Characterization of a prolactin binding protein in rat serum.
To determine the presence of a PRL-binding protein (PRL-BP) in rat serum, female rats were ovariectomized and administered sc estradiol benzoate capsules. Serum was incubated with [125I]rat PRL (rPRL) for 1 h at 37 C with or without different doses of rPRL, ovine PRL (oPRL), or rGH. Separation of the [125I]rPRL-PRL-BP complex from unbound [125I]rPRL was accomplished by Sephadex G100 chromatography or by precipitation with an antibody against rat liver PRL receptor. Results showed that a protein was able to bind specifically to rPRL or oPRL, but not to rGH. Scatchard plots gave an affinity constant of 1.18 +/- 0.7 10(9) M-1 and a capacity of 11.24 +/- 1.4 nM. The complex [125I]rPRL-PRL-BP migrated in the void volume of sephadex G100 column, but with an apparent mol wt of 80K after cross-linking and electrophoresis under reducing conditions. PRL BP was purified from estradiol-treated ovariectomized females by an oPRL sepharose 4B affinity column. Purified protein migrated under reducing conditions with apparent mol wt of 50K and 27K but of 160K under nonreducing conditions. After transfer on nitrocellulose filter, the 160K, 50K and 27K forms were able to bind to monoclonal antibodies directed against rat liver PRL receptor. The 160K and 50K were still able to bind to [125I]oPRL, but not the 27K. These results showed that rat serum contained a PRL-BP, which presented a strong homology to PRL receptor, at least for the immunological and binding characteristics.
INSERM U.352 Institut National des Sciences Apliquees, Villeurbanne, France.
H
H
Cohen
P
P
Guillaumot
I
I
Sabbagh
D000818:Animals; D002352:Carrier Proteins / Q000097:blood* / Q000737:chemistry; D002850:Chromatography, Gel; D003432:Cross-Linking Reagents; D004591:Electrophoresis, Polyacrylamide Gel; D004958:Estradiol / Q000494:pharmacology; D005260:Female; D008970:Molecular Weight; D010052:Ovariectomy; D011388:Prolactin / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley
D002352:Carrier Proteins; D003432:Cross-Linking Reagents; C029702:prolactin-binding protein; D004958:Estradiol; D011388:Prolactin