Scalable Production of HPV16 L1 Protein and VLPs from Tobacco Leaves.
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 le...
| Main Authors: | , , , , , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
Public Library of Science (PLoS)
2016-01-01
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| Series: | PLoS ONE |
| Online Access: | http://europepmc.org/articles/PMC4982596?pdf=render |
| _version_ | 1818410080367804416 |
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| author | Maryam Zahin Joongho Joh Sujita Khanal Adam Husk Hugh Mason Heribert Warzecha Shin-Je Ghim Donald M Miller Nobuyuki Matoba Alfred Bennett Jenson |
| author_facet | Maryam Zahin Joongho Joh Sujita Khanal Adam Husk Hugh Mason Heribert Warzecha Shin-Je Ghim Donald M Miller Nobuyuki Matoba Alfred Bennett Jenson |
| author_sort | Maryam Zahin |
| collection | DOAJ |
| description | 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. |
| first_indexed | 2024-12-14T10:09:50Z |
| format | Article |
| id | doaj.art-eca84e183f284eadbbd1707ce7fa912b |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-14T10:09:50Z |
| publishDate | 2016-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-eca84e183f284eadbbd1707ce7fa912b2022-12-21T23:07:01ZengPublic Library of Science (PLoS)PLoS ONE1932-62032016-01-01118e016099510.1371/journal.pone.0160995Scalable Production of HPV16 L1 Protein and VLPs from Tobacco Leaves.Maryam ZahinJoongho JohSujita KhanalAdam HuskHugh MasonHeribert WarzechaShin-Je GhimDonald M MillerNobuyuki MatobaAlfred Bennett JensonCervical 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.http://europepmc.org/articles/PMC4982596?pdf=render |
| spellingShingle | Maryam Zahin Joongho Joh Sujita Khanal Adam Husk Hugh Mason Heribert Warzecha Shin-Je Ghim Donald M Miller Nobuyuki Matoba Alfred Bennett Jenson Scalable Production of HPV16 L1 Protein and VLPs from Tobacco Leaves. PLoS ONE |
| title | Scalable Production of HPV16 L1 Protein and VLPs from Tobacco Leaves. |
| title_full | Scalable Production of HPV16 L1 Protein and VLPs from Tobacco Leaves. |
| title_fullStr | Scalable Production of HPV16 L1 Protein and VLPs from Tobacco Leaves. |
| title_full_unstemmed | Scalable Production of HPV16 L1 Protein and VLPs from Tobacco Leaves. |
| title_short | Scalable Production of HPV16 L1 Protein and VLPs from Tobacco Leaves. |
| title_sort | scalable production of hpv16 l1 protein and vlps from tobacco leaves |
| url | http://europepmc.org/articles/PMC4982596?pdf=render |
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