Resurrecting inactive antimicrobial peptides from LPS trap.

Abstract

An important component on the outer membrane of Gram-negative bacteria is LPS (Lipopolysaccharide). LPS is an endotoxin, responsible for causing septic shock. It also serves as a barrier to exclude various antimicrobial agents. Recent studies on a membrane active antimicrobial peptide temporin A revealed that it is active towards Gram-positive bacteria. However, it aggregates in LPS of Gram-negative bacteria, hindering its translocation through the outer membrane. A previously described β-boomerang antiendotoxic peptide contains the LPS-binding motif GG8WF. In this project, we demonstrated GG8WF ability to rescue temporin A, by preventing its aggregation in LPS. This new conjugated peptide, FG21, showed antimicrobial activity towards both Gram-negative and Gram-positive bacteria. Tryptophan fluorescence studies, FITC-LPS fluorescence assay and dynamic light scattering measurements indicated that FG21 disaggregates LPS micelles. Zeta potential measurements proved that FG21 is binding to bacterial membrane. NPN and SYTOX uptake assays illustrated FG21 potential in breaking down membrane integrity. Results from these biophysical studies supported the hypothesis that FG21 is antiendotoxic and antimicrobial, able to bind LPS and permeabilize bacterial membrane. In conclusion, we are able to demonstrate that results from this project contributed knowledge towards rational design of the relatively new class of peptide antibiotics.