Selection, biophysical and structural analysis of synthetic nanobodies that effectively neutralize SARS-CoV-2

• Published in: bioRxiv 2020-06
• Main Authors: Custodio, Tania F, Das, Hrishikesh, Sheward, Daniel J, Hanke, Leo, Pazicky, Samuel, Pieprzyk, Joanna, Sorgenfrei, Michele, Schroer, Martin, Gruzinov, Andrey, Jeffries, Cy, Graewert, Melissa, Svergun, Dmitri I, Dobrev, Nikolay, Remans, Kim, Seeger, Markus, Mcinerney, Gerald M, Murrell, Ben, B Martin Hällberg, Löw, Christian
• Format: Article
• Language: English
• Subjects: ACE2; Angiotensin-converting enzyme 2; Binding sites; Coronaviruses; COVID-19; Nanobodies; Pandemics; Severe acute respiratory syndrome coronavirus 2; Spike protein
• DOI: 10.1101/2020.06.23.165415

The coronavirus SARS-CoV-2 is the cause of the ongoing COVID-19 pandemic. Therapeutic neutralizing antibodies constitute a key short-to-medium term approach to tackle COVID-19. However, traditional antibody production is hampered by long development times and costly production. Here, we report the rapid isolation and characterization of nanobodies from a synthetic library, known as sybodies (Sb), that target the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein. Several binders with low nanomolar affinities and efficient neutralization activity were identified of which Sb23 displayed high affinity and neutralized pseudovirus with an IC50 of 0.6 ug/ml. A cryo-EM structure of the spike bound to Sb23 showed that Sb23 binds competitively in the ACE2 binding site. Furthermore, the cryo-EM reconstruction revealed a novel conformation of the spike where two RBDs are in the 'up' ACE2-binding conformation. The combined approach represents an alternative, fast workflow to select binders with neutralizing activity against newly emerging viruses. Competing Interest Statement The authors have declared no competing interest.