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 appa...