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Structure-functional studies have recently revealed a spectrum of diverse high-affinity nanobodies with efficient neutralizing capacity against SARS-CoV-2 virus and resilience against mutational escape. In this study, we combine atomistic simulations with the ensemble-based mutational profiling of binding for the SARS-CoV-2 S-RBD complexes with a wide range of nanobodies to identify dynamic and binding affinity fingerprints and characterize the energetic determinants of nanobody-escaping mutations. Using an in silico mutational profiling approach for probing the protein stability and binding, we examine dynamics and energetics of the SARS-CoV-2 complexes with single nanobodies Nb6 and Nb20, VHH E, a pair combination VHH E + U, a biparatopic nanobody VHH VE, and a combination of the CC12.3 antibody and VHH V/W nanobodies. This study characterizes the binding energy hotspots in the SARS-CoV-2 protein and complexes with nanobodies providing a quantitative analysis of the effects of circulating variants and escaping mutations on binding that is consistent with a broad range of biochemical experiments. The results suggest that mutational escape may be controlled through structurally adaptable binding hotspots in the receptor-accessible binding epitope that are dynamically coupled to the stability centers in the distant binding epitope targeted by VHH U/V/W nanobodies. This study offers a plausible mechanism in which through cooperative dynamic changes, nanobody combinations and biparatopic nanobodies can elicit the increased binding affinity response and yield resilience to common escape mutants.


This article was originally published in ACS Omega in 2021.

This scholarship is part of the Chapman University COVID-19 Archives.

ao1c03558_si_001.pdf (1004 kB)
Domains in the full-length SARS-CoV-2 S protein and a detailed structural organization of the S-RBD, cysteine residues that form disulfide linkages in the S-RBD protein, structural organization of cysteine clusters in the S2 subdomain of the SARS-CoV-2 spike prefusion structure, covariance matrices of residue fluctuations in the S-RBD complexes with nanobodies, and structural mapping of protein stability hotspots for the panel of studied nanobodies (PDF)

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This work is licensed under a Creative Commons Attribution-Noncommercial-No Derivative Works 4.0 License.



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