IDENTIFICATION OF BIONEUTRALIZATION EPITOPES OF HUMAN FOLLICLE-STIMULATING-HORMONE IN THE REGION-31-52 AND REGION-66-75 OF ITS BETA-SUBUNIT

The crucial role played by follicle stimulating hormone (FSH) in regul ating both male and female reproduction and the possibilities of devel oping contraceptive methods for males by blocking the function of the hormone, makes it important to delineate the hormone-specific bioneutr alization epitopes of human follicle stimulating hormone (hFSH) on its beta-subunit. Predictive methods were used to identify the potential surface-oriented regions of hFSH-beta. Peptides corresponding to these regions, i.e. 31-52, 66-75 and 86-95 hFSH-beta, were synthesized, ant i-peptide antibodies were elicited in rabbits and the properties of th ese antisera to bind hFSH and neutralize its biological activity were assessed. Anti-31-52 hFSH-beta antisera bound hFSH specifically, where as anti-66-75 and anti-86-95 hFSH-beta antisera did not show any detec table binding, proving the region 31-52 hFSH-beta to be a specific ant igenic determinant of hFSH. The bioneutralizing abilities of the anti- peptide antibodies were assessed by measuring the hFSH-induced progest erone secretion by rat granulosa cells in vitro. Antibodies to 31-52 a nd 66-75 hFSH-beta neutralized the bioactivity of hFSH, but anti-86-95 hFSH-beta antibodies did not. Furthermore, the three linear peptides and two disulphide looped peptides of 31-52 hFSH-beta and 86-95 hFSH-b eta were also subjected to the in-vitro granulosa cell assay. The line ar peptides 31-52 hFSH-beta and 66-75 hFSH-beta and the cyclic 31-52 h FSH-beta disulphide loop peptide significantly inhibited the hFSH-indu ced progesterone secretion by rat granulosa cells, but the linear 86-9 5 hFSH-beta peptide and the corresponding cyclic disulphide loop pepti de did not. The results clearly show that the regions 31-52 and 66-75 of hFSH-beta harbor bioneutralization epitopes of the hormone. The stu dies also indicate that cyclization of the linear 31-52 hFSH-beta pept ide greatly enhances receptor recognition and that the region 66-75 hF SH-beta may also be involved in hormone-receptor interaction. (C) 1997 Elsevier Science Ireland Ltd.