Removal of the N-Glycosylation Sequon at Position N116 Located in p27 of the Respiratory Syncytial Virus Fusion Protein Elicits Enhanced Antibody Responses after DNA Immunization
Abstract
:1. Introduction
2. Materials and Methods
2.1. Cells, Virus and Antibodies
2.2. Construction and Expression of Recombinant RSV F Proteins
2.3. Western Blot Analysis
2.4. Immunofluorescence Analysis of Surface Expression
2.5. Flow Cytometric Analysis
2.6. Fusion Assay
2.7. Immunization of Mice with Plasmid DNA and RSV Challenge
2.8. Antibody Responses and Neutralization Assay
2.9. Determination of Lung Viral Titre by qRT-PCR
2.10. Immunohistochemistry
2.11. Statistical Analysis
3. Results
3.1. Expression Analysis of the RSV F Glycomutants and the Effect of N-Sequon Removal on RSV F-Specific mAb Recognition
3.2. RSV F Fusion Activity Is Determined by the N-Glycan at Position N500
3.3. Effect of Loss of RSV F N-Glycosylation Sites on Antibody Responses
3.4. Immunization with F N116Q DNA Results in Enhanced Neutralizing Antibody Responses and Protection upon Challenge
4. Discussion
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Surface Expression | Total Expression | % Transfected Cells | Fusion Capacity | Total Ab Response | Neutralizing Ab Response | |
---|---|---|---|---|---|---|
F N27Q | = | = | = | = | = | = |
F N70Q | – | – | = | = 1 | = | − |
F N116Q | = | = | = | = 1 | = | + |
F N126Q | = | − | = | = | = | = |
F N500Q | – | − | – | − | = | = |
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Leemans, A.; Boeren, M.; Van der Gucht, W.; Pintelon, I.; Roose, K.; Schepens, B.; Saelens, X.; Bailey, D.; Martinet, W.; Caljon, G.; et al. Removal of the N-Glycosylation Sequon at Position N116 Located in p27 of the Respiratory Syncytial Virus Fusion Protein Elicits Enhanced Antibody Responses after DNA Immunization. Viruses 2018, 10, 426. https://doi.org/10.3390/v10080426
Leemans A, Boeren M, Van der Gucht W, Pintelon I, Roose K, Schepens B, Saelens X, Bailey D, Martinet W, Caljon G, et al. Removal of the N-Glycosylation Sequon at Position N116 Located in p27 of the Respiratory Syncytial Virus Fusion Protein Elicits Enhanced Antibody Responses after DNA Immunization. Viruses. 2018; 10(8):426. https://doi.org/10.3390/v10080426
Chicago/Turabian StyleLeemans, Annelies, Marlies Boeren, Winke Van der Gucht, Isabel Pintelon, Kenny Roose, Bert Schepens, Xavier Saelens, Dalan Bailey, Wim Martinet, Guy Caljon, and et al. 2018. "Removal of the N-Glycosylation Sequon at Position N116 Located in p27 of the Respiratory Syncytial Virus Fusion Protein Elicits Enhanced Antibody Responses after DNA Immunization" Viruses 10, no. 8: 426. https://doi.org/10.3390/v10080426