Characterization of epitope regions of thyrotropin beta-subunit recognizedby the monoclonal antibodies mAb279 and mAb299: a chimeric peptide approach

This investigation describes the design, synthesis and evaluation of chimer ic peptides related to the bovine thyrotropin beta-subunit, bTSH beta. The structures of these chimeric peptides were derived from investigations with linear peptides and sequence alignment studies, in association with a homo logy model of TSH beta developed from the hCG X-ray crystallographic struct ure. The structures of these chimeric peptides comprised beta-turn regions of loop LI [bTSH beta(14-20)] and loop L3 [bTSH beta(65-72)] held in close proximity by a bis-beta-alanine linker and the disulfide bond bTSH beta[Cys (16)-Cys(67)]. Linear and cyclic chimeric peptides were evaluated in immuno chemical assays for their ability to inhibit the binding of radio-iodinated bTSH beta [I-125-bTSH beta] to the monoclonal antibodies, mAb279 and mAb29 9. Previously, mAb279 and mAb299 have been shown to recognize epitopes acce ssible on the surface of TSH beta that lie in close proximity to the TSH re ceptor-binding site. The results indicate that these chimeric peptides can specifically inhibit in a dose-dependent manner the binding of I-125-bTSH b eta to mAb299, while having a lesser effect on the binding with mAb279. Bas ed on these results, it can be concluded that the bTSH beta-epitope recogni zed by mAb299 involves contributions from amino residues from the beta-turn regions of the L1 and L3 loops of TSH beta, and that these loop regions fl ank part of the receptor binding site of the bTSH beta-subunit.