Equine follicle-stimulating hormone: Molecular cloning of beta subunit andbiological role of the asparagine-linked oligosaccharide at asparagine(56)of alpha subunit

Equine FSH (eFSH) and eCG are members of the glycoprotein hormone family. T hese proteins are heterodimeric, composed of noncovalently associated alpha and 0 subunits. We have previously reported that recombinant eCG has poten t LH- and FSH-like activities and that the oligosaccharide at Asn(56) of th e alpha subunit plays an indispensable role in expressing LH- but not FSH-l ike activity. In the present study, we cloned eFSH beta subunit cDNA and ex pressed wild-type recombinant eFSH and a partially deglycosylated mutant FS H (eFSH alpha 56/beta) to investigate the biological role of the oligosacch aride at Asn(56) in FSH activity. The wild-type eFSH and eCG stimulated est radiol production in a dose-dependent manner in the primary cultures of rat granulosa cells, indicating that these equine gonadotropins have FSH activ ity. Partially deglycosylated eCG (eCG alpha 56/beta) also stimulated estra diol production, confirming that the FSH-like activity of eCG is resistant to the removal of the N-linked oligosaccharide. Partially deglycosylated eF SH (eFSH alpha 56/beta), however, did not show any FSH activity, indicating that the oligosaccharide at Asn(56) was necessary for eFSH. Thus, FSH-like activities of two gonadotropins, eCG and eFSH, are evoked through the dist inct molecular mechanisms regarding the biological role of oligosaccharide at Asn(56) of the alpha subunit.