High-affinity folate binding in human mammary gland.
High-affinity 3H-folate binding in Triton X-100 solubilized human mammary gland tissue displayed characteristics, e.g. apparent positive cooperativity and increasing affinity with decreasing concentration of folate binding protein, shown to be typical of specific folate binding. Radioligand dissociation was slow at pH 7.4. A major fraction of the bound radioligand dissociated rapidly at pH 3.5, while a residual binding of 20% persisted even after prolonged dialysis at pH 3.5. Gel chromatography revealed two major folate binding proteins (M(r) approximately 100 kDa and 25 kDa). However, only one single band was detectable on SDS-PAGE immunoblotting. The highest folate binding activity per g protein was associated with the upper triglyceride-containing layer of the 1000 g supernatant of the homogenate. The folate binding protein extracted from this layer had a low cross-reactivity (< 5%) with rabbit antibodies against 25 kDa human milk folate binding protein. The folate binding protein in the 1000 g pellet and the aqueous phase of the 1000 g supernatant was present at a low concentration and had a cross-reactivity of 100%.
Holm
J
J
Department of Clinical Chemistry, Central Hospital Nykøbing Falster, Denmark.
Hansen
S I
SI
Høier-Madsen
M
M
D001940:Breast / Q000737:chemistry* / Q000378:metabolism; D002352:Carrier Proteins / Q000032:analysis*; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D058973:Folate Receptors, GPI-Anchored; D005492:Folic Acid / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D015151:Immunoblotting; D008895:Milk, Human / Q000737:chemistry; D011869:Radioligand Assay; D011956:Receptors, Cell Surface; D014316:Tritium
D002352:Carrier Proteins; D058973:Folate Receptors, GPI-Anchored; D011956:Receptors, Cell Surface; D014316:Tritium; D005492:Folic Acid