AFM Study of Nanoscale Membrane Perturbation Induced by Antimicrobial Lipopeptide C14 KYR.

Sawinee Nasompag,Saengrawee Thammawithan,Pawinee Siritongsuk,Panchika Prangkio,Rina Patramanon,Chomdao Sinthuvanich,Oranee Srichaiyapol,Terri A Camesano

doi:10.3390/membranes11070495

Abstract

Lipopeptides have been extensively studied as potential antimicrobial agents. In this study, we focused on the C14-KYR lipopeptide, a modified version of the KYR tripeptide with myristic acid at the N-terminus. Here, membrane perturbation of live E. coli treated with the parent KYR and C14-KYR peptides was compared at the nanoscale level using AFM imaging. AFM analyses, including average cellular roughness and force spectroscopy, revealed the severe surface disruption mechanism of C14-KYR. A loss of surface roughness and changes in topographic features included membrane shrinkage, periplasmic membrane separation from the cell wall, and cytosolic leakage. Additional evidence from synchrotron radiation FTIR microspectroscopy (SR-FTIR) revealed a marked structural change in the membrane component after lipopeptide attack. The average roughness of the E. coli cell before and after treatment with C14-KYR was 129.2 ± 51.4 and 223.5 ± 14.1 nm, respectively. The average rupture force of the cell treated with C14-KYR was 0.16 nN, four times higher than that of the untreated cell. Our study demonstrates that the mechanistic effect of the lipopeptide against bacterial cells can be quantified through surface imaging and adhesion force using AFM.

Highlights

Natural lipopeptides (LiPs) are molecules consisting of a peptide linked to a lipid moiety commonly produced non-ribosomally in bacteria and fungi during cultivation in the presence of carbon sources
We focused on the C14 KYR lipopeptide, a KYR peptide with amidation at the C-terminus and acylation with myristic acid at the N-terminus
The E. coli HB101 was chosen as a bacterial model for studying changes in the bacterial surface after treatment with LiP

Summary

Introduction

Natural lipopeptides (LiPs) are molecules consisting of a peptide linked to a lipid moiety commonly produced non-ribosomally in bacteria and fungi during cultivation in the presence of carbon sources. Compared to conventional antibiotics or antimicrobial peptides, bacterial resistance is generally rare in LiPs [1,2,3,4]. Synthetic LiPs have recently attracted considerable interest as the alternative to antibiotics. Most LiPs consist of a short linear or cyclic peptide covalently linked to a fatty acid (8–18 carbon chain) tail at the N-terminus. Among synthetic LiPs tested in the literature, the shortest LiPs with the highest bactericidal activity is composed of 2–4 amino acids linked to a fatty acid with a 8- to 16-carbon chain length [1,4,5]. The interaction between the amphiphilic peptide and the hydrophilic head of the fatty acyl chains of phospholipids leads to bacterial membrane insertion, and disruption of the physical integrity of the membrane, resulting in the leakage of cellular materials and eventually cell death [1,4]

Methods

Results

Conclusion