Folate binding in intestinal brush border membranes: evidence for the presence of two binding activities

Binding of [(3)H]folic acid by isolated rat jejunal brush border membranes (BBMs) was analyzed by chromatography on small Biogel P-30 columns. Folic acid binding to BBMs exhibited a prominent pH effect with a sharp maximum at pH 5.5 to 6.0. After acid treatment to strip the BBMs of bound folate, the membranes demonstrated a wider pH optimum (5.5 to 7.5) of folate binding and a higher binding capacity. Scatchard analysis of binding experiments performed at pH 6.0 revealed the existence of two components: one with a high affinity (kd = 12 to 25 nM) and low capacity (V(max) for non-acidified BBMs = 0.259 to 0.264 pmol/mg protein, V(max) for acidified BBMs = 0.41 to 0.71 pmol/mg protein) and the other with a low affinity (kd = 1.1 to 5.1 microM and high capacity (V(max) for non-acidified BBMs = 0.93 to 1.93 pmol/mg protein, V(max) for acidified BBMs = 4.05 to 7.69 pmol/mg protein). Phosphatidylinositol-specific phospholipase C preferentially detached the high affinity component from jejunal BBMs. Phosphatidylinositol-specific phospholipase C-released folate binding protein was precipitated by antibodies to the high-affinity folate-binding protein from rat kidney. These data suggest the existence of two different folate-binding proteins in isolated rat jejunal BBMs. The high-affinity folate-binding protein shares epitopes with the folate-binding protein in the kidney.