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Scalable Production of HPV16 L1 Protein and VLPs from Tobacco Leaves.

Zahin, Maryam and Joh, Joongho and Khanal, Sujita and Husk, Adam and Mason, Hugh and Warzecha, Heribert and Ghim, Shin-Je and Miller, Donald M. and Matoba, Nobuyuki and Jenson, Alfred Bennett (2016):
Scalable Production of HPV16 L1 Protein and VLPs from Tobacco Leaves.
11, In: PloS one, (8), pp. e0160995. ISSN 1932-6203,
[Article]

Abstract

Cervical cancer is the most common malignancy among women particularly in developing countries, with human papillomavirus (HPV) 16 causing 50% of invasive cervical cancers. A plant-based HPV vaccine is an alternative to the currently available virus-like particle (VLP) vaccines, and would be much less expensive. We optimized methods to express HPV16 L1 protein and purify VLPs from tobacco (Nicotiana benthamiana) leaves transfected with the magnICON deconstructed viral vector expression system. L1 proteins were extracted from agro-infiltrated leaves using a series of pH and salt mediated buffers. Expression levels of L1 proteins and VLPs were verified by immunoblot and ELISA, which confirmed the presence of sequential and conformational epitopes, respectively. Among three constructs tested (16L1d22, TPL1d22, and TPL1F), TPL1F, containing a full-length L1 and chloroplast transit peptide, was best. Extraction of HPV16 L1 from leaf tissue was most efficient (> 2.5% of total soluble protein) with a low-salt phosphate buffer. VLPs were purified using both cesium chloride (CsCl) density gradient and size exclusion chromatography. Electron microscopy studies confirmed the presence of assembled forms of HPV16 L1 VLPs. Collectively; our results indicated that chloroplast-targeted transient expression in tobacco plants is promising for the production of a cheap, efficacious HPV16 L1 VLP vaccine. Studies are underway to develop plant VLPs for the production of a cervical cancer vaccine.

Item Type: Article
Erschienen: 2016
Creators: Zahin, Maryam and Joh, Joongho and Khanal, Sujita and Husk, Adam and Mason, Hugh and Warzecha, Heribert and Ghim, Shin-Je and Miller, Donald M. and Matoba, Nobuyuki and Jenson, Alfred Bennett
Title: Scalable Production of HPV16 L1 Protein and VLPs from Tobacco Leaves.
Language: English
Abstract:

Cervical cancer is the most common malignancy among women particularly in developing countries, with human papillomavirus (HPV) 16 causing 50% of invasive cervical cancers. A plant-based HPV vaccine is an alternative to the currently available virus-like particle (VLP) vaccines, and would be much less expensive. We optimized methods to express HPV16 L1 protein and purify VLPs from tobacco (Nicotiana benthamiana) leaves transfected with the magnICON deconstructed viral vector expression system. L1 proteins were extracted from agro-infiltrated leaves using a series of pH and salt mediated buffers. Expression levels of L1 proteins and VLPs were verified by immunoblot and ELISA, which confirmed the presence of sequential and conformational epitopes, respectively. Among three constructs tested (16L1d22, TPL1d22, and TPL1F), TPL1F, containing a full-length L1 and chloroplast transit peptide, was best. Extraction of HPV16 L1 from leaf tissue was most efficient (> 2.5% of total soluble protein) with a low-salt phosphate buffer. VLPs were purified using both cesium chloride (CsCl) density gradient and size exclusion chromatography. Electron microscopy studies confirmed the presence of assembled forms of HPV16 L1 VLPs. Collectively; our results indicated that chloroplast-targeted transient expression in tobacco plants is promising for the production of a cheap, efficacious HPV16 L1 VLP vaccine. Studies are underway to develop plant VLPs for the production of a cervical cancer vaccine.

Journal or Publication Title: PloS one
Volume: 11
Number: 8
Divisions: 10 Department of Biology
10 Department of Biology > Plant Biotechnology and Metabolic Engineering
Date Deposited: 30 Aug 2016 10:21
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