Effect of mutations in capsid shell protein on the assembly of BmCPV virus-like particles
Effect of mutations in capsid shell protein on the assembly of BmCPV virus-like particles
复制标题
衣壳壳蛋白突变对 BmCPV 病毒样颗粒组装的影响。
DOI：
10.1099/jgv.0.001542
复制
发表时间：
2021-01-01
影响因子：
3.8
通讯作者：
Sun，Jingchen
中科院分区：
文献类型：
--
作者：
Ren，Feifei;Swevers，Luc;Sun，Jingchen
家蚕胞质多角体病毒(BmCPV)属于呼肠孤病毒科CypoVirus属，是一种典型的单层衣壳dsRNA病毒。冷冻电子显微镜观察结果表明，BmCPV衣壳由60个不对称单位组成，每个不对称单位含有一个转塔蛋白(TP)、两个大突起蛋白(LPP)和两个衣壳蛋白(CSP)。CSP具有自组装成病毒样颗粒(VLP)的能力，CSP中的小突起结构域(SPD)可能在病毒衣壳的组装中发挥重要作用。本研究利用聚合酶链式反应循环突变的原理，对SPD中的D828、S829和V945三个关键氨基酸位点进行了点突变。此外，利用多基因表达系统Ac-MultiBac杆状病毒在体外产生了8种不同的重组VLP。透射电子显微镜显示，单、双位点突变对VLP的组装效率和形态影响不大。尽管如此，这三个位点的同时突变还是产生了重大影响。实验结果表明，CSP的SPD在病毒衣壳的组装中起着至关重要的作用，为进一步分析BmCPV衣壳组装的分子和结构机制奠定了基础。
Bombyx mori cytoplasmic polyhedrosis virus (BmCPV) is a typical single-layer capsid dsRNA virus belonging to the genus Cypovirus in the family Reoviridae. The results of cryo-electron microscopy showed that the BmCPV capsid consists of 60 asymmetric units, and each asymmetric unit contains one turret protein (TP), two large protrusion proteins (LPP) and two capsid shell proteins (CSP). CSP has the ability to self-assemble into virus-like particles (VLPs), and the small protrusion domain (SPD) in CSP may play an essential role in the assembly of viral capsids. In this study, three critical amino acid sites, D828, S829 and V945, in the SPD were efficiently mutated (point mutation) based on the principle of PCR circular mutagenesis. Moreover, a multi-gene expression system, Ac-MultiBac baculovirus, was used to produce eight different recombinant VLPs in vitro. Transmission electron microscopy showed that the single site and double site mutations had little effect on the efficiency and morphology of the assembly of VLPs. Still, the simultaneous mutation of the three sites had a significant impact. The experimental results demonstrate that the SPD of CSP plays an essential role in assembly of the viral capsid, which lays the foundation for further analysis of the molecular and structural mechanism of BmCPV capsid assembly.