High-affinity folate binding in leukocyte lysate from normal subjects: effect of concentration of binding protein,...

Folate binding in lysate prepared from leukocytes containing large amounts of the folate-binding protein was studied in equilibrium dialysis experiments (pH 7.4, 37 degrees C). Binding displayed positive cooperativity, and the binding affinity increased with decreasing concentration of binding protein (affinity constants in the range of 5 X 10(9) l/mol to 7 X 10(10) l/mol). Both phenomena could be interpreted in terms of ligand binding to a polymerizing system where the affinity of ligand for the oligomer is greater than its affinity for the polymer prevailing at high concentrations of the binding protein. Binding affinity was decreased at a low temperature (7 degrees C) and particularly at pH 5.0 where positive cooperativity also disappeared. The binder (Mr approximately 25,000-30,000) eluted in the front effluent after DEAE-Sepharose CL-6B chromatography of the lysate at pH 6.3.