Selection of mimotopes of pemphigus vulgaris antigen from a phage-displayed random nonapeptide library / 中华皮肤科杂志

ABSTRACT

Objective To screen the mimotopes ofpemphigus vulgaris (PV) antigen, desmoglein3 (Dsg3) with a phage-displayed random nonapeptide library, so as to update the knowledge on the patho-genesis of PV. Methods Recombinant fusion protein of extracellular domain 1-2 (EC1-2) of Dsg3 and glutathione transferase was expressed by E.coli BL21, and used to purify polyclonal autoantibody binding to recombinant EC 1-2 from the sera of patients with PV. Then, selected autoantibody was applied as a ligand for biopanning of a phage-displayed linear random nonapeptide library and circular random nonapeptide library. Monoclonal phages were selected by immunoscreening and tested with ELISA and competitive ELISA. Results After two rounds ofbiopanning, a population ofpeptide-displaying phages binding to autoan- tidody were highly enriched. Sixty individual phage clones selected by immunosereening were further sub-jected to screening with ELISA and competitive ELISA. Finally, three positive phage clones were obtained. As shown by ELISA and competitive ELISA, they reacted with serum from patients with PV but not with that from normal human controls, and blocked the interaction between patients' sera and recombinant fusion protein of EC1-2. Conclusion Three mimotopes closely associated with PV antigen were successfully selected from a phage-displayed random nonapeptide library.