Investigations of microbial adaptation to singular, binary, and fully formulated quaternary ammonium compounds.

The study was conducted to inform risk assessments concerning microbial exposure to quaternary ammonium biocides (QUATs) by investigating their effects on 10 microbial strains of hygiene relevance. Biocides were divided into three categories: simple aqueous solutions, biocide mixtures, and formulated biocides. Organisms were grown in the presence of biocides for 10 generations and then subsequently for another 10 generations in biocide-free media. Control organisms were passaged 20 times in biocide-free media. Strains were then assessed for biocide and antibiotic susceptibility, changes in growth dynamics, and single nucleotide polymorphisms (SNPs). Biocide mixtures demonstrated greater antimicrobial potency than singular and formulated biocides. Susceptibility changes of under twofold were observed for all biocides tested. Susceptibility decreased significantly for organisms passaged with singular biocides (1.29- to 4.35-fold) and biocide mixtures (1.4- to 1.5-fold), but not for formulated biocides (1.3- to 1.84-fold) compared to controls. Antibiotic susceptibility both increased and decreased in passaged organisms, with heightened susceptibility occurring more frequently in the singular biocide group. Changes in susceptibility and growth dynamics were similar in the passaged and unexposed controls for fitness measures of adapted bacteria; there were no significant differences between biocide groups, but significantly lower generation and doubling times in organisms exposed to singular biocides. Similar frequencies in SNPs occurred for the three biocide groups and unexposed controls. While some adaptations occurred, particularly with singular biocides, the impact on antibiotic resistance and genomic mutations was limited. These findings suggest that the use of formulated QUATs may pose a comparatively lower risk for antimicrobial resistance.IMPORTANCEBiocides are used globally to control microbial growth and effective assessment of the risks and benefits of their use is therefore a high priority. Much of the data used to assess risk has been based on sub-lethal exposure of bacteria to singular biocides in simple aqueous solutions. This work builds on limited prior realism-based studies to demonstrate enhanced potency in biocidal mixtures; the mitigation of resistance selection by formulations and inconsistent cross-resistance effects with both increases and decreases in susceptibility for a wide range of antibiotics. These data can be used to better inform risk assessments of biocide deployment.

Chiral inversion induced by aromatic interactions in short peptide assembly.

Although hydrophobic interactions provide the main driving force for initial peptide aggregation, their role in regulating suprastructure handedness of higher-order architectures remains largely unknown. We here interrogate the effects of hydrophobic amino acids on handedness at various assembly stages of peptide amphiphiles. Our studies reveal that relative to aliphatic side chains, aromatic side chains set the twisting directions of single ß-strands due to their strong steric repulsion to the backbone, and upon packing into multi-stranded ß-sheets, the side-chain aromatic interactions between strands form the aromatic ladders with a directional preference. This ordering not only leads to parallel ß-sheet arrangements but also induces the chiral flipping over of single ß-strands within a ß-sheet. In contrast, the lack of orientational hydrophobic interactions in the assembly of aliphatic peptides implies no chiral inversion upon packing into ß-sheets. This study opens an avenue to harness peptide aggregates with targeted handedness via aromatic side-chain interactions.

Adsorption of monoclonal antibody fragments at the water-oil interface: A coarse-grained molecular dynamics study.

Monoclonal antibodies (mAbs) can undergo structural changes due to interaction with oil-water interfaces during storage. Such changes can lead to aggregation, resulting in a loss of therapeutic efficacy. Therefore, understanding the microscopic mechanism controlling mAb adsorption is crucial to developing strategies that can minimize the impact of interfaces on the therapeutic properties of mAbs. In this study, we used MARTINI coarse-grained molecular dynamics simulations to investigate the adsorption of the Fab and Fc domains of the monoclonal antibody COE3 at the oil-water interface. Our aim was to determine the regions on the protein surface that drive mAb adsorption. We also investigate the role of protein concentration on protein orientation and protrusion to the oil phase. While our structural analyses compare favorably with recent neutron reflectivity measurements, we observe some differences. Unlike the monolayer at the interface predicted by neutron reflectivity experiments, our simulations indicate the presence of a secondary diffused layer near the interface. We also find that under certain conditions, protein-oil interaction can lead to a considerable distortion in the protein structure, resulting in enhanced adsorption behavior.

Unlocking roles of cationic and aromatic residues in peptide amphiphiles in treating drug-resistant gram-positive pathogens.

Multidrug resistance (MDR) is a rising threat to global health because the number of essential antibiotics used for treating MDR infections is increasingly compromised. In this work we report a group of new amphiphilic peptides (AMPs) derived from the well-studied G3 (G(IIKK)3I-NH2) to fight infections from Gram-positive bacteria including susceptible Staphylococcus aureus and methicillin-resistant Staphylococcus aureus (MRSA), focusing on membrane interactions. Time-dependent killing experiments revealed that substitutions of II by WW (GWK), II by FF (GFK) and KK by RR (GIR) resulted in improved bactericidal efficiencies compared to G3 (GIK) on both S. aureus and MRSA, with the order of GWK > GIR > GFK > GIK. Electronic microscopy imaging revealed structural disruptions of AMP binding to bacterial cell walls. Fluorescence assays including AMP binding to anionic lipoteichoic acids (LTA) in cell-free and cell systems indicated concentration and time-dependent membrane destabilization associated with bacterial killing. Furthermore, AMP's binding to anionic plasma membrane via similar fluorescence assays revealed a different extent of membrane depolarization and leakage. These observations were supported by the penetration of AMPs into the LTA barrier and the subsequent structural compromise to the cytoplasmic membrane as revealed from SANS (small angle neutron scattering). Both experiments and molecular dynamics (MD) simulations revealed that GWK and GIR could make the membrane more rigid but less effective in diffusive efficiency than GIK and GFK through forming intramembrane peptide nanoaggregates associated with hydrophobic mismatch and formation of fluidic and rigid patches. The reported peptide-aggregate-induced phase-separation emerged as a crucial factor in accelerated membrane disintegration and fast bacterial killing. This work has demonstrated the importance of membrane interactions to the development of more effective AMPs and the relevance of the approaches as reported in assisting this area of research.

The viscoelasticity of high concentration monoclonal antibodies using particle tracking microrheology.

The viscoelasticity of monoclonal antibodies (mAbs) is important during their production, formulation, and drug delivery. High concentration mAbs can provide higher efficacy therapeutics (e.g., during immunotherapy) and improved efficiency during their production (economy of scale during processing). Two humanized mAbs were studied (mAb-1 and mAb-2) with differing isoelectric points. Using high speed particle tracking microrheology, we demonstrated that the mAb solutions have significant viscoelasticities above concentrations of 40 mg/ml. Power law viscoelasticity was observed over the range of time scales (10-4-1 s) probed for the high concentration mAb suspensions. The terminal viscosity demonstrated an exponential dependence on mAb concentration (a modified Mooney relationship) as expected for charged stabilized Brownian colloids. Gelation of the mAbs was explored by lowering the pH of the buffer and a power law scaling of the gelation transition was observed, i.e., the exponent of the anomalous diffusion of the probe particles scaled inversely with the gelation time.

Unraveling the Microscopic Mechanism of Molecular Ion Interaction with Monoclonal Antibodies: Impact on Protein Aggregation.

Understanding and predicting protein aggregation represents one of the major challenges in accelerating the pharmaceutical development of protein therapeutics. In addition to maintaining the solution pH, buffers influence both monoclonal antibody (mAb) aggregation in solution and the aggregation mechanisms since the latter depend on the protein charge. Molecular-level insight is necessary to understand the relationship between the buffer-mAb interaction and mAb aggregation. Here, we use all-atom molecular dynamics simulations to investigate the interaction of phosphate (Phos) and citrate (Cit) buffer ions with the Fab and Fc domains of mAb COE3. We demonstrate that Phos and Cit ions feature binding mechanisms, with the protein that are very different from those reported previously for histidine (His). These differences are reflected in distinctive ion-protein binding modes and adsorption/desorption kinetics of the buffer molecules from the mAb surface and result in dissimilar effects of these buffer species on mAb aggregation. While His shows significant affinity toward hydrophobic amino acids on the protein surface, Phos and Cit ions preferentially bind to charged amino acids. We also show that Phos and Cit anions provide bridging contacts between basic amino acids in neighboring proteins. The implications of such contacts and their connection to mAb aggregation in therapeutic formulations are discussed.

Combination of a pH-responsive peptide amphiphile and a conventional antibiotic in treating Gram-negative bacteria.

BACKGROUND: Clinical treatments ofgastric infections using antibiotics suffer from the undesired killing of commensal bacteria and emergence of antibiotic resistance. It is desirable to develop pH-responsive antimicrobial peptides (AMPs) that kill pathogenic bacteria such as H. pyloriand resistant E. coli under acidic environment with minimal impact to commensal bacteria whilst not causing antibiotic resistance. EXPERIMENTS: Using a combined approach of cell assays, molecular dynamics (MD) simulations and membrane models facilitating biophysical and biochemical measurements including small angle neutron scattering (SANS), we have characterized the pH-responsive physiochemical properties and antimicrobial performance of two amphiphilic AMPs, GIIKDIIKDIIKDI-NH2 and GIIKKIIDDIIKKI-NH2 (denoted as 3D and 2D, respectively), that were designed by selective substitutions of cationic residues of Lys (K) in the extensively studied AMP G(IIKK)3I-NH2 with anionic residue Asp (D). FINDINGS: Whilst 2D kept non-ordered coils across the entire pH range studied, 3D displayed a range of secondary structures when pH was shifted from basic to acidic, with distinct self-assembly into nanofibers in aqueous environment. Further experimental and modeling studies revealed that the AMPs interacted differently with the inner and outer membranes of Gram-negative bacteria in a pH-responsive manner and that the structural features characterized by membrane leakage and intramembrane nanoaggregates revealed from fluorescence spectroscopy and SANS were well linked to antimicrobial actions. Different antimicrobial efficacies of 2D and 3D were underlined by the interplay between their ability to bind to the outer membrane lipid LPS (lipopolysaccharide), outer membrane permeability change and inner membrane depolarization and leakage. Furthermore, AMP's binding with the inner membrane under acidic condition caused both the dissipation of membrane potential (Δψ) and the continuous dissipation of transmembrane ΔpH, with Δψ and ΔpH being the key components of the proton motive force. Combinations of antibiotic (Minocycline) with the pH-responsive AMP generated the synergistic effects against Gram-negative bacteria only under acidic condition. These features are crucial to target applications to gastric infections, anti-acne and wound healing.

Mechanistic Insights into the Adsorption of Monoclonal Antibodies at the Water/Vapor Interface.

Monoclonal antibodies (mAbs) are active components of therapeutic formulations that interact with the water-vapor interface during manufacturing, storage, and administration. Surface adsorption has been demonstrated to mediate antibody aggregation, which leads to a loss of therapeutic efficacy. Controlling mAb adsorption at interfaces requires a deep understanding of the microscopic processes that lead to adsorption and identification of the protein regions that drive mAb surface activity. Here, we report all-atom molecular dynamics (MD) simulations of the adsorption behavior of a full IgG1-type antibody at the water/vapor interface. We demonstrate that small local changes in the protein structure play a crucial role in promoting adsorption. Also, interfacial adsorption triggers structural changes in the antibody, potentially contributing to the further enhancement of surface activity. Moreover, we identify key amino acid sequences that determine the adsorption of antibodies at the water-air interface and outline strategies to control the surface activity of these important therapeutic proteins.

Unraveling How Membrane Nanostructure Changes Impact the Eye Irritation of Nonionic Alkyl Ethoxylate Surfactants.

Nonionic surfactants used in agri-spraying processes may cause varying degrees of corneal irritation when they come in direct contact with farmers' eyes, and the exact irritations are thought to be determined by how surfactants interact with corneal cell membranes. However, how nonionic surfactants interact with cell membranes at the molecular and nano levels remains largely unexplored. In this study, the interactions between nonionic surfactants (alkyl ethoxylate, C12Em) and lipid membranes were examined by membrane permeability measurement, quartz crystal microbalance with dissipation, dual polarization interferometry, confocal laser scanning microscopy, and neutron reflection, aiming to reveal complementary structural features at the molecular and nano levels. Apart from the extremely hydrophobic surfactant C12E2, all nonionic surfactants studied could penetrate the model cell membrane composed of a phosphocholine lipid bilayer. Nonionic surfactants with intermediate amphiphilicity (C12E6) rapidly fused into the lipid membrane and stimulated the formation of pores across the lipid bilayer, consistent with the cytoplasm leakage and fast cell necrosis observed from the cytotoxicity study of corneal cells. In comparison, while hydrophobic and hydrophilic surfactants [those with long and short ethoxylates (C12E4,12,23)] could cause mild structural alteration to the outer lipid layer of the membrane, these structural changes were insufficient to elicit large cytoplasmic leakage rapidly and instead cell death occurred over longer periods of time due to changes in the membrane permeability. These results reveal the strong link of surfactant-lipid membrane interactions to surfactant cytotoxicity and the association with amphiphilicity of nonionic surfactants.

Structure and interaction of therapeutic proteins in solution: a combined simulation and experimental study.

The aggregation of therapeutic proteins in solution has attracted significant interest, driving efforts to understand the relationship between microscopic structural changes and protein-protein interactions determining aggregation processes in solution. Additionally, there is substantial interest in being able to predict aggregation based on protein structure as part of molecular developability assessments. Molecular Dynamics provides theoretical tools to complement experimental studies and to interrogate and identify the microscopic mechanisms determining aggregation. Here we perform all-atom MD simulations to study the structure and inter-protein interaction of the Fab and Fc fragments of the monoclonal antibody (mAb) COE3. We unravel the role of ion-protein interactions in building the ionic double layer and determining effective inter-protein interaction. Further, we demonstrate, using various state-of-the-art force fields (charmm, gromos, amber, opls/aa), that the protein solvation, ionic structure and protein-protein interaction depend significantly on the force field parameters. We perform SANS and Static Light Scattering experiments to assess the accuracy of the different forcefields. Comparison of the simulated and experimental results reveal significant differences in the forcefields' performance, particularly in their ability to predict the protein size in solution and inter-protein interactions quantified through the second virial coefficients. In addition, the performance of the forcefields is correlated with the protein hydration structure.

Neutron reflection and scattering in characterising peptide assemblies.

Self-assemblies of de novo designed short peptides at interface and in bulk solution provide potential platforms for developing applications in many medical and technological areas. However, characterising how bioinspired supramolecular nanostructures evolve with dynamic self-assembling processes and respond to different stimuli remains challenging. Neutron scattering technologies including small angle neutron scattering (SANS) and neutron reflection (NR) can be advantageous and complementary to other state-of-the-art techniques in tracing structural changes under different conditions. With more neutron sources now available, SANS and NR are becoming increasingly popular in studying self-assembling processes of diverse peptide and protein systems, but the difficulty in experimental manipulation and data analysis can deter beginners. This review will introduce the basic theory, general experimental setup and data analysis of SANS and NR, followed by provision of their applications in characterising interfacial and solution self-assemblies of representative peptides and proteins. SANS and NR are remarkably effective in determining the morphological features self-assembled short peptides, especially size and shape transitions as a result of either sequence changes or in response to environmental stimuli, demonstrating the unique capability of NR and SANS in unravelling the interactive processes. These examples highlight the potential of NR and SANS in supporting the development of novel short peptides and proteins as biopharmaceutical candidates in the fight against many diseases and infections that share common features of membrane interactive processes.

Antibacterial effects in blood irradiated with a polychromatic device mediated through reactive oxygen species: possible involvement of haem.

The antibacterial effects of a polychromatic light device designed for intravenous application were assessed in vitro. Staphylococcus aureus, Klebsiella pneumoniae, or Escherichia coli were exposed to a 60-min sequential light cycle comprising 365, 530, and 630 nm wavelengths in circulated sheep blood. Bacteria were quantified by viable counting. The potential involvement of reactive oxygen species in the antibacterial effect was assessed using the antioxidant N-acetylcysteine-amide. A modified device was then used to determine the effects of the individual wavelengths. Exposure of blood to the standard wavelength sequence caused small (c. 0.5 Log 10 CFU) but statistically significant reductions in viable counts for all three bacteria, which were prevented by the addition of N-acetylcysteine-amide. Bacterial inactivation did not occur in blood-free medium, but supplementation with haem restored the moderate bactericidal effect. In single-wavelength experiments, bacterial inactivation occurred only with red (630 nm) light. Concentrations of reactive oxygen species were significantly higher under light stimulation than in unstimulated controls. In summary, exposure of bacteria within blood to a cycle of visible light wavelengths resulted in small but statistically significant bacterial inactivation apparently mediated by a 630 nm wavelength only, via reactive oxygen species possibly generated by excitation of haem groups.

Bioinspired Interfacial Spontaneous Growth of ZnO Nanocatalysts onto Recycled Textiles as a Sustainable Approach for Water Purification.

Zinc oxide, as a commonly used photocatalytic degradation of organic pollutants, typically shows limitations in wastewater treatment, such as aggregation and recycling problems caused by nanoscale dimensions and inappropriate substrates. Anchoring ZnO on substrates is a strategy to obtain stable catalytic performance. Particularly, natural fibers with hollow structures are an attractive alternative for ecological and economical ZnO loading templates, but depositing ZnO onto hollowed fiber surfaces presents a challenge. Here, a straightforward in situ growth method for producing nanostructured zinc oxide on cotton fibers from recycled garments is reported. The modified polydopamine on the fiber surface captures the catalyst required for in situ growth of ZnO and serves as a platform for spontaneous catalytic crystal growth on the fiber surface. The ZnO nanocrystals are uniformly dispersed on the outer and inner walls of cotton fibers, demonstrating excellent durability in wastewater treatments. Moreover, the photocatalytic performance of functional fibers is optimized by doping Ag nanoparticles to improve degradation efficiency. This can extend the prospect of further applications of developed ZnO/fibers in optoelectronics, spintronics, and provide inspiration for recycling and upgrading of used garments.

Investigating the Orientation of an Interfacially Adsorbed Monoclonal Antibody and Its Fragments Using Neutron Reflection.

Interfacial adsorption is a molecular process occurring during the production, purification, transport, and storage of antibodies, with a direct impact on their structural stability and subsequent implications on their bioactivities. While the average conformational orientation of an adsorbed protein can be readily determined, its associated structures are more complex to characterize. Neutron reflection has been used in this work to investigate the conformational orientations of the monoclonal antibody COE-3 and its Fab and Fc fragments at the oil/water and air/water interfaces. Rigid body rotation modeling was found to be suitable for globular and relatively rigid proteins such as the Fab and Fc fragments but less so for relatively flexible proteins such as full COE-3. Fab and Fc fragments adopted a 'flat-on' orientation at the air/water interface, minimizing the thickness of the protein layer, but they adopted a substantially tilted orientation at the oil/water interface with increased layer thickness. In contrast, COE-3 was found to adsorb in tilted orientations at both interfaces, with one fragment protruding into the solution. This work demonstrates that rigid-body modeling can provide additional insights into protein layers at various interfaces relevant to bioprocess engineering.

Exploiting terminal charged residue shift for wide bilayer nanotube assembly.

HYPOTHESIS: Self-assembly of peptides is influenced by both molecular structure and external conditions, which dictate the delicate balance of different non-covalent interactions that driving the self-assembling process. The shifting of terminal charge residue is expected to influence the non-covalent interactions and their interplay, thereby affecting the morphologies of self-assemblies. Therefore, the morphology transition can be realized by shifting the position of the terminal charge residue. EXPERIMENTS: The structure transition from thin nanofibers to giant nanotubes is realized by simply shifting the C-terminal lysine of ultrashort Ac-I3K-NH2 to its N-terminus. The morphologies and detailed structure information of the self-assemblies formed by these two peptides are investigated systemically by a combination of different experimental techniques. The effect of terminal residue on the morphologies of the self-assemblies is well presented and the underlying mechanism is revealed. FINDINGS: Giant nanotubes with a bilayer shell structure can be self-assembled by the ultrashort peptide Ac-KI3-NH2 with the lysine residue close to the N-terminal. The Ac-KI3-NH2 dimerization through intermolecular C-terminal H-bonding promotes the formation of a bola-form geometry, which is responsible for the wide nanotube assembly formation. The evolution process of Ac-KI3-NH2 nanotubes follows the "growing width" model. Such a morphological transformation with the terminal lysine shift is applicable to other analogues and thus provides a facile approach for the self-assembly of wide peptide nanotubes, which can expand the library of good template structures for the prediction of peptide nanostructures.

Impacts of chain and head lengths of nonionic alkyl ethoxylate surfactants on cytotoxicity to human corneal and skin cells in agri-spraying processes.

HYPOTHESIS: Nonionic surfactants are widely used as co-formulants in agrochemical sprays. During spraying, they may come into direct contact with humans and animals, causing irritation in different tissues. However, how the molecular structures of these surfactants affect their toxicity towards human eye and skin at the cellular level has not been well characterised. EXPERIMENT: In this study, the cytotoxicities of two sets of nonionic surfactants (alkyl ethoxylate, CnEm) against human corneal and skin cell lines were examined, with one set composed of varied surfactant head length but fixed tail length (C12E4-23) and the other set oppositely composed (C10-16E6). The cell viability and morphology against different nonionic surfactants for varied exposure times were studied, followed by characterisation of their membrane-lytic ability. FINDING: Nonionic surfactants with intermediate amphiphilicity killed cells rapidly due to their strong membrane-lytic power. Those with weak or strong hydrophobicity exhibited low cytotoxicity but had different modes of action depending on their hydrophobicity. Hydrophobic surfactants were found to adsorb on to cell membranes with no observed structural damage for 2 hr. Hydrophilic surfactants were also found to adsorb on to cell membranes but did cause mild structural changes. While the changes were not sufficient to elicit large cytoplasmic leakage over short periods of time, membrane associations did cause cell shrinkage which eventually resulted in cell death over longer exposure periods. These results revealed that the specific amphiphilic nature of nonionic surfactants played a crucial role in determining their cytotoxicity. This work provided a useful basis for the assessment of amphiphilicity of the nonionic surfactants used in agrochemical sprays by balancing their efficiency, toxicity and environmental impact.

Understanding the Stabilizing Effect of Histidine on mAb Aggregation: A Molecular Dynamics Study.

Histidine, a widely used buffer in monoclonal antibody (mAb) formulations, is known to reduce antibody aggregation. While experimental studies suggest a nonelectrostatic, nonstructural (relating to secondary structure preservation) origin of the phenomenon, the underlying microscopic mechanism behind the histidine action is still unknown. Understanding this mechanism will help evaluate and predict the stabilizing effect of this buffer under different experimental conditions and for different mAbs. We have used all-atom molecular dynamics simulations and contact-based free energy calculations to investigate molecular-level interactions between the histidine buffer and mAbs, which lead to the observed stability of therapeutic formulations in the presence of histidine. We reformulate the Spatial Aggregation Propensity index by including the buffer-protein interactions. The buffer adsorption on the protein surface leads to lower exposure of the hydrophobic regions to water. Our analysis indicates that the mechanism behind the stabilizing action of histidine is connected to the shielding of the solvent-exposed hydrophobic regions on the protein surface by the buffer molecules.

Contrasting impacts of mixed nonionic surfactant micelles on plant growth in the delivery of fungicide and herbicide.

HYPOTHESIS: Nonionic alkyl ethoxylate surfactants are widely used in agrochemicals to facilitate the permeation of systemic herbicides and fungicides across the plant waxy film. Industrial grade surfactants are often highly mixed and how the mixing affects their interactions with pesticides and wax films remains largely unexplored. A better understanding could enable design of mixed nonionic surfactants for herbicides and fungicides to maximize their efficiency and reduce wastage whilst controlling their impact on plant wax films. EXPERIMENT: In this study, nonionic surfactants with general structure n-oxyethylene glycol monododecyl ether (C12En) were used to form surfactant mixtures with the same average ethoxylate numbers but different hydrophilic-lipophilic balance (HLB) values. Their mixed micellar systems were then used to solubilize a herbicide diuron (DN) and a fungicide cyprodinil (CP), followed by plant wax solubilization upon contact with wax films. These processes were monitored by 1H NMR and SANS. FINDING: Pesticide solubilization made surfactant micelles effectively more hydrophobic but subsequent wax dissolution caused pesticide release and the restoration of the micellar amphiphilicity. Nonionic surfactants with lower HLBs form larger nanoaggregates, show enhanced wettability, and have better ability to solubilize and permeate pesticides across the wax film, but may cause significant damage to plant growth. These observations help explain why herbicides applied on weeds would benefit from surfactants with lower HLB values while fungicides require surfactants with HLBs to balance between delivery efficiency and potential phytotoxicity risks.

How do terminal modifications of short designed IIKK peptide amphiphiles affect their antifungal activity and biocompatibility?

HYPOTHESIS: The widespread and prolonged use of antifungal antibiotics has led to the rapid emergence of multidrug resistant Candida species that compromise current treatments. Natural and synthetic antimicrobial peptides (AMPs) offer potential alternatives but require further development to overcome some of their current drawbacks. AMPs kill pathogenic fungi by permeabilising their membranes but it remains unclear how AMPs can be designed to maximise their antifungal potency whilst minimising their toxicity to host cells. EXPERIMENTS: We have designed a group of short (IIKK)3 AMPs via selective terminal modifications ending up with different amphiphilicities. Their antifungal performance was assessed by minimum inhibition concentration (MICs) and dynamic killing to 4 Candida strains and Cryptococcus neoformans, and the minimum biofilm-eradicating concentrations to kill 95% of the C. albicans biofilms (BEC95). Different antifungal actions were interpreted on the basis of structural disruptions of the AMPs to small unilamellar vesicles from fluorescence leakage, Zeta potential, small angle neutron scattering (SANS) and molecular dynamics simulations (MD). FINDING: AMPs possess high antifungal activities against the Candida species and Cryptococcus neoformans; some of them displayed faster dynamic killing than antibiotics like amphotericin B. G(IIKK)3I-NH2 and (IIKK)3II-NH2 were particularly potent against not only planktonic microbes but also fungal biofilms with low cytotoxicity to host cells. It was found that their high selectivity and fast action were well correlated to their fast membrane lysis, evident from data measured from Zeta potential measurements, SANS and MD, and also consistent with the previously observed antibacterial and anticancer performance. These studies demonstrate the important role of colloid and interface science in further developing short, potent and biocompatible AMPs towards clinical treatments via structure design and optimization.

Implications of surfactant hydrophobic chain architecture on the Surfactant-Skin lipid model interaction.

Although surfactants have been widely used in skin care and other related applications, our knowledge about how surfactants interact with stratum corneum (SC) lipids remains limited. This work reports how surfactants interact with a lipid SC model by neutron diffraction and molecular dynamics (MD) simulations, focusing on examining the impact of surfactant molecular architecture. The surfactant-SC mixed membrane was constructed by an equimolar mixture of ceramide/cholesterol/fatty acids and surfactant at 1% molar ratio of total lipids. The arrangements of water and surfactant molecules in the membrane were obtained through neutron scattering length density (NSLD) profiles via contrast variation method, meanwhile, MD simulation clearly demonstrated the mechanism of hydration change in the surfactant-model SC mixed membrane. No drastic difference was detected in the repeating distance of the short periodicity phase (SPP) upon adding surfactants, however, it significantly enhanced the membrane hydration and reduced the amount of phase separated crystalline cholesterol, showing a strong dependence on surfactant chain length, branching and double bond. This work clearly demonstrates how surfactant architecture affects its interaction with the SC membrane, providing useful guidance for either choosing an existing surfactant or designing a new one for surfactant-based transdermal application.