Combating the worldwide spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and the emergence of new variants demands understanding of the structural basis of the interaction of antibodies with the SARS-CoV-2 receptor-binding domain (RBD). Here, we report five X-ray crystal structures of sybodies (synthetic nanobodies) including those of binary and ternary complexes of Sb16-RBD, Sb45-RBD, Sb14-RBD-Sb68, and Sb45-RBD-Sb68, as well as unliganded Sb16. These structures reveal that Sb14, Sb16, and Sb45 bind the RBD at the angiotensin-converting enzyme 2 interface and that the Sb16 interaction is accompanied by a large conformational adjustment of complementarity-determining region 2. In contrast, Sb68 interacts at the periphery of the SARS-CoV-2 RBD-angiotensin-converting enzyme 2 interface. We also determined cryo-EM structures of Sb45 bound to the SARS-CoV-2 spike protein. Superposition of the X-ray structures of sybodies onto the trimeric spike protein cryo-EM map indicates that some sybodies may bind in both "up" and "down" configurations, but others may not. Differences in sybody recognition of several recently identified RBD variants are explained by these structures.

中文翻译：

---

合成纳米抗体-SARS-CoV-2受体结合域复合物的结构揭示了不同的相互作用位点。

抗击严重急性呼吸综合征冠状病毒 2 (SARS-CoV-2) 的全球传播和新变种的出现，需要了解抗体与 SARS-CoV-2 受体结合域 (RBD) 相互作用的结构基础。在这里，我们报告了 sybodies（合成纳米抗体）的五种 X 射线晶体结构，包括 Sb16-RBD、Sb45-RBD、Sb14-RBD-Sb68 和 Sb45-RBD-Sb68 的二元和三元复合物以及未配位的 Sb16 。这些结构表明，Sb14、Sb16 和 Sb45 在血管紧张素转换酶 2 界面处与 RBD 结合，并且 Sb16 相互作用伴随着互补决定区 2 的大幅构象调整。相反，Sb68 在SARS-CoV-2 RBD-血管紧张素转换酶 2 界面。我们还确定了与 SARS-CoV-2 刺突蛋白结合的 Sb45 的冷冻电镜结构。sybodies 的 X 射线结构在三聚刺突蛋白冷冻电镜图上的叠加表明，一些 sybodies 可能以“向上”和“向下”构型结合，但其他可能不会。这些结构解释了最近发现的几个 RBD 变体的 sybody 识别差异。