Zhou Ji-Yong, Shang Shao-Bin, Gong Hui, Chen Qing-Xin, Wu Jian-Xiang, Shen Hui-Gang, Chen Ting-Fei, Guo Jun-Qing
Institute of Preventive Veterinary Medicine, College of Animal Sciences, Zhejiang University, 268 Kaixuan Road, Hangzhou, Zhejiang 310029, China.
J Biotechnol. 2005 Aug 4;118(2):201-11. doi: 10.1016/j.jbiotec.2005.02.017.
We expressed firstly the Capsid protein gene defecting the nuclear localization signal (NLS) of Porcine circovirus type II (PCV2) in Escherichia coli as a fusion protein with glutathione S-transferase (rGST-dCap protein). The purified rGST-dCap protein and the recombinant NLS-defected Cap protein of PCV2 (rdCap protein) from the purified rGST-dCap protein reacted specifically with swine antiserum to PCV2. Furthermore, the obtained monoclonal antibodies (mAbs) to rdCap protein were shown to bind to PCV2 particles replicated in PK15 cell and capsid protein (Cap protein) of PCV2 expressed in PK15 cells, respectively. mAbs to rdCap protein also revealed the neutralizing ability to PCV2 particles. These results demonstrated that rGST-dCap protein expressed in E. coli was folded correctly or at least partly, and mAbs to rdCap protein possessed the binding epitopes of PCV2 particles whereas mAbs 4C4 and 3F6 to rdCap protein remained the neutralization epitope of PCV2 particle, showing a possibility of neutralizing mAb to rdCap protein as an immnuotherapeutic agent and a potential of rGST-dCap protein as a vaccine antigen or serodiagnostic reagent.
我们首先在大肠杆菌中表达了缺失猪圆环病毒2型（PCV2）核定位信号（NLS）的衣壳蛋白基因，使其作为与谷胱甘肽S-转移酶的融合蛋白（rGST-dCap蛋白）。纯化的rGST-dCap蛋白以及从纯化的rGST-dCap蛋白中获得的重组NLS缺失的PCV2衣壳蛋白（rdCap蛋白）与猪抗PCV2血清发生特异性反应。此外，所获得的针对rdCap蛋白的单克隆抗体（mAbs）分别显示与在PK15细胞中复制的PCV2颗粒以及在PK15细胞中表达的PCV2衣壳蛋白（Cap蛋白）结合。针对rdCap蛋白的mAbs也显示出对PCV2颗粒的中和能力。这些结果表明，在大肠杆菌中表达的rGST-dCap蛋白能够正确折叠或至少部分正确折叠，针对rdCap蛋白的mAbs具有PCV2颗粒的结合表位，而针对rdCap蛋白的mAbs 4C4和3F6保留了PCV2颗粒的中和表位，这表明针对rdCap蛋白的中和性mAb有可能作为免疫治疗剂，并且rGST-dCap蛋白有作为疫苗抗原或血清学诊断试剂的潜力。
