Evidence for the existence of LH-RH binding protein

LH-RH in rat hypothalamic extract (HE) emerges in 2 peaks after gel filtration on Sephadex G-25 with pyridine acetate buffer (PAB), pH 5.8, 1 peak in the unretarded protein fraction and the other near the salt region. A synthetic LH-RH marker was found only in the peak near the salt region by elution with acid. Using a dialysis technique, a protein fraction, from which LH-RH and other small molecular weight material were dissociated by gel filtration on Sephadex G-25 with acid, showed affinity for synthetic LH-RH. Synthetic LH-RH in the hypothalamic protein solution was retained in the dialysis bags more effectively than other types of protein solutions. This binding protein is of relatively small molecular size according to gel filtration behavior on Sephadex G-75. Since the dialysis experiment showed extremely poor recovery of total LH-RH, we determined the rate of loss of LH-RH under several conditions. The rate of disappearance was significantly less in solutions containing protein derived from cerebellum and hypothalamus, compared to bovine serum albumin (BSA) solution or salt solution. The ability of the hypothalamic protein to retard the rate of disappearance of LH-RH is further evidence for a special affinity between LH-RH and the protein factor. It is concluded that the protein LH-RH complex is dissociated by lowering the pH and that the dissociated protein and synthetic LH-RH can be re-associated to form a protein-hormone (PrH) complex at neutral pH.