Endotoxin (Lipopolysaccharide) Neutralization by Innate Immunity Host-Defense Peptides: PEPTIDE PROPERTIES AND PLAUSIBLE MODES OF ACTION

Yosef Rosenfeld,Niv Papo,Yechiel Shai

doi:10.1074/jbc.m504327200

Yosef Rosenfeld, Niv Papo + Show 1 more

Open Access

https://doi.org/10.1074/jbc.m504327200

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Abstract

Binding of lipopolysaccharide (LPS) to macrophages results in proinflammatory cytokine secretion. In extreme cases it leads to endotoxic shock. A few innate immunity antimicrobial peptides (AMPs) neutralize LPS activity. However, the underlying mechanism and properties of the peptides are not yet clear. Toward meeting this goal we investigated four AMPs and their fluorescently labeled analogs. These AMPs varied in composition, length, structure, and selectivity toward cells. The list included human LL-37 (37-mer), magainin (24-mer), a 15-mer amphipathic alpha-helix, and its D,L-amino acid structurally altered analog. The peptides were investigated for their ability to inhibit LPS-mediated cytokine release from RAW264.7 and bone marrow-derived primary macrophages, to bind LPS in solution, and when LPS is already bound to macrophages (fluorescence spectroscopy and confocal microscopy), to compete with LPS for its binding site on the CD14 receptor (flow cytometry) and affect LPS oligomerization. We conclude that a strong binding of a peptide to LPS aggregates accompanied by aggregate dissociation prevents LPS from binding to the carrier protein lipopolysaccharide-binding protein, or alternatively to its receptor, and hence inhibits cytokine secretion.

Highlights

Lipopolysaccharide (LPS),2 termed endotoxin, is an integral structural component of the outer membrane of Gram-negative bacteria [1]
The list includes two native antimicrobial peptides, human LL-37 (37-mer) and magainin (24-mer), as well as a 15-mer amphipathic ␣-helical peptide composed of Leu and Lys, and its D,L-amino acid diastereomer, a structurally altered analog
The list includes human LL-37, magainin, a model 15-mer peptide composed of only Lys and Leu, and its D,L-amino acid analog (Table 1)

Summary

Introduction

Lipopolysaccharide (LPS), termed endotoxin, is an integral structural component of the outer membrane of Gram-negative bacteria [1]. Recent studies have shown that a few antimicrobial peptides (AMPs) have the potential to neutralize LPS-induced endotoxic effects. These peptides are important components of the innate defense system of all species of life [21]. Several AMPs prevent LPS-dependent cytokine induction in macrophages and block sepsis in animal models [23,24,25,26,27] These studies showed a direct correlation between the ability of AMPs to bind LPS and their antimicrobial activity. To better understand the mechanism by which AMPs block LPS-dependent cytokine induction and the peptide properties required for this activity, we investigated four AMPs and their fluorescently labeled analogs. The results are discussed in view of the essential properties of a particular peptide required for LPS detoxification, as well as plausible modes of action

Results

Discussion

Conclusion