Membrane Charge and Packing Govern the Interfacial Insertion, Structure, and Dynamics of Ctx(Ile21)-Ha and Its TOAC-Labeled Analogues in Biomimetic Liposomes
4 affiliations
- 1State University of Northern Rio de Janeiro (UENF) , , Campos dos Goytacazes , ,
- 2São Paulo State University (UNESP) , , São José do Rio Preto, ,
- 3São Paulo State University (UNESP) , , Jaboticabal, ,
- 4São Paulo State University (UNESP) , , Tupã, ,
Abstract
Cationic antimicrobial peptides are promising candidates to combat antifungal resistance. However, a detailed understanding of their membranotropic activity is required for targeted applications and sequence engineering. Here, we investigated the native peptide Ctx(Ile21)-Ha, derived from the Brazilian Cerrado tree frog Boana albopunctata, and its analogues site-specifically labeled with the paramagnetic amino acid TOAC (2,2,6,6-tetramethylpiperidine-N-oxyl-4-amino-4-carboxylic acid). Peptide interactions with biomimetic monolayers and liposomes mimicking human, fungal, and bacterial membranes were investigated using Langmuir monolayers, circular dichroism, and electron spin resonance (ESR). Anionic lipids, particularly palmitoyl-oleoyl-phosphatidylglycerol (POPG) and palmitoyl-oleoyl-phosphatidylserine (POPS), strongly enhanced peptide penetration into lipid monolayers and promoted substantial α-helical folding in large unilamellar vesicles (LUVs). The distinct monolayer responses of POPG- and POPS-containing membranes, despite their comparable α-helical contents, indicate that anionic headgroup chemistry modulates peptide–membrane interactions beyond electrostatic attraction. Conversely, cholesterol and ergosterol reduced peptide penetration and attenuated membrane-induced helix formation due to sterol-induced membrane packing. Site-directed ESR revealed lipid-dependent motional regimes for [TOAC2]Ctx(Ile21)-Ha and [TOAC13]Ctx(Ile21)-Ha, with the central region of the peptide becoming substantially more immobilized than the N-terminus upon membrane interaction. CD-derived membrane-association isotherms further showed that anionic membranes favor the coupled association/folding response, whereas comparison across the complementary techniques indicates that membrane association, peptide folding, and membrane penetration are mechanistically related but distinct steps in peptide–membrane interactions. Overall, the membrane activity of Ctx(Ile21)-Ha is best described as a charge- and packing-modulated process, in which electrostatic interactions favor peptide binding and folding, whereas membrane order modulates membrane penetration and local peptide dynamics.
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BibTeX
@article{magalhaes2026membrane,
title = {{Membrane Charge and Packing Govern the Interfacial Insertion, Structure, and Dynamics of Ctx(Ile21)-Ha and Its TOAC-Labeled Analogues in Biomimetic Liposomes}},
author = {Fadi S. S. Magalhães and Dayane S. Alvares and Rayssa P. R. Costa and Luis G. M. Basso and Eduardo F. Vicente},
journal = {ACS Omega},
year = {2026},
month = oct,
doi = {10.1021/acsomega.6c07092},
url = {https://pubs.acs.org/acsodf/article/doi/10.1021/acsomega.6c07092/5453286/Membrane-Charge-and-Packing-Govern-the-Interfacial},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Membrane Charge and Packing Govern the Interfacial Insertion, Structure, and Dynamics of Ctx(Ile21)-Ha and Its TOAC-Labeled Analogues in Biomimetic Liposomes
AU - Fadi S. S. Magalhães
AU - Dayane S. Alvares
AU - Rayssa P. R. Costa
AU - Luis G. M. Basso
AU - Eduardo F. Vicente
PY - 2026
DA - 2026/10/17
JO - ACS Omega
DO - 10.1021/acsomega.6c07092
UR - https://pubs.acs.org/acsodf/article/doi/10.1021/acsomega.6c07092/5453286/Membrane-Charge-and-Packing-Govern-the-Interfacial
AB - Cationic antimicrobial peptides are promising candidates to combat antifungal resistance. However, a detailed understanding of their membranotropic activity is required for targeted applications and sequence engineering. Here, we investigated the native peptide Ctx(Ile21)-Ha, derived from the Brazilian Cerrado tree frog Boana albopunctata, and its analogues site-specifically labeled with the paramagnetic amino acid TOAC (2,2,6,6-tetramethylpiperidine-N-oxyl-4-amino-4-carboxylic acid). Peptide interactions with biomimetic monolayers and liposomes mimicking human, fungal, and bacterial membranes were investigated using Langmuir monolayers, circular dichroism, and electron spin resonance (ESR). Anionic lipids, particularly palmitoyl-oleoyl-phosphatidylglycerol (POPG) and palmitoyl-oleoyl-phosphatidylserine (POPS), strongly enhanced peptide penetration into lipid monolayers and promoted substantial α-helical folding in large unilamellar vesicles (LUVs). The distinct monolayer responses of POPG- and POPS-containing membranes, despite their comparable α-helical contents, indicate that anionic headgroup chemistry modulates peptide–membrane interactions beyond electrostatic attraction. Conversely, cholesterol and ergosterol reduced peptide penetration and attenuated membrane-induced helix formation due to sterol-induced membrane packing. Site-directed ESR revealed lipid-dependent motional regimes for [TOAC2]Ctx(Ile21)-Ha and [TOAC13]Ctx(Ile21)-Ha, with the central region of the peptide becoming substantially more immobilized than the N-terminus upon membrane interaction. CD-derived membrane-association isotherms further showed that anionic membranes favor the coupled association/folding response, whereas comparison across the complementary techniques indicates that membrane association, peptide folding, and membrane penetration are mechanistically related but distinct steps in peptide–membrane interactions. Overall, the membrane activity of Ctx(Ile21)-Ha is best described as a charge- and packing-modulated process, in which electrostatic interactions favor peptide binding and folding, whereas membrane order modulates membrane penetration and local peptide dynamics.
SN - 2470-1343
ER -