Max Planck Institute for Biophysical Chemistry, Department of Molecular Biology, Am Fassberg 11, Göttingen, 37077, Germany.
Department of Cellular Biochemistry, University Medical Center Göttingen, Humboldtallee 23, Göttingen, 37073, Germany.
Nat Commun. 2021 Jan 12;12(1):279. doi: 10.1038/s41467-020-20542-0.
Remdesivir is the only FDA-approved drug for the treatment of COVID-19 patients. The active form of remdesivir acts as a nucleoside analog and inhibits the RNA-dependent RNA polymerase (RdRp) of coronaviruses including SARS-CoV-2. Remdesivir is incorporated by the RdRp into the growing RNA product and allows for addition of three more nucleotides before RNA synthesis stalls. Here we use synthetic RNA chemistry, biochemistry and cryo-electron microscopy to establish the molecular mechanism of remdesivir-induced RdRp stalling. We show that addition of the fourth nucleotide following remdesivir incorporation into the RNA product is impaired by a barrier to further RNA translocation. This translocation barrier causes retention of the RNA 3'-nucleotide in the substrate-binding site of the RdRp and interferes with entry of the next nucleoside triphosphate, thereby stalling RdRp. In the structure of the remdesivir-stalled state, the 3'-nucleotide of the RNA product is matched and located with the template base in the active center, and this may impair proofreading by the viral 3'-exonuclease. These mechanistic insights should facilitate the quest for improved antivirals that target coronavirus replication.
瑞德西韦是唯一获得 FDA 批准用于治疗 COVID-19 患者的药物。瑞德西韦的活性形式作为核苷类似物，可抑制包括 SARS-CoV-2 在内的冠状病毒的 RNA 依赖性 RNA 聚合酶（RdRp）。瑞德西韦被 RdRp 掺入到正在生长的 RNA 产物中，并允许在 RNA 合成暂停之前再添加三个核苷酸。在这里，我们使用合成 RNA 化学、生物化学和低温电子显微镜来建立瑞德西韦诱导的 RdRp 停滞的分子机制。我们表明，在 RNA 产物中掺入瑞德西韦后，添加第四个核苷酸受到进一步 RNA 易位的障碍的阻碍。这种易位障碍导致 RNA 3'-核苷酸在 RdRp 的底物结合位点中保留，并干扰下一个核苷三磷酸的进入，从而使 RdRp 停滞。在瑞德西韦停滞状态的结构中，RNA 产物的 3'-核苷酸与活性中心的模板碱基匹配和定位，这可能会损害病毒 3'-核酸外切酶的校对。这些机制上的见解应该有助于寻找针对冠状病毒复制的改进型抗病毒药物。
