Molecular cloning and functional expression of a human intestinal lactoferrin receptor

Y. A. Suzuki, K. Shin, B. Lönnerdal

Research output: Contribution to journalArticle

249 Citations (Scopus)

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 Ca2+ 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 Kd of @O360 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.

Original languageEnglish (US)
Pages (from-to)15771-15779
Number of pages9
JournalBiochemistry
Volume40
Issue number51
DOIs
StatePublished - Dec 25 2001

Fingerprint

Cloning
Molecular Cloning
Lactoferrin
Iron
Caco-2 Cells
Genes
Iron-Binding Proteins
Affinity chromatography
Immune system
Cell proliferation
Molecular mass
Programmable logic controllers
Metabolism
Nutrients
Small Intestine
Complementary DNA
Newborn Infant
Amino Acids
Polymerase Chain Reaction
Baculoviridae

ASJC Scopus subject areas

  • Biochemistry

Cite this

Molecular cloning and functional expression of a human intestinal lactoferrin receptor. / Suzuki, Y. A.; Shin, K.; Lönnerdal, B.

In: Biochemistry, Vol. 40, No. 51, 25.12.2001, p. 15771-15779.

Research output: Contribution to journalArticle

Suzuki, Y. A. ; Shin, K. ; Lönnerdal, B. / Molecular cloning and functional expression of a human intestinal lactoferrin receptor. In: Biochemistry. 2001 ; Vol. 40, No. 51. pp. 15771-15779.
@article{25739110808f4798bc9fc67a6129943c,
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 Ca2+ 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 Kd of @O360 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.",
author = "Suzuki, {Y. A.} and K. Shin and B. L{\"o}nnerdal",
year = "2001",
month = "12",
day = "25",
doi = "10.1021/bi0155899",
language = "English (US)",
volume = "40",
pages = "15771--15779",
journal = "Biochemistry",
issn = "0006-2960",
publisher = "American Chemical Society",
number = "51",

}

TY - JOUR

T1 - Molecular cloning and functional expression of a human intestinal lactoferrin receptor

AU - Suzuki, Y. A.

AU - Shin, K.

AU - Lönnerdal, B.

PY - 2001/12/25

Y1 - 2001/12/25

N2 - 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 Ca2+ 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 Kd of @O360 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.

AB - 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 Ca2+ 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 Kd of @O360 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.

UR - http://www.scopus.com/inward/record.url?scp=0035951070&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0035951070&partnerID=8YFLogxK

U2 - 10.1021/bi0155899

DO - 10.1021/bi0155899

M3 - Article

VL - 40

SP - 15771

EP - 15779

JO - Biochemistry

JF - Biochemistry

SN - 0006-2960

IS - 51

ER -