Abstract
It is important to understand the amino acid residues that govern the properties of the binding between antibodies and ligands. We studied the binding of two anti-norfloxacins, anti-nor 132 and anti-nor 155, and the fluoroquinolones norfloxacin, enrofloxacin, ciprofloxacin, and ofloxacin. Binding cross-reactivities tested by an indirect competitive enzyme-linked immunosorbent assay indicated that anti-nor 132 (22–100%) had a broader range of cross-reactivity than anti-nor 155 (62–100%). These cross-reactivities correlated with variations in the numbers of interacting amino acid residues and their positions. Molecular docking was employed to investigate the molecular interactions between the fluoroquinolones and the monoclonal antibodies. Homology models of the heavy chain and light chain variable regions of each mAb 3D structure were docked with the fluoroquinolones targeting the crucial part of the complementarity-determining regions. The fluoroquinolone binding site of anti-nor 155 was a region of the HCDR3 and LCDR3 loops in which hydrogen bonds were formed with TYR (H:35), ASN (H:101), LYS (H:106), ASN (L:92), and ASN (L:93). These regions were further away in anti-nor 132 and could not contact the fluoroquinolones. Another binding region consisting of HIS (L:38) and ASP (H:100) was found for norfloxacin, enrofloxacin, and ciprofloxacin, whereas only ASP (H:100) was found for ofloxacin.
Highlights
It is important to understand the amino acid residues that govern the properties of the binding between antibodies and ligands
Each antibody consists of four polypeptides—two heavy chains (H) and two light chains (L) joined to form a “Y”-shaped molecule
Detection based on immunological methods such as enzyme-linked immunosorbent assay (ELISA) and lateral flow immunoassays has been widely used for screening in food safety applications[20,21]
Summary
It is important to understand the amino acid residues that govern the properties of the binding between antibodies and ligands. Binding cross-reactivities tested by an indirect competitive enzyme-linked immunosorbent assay indicated that anti-nor 132 (22–100%) had a broader range of cross-reactivity than anti-nor 155 (62–100%) These cross-reactivities correlated with variations in the numbers of interacting amino acid residues and their positions. A change in the amino acid sequence in this region can greatly affect the specificity and affinity of the binding between the antibody and antigens. Detection based on immunological methods such as enzyme-linked immunosorbent assay (ELISA) and lateral flow immunoassays has been widely used for screening in food safety applications[20,21] In these methods, either an antibody with broad specificity to FQs or several specific antibodies for each antibiotic are required
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