Identification of the Receptor-Binding Domain of the Spike Glycoprotein of Human Betacoronavirus HKU1
Identification of the Receptor-Binding Domain of the Spike Glycoprotein of Human Betacoronavirus HKU1
复制标题
人乙型冠状病毒 HKU1 刺突糖蛋白受体结合域的鉴定
DOI：
10.1128/jvi.03737-14
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发表时间：
2015-09-01
影响因子：
5.4
通讯作者：
Dominguez, Samuel R.
中科院分区：
文献类型：
--
作者：
Qian, Zhaohui;Ou, Xiuyuan;Dominguez, Samuel R.
冠状病毒刺突(S)糖蛋白介导受体结合、膜融合和病毒进入并决定宿主范围。A组鼠β-冠状病毒（β-CoV）利用S蛋白的n端结构域（NTD）与其受体结合，而B组β-冠状病毒（β-CoV）和C组中东呼吸综合征冠状病毒（β-CoV）以及几种α-冠状病毒（α-CoV）利用其S蛋白的下游C结构域识别其受体蛋白。为了鉴定A组人β-冠状病毒HKU1刺突蛋白的受体结合域，我们生成了一组单克隆抗体（mab），并将其定位到HKU1刺突蛋白的外域。它们没有与其他冠状病毒的S蛋白发生交叉反应。大多数HKU1刺突单克隆抗体识别的表位位于535和673氨基酸之间的C结构域，表明该区域具有免疫优势。其中两种单克隆抗体可阻断HKU1病毒对人原代气管支气管上皮细胞的感染。含有C结构域的截断的HKU1 S蛋白的HTBE细胞预孵育可阻断HKU1病毒的感染，但仅含有NTD的截断的S蛋白的HTBE细胞预孵育不能阻断感染。这些数据表明，HKU1刺突蛋白的受体结合域（receptor-binding domain， RBD）位于C结构域，B组α- cov和C组β- cov的刺突蛋白在此与其特异性受体蛋白结合。因此，A组中的两种β-冠状病毒HKU1和小鼠冠状病毒已经进化到使用其刺突糖蛋白的不同区域来识别各自的受体蛋白。a组中的β-冠状病毒小鼠肝炎病毒利用刺突蛋白中的半乳糖凝集素样NTD与其受体蛋白结合，而a组中的另一种β-冠状病毒HCoV-OC43则利用NTD与其含唾液酸的受体结合。与此形成鲜明对比的是，同样属于A组的人呼吸道β-CoV HKU1刺突糖蛋白的NTD不结合糖。在本研究中，我们发现对于HKU1的刺突蛋白，纯化的NTD下游的C结构域可以阻断HKU1病毒对人呼吸道上皮细胞的感染，并且几种映射到C结构域的单克隆抗体可以中和病毒的感染。因此，HKU1刺突糖蛋白的受体结合域位于C结构域。令人惊讶的是，A组中的两种β-冠状病毒，小鼠肝炎病毒和HKU1，已经进化到使用其刺突糖蛋白的不同区域来识别各自的受体。
Coronavirus spike (S) glycoproteins mediate receptor binding, membrane fusion, and virus entry and determine host range. Murine betacoronavirus (β-CoV) in group A uses the N-terminal domain (NTD) of S protein to bind to its receptor, whereas the β-CoVs severe acute respiratory syndrome CoV in group B and Middle East respiratory syndrome CoV in group C and several α-CoVs use the downstream C domain in their S proteins to recognize their receptor proteins. To identify the receptor-binding domain in the spike of human β-CoV HKU1 in group A, we generated and mapped a panel of monoclonal antibodies (MAbs) to the ectodomain of HKU1 spike protein. They did not cross-react with S proteins of any other CoV tested. Most of the HKU1 spike MAbs recognized epitopes in the C domain between amino acids 535 and 673, indicating that this region is immunodominant. Two of the MAbs blocked HKU1 virus infection of primary human tracheal-bronchial epithelial (HTBE) cells. Preincubation of HTBE cells with a truncated HKU1 S protein that includes the C domain blocked infection with HKU1 virus, but preincubation of cells with truncated S protein containing only the NTD did not block infection. These data suggest that the receptor-binding domain (RBD) of HKU1 spike protein is located in the C domain, where the spike proteins of α-CoVs and β-CoVs in groups B and C bind to their specific receptor proteins. Thus, two β-CoVs in group A, HKU1 and murine CoV, have evolved to use different regions of their spike glycoproteins to recognize their respective receptor proteins.IMPORTANCEMouse hepatitis virus, a β-CoV in group A, uses the galectin-like NTD in its spike protein to bind its receptor protein, while HCoV-OC43, another β-CoV in group A, uses the NTD to bind to its sialic-acid containing receptor. In marked contrast, the NTD of the spike glycoprotein of human respiratory β-CoV HKU1, which is also in group A, does not bind sugar. In this study, we showed that for the spike protein of HKU1, the purified C domain, downstream of the NTD, could block HKU1 virus infection of human respiratory epithelial cells, and that several monoclonal antibodies that mapped to the C domain neutralized virus infectivity. Thus, the receptor-binding domain of HKU1 spike glycoprotein is located in the C domain. Surprisingly, two β-CoVs in group A, mouse hepatitis virus and HKU1, have evolved to use different regions of their spike glycoproteins to recognize their respective receptors.