High and low affinity binding of folate to proteins in serum of pregnancy women.
Binding of [3H]folate to proteins in serum of pregnant women was studied in equilibrium dialysis experiments (pH 7.4, 37 degrees C). A Scatchard analysis revealed the presence of high-affinity (Kass = 10(10) M(-1), N = 0.4 nM folate) and low-affinity sites. The high-affinity folate binding protein (Mr approximately 30 000--35 000) appeared in front effluent after application of serum to a DEAE-Sepharose CL-6B column equilibrated with 0.05 M imidazole buffer (pH 6.3)/30 mM NaCl. Low-affinity binding protein eluted from the column after a rise in NaCl concentration to 1 M was mainly similar to albumin. A minor part was, however, associated with a large molecular size (Mr > 200 000) protein, probably alpha 2-macroglobulin. High-affinity binding which displayed positive cooperativity was saturated at folate concentrations above 10(-10) M. Folate dissociation was a complex process consisting of an initial rapid phase (terminated within 48 h) followed by a slow release. At pH 3.5 dissociation became rapid and complete. Purified methotrexate had no effect on high-affinity binding, whereas N10-methylfolate (an impurity in the methotrexate preparation) acted as a potent inhibitor. Low-affinity binding was proportional to the folate concentration within the range 10(-10)--10(-7) M. Dissociation of folate was rapid.
D001665:Binding Sites; D001667:Binding, Competitive; D001798:Blood Proteins / Q000302:isolation & purification / Q000378:metabolism*; D005260:Female; D005492:Folic Acid / Q000031:analogs & derivatives / Q000097:blood*; D006801:Humans; D008727:Methotrexate / Q000494:pharmacology; D011247:Pregnancy; D011485:Protein Binding
D001798:Blood Proteins; D005492:Folic Acid; D008727:Methotrexate