Conversion of human choriogonadotropin into a follitropin by protein engineering.
Human reproduction is dependent upon the actions of follicle-stimulating hormone (hFSH), luteinizing hormone (hLH), and chorionic gonadotropin (hCG). While the alpha subunits of these heterodimeric proteins can be interchanged without effect on receptor-binding specificity, their beta subunits differ and direct hormone binding to either LH/CG or FSH receptors. Previous studies employing chemical modifications of the hormones, monoclonal antibodies, or synthetic peptides have implicated hCG beta-subunit residues between Cys-38 and Cys-57 and corresponding regions of hLH beta and hFSH beta in receptor recognition and activation. Since the beta subunits of hCG or hLH and hFSH exhibit very little sequence similarity in this region, we postulated that these residues might contribute to hormone specificity. To test this hypothesis we constructed chimeric hCG/hFSH beta subunits, coexpressed them with the human alpha subunit, and examined their ability to interact with LH and FSH receptors and hormone-specific monoclonal antibodies. Surprisingly, substitution of hFSH beta residues 33-52 for hCG beta residues 39-58 had no effect on receptor binding or stimulation. However, substitution of hFSH beta residues 88-108 in place of the carboxyl terminus of hCG beta (residues 94-145) resulted in a hormone analog identical to hFSH in its ability to bind and stimulate FSH receptors. The altered binding specificity displayed by this analog is not attributable solely to the replacement of hCG beta residues 108-145 or substitution of residues in the "determinant loop" located between hCG beta residues 93 and 100.
Campbell
R K
RK
Department of Obstetrics & Gynecology, University of Medicine and Dentistry of New Jersey, Robert Wood Johnson Medical School, Piscataway 08854.
Dean-Emig
D M
DM
Moyle
W R
WR
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D006063:Chorionic Gonadotropin / Q000235:genetics* / Q000378:metabolism / Q000494:pharmacology; D003338:Corpus Luteum / Q000378:metabolism; D005260:Female; D005640:Follicle Stimulating Hormone / Q000235:genetics* / Q000378:metabolism / Q000494:pharmacology; D005818:Genetic Engineering / Q000379:methods*; D006801:Humans; D007700:Kinetics; D007985:Leydig Cells / Q000187:drug effects / Q000378:metabolism; D008297:Male; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D010957:Plasmids; D051381:Rats; D011974:Receptors, LH / Q000187:drug effects / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism / Q000494:pharmacology; D012689:Sequence Homology, Nucleic Acid; D013739:Testosterone / Q000096:biosynthesis; D014162:Transfection
D006063:Chorionic Gonadotropin; D011974:Receptors, LH; D011994:Recombinant Proteins; D013739:Testosterone; D005640:Follicle Stimulating Hormone