PARTIAL PURIFICATION AND CHARACTERIZATION OF A BINDING PROTEIN FOR INSULIN-LIKE ACTIVITY (ILAs) IN HUMAN AMNIOTIC FLUID: A POSSIBLE INHIBITOR OF INSULIN-LIKE ACTIVITY

Abstract

An insulin radioreceptor assay (INS-RRA) and an insulin-like activity radioreceptor assay (ILA-RRA) were utilized to partially purify and characterize a protein from human amniotic fluid with ILA-RRA reactivity. An acid-ethanol soluble protein with an apparent MW of 34,500 daltons by calibrated Sephadex chromatography and an isoelectric point (pI) of 4.7 accounted for all the ILA-RRA reactivity present in human amniotic fluid. Since this protein is a binding protein for ILA, but not for insulin, it was termed amniotic fluid binding protein or AFBP. AFBP was reactive in a non-parallel manner in the ILA-RRA and totally inactive in the INS-RRA. The activity of AFBP in the ILA-RRA was due to the competition of AFBP with the placental membrane receptor for the [125I]ILA, tracer employed in the ILA-RRA. AFBP inhibited the activity of added ILA, but not of added insulin, in the INS-RRA, presumably by binding ILA, while being inactive itself. In 2 biological assays studied to date, the rat epidiymal fat pad assay and the rabbit chondrocyte sulfation assay, AFBP also inhibited the activity of added ILA. These observations raise the possibility that binding protein(s) for insulin-like peptides may function as inhibitors of their bioactivity in different physiologic and pathologic states. The relation of AFBP to binding protein(s) in human plasma remains to be clarified.