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Genetic Engineering and Characterization of Cowpea Mosaic Virus Empty Virus-Like Particles

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Virus Hybrids as Nanomaterials

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1108))

Abstract

The development of methods for the production of empty Cowpea mosaic virus (CPMV) virus-like particles (VLPs) that are devoid of RNA, eVLPs, has renewed promise in CPMV capsid technologies. The recombinant nature of CPMV eVLP production means that the extent and variety of genetic modifications that may be incorporated into the particles is theoretically much greater than those that can be made to infectious CPMV virions due to restrictions on viral propagation of the latter. Free of the infectious agent, the genomic RNA, these particles are now finding potential uses in vaccine development, in vivo imaging, drug delivery, and other nanotechnology applications that make use of internal loading of the empty particles. Here we describe methods for the genetic modification and production of CPMV eVLPs and describe techniques useful for their characterization.

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References

  1. Porta C, Spall VE, Loveland J, Johnson JE, Barker PJ, Lomonossoff GP (1994) Development of cowpea mosaic virus as a high-yielding system for the presentation of foreign peptides. Virology 202:949–955

    Article  CAS  Google Scholar 

  2. Leong HS, Steinmetz NF, Ablack A, Destito G, Zijlstra A, Stuhlmann H, Manchester M, Lewis JD (2010) Intravital imaging of embryonic and tumor neovasculature using viral nanoparticles. Nat Protoc 5:1406–1417

    Article  CAS  Google Scholar 

  3. Steinmetz NF, Lin T, Lomonossoff GP, Johnson JE (2009) Structure-based engineering of an icosahedral virus for nanomedicine and nanotechnology. Curr Top Microbiol Immunol 327:23–58

    Article  CAS  Google Scholar 

  4. Wang Q, Kaltgrad E, Lin TW, Johnson JE, Finn MG (2002) Natural supramolecular building blocks: wild-type cowpea mosaic virus. Chem Biol 9:805–811

    Article  CAS  Google Scholar 

  5. Steinmetz NF, Lomonossoff GP, Evans DJ (2006) Cowpea mosaic virus for material fabrication: addressable carboxylate groups on a programmable nanoscaffold. Langmuir 22: 3488–3490

    Article  CAS  Google Scholar 

  6. Aljabali AAA, Shah SN, Evans-Gowing R, Lomonossoff GP, Evans DJ (2011) Chemically-coupled-peptide-promoted virus nanoparticle templated mineralization. Integr Biol 3:119–125

    Article  CAS  Google Scholar 

  7. Aljabali AAA, Barclay JE, Steinmetz NF, Lomonossoff GP, Evans DJ (2012) Controlled immobilisation of active enzymes on the cowpea mosaic virus capsid. Nanoscale 4: 5640–5645

    Article  CAS  Google Scholar 

  8. Aljabali AAA, Barclay JE, Lomonossoff GP, Evans DJ (2010) Virus templated metallic nanoparticles. Nanoscale 2:2596–2600

    Article  CAS  Google Scholar 

  9. Lin TW, Chen ZG, Usha R, Stauffacher CV, Dai JB, Schmidt T, Johnson JE (1999) The refined crystal structure of cowpea mosaic virus at 2.8 A resolution. Virology 265:20–34

    Article  CAS  Google Scholar 

  10. Lomonossoff GP, Johnson JE (1991) The synthesis and structure of comovirus capsids. Prog Biophys Mol Biol 55:107–137

    Article  CAS  Google Scholar 

  11. Shah SN, Steinmetz NF, Aljabali AA, Lomonossoff GP, Evans DJ (2009) Environmentally benign synthesis of virus-templated, monodisperse, iron-platinum nanoparticles. Dalton Trans 8479–8480

    Google Scholar 

  12. Blum AS, Soto CM, Wilson CD, Brower TL, Pollack SK, Schull TL, Chatterji A, Lin T, Johnson JE, Amsinck C, Franzon P, Shashidhar R, Ratna BR (2005) An engineered virus as a scaffold for three-dimensional self-assembly on the nanoscale. Small 1:702–706

    Article  CAS  Google Scholar 

  13. Sainsbury F, Canizares MC, Lomonossoff GP (2010) Cowpea mosaic virus: the plant virus-based biotechnology workhorse. Annu Rev Phytopathol 48:437–455

    Article  CAS  Google Scholar 

  14. Porta C, Spall VE, Findlay KC, Gergerich RC, Farrance CE, Lomonossoff GP (2003) Cowpea mosaic virus-based chimaeras: effects of inserted peptides on the phenotype, host range, and transmissibility of the modified viruses. Virology 310:50–63

    Article  CAS  Google Scholar 

  15. Saunders K, Sainsbury F, Lomonossoff GP (2009) Efficient generation of cowpea mosaic virus empty virus-like particles by the proteolytic processing of precursors in insect cells and plants. Virology 393:329–337

    Article  CAS  Google Scholar 

  16. Aljabali AAA, Sainsbury F, Lomonossoff GP, Evans DJ (2010) Cowpea mosaic virus unmodified empty viruslike particles loaded with metal and metal oxide. Small 6:818–821

    Article  CAS  Google Scholar 

  17. Sainsbury F, Saunders K, Aljabali AAA, Evans DJ, Lomonossoff GP (2011) Peptide-controlled access to the interior surface of empty virus nanoparticles. Chembiochem 12: 2435–2440

    Article  CAS  Google Scholar 

  18. Wen AM, Shukla S, Saxena P, Aljabali AAA, Yildiz I, Dey S, Mealy JE, Yang AC, Evans DJ, Lomonossoff GP, Steinmetz NF (2012) Interior engineering of a viral nanoparticle and its tumor homing properties. Biomacromolecules 13: 3990–4001

    Article  CAS  Google Scholar 

  19. Sainsbury F, Lomonossoff GP (2008) Extremely high-level and rapid transient protein production in plants without the use of viral replication. Plant Physiol 148:1212–1218

    Article  CAS  Google Scholar 

  20. Sainsbury F, Thuenemann EC, Lomonossoff GP (2009) pEAQ: versatile expression vectors for easy and quick transient expression of heterologous proteins in plants. Plant Biotechnol J 7:682–693

    Article  CAS  Google Scholar 

  21. Montague NP, Thuenemann EC, Saxena P, Saunders K, Lenzi P, Lomonossoff GP (2011) Recent advances of Cowpea mosaic virus-based particle technology. Hum Vaccin 7: 383–390

    Article  CAS  Google Scholar 

  22. Gelvin SB (2003) Agrobacterium-mediated plant transformation: the biology behind the "gene-jockeying" tool. Microbiol Mol Biol Rev 67:16–37

    Article  CAS  Google Scholar 

  23. Johnson J, Lin T, Lomonossoff GP (1997) Presentation of heterologous peptides on plant viruses: genetics, structure, and function. Annu Rev Phytopathol 35:67–86

    Article  CAS  Google Scholar 

  24. Mattanovich D, Rüker F, da Câmara Machado A, Laimer M, Regner F, Stein-kellner H, Himmler G, Katinger H (1989) Efficient transformation of Agro-bacterium spp. by electroporation. Nucleic Acids Res 17:6747

    Article  CAS  Google Scholar 

  25. Saunders K, Sainsbury F, Lomonossoff GP (2010) Production of viral capsids. UK Patent application No. GB2010/001183, PCT Publication No. WO2010/146359, 23 Dec 2010

    Google Scholar 

  26. van Kammen A (1967) Purification and properties of the components of cowpea mosaic virus. Virology 31:633–642

    Article  Google Scholar 

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Acknowledgments

This work was supported by grant BB/J004561/1 and a Doctoral Training Grant from the Biotechnology and Biological Research Council (BBSRC) UK and the John Innes Foundation.

Author information

Authors and Affiliations

  1. Australian Institute for Bioengineering and Nanotechnology, University of Queensland, St-Lucia, QLD, Australia

    Frank Sainsbury

  2. Department of Biological Chemistry, John Innes Centre, Norwich, UK

    Pooja Saxena, Keith Saunders & George P. Lomonossoff

  3. Department of Chemistry, University of Hull, Hull, UK

    Alaa A. A. Aljabali & David J. Evans

Authors
  1. Frank Sainsbury
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  2. Pooja Saxena
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  3. Alaa A. A. Aljabali
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  4. Keith Saunders
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  5. David J. Evans
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  6. George P. Lomonossoff
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Editor information

Editors and Affiliations

  1. Bio/Molecular Science and Engineering, US Naval Research Laboratory, Washington, District of Columbia, USA

    Baochuan Lin

  2. Bio/Molecular Science and Engineering, US Naval Research Laboratory, Washington, District of Columbia, USA

    Banahalli Ratna

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Sainsbury, F., Saxena, P., Aljabali, A.A.A., Saunders, K., Evans, D.J., Lomonossoff, G.P. (2014). Genetic Engineering and Characterization of Cowpea Mosaic Virus Empty Virus-Like Particles. In: Lin, B., Ratna, B. (eds) Virus Hybrids as Nanomaterials. Methods in Molecular Biology, vol 1108. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-751-8_11

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  • DOI: https://doi.org/10.1007/978-1-62703-751-8_11

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-750-1

  • Online ISBN: 978-1-62703-751-8

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