Guo Yukun, Guo Ruizhen, Ma Yingxian, Chang Wenru, Ming Shengli, Yang Guoyu, Guo Yujie
College of Veterinary Medicine, Henan Agricultural University, Zhengzhou 450046, China.
Curr Top Med Chem. 2021 Oct 5;21(14):1235-1250. doi: 10.2174/1568026621666210618145411.
Virus-like Particles (VLPs) are non-genetic multimeric nanoparticles synthesized through in vitro or in vivo self-assembly of one or more viral structural proteins. Immunogenicity and safety of VLPs make them ideal candidates for vaccine development and efficient nanocarriers for foreign antigens or adjuvants to activate the immune system.
The present study aimed to design and synthesize a chimeric VLP vaccine of the phage Qbeta (Qβ) coat protein presenting the universal epitope of the coronavirus.
The RNA phage Qβ coat protein was designed and synthesized, denoted as Qbeta. The CoV epitope, a universal epitope of coronavirus, was inserted into the C-terminal of Qbeta using genetic recombination, designated as Qbeta-CoV. The N-terminal of Qbeta-CoV was successively inserted into the TEV restriction site using mCherry red fluorescent label and modified affinity purified histidine label 6xHE, which was denoted as HE-Qbeta-CoV. Isopropyl β-D-1-thiogalactopyranoside (IPTG) assessment revealed the expression of Qbeta, Qbeta-CoV, and HE-Qbeta-CoV in the BL21 (DE3) cells. The fusion protein was purified by salting out using ammonium sulfate and affinity chromatography. The morphology of particles was observed using electron microscopy. The female BALB/C mice were immunized intraperitoneally with the Qbeta-CoV and HE-Qbeta-- CoV chimeric VLPs vaccines and their sera were collected for the detection of antibody level and antibody titer using ELISA. The serum is used for the neutralization test of the three viruses of MHV, PEDV, and PDCoV.
The results revealed that the fusion proteins Qbeta, Qbeta-CoV, and HE-Qbeta-CoV could all obtain successful expression. Particles with high purity were obtained after purification; the chimeric particles of Qbeta-CoV and HE-Qbeta-CoV were found to be similar to Qbeta particles in morphology and formed chimeric VLPs. In addition, two chimeric VLP vaccines induced specific antibody responses in mice and the antibodies showed certain neutralizing activity.
The successful construction of the chimeric VLPs of the phage Qβ coat protein presenting the universal epitope of coronavirus provides a vaccine form with potential clinical applications for the treatment of coronavirus disease.
病毒样颗粒（VLPs）是通过一种或多种病毒结构蛋白在体外或体内自组装合成的非基因多聚体纳米颗粒。VLPs的免疫原性和安全性使其成为疫苗开发的理想候选者，以及用于激活免疫系统的外源抗原或佐剂的高效纳米载体。
本研究旨在设计并合成一种呈现冠状病毒通用表位的噬菌体Qβ（Qβ）衣壳蛋白嵌合VLP疫苗。
设计并合成RNA噬菌体Qβ衣壳蛋白，命名为Qbeta。利用基因重组将冠状病毒通用表位CoV表位插入Qbeta的C端，命名为Qbeta-CoV。使用mCherry红色荧光标签和修饰的亲和纯化组氨酸标签6xHE将Qbeta-CoV的N端依次插入TEV酶切位点，命名为HE-Qbeta-CoV。异丙基-β-D-硫代半乳糖苷（IPTG）评估显示Qbeta、Qbeta-CoV和HE-Qbeta-CoV在BL21（DE3）细胞中表达。融合蛋白通过硫酸铵盐析和亲和层析进行纯化。使用电子显微镜观察颗粒形态。雌性BALB/C小鼠腹腔注射Qbeta-CoV和HE-Qbeta-CoV嵌合VLP疫苗，收集其血清，采用ELISA检测抗体水平和抗体效价。血清用于对小鼠肝炎病毒（MHV）、猪流行性腹泻病毒（PEDV）和猪德尔塔冠状病毒（PDCoV）三种病毒进行中和试验。
结果显示，融合蛋白Qbeta、Qbeta-CoV和HE-Qbeta-CoV均能成功表达。纯化后获得了高纯度的颗粒；发现Qbeta-CoV和HE-Qbeta-CoV的嵌合颗粒在形态上与Qbeta颗粒相似，形成了嵌合VLPs。此外，两种嵌合VLP疫苗在小鼠中诱导了特异性抗体反应，且抗体表现出一定的中和活性。
成功构建了呈现冠状病毒通用表位的噬菌体Qβ衣壳蛋白嵌合VLPs，为冠状病毒病的治疗提供了一种具有潜在临床应用价值的疫苗形式。
