Modified tobacco mosaic virus particles as scaffolds for display of protein antigens for vaccine applications.
Modified tobacco mosaic virus particles as scaffolds for display of protein antigens for vaccine applications.
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DOI：
10.1016/j.virol.2005.12.039
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发表时间：
2006
期刊：
影响因子：
3.7
通讯作者：
Mark L. Smith;J. Lindbo;S. Dillard-Telm;P. Brosio;Amanda B. Lasnik;Alison A McCormick;L. Nguyen;K. Palmer
中科院分区：
文献类型：
--
作者：
Mark L. Smith;J. Lindbo;S. Dillard-Telm;P. Brosio;Amanda B. Lasnik;Alison A McCormick;L. Nguyen;K. Palmer
已知肽或蛋白质以有序的重复阵列展示，例如在病毒样颗粒的表面上展示，相对于用“游离”蛋白质抗原接种，诱导增强的免疫应答。烟草花叶病毒（TMV）的外壳蛋白可以容纳插入到一级序列中的短肽，但是作为与衣壳蛋白的遗传融合物的较大蛋白部分的展示是不可能的。我们采用了一种随机文库的方法，在外壳蛋白的外部氨基末端引入反应性赖氨酸，这有助于衣壳的生物素化。为了表征病毒粒子表面上异源蛋白的展示，我们将模型抗原（从植物表达和纯化的绿色荧光蛋白（GFP）-链霉亲和素（SA））结合到生物素化的TMV颗粒，产生GFP-SA装饰的病毒颗粒。获得26%的GFP-SA四聚体加载，对应于每个完整病毒体展示约2200个GFP部分。我们评估了GFP修饰的病毒粒子在小鼠和豚鼠中的免疫原性，发现相对于未结合的GFP-SA四聚体，两种物种中的体液IgG滴度增加。接下来，我们将犬口腔乳头瘤病毒L2蛋白的N-末端片段融合到链霉亲和素上。与TMV展示，L2蛋白片段是显着更强的免疫原性比未偶联抗原在小鼠中进行测试时。通过展示完整蛋白的呈现，这项研究扩展了TMV作为疫苗支架的实用性，超出了遗传操作的可能性。
Display of peptides or proteins in an ordered, repetitive array, such as on the surface of a virus-like particle, is known to induce an enhanced immune response relative to vaccination with the “free” protein antigen. The coat protein of Tobacco mosaic virus (TMV) can accommodate short peptide insertions into the primary sequence, but the display of larger protein moieties as genetic fusions to the capsid protein has not been possible. We employed a randomized library approach to introduce a reactive lysine at the externally located amino terminus of the coat protein, which facilitated biotinylation of the capsid. To characterize display of heterologous proteins on the virion surface, we bound a model antigen (green fluorescent protein (GFP)–streptavidin (SA), expressed and purified from plants) to the biotinylated TMV particles, creating a GFP-SA decorated virus particle. A GFP-SA tetramer loading of 26% was obtained, corresponding to approximately 2200 GFP moieties displayed per intact virion. We evaluated the immunogenicity of GFP decorated virions in both mice and guinea pigs and found augmented humoral IgG titers in both species, relative to unbound GFP-SA tetramer. Next, we fused an N-terminal fragment of the Canine oral papillomavirus L2 protein to streptavidin. With TMV display, the L2 protein fragment was significantly more immunogenic than uncoupled antigen when tested in mice. By demonstrating the presentation of whole proteins, this study expands the utility of TMV as a vaccine scaffold beyond that which is possible by genetic manipulation.