LACTOFERRIN INHIBITS THE ENDOTOXIN INTERACTION WITH CD14 BY COMPETITION WITH THE LIPOPOLYSACCHARIDE-BINDING PROTEIN

Human lactoferrin (hLf), a glycoprotein released from neutrophil granu les during inflammation, and the lipopolysaccharide (LPS)-binding prot ein (LBP), an acute-phase serum protein, are known to bind to the lipi d A of LPS, The LPS-binding sites are located in the N-terminal region s of both proteins, at amino acid residues 28 to 34 of hLf and 91 to 1 08 of LBP. Both of these proteins modulate endotoxin activities, but t hey possess biologically antagonistic properties, In this study, we ha ve investigated the competition between hLf and recombinant human LBP (rhLBP) for the binding of Escherichia coli 055:B5 LPS to the differen tiated monocytic THP-1 cell line, Our studies revealed that hLf preven ted the rhLBP-mediated binding of LPS to the CD14 receptor on cells, M aximal inhibition of LPS-cell interactions by hLf was raised when both hLf and rhLBP were simultaneously added to LPS or when hLf and LPS we re mixed with cells 30 min prior to the incubation with rhLBP, However , when hLf was added 30 min after the interaction of rhLBP with LPS, t he binding of the rhLPS-LBP complex to CD14 could not be reversed, The se observations indicate that hLf competes with rhLBP for the LPS bind ing and therefore interferes with the interaction of LPS with CD14, Fu rthermore, experiments involving competitive binding of the rhLBP-LPS complex to cells with two recombinant mutated hLfs show that in additi on to residues 28 to 34, another basic cluster which contains residues 1 to 5 of hLf competes for the binding to LPS, Basic sequences homolo gous to residues 28 to 34 of hLf were evidenced on LPS-binding protein s such as LBP, bactericidal/permeability-increasing protein, and Limul us anti-LPS factor.