CHARACTERISTICS OF IRON-BINDING TO SOLUBILIZED BRUSH-BORDER MEMBRANE OF THE RAT INTESTINE

Characteristics of iron binding to the solubilized brush border membra ne (BBM) of rat intestine were studied. Specific Fe(II) and Fe(III) bi nding sites were detected by iron binding assay using immobilized BBM from the upper intestine on a nitrocellulose sheet and the binding to Fe(II) was considerably higher than that to Fe(III). The dissociation constants for Fe(II) were 0.26+/-0.02 nM (M+/-SE) for high-affinity bi nding sites and 2.70+/-0.26 nM for low-affinity binding sites. For Fe( II), the number of high-affinity binding sites was 0.35+/-0.04X10(16)/ mg of protein (M+/-SE) and that of low-affinity binding sites was 1.07 +/-0.11X10(16)/mg of protein. Bound Fe(II) could not be replaced with Fe (II), Fe(III) or transferrin-bound iron and approximately 50% of bo und Fe(II) was removed by chelators. The apparent molecular weights of Fe (II) binding peaks were 250 and 120 kDa by gel filtration. SDS-pol yacrylamide gel electrophoresis (SDS-PAGE) detected three bands with i ron-binding activity. The distribution of the Fe(II) binding sites sho wed that the number of low-affinity sites was significantly higher in the proximal third of the intestine compared with the rest of the inte stine. Iron deficiency increased the number of Fe(II) binding sites. T hese findings suggest that the Fe(II) binding sites might play a physi ological role in iron absorption in the rat.