Zahin, Maryam ; Joh, Joongho ; Khanal, Sujita ; Husk, Adam ; Mason, Hugh ; Warzecha, Heribert ; Ghim, Shin-Je ; Miller, Donald M. ; Matoba, Nobuyuki ; Jenson, Alfred Bennett (2016)
Scalable Production of HPV16 L1 Protein and VLPs from Tobacco Leaves.
In: PloS one, 11 (8)
Artikel, Bibliographie
Kurzbeschreibung (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.
Typ des Eintrags: | Artikel |
---|---|
Erschienen: | 2016 |
Autor(en): | Zahin, Maryam ; Joh, Joongho ; Khanal, Sujita ; Husk, Adam ; Mason, Hugh ; Warzecha, Heribert ; Ghim, Shin-Je ; Miller, Donald M. ; Matoba, Nobuyuki ; Jenson, Alfred Bennett |
Art des Eintrags: | Bibliographie |
Titel: | Scalable Production of HPV16 L1 Protein and VLPs from Tobacco Leaves. |
Sprache: | Englisch |
Publikationsjahr: | 2016 |
Titel der Zeitschrift, Zeitung oder Schriftenreihe: | PloS one |
Jahrgang/Volume einer Zeitschrift: | 11 |
(Heft-)Nummer: | 8 |
Kurzbeschreibung (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. |
Fachbereich(e)/-gebiet(e): | 10 Fachbereich Biologie 10 Fachbereich Biologie > Plant Biotechnology and Metabolic Engineering |
Hinterlegungsdatum: | 30 Aug 2016 10:21 |
Letzte Änderung: | 30 Aug 2016 10:21 |
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