High-level expression and enrichment of norovirus virus-like particles in plants using modified geminiviral vectors

Andrew G. Diamos, Hugh Mason

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

Recombinant virus-like particles (VLPs) are proven to be safe and effective vaccine candidates. We have previously described a plant-based recombinant protein expression system based on agroinfiltration of a replicating vector derived from the geminivirus bean yellow dwarf virus (BeYDV). The system has been systematically optimized to improve expression and reduce cell death in Nicotiana benthamiana leaves. Using these modifications, we show that VLPs derived from genotype GII.4 norovirus, the leading cause of acute gastroenteritis worldwide, can be produced at >1 mg/g leaf fresh weight (LFW), over three times the highest level ever reported in plant-based systems. We also produced norovirus GI VLPs at 2.3 mg/g LFW. Treatment of VLP-containing crude leaf extracts with acid, detergent, or heat enhanced recovery and allowed selective enrichment of norovirus VLPs. Optimal treatment conditions allowed removal of >90% of endogenous plant proteins without any loss of norovirus VLPs. Selective enrichment of hepatitis B core antigen (HBcAg) VLPs by acid treatment was also demonstrated, with some losses in yield that were partially mitigated in the presence of detergent. Sedimentation analysis confirmed that acid and detergent did not inhibit proper assembly of norovirus VLPs, although heat treatment had a small negative effect. These results demonstrate that milligram quantities of norovirus VLPs can be obtained and highly enriched in a matter of days from a single plant leaf using the BeYDV plant expression system.

Original languageEnglish (US)
Pages (from-to)86-92
Number of pages7
JournalProtein Expression and Purification
Volume151
DOIs
StatePublished - Nov 1 2018

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Norovirus
Virion
Detergents
Acids
Hot Temperature
Geminiviridae
Hepatitis B Core Antigens
Plant Viruses
Weights and Measures
Plant Proteins
Plant Leaves
Gastroenteritis
Complex Mixtures
Recombinant Proteins
Tobacco
Cell Death
Vaccines
Genotype
Viruses

ASJC Scopus subject areas

  • Biotechnology

Cite this

High-level expression and enrichment of norovirus virus-like particles in plants using modified geminiviral vectors. / Diamos, Andrew G.; Mason, Hugh.

In: Protein Expression and Purification, Vol. 151, 01.11.2018, p. 86-92.

Research output: Contribution to journalArticle

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