A single amino acid residue replacement in the beta subunit of human chorionic gonadotrophin results in the loss of biological activity.
The heterodimer, human chorionic gonadotrophin (hCG), contains an alpha subunit that is common to the glycoprotein hormones and a hormone-specific beta subunit. A comparison of all known beta amino acid sequences shows that an aspartic acid at position 99 (with the numbering scheme for hCG-beta) is one of the seven non-Cys invariant residues. Using site-directed mutagenesis we have replaced hCG-beta Asp99 with Arg. Chinese hamster ovary cells, containing a stably integrated gene for bovine alpha subunit, were transiently transfected with plasmids containing wild-type and mutant hCG-beta cDNAs. The Arg99 beta mutant associated with the alpha subunit, but the resulting heterodimer failed to enhance intracellular cyclic AMP production in a gonadotrophin-responsive transformed murine Leydig cell line. Thus, a single amino acid residue replacement in this glycosylated heterodimer containing 237 amino acid residues is sufficient to abolish activity.
Chen
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F
Department of Biochemistry and Molecular Biology, University of Miami School of Medicine, Florida 33101.
Puett
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D
D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D006063:Chorionic Gonadotropin / Q000235:genetics* / Q000494:pharmacology; D003001:Cloning, Molecular; D006224:Cricetinae; D000242:Cyclic AMP / Q000096:biosynthesis; D006801:Humans; D007985:Leydig Cells / Q000187:drug effects / Q000378:metabolism; D008297:Male; D016297:Mutagenesis, Site-Directed; D013329:Structure-Activity Relationship
D006063:Chorionic Gonadotropin; D000242:Cyclic AMP