Transplastomic expression of a modified human papillomavirus L1 protein leading to the assembly of capsomeres in tobacco
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ORIGINAL PAPER
Transplastomic expression of a modified human papillomavirus L1 protein leading to the assembly of capsomeres in tobacco: a step towards cost-effective second-generation vaccines M. Tahir Waheed • Nadja Tho¨nes • Martin Mu¨ller S. Waqas Hassan • N. Mona Razavi • Elke Lo¨ssl • Hans-Peter Kaul • Andreas G. Lo¨ssl
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Received: 22 January 2010 / Accepted: 2 June 2010 / Published online: 19 June 2010 Ó Springer Science+Business Media B.V. 2010
Abstract Certain types of human papillomaviruses (HPV) are causatively associated with cervical carcinoma, the second most common cancer in women worldwide. Due to limitations in the availability of currently used virus-like particle (VLP)-based vaccines against HPV to women of developing countries, where most cases of cervical cancer occur, the development of a cost-effective second-generation vaccine is a necessity. Capsomeres have recently been demonstrated to be highly immunogenic and to have a number of advantages as a potential cost-effective alternative to VLP-based HPV vaccines. We have expressed a mutated HPV-16 L1 (L1_2xCysM) gene that retained the ability to assemble L1 protein to capsomeres in tobacco chloroplasts. The recombinant protein yielded up to 1.5% of total soluble protein. The assembly of capsomeres was examined and verified by
Electronic supplementary material The online version of this article (doi:10.1007/s11248-010-9415-4) contains supplementary material, which is available to authorized users. M. T. Waheed S. W. Hassan N. M. Razavi E. Lo¨ssl H.-P. Kaul A. G. Lo¨ssl (&) Department of Applied Plant Sciences and Plant Biotechnology (DAPP), University of Natural Resources and Applied Life Sciences (BOKU), Gregor-MendelStrasse 33, 1180 Vienna, Austria e-mail: [email protected] N. Tho¨nes M. Mu¨ller Deutsches Krebsforschungszentrum, F035, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany
cesium chloride density gradient centrifugation and sucrose sedimentation analysis. An antigen capture enzyme-linked immunosorbent assay confirmed the formation of capsomeres by using a conformationspecific monoclonal antibody which recognized the conformational epitopes. Transplastomic tobacco plants exhibited normal growth and morphology, but all such lines showed male sterility in the T0, T1 and T2 generations. Taken together, these results indicate the possibility of producing a low-cost capsomere-based vaccine by plastids. Keywords HPV-16 L1_2xCysM gene Capsomeres Plastids ELISA Male sterility
Introduction Cervical cancer is the second most common cancer in women worldwide, with approximately 493,000 new cases and 274,000 deaths occurring annually around the globe (Parkin and Bray 2006). Human papillomavirus (HPV) types 16 and 18 have been identified to be responsible for approximately 70% of invasive cervical cancers, with HPV-16 being by far the most prevalent type and found in about 54% of all cervical cancer cases (Smith et al. 2007). L1 is the major capsid protein of HPV and self assembles into higher order molecular
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