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Antibiotics are losing effectiveness as bacteria become resistant to conventional drugs. To find new alternatives, antimicrobial peptides (AMPs) are rationally designed with different lengths, charges, hydrophobicities (H), and hydrophobic moments (μH), containing only three types of amino acids: arginine, tryptophan, and valine. Six AMPs with low minimum inhibitory concentrations (MICs) and <25% toxicity to mammalian cells are selected for biophysical studies. Their secondary structures are determined using circular dichroism (CD), which finds that the % α‐helicity of AMPs depends on composition of the lipid model membranes (LMMs): gram‐negative (G(−)) inner membrane (IM) >gram‐positive (G(+))> Euk33 (eukaryotic with 33 mol% cholesterol). The two most effective peptides, E2‐35 (16 amino acid [AA] residues) and E2‐05 (22 AAs), are predominantly helical in G(–) IM and G(+) LMMs. AMP/membrane interactions such as membrane elasticity, chain order parameter, and location of the peptides in the membrane are investigated by low‐angle and wide‐angle X‐ray diffuse scattering (XDS). It is found that headgroup location correlates with efficacy and toxicity. The membrane bending modulusKCdisplays nonmonotonic changes due to increasing concentrations of E2‐35 and E2‐05 in G(–) and G(+) LMMs, suggesting a bacterial killing mechanism where domain formation causes ion and water leakage.
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Gram-Positive Bacterial Membrane-Based Biosensor for Multimodal Investigation of Membrane–Antibiotic Interactions
As membrane-mediated antibiotic resistance continues to evolve in Gram-positive bacteria, the development of new approaches to elucidate the membrane properties involved in antibiotic resistance has become critical. Membrane vesicles (MVs) secreted by the cytoplasmic membrane of Gram-positive bacteria contain native components, preserving lipid and protein diversity, nucleic acids, and sometimes virulence factors. Thus, MV-derived membrane platforms present a great model for Gram-positive bacterial membranes. In this work, we report the development of a planar bacterial cytoplasmic membrane-based biosensor using MVs isolated from the Bacillus subtilis WT strain that can be coated on multiple surface types such as glass, quartz crystals, and polymeric electrodes, fostering the multimodal assessment of drug–membrane interactions. Retention of native membrane components such as lipoteichoic acids, lipids, and proteins is verified. This biosensor replicates known interaction patterns of the antimicrobial compound, daptomycin, with the Gram-positive bacterial membrane, establishing the applicability of this platform for carrying out biophysical characterization of the interactions of membrane-acting antibiotic compounds with the bacterial cytoplasmic membrane. We report changes in membrane viscoelasticity and permeability that correspond to partial membrane disruption when calcium ions are present with daptomycin but not when these ions are absent. This biomembrane-based biosensing platform enables an assessment of membrane biophysical characteristics during exposure to antibiotic drug candidates to aid in identifying compounds that target membrane disruption as a mechanism of action.
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- Award ID(s):
- 1719875
- PAR ID:
- 10549136
- Publisher / Repository:
- MDPI
- Date Published:
- Journal Name:
- Biosensors
- Volume:
- 14
- Issue:
- 1
- ISSN:
- 2079-6374
- Page Range / eLocation ID:
- 45
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.3390/bios14010045
