Recombinant factor C competes against LBP to bind lipopolysaccharide and neutralizes the endotoxicity.
Endotoxin-mediated inflammation and septic shock remains a grave challenge to human healthcare management. It is, therefore, a worthwhile effort to develop anti-lipopolysaccharide (LPS) strategies to prevent downstream effects. Here, we demonstrate that purified recombinant Factor C (rFC) cloned from the horseshoe crab, binds LPS with high affinity, preventing it from binding a peptide derived from the human LPS-binding protein (LBP). Factor C is an innate immune defense protein present in the horseshoe crab hemocytes. The full-length rFC was found to be more efficacious in blocking LBP-mediated downstream effects than either of the individual LPS-binding peptides (Sushi 1 and Sushi 3) derived from rFC. When added to human macrophage culture, rFC blocks the LPS-induced phosphorylation of p38, which, in turn, inhibits the consequential overexpression of TNF-alpha and IL-8. The tandem arrangement of the LPS-binding Sushi domains in the Factor C molecule appears to be required for the synergy and amplification of LPS-binding in vivo to achieve such high affinity for LPS. Thus, rFC binds and neutralizes LPS to arrest signal transduction via the p38 pathway. The rFC does not show acute cytotoxicity and could be a potential lead for further development into an endotoxin-antagonist to inhibit inflammation and septic shock.
Li
Peng
P
Department of Biological Sciences, National University of Singapore, Singapore.
Ho
Bow
B
Ding
Jeak Ling
JL
D000209:Acute-Phase Proteins / Q000378:metabolism*; D060829:Arthropod Proteins; D001665:Binding Sites; D001667:Binding, Competitive; D002352:Carrier Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D004731:Endotoxins / Q000633:toxicity; D004792:Enzyme Precursors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008070:Lipopolysaccharides / Q000633:toxicity*; D008264:Macrophages / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism*; D009000:Monocytes / Q000166:cytology / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D012697:Serine Endopeptidases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D000099231:Lipopolysaccharide-Binding Protein
D000209:Acute-Phase Proteins; D060829:Arthropod Proteins; D002352:Carrier Proteins; D004731:Endotoxins; D004792:Enzyme Precursors; D008070:Lipopolysaccharides; D008562:Membrane Glycoproteins; D011994:Recombinant Proteins; D012697:Serine Endopeptidases; D014409:Tumor Necrosis Factor-alpha; D000099231:Lipopolysaccharide-Binding Protein; C057151:Limulus clotting factor C