An ultrapotent synthetic nanobody neutralizes SARS-CoV-2 by stabilizing inactive Spike.

Michael Schoof ,Melody G Campbell ,Li Yang ,Sergei Pourmal ,Oren S Rosenberg ,Henry C Nguyen ,Reuben A Saunders ,Caleigh M Azumaya ,Axel F Brilot ,Ming Sun ,Yifan Cheng ,Sayan Gupta ,Jiahao Liang ,Nick Hoppe ,Peter Walter ,Camille R Simoneau ,Feng Wang ,Kris M White ,Alisa Bowen ,Niv Dobzinski ,Liam Mckay ,Kaihua Zhang ,Arthur A Melo ,Almarie Joves ,Arceli Joves ,Raphael Trenker ,Eric Tse ,Amber M Smith ,Alexandrea N Rizo ,Mali Safari ,Danielle L Swaney ,Loan Doan ,Mariano Tabios ,Jessica K Peters ,Kyle E Lopez ,Daniel Asarnow ,Joana Paulino ,Kliment A Verba ,Yen-Li Li ,Tristan W Owens ,Vladislav Belyy ,Fĕi Li ,Amy Diallo ,Megan Lo ,Benjamin Barsi‐Rhyne ,Jenny Chen ,Kate Kim ,Andrew W Barile-Hill ,Kaitlin Schaefer ,Sebastián Flores ,Nevan J Krogan ,Zanlin Yu ,Ishan Deshpande ,Junrui Li ,Morgane Boone ,Gregory E Merz ,J.t Biel ,Stephanie A Wankowicz ,Phuong Nguyen ,Veronica V Rezelj ,Fengbo Zhou ,Smriti Sangwan ,Daniel R Southworth ,Yanxin Liu ,Ursula Schulze‐Gahmen ,Corie Y Ralston ,Tanja Kortemme ,Aashish Manglik ,Aditya Anand ,Carlos Nowotny ,Aye C Thwin ,Liu Xi ,Fei Li ,Michael C Thompson ,Jianhua Zhao ,Tsz Kin Martin Tsui ,Adolfo Garcı́a-Sastre ,Frank R Moss ,Beth Shoshana Zha ,Julian R Braxton ,Yang Zhang ,Miles Sasha Dickinson ,Devan Diwanji ,Cristina Puchades ,David Bulkley ,Adam Frost ,Melanie Ott ,Natalia Jura ,Yuwei Liu ,Un Seng Chio ,Victor Lam ,Christian B Billesbølle ,M Zimanyi ,Natalie Whitis ,Silke Nock ,James S Fraser ,Iris D Young ,Marco Vignuzzi ,Meghna Gupta ,Huong T Kratochvil ,Bryan Faust ,Donovan Trinidad ,Michelle Moritz ,Cynthia M Chio ,Paul Thomas ,Erron W Titus ,Nadia Herrera ,Kristoffer E Leon ,David A Agard ,Thomas H Pospiech ,Mingliang Jin ,Robert M Stroud

doi:10.1126/science.abe3255

Abstract

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus enters host cells via an interaction between its Spike protein and the host cell receptor angiotensin-converting enzyme 2 (ACE2). By screening a yeast surface-displayed library of synthetic nanobody sequences, we developed nanobodies that disrupt the interaction between Spike and ACE2. Cryo-electron microscopy (cryo-EM) revealed that one nanobody, Nb6, binds Spike in a fully inactive conformation with its receptor binding domains locked into their inaccessible down state, incapable of binding ACE2. Affinity maturation and structure-guided design of multivalency yielded a trivalent nanobody, mNb6-tri, with femtomolar affinity for Spike and picomolar neutralization of SARS-CoV-2 infection. mNb6-tri retains function after aerosolization, lyophilization, and heat treatment, which enables aerosol-mediated delivery of this potent neutralizer directly to the airway epithelia.

Highlights

Coronavirus virions are bounded by a membrane that contains the homotrimeric transmembrane glycoprotein Spike responsible for virus entry into the host cell [8, 9]
Contained within S1 is the receptor binding domain (RBD), which directly binds to angiotensin converting enzyme 2 (ACE2), and the N terminal domain (NTD)
A prototypical example of this class is nanobody Nb6, which binds to SpikeS2P and to RBD alone with a KD of 210nM and 41nM, respectively (Fig. 1C and table S1)

Summary

Introduction

Coronavirus virions are bounded by a membrane that contains the homotrimeric transmembrane glycoprotein Spike responsible for virus entry into the host cell [8, 9]. A prototypical example of this class is nanobody Nb6, which binds to SpikeS2P and to RBD alone with a KD of 210nM and 41nM, respectively (Fig. 1C and table S1). Class II, exemplified by nanobody Nb3, binds to SpikeS2P (KD=61nM), but displays no binding to RBD alone (Fig. 1C and table S1).

Results

Conclusion