Plant Pathology Department, Faculty of Agriculture, Tarbiat Modares University, Pajouhesh Blvd., Tehran to Karaj highway, Tehran, Iran.
Laboratory for the Analysis of Medicines (LAM), Department of Pharmaceutical Sciences, CIRM, University of Liège, Quartier Hôpital, B36, Tower 4, Avenue Hippocrate, 15, 4000, Liège, Belgium.
BMC Biotechnol. 2019 Nov 21;19(1):81. doi: 10.1186/s12896-019-0566-y.
Virus-like particle (VLP) platform represents a promising approach for the generation of efficient and immunogenic subunit vaccines. Here, the feasibility of using grapevine fanleaf virus (GFLV) VLPs as a new carrier for the presentation of human papillomavirus (HPV) L2 epitope was studied. To achieve this goal, a model of the HPV L2 epitope secondary structure was predicted and its insertion within 5 external loops in the GFLV capsid protein (CP) was evaluated.
The epitope sequence was genetically inserted in the αB-αB domain C of the GFLV CP, which was then over-expressed in Pichia pastoris and Escherichia coli. The highest expression yield was obtained in E. coli. Using this system, VLP formation requires a denaturation-refolding step, whereas VLPs with lower production yield were directly formed using P. pastoris, as confirmed by electron microscopy and immunostaining electron microscopy. Since the GFLV L2 VLPs were found to interact with the HPV L2 antibody under native conditions in capillary electrophoresis and in ELISA, it can be assumed that the inserted epitope is located at the VLP surface with its proper ternary structure.
The results demonstrate that GFLV VLPs constitute a potential scaffold for surface display of the epitope of interest.
病毒样颗粒（VLP）平台代表了一种生成高效和免疫原性亚单位疫苗的有前途的方法。在这里，研究了使用葡萄扇叶病毒（GFLV）VLPs 作为展示人乳头瘤病毒（HPV）L2 表位的新型载体的可行性。为了实现这一目标，预测了 HPV L2 表位的二级结构模型，并评估了其在 GFLV 衣壳蛋白（CP）5 个外部环中的插入。
该表位序列被遗传插入 GFLV CP 的αB-αB 结构域 C 中，然后在毕赤酵母和大肠杆菌中过表达。在大肠杆菌中获得了最高的表达产量。使用该系统，VLP 的形成需要变性-复性步骤，而使用毕赤酵母则可以直接形成产量较低的 VLPs，这通过电子显微镜和免疫染色电子显微镜得到证实。由于在毛细管电泳和 ELISA 中发现 GFLV L2 VLPs 在天然条件下与 HPV L2 抗体相互作用，因此可以假设插入的表位位于 VLP 表面，具有适当的三元结构。
结果表明，GFLV VLPs 构成了展示感兴趣表位的潜在支架。
