Department of Biological Sciences, National University of Singapore, Singapore, Singapore.
Bioinformatics Institute, Agency for Science, Technology, and Research (A*STAR), Singapore, Singapore.
Elife. 2021 Feb 8;10:e63646. doi: 10.7554/eLife.63646.
The spike (S) protein is the main handle for SARS-CoV-2 to enter host cells via surface angiotensin-converting enzyme 2 (ACE2) receptors. How ACE2 binding activates proteolysis of S protein is unknown. Here, using amide hydrogen-deuterium exchange mass spectrometry and molecular dynamics simulations, we have mapped the S:ACE2 interaction interface and uncovered long-range allosteric propagation of ACE2 binding to sites necessary for host-mediated proteolysis of S protein, critical for viral host entry. Unexpectedly, ACE2 binding enhances dynamics at a distal S1/S2 cleavage site and flanking protease docking site ~27 Å away while dampening dynamics of the stalk hinge (central helix and heptad repeat [HR]) regions ~130 Å away. This highlights that the stalk and proteolysis sites of the S protein are dynamic hotspots in the prefusion state. Our findings provide a dynamics map of the S:ACE2 interface in solution and also offer mechanistic insights into how ACE2 binding is allosterically coupled to distal proteolytic processing sites and viral-host membrane fusion. Thus, protease docking sites flanking the S1/S2 cleavage site represent alternate allosteric hotspot targets for potential therapeutic development.
刺突（S）蛋白是 SARS-CoV-2 通过表面血管紧张素转换酶 2（ACE2）受体进入宿主细胞的主要把手。ACE2 结合如何激活 S 蛋白的蛋白水解尚不清楚。在这里，我们使用酰胺氢氘交换质谱和分子动力学模拟，绘制了 S:ACE2 相互作用界面，并揭示了 ACE2 结合到宿主介导的 S 蛋白蛋白水解所必需的位点的长程变构传播，这对于病毒进入宿主至关重要。出乎意料的是，ACE2 结合增强了距离 S1/S2 切割位点和侧翼蛋白酶结合位点约 27 Å 的位点的动力学，同时抑制了距离茎铰链（中央螺旋和七肽重复 [HR]）区域约 130 Å 的位点的动力学。这突出表明 S 蛋白的茎和蛋白水解位点在预融合状态下是动态热点。我们的研究结果提供了 S:ACE2 界面在溶液中的动力学图谱，并为 ACE2 结合如何变构偶联到远端蛋白水解加工位点和病毒-宿主膜融合提供了机制见解。因此，S1/S2 切割位点侧翼的蛋白酶结合位点代表潜在治疗开发的替代变构热点靶标。
