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Published on: August 11, 2018
Antimicrobial peptide magainin 2-induced rupture of single giant unilamellar vesicles comprising E. coli polar lipids
Md Masum Billah1, Md Mamun Or Rashid1, Marzuk Ahmed1
1Integrated Bioscience Section, Graduate School of Science and Technology, Shizuoka University, Shizuoka 422-8529, Japan.
Abstract:
Most antimicrobial peptides (AMPs) damage the cell membrane of bacterial cells and induce rapid leakage of the internal cell contents, which is a main cause of their bactericidal activity. One of the AMPs, magainin 2 (Mag), forms nanopores in giant unilamellar vesicles (GUVs) comprising phosphatidylcholine (PC) and phosphatidylglycerol (PG), inducing leakage of fluorescent probes. In this study, to elucidate the Mag-induced pore formation in lipid bilayer region in E. coli cell membrane, we examined the interaction of Mag with single GUVs comprising E. coli polar lipids (E. coli-lipid-GUVs). First, we investigated the Mag-induced leakage of a fluorescent probe AF488 from single E. coli-lipid-GUVs, and found that Mag caused rupture of GUVs, inducing rapid AF488 leakage. The rate constant of Mag-induced GUV rupture increased with the Mag concentration. Using fluorescence microscopy with a time resolution of 5 ms, we revealed the GUV rupture process: first, a small micropore was observed in the GUV membrane, then the pore radius increased within 50 ms without changing the GUV diameter, the thickness of the membrane at the pore rim concomitantly increased, and eventually membrane aggregates were formed. Mag bound to only the outer monolayer of the GUV before GUV rupture, which increased the area of the GUV bilayer. We also examined the physical properties of E. coli-lipid-GUVs themselves. We found that the rate constant of the constant tension-induced rupture of E. coli-lipid-GUVs was higher than that of PG/PC-GUVs. Based on these results, we discussed the Mag-induced rupture of E. coli-lipid-GUVs and its mechanism.
Insights
Antimicrobial peptide magainin 2 (Mag) ruptures bacterial membranes by forming pores, causing rapid leakage of cellular contents. This study reveals Mag
Area of Science:
- Membrane biophysics
- Antimicrobial peptide research
- Bacterial cell membrane interactions
Background:
- Antimicrobial peptides (AMPs) are crucial in innate immunity and exhibit bactericidal activity primarily through cell membrane damage.
- Magainin 2 (Mag), a well-studied AMP, is known to form nanopores in lipid bilayers, leading to leakage and cell death.
- Understanding the precise mechanism of AMP-induced membrane disruption is vital for developing new antimicrobial strategies.
Purpose of the Study:
- To elucidate the mechanism of magainin 2 (Mag)-induced pore formation and rupture in *E. coli* model cell membranes.
- To investigate the interaction of Mag with giant unilamellar vesicles (GUVs) composed of *E. coli* polar lipids.
- To characterize the dynamics of GUV rupture and the physical changes in the membrane upon Mag interaction.
Main Methods:
- Utilized giant unilamellar vesicles (GUVs) composed of *E. coli* polar lipids to mimic bacterial membranes.
- Investigated Mag-induced leakage of the fluorescent probe AF488 from single *E. coli*-lipid-GUVs.
- Employed time-resolved fluorescence microscopy (5 ms resolution) to observe the dynamic process of GUV rupture.
- Measured physical properties of *E. coli*-lipid-GUVs, including tension-induced rupture rates.
Main Results:
- Mag induced rapid rupture of *E. coli*-lipid-GUVs, leading to significant leakage of AF488.
- The rate of GUV rupture increased with Mag concentration.
- Microscopy revealed a dynamic pore formation process: initial micropore formation, rapid pore expansion, increased membrane thickness at the pore rim, and eventual membrane aggregate formation.
- Mag binding to the outer monolayer preceded rupture and increased the bilayer area.
- *E. coli*-lipid-GUVs exhibited a higher rate constant for tension-induced rupture compared to standard PC/PG GUVs.
Conclusions:
- Mag induces rapid rupture of *E. coli* model membranes through a dynamic pore formation mechanism.
- The observed membrane dynamics, including pore expansion and thickening, are key to Mag's bactericidal action.
- The distinct physical properties of *E. coli* membranes may influence their susceptibility to AMP-induced disruption.
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