Immunochemical mapping of antigenic regions on the human thyrotropin beta-subunit by antipeptide antibodies.
To study antigenic sites present in the beta-subunit of human thyrotropin (hTSH), we produced site-specific antibodies directed against synthetic peptides analogous to the 1-18, 44-59, and 85-112 regions of the thyrotropin beta-subunit. The hTSH beta(1-18) peptide-carrier conjugate elicited antisera capable of binding to both radiolabeled hTSH and its beta-subunit whereas antibodies elicited against the hTSH beta(44-59) peptide-carrier conjugate bound only to the peptide. Thus, the NH2-terminal region of hTSH beta appears to be accessible at the surface of the hormone whereas the hTSH beta(44-59) region may be poorly accessible. Two monoclonal antipeptide antibodies that bound to 125I-hTSH beta, designated as TS01 and TS02, were selected after immunization with the hTSH beta(85-112) peptide-carrier conjugate. The antigenic site recognized by TS01 was located on the eight COOH-terminal(105-112) amino acid residues. TS02 antibody bound to an antigenic region included within Cys95 and Cys105. Both antigenic sites appeared to be more accessible on the free hTSH beta than on the hormone. Immunoblots performed on various preparations containing TSH revealed that TS02 antibody detected the beta-subunit from both the human and bovine species but not the rat TSH beta. Under reducing conditions, a low molecular weight material was identified in hTSH beta, likely caused by intrachain nicking.
Département de Biologie Clinique, Institut Gustave-Roussy, Villejuif, France.
J M
JM
Bidart
F
F
Troalen
P
P
Ghillani
A
A
Puisieux
C
C
Bohuon
D
D
Bellet
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000941:Antigens / Q000378:metabolism*; D015153:Blotting, Western; D002417:Cattle; D006801:Humans; D007120:Immunochemistry; D008969:Molecular Sequence Data; D011863:Radioimmunoassay; D051381:Rats; D013552:Swine; D013972:Thyrotropin / Q000276:immunology* / Q000378:metabolism
D000911:Antibodies, Monoclonal; D000941:Antigens; D013972:Thyrotropin