鸭坦布苏病毒HN1株囊膜E蛋白抑制肽的筛选及鉴定

Screening and identification of specific binding peptides to duck Tembusu virus E protein by phage display peptide library

鸭坦布苏病毒(DTMUV)是一个新发现的病毒,主要引起鸭产蛋严重下降为特征,为鉴定与DTMUV囊膜E蛋白特异性结合的多肽,本试验利用噬菌体展示技术以DTMUV HN1株E蛋白为靶标,进行随机12肽库3轮筛选,结合ELISA试验和竞争抑制试验,成功筛选了2个能够与DTMUV HN1株E蛋白特异性结合的多肽。测序结果显示,其氨基酸序列分别为:HWSTRQGSTRWN(P3-12)和THRSWQGNSWYM(P8-12)。进一步分析多肽P3-12和P8-12对DTMUV在鸭胚成纤维细胞增殖方面的影响,结果显示,E蛋白抑制肽P3-12和P8-12本身对鸭胚成纤维细胞的增殖无明显影响,不同质量浓度的的抑制肽P3-12和P8-12(0.02、0.2、2、20mg/L)均能显著降低DTMUV在鸭胚成纤维细胞中的病毒滴度和病毒拷贝数,以上结果表明,抑制肽P3-12和P8-12均能抑制DTMUV在鸭胚成纤维细胞的增殖,本试验为进一步探明DTMUV与宿主的相互作用及抗病毒制剂的开发提供了理论依据。

Duck Tembusu virus （DTMUV） is newly emerged which causes severe egg-drop syndrome in duck. To identify the peptides that specifically bind to DTMUV E protein, in this study, E protein was used to screen its binding peptides from the 12-mer random phage display peptide library. After three rounds of biopanning, ELISA and competitive inhibition test, two positive phage clones were found and the sequencing results showed that they encoded the peptides of HWSTRQGSTRWN （P3-12）, and THRSWQGNSWYM （P8-12）. Furthermore, TCID50 and quantitative RT-PCR were used to detect the effect of E protein inhibitory peptide on the proliferation of DTMUV in duck embryo fibroblast （DEF）. The results indicated that P3-12 and P8-12 peptide had no significant effect on the proliferation of DEF. However,P3-12 and P8-12 peptides（0. 02,0. 2,2,20 g/mL） could significantly reduce the virus titer and virus copes of DTMUV in DEF. Taken together, this study provides novel insights on the interaction mechanism of Duck Tembusu virus and host cells.