Due to the high prevalence and extensive genetic diversity of foot-and-mouth disease virus (FMDV), an RNA virus, substantial annual economic losses occur in Iran’s livestock farming and food production sectors. Considering the important role of the G–H loop, particularly the conserved RGD motif within the VP1 capsid protein, in eliciting immune responses, recombinant monoclonal antibodies were designed using bioinformatics approaches.
The epitopes and paratopes of protein sequences and antibody structures binding to FMDV VP1 were edited in Chimera and subsequently analyzed using web-based servers and molecular docking tools. Next, post-translational modification features and antibody-binding properties were evaluated. The final engineered scFv construct, consisting of light and heavy chains connected by a linker and containing a signal peptide and His-tag, was optimized for expression in E. coli.
Crystallographic structure analysis showed that three amino acids, R3141, G3142, and D3143, from the G–H loop were in direct contact with both the light and heavy chains of the antibody. The strongest interaction was observed between R3141 and D2098 of the light chain, with a distance of less than 4 Å. In addition, no N-glycosylation sites were detected in either the heavy or light chain. The numbers of interface residues in the heavy and light chains were 41 and 33, respectively, while the numbers of hydrogen bonds were 2 and 1, respectively.
The final construct consisted of 473 amino acids and had a calculated molecular weight of 50.3 kDa, an isoelectric point of 6.9, and a GRAVY value of 0.29. Finally, codon optimization was performed for expression in E. coli, resulting in a CAI value of 0.91 and a GC content of 54%. The designed construct was virtually cloned into the pET28a vector and subsequently submitted to the manufacturer for synthesis and cloning.

Keywords: Foot-and-mouth disease virus, Recombinant monoclonal antibody, Bioinformatics design, VP1 protein