Receptor Binding Domain
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Michael J Peluso ◽  
Sadie E Munter ◽  
Kara L Lynch ◽  
Cassandra Yun ◽  
Leonel Torres ◽  

Abstract We report a patient with connective tissue disease who developed modest SARS-CoV-2 receptor binding domain-specific antibody levels and a lack of neutralization capacity, despite having received three mRNA COVID-19 vaccines and holding anti-B cell therapy for >7 months prior to vaccination. The patient developed virus-specific T cell responses.

2021 ◽  
Davida S Smyth ◽  
Monica Trujillo ◽  
Devon A Gregory ◽  
Kristen Cheung ◽  
Anna Gao ◽  

Tracking SARS-CoV-2 genetic diversity is strongly indicated because diversifying selection may lead to the emergence of novel variants resistant to naturally acquired or vaccine-induced immunity. To monitor New York City (NYC) for the presence of novel variants, we amplified regions of the SARS-CoV-2 Spike protein gene from RNA acquired from all 14 NYC wastewater treatment plants (WWTPs) and ascertained the diversity of lineages from these samples using high throughput sequencing. Here we report the detection and increasing frequencies of novel SARS-CoV-2 lineages not recognized in GISAIDs EpiCoV database. These lineages contain mutations rarely observed in clinical samples, including Q493K, Q498Y, H519N and T572N. Many of these mutations were found to expand the tropism of SARS-CoV-2 pseudoviruses by allowing infection of cells expressing the human, mouse, or rat ACE2 receptor. In addition, pseudoviruses containing the Spike amino acid sequence of these lineages were found to be resistant to many different classes of receptor binding domain (RBD) binding neutralizing monoclonal antibodies. We offer several hypotheses for the anomalous presence of these mutations, including the possibility of a non-human animal reservoir. Although wastewater sampling cannot provide direct inference of SARS-CoV-2 clinical sequences, our research revealed several lineages that could be relevant to public health and they would not have been discovered if not for wastewater surveillance.

2021 ◽  
Geert V.T. Roozen ◽  
Manon Prins ◽  
Rob van Binnendijk ◽  
Gerco den ◽  
Vincent Kuiper ◽  

Background There is an urgent need for fair and equitable access to safe and effective vaccines to end the COVID-19 pandemic. Shortages in reagents and vaccines are a major challenge, as well as limited knowledge on dose response relationship with mRNA COVID-19 vaccines. We explored intradermal fractional dose administration of a mRNA SARS-CoV-2/COVID-19 vaccine as a potential dose-sparing strategy. Methods We conducted a proof-of-concept, dose-escalation, open-label, randomised-controlled vaccine trial (IDSCOVA) in healthy adults aged 18-30 years. To test initial safety, ten participants received 10 μg mRNA-1273 vaccine through intradermal injection at day 1 and 29. Following a favourable safety review, thirty participants were 1:1 randomised to receive 20 μg mRNA-1273 either intradermally or intramuscularly. The primary endpoint was tolerability and safety. The secondary endpoint was seroconversion and specific IgG concentration against SARS-CoV-2 spike S1 and Receptor Binding Domain (RBD) after the second dose at day 43. We compared results to two historical cohorts of non-hospitalised COVID-19 patients and vaccinated individuals. Findings Thirty-eight of forty included participants (median age 25 years) completed the study. There were no serious adverse events. Self-reported local adverse reactions after intradermal delivery were mild, both in the 10 μg and the 20 μg group. In the higher dose group, systemic adverse reactions were more common, but still well tolerated. All 38 participants mounted substantially higher IgG-anti-S1 and IgG-anti-RBD concentrations at day 43 than COVID-19 controls. At day 43, anti-S1 (95% CI) was 1,696 (1,309-2,198) BAU/mL for the 10 μg intradermal group, 1,406 (953.5-2,074) BAU/mL for the 20 μg intramuscular group and 2,057 (1,421-2,975) BAU/mL for the 20 μg intradermal group. Anti-S1 was 107.2 (63-182.2) BAU/mL for the convalescent plasma control group and 1,558 (547.8-4,433) BAU/mL for the individuals vaccinated with 100 μg mRNA-1273. Interpretation Intradermal administration of 10 μg and 20 μg mRNA-1273 vaccine was well tolerated and safe, and resulted in a robust antibody response. Intradermal vaccination has the potential to be deployed for vaccine dose-sparing.

Vaccines ◽  
2021 ◽  
Vol 9 (8) ◽  
pp. 829
Junping Hong ◽  
Qian Wang ◽  
Qian Wu ◽  
Junyu Chen ◽  
Xijing Wang ◽  

To date, SARS-CoV-2 pandemic has caused more than 188 million infections and 4.06 million deaths worldwide. The receptor-binding domain (RBD) of the SARS-CoV-2 spike protein has been regarded as an important target for vaccine and therapeutics development because it plays a key role in binding the human cell receptor ACE2 that is required for viral entry. However, it is not easy to detect RBD in Western blot using polyclonal antibody, suggesting that RBD may form a complicated conformation under native condition and bear rare linear epitope. So far, no linear epitope on RBD is reported. Thus, a monoclonal antibody (mAb) that recognizes linear epitope on RBD will become valuable. In the present study, an RBD-specific rabbit antibody named 9E1 was isolated from peripheral blood mononuclear cells (PBMC) of immunized rabbit by RBD-specific single B cell sorting and mapped to a highly conserved linear epitope within twelve amino acids 480CNGVEGFNCYFP491 on RBD. 9E1 works well in Western blot on S protein and immunohistochemistry on the SARS-CoV-2 infected tissue sections. The results demonstrated that 9E1 can be used as a useful tool for pathological and functional studies of SARS-CoV-2.

2021 ◽  
Michelle Lubetzky ◽  
Ashely Sukhu ◽  
Zhen Zhao ◽  
Sophie Rand ◽  
Vijay Sharma ◽  

Abstract The response of the immune system to COVID-19 in end stage kidney disease patients who undergo kidney transplantation has yet to be described. We report data on 72 patients who underwent SARS-CoV-2 antibody testing both before and after kidney transplantation and were followed for a median of 186 days (range 83, 277). Of the 25 patients with a positive antibody test at the time of transplant, 17 (68%) remained positive after transplantation. Patients were significantly more likely to have a persistently positive test if they reported a symptomatic COVID-19 infection prior to transplant (p=0.01). SARS-CoV-2 IgG index values were measured in a subset of kidney transplant recipients and compared to wait -listed dialysis patients. These assays demonstrated a more significant decline in IgG (58% versus 14% p = 0.008) in transplant recipients when compared to dialysis patients tested during the same time period. Additional analysis of the quality of the immune response measuring the binding of SARS-CoV-2 antibodies to the receptor-binding domain (RBD binding), the antibody neutralizing capability, and the antibody avidity demonstrated a more pronounced effect when comparing pre-transplant values to post-induction therapy/post transplant values. The attenuated IgG response seen in transplant patients compared to dialysis patients after induction therapy requires further study. These data have important implications for post-transplant management of vaccinated dialysis patients.

2021 ◽  
Vol 8 ◽  
Yashpal S. Malik ◽  
Prashant Kumar ◽  
Mohd Ikram Ansari ◽  
Maged G. Hemida ◽  
Mohamed E. El Zowalaty ◽  

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) led to coronavirus disease 2019 (COVID-19) pandemic affecting nearly 71.2 million humans in more than 191 countries, with more than 1.6 million mortalities as of 12 December, 2020. The spike glycoprotein (S-protein), anchored onto the virus envelope, is the trimer of S-protein comprised of S1 and S2 domains which interacts with host cell receptors and facilitates virus-cell membrane fusion. The S1 domain comprises of a receptor binding domain (RBD) possessing an N-terminal domain and two subdomains (SD1 and SD2). Certain regions of S-protein of SARS-CoV-2 such as S2 domain and fragment of the RBD remain conserved despite the high selection pressure. These conserved regions of the S-protein are extrapolated as the potential target for developing molecular diagnostic techniques. Further, the S-protein acts as an antigenic target for different serological assay platforms for the diagnosis of COVID-19. Virus-specific IgM and IgG antibodies can be used to detect viral proteins in ELISA and lateral flow immunoassays. The S-protein of SARS-CoV-2 has very high sequence similarity to SARS-CoV-1, and the monoclonal antibodies (mAbs) against SARS-CoV-1 cross-react with S-protein of SARS-CoV-2 and neutralize its activity. Furthermore, in vitro studies have demonstrated that polyclonal antibodies targeted against the RBD of S-protein of SARS-CoV-1 can neutralize SARS-CoV-2 thus inhibiting its infectivity in permissive cell lines. Research on coronaviral S-proteins paves the way for the development of vaccines that may prevent SARS-CoV-2 infection and alleviate the current global coronavirus pandemic. However, specific neutralizing mAbs against SARS-CoV-2 are in clinical development. Therefore, neutralizing antibodies targeting SARS-CoV-2 S-protein are promising specific antiviral therapeutics for pre-and post-exposure prophylaxis and treatment of SARS-CoV-2 infection. We hereby review the approaches taken by researchers across the world to use spike gene and S-glycoprotein for the development of effective diagnostics, vaccines and therapeutics against SARA-CoV-2 infection the COVID-19 pandemic.

2021 ◽  
Vol 7 (1) ◽  
Yongbing Pan ◽  
Jianhui Du ◽  
Jia Liu ◽  
Hai Wu ◽  
Fang Gui ◽  

AbstractAs the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to threaten public health worldwide, the development of effective interventions is urgently needed. Neutralizing antibodies (nAbs) have great potential for the prevention and treatment of SARS-CoV-2 infection. In this study, ten nAbs were isolated from two phage-display immune libraries constructed from the pooled PBMCs of eight COVID-19 convalescent patients. Eight of them, consisting of heavy chains encoded by the immunoglobulin heavy-chain gene-variable region (IGHV)3-66 or IGHV3-53 genes, recognized the same epitope on the receptor-binding domain (RBD), while the remaining two bound to different epitopes. Among the ten antibodies, 2B11 exhibited the highest affinity and neutralization potency against the original wild-type (WT) SARS-CoV-2 virus (KD = 4.76 nM for the S1 protein, IC50 = 6 ng/mL for pseudoviruses, and IC50 = 1 ng/mL for authentic viruses), and potent neutralizing ability against B.1.1.7 pseudoviruses. Furthermore, 1E10, targeting a distinct epitope on RBD, exhibited different neutralization efficiency against WT SARS-CoV-2 and its variants B.1.1.7, B.1.351, and P.1. The crystal structure of the 2B11–RBD complexes revealed that the epitope of 2B11 highly overlaps with the ACE2-binding site. The in vivo experiment of 2B11 using AdV5-hACE2-transduced mice showed encouraging therapeutic and prophylactic efficacy against SARS-CoV-2. Taken together, our results suggest that the highly potent SARS-CoV-2-neutralizing antibody, 2B11, could be used against the WT SARS-CoV-2 and B.1.1.7 variant, or in combination with a different epitope-targeted neutralizing antibody, such as 1E10, against SARS-CoV-2 variants.

2021 ◽  
Vol 8 ◽  
Yaoqiang Du ◽  
Hao Wang ◽  
Linjie Chen ◽  
Quan Fang ◽  
Biqin Zhang ◽  

The emergence of novel coronavirus mutants is a main factor behind the deterioration of the epidemic situation. Further studies into the pathogenicity of these mutants are thus urgently needed. Binding of the spinous protein receptor binding domain (RBD) of SARS-CoV-2 to the angiotensin-converting enzyme 2 (ACE2) receptor was shown to initiate coronavirus entry into host cells and lead to their infection. The receptor-binding motif (RBM, 438–506) is a region that directly interacts with ACE2 receptor in the RBD and plays a crucial role in determining affinity. To unravel how mutations in the non-RBM regions impact the interaction between RBD and ACE2, we selected three non-RBM mutant systems (N354D, D364Y, and V367F) from the documented clinical cases, and the Q498A mutant system located in the RBM region served as the control. Molecular dynamics simulation was conducted on the mutant systems and the wild-type (WT) system, and verified experiments also performed. Non-RBM mutations have been shown not only to change conformation of the RBM region but also to significantly influence its hydrogen bonding and hydrophobic interactions. In particular, the D364Y and V367F systems showed a higher affinity for ACE2 owing to their electrostatic interactions and polar solvation energy changes. In addition, although the binding free energy at this point increased after the mutation of N354D, the conformation of the random coil (Pro384-Asp389) was looser than that of other systems, and the combined effect weakened the binding free energy between RBD and ACE2. Interestingly, we also found a random coil (Ala475-Gly485). This random coil is very sensitive to mutations, and both types of mutations increase the binding free energy of residues in this region. We found that the binding loop (Tyr495-Tyr505) in the RBD domain strongly binds to Lys353, an important residue of the ACE2 domain previously identified. The binding free energy of the non-RBM mutant group at the binding loop had positive and negative changes, and these changes were more obvious than that of the Q498A system. The results of this study elucidate the effect of non-RBM mutation on ACE2-RBD binding, and provide new insights for SARS-CoV-2 mutation research.

mBio ◽  
2021 ◽  
Fatima Amanat ◽  
Shirin Strohmeier ◽  
Wen-Hsin Lee ◽  
Sandhya Bangaru ◽  
Andrew B. Ward ◽  

Cross-neutralization of SARS-CoV-2 variants by RBD-targeting antibodies is still not well understood, and very little is known about the potential protective effect of nonneutralizing antibodies in vivo . Using a panel of mouse monoclonal antibodies, we investigate both of these points.

2021 ◽  
Jin-Kui Yang ◽  
Miao-Miao Zhao ◽  
Yun Zhu ◽  
Li Zhang ◽  
Gongxun Zhong ◽  

Abstract The spike (S) protein of SARS coronavirus 2 (SARS-CoV-2) is an ideal target for the development of specific vaccines or drugs. However, treatments targeting viruses with mutant S proteins that have recently emerged in many countries are limited. Cleavage of the S protein by host proteases is essential for viral infection. Here, we discovered two novel sites (CS-1 and CS-2) in the S protein for cleavage by the protease Cathepsin L (CTSL). Both sites are highly conserved among all SARS-CoV-2 variants of concern. Cryo-electron microscopy structural studies revealed that CTSL cleavage increases the dynamics of the receptor binding domain of S and induces novel conformations. In our pseudovirus (PsV) infection experiment, alteration of the cleavage site significantly reduced the infection efficiency, and CTSL inhibitors markedly inhibited infection with PsVs of both the wild-type and emerged SARS-CoV-2 variants. Furthermore, six highly efficient CTSL inhibitors were found to effectively inhibit live virus infection in human cells in vitro, and two of these were further confirmed to prevent live virus infection in human ACE2 transgenic mice in vivo. Our work suggested that the CTSL cleavage sites in SARS-CoV-2 S are emerging new but effective targets for the development of mutation-resistant vaccines and drugs.

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