Targeted mutagenesis on PDGFRα-Fc identifies amino acid modifications that allow efficient inhibition of HCMV infection while abolishing PDGF sequestration
| Autor: | Feldmann, Svenja, Grimm, Immanuel, St��hr, Dagmar, Antonini, Chiara, Lischka, Peter, Sinzger, Christian, Stegmann, Cora |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2021 |
| Předmět: |
Cytomegalovirus Infection
Viral Diseases Receptor Platelet-Derived Growth Factor alpha Physiology Alanin Becaplermin Cytomegalovirus Pathology and Laboratory Medicine Biochemistry DDC 570 / Life sciences Medical Conditions Cell Signaling Animal Cells Immune Physiology Medicine and Health Sciences Enzyme-linked immunoassays Amino Acids Enzyme-Linked Immunoassays Biology (General) Connective Tissue Cells Immune System Proteins Alanine Organic Compounds Growth factor Signal inhibition Chemistry Infectious Diseases Connective Tissue Medical Microbiology Viral Pathogens Viruses Physical Sciences Cytomegalovirus Infections Intercellular Signaling Peptides and Proteins Fibroblast Human Cytomegalovirus Cellular Types Anatomy Pathogens Growth factors Research Article Signal Transduction Herpesviruses Signal Inhibition QH301-705.5 Immunoblotting Immunology Fibroblastenwachstumsfaktor Research and Analysis Methods Immunoblot Microbiology Antibodies ddc:570 Humans ddc:610 Immunoassays Microbial Pathogens Organic Chemistry Organisms Chemical Compounds Endothelial Cells Biology and Life Sciences Proteins Neural Inhibition Cell Biology Fibroblasts RC581-607 Immunoglobulin Fc Fragments Biological Tissue HEK293 Cells Aliphatic Amino Acids Immunologic Techniques Mutagenesis Site-Directed Immunologic diseases. Allergy DNA viruses DDC 610 / Medicine & health |
| Zdroj: | PLoS Pathogens, Vol 17, Iss 3, p e1009471 (2021) PLoS Pathogens |
| ISSN: | 1553-7374 1553-7366 |
| Popis: | Platelet-derived growth factor receptor alpha (PDGFR��) serves as an entry receptor for the human cytomegalovirus (HCMV), and soluble PDGFR��-Fc can neutralize HCMV at a halfmaximal effective concentration (EC50) of about 10 ng/ml. While this indicates a potential for usage as an HCMV entry inhibitor PDGFR��-Fc can also bind the physiological ligands of PDGFR�� (PDGFs), which likely interferes with the respective signaling pathways and represents a potential source of side effects. Therefore, we tested the hypothesis that interference with PDGF signaling can be prevented by mutations in PDGFR��-Fc or combinations thereof, without losing the inhibitory potential for HCMV. To this aim, a targeted mutagenesis approach was chosen. The mutations were quantitatively tested in biological assays for interference with PDGF-dependent signaling as well as inhibition of HCMV infection and biochemically for reduced affinity to PDGF-BB, facilitating quantification of PDGFR��-Fc selectivity for HCMV inhibition. Mutation of Ile 139 to Glu and Tyr 206 to Ser strongly reduced the affinity for PDGF-BB and hence interference with PDGF-dependent signaling. Inhibition of HCMV infection was less affected, thus increasing the selectivity by factor 4 and 8, respectively. Surprisingly, the combination of these mutations had an additive effect on binding of PDGF-BB but not on inhibition of HCMV, resulting in a synergistic 260fold increase of selectivity. In addition, a recently reported mutation, Val 242 to Lys, was included in the analysis. PDGFR��-Fc with this mutation was fully effective at blocking HCMV entry and had a drastically reduced affinity for PDGF-BB. Combining Val 242 to Lys with Ile 139 to Glu and/or Tyr 206 to Ser further reduced PDGF ligand binding beyond detection. In conclusion, this targeted mutagenesis approach identified combinations of mutations in PDGFR��-Fc that prevent interference with PDGF-BB but maintain inhibition of HCMV, which qualifies such mutants as candidates for the development of HCMV entry inhibitors. publishedVersion |
| Databáze: | OpenAIRE |
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