Lactoferrin‐binding sites at the surface of HT29‐D4 cells

Abstract

The binding of '251-lactoferrin to HT29-D4 cells, a clone of HT29 cells, was studied and compared to the binding of '"1-transferrin to the same cells. The binding of the two iron-transport proteins is saturable and reversiblc suggesting the presence of specific reccptors for each protein. Scatchard analysis suggests the existence of binding sites for lactoferrin with the relatively high equilibrium dissociation constant, K d l of 408 nM. Additionally, the cell is capable of binding large amounts of lactoferrin with very low affinity, probably in a non-receptor intermediale fashion. The dissociation constant of transferrill and its receptor was calculated 9.29 nM which corresponds well to values found in the literature. In contrast to lactoferrin, the cell was capable of binding only low amounts of transferrin in a non-receptor intermediate fashion. Aftcr chemical crosslinking of lactoferrin to the cell surface, the radiolabeled lactoferrin was found in a complex of molecular inass 300 kDa. Crosslinking of transferrin resulted in a complex of much higher molecular mass.These data clearly show a binding site for lactoferrin different from the transferrin receptor. Only if competition cxperiments were performed with a high molar excess of both ligand proteins did a small percentage of either of [he two ligands crossreact with the receptor for glycoproteins.Iron plays a critical role in many biological processes. Free iron under physiological conditions is in the hydroxide[Fe(OH)3] form and is insoluble. Iron is transported after chelating by a carrier system of iron-binding proteins. Lactoferrin, also known as lactotransferrin [l], is a member of the family or iron-binding proteins, that also includes transferrin, for which a critical role in iron metabolisin in higher animals has been shown. Transferrin is essential for the growth of many cell lines in serum-free medium and cellsurface receptors for transferrin are expressed on proliferating cclls in vitro and irz vivo [2, 31.The biological functions of lactoferrin, an important component excreted by granulocytes and several other cell types, are still poorly understood. Only its bacteriostatic role, attributed to its high affinity for iron (K, = 10' ' M-I) depriving bacteria of iron essential for growth, is well described 151.the other, possibly due to a structural similarity of the two These data suggest that lactoferrin may play the role of iron-carrier protein in this model. In this publication we present evidence for the presence of specific lactoferrin receptors on HT29 cells by comparing lactoferrin and transferrin interactions with the cells.

Effect of lactoferrin on the production of tumor necrosis factor-? and nitric oxide

“…If lactoferrin itself acts as a transcriptional factor that binds to the promoter region of the specific gene, it must first be established that lactoferrin can be transported to the nucleus from the outside of the cell. Several researchers have reported on the lactoferrin-specific receptor to which lactoferrin binds, as well as the translocation of the intact lactoferrin into nucleus [Garre et al, 1992;Hu et al, 1990;Roiron et al, 1989;Mazurier et al, 1989]. Once it can be translocated into the nucleus, it binds easily to the DNA [Ravazzollo et al, 1988].…”

Influence of lactoferrin on the entry process of Escherichia coli HB101(pRI203) in HeLa cells

“…Lf has been evidenced in all epithelial cells at the luminal surface as an autochthonous product of the enterocytes [26], and the presence of Lf receptors has been demonstrated on the enteric mucosa of several species [11,16,25].…”

Evaluation of the Bioactivity of Recombinant Human Lactoferrins Toward Murine Osteoblast-Like Cells for Bone Tissue Engineering

“…This most likely occur due to the structural similarity of the two glycoproteins. [51][52][53] With regard to transferrin and iron, the transferrin family is known to be potent regulators of iron homeostasis, 54 and Tfrc 1 is the primary target of transferrin in the iron transport system. It has been reported that low iron concentrations can lead to upregulation of this protein, 55 which may explain why Tfrc was differentially regulated by the iron unsaturated and saturated forms.…”