title: Molecular cloning and functional expression of a human intestinal lactoferrin receptor.
abstract: Lactoferrin (Lf), a major iron-binding protein in human milk, has been suggested to have multiple biological roles such as facilitating iron absorption, modulating the immune system, embryonic development, and cell proliferation. Our previous binding studies suggested the presence of a specific receptor for Lf (LfR) in the small intestine of newborn infants, which may facilitate iron absorption. We here report the cloning and the functional expression of the human intestinal LfR and the evidence of its involvement in iron metabolism. The entire coding region of the LfR cDNA was cloned by PCR based on amino acid sequences of the purified native LfR (nLfR). The recombinant LfR (rLfR) was then expressed in a baculovirus-insect cell system and purified by immobilized human Lf (hLf) affinity chromatography where binding of hLf to the rLfR was partially Ca(2+) dependent. The apparent molecular mass was 136 kDa under nonreducing conditions and 34 kDa under reducing conditions. 125I-hLf bound to the rLfR with an apparent K(d) of approximately 360 nM. These biochemical properties of the rLfR are similar to those of the nLfR. RT-PCR revealed that the gene was expressed at high levels in fetal small intestine and in adult heart and at lower levels in Caco-2 cells. PI-PLC treatment of Caco-2 cells indicated that the LfR is GPI anchored. In Caco-2 cells transfected with the LfR gene, 125I-hLf binding and 59Fe-hLf uptake were increased by 1.7 and 3.4 times, respectively, compared to those in mock-transfected cells. Our findings demonstrate the presence of a unique receptor-mediated mechanism for nutrient uptake by the newborn.
authors.affiliation: Department of Nutrition, University of California, Davis, One Shields Avenue, Davis, California 95616, USA.
authors.forename: Y A
authors.initials: YA
authors.lastname: Suzuki
authors.forename: K
authors.initials: K
authors.lastname: Shin
authors.forename: B
authors.initials: B
authors.lastname: Lönnerdal
mesh_terms: D000328:Adult; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D018938:Caco-2 Cells; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000302:isolation & purification; D005333:Fetus; D015870:Gene Expression; D006801:Humans; D007421:Intestine, Small / Q000378:metabolism* / Q000502:physiology; D007457:Iodine Radioisotopes; D007781:Lactoferrin / Q000378:metabolism*; D008969:Molecular Sequence Data; D009036:Moths / Q000235:genetics; D009928:Organ Specificity / Q000235:genetics; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054801:Phosphoinositide Phospholipase C; D011485:Protein Binding; D011956:Receptors, Cell Surface / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D010738:Type C Phospholipases / Q000378:metabolism
chemical_list: D018076:DNA, Complementary; D007457:Iodine Radioisotopes; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; C038340:lactoferrin receptors; D010738:Type C Phospholipases; D054801:Phosphoinositide Phospholipase C; D007781:Lactoferrin; D043265:Phosphatidylinositol Diacylglycerol-Lyase