The entry of the coronavirus SARS-CoV-2 into human lung cells can be inhibited by the approved drugs camostat and nafamostat. Here we elucidate the molecular mechanism of these drugs by combining experiments and simulations. In vitro assays confirm that both drugs inhibit the human protein TMPRSS2, a SARS-Cov-2 spike protein activator. As no experimental structure is available, we provide a model of the TMPRSS2 equilibrium structure and its fluctuations by relaxing an initial homology structure with extensive 330 microseconds of all-atom molecular dynamics (MD) and Markov modeling. Through Markov modeling, we describe the binding process of both drugs and a metabolic product of camostat (GBPA) to TMPRSS2, reaching a Michaelis complex (MC) state, which precedes the formation of a long-lived covalent inhibitory state. We find that nafamostat has a higher MC population than camostat and GBPA, suggesting that nafamostat is more readily available to form the stable covalent enzyme–substrate intermediate, effectively explaining its high potency. This model is backed by our in vitro experiments and consistent with previous virus cell entry assays. Our TMPRSS2–drug structures are made public to guide the design of more potent and specific inhibitors.

中文翻译：

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卡莫司他和萘莫司他抑制 SARS-CoV-2 细胞进入促进剂 TMPRSS2 的分子机制

批准的药物卡莫司他和萘莫司他可以抑制冠状病毒 SARS-CoV-2 进入人肺细胞。在这里，我们通过结合实验和模拟来阐明这些药物的分子机制。体外化验证实，这两种药物都能抑制人类蛋白质 TMPRSS2，这是一种 SARS-Cov-2 刺突蛋白激活剂。由于没有可用的实验结构，我们通过放松具有广泛的 330 微秒全原子分子动力学 (MD) 和马尔可夫建模的初始同源结构来提供 TMPRSS2 平衡结构及其波动的模型。通过马尔科夫模型，我们描述了药物和卡莫司他 (GBPA) 的代谢产物与 TMPRSS2 的结合过程，达到了米氏复合体 (MC) 状态，该状态先于长寿命共价抑制状态的形成。我们发现萘莫司他的 MC 数量高于卡莫司他和 GBPA，这表明萘莫司他更容易形成稳定的共价酶-底物中间体，有效地解释了其高效力。该模型由我们的支持体外实验并与之前的病毒细胞进入试验一致。我们的 TMPRSS2 药物结构已公开，以指导设计更有效和特异性的抑制剂。