Generation and characterization of a recombinant antibody fragment that binds to the coat protein of grapevine leafroll-associated virus 3
| Autor: | Greta Nölke, Kerstin Uhde-Holzem, Mariangela Dell’Orco, Martin Anibal Orecchia, Rainer Fischer, Giovanni P. Martelli, Markus Sack, Stefan Schillberg, Pasquale Saldarelli |
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| Rok vydání: | 2008 |
| Předmět: |
Phage display
Antibody Affinity Nicotiana benthamiana Cross Reactions Antibodies Viral scFv Epitope Virus law.invention Epitopes GLRaV-3 Peptide Library law Virology Tobacco Vitis Closteroviridae Immunoglobulin Fragments Plant Diseases biology fungi General Medicine biology.organism_classification Molecular biology Recombinant Proteins Pepscan biology.protein Recombinant DNA Capsid Proteins ELISA Antibody |
| Zdroj: | Archives of virology 153 (2008): 1075–1084. doi:10.1007/s00705-008-0100-3 info:cnr-pdr/source/autori:Orecchia M.; Nolke G.; Saldarelli P.; Dell'Orco M.; Uhde-Holzem K.; Sack M.; Martelli G.P.; Fischer R.; Schillberg S./titolo:Generation and characterization of a recombinant antibody fragment that binds to the coat protein of grapevine leafroll-associated virus3/doi:10.1007%2Fs00705-008-0100-3/rivista:Archives of virology/anno:2008/pagina_da:1075/pagina_a:1084/intervallo_pagine:1075–1084/volume:153 |
| ISSN: | 1432-8798 0304-8608 |
| Popis: | Pathogen-specific recombinant antibodies have been used to characterize pathogen infections and to engineer resistance in crops. We selected a single-chain antibody fragment (scFvLR3cp-1) specific for the coat protein of grapevine leafroll-associated virus 3 (GLRaV-3), one of the agents of grapevine leafroll (GLR) disease, from a phage display library. The antibody binds specifically to the entire length of GLRaV-3 particles and has a high binding affinity value (K D) of 42 nM. The amino acid motif AQEPPRQ located at the N terminus of the GLRaV-3 coat protein was identified as the antibody-binding epitope by PEPSCAN analysis. To evaluate scFv stability in the reducing environment of the plant cell cytosol, transient expression assays were performed using Nicotiana benthamiana as a model plant. Capture ELISA demonstrated that the scFv fragment was produced and retained its antigen-binding capacity in the plant cytosol. Further functional assays showed that scFvLR3cp-1 binds with high specificity to at least four members of the family Closteroviridae. Therefore, the GLRaV-3-specific scFv fragment could be an ideal candidate for mediating broad-spectrum virus resistance if produced in transgenic grapevine plants. |
| Databáze: | OpenAIRE |
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