogle Scholar
  33. 33.
    Chen C, Daniel MC, Quinkert ZT et al (2006) Nanoparticle-templated assembly of viral protein cages. Nano Lett 6:611–615PubMedCrossRefGoogle Scholar
  34. 34.
    Loo L, Guenther RH, Basnayake VR et al (2006) Controlled encapsidation of gold nanoparticles by a viral protein shell. J Am Chem Soc 128:4502–4503PubMedCrossRefGoogle Scholar
  35. 35.
    Wang TJ, Zhang ZP, Gao D et al (2011) Encapsulation of gold nanoparticles by simian virus 40 capsids. Nanoscale 3:4275–4282PubMedCrossRefGoogle Scholar
  36. 36.
    Aniagyei SE, Kennedy CJ, Stein B et al (2009) Synergistic effects of mutations and nanoparticle templating in the self-assembly of cowpea chlorotic mottle virus capsids. Nano Lett 9:393–398PubMedCrossRefPubMedCentralGoogle Scholar
  37. 37.
    Bancroft JB, Hiebert E, Bracker CE (1969) The effects of various polyanions on shell formation of some spherical viruses. Virology 39:924–930PubMedCrossRefGoogle Scholar
  38. 38.
    Goicochea NL, De M, Rotello VM et al (2007) Core-like particles of an enveloped animal virus can self-assemble efficiently on artificial templates. Nano Lett 7:2281–2290PubMedCrossRefGoogle Scholar
  39. 39.
    Goicochea NL, Datta SAK, Ayaluru M et al (2011) Structure and stoichiometry of template-directed recombinant HIV-1 Gag particles. J Mol Biol 410:667–680PubMedCrossRefPubMedCentralGoogle Scholar
  40. 40.
    Hiebert E, Bancroft JB, Bracker CE (1968) The assembly in vitro of some small spherical viruses, hybrid viruses, and other nucleoproteins. Virology 34:492–508PubMedCrossRefGoogle Scholar
  41. 41.
    Caspar D, Klug A (1962) Physical principles in the construction of regular viruses. Cold Spring Harb Symp Quant Biol 27:1–24PubMedCrossRefGoogle Scholar
  42. 42.
    Lucas R, Larson S, McPherson A (2002) The crystallographic structure of brome mosaic virus. J Mol Biol 317:95–108PubMedCrossRefGoogle Scholar
  43. 43.
    Cuillel M, Berthetcolominas C, Timmins PA et al (1987) Reassembly of Brome Mosaic-virus from dissociated virus - a neutron-scattering study. Eur Biophys J 15:169–176CrossRefGoogle Scholar
  44. 44.
    Lavelle L, Gingery M, Phillips M et al (2009) Phase diagram of self-assembled viral capsid protein polymorphs. J Phys Chem B 113:3813–3819PubMedCrossRefGoogle Scholar
  45. 45.
    Bahadur RP, Rodier F, Janin J (2007) A dissection of the protein–protein interfaces in icosahedral virus capsids. J Mol Biol 367:574–590PubMedCrossRefGoogle Scholar
  46. 46.
    Rao ALN (2006) Genome packaging by spherical plant RNA viruses. Annu Rev Phytopathol 44:61–87PubMedCrossRefGoogle Scholar
  47. 47.
    Choi YG, Rao ALN (2003) Packaging of brome mosaic virus RNA3 is mediated through a bipartite signal. J Virol 77:9750–9757PubMedCrossRefPubMedCentralGoogle Scholar
  48. 48.
    Ni P, Wang Z, Ma X et al (2012) An examination of the electrostatic interactions between the N-terminal tail of the brome mosaic virus coat protein and encapsidated RNAs. J Mol Biol 419:284–300PubMedCrossRefPubMedCentralGoogle Scholar
  49. 49.
    Sun J, DuFort C, Daniel M-C et al (2007) Core-controlled polymorphism in virus-like particles. Proc Natl Acad Sci 104:1354–1359PubMedCrossRefPubMedCentralGoogle Scholar
  50. 50.
    Chen C, Kwak ES, Stein B et al (2005) Packaging of gold particles in viral capsids. J Nanosci Nanotechnol 5:2029–2033PubMedCrossRefGoogle Scholar
  51. 51.
    Strauss JH, Strauss EG (1994) The alphaviruses: gene expression, replication, and evolution. Microbiol Rev 58:491–562PubMedPubMedCentralGoogle Scholar
  52. 52.
    Cheng RH, Kuhn RJ, Olson NH et al (1995) Nucleocapsid and glycoprotein organization in an enveloped virus. Cell 80:621–630PubMedCrossRefPubMedCentralGoogle Scholar
  53. 53.
    Lopez S, Yao JS, Kuhn RJ et al (1994) Nucleocapsid-glycoprotein interactions required for assembly of alphaviruses. J Virol 68:1316–1323PubMedPubMedCentralGoogle Scholar
  54. 54.
    Goicochea NL (2010) Nanoparticle-directed assembly of enveloped virus components and applications. Ph.D. Thesis, Indiana UniversityGoogle Scholar
  55. 55.
    Ganser-Pornillos BK, Yeager M, Sundquist WI (2008) The structural biology of HIV assembly. Curr Opin Struct Biol 18:203–217PubMedCrossRefPubMedCentralGoogle Scholar
  56. 56.
    Briggs JAG, Riches JD, Glass B et al (2009) Structure and assembly of immature HIV. Proc Natl Acad Sci 106:11090–11095PubMedCrossRefPubMedCentralGoogle Scholar
  57. 57.
    Wright ER, Schooler JB, Ding HJ et al (2007) Electron cryotomography of immature HIV-1 virions reveals the structure of the CA and SP1 Gag shells. EMBO J 26:2218–2226PubMedCrossRefPubMedCentralGoogle Scholar
  58. 58.
    Briggs JAG, Kräusslich H-G (2011) The molecular architecture of HIV. J Mol Biol 410:491–500PubMedCrossRefGoogle Scholar
  59. 59.
    Datta SK, Rein A (2009) Preparation of recombinant HIV-1 Gag protein and assembly of virus-like particles in vitro. In: Prasad V, Kalpana G (eds) HIV protocols, vol 485. Humana Press, New York, pp 197–208CrossRefGoogle Scholar
  60. 60.
    Wilk T, Gross I, Gowen BE et al (2001) Organization of immature human immunodeficiency virus type 1. J Virol 75:759–771PubMedCrossRefPubMedCentralGoogle Scholar
  61. 61.
    Slot JW, Geuze HJ (1985) A new method of preparing gold probes for multiple-labeling cytochemistry. Eur J Cell Biol 38:87–93PubMedGoogle Scholar
  62. 62.
    Frens G (1973) Controlled nucleation for the regulation of the particle size in monodisperse gold suspensions. Nature Phys Sci 241:20–22CrossRefGoogle Scholar
  63. 63.
    Hurst SJ, Lytton-Jean AKR, Mirkin CA (2006) Maximizing DNA loading on a range of gold nanoparticle sizes. Anal Chem 78:8313–8318PubMedCrossRefPubMedCentralGoogle Scholar
  64. 64.
    Gopinath K, Kao CC (2007) Replication-independent long-distance trafficking by viral RNAs in Nicotiana benthamiana. Plant Cell 19:1179–1191PubMedCrossRefPubMedCentralGoogle Scholar
  65. 65.
    Cuillel M, Zulauf M, Jacrot B (1983) Self-assembly of brome mosaic virus protein into capsids: initial and final states of aggregation. J Mol Biol 164:589–603PubMedCrossRefGoogle Scholar
  66. 66.
    Yamazaki H, Kaesberg P (1963) Degradation of bromegrass mosaic virus with calcium chloride and isolation of its protein and nucleic acid. J Mol Biol 7:760–762PubMedCrossRefGoogle Scholar
  67. 67.
    Mukhopadhyay S, Chipman PR, Hong EM et al (2002) In vitro-assembled alphavirus core-like particles maintain a structure similar to that of nucleocapsid cores in mature virus. J Virol 76:11128–11132PubMedCrossRefPubMedCentralGoogle Scholar
  68. 68.
    Garmann RF, Comas-Garcia M, Gopal A et al (2014) The assembly pathway of an icosahedral single-stranded RNA virus depends on the strength of inter-subunit attractions. J Mol Biol 426:1050–1060PubMedCrossRefGoogle Scholar
  69. 69.
    Datta SAK, Curtis JE, Ratcliff W et al (2007) Conformation of the HIV-1 Gag protein in solution. J Mol Biol 365:812–824PubMedCrossRefPubMedCentralGoogle Scholar
  70. 70.
    Ludtke S, Baldwin P, Chiu W (1999) EMAN: semiautomated software for high-resolution single-particle reconstructions. J Struct Biol 128:82–97PubMedCrossRefGoogle Scholar
  71. 71.
    Pettersen EF, Goddard TD, Huang CC et al (2004) UCSF Chimera–a visualization system for exploratory research and analysis. J Comput Chem 25:1605–1612PubMedCrossRefGoogle Scholar
  72. 72.
    Chen S, Kimura K (1999) Synthesis and characterization of carboxylate-modified gold nanoparticle powders dispersible in water. Langmuir 15:1075–1082CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  1. 1.Department of ChemistryIndiana UniversityBloomingtonUSA

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