Department of Biochemistry , University of Oxford , South Parks Road , Oxford OX1 3QU , United Kingdom.
Jenner Institute , University of Oxford , Oxford OX3 7DQ , United Kingdom.
ACS Nano. 2018 Sep 25;12(9):8855-8866. doi: 10.1021/acsnano.8b02805. Epub 2018 Jul 26.
Nanoscale organization is crucial to stimulating an immune response. Using self-assembling proteins as multimerization platforms provides a safe and immunogenic system to vaccinate against otherwise weakly immunogenic antigens. Such multimerization platforms are generally based on icosahedral viruses and have led to vaccines given to millions of people. It is unclear whether synthetic protein nanoassemblies would show similar potency. Here we take the computationally designed porous dodecahedral i301 60-mer and rationally engineer this particle, giving a mutated i301 (mi3) with improved particle uniformity and stability. To simplify the conjugation of this nanoparticle, we employ a SpyCatcher fusion of mi3, such that an antigen of interest linked to the SpyTag peptide can spontaneously couple through isopeptide bond formation (Plug-and-Display). SpyCatcher-mi3 expressed solubly to high yields in Escherichia coli, giving more than 10-fold greater yield than a comparable phage-derived icosahedral nanoparticle, SpyCatcher-AP205. SpyCatcher-mi3 nanoparticles showed high stability to temperature, freeze-thaw, lyophilization, and storage over time. We demonstrate approximately 95% efficiency coupling to different transmission-blocking and blood-stage malaria antigens. Plasmodium falciparum CyRPA was conjugated to SpyCatcher-mi3 nanoparticles and elicited a high avidity antibody response, comparable to phage-derived virus-like particles despite their higher valency and RNA cargo. The simple production, precise derivatization, and exceptional ruggedness of this nanoscaffold should facilitate broad application for nanobiotechnology and vaccine development.
纳米级组织对于刺激免疫反应至关重要。使用自组装蛋白作为多聚化平台为针对弱免疫原性抗原的疫苗接种提供了一种安全且具有免疫原性的系统。这种多聚化平台通常基于二十面体病毒，并已导致为数百万人接种疫苗。目前尚不清楚合成蛋白纳米组装体是否具有类似的效力。在这里，我们采用计算设计的多孔十二面体 i301 60 聚体，并对该颗粒进行合理工程改造，得到突变的 i301（mi3），其颗粒均匀性和稳定性得到改善。为了简化该纳米颗粒的缀合，我们采用 mi3 的 SpyCatcher 融合，使得与 SpyTag 肽连接的感兴趣抗原可以通过异肽键形成（Plug-and-Display）自动偶联。可溶性表达的 SpyCatcher-mi3 在大肠杆菌中以高产量表达，产量比可比的噬菌体衍生的二十面体纳米颗粒 SpyCatcher-AP205 高 10 倍以上。SpyCatcher-mi3 纳米颗粒对温度、冻融、冻干和长期储存具有很高的稳定性。我们证明了与不同的传播阻断和血期疟原抗原的偶联效率约为 95%。将 PfCyRPA 与 SpyCatcher-mi3 纳米颗粒缀合后，可引发高亲和力的抗体反应，与噬菌体衍生的病毒样颗粒相当，尽管它们的价数和 RNA 载量更高。这种纳米支架的简单生产、精确衍生化和卓越的坚固性应有助于纳米生物技术和疫苗开发的广泛应用。
