Characterization of epitope regions of thyrotropin β-subunit recognized by the monoclonal antibodies mAB279 and mAB299: A chimeric peptide approach

This investigation describes the design, synthesis and evaluation of chimeric peptides related to the bovine thyrotropin β-subunit, bTSHβ. The structures of these chimeric peptides were derived from investigations with linear peptides and sequence alignment studies, in association with a homology model of TSHβ developed from the hCG X-ray crystallographic structure. The structures of these chimeric peptides comprised β-turn regions of loop L1 [bTSHβ(14-20)] and loop L3 [bTSHβ(65-72)] held in close proximity by a bis-β-alanine linker and the disulfide bond bTSHβ[Cys¹⁶- Cys⁶⁷]. Linear and cyclic chimeric peptides were evaluated in immunochemical assays for their ability to inhibit the binding of radio- iodinated bTSHβ [¹²⁵I-bTSHβ] to the monoclonal antibodies, mAb279 and mAb299. Previously, mAb279 and mAb299 have been shown to recognize epitopes accessible on the surface of TSHβ that lie in close proximity to the TSH receptor-binding site. The results indicate that these chimeric peptides can specifically inhibit in a dose-dependent manner the binding of ¹²⁵I-bTSHβ to mAb299, while having a lesser effect on the binding with mAb279. Based on these results, it can be concluded that the bTSHβ-epitope recognized by mAb299 revolves contributions from amino residues from the β-turn regions of the L1 and L3 loops of TSHβ, and that these loop regions flank part of the receptor binding site of the bTSH β-subunit.