ISSN:
0263-6484
Keywords:
Hormones
;
GnRH
;
receptor
;
subcellular distribution
;
positive cooperativity
;
Chemistry
;
Biochemistry and Biotechnology
Source:
Wiley InterScience Backfile Collection 1832-2000
Topics:
Biology
,
Medicine
Notes:
Specific binding of a fully biologically active 125I-gonadotrophin releasing hormone (GnRH) to isolated anterior pituitary cells is time dependent, saturable and the concentration dependent binding curves exhibit positive cooperativity. Binding to intact or solubilized plasma membranes and an affinity purified GnRH receptor protein reveals in all instances multiple high affinity binding sites. Thus, GnRH receptor protein appears to be an intrinsic constituent of the cell membrane, and perhaps, other membranous organelles. To investigate the latter, the binding of 125I-GnRH to various subcellular fractions was studied and its affinity and time requirements determined. GnRH binding to plasma membranes and secretory granules was to multiple high affinity sites, while that to nuclei and microsomes was to a single high affinity site. Binding was 1.83 ± 0.07, 0.78 ± 0.04, 0.31 ± 0.03 and 0.27 ± 0.03 fmol μg-1 protein for isolated plasma membranes, secretory granules, microsomes and nuclei, respectively, after 30 min incubation with 10-9 M GnRH. The magnitude of binding to microsomes did not change during the incubation period. It did not show any decrease (p 〉 0.5) in isolated nuclei and plasma membranes, except for the 24 h time period, when a significant drop (p 〈 0.001) was seen. Binding to the secretory granule fraction culminated at 15 min and then decreased (p 〈 0.001) steadily to a non-detectable level at 24 h. Thus GnRH receptor protein or its portion may be an integral part of some membranous particles in the anterior pituitary cells. A single, low-capacity binding site may, or may not suggest the presence of a structurally incomplete form of the receptor protein in microsomes and nuclei. Binding to the secretory granules fraction exhibited only a relatively minor temporal difference compared to the plasma membrane, which may have resulted from an inappropriate conformational state of the receptor protein. Only the binding to the plasma membrane exhibited appropriately both the affinity and temporal requirements of the intact GnRH receptor protein in vitro and in vivo.
Additional Material:
5 Ill.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1002/cbf.290020405
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