scholarly journals Effects of common mutations in the SARS-CoV-2 Spike RBD and its ligand, the human ACE2 receptor on binding affinity and kinetics

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Michael I Barton ◽  
Stuart A MacGowan ◽  
Mikhail A Kutuzov ◽  
Omer Dushek ◽  
Geoffrey John Barton ◽  
...  
Keyword(s):  
Cell Surface ◽  
Binding Site ◽  
Receptor Binding ◽  
Immune Escape ◽  
Surface Protein ◽  
Spike Protein ◽  
Receptor Binding Domain ◽  
Cell Surface Protein ◽  
Kinetics Analysis ◽  
Protein Receptor

The interaction between the SARS-CoV-2 virus Spike protein receptor binding domain (RBD) and the ACE2 cell surface protein is required for viral infection of cells. Mutations in the RBD are present in SARS-CoV-2 variants of concern that have emerged independently worldwide. For example, the B.1.1.7 lineage has a mutation (N501Y) in its Spike RBD that enhances binding to ACE2. There are also ACE2 alleles in humans with mutations in the RBD binding site. Here we perform a detailed affinity and kinetics analysis of the effect of five common RBD mutations (K417N, K417T, N501Y, E484K, and S477N) and two common ACE2 mutations (S19P and K26R) on the RBD/ACE2 interaction. We analysed the effects of individual RBD mutations and combinations found in new SARS-CoV-2 Alpha (B.1.1.7), Beta (B.1.351), and Gamma (P1) variants. Most of these mutations increased the affinity of the RBD/ACE2 interaction. The exceptions were mutations K417N/T, which decreased the affinity. Taken together with other studies, our results suggest that the N501Y and S477N mutations enhance transmission primarily by enhancing binding, the K417N/T mutations facilitate immune escape, and the E484K mutation enhances binding and immune escape.

scholarly journals Effects of common mutations in the SARS-CoV-2 Spike RBD domain and its ligand the human ACE2 receptor on binding affinity and kinetics

2021 ◽  
Author(s):  
Michael I Barton ◽  
Stuart MacGowan ◽  
Mikhail A Kutuzov ◽  
Omer Dushek ◽  
Geoffrey J Barton ◽  
...  
Keyword(s):  
South Africa ◽  
Cell Surface ◽  
Immune Escape ◽  
Surface Protein ◽  
Spike Protein ◽  
Receptor Binding Domain ◽  
Cell Surface Protein ◽  
Kinetics Analysis ◽  
Protein Receptor ◽  
The Uk

The interaction between the SARS-CoV-2 virus Spike protein receptor binding domain (RBD) and the ACE2 cell surface protein is required for viral infection of cells. Mutations in the RBD domain are present in SARS-CoV-2 variants of concern that have emerged independently worldwide. For example, the more transmissible B.1.1.7 lineage has a mutation (N501Y) in its Spike RBD domain that enhances binding to ACE2. There are also ACE2 alleles in humans with mutations in the RBD binding site. Here we perform a detailed affinity and kinetics analysis of the effect of five common RBD mutations (K417N, K417T, N501Y, E484K and S477N) and two common ACE2 mutations (S19P and K26R) on the RBD/ACE2 interaction. We analysed the effects of individual RBD mutations, and combinations found in new SARS-CoV-2 variants first identified in the UK (B.1.1.7), South Africa (B.1.351) and Brazil (P1). Most of these mutations increased the affinity of the RBD/ACE2 interaction. The exceptions were mutations K417N/T, which decreased the affinity. Taken together with other studies, our results suggest that the N501Y and S477N mutations primarily enhance transmission, the K417N/T mutations facilitate immune escape, and the E484K mutation facilitates both transmission and immune escape.

scholarly journals Neutralization of recombinant RBD-subunit vaccine ZF2001-elicited antisera to SARS-CoV-2 variants including Delta

2021 ◽  
Author(s):  
Xin Zhao ◽  
Anqi Zheng ◽  
Dedong Li ◽  
Rong Zhang ◽  
Huan Sun ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Tandem Repeat ◽  
Dose Regimen ◽  
Subunit Vaccine ◽  
Immune Escape ◽  
Spike Protein ◽  
Receptor Binding Domain ◽  
Protein Receptor ◽  
A Subunit ◽  
Alpha Beta

SARS-CoV-2 variants brought new waves of infection worldwide. In particular, Delta variant (B.1.617.2 lineage) has become predominant in many countries. These variants raised the concern for their potential immune escape to the currently approved vaccines. ZF2001 is a subunit vaccine received emergency use authorization (EUA) in both China and Uzbekistan, with more than 100-million doses administrated with a three-dose regimen. The tandem-repeat dimer of SARS-CoV-2 spike protein receptor binding domain (RBD) was used as the antigen. In this work, we evaluated the neutralization of ZF2001-elicited antisera to SARS-CoV-2 variants including all four variants of concern (Alpha, Beta, Gamma and Delta) and other three variants of interest (Epsilon, Eta and Kappa) by pseudovirus-based assay. We found antisera preserved majority of the neutralizing activity against these variants. E484K/Q substitution is the key mutation to reduce the RBD-elicited sera neutralization. Moreover, ZF2001-elicited sera with a prolonged intervals between the second and third dose enhanced the neutralizing titers and resilience to SARS-CoV-2 variants.

scholarly journals Conjugation of Human β-Defensin 2 to Spike Protein Receptor-Binding Domain Induces Antigen-Specific Protective Immunity against Middle East Respiratory Syndrome Coronavirus Infection in Human Dipeptidyl Peptidase 4 Transgenic Mice

Vaccines ◽  
2020 ◽  
Vol 8 (4) ◽  
pp. 635
Author(s):  
Ju Kim ◽  
Ye Lin Yang ◽  
Yongsu Jeong ◽  
Yong-Suk Jang
Keyword(s):  
Middle East ◽  
Immune Responses ◽  
Receptor Binding ◽  
Dipeptidyl Peptidase ◽  
Binding Domain ◽  
Middle East Respiratory Syndrome ◽  
Spike Protein ◽  
Receptor Binding Domain ◽  
Dipeptidyl Peptidase 4 ◽  
Protein Receptor

Middle East respiratory syndrome coronavirus (MERS-CoV) causes severe acute respiratory symptoms. Due to the lack of medical countermeasures, effective and safe vaccines against MERS-CoV infection are urgently required. Although different types of candidate vaccines have been developed, their immunogenicity is limited, and the dose and administration route need optimization to achieve optimal protection. We here investigated the potential use of human β-defensin 2 (HBD 2) as an adjuvant to enhance the protection provided by MERS-CoV vaccination. We found that immunization of human dipeptidyl peptidase 4 (hDPP4)-transgenic (hDPP4-Tg) mice with spike protein receptor-binding domain (S RBD) conjugated with HBD 2 (S RBD-HBD 2) induced potent antigen (Ag)-specific adaptive immune responses and protected against MERS-CoV infection. In addition, immunization with S RBD-HBD 2 alleviated progressive pulmonary fibrosis in the lungs of MERS-CoV-infected hDPP4-Tg mice and suppressed endoplasmic reticulum stress signaling activation upon viral infection. Compared to intramuscular administration, intranasal administration of S RBD-HBD 2 induced more potent mucosal IgA responses and was more effective for protecting against intranasal MERS-CoV infection. In conclusion, our findings suggest that HBD 2 potentiates Ag-specific immune responses against viral Ag and can be used as an adjuvant enhancing the immunogenicity of subunit vaccine candidates against MERS-CoV.

scholarly journals Combining a Fusion Inhibitory Peptide Targeting the MERS-CoV S2 Protein HR1 Domain and a Neutralizing Antibody Specific for the S1 Protein Receptor-Binding Domain (RBD) Showed Potent Synergism against Pseudotyped MERS-CoV with or without Mutations in RBD

Viruses ◽  
2019 ◽  
Vol 11 (1) ◽  
pp. 31 ◽  
Author(s):  
Cong Wang ◽  
Chen Hua ◽  
Shuai Xia ◽  
Weihua Li ◽  
Lu Lu ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Neutralizing Antibody ◽  
Binding Domain ◽  
Spike Protein ◽  
Receptor Binding Domain ◽  
Inhibitory Peptide ◽  
Protein Receptor ◽  
Neutralizing Monoclonal Antibody ◽  
Peptide Targeting ◽  
Combinatorial Strategy

Middle East respiratory syndrome coronavirus (MERS-CoV) has continuously posed a threat to public health worldwide, yet no therapeutics or vaccines are currently available to prevent or treat MERS-CoV infection. We previously identified a fusion inhibitory peptide (HR2P-M2) targeting the MERS-CoV S2 protein HR1 domain and a highly potent neutralizing monoclonal antibody (m336) specific to the S1 spike protein receptor-binding domain (RBD). However, m336 was found to have reduced efficacy against MERS-CoV strains with mutations in RBD, and HR2P-M2 showed low potency, thus limiting the clinical application of each when administered separately. However, we herein report that the combination of m336 and HR2P-M2 exhibited potent synergism in inhibiting MERS-CoV S protein-mediated cell–cell fusion and infection by MERS-CoV pseudoviruses with or without mutations in the RBD, resulting in the enhancement of antiviral activity in contrast to either one administered alone. Thus, this combinatorial strategy could be used in clinics for the urgent treatment of MERS-CoV-infected patients.

scholarly journals The Cell Surface Protein Geneecm33+Is a Target of the Two Transcription Factors Atf1 and Mbx1 and Negatively Regulates Pmk1 MAPK Cell Integrity Signaling in Fission Yeast

2010 ◽  
Vol 21 (4) ◽  
pp. 674-685 ◽  
Author(s):  
Hirofumi Takada ◽  
Aiko Nishida ◽  
Mitsuhiro Domae ◽  
Ayako Kita ◽  
Yuki Yamano ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Cell Surface ◽  
Binding Site ◽  
Fission Yeast ◽  
Tandem Repeats ◽  
Surface Protein ◽  
Luciferase Reporter ◽  
Cell Surface Protein ◽  
Cell Integrity ◽  
Negative Feedback Regulation

The highly conserved fission yeast Pmk1 MAPK pathway plays a key role in cell integrity by regulating Atf1, which belongs to the ATF/cAMP-responsive element-binding (CREB) protein family. We identified and characterized ecm33+, which encodes a glycosyl-phosphatidylinositol (GPI)-anchored cell surface protein as a transcriptional target of Pmk1 and Atf1. We demonstrated that the gene expression of Ecm33 is regulated by two transcription factors Atf1 and a MADS-box-type transcription factor Mbx1. We identified a putative ATF/CREB-binding site and an RLM1-binding site in the ecm33+promoter region and monitored the transcriptional activity of Atf1 or Mbx1 in living cells using a destabilized luciferase reporter gene fused to three tandem repeats of the CRE and six tandem repeats of the Rlm1-binding sequence, respectively. These reporter genes reflect the activation of the Pmk1 pathway by various stimuli, thereby enabling the real-time monitoring of the Pmk1 cell integrity pathway. Notably, the Δecm33 cells displayed hyperactivation of the Pmk1 signaling together with hypersensitivity to Ca2+and an abnormal morphology, which were almost abolished by simultaneous deletion of the components of the Rho2/Pck2/Pmk1 pathway. Our results suggest that Ecm33 is involved in the negative feedback regulation of Pmk1 cell integrity signaling and is linked to cellular Ca2+signaling.

scholarly journals Can the SARS-CoV-2 Spike Protein Bind Integrins Independent of the RGD Sequence?

2021 ◽  
Vol 11 ◽  
Author(s):  
Christopher A. Beaudoin ◽  
Samir W. Hamaia ◽  
Christopher L.-H. Huang ◽  
Tom L. Blundell ◽  
Antony P. Jackson
Keyword(s):  
Receptor Binding ◽  
Conformational Changes ◽  
Sequence Similarity ◽  
Binding Domain ◽  
Spike Protein ◽  
Receptor Binding Domain ◽  
Binding Motifs ◽  
Rgd Motif ◽  
Rgd Sequence ◽  
Protein Receptor

The RGD motif in the Severe Acute Syndrome Coronavirus 2 (SARS-CoV-2) spike protein has been predicted to bind RGD-recognizing integrins. Recent studies have shown that the spike protein does, indeed, interact with αVβ3 and α5β1 integrins, both of which bind to RGD-containing ligands. However, computational studies have suggested that binding between the spike RGD motif and integrins is not favourable, even when unfolding occurs after conformational changes induced by binding to the canonical host entry receptor, angiotensin-converting enzyme 2 (ACE2). Furthermore, non-RGD-binding integrins, such as αx, have been suggested to interact with the SARS-CoV-2 spike protein. Other viral pathogens, such as rotaviruses, have been recorded to bind integrins in an RGD-independent manner to initiate host cell entry. Thus, in order to consider the potential for the SARS-CoV-2 spike protein to bind integrins independent of the RGD sequence, we investigate several factors related to the involvement of integrins in SARS-CoV-2 infection. First, we review changes in integrin expression during SARS-CoV-2 infection to identify which integrins might be of interest. Then, all known non-RGD integrin-binding motifs are collected and mapped to the spike protein receptor-binding domain and analyzed for their 3D availability. Several integrin-binding motifs are shown to exhibit high sequence similarity with solvent accessible regions of the spike receptor-binding domain. Comparisons of these motifs with other betacoronavirus spike proteins, such as SARS-CoV and RaTG13, reveal that some have recently evolved while others are more conserved throughout phylogenetically similar betacoronaviruses. Interestingly, all of the potential integrin-binding motifs, including the RGD sequence, are conserved in one of the known pangolin coronavirus strains. Of note, the most recently recorded mutations in the spike protein receptor-binding domain were found outside of the putative integrin-binding sequences, although several mutations formed inside and close to one motif, in particular, may potentially enhance binding. These data suggest that the SARS-CoV-2 spike protein may interact with integrins independent of the RGD sequence and may help further explain how SARS-CoV-2 and other viruses can evolve to bind to integrins.

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