Characterization and Antimicrobial Activity of Amphiphilic Peptide AP3 and Derivative Sequences

Abstract

The continued emergence of new antibiotic resistant bacterial strains has resulted in great interest in the development of new antimicrobial treatments. Antimicrobial peptides (AMPs) are one of many potential classes of molecules to help meet this emerging need. AMPs are naturally derived sequences, which act as part of the innate immune system of organisms ranging from insects through humans. We investigated the antimicrobial peptide AP3, which is originally isolated from the winter flounder Pleuronectes americanus. This peptide is of specific interest because it does not exhibit the canonical facially amphiphilic orientation of side chains when in a helical orientation. Different analogs of AP3 were synthesized in which length, charge identity, and Trp position were varied to investigate the sequence-structure and activity relationship. We performed biophysical and microbiological characterization using fluorescence spectroscopy, CD spectroscopy, vesicle leakage assays, bacterial membrane permeabilization assays, and minimal inhibitory concentration (MIC) assays. Fluorescence spectroscopy showed that the peptides bind to lipid bilayers to similar extents, while CD spectra show the peptides adopt helical conformations. All five peptides tested in this study exhibited binding to model lipid membranes, while the truncated peptides showed no measurable antimicrobial activity. The most active peptide proved to be the parent peptide AP3 with the highest degree of leakage and bacterial membrane permeabilization. Moreover, it was found that the ability to permeabilize model and bacterial membranes correlated most closely with the ability to predict antimicrobial activity.

Investigation of the structure‐activity relationship in ponericin L1 from Neoponera goeldii

“…Like L1, numerous venom peptides do not have a strictly cationic face, but instead a polar face with cationic and polar‐uncharged residues. [ 40,48,49 ]…”

“…Using a series of well‐characterized biophysical/spectroscopic measurements as well as microbiological and cellular assays, the activity of these L1 sequences was investigated. [ 14,17,38‐44 ] Due to the inherent environmental sensitivity of Trp fluorescence emission, the native Trp in the L1 sequence (W6) was exploited as a spectroscopic probe for the oligomerization in solution, bilayer binding, and bilayer orientation of peptides. Antibacterial, bacterial membrane permeabilization, and hemolysis assays were also conducted to evaluate the activity of these sequences against natural biological membranes.…”

Correlating Lipid Membrane Permeabilities of Imidazolium Ionic Liquids with Their Cytotoxicities on Yeast, Bacterial, and Mammalian Cells

“…Vaden and coworkers have previously reported a fluorescence lifetime-based leakage assay to quantify IL permeabilities in model lipid bilayer vesicles [11]. This assay, in which the environment-sensitive fluorescence lifetime of a fluorophore entrapped in vesicles reports on membrane leakage, is similar to the well-known calcein assay [41,42] but is very sensitive to small amounts of leakage. To understand IL membrane permeability for real cellular applications, model lipid vesicle membrane permeability results should be correlated with permeability, cytotoxicity, and antibiotic activities.…”

Quaternized Amphiphilic Block Copolymers as Antimicrobial Agents

“…It is known that nonionic surfactants such as Triton can exist as both micelles and monomers in solution and interfere with protein-lipid and lipid-lipid interaction of cell membrane as lysis agents [33]. On the other hand, the prepared PS-b-PIN, as an amphiphilic functional group, is theoretically stronger lysis agent for RBCs membranes, as it did for NIH 3T3 cell membranes [34]. The ability to destroy the RBCs and NIH 3T3 cell membranes might be attributed to the difference in the membrane structure and composition.…”