Wang Heng, Liu Rongchang, Cui Jin, Deng Shengchao, Xie Jiexiong, Nin Zhangyong, Zhang Guihong
College of Veterinary Medicine, South China Agricultural University, Guangzhou, Guangdong Province 510642, People's Republic of China; Guangdong Provincial Key Laboratory of Prevention and Control for Severe Clinical Animal Diseases, Guangzhou, Guangdong Province 510642, People's Republic of China; MOA Key Laboratory of Animal Vaccine Development, Guangzhou, Guangdong Province 510642, People's Republic of China.
College of Veterinary Medicine, South China Agricultural University, Guangzhou, Guangdong Province 510642, People's Republic of China; Guangdong Provincial Key Laboratory of Prevention and Control for Severe Clinical Animal Diseases, Guangzhou, Guangdong Province 510642, People's Republic of China; MOA Key Laboratory of Animal Vaccine Development, Guangzhou, Guangdong Province 510642, People's Republic of China.
J Virol Methods. 2015 Sep 15;222:231-41. doi: 10.1016/j.jviromet.2015.04.026. Epub 2015 May 2.
High-affinity peptides to porcine reproductive and respiratory syndrome virus (PRRSV) nonstructural protein (nsp) 7 were identified using phage-display technology. Five 12-amino-acid peptide sequences were identified after six rounds of biopanning. A putative CD##WC motif was found in two different consensus peptides borne by phages 4 and 5. The peptides borne by phages 4, 5, and 6 were synthesized for subsequent experiments, according to the results of the binding assays. Immunofluorescence assay revealed that all these peptides recognized nsp7 in PRRSV-infected cells. Furthermore, the peptides demonstrated antiviral activities, with peptides 5 and 6 showing effective inhibition. Early peptide stimulation was associated with strong antiviral activity, and the inhibitory effects of the peptides were dose-dependent at 36 and 48 h post-infection. Peptide 5 was selected to detect the intracellular localization of nsp7 by confocal microscopy. This peptide had a similar effect to anti-nsp7 monoclonal antibody on nsp7. These results suggest that high-affinity peptides to PRRSV nsp7 could mimic the potential of nsp7 antibody as a diagnostic reagent for virus detection. Moreover, the peptides selected in this study represented a potentially effective antiviral candidate to inhibit PRRSV.
利用噬菌体展示技术鉴定了针对猪繁殖与呼吸综合征病毒（PRRSV）非结构蛋白（nsp）7的高亲和力肽。经过六轮生物淘选后，鉴定出了五个12氨基酸的肽序列。在噬菌体4和5携带的两种不同共有肽中发现了一个假定的CD##WC基序。根据结合试验结果，合成了噬菌体4、5和6携带的肽用于后续实验。免疫荧光试验表明，所有这些肽都能识别PRRSV感染细胞中的nsp7。此外，这些肽表现出抗病毒活性，其中肽5和6表现出有效的抑制作用。早期肽刺激与强大的抗病毒活性相关，并且在感染后36小时和48小时，肽的抑制作用呈剂量依赖性。选择肽5通过共聚焦显微镜检测nsp7的细胞内定位。该肽对nsp7的作用与抗nsp7单克隆抗体相似。这些结果表明，针对PRRSV nsp7的高亲和力肽可模拟nsp7抗体作为病毒检测诊断试剂的潜力。此外，本研究中筛选出的肽代表了一种潜在有效的抑制PRRSV的抗病毒候选物。
