The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic continues to spread with devastating consequences. For passive immunization efforts, nanobodies have size and cost advantages over conventional antibodies. Here, we generated four neutralizing nanobodies that target the receptor-binding domain of the SARS-CoV-2 spike protein. We defined two distinct binding epitopes using x-ray crystallography and cryo-electron microscopy. Based on the structures, we engineered multivalent nanobodies with more than 100-fold improved neutralizing activity than monovalent nanobodies. Biparatopic nanobody fusions suppressed the emergence of escape mutants. Several nanobody constructs neutralized through receptor-binding competition, while other monovalent and biparatopic nanobodies triggered aberrant activation of the spike fusion machinery. These premature conformational changes in the spike protein forestalled productive fusion, and rendered the virions non-infectious.

中文翻译：

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结构引导的多价纳米抗体可阻断 SARS-CoV-2 感染并抑制突变逃逸


严重急性呼吸综合征冠状病毒 2 (SARS-CoV-2) 大流行继续蔓延，造成了毁灭性后果。对于被动免疫工作，纳米抗体比传统抗体具有尺寸和成本优势。在这里，我们生成了四种针对 SARS-CoV-2 刺突蛋白受体结合域的中和纳米抗体。我们使用 X 射线晶体学和冷冻电子显微镜定义了两个不同的结合表位。基于这些结构，我们设计了多价纳米抗体，其中和活性比单价纳米抗体提高了 100 倍以上。双互补位纳米抗体融合抑制了逃逸突变体的出现。几种纳米抗体构建体通过受体结合竞争而被中和，而其他单价和双互补位纳米抗体则引发刺突融合机制的异常激活。刺突蛋白中的这些过早构象变化阻止了有效的融合，并使病毒颗粒不具有感染性。