The role of pathologic auto-antibodies against myelin oligodendrocyte glycoprotein (MOG) in multiple sclerosis is a highly contro- versial matter. As the use of animal models may enable to unravel the molecular mechanisms of the human disorder, numerous studies on multiple sclerosis are carried out using experimental autoimmune encephalomyelitis (EAE). In particular, the most exten- sively used EAE model is obtained by immunizing C57BL/6 mice with the immunodominant peptide MOG(35–55). In this scenario, we analyzed the anti-MOG antibody response in this model using the recombinant refolded extracellular domain of the protein, MOG(1–117). To assess the presence of a B-cell intramolecular epitope spreading mechanism, we tested also five synthetic peptides mapping the 1–117 sequence of MOG, including MOG(35–55). For this purpose, we cloned, expressed in Escherichia coli and on- column refolded MOG(1–117), and we applied an optimized microwave-assisted solid-phase synthetic strategy to obtain the de- signed peptide sequences. Subsequently, we set up a solid-phase immunoenzymatic assay testing both naïve and EAE mice sera and using MOG protein and peptides as antigenic probes. The results obtained disclose an intense IgG antibody response against both the recombinant protein and the immunizing peptide, while no response was observed against the other synthetic fragments, thus excluding the presence of an intramolecular epitope spreading mechanism. Furthermore, as the properly refolded recombinant probe is able to bind antibodies with greater efficiency compared with MOG(35–55), we hypothesize the presence of both linear and conformational epitopes on MOG(35–55) sequence.

Epitope mapping of anti-myelin oligodendrocyte glycoprotein (MOG) antibodies in a mouse model of multiple sclerosis: Microwave-assisted synthesis of the peptide antigens and ELISA screening / Pacini, Giulia; Ieronymaki, Matthaia; Nuti, Francesca; Sabatino, Giuseppina; Larregola, Maud; Aharoni, Rina; Papini, Anna Maria; Rovero, Paolo. - In: JOURNAL OF PEPTIDE SCIENCE. - ISSN 1075-2617. - ELETTRONICO. - 22:(2016), pp. 52-58. [10.1002/psc.2839]

Epitope mapping of anti-myelin oligodendrocyte glycoprotein (MOG) antibodies in a mouse model of multiple sclerosis: Microwave-assisted synthesis of the peptide antigens and ELISA screening

Pacini, Giulia;NUTI, FRANCESCA;SABATINO, GIUSEPPINA;PAPINI, ANNA MARIA;ROVERO, PAOLO
2016

Abstract

The role of pathologic auto-antibodies against myelin oligodendrocyte glycoprotein (MOG) in multiple sclerosis is a highly contro- versial matter. As the use of animal models may enable to unravel the molecular mechanisms of the human disorder, numerous studies on multiple sclerosis are carried out using experimental autoimmune encephalomyelitis (EAE). In particular, the most exten- sively used EAE model is obtained by immunizing C57BL/6 mice with the immunodominant peptide MOG(35–55). In this scenario, we analyzed the anti-MOG antibody response in this model using the recombinant refolded extracellular domain of the protein, MOG(1–117). To assess the presence of a B-cell intramolecular epitope spreading mechanism, we tested also five synthetic peptides mapping the 1–117 sequence of MOG, including MOG(35–55). For this purpose, we cloned, expressed in Escherichia coli and on- column refolded MOG(1–117), and we applied an optimized microwave-assisted solid-phase synthetic strategy to obtain the de- signed peptide sequences. Subsequently, we set up a solid-phase immunoenzymatic assay testing both naïve and EAE mice sera and using MOG protein and peptides as antigenic probes. The results obtained disclose an intense IgG antibody response against both the recombinant protein and the immunizing peptide, while no response was observed against the other synthetic fragments, thus excluding the presence of an intramolecular epitope spreading mechanism. Furthermore, as the properly refolded recombinant probe is able to bind antibodies with greater efficiency compared with MOG(35–55), we hypothesize the presence of both linear and conformational epitopes on MOG(35–55) sequence.
2016
22
52
58
Pacini, Giulia; Ieronymaki, Matthaia; Nuti, Francesca; Sabatino, Giuseppina; Larregola, Maud; Aharoni, Rina; Papini, Anna Maria; Rovero, Paolo...espandi
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Utilizza questo identificatore per citare o creare un link a questa risorsa: https://hdl.handle.net/2158/1028297
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