Guo Ying, Tisoncik Jennifer, McReynolds Susanna, Farzan Michael, Prabhakar Bellur S, Gallagher Thomas, Rong Lijun, Caffrey Michael
Department of Microbiology and Immunology, College of Medicine, University of Illinois at Chicago, Chicago, IL 60612, USA.
J Mol Biol. 2009 Dec 11;394(4):600-5. doi: 10.1016/j.jmb.2009.10.032. Epub 2009 Oct 21.
Infection by severe acute respiratory syndrome coronavirus (SARS-CoV) is initiated by specific interactions between the SARS-CoV spike (S) protein and its receptor ACE2. In this report, we screened a peptide library representing the SARS-CoV S protein sequence using a human immunodeficiency virus-based pseudotyping system to identify specific regions that affect viral entry. One of the 169 peptides screened, peptide 9626 (S residues 217-234), inhibited SARS-CoV S-mediated entry of the pseudotyped virions in 293T cells expressing a functional SARS-CoV receptor (human angiotensin-converting enzyme 2) in a dose-dependent manner (IC(50) approximately 11 microM). Alanine scanning mutagenesis was performed to assess the roles of individual residues within this region of S, which was previously uncharacterized. The effects included significant reductions in expression (K223A), viral incorporation (L218A, I230A, and N232A), and reduced viral entry (L224A, L226A, I228A, T231A, and F233A). Taken together, these results reveal a new region of the S protein that is crucial for SARS-CoV entry.
严重急性呼吸综合征冠状病毒（SARS-CoV）的感染是由SARS-CoV刺突（S）蛋白与其受体血管紧张素转换酶2（ACE2）之间的特异性相互作用引发的。在本报告中，我们使用基于人类免疫缺陷病毒的假型系统筛选了一个代表SARS-CoV S蛋白序列的肽库，以鉴定影响病毒进入的特定区域。在筛选的169种肽中，肽9626（S蛋白第217 - 234位氨基酸残基）以剂量依赖性方式抑制了SARS-CoV S介导的假型病毒粒子进入表达功能性SARS-CoV受体（人类血管紧张素转换酶2）的293T细胞（半数抑制浓度约为11微摩尔）。进行丙氨酸扫描诱变以评估S蛋白这一此前未被表征区域内各个残基的作用。这些作用包括表达显著降低（K223A）、病毒掺入减少（L218A、I230A和N232A）以及病毒进入减少（L224A、L226A、I228A、T231A和F233A）。综上所述，这些结果揭示了S蛋白中一个对SARS-CoV进入至关重要的新区域。
