| In this study, the metabolism of gonadotropin-releasing hormone (GnRH) and the involvement of a specific GnRH binding protein in goldfish serum were investigated. The binding of salmon GnRH (sGnRH) and its superactive analog, (D-Arg$sp6,$Pro$sp9$-NEt) -sGnRH to a macromolecular component of goldfish serum was characterized, using $sp{125}$I-sGnRH and $sp{125}$I-(D-Arg6-Pro9-NEt) -sGnRH as labeled ligand. Bound was separated from free labeled ligand by gel filtration with Sephadex G-50 mini-columns. The binding of labeled ligand to goldfish serum was dose-dependent, reversible and specific. The binding was associated with a serum protein, which was isolated from goldfish serum using a nondenaturing polyacrylamide gel electrophoresis (PAGE) method. This binding protein was concentrated in a single band, separated from all major components of serum proteins. The molecular weight of this GnRH binding protein was approximately 40,000 dalton, measured by SDS-PAGE. The isolated GnRH binding protein was found to specifically bind with sGnRH and chicken-II GnRH, the circulating forms of GnRH in goldfish serum identified using the combination of high pressure liquid chromatography and specific radioimmunoassays, but not with mammalian GnRH (mGnRH), chicken-I GnRH or lamprey GnRH. The GnRH binding protein was detected in both sexes of goldfish at different stages of the reproductive cycle, and its titer in serum was not affected by long term treatment of (D-Arg$sp6,$Pro$sp9$-NEt) -sGnRH or E$sb2$ implantation. The binding of GnRH to the binding protein in goldfish serum may affect the clearance of GnRH from circulation. There was good correlation between the half-lives of mGnRH, sGnRH, (D-Arg6,Pro9-NEt) -sGnRH and their affinity towards the binding protein, with the superactive analog (D-Arg6,Pro9-NEt) -sGnRH showing the highest affinity and longest half-life. |
