Overexpression and self-assembly of virus-like particles in Nicotiana benthamiana by a single-vector DNA replicon system

Ki Beom Moon, Jisu Lee, Sebyung Kang, Moonil Kim, Hugh Mason, Jae Heung Jeon, Hyun Soon Kim

Research output: Contribution to journalArticle

13 Scopus citations

Abstract

Based on recent developments, virus-like particles (VLPs) are considered to be perfect candidates as nanoplatforms for applications in materials science and medicine. To succeed, mass production of VLPs and self-assembly into a correct form in plant systems are key factors. Here, we report expression of synthesized coat proteins of the three viruses, Brome mosaic virus, Cucumber mosaic virus, and Maize rayado fino virus, in Nicotiana benthamiana and production of self-assembled VLPs by transient expression system using agroinfiltration. Each coat protein was synthesized and cloned into a pBYR2fp single replicon vector. Target protein expression in cells containing p19 was fourfold higher than that of cells lacking p19. After agroinfiltration, protein expression was analyzed by SDS-PAGE and quantitative image analyzer. Quantitative analysis showed that BMVCP, CMVCP, and MRFVCP concentrations were 0.5, 1.0, and 0.8 mg · g−1 leaf fresh weight, respectively. VLPs were purified by sucrose cushion ultracentrifugation and then analyzed by transmission electron microscopy. Our results suggested that BMVCP and CMVCP proteins expressed in N. benthamiana leaves were able to correctly self-assemble into particles. Moreover, we evaluated internal cavity accessibility of VLPs to load foreign molecules. Finally, plant growth conditions after agroinfiltration are critical for increasing heterologous protein expression levels in a transient expression system.

Original languageEnglish (US)
Pages (from-to)8281-8290
Number of pages10
JournalApplied Microbiology and Biotechnology
Volume98
Issue number19
DOIs
StatePublished - Jan 1 2014

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Keywords

  • Bionanomaterial
  • Plant virus
  • Self-assembly
  • Virus-like particle
  • pBYR2fp viral vector

ASJC Scopus subject areas

  • Biotechnology
  • Applied Microbiology and Biotechnology

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