Assembly-Induced Membrane Selectivity of Artificial Model Peptides through Entropy-Enthalpy Competition.

Peptide design and drug development offer a promising solution for combating serious diseases or infections. In this study, using an AI-human negotiation approach, we have designed a class of minimal model peptides against tuberculosis (TB), among which K7W6 exhibits potent efficacy attributed to its assembly-induced function. Comprising lysine and tryptophan with an amphiphilic α-helical structure, the K7W6 sequence exhibits robust activity against various infectious bacteria causing TB (including clinically isolated and drug-resistant strains) both in vitro and in vivo. Moreover, it synergistically enhances the effectiveness of the first-line antibiotic rifampicin while displaying low potential for inducing drug resistance and minimal toxicity toward mammalian cells. Biophysical experiments and simulations elucidate that K7W6's exceptional performance can be ascribed to its highly selective and efficient membrane permeabilization activity induced by its distinctive self-assembly behavior. Additionally, these assemblies regulate the interplay between enthalpy and entropy during K7W6-membrane interaction, leading to the peptide's two-step mechanism of membrane interaction. These findings provide valuable insights into rational design principles for developing advanced peptide-based drugs while uncovering the functional role played by assembly.

Heterogeneous Structural Disturbance of Cell Membrane by Peptides with Modulated Hydrophobic Properties.

Extensive effort has been devoted to developing new clinical therapies based on membrane-active peptides (MAPs). Previous models on the membrane action mechanisms of these peptides mostly focused on the MAP−membrane interactions in a local region, while the influence of the spatial heterogeneity of the MAP distribution on the membrane was much ignored. Herein, three types of natural peptide variants, AS4-1, AS4-5, and AS4-9, with similar amphiphilic α-helical structures but distinct hydrophobic degrees (AS4-1 < AS4-5 < AS4-9) and net charges (+9 vs. +7 vs. +5), were used to interact with a mixed phosphatidylcholine (PC) and phosphatidylglycerol (PG) membrane. A combination of giant unilamellar vesicle (GUV) leakage assays, atomic force microscopy (AFM) characterizations, and molecular dynamics (MD) simulations demonstrated the coexistence of multiple action mechanisms of peptides on a membrane, probably due to the spatially heterogeneous distribution of peptides on the membrane surface. Specifically, the most hydrophobic peptide (i.e., AS4-9) had the strongest membrane binding, perturbation, and permeabilization effects, leading to the formation of large peptide−lipid aggregates (10 ± 5 nm in height and 150 ± 50 nm in size), as well as continuous fragments and ridges on the supported membrane surface. The AS4-5 peptides, with a half-hydrophilic and half-hydrophobic structure, induced membrane lysis in addition to reconstruction. The most hydrophilic peptide AS4-1 only exhibited unstable binding on the supported membrane surface. These results demonstrate the heterogeneous structural disturbance of model cell membranes by amphiphilic α-helical peptides, which could be significantly strengthened by increasing the degree of hydrophobicity and/or local number density of peptides. This work provides support for the modulation of the membrane activity of MAPs by adjusting their hydrophobicity and local concentration.

A real-time and in-situ monitoring of the molecular interactions between drug carrier polymers and a phospholipid membrane.

The dynamic interactions between drug carrier molecules and a cell membrane can not be ignored in their clinical use. Here a simple, label-free and non-invasive approach, photo-voltage transient method, combined with the atomic force microscopy, dynamic giant unilamellar vesicle leakage assay and cytotoxicity method, was employed for a real-time monitoring of the interaction process. Two representative polymer molecules, polyoxyethylene (35) lauryl ether (Brij35) and polyvinylpyrrolidone (PVPk30), were taken as examples to interact with a phospholipid bilayer membrane in a low ionic strength and neutral pH condition. Brij35 demonstrated an adsorption-accumulation-permeabilization dominated process under the modulation of polymer concentration in the solution. In contrast, PVPk30 performed a dynamic balance between adsorption-desorption of the molecules and/or permeabilization-resealing of the membrane. Such difference explains the high and low cytotoxicity of them, respectively, in the living cell tests. Briefly, through combining the photo-voltage approach with conventional fluorescent microscopy method, this work demonstrates new ideas on the time and membrane actions of polymer surfactants which should be taken into account for their biomedical applications.

Hydrophobicity Determines the Bacterial Killing Rate of α-Helical Antimicrobial Peptides and Influences the Bacterial Resistance Development.

Rapid antimicrobial action is an important advantage of antimicrobial peptides (AMPs) over antibiotics, which is also a reason for AMPs being less likely to induce bacterial resistance. However, the structural parameters and underlying mechanisms affecting the bacterial killing rate of AMPs remain unknown. In this study, we performed a structure-activity relationship (SAR) study using As-CATH4 and 5 as templates. We revealed that hydrophobicity, rather than other characteristics, is the critical structural parameter determining the bacterial killing rate of α-helical AMPs. With the hydrophobicity increase, the action rates of AMPs including bacterial binding, lipopolysaccharides neutralization, and outer and inner membrane permeabilization increased. Additionally, the higher hydrophobic AMPs with enhanced bacterial killing rates possess better in vivo therapeutic potency and a lower propensity to induce bacterial resistance. These findings revealed the importance of the bacterial killing rate for AMPs and are of great significance to the design and optimization of AMP-related drugs.