Structural Biology of Nanobodies against the Spike Protein of SARS-CoV-2

Tang, Qilong; Owens, Raymond J.; Naismith, James H.

doi:10.3390/v13112214

Cited by 25 publications

(20 citation statements)

References 72 publications

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“…Recent research efforts in the design, engineering, and structure-functional characterization of nanobodies and their binding with SARS-CoV-2 S proteins reflect a growing realization that nanobody combinations could deliver a powerful array of neutralizing and escape mutation resistant molecular assemblies capable of rationally exploiting distinct binding epitopes and the intrinsic plasticity of the SARS-CoV-2 S protein. Structural aspects and classification of the nanobodies binding with the SARS-CoV-2 S were recently discussed in a review [42], highlighting several classes of high-affinity nanobodies An ultra-potent synthetic nanobody, Nb6, neutralizes SARS-CoV-2 by stabilizing the fully inactive down S conformation preventing binding with ACE2 receptor [43]. A high-affinity trivalent nanobody, mNb6-tri, can simultaneously bind to all three RBDs and inhibit the interactions with the host receptor by occupying the binding site and locking the S protein in the inactive state [43].…”

Section: Introductionmentioning

confidence: 99%

Allosteric Determinants of the SARS-CoV-2 Spike Protein Binding with Nanobodies: Examining Mechanisms of Mutational Escape and Sensitivity of the Omicron Variant

Verkhivker

2022

IJMS

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Structural and biochemical studies have recently revealed a range of rationally engineered nanobodies with efficient neutralizing capacity against the SARS-CoV-2 virus and resilience against mutational escape. In this study, we performed a comprehensive computational analysis of the SARS-CoV-2 spike trimer complexes with single nanobodies Nb6, VHH E, and complex with VHH E/VHH V nanobody combination. We combined coarse-grained and all-atom molecular simulations and collective dynamics analysis with binding free energy scanning, perturbation-response scanning, and network centrality analysis to examine mechanisms of nanobody-induced allosteric modulation and cooperativity in the SARS-CoV-2 spike trimer complexes with these nanobodies. By quantifying energetic and allosteric determinants of the SARS-CoV-2 spike protein binding with nanobodies, we also examined nanobody-induced modulation of escaping mutations and the effect of the Omicron variant on nanobody binding. The mutational scanning analysis supported the notion that E484A mutation can have a significant detrimental effect on nanobody binding and result in Omicron-induced escape from nanobody neutralization. Our findings showed that SARS-CoV-2 spike protein might exploit the plasticity of specific allosteric hotspots to generate escape mutants that alter response to binding without compromising activity. The network analysis supported these findings showing that VHH E/VHH V nanobody binding can induce long-range couplings between the cryptic binding epitope and ACE2-binding site through a broader ensemble of communication paths that is less dependent on specific mediating centers and therefore may be less sensitive to mutational perturbations of functional residues. The results suggest that binding affinity and long-range communications of the SARS-CoV-2 complexes with nanobodies can be determined by structurally stable regulatory centers and conformationally adaptable hotspots that are allosterically coupled and collectively control resilience to mutational escape.

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Section: Introductionmentioning

confidence: 99%

Allosteric Determinants of the SARS-CoV-2 Spike Protein Binding with Nanobodies: Examining Mechanisms of Mutational Escape and Sensitivity of the Omicron Variant

Verkhivker

2022

IJMS

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“…Nanobodies also facilitate protein crystallization and are widely used in structural biology. Structural aspects and classification of the nanobodies binding with the SARS-CoV-2 S were recently discussed in a review [81], highlighting several classes of high affinity nanobodies (Figure 1). Here, we review in detail some of the recent structural and biophysical studies of high affinity nanobody complexes with SARS-CoV-2 S protein with a focus on classification, characterization of binding epitopes, and binding affinities and specificities.…”

Section: Structural Biology Of the Sars-cov-2 S Protein Complexes Wit...mentioning

confidence: 99%

Section: Structural Biology Of the Sars-cov-2 S Protein Complexes Wit...mentioning

confidence: 99%

“…The binding interface for Sb68 is fairly small and structural analysis showed that Sb68 is not fully optimal for the binding cleft. The experimentally observed binding affinity of ~40 nM is lower than that of other nanobodies in this class [ 81 , 94 ]. A similar binding mode and interaction interface can be seen for VHH U nanobody in the complex with SARS-CoV-2 S protein [ 88 ].…”

Section: Exploring Computational Approaches For Mutational Profiling ...mentioning

confidence: 99%

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Structural and Computational Studies of the SARS-CoV-2 Spike Protein Binding Mechanisms with Nanobodies: From Structure and Dynamics to Avidity-Driven Nanobody Engineering

Verkhivker

2022

IJMS

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Nanobodies provide important advantages over traditional antibodies, including their smaller size and robust biochemical properties such as high thermal stability, high solubility, and the ability to be bioengineered into novel multivalent, multi-specific, and high-affinity molecules, making them a class of emerging powerful therapies against SARS-CoV-2. Recent research efforts on the design, protein engineering, and structure-functional characterization of nanobodies and their binding with SARS-CoV-2 S proteins reflected a growing realization that nanobody combinations can exploit distinct binding epitopes and leverage the intrinsic plasticity of the conformational landscape for the SARS-CoV-2 S protein to produce efficient neutralizing and mutation resistant characteristics. Structural and computational studies have also been instrumental in quantifying the structure, dynamics, and energetics of the SARS-CoV-2 spike protein binding with nanobodies. In this review, a comprehensive analysis of the current structural, biophysical, and computational biology investigations of SARS-CoV-2 S proteins and their complexes with distinct classes of nanobodies targeting different binding sites is presented. The analysis of computational studies is supplemented by an in-depth examination of mutational scanning simulations and identification of binding energy hotspots for distinct nanobody classes. The review is focused on the analysis of mechanisms underlying synergistic binding of multivalent nanobodies that can be superior to single nanobodies and conventional nanobody cocktails in combating escape mutations by effectively leveraging binding avidity and allosteric cooperativity. We discuss how structural insights and protein engineering approaches together with computational biology tools can aid in the rational design of synergistic combinations that exhibit superior binding and neutralization characteristics owing to avidity-mediated mechanisms.

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“…In addition, antibodies targeting NTD could also have neutralization efficacy [17,18]. The mechanism of anti-RBD neutralising antibodies could also be classified into two groups [19]. The first group of antibodies bind to RBD and blocked its interaction with ACE2 directly, thus protecting the host cells from virus infection [10,[20][21][22][23].…”

mentioning

confidence: 99%

A potent synthetic nanobody with broad-spectrum activity neutralizes SARS-Cov-2 virus and Omicron variant through a unique binding mode

Huang¹,

Tao

et al. 2022

Preprint

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COVID-19 continues to be a severe public health thread worldwide. The major challenge to control this pandemic is the rapid mutation rate of the SARS-Cov-2 virus, leading to the escape of the protection of vaccines and most of the neutralizing antibodies to date. Thus, it is pivotal to develop neutralizing antibodies with broad-spectrum activity targeting multiple SARS-Cov-2 variants. In this study, we first got a synthetic nanobody (named C5) which could interfere with ACE2 binding to SARS-Cov-2 spike protein and its RBD domain. The affinity matured format of C5 clone, named C5G2, has a single digit nanomolar affinity to RBD domain and inhibit its binding to ACE2 with an IC50 of 3.7 nM. Pseudovirus assay indicated that the monovalent C5G2 could protect the cells from the infection of SARS-Cov-2 wild type virus as well as most of the virus of concern, i.e. Alpha, Beta, Gamma and Omicron variants. Strikingly, C5G2 has the highest potency against omicron among all the variants with the IC50 of 4.9ng/ml (0.3 nM). We further solved the Cryo-EM structure of C5G2 in complex with the Spike trimer. The structure data showed that C5G2 bind to RBD mainly through its CDR3 at a vast region that not overlapping with the ACE2 binding surface. Surprisingly, C5G2 also bind to a distinct epitope residing in the NTD domain of spike protein through the same CDR3 loop, which may further increase its potency against the virus infection. Thus, this bi-paratopic nanobody may be served as an effective drug for the prophylaxis and therapy of the Omicron infection.

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Structural Biology of Nanobodies against the Spike Protein of SARS-CoV-2

Cited by 25 publications

References 72 publications

Allosteric Determinants of the SARS-CoV-2 Spike Protein Binding with Nanobodies: Examining Mechanisms of Mutational Escape and Sensitivity of the Omicron Variant

Allosteric Determinants of the SARS-CoV-2 Spike Protein Binding with Nanobodies: Examining Mechanisms of Mutational Escape and Sensitivity of the Omicron Variant

Structural and Computational Studies of the SARS-CoV-2 Spike Protein Binding Mechanisms with Nanobodies: From Structure and Dynamics to Avidity-Driven Nanobody Engineering

A potent synthetic nanobody with broad-spectrum activity neutralizes SARS-Cov-2 virus and Omicron variant through a unique binding mode

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