State Key Laboratory of Veterinary Etiological Biology, National/OIE Foot and Mouth Disease Reference Laboratory, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, 730046, People's Republic of China.
Department of Veterinary Basics and Diagnostic Sciences, College of Veterinary Science, Mekelle University, 2084, Mekelle, Tigray, Ethiopia.
Virol J. 2024 Apr 19;21(1):87. doi: 10.1186/s12985-024-02322-0.
Bovine parvovirus (BPV) is an autonomous DNA virus with a smaller molecular size and subtle differences in its structural proteins, unlike other animal parvoviruses. More importantly, this virus has the potential to produce visible to silent economic catastrophes in the livestock business, despite receiving very little attention. Parvoviral virus-like particles (VLPs) as vaccines and as logistical platforms for vaccine deployment are well studied. However, no single experimental report on the role of VP1 in the assembly and stability of BPV-VLPs is available. Furthermore, the self-assembly, integrity and stability of the VLPs of recombinant BPV VP2 in comparison to VP1 VP2 Cap proteins using any expression method has not been studied previously. In this study, we experimentally evaluated the self-assembling ability with which BPV virus-like particles (VLPs) could be synthesized from a single structural protein (VP2) and by integrating both VP2 and VP1 amino acid sequences.
In silico and experimental cloning methods were carried out. His-tagged and without-His-tag VP2 and V1VP2-encoding amino acid sequences were cloned and inserted into pFastbacdual, and insect cell-generated recombinant protein was evaluated by SDS‒PAGE and western blot. Period of infectivity and expression level were determined by IFA. The integrity and stability of the BPV VLPs were evaluated by transmission electron microscopy. The secondary structure of the BPV VLPs from both VP2 and V1VP2 was analyzed by circular dichroism.
Our findings show that VP2 alone was equally expressed and purified into detectable proteins, and the stability at different temperatures and pH values was not appreciably different between the two kinds of VLPs. Furthermore, BPV-VP2 VLPs were praised for their greater purity and integrity than BPV-VP1VP2 VLPs, as indicated by SDS‒PAGE. Therefore, our research demonstrates that the function of VP1 has no bearing on the stability or integrity of BPV-VLPs.
In summary, incredible physiochemically stable BPV VP2-derived VLPs have been found to be promising candidates for the development of multivalent vaccines and immunodiagnostic kits against enteric viruses and to carry heterogeneous epitopes for various economically important livestock diseases.
牛细小病毒 (BPV) 是一种自主 DNA 病毒，其结构蛋白的分子大小较小，结构上略有差异，与其他动物细小病毒不同。更重要的是，尽管这种病毒受到的关注很少，但它有可能在畜牧业中造成明显或隐性的经济灾难。细小病毒病毒样颗粒 (VLPs) 作为疫苗和疫苗部署的后勤平台已得到充分研究。然而，目前尚无关于 VP1 在 BPV-VLPs 组装和稳定性中的作用的单一实验报告。此外，以前从未研究过使用任何表达方法，重组 BPV VP2 的自组装、完整性和稳定性与 VP1 VP2 Cap 蛋白相比。在这项研究中，我们通过实验评估了从单个结构蛋白 (VP2) 合成 BPV 病毒样颗粒 (VLPs) 的自组装能力，以及整合 VP2 和 VP1 氨基酸序列的能力。
进行了计算机模拟和实验克隆方法。将带有 His 标签和不带 His 标签的 VP2 和 V1VP2 编码的氨基酸序列克隆并插入 pFastbacdual 中，并通过 SDS-PAGE 和 Western blot 评估昆虫细胞生成的重组蛋白。通过 IFA 确定感染期和表达水平。通过透射电子显微镜评估 BPV VLPs 的完整性和稳定性。通过圆二色性分析评估来自 VP2 和 V1VP2 的 BPV VLPs 的二级结构。
我们的研究结果表明，VP2 单独表达并可纯化出可检测的蛋白质，两种 VLPs 的稳定性在不同温度和 pH 值下没有明显差异。此外，与 BPV-VP1VP2 VLPs 相比，BPV-VP2 VLPs 具有更高的纯度和完整性，这一点通过 SDS-PAGE 得到了证明。因此，我们的研究表明 VP1 的功能与 BPV-VLPs 的稳定性或完整性无关。
综上所述，我们发现具有令人难以置信的物理化学稳定性的 BPV VP2 衍生 VLPs 是开发针对肠病毒的多价疫苗和免疫诊断试剂盒以及携带各种重要经济动物疾病异质表位的有前途的候选物。
