receptor binding domain
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Viruses ◽  
2022 ◽  
Vol 14 (1) ◽  
pp. 144
Author(s):  
Debashree Chatterjee ◽  
Alexandra Tauzin ◽  
Annemarie Laumaea ◽  
Shang Yu Gong ◽  
Yuxia Bo ◽  
...  

The rapid emergence of SARS-CoV-2 variants is fueling the recent waves of the COVID-19 pandemic. Here, we assessed ACE2 binding and antigenicity of Mu (B.1.621) and A.2.5 Spikes. Both these variants carry some mutations shared by other emerging variants. Some of the pivotal mutations such as N501Y and E484K in the receptor-binding domain (RBD) detected in B.1.1.7 (Alpha), B.1.351 (Beta) and P.1 (Gamma) are now present within the Mu variant. Similarly, the L452R mutation of B.1.617.2 (Delta) variant is present in A.2.5. In this study, we observed that these Spike variants bound better to the ACE2 receptor in a temperature-dependent manner. Pseudoviral particles bearing the Spike of Mu were similarly neutralized by plasma from vaccinated individuals than those carrying the Beta (B.1.351) and Delta (B.1.617.2) Spikes. Altogether, our results indicate the importance of measuring critical parameters such as ACE2 interaction, plasma recognition and neutralization ability of each emerging variant.


2022 ◽  
Author(s):  
Arinjay Banerjee ◽  
Jocelyne Lew ◽  
Andrea Kroeker ◽  
Kaushal Baid ◽  
Patryk Aftanas ◽  
...  

The omicron variant of concern (VOC) of SARS-CoV-2 was first reported in November 2021 in Botswana and South Africa. Omicron variant has evolved multiple mutations within the spike protein and the receptor binding domain (RBD), raising concerns of increased antibody evasion. Here, we isolated infectious omicron from a clinical specimen obtained in Canada. The neutralizing activity of sera from 65 coronavirus disease (COVID-19) vaccine recipients and convalescent individuals against clinical isolates of ancestral SARS-CoV-2, beta, delta, and omicron VOCs was assessed. Convalescent sera from unvaccinated individuals infected by the ancestral virus during the first wave of COVID-19 in Canada (July, 2020) demonstrated reduced neutralization against beta, delta and omicron VOCs. Convalescent sera from unvaccinated individuals infected by the delta variant (May-June, 2021) neutralized omicron to significantly lower levels compared to the delta variant. Sera from individuals that received three doses of the Pfizer or Moderna vaccines demonstrated reduced neutralization of both delta and omicron variants relative to ancestral SARS-CoV-2. Sera from individuals that were naturally infected with ancestral SARS-CoV-2 and subsequently received two doses of the Pfizer vaccine induced significantly higher neutralizing antibody levels against ancestral virus and all VOCs. Importantly, infection alone, either with ancestral SARS-CoV-2 or the delta variant was not sufficient to induce high neutralizing antibody titers against omicron. This data will inform current booster vaccination strategies and we highlight the need for additional studies to identify longevity of immunity against SARS-CoV-2 and optimal neutralizing antibody levels that are necessary to prevent infection and/or severe COVID-19.


2022 ◽  
Vol 12 (1) ◽  
Author(s):  
◽  
Paula M. Berguer ◽  
Matías Blaustein ◽  
Luis M. Bredeston ◽  
Patricio O. Craig ◽  
...  

AbstractThe receptor binding domain (RBD) of the Spike protein from SARS-CoV-2 is a promising candidate to develop effective COVID-19 vaccines since it can induce potent neutralizing antibodies. We have previously reported the highly efficient production of RBD in Pichia pastoris, which is structurally similar to the same protein produced in mammalian HEK-293T cells. In this work we designed an RBD multimer with the purpose of increasing its immunogenicity. We produced multimeric particles by a transpeptidation reaction between RBD expressed in P. pastoris and Lumazine Synthase from Brucella abortus (BLS), which is a highly immunogenic and very stable decameric 170 kDa protein. Such particles were used to vaccinate mice with two doses 30 days apart. When the particles ratio of RBD to BLS units was high (6–7 RBD molecules per BLS decamer in average), the humoral immune response was significantly higher than that elicited by RBD alone or by RBD-BLS particles with a lower RBD to BLS ratio (1–2 RBD molecules per BLS decamer). Remarkably, multimeric particles with a high number of RBD copies elicited a high titer of neutralizing IgGs. These results indicate that multimeric particles composed of RBD covalent coupled to BLS possess an advantageous architecture for antigen presentation to the immune system, and therefore enhancing RBD immunogenicity. Thus, multimeric RBD-BLS particles are promising candidates for a protein-based vaccine.


2022 ◽  
Vol 119 (4) ◽  
pp. e2113118119
Author(s):  
Juan Rodriguez-Rivas ◽  
Giancarlo Croce ◽  
Maureen Muscat ◽  
Martin Weigt

The emergence of new variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a major concern given their potential impact on the transmissibility and pathogenicity of the virus as well as the efficacy of therapeutic interventions. Here, we predict the mutability of all positions in SARS-CoV-2 protein domains to forecast the appearance of unseen variants. Using sequence data from other coronaviruses, preexisting to SARS-CoV-2, we build statistical models that not only capture amino acid conservation but also more complex patterns resulting from epistasis. We show that these models are notably superior to conservation profiles in estimating the already observable SARS-CoV-2 variability. In the receptor binding domain of the spike protein, we observe that the predicted mutability correlates well with experimental measures of protein stability and that both are reliable mutability predictors (receiver operating characteristic areas under the curve ∼0.8). Most interestingly, we observe an increasing agreement between our model and the observed variability as more data become available over time, proving the anticipatory capacity of our model. When combined with data concerning the immune response, our approach identifies positions where current variants of concern are highly overrepresented. These results could assist studies on viral evolution and future viral outbreaks and, in particular, guide the exploration and anticipation of potentially harmful future SARS-CoV-2 variants.


2022 ◽  
Author(s):  
Jun Zhang ◽  
Yongfei Cai ◽  
Christy Lavine ◽  
Hanqin Peng ◽  
Haisun Zhu ◽  
...  

The Omicron variant of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), bearing an unusually high number of mutations, has become a dominant strain in many countries within several weeks. We report here structural, functional and antigenic properties of its full-length spike (S) protein with a native sequence in comparison with those of previously prevalent variants. Omicron S requires a substantially higher level of host receptor ACE2 for efficient membrane fusion than other variants, possibly explaining its unexpected cellular tropism. Mutations not only remodel the antigenic structure of the N-terminal domain of the S protein, but also alter the surface of the receptor-binding domain in a way not seen in other variants, consistent with its remarkable resistance to neutralizing antibodies. These results suggest that Omicron S has acquired an extraordinary ability to evade host immunity by excessive mutations, which also compromise its fusogenic capability.


2022 ◽  
Author(s):  
Jesse Wilson ◽  
Aivett Bilbao ◽  
Juan Wang ◽  
Yen-Chen Liao ◽  
Dusan Velickovic ◽  
...  

SARS-CoV-2 cellular infection is mediated by the heavily glycosylated spike protein. Recombinant versions of the spike protein and the receptor binding domain (RBD) are necessary for seropositivity assays and can potentially serve as vaccines against viral infection. RBD plays key roles in the spike protein’s structure and function, and thus comprehensive characterization of recombinant RBD is critically important for biopharmaceutical applications. Liquid-chromatography coupled to mass spectrometry (LCMS) has been widely used to characterize post-translational modifications in proteins including glycosylation. Most studies of RBDs were performed at the proteolytic peptide (bottom-up proteomics) or released glycan level because of the technical challenges in resolving highly heterogenous glycans at the intact protein level. Herein, we evaluated several online separation techniques: 1. C2 reverse-phase liquid chromatography (RPLC), 2. capillary zone electrophoresis (CZE), and 3. acrylamide-based monolithic hydrophilic interaction chromatography (HILIC) to separate intact recombinant RBDs with varying combinations of glycosylations (glycoforms) for top-down MS. Within the conditions we explored, the HILIC method was superior to RPLC and CZE at separating RBD glycoforms, which differ significantly in neutral glycan groups. In addition, our top-down analysis readily captured unexpected modifications (e.g., cysteinylation, N-terminal sequence variation) and low abundance, heavily glycosylated proteoforms that may be missed by using glycopeptide data alone. The HILIC top-down MS platform holds great potential in resolving heterogenous glycoproteins for facile comparison of biosimilars in quality control applications.


Vaccines ◽  
2022 ◽  
Vol 10 (1) ◽  
pp. 96
Author(s):  
Iuliia A. Merkuleva ◽  
Dmitry N. Shcherbakov ◽  
Mariya B. Borgoyakova ◽  
Daniil V. Shanshin ◽  
Andrey P. Rudometov ◽  
...  

The receptor-binding domain (RBD) of the protein S SARS-CoV-2 is considered to be one of the appealing targets for developing a vaccine against COVID-19. The choice of an expression system is essential when developing subunit vaccines, as it ensures the effective synthesis of the correctly folded target protein, and maintains its antigenic and immunogenic properties. Here, we describe the production of a recombinant RBD protein using prokaryotic (pRBD) and mammalian (mRBD) expression systems, and compare the immunogenicity of prokaryotic and mammalian-expressed RBD using a BALB/c mice model. An analysis of the sera from mice immunized with both variants of the protein revealed that the mRBD expressed in CHO cells provides a significantly stronger humoral immune response compared with the RBD expressed in E.coli cells. A specific antibody titer of sera from mice immunized with mRBD was ten-fold higher than the sera from the mice that received pRBD in ELISA, and about 100-fold higher in a neutralization test. The data obtained suggests that mRBD is capable of inducing neutralizing antibodies against SARS-CoV-2.


2022 ◽  
Vol 23 (1) ◽  
Author(s):  
Shijulal Nelson-Sathi ◽  
P. K. Umasankar ◽  
E. Sreekumar ◽  
R. Radhakrishnan Nair ◽  
Iype Joseph ◽  
...  

Abstract Background SARS-CoV-2, the causative agent of COVID-19 pandemic is a RNA virus prone to mutations. Formation of a stable binding interface between the Receptor Binding Domain (RBD) of SARS-CoV-2 Spike (S) protein and Angiotensin-Converting Enzyme 2 (ACE2) of host is pivotal for viral entry. RBD has been shown to mutate frequently during pandemic. Although, a few mutations in RBD exhibit enhanced transmission rates leading to rise of new variants of concern, most RBD mutations show sustained ACE2 binding and virus infectivity. Yet, how all these mutations make the binding interface constantly favourable for virus remain enigmatic. This study aims to delineate molecular rearrangements in the binding interface of SARS-CoV-2 RBD mutants. Results Here, we have generated a mutational and structural landscape of SARS-CoV-2 RBD in first six months of the pandemic. We analyzed 31,403 SARS-CoV-2 genomes randomly across the globe, and identified 444 non-synonymous mutations in RBD that cause 49 distinct amino acid substitutions in contact and non-contact amino acid residues. Molecular phylogenetic analysis suggested independent emergence of RBD mutants. Structural mapping of these mutations on the SARS-CoV-2 Wuhan reference strain RBD and structural comparison with RBDs from bat-CoV, SARS-CoV, and pangolin-CoV, all bound to human or mouse ACE2, revealed several changes in the interfacial interactions in all three binding clusters. Interestingly, interactions mediated via N487 residue in cluster-I and Y449, G496, T500, G502 residues in cluster-III remained largely unchanged in all RBD mutants. Further analysis showed that these interactions are evolutionarily conserved in sarbecoviruses which use ACE2 for entry. Importantly, despite extensive changes in the interface, RBD-ACE2 stability and binding affinities were maintained in all the analyzed mutants. Taken together, these findings reveal how SARS-CoV-2 uses its RBD residues to constantly remodel the binding interface. Conclusion Our study broadly signifies understanding virus-host binding interfaces and their alterations during pandemic. Our findings propose a possible interface remodelling mechanism used by SARS-CoV-2 to escape deleterious mutations. Future investigations will focus on functional validation of in-silico findings and on investigating interface remodelling mechanisms across sarbecoviruses. Thus, in long run, this study may provide novel clues to therapeutically target RBD-ACE2 interface for pan-sarbecovirus infections.


2022 ◽  
Author(s):  
Camryn Carter ◽  
Justin Airas ◽  
Carol A. Parish

SARS-CoV-2 is a coronavirus that has created a global pandemic. The virus contains a spike protein which has been shown to bind to the ACE2 receptor on the surface of human cells. Vaccines have been developed that recognize elements of the SARS-CoV-2 spike protein and they have been successful in preventing infection. Recently, the omicron variant of the SARS-CoV-2 virus was reported and quickly became a variant of concern due to its transmissibility. This variant contained an unusually large number (32) of point mutations, of which 15 of those mutations are in the receptor binding domain of the spike protein. In order to assess the differential binding ability of the wild type and omicron variant of the RBD spike protein to human ACE2 receptors, we conducted 2 μs of molecular dynamics simulation to estimate the binding affinities and behaviors. Based upon MM-GBSA binding affinity, center of mass distance measurements, ensemble clustering, pairwise residue decomposition and hydrogen bonding analysis, we can conclude that the 15 point mutations in the receptor binding domain do not increase the affinity of the spike protein for the human ACE2 receptor. The MM-GBSA binding estimations over a 2 μs trajectory, suggest that the wild type binds to ACE2 with a value of -29.69 kcal/mol while the omicron mutant binds with an energy value of -26.67 kcal/mol. These values are within the error estimates of the MM-GBSA method. While some mutations increase binding, more mutations diminish binding, leading to an overall similar picture of binding.


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