Isolation and characterization of porcine monoclonal antibodies revealed two distinct serotype-independent epitopes on VP2 of foot-and-mouth disease virus
Isolation and characterization of porcine monoclonal antibodies revealed two distinct serotype-independent epitopes on VP2 of foot-and-mouth disease virus
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猪单克隆抗体的分离和表征揭示了口蹄疫病毒 VP2 上两个不同的血清型独立表位
DOI：
10.1099/jgv.0.001608
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发表时间：
2021-01-01
影响因子：
3.8
通讯作者：
Lu, Zengjun
中科院分区：
文献类型：
--
作者：
Li, Kun;Zhu, Guoqiang;Lu, Zengjun
猪对口蹄疫病毒（FMDV）易感，体液免疫应答在保护猪免受FMDV感染中起重要作用。然而，很少有关于来自猪的单个B细胞的FMDV特异性mAb的信息。本研究的目的是确定口蹄疫病毒的抗原特性，被识别的抗体从猪。因此，使用基于荧光的单B细胞抗体技术开发了一组猪源性抗FMDV的mAb。Western blotting结果显示，其中3株抗体（1C 6、P2- 7 E和P2- 8 G）均能识别VP 2上的保守抗原表位，并对A型和O型口蹄疫病毒均表现出较强的免疫反应性。丙氨酸取代扫描测定和序列保守性分析阐明，这些猪mAb识别VP 2上的两个保守表位：N末端的线性表位（（KKTEETTLL 10）-K-2）和依赖于VP 2完整性的两个β折叠（B折叠和C折叠）上涉及残基K63、H65、L 66、F67、D 68和L 81的构象表位。IgG的重链和轻链的随机配对证实重链主要参与与抗原的结合。猪IgG的轻链有助于对抗原的结合亲和力，并且可以用作抗体稳定性的支持平台。总之，本研究首次揭示了猪B细胞识别的FMDV保守抗原谱，并提供了一种新的方法，用于在克隆水平上分析其天然宿主（即猪）中针对FMDV的抗体应答。
Pigs are susceptible to foot-and-mouth disease virus (FMDV), and the humoral immune response plays an essential role in protection against FMDV infection. However, little information is available about FMDV-specific mAbs derived from single B cells of pigs. This study aimed to determine the antigenic features of FMDV that are recognized by antibodies from pigs. Therefore, a panel of pig-derived mAbs against FMDV were developed using fluorescence-based single B cell antibody technology. Western blotting revealed that three of the antibodies (1C6, P2-7E and P2-8G) recognized conserved antigen epitopes on capsid protein VP2, and exhibited broad reactivity against both FMDV serotypes A and O. An alanine-substitution scanning assay and sequence conservation analysis elucidated that these porcine mAbs recognized two conserved epitopes on VP2: a linear epitope ((KKTEETTLL10)-K-2) in the N terminus and a conformational epitope involving residues K63, H65, L66, F67, D68 and L81 on two beta-sheets (B-sheet and C-sheet) that depended on the integrity of VP2. Random parings of heavy and light chains of the IgGs confirmed that the heavy chain is predominantly involved in binding to antigen. The light chain of porcine IgG contributes to the binding affinity toward an antigen and may function as a support platform for antibody stability. In summary, this study is the first to reveal the conserved antigenic profile of FMDV recognized by porcine B cells and provides a novel method for analysing the antibody response against FMDV in its natural hosts (i.e. pigs) at the clonal level.