Kueh Chare Li, Yong Chean Yeah, Masoomi Dezfooli Seyedehsara, Bhassu Subha, Tan Soon Guan, Tan Wen Siang
Dept. of Cell and Molecular Biology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400 Serdang, Selangor, Malaysia.
Dept. of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400 Serdang, Selangor, Malaysia.
Biotechnol Prog. 2017 Mar;33(2):549-557. doi: 10.1002/btpr.2409. Epub 2016 Nov 29.
Macrobrachium rosenbergii nodavirus (MrNV) is a virus native to giant freshwater prawn. Recombinant MrNV capsid protein has been produced in Escherichia coli, which self-assembled into virus-like particles (VLPs). However, this recombinant protein is unstable, degrading and forming heterogenous VLPs. In this study, MrNV capsid protein was produced in insect Spodoptera frugiperda (Sf9) cells through a baculovirus system. Dynamic light scattering (DLS) and transmission electron microscopy (TEM) revealed that the recombinant protein produced by the insect cells self-assembled into highly stable, homogenous VLPs each of approximately 40 nm in diameter. Enzyme-linked immunosorbent assay (ELISA) showed that the VLPs produced in Sf9 cells were highly antigenic and comparable to those produced in E. coli. In addition, the Sf9 produced VLPs were highly stable across a wide pH range (2-12). Interestingly, the Sf9 produced VLPs contained DNA of approximately 48 kilo base pairs and RNA molecules. This study is the first report on the production and characterization of MrNV VLPs produced in a eukaryotic system. The MrNV VLPs produced in Sf9 cells were about 10 nm bigger and had a uniform morphology compared with the VLPs produced in E. coli. The insect cell production system provides a good source of MrNV VLPs for structural and immunological studies as well as for host-pathogen interaction studies. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:549-557, 2017.
罗氏沼虾诺达病毒（MrNV）是一种原产于巨型淡水虾的病毒。重组MrNV衣壳蛋白已在大肠杆菌中产生，其自组装成病毒样颗粒（VLP）。然而，这种重组蛋白不稳定，会降解并形成异质VLP。在本研究中，MrNV衣壳蛋白通过杆状病毒系统在昆虫草地贪夜蛾（Sf9）细胞中产生。动态光散射（DLS）和透射电子显微镜（TEM）显示，昆虫细胞产生的重组蛋白自组装成高度稳定、均匀的VLP，每个直径约为40 nm。酶联免疫吸附测定（ELISA）表明，Sf9细胞中产生的VLP具有高度抗原性，与大肠杆菌中产生的VLP相当。此外，Sf9产生的VLP在很宽的pH范围（2-12）内都非常稳定。有趣的是，Sf9产生的VLP含有约48千碱基对的DNA和RNA分子。本研究是关于在真核系统中产生和表征MrNV VLP的首次报道。与大肠杆菌中产生的VLP相比，Sf9细胞中产生的MrNV VLP大约大10 nm，并且具有均匀的形态。昆虫细胞生产系统为结构和免疫学研究以及宿主-病原体相互作用研究提供了良好的MrNV VLP来源。© 2016美国化学工程师学会生物技术进展，33:549-557，2017。
