Virology Laboratory, College of Biological Science and Technology, Heilongjiang Bayi Agricultural University, 5 Xinfeng road, Daqing, 163319, China.
Pharmacology laboratory, Heilongjiang Bayi Agricultural University, 5 Xinfeng road, Daqing, 163319, China.
Sci Rep. 2019 Nov 6;9(1):16094. doi: 10.1038/s41598-019-52446-5.
Previously, we successfully prepared a monoclonal antibody (mAb) named 2E4, that directly recognizes the major envelope protein B2L of the orf virus (ORFV), but there is little information about its epitope. Here, we meticulously mapped the 2E4 epitope through combinatorial programs and identified the functional binding domain and a key amino acid residue. Briefly, the simulated epitope peptide closely resembles VDVQSKDKDADELR located at the N-terminus of B2L, strongly suggesting that the epitope is conformationally or spatially structure-dependent. Subsequently, we combined these findings with the results from the antigenicity prediction of B2L to design three truncated fragments of B2L (F1, F2 and F3) selected using 2E4, and only the F1 fragment was found to be eligible for the advanced stage. Alanine-scanning mutagenesis suggested that the D residue is structurally crucial for the 2E4 epitope. The other participating residues, including K, E, and D, together with D were responsible for enabling 2E4 binding and served as factors that synergistically enabled binding to the whole 2E4 epitope. In this paper, we describe, for the first time, the architecture of an ORFV conformational epitope, and it is also expected that mAb 2E4 and its epitope can be used for applications relating to orf control.
先前，我们成功制备了一种单克隆抗体（mAb），命名为 2E4，该抗体可直接识别口疮病毒（ORFV）的主要包膜蛋白 B2L，但关于其表位的信息较少。在这里，我们通过组合程序仔细绘制了 2E4 表位，并确定了功能结合域和关键氨基酸残基。简而言之，模拟的表位肽与 B2L 氨基端的 VDVQSKDKDADELR 非常相似，强烈表明该表位是构象或空间结构依赖性的。随后，我们将这些发现与 B2L 的抗原性预测结果相结合，设计了三个使用 2E4 选择的 B2L 截断片段（F1、F2 和 F3），只有 F1 片段被发现符合高级阶段的要求。丙氨酸扫描诱变表明，D 残基对 2E4 表位具有结构重要性。其他参与的残基，包括 K、E 和 D，与 D 一起共同负责 2E4 结合，并作为协同结合整个 2E4 表位的因素。在本文中，我们首次描述了 ORFV 构象表位的结构，预计 mAb 2E4 及其表位可用于与口蹄疫控制相关的应用。
