The COVID-19 pandemic has prompted a rapid response in vaccine and drug development. Herein, we modeled a complete membrane-embedded SARS-CoV-2 spike glycoprotein and used molecular dynamics simulations with benzene probes designed to enhance discovery of cryptic pockets. This approach recapitulated lipid and host metabolite binding sites previously characterized by cryo-electron microscopy, revealing likely ligand entry routes, and uncovered a novel cryptic pocket with promising druggable properties located underneath the 617–628 loop. A full representation of glycan moieties was essential to accurately describe pocket dynamics. A multi-conformational behavior of the 617–628 loop in simulations was validated using hydrogen-deuterium exchange mass spectrometry experiments, supportive of opening and closing dynamics. The pocket is the site of multiple mutations associated with increased transmissibility found in SARS-CoV-2 variants of concern including Omicron. Collectively, this work highlights the utility of the benzene mapping approach in uncovering potential druggable sites on the surface of SARS-CoV-2 targets.

COVID-19 大流行促使疫苗和药物开发迅速做出反应。在此，我们对完整的膜嵌入 SARS-CoV-2 刺突糖蛋白进行了建模，并使用苯探针进行分子动力学模拟，旨在增强对隐秘袋的发现。这种方法重现了之前通过冷冻电子显微镜表征的脂质和宿主代谢物结合位点，揭示了可能的配体进入途径，并发现了位于 617-628 环下方的具有有希望的可成药特性的新型神秘口袋。聚糖部分的完整表示对于准确描述口袋动力学至关重要。使用氢-氘交换质谱实验验证了模拟中 617-628 环的多构象行为，支持打开和关闭动力学。该口袋是多个突变位点，这些突变与 Omicron 等受关注的 SARS-CoV-2 变体中发现的传播性增加相关。总的来说，这项工作强调了苯作图方法在发现 SARS-CoV-2 靶标表面潜在可成药位点方面的效用。