Andoh Kiyohiko, Ashikaga Kanako, Suenaga Kiyotaka, Endo Shun, Yamazaki Kenichi
A Animal Pharmaceuticals Division, Chemo-Sero-Therapeutic Research Institute, 1-6-1 Okubo, Kita-ku, Kumamoto, Kumamoto 860-8568, Japan.
Avian Dis. 2018 Jun;62(2):210-217. doi: 10.1637/11796-011518-Reg.1.
We identified novel linear epitopes on the infectious bronchitis virus (IBV) spike S2 region. The conformational structure of the IBV spike protein was predicted from a homologous protein, human coronavirus NL63 spike. Although the obtained structure was incomplete, most of the IBV spike protein structure was predicted; the N-terminus of the S1 region could not be predicted due to its variability. In the model, the region located in the proximity of the fusion peptide appeared to be well conserved, and we evaluated the antigenicity of these domains, which are involved in the membrane fusion machinery. Western blotting revealed that IBV TM86 spike residues 686-723 were antigenic. Epitope mapping analysis using synthesized peptides revealed that IBV TM86 spike 669-685 (SNFSTGAFNISLLLTPP), 686-697 (SNPRGRSFIEDL), and 692-703 (SFIEDLLFTSVE) residues were major linear epitopes; two identified epitopes (686-697 and 692-703) were covered by the fusion peptide, and the other epitope (669-685) was adjacent to the fusion peptide. Although the identified epitopes are identically located as the neutralizing epitope in severe acute respiratory syndrome coronavirus, the recombinant protein that includes those epitopes could not elicit neutralizing antibodies against IBV. This is the first report describing IBV spike S2 epitopes located in the proximity of the fusion peptide, and it is suggested that the spike fusion machinery of IBV may differ from that of severe acute respiratory syndrome coronavirus, or, alternatively, IBV may have another mechanism to penetrate the cell membrane.
我们在传染性支气管炎病毒（IBV）刺突S2区域鉴定出了新的线性表位。IBV刺突蛋白的构象结构是根据同源蛋白人冠状病毒NL63刺突预测的。尽管获得的结构不完整，但大部分IBV刺突蛋白结构已被预测；由于S1区域N端的变异性，该区域无法预测。在模型中，位于融合肽附近的区域似乎高度保守，我们评估了这些参与膜融合机制的结构域的抗原性。蛋白质印迹分析显示，IBV TM86刺突蛋白的686 - 723位残基具有抗原性。使用合成肽进行的表位作图分析表明，IBV TM86刺突蛋白的669 - 685位（SNFSTGAFNISLLLTPP）、686 - 697位（SNPRGRSFIEDL）和692 - 703位（SFIEDLLFTSVE）残基是主要的线性表位；两个已鉴定的表位（686 - 697和692 - 703）被融合肽覆盖，另一个表位（669 - 685）与融合肽相邻。尽管所鉴定的表位与严重急性呼吸综合征冠状病毒中的中和表位位置相同，但包含这些表位的重组蛋白不能引发针对IBV的中和抗体。这是第一份描述位于融合肽附近的IBV刺突S2表位的报告，提示IBV的刺突融合机制可能与严重急性呼吸综合征冠状病毒不同，或者IBV可能有另一种穿透细胞膜的机制。
