Purification of Two Types of Gonadotropin Receptors From Carp Ovarian Follicles: Overlapping Recognition by Two Different Ligands
IR@IICB: CSIR-Indian Institute of Chemical Biology, KolkataView Archive Info
Purification of Two Types of Gonadotropin Receptors From
Carp Ovarian Follicles: Overlapping Recognition by Two Different Ligands
Cell Biology & Physiology
Teleostean gonadotropin receptors were solubilized from the plasma membrane preparation of carp ovarian follicles by lithium
diiodosalicylate and Triton X-100. Solubilization resulted fourfold increase in GTH binding activity as compared to the crude
plasma membrane preparation. An addition of 25% glycerol and protease inhibitors to the solubilized receptor retained more than
90% original activity at )20 �C for 30 days. Gel filtration through Sephadex G 100 significantly increased the specific binding capacity
from 70 fmol/mg protein (soluble receptor) to 250 fmol/mg protein. Peak I of gel filtration showing the receptor activity was
further purified by affinity chromatography on purified salmon GTH II — Sepharose with a remarkable increase in specific binding
capacity from 250 fmol/mg protein (gel filtration peak) to 2300 pmol/mg protein for salmon GTH I ligand and 2800 pmol/mg protein
for GTH II ligand. In SDS–PAGE affinity eluate the active peak showed two distinct bands corresponding to 66 and 62 kDa
molecular masses. These two proteins were clearly separated in FPLC Mono S chromatography, 62 kDa as GTHR I and 66 kDa as
GTHR II. The former preferably binds to GTH I ligand, while the latter to GTH II, although both demonstrated overlapping
recognition to both the ligands. GTH receptor protein I (GTHR I) and II (GTHR II) were purified 42,000- and 54,000-fold, respectively.
Competitive binding inhibition studies indicate GTH I and ovine FSH a better ligand for GTHR I, while GTH II and
ovine LH were preferable ligands for GTHR II. Biological relevance of these two receptor proteins was ascertained by monitoring
the specific binding capacity of GTHR I and II at different stages of the annual reproductive cycle. GTHR I–GTH I was a prevailing
complex during preparatory and pre-spawning stages, while GTHR II–GTH II was the dominant one at the maturational and final
maturational stages. It may be concluded there are two GTH receptor proteins, each having a preferred ligand. The overlapping
ligand-binding activity and profile of each receptor ligand complex suggest their link to seasonal development and maturation of
carp ovarian follicle.
Basu, Dipanjan and Bhattacharya, Samir (2002) Purification of Two Types of Gonadotropin Receptors From Carp Ovarian Follicles: Overlapping Recognition by Two Different Ligands. General and Comparative Endocrinology, 129 (3). pp. 152-162.