Localization of residues that confer antibody binding specificity using human chorionic gonadotropin/luteinizing hormone beta subunit chimeras and mutants.
The glycoprotein hormones are a family of conserved heterodimeric proteins which share a common alpha subunit but differ in their hormone-specific beta subunits. We used chimeras of human chorionic gonadotropin (hCG) and luteinizing hormone (hLH) beta subunits to identify residues which enable monoclonal antibodies (mAb) to distinguish the two hormones. The LH beta-CG beta chimeras appeared to fold similar to hCG beta, since they combined with hCG alpha and, depending on their sequences, were recognized by hCG-selective mAbs. Amino acid residues Arg8-Arg10,Gly47-Ala51, and Gln89-Leu92 form a major epitope region and appear to be adjacent to each other on the surface of hCG beta. Gly47-Ala51 and Gln89-Leu92 are recognized by dimer-specific mAbs while Arg8-Arg10 is recognized by mAbs which have highest affinity for the free beta subunit. These observations suggest that the conformation of this region of the beta subunit changes when the alpha and beta subunits combine. Residues which are C-terminal of Asp112 form a second epitope domain. mAbs to the third domain distinguish hCG beta and hLH beta by the presence of Asn77 in hCG beta and can be detected after hCG binds to receptors. These findings were used to develop a model of hCG beta which predicts the locations of these residues and their positions relative to the alpha subunit and receptor interfaces.
Department of Obstetrics/Gynecology, Robert Wood Johnson (Rutgers) Medical School, Piscataway, New Jersey 08854.
W R
WR
Moyle
M M
MM
Matzuk
R K
RK
Campbell
E
E
Cogliani
D M
DM
Dean-Emig
A
A
Krichevsky
R W
RW
Barnett
I
I
Boime
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D015153:Blotting, Western; D002678:Chimera; D006063:Chorionic Gonadotropin / Q000235:genetics* / Q000276:immunology; D018997:Chorionic Gonadotropin, beta Subunit, Human; D006801:Humans; D007118:Immunoassay; D007986:Luteinizing Hormone / Q000235:genetics* / Q000276:immunology; D008958:Models, Molecular; D008961:Models, Structural; D008969:Molecular Sequence Data; D009154:Mutation; D010446:Peptide Fragments / Q000235:genetics* / Q000276:immunology; D011487:Protein Conformation; D012689:Sequence Homology, Nucleic Acid
D000911:Antibodies, Monoclonal; D006063:Chorionic Gonadotropin; D018997:Chorionic Gonadotropin, beta Subunit, Human; D010446:Peptide Fragments; D007986:Luteinizing Hormone