Lactoferrin-lipid A-lipopolysaccharide interaction: inhibition by anti-human lactoferrin monoclonal antibody AGM 10.14.
Lactoferrin (LF) is a glycoprotein that exerts both bacteriostatic and bactericidal activities. The interaction of LF with lipopolysaccharide (LPS) of gram-negative bacteria seems to play a crucial role in the bactericidal effect. In this study, we evaluated, by means of an enzyme-linked immunosorbent assay, the binding of biotinylated LF to the S (smooth) and R (rough) (Ra, Rb, Rc, Rd1, Rd2, and Re) forms of LPS and different lipid A preparations. In addition, the effects of two monoclonal antibodies (AGM 10.14, an immunoglobulin G1 [IgG1] antibody, and AGM 2.29, an IgG2b antibody), directed against spatially distant epitopes of human LF, on the LF-lipid A or LF-LPS interaction were evaluated. The results showed that biotinylated LF specifically binds to solid-phase lipid A, as this interaction was prevented in a dose-dependent fashion by either soluble uncoupled LF or lipid A. The binding of LF to S-form LPS was markedly weaker than that to lipid A. Moreover, the rate of LF binding to R-form LPS was inversely related to core length. The results suggest that the polysaccharide O chain as well as oligosaccharide core structures may interfere with the LF-lipid A interaction. In addition, we found that soluble lipid A also inhibited LF binding to immobilized LPS, demonstrating that, in the whole LPS structure, the lipid A region contains the major determinant recognized by LF. AGM 10.14 inhibited LF binding to lipid A and LPS in a dose-dependent fashion, indicating that this monoclonal antibody recognizes an epitope involved in the binding of LF to lipid A or some epitope in its close vicinity. In contrast, AGM 2.29, even in a molar excess, did not prevent the binding of LF to lipid A or LPS. Therefore, AGM 10.14 may represent a useful tool for neutralizing selectively the binding of LF to lipid A. In addition, the use of such a monoclonal antibody could allow better elucidation of the consequences of the LF-lipid A interaction.
Caccavo
D
D
Department of Clinical Medicine, Immunology, and Infectious Diseases, University of Bari, Bari, Italy.
Afeltra
A
A
Pece
S
S
Giuliani
G
G
Freudenberg
M
M
Galanos
C
C
Jirillo
E
E
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D006801:Humans; D007781:Lactoferrin / Q000276:immunology / Q000378:metabolism*; D008050:Lipid A / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000378:metabolism*; D051379:Mice; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D008050:Lipid A; D008070:Lipopolysaccharides; D007781:Lactoferrin