scholarly journals Identification of Two Novel Linear Neutralizing Epitopes within the Hexon Protein of Canine Adenovirus Using Monoclonal Antibodies

Vaccines ◽  
2021 ◽  
Vol 9 (2) ◽  
pp. 135
Author(s):  
Shujie Wang ◽  
Chunsheng Wang ◽  
Xiao Ren ◽  
Wenjiao Xue ◽  
Haijuan He ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Neutralizing Antibodies ◽  
3D Structure ◽  
Surface Region ◽  
Neutralization Titer ◽  
Random Peptide ◽  
Hexon Protein ◽  
Potential Applications ◽  
Alignment Analysis ◽  
Neutralizing Epitopes

Canine adenovirus (CAdV) has a high prevalence in canine populations. High affinity neutralizing antibodies against conserved epitopes can provide protective immunity against CAdV and protect against future outbreaks. In this study, we identified two CAdV-2-specific neutralizing monoclonal antibodies (mAbs), 2C1 and 7D7, which recognized two linear-dependent epitopes. MAb 2C1 potently neutralized CAdV-2 with a 50% neutralization titer (NT50) of 4096, and mAb 7D7 partially neutralized CAdV-2 with a 50% NT50 of 64. Immunoprecipitation, Western blot and protein spectral analysis indicated that both neutralizing mAbs recognized the hexon protein (Hex) of CAdV-2. Through a 12-mer random peptide phage display and synthetic peptides analysis, we finely mapped the neutralizing epitopes to two 10-amino acid (aa) peptides within the CAdV Hex: 634RIKQRETPAL643 located on the surface region; and 736PESYKDRMYS745 located in the inner region of the expected 3D structure of trimeric Hex. Importantly, the two epitopes are highly conserved among all CAdV isolates by sequence alignment analysis. Thus, these results provide insights into the interaction between virus and mAbs at the aa level and may have potential applications in the development of novel therapeutic or epitope-based vaccines, antibody therapeutics and a diagnostic method suitable for the rapid detection of all CAdVs.

scholarly journals Development of Neutralizing Monoclonal Antibodies for Oncogenic Human Papillomavirus Types 31, 33, 45, 52, and 58

2014 ◽  
Vol 21 (4) ◽  
pp. 587-593 ◽  
Author(s):  
Martha J. Brown ◽  
Hanna Seitz ◽  
Victoria Towne ◽  
Martin Müller ◽  
Adam C. Finnefrock
Keyword(s):  
Monoclonal Antibodies ◽  
Human Papillomavirus ◽  
Immune Responses ◽  
Neutralizing Antibodies ◽  
Cervical Cancers ◽  
Hpv Types ◽  
Oncogenic Hpv ◽  
Neutralizing Epitopes ◽  
Selection Of

ABSTRACTHuman papillomavirus (HPV) is the etiological agent for all cervical cancers, a significant number of other anogenital cancers, and a growing number of head and neck cancers. Two licensed vaccines offer protection against the most prevalent oncogenic types, 16 and 18, responsible for approximately 70% of cervical cancer cases worldwide and one of these also offers protection against types 6 and 11, responsible for 90% of genital warts. The vaccines are comprised of recombinantly expressed major capsid proteins that self-assemble into virus-like particles (VLPs) and prevent infection by eliciting neutralizing antibodies. Adding the other frequently identified oncogenic types 31, 33, 45, 52, and 58 to a vaccine would increase the coverage against HPV-induced cancers to approximately 90%. We describe the generation and characterization of panels of monoclonal antibodies to these five additional oncogenic HPV types, and the selection of antibody pairs that were high affinity and type specific and recognized conformation-dependent neutralizing epitopes. Such characteristics make these antibodies useful tools for monitoring the production and potency of a prototype vaccine as well as monitoring vaccine-induced immune responses in the clinic.

Impact of the B.1.1.7 variant on neutralizing monoclonal antibodies recognizing diverse epitopes on SARS–CoV–2 Spike

2021 ◽  
Author(s):  
Carl Graham ◽  
Jeffrey Seow ◽  
Isabella Huettner ◽  
Hataf Khan ◽  
Neophytos Kouphou ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Viral Entry ◽  
Neutralizing Antibodies ◽  
Infectious Virus ◽  
Antigenic Drift ◽  
Host Cells ◽  
Receptor Binding Domain ◽  
Neutralization Activity ◽  
Neutralizing Activity ◽  
Neutralizing Epitopes

The interaction of the SARS–CoV–2 Spike receptor binding domain (RBD) with the ACE2 receptor on host cells is essential for viral entry. RBD is the dominant target for neutralizing antibodies and several neutralizing epitopes on RBD have been molecularly characterized. Analysis of circulating SARS–CoV–2 variants has revealed mutations arising in the RBD, the N–terminal domain (NTD) and S2 subunits of Spike. To fully understand how these mutations affect the antigenicity of Spike, we have isolated and characterized neutralizing antibodies targeting epitopes beyond the already identified RBD epitopes. Using recombinant Spike as a sorting bait, we isolated >100 Spike–reactive monoclonal antibodies from SARS–CoV–2 infected individuals. ≈45% showed neutralizing activity of which ≈20% were NTD–specific. None of the S2–specific antibodies showed neutralizing activity. Competition ELISA revealed that NTD–specific mAbs formed two distinct groups: the first group was highly potent against infectious virus, whereas the second was less potent and displayed glycan–dependant neutralization activity. Importantly, mutations present in B.1.1.7 Spike frequently conferred resistance to neutralization by the NTD–specific neutralizing antibodies. This work demonstrates that neutralizing antibodies targeting subdominant epitopes need to be considered when investigating antigenic drift in emerging variants.

scholarly journals Characterization of Neutralizing Antibodies and Identification of Neutralizing Epitope Mimics on the Clostridium botulinum Neurotoxin Type A

2001 ◽  
Vol 67 (7) ◽  
pp. 3201-3207 ◽  
Author(s):  
Han-Chung Wu ◽  
Chia-Tsui Yeh ◽  
Yue-Ling Huang ◽  
Lih-Jeng Tarn ◽  
Chien-Cheng Lung
Keyword(s):  
Monoclonal Antibodies ◽  
Neutralizing Antibodies ◽  
Clostridium Botulinum ◽  
Neuromuscular Junctions ◽  
Competitive Inhibition ◽  
Lethal Dose ◽  
Humoral Response ◽  
Type A ◽  
Phage Clone ◽  
Neutralizing Epitopes

ABSTRACT Clostridium botulinum neurotoxin type A (BTx-A) is known to inhibit the release of acetylcholine at the neuromuscular junctions and synapses and to cause neuroparalysis and death. In this study, we have identified two monoclonal antibodies, BT57-1 and BT150-3, which protect ICR mice against lethal doses of BTx-A challenge. The neutralizing activities for BT57-1 and BT150-3 were 103 and 104 times the 50% lethal dose, respectively. Using immunoblotting analysis, BT57-1 was recognized as a light chain and BT150-3 was recognized as a heavy chain of BTx-A. Also, applying the phage display method, we investigated the antibodies' neutralizing B-cell epitopes. These immunopositive phage clones displayed consensus motifs, Asp-Pro-Leu for BT57-1 and Cys-X-Asp-Cys for BT150. The synthetic peptide P4M (KGTFDPLQEPRT) corresponded to the phage-displayed peptide selected by BT57-1 and was able to bind the antibodies specifically. This peptide was also shown by competitive inhibition assay to be able to inhibit phage clone binding to BT57-1. Aspartic acid (D5) in P4M was crucial to the binding of P4M to BT57-1, since its binding activity dramatically decreased when it was changed to lysine (K5). Finally, immunizing mice with the selected phage clones elicited a specific humoral response against BTx-A. These results suggest that phage-displayed random-peptide libraries are useful in identifying the neutralizing epitopes of monoclonal antibodies. In the future, the identification of the neutralizing epitopes of BTx-A may provide important information for the identification of the BTx-A receptor and the design of a BTx-A vaccine.

scholarly journals Escape from neutralizing antibodies by SARS-CoV-2 spike protein variants

2020 ◽  
Author(s):  
Yiska Weisblum ◽  
Fabian Schmidt ◽  
Fengwen Zhang ◽  
Justin DaSilva ◽  
Daniel Poston ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Receptor Binding ◽  
Neutralizing Antibodies ◽  
Spike Protein ◽  
Receptor Binding Domain ◽  
Reporter Virus ◽  
Low Frequencies ◽  
Protein Variants ◽  
Prior Infection ◽  
Neutralizing Epitopes

AbstractNeutralizing antibodies elicited by prior infection or vaccination are likely to be key for future protection of individuals and populations against SARS-CoV-2. Moreover, passively administered antibodies are among the most promising therapeutic and prophylactic anti-SARS-CoV-2 agents. However, the degree to which SARS-CoV-2 will adapt to evade neutralizing antibodies is unclear. Using a recombinant chimeric VSV/SARS-CoV-2 reporter virus, we show that functional SARS-CoV-2 S protein variants with mutations in the receptor binding domain (RBD) and N-terminal domain that confer resistance to monoclonal antibodies or convalescent plasma can be readily selected. Notably, SARS-CoV-2 S variants that resist commonly elicited neutralizing antibodies are now present at low frequencies in circulating SARS-CoV-2 populations. Finally, the emergence of antibody-resistant SARS-CoV-2 variants that might limit the therapeutic usefulness of monoclonal antibodies can be mitigated by the use of antibody combinations that target distinct neutralizing epitopes.

scholarly journals Isolation of Porcine Reproductive and Respiratory Syndrome Virus GP5-Specific, Neutralizing Monoclonal Antibodies From Hyperimmune Sows

2021 ◽  
Vol 12 ◽  
Author(s):  
Jordan E. Young ◽  
Cheryl M. T. Dvorak ◽  
Simon P. Graham ◽  
Michael P. Murtaugh
Keyword(s):  
Monoclonal Antibodies ◽  
B Cells ◽  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Growing Pigs ◽  
Respiratory Syndrome Virus ◽  
Broadly Neutralizing Antibodies ◽  
Complete Protection ◽  
Broadly Neutralizing Antibody ◽  
Neutralizing Epitopes

Porcine reproductive and respiratory syndrome (PRRS) is a devastating disease which impacts the pig industry worldwide. The disease is caused by PRRS viruses (PRRSV-1 and -2) which leads to abortions and other forms of reproductive failure in sows and severe respiratory disease in growing pigs. Current PRRSV vaccines provide limited protection; only providing complete protection against closely related strains. The development of improved PRRSV vaccines would benefit from an increased understanding of epitopes relevant to protection, including those recognized by antibodies which possess the ability to neutralize distantly related strains. In this work, a reverse vaccinology approach was taken; starting first with pigs known to have a broadly neutralizing antibody response and then investigating the responsible B cells/antibodies through the isolation of PRRSV neutralizing monoclonal antibodies (mAbs). PBMCs were harvested from pigs sequentially exposed to a modified-live PRRSV-2 vaccine as well as divergent PRRSV-2 field isolates. Memory B cells were immortalized and a total of 5 PRRSV-specific B-cell populations were isolated. All identified PRRSV-specific antibodies were found to be broadly binding to all PRRSV-2 isolates tested, but not PRRSV-1 isolates. Antibodies against GP5 protein, commonly thought to possess a dominant PRRSV neutralizing epitope, were found to be highly abundant, as four out of five B cells populations were GP5 specific. One of the GP5-specific mAbs was shown to be neutralizing but this was only observed against homologous and not heterologous PRRSV strains. Further investigation of these antibodies, and others, may lead to the elucidation of conserved neutralizing epitopes that can be exploited for improved vaccine design and lays the groundwork for the study of broadly neutralizing antibodies against other porcine pathogens.

scholarly journals Escape from neutralizing antibodies by SARS-CoV-2 spike protein variants

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Yiska Weisblum ◽  
Fabian Schmidt ◽  
Fengwen Zhang ◽  
Justin DaSilva ◽  
Daniel Poston ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Receptor Binding ◽  
Neutralizing Antibodies ◽  
Spike Protein ◽  
Receptor Binding Domain ◽  
Reporter Virus ◽  
Low Frequencies ◽  
Protein Variants ◽  
Prior Infection ◽  
Neutralizing Epitopes

Neutralizing antibodies elicited by prior infection or vaccination are likely to be key for future protection of individuals and populations against SARS-CoV-2. Moreover, passively administered antibodies are among the most promising therapeutic and prophylactic anti-SARS-CoV-2 agents. However, the degree to which SARS-CoV-2 will adapt to evade neutralizing antibodies is unclear. Using a recombinant chimeric VSV/SARS-CoV-2 reporter virus, we show that functional SARS-CoV-2 S protein variants with mutations in the receptor-binding domain (RBD) and N-terminal domain that confer resistance to monoclonal antibodies or convalescent plasma can be readily selected. Notably, SARS-CoV-2 S variants that resist commonly elicited neutralizing antibodies are now present at low frequencies in circulating SARS-CoV-2 populations. Finally, the emergence of antibody-resistant SARS-CoV-2 variants that might limit the therapeutic usefulness of monoclonal antibodies can be mitigated by the use of antibody combinations that target distinct neutralizing epitopes.

scholarly journals Passive transfer of modest titers of potent and broadly neutralizing anti-HIV monoclonal antibodies block SHIV infection in macaques

2014 ◽  
Vol 211 (10) ◽  
pp. 2061-2074 ◽  
Author(s):  
Masashi Shingai ◽  
Olivia K. Donau ◽  
Ronald J. Plishka ◽  
Alicia Buckler-White ◽  
John R. Mascola ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Neutralizing Antibodies ◽  
Rhesus Macaques ◽  
Passive Transfer ◽  
Neutralization Titer ◽  
Viral Pathogens ◽  
Shiv Infection ◽  
Anti Hiv ◽  
Sterilizing Immunity ◽  
Hiv 1

It is widely appreciated that effective human vaccines directed against viral pathogens elicit neutralizing antibodies (NAbs). The passive transfer of anti–HIV-1 NAbs conferring sterilizing immunity to macaques has been used to determine the plasma neutralization titers, which must be present at the time of exposure, to prevent acquisition of SIV/HIV chimeric virus (SHIV) infections. We administered five recently isolated potent and broadly acting anti-HIV neutralizing monoclonal antibodies (mAbs) to rhesus macaques and challenged them intrarectally 24 h later with either of two different R5-tropic SHIVs. By combining the results obtained from 60 challenged animals, we determined that the protective neutralization titer in plasma preventing virus infection in 50% of the exposed monkeys was relatively modest (∼1:100) and potentially achievable by vaccination.

scholarly journals A Novel Pseudomonas geniculata AGE Family Epimerase/Isomerase and Its Application in d-Mannose Synthesis

Foods ◽  
2020 ◽  
Vol 9 (12) ◽  
pp. 1809
Author(s):  
Zhanzhi Liu ◽  
Ying Li ◽  
Jing Wu ◽  
Sheng Chen
Keyword(s):  
3D Structure ◽  
Three Dimensional ◽  
Optimal Temperature ◽  
Reaction Conditions ◽  
Enzymatic Production ◽  
Physiological Properties ◽  
Potential Applications ◽  
Kinetics Of ◽  
Optimal Reaction ◽  
Gene Age

d-mannose has exhibited excellent physiological properties in the food, pharmaceutical, and feed industries. Therefore, emerging attention has been applied to enzymatic production of d-mannose due to its advantage over chemical synthesis. The gene age of N-acetyl-d-glucosamine 2-epimerase family epimerase/isomerase (AGEase) derived from Pseudomonas geniculata was amplified, and the recombinant P. geniculata AGEase was characterized. The optimal temperature and pH of P. geniculata AGEase were 60 °C and 7.5, respectively. The Km, kcat, and kcat/Km of P. geniculata AGEase for d-mannose were 49.2 ± 8.5 mM, 476.3 ± 4.0 s−1, and 9.7 ± 0.5 s−1·mM−1, respectively. The recombinant P. geniculata AGEase was classified into the YihS enzyme subfamily in the AGE enzyme family by analyzing its substrate specificity and active center of the three-dimensional (3D) structure. Further studies on the kinetics of different substrates showed that the P. geniculata AGEase belongs to the d-mannose isomerase of the YihS enzyme. The P. geniculata AGEase catalyzed the synthesis of d-mannose with d-fructose as a substrate, and the conversion rate was as high as 39.3% with the d-mannose yield of 78.6 g·L−1 under optimal reaction conditions of 200 g·L−1d-fructose and 2.5 U·mL−1P. geniculata AGEase. This novel P. geniculata AGEase has potential applications in the industrial production of d-mannose.

scholarly journals The Impact on Infectivity and Neutralization Efficiency of SARS-CoV-2 Lineage B.1.351 Pseudovirus

Viruses ◽  
2021 ◽  
Vol 13 (4) ◽  
pp. 633
Author(s):  
Yeong Jun Kim ◽  
Ui Soon Jang ◽  
Sandrine M. Soh ◽  
Joo-Youn Lee ◽  
Hye-Ra Lee
Keyword(s):  
South Africa ◽  
Monoclonal Antibodies ◽  
Cell Fusion ◽  
Receptor Binding ◽  
Neutralizing Antibodies ◽  
Receptor Binding Domain ◽  
Viral Spread ◽  
New Variant ◽  
Global Concern ◽  
The Impact

A new variant of SARS-CoV-2 B.1.351 lineage (first found in South Africa) has been raising global concern due to its harboring of multiple mutations in the spike that potentially increase transmissibility and yield resistance to neutralizing antibodies. We here tested infectivity and neutralization efficiency of SARS-CoV-2 spike pseudoviruses bearing particular mutations of the receptor-binding domain (RBD) derived either from the Wuhan strains (referred to as D614G or with other sites) or the B.1.351 lineage (referred to as N501Y, K417N, and E484K). The three different pseudoviruses B.1.351 lineage related significantly increased infectivity compared with other mutants that indicated Wuhan strains. Interestingly, K417N and E484K mutations dramatically enhanced cell–cell fusion than N501Y even though their infectivity were similar, suggesting that K417N and E484K mutations harboring SARS-CoV-2 variant might be more transmissible than N501Y mutation containing SARS-CoV-2 variant. We also investigated the efficacy of two different monoclonal antibodies, Casirivimab and Imdevimab that neutralized SARS-CoV-2, against several kinds of pseudoviruses which indicated Wuhan or B.1.351 lineage. Remarkably, Imdevimab effectively neutralized B.1.351 lineage pseudoviruses containing N501Y, K417N, and E484K mutations, while Casirivimab partially affected them. Overall, our results underscore the importance of B.1.351 lineage SARS-CoV-2 in the viral spread and its implication for antibody efficacy.

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