Amino acids 270 to 510 of the severe acute respiratory syndrome coronavirus spike protein are required for interaction with receptor
Amino acids 270 to 510 of the severe acute respiratory syndrome coronavirus spike protein are required for interaction with receptor
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DOI：
10.1128/jvi.78.9.4552-4560.2004
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发表时间：
2004-05-01
影响因子：
5.4
通讯作者：
Ambrosino, DM
中科院分区：
文献类型：
--
作者：
Babcock, GJ;Esshaki, DJ;Ambrosino, DM
一种新型冠状病毒，严重急性呼吸综合征冠状病毒（SARS-CoV），最近被确定为严重急性呼吸综合征（SARS）的病原体。SARS-CoV在病毒粒子结构和基因组组织方面与其他冠状病毒相似。已知对于其他冠状病毒，刺突（S）糖蛋白是病毒附着至容许细胞以及病毒包膜与宿主细胞膜融合所需的。在这里，我们描述了一个可溶性的密码子优化的SARS-CoV S糖蛋白，包括天然S糖蛋白（S-1190）的前1190个氨基酸的构建和表达。密码子优化的和天然的S糖蛋白表现出相似的分子量，通过Western印迹分析确定，表明合成的S糖蛋白在哺乳动物表达系统中被正确修饰。荧光激活细胞分选仪分析表明，S-1190与Vero E6细胞（一种允许感染的细胞）表面结合，表明S-1190保持了天然S糖蛋白中存在的生物活性。这种相互作用被从恢复的SARS患者中获得的血清阻断，表明这种结合是特异性的。为了绘制SARS冠状病毒S糖蛋白的配体结合结构域，构建了S-1190糖蛋白的羧基末端和氨基末端截短。氨基酸270至510是SARS-CoV S糖蛋白的最小受体结合区域，如通过流式细胞术测定的。我们推测SARS-CoV S糖蛋白的氨基酸1至510代表了包含受体结合位点（氨基酸270至510）的独特结构域，类似于其他冠状病毒S糖蛋白的S1亚基。
A novel coronavirus, severe acute respiratory syndrome coronavirus (SARS-CoV), has recently been identified as the causative agent of severe acute respiratory syndrome (SARS). SARS-CoV appears similar to other coronaviruses in both virion structure and genome organization. It is known for other coronaviruses that the spike (S) glycoprotein is required for both viral attachment to permissive cells and for fusion of the viral envelope with the host cell membrane. Here we describe the construction and expression of a soluble codon-optimized SARS-CoV S glycoprotein comprising the first 1,190 amino acids of the native S glycoprotein (S-1190). The codon-optimized and native S glycoproteins exhibit similar molecular weight as determined by Western blot analysis, indicating that synthetic S glycoprotein is modified correctly in a mammalian expression system. S-1190 binds to the surface of Vero E6 cells, a cell permissive to infection, as demonstrated by fluorescence-activated cell sorter analysis, suggesting that S-1190 maintains the biologic activity present in native S glycoprotein. This interaction is blocked with serum obtained from recovering SARS patients, indicating that the binding is specific. In an effort to map the ligand-binding domain of the SARS-CoV S glycoprotein, carboxy- and amino-terminal truncations of the S-1190 glycoprotein were constructed. Amino acids 270 to 510 were the minimal receptor-binding region of the SARS-CoV S glycoprotein as determined by How cytometry. We speculate that amino acids 1 to 510 of the SARS-CoV S glycoprotein represent a unique domain containing the receptor-binding site (amino acids 270 to 510), analogous to the S1 subunit of other coronavirus S glycoproteins.