Search Results (463)

Erythrocyte (RBC) aging involves significant structural and nanomechanical alterations crucial to their function. This study aims to bridge the gap between analyses based on statistical morphometric parameters, e.g., membrane roughness, and those based on point-dependent nanomechanical properties, e.g., stiffness or Young’s modulus. Using Atomic Force Microscopy, we investigated morphology, membrane roughness, and nanomechanical properties on the very same RBCs under dehydrated (air) and hydrated (physiological buffer) conditions. The cells were studied at different stages of in vitro aging: one, seven, and 12 days. Our results quantitatively show that across dehydration, as well as along the aging pathway, RBCs become progressively more rigid while their membrane roughness decreases, a trend observed in both environments. Notably, the differences between the hydrated and dehydrated states were large in young cells but diminished when erythrocytes aged. Despite these parallel trends, high-resolution mapping on the nanoscale revealed that roughness and Young’s modulus do not correlate, indicating that these parameters are linked to different properties. In conclusion, this work provides a comprehensive protocol for a biophysical description of RBC aging and establishes that the simultaneous measurement of membrane roughness and nanomechanical properties offers a complementary approach, yielding a more complete characterization of cellular properties. Full article

Background/Objectives: Viral respiratory infections have a significant impact on global health and the economy. While vaccines are effective in preventing infection, they might not be available or sufficient when used alone and must be complemented by specific therapeutic strategies. The development of new antiviral agents is increasingly important due to the continual emergence of novel respiratory pathogens. Previously we identified bovine lactoferrin (bLf)-derived tetrapeptides and peptidomimetics that showed potent in vitro activity against the influenza A virus in the picomolar range. Methods: Inspired by these results, in this study, we evaluated the antiviral potential of these compounds against HCoV-229E, a human coronavirus that can cause severe disease in immunocompromised individuals, using a compound repositioning approach. Results: Functional studies revealed that SK(N-Me)HS (3) interferes with viral entry and replication, while compound SNKHS (5) primarily blocks infection in the early stages. Biophysical analyses confirmed the occurrence of high-affinity binding to the viral spike protein, and computational studies suggested that the compounds target a region involved in conformational changes necessary for membrane fusion. Conclusions: These findings highlight these compounds as promising candidates for coronavirus entry inhibition and underscore the value of compound repurposing in antiviral development. Full article

Tellurite (TeO₃²⁻) is a highly soluble and toxic oxyanion that inhibits the growth of Escherichia coli at concentrations as low as ~1 µg/mL. This toxicity has been primarily attributed to the generation of reactive oxygen species (ROS) during its intracellular reduction by thiol-containing molecules and NAD(P)H-dependent enzymes. However, under anaerobic conditions, E. coli exhibits significantly increased tellurite tolerance—up to 100-fold in minimal media—suggesting the involvement of additional, ROS-independent mechanisms. In this study, we combined chemical-genomic screening, untargeted metabolomics, and targeted biochemical assays to investigate the effects of tellurite under both aerobic and anaerobic conditions. Our findings reveal that tellurite perturbs amino acid and nucleotide metabolism, leading to intracellular imbalances that impair protein synthesis. Additionally, tellurite induces notable changes in membrane lipid composition, particularly in phosphatidylethanolamine derivatives, which may influence biophysical properties of the membrane, such as fluidity or curvature. This membrane remodeling could contribute to the increased resistance observed under anaerobic conditions, although direct evidence of altered membrane fluidity remains to be established. Overall, these results demonstrate that tellurite toxicity extends beyond oxidative stress, impacting central metabolic pathways and membrane-associated functions regardless of oxygen availability. Full article

Cell membrane capacitance (C_m) is a potential biomarker that reflects the structural and functional integrity of cell membranes. It is essential for physiological processes such as signal transduction, ion transport, and cellular homeostasis. In clinical practice, C_m can be determined using bioimpedance spectroscopy (BIS), a non-invasive technique for analysing the intrinsic electrical properties of biological tissues across a range of frequencies. C_m may be relevant in various clinical fields, where high capacitance is associated with healthy and intact membranes, while low capacitance indicates cellular damage or disease. Despite its promise as a prognostic indicator, several knowledge gaps limit the broader clinical application of C_m. These include variability in measurement techniques (e.g., electrode placement, frequency selection), the lack of standardised measurement protocols, uncertainty on how C_m is related to pathology, and the relatively low amount of C_m research. By addressing these gaps, C_m may become a valuable tool for examining cellular health, early disease detection, and evaluating treatment efficacy in clinical practice. This review explores the fundamental principles of C_m measured with the BIS technique, its mathematical basis and relationship to the biophysical Cole model, and its potential clinical applications. It identifies current gaps in our knowledge and outlines future research directions to enhance the understanding and use of C_m. For example, C_m has shown promise in identifying membrane degradation in sepsis, predicting malnutrition in anorexia nervosa, and as a prognostic factor in cancer. Full article

Peptide-induced disruption of lipid membranes is central to both amyloid diseases and the activity of antimicrobial peptides. Here, we combine all-atom molecular dynamics simulations with biophysical experiments to investigate how four amphipathic peptides interact with lipid bilayers. All peptides adsorb on the membrane surface. Peptide M01 [Ac-(FKFE)₂-NH₂] self-assembles into $\beta$-sheet nanofibrils that span both leaflets of the membrane, creating water-permeable channels. The other three peptides adopt $\alpha$-helical structures at the water–lipid interface. Peptide M02 [Ac-FFKKFFEE-NH₂], a sequence isomer of M01, does not form $\beta$-sheet aggregates and is too short to span the bilayer, resulting in no observable water permeation across the membrane. Peptides M03 and M04 are $\alpha$-helical isomers long enough to span the bilayer, with a polar face that allows the penetration of water deep inside the membrane. For the M03 peptide [Ac-(FFKKFFEE)₂-NH₂], insertion into the bilayer starts with the nonpolar N-terminal amino acids penetrating the hydrophobic core of the bilayer, while electrostatic interactions hold negative residues at the C-terminus on the membrane surface. The M04 peptide, [Ac-FFKKFFEEFKKFFEEF-NH₂], is made by relocating a single nonpolar residue from the central region of M03 to the C-terminus. This nonpolar residue becomes unfavorably exposed to the solvent upon insertion of the N-terminal region of the peptide into the membrane. Consequently, higher concentrations of M04 peptides are required to induce water permeation compared to M03. Overall, our comparative analysis reveals how subtle rearrangements of polar and nonpolar residues modulate peptide-induced water permeation. This provides mechanistic insights relevant to amyloid pathology and antimicrobial peptide design. Full article

Background/Objectives: The increasing prevalence of multidrug-resistant bacteria presents a major global health challenge, prompting a search for innovative antimicrobial strategies. This study aimed to develop and evaluate a novel nanobiostructure combining alumina nanoparticles (NPs) with the antimicrobial peptide lunatin-1 (Lun-1), forming peptide-functionalized nanofilaments. The main objective was to investigate how the site of peptide functionalization (C-terminal vs. N-terminal) affects membrane interactions and antibacterial activity. Methods: NP–peptide conjugates were synthesized via covalent bonding between lun-1 and alumina NP and characterized using transmission electron microscopy (TEM), X-ray diffraction (XRD), zeta potential analysis, dynamic light scattering (DLS), Fourier-transform infrared (FTIR), and solid-state ¹³C NMR. Antibacterial activities were assessed against different Gram-positive and Gram-negative strains. Biophysical analyses, including circular dichroism (CD), isothermal titration calorimetry (ITC), differential scanning calorimetry (DSC), and solid-state ²H NMR, were employed to evaluate peptide–membrane interactions in the presence of membrane-mimetic vesicles composed of POPC:POPG (3:1) and DMPC:DMPG (3:1). Results: Characterization confirmed the successful formation of NP–peptide nanofilaments. Functionalization at the N-terminal significantly influenced both antibacterial activity and peptide conformation compared to C-terminal attachment. Biophysical data demonstrated stronger membrane interaction and greater membrane disruption when lun-1 was conjugated at the N-terminal. Conclusions: The site of peptide conjugation plays a crucial role in modulating the biological and biophysical properties of NP–lunatin-1 conjugates. C-terminal attachment of lunatin-1 retains both membrane interaction and antibacterial efficacy, making it a promising strategy for the design of peptide-based nanotherapeutics targeting resistant pathogens. Full article

Cell membranes contain a variety of biomolecules, especially various kinds of lipids and proteins, which constantly change with fluidity and environmental stimuli. Though Helfrich curvature elastic energy has successfully explained many phenomena for single-component membranes, a new theoretical framework for multicomponent membranes is still a challenge. In this work, we propose a generalized Helfrich free-energy functional describe equilibrium shapes and phase behaviors related to membrane heterogeneity with via curvature-component coupling in a unified framework. For multicomponent membranes, a new but important Laplace–Beltrami operator is derived from the variational calculation on the integral of Gaussian curvature and applied to explain the spontaneous nanotube formation of an asymmetric glycolipid vesicle. Therefore, our general mathematical framework shows a predictive capabilities beyond the existing multicomponent membrane models. The set of new curvature-component coupling EL equations have been derived for global vesicle shapes associated with the composition redistribution of multicomponent membranes for the first time and specified into several typical geometric shape equations. The equilibrium radii of isotonic vesicles for both spherical and cylindrical geometries are calculated. The analytical solution for isotonic vesicles reveals that membrane stability requires distinct elastic moduli among components ($k_{\mathrm{A}}\ne k_{\mathrm{B}}$, ${\overline{k}}_{\mathrm{A}}\ne {\overline{k}}_{\mathrm{B}}$), which is consistent with experimental observations of coexisting lipid domains. Furthermore, we elucidate the biophysical implications of the derived shape equations, linking them to experimentally observed membrane remodeling processes. Our new free-energy framework provides a baseline for more detailed microscopic membrane models. Full article

Cholesterol plays an essential role in biological membranes and is crucial for maintaining their stability and functionality. In addition to biological membranes, cholesterol is also used in various synthetic lipid-based structures such as liposomes, proteoliposomes, and nanodiscs. Cholesterol regulates membrane properties by influencing the density of lipids, phase separation into liquid-ordered (Lo) and liquid-disordered (Ld) areas, and stability of protein–membrane interactions. For planar bilayers, cholesterol thickens the membrane, decreases permeability, and brings lipids into well-ordered domains, thereby increasing membrane rigidity by condensing lipid packing, while maintaining lateral lipid mobility in disordered regions to preserve overall membrane fluidity. It modulates membrane curvature in curved bilayers and vesicles, and stabilizes low-curvature regions, which are important for structural integrity. In liposomes, cholesterol facilitates drug encapsulation and release by controlling bilayer flexibility and stability. In nanodiscs, cholesterol enhances structural integrity and protein compatibility, which enables the investigation of protein–lipid interactions under physiological conditions. In proteoliposomes, cholesterol regulates the conformational stability of embedded proteins that have implications for protein–lipid interaction. Developments in molecular dynamics (MD) techniques, from coarse-grained to all-atom simulations, have shown how cholesterol modulates lipid tail ordering, membrane curvature, and flip-flop behavior in response to concentration. Such simulations provide insights into the mechanisms underlying membrane-associated diseases, aiding in the design of efficient drug delivery systems. In this review, we combine results from MD simulations to provide a synoptic explanation of cholesterol’s complex function in regulating membrane behavior. This synthesis combines fundamental biophysical information with practical membrane engineering, underscoring cholesterol’s important role in membrane structure, dynamics, and performance, and paving the way for rational design of stable and functional lipid-based systems to be used in medicine. In this review, we gather evidence from MD simulations to provide an overview of cholesterol’s complex function regulating membrane behavior. This synthesis connects the fundamental biophysical science with practical membrane engineering, which highlights cholesterol’s important role in membrane structure, dynamics, and function and helps us rationally design stable and functional lipid-based systems for therapeutic purposes. Full article

Dipalmitoylphosphatidylcholine (DPPC), is one of the key bilayer membranes of the phosphatidylcholine (PC) family which constitutes 40–50% of total cellular phospholipids in mammal cells. We investigate the behavior of an initially planar DPPC membrane under lateral pressures from −200 to 150 bar at 323 K using microsecond-scale simulations. We identify, with very high precision, the pressure range for the occurrence of critical phenomena, mainly undulation and rupture. Notably, under compression, the membrane initially thickens, leading to a phase transition to an undulated state between 40 and 50 bar, as gauged by the sharp changes in the diverse structural metrics. Stretching induces systematic membrane thinning, with rupture becoming probable at −170 bar and certain at −200 bar. The reverse compression cycle shows pressure hysteresis with a 10-bar shift, while the reverse stretching cycle retraces the pathway. System size has a minimal impact on the observed trends. Under extreme mechanical stress, particularly near critical phenomena, simulation times on the order of microsecond are needed to accurately capture phase behavior and structural alterations. This work provides important insights into understanding membrane behavior under extreme conditions, which are relevant to numerous biological and technological applications. Full article

As a surface-located molecule in biological membranes, cholesterol sulphate (CholS) plays a major role in membrane-driven cell–cell processes and events including platelet-cell adhesion, T-cell receptor signalling, sperm–egg interaction, membrane fusion, and skin differentiation. Despite this, little is known about the biophysical implications of CholS at the membrane in cells and organelles. In this study, we investigate the effect of increasing the content of CholS on the biophysical properties in cholesterol-poor and cholesterol-rich biomimetic models. Data obtained show that increasing amounts of CholS result in a slight increase in anisotropy, evidence for decreased membrane fluidity at higher CholS content (10 mol%) in cholesterol-poor systems but only negligible in rigidified epithelial-like cholesterol-rich systems. On the other hand, incorporation of CholS has an overall increasing ordering effect on membrane organisation and on-surface potential that is influenced by the lipid composition and cholesterol content. Though further research is needed to gain better insights on the (patho)physiological levels of CholS in cells and organelles, our findings are discussed in the context of diet–microbiota–host interactions in membrane-driven events in inflammatory-related disorders. Full article

The rapid regulatory mechanism of light-induced state transitions (STs) in oxygenic photosynthesis is particularly appealing for membrane-based applications. This interest stems from the unique ability of the thylakoid membrane protein cytochrome b₆f (cytb₆f) to increase or decrease its hydrophobic thickness (d_P) in parallel with the reduction or oxidation of the PQ pool induced by changes in light quality. This property appears to be the long-sought biophysical driver behind the reorganizations of membrane proteins during STs. This study decisively advances the hydrophobic mismatch (HMM) model for cytb₆f-driven STs by thoroughly analyzing thirteen X-ray crystal and eight cryo-electron microscopy cytb₆f structures. It uncovers the lipid nanoenvironments that cytb₆f, with different hydrophobic thicknesses, selectively attracts. Under optimal, stationary conditions for photosynthesis in low light, when there is hydrophobic matching between the hydrophobic thicknesses of cytb₆f d_P and that of the bulk thylakoid lipid phase d_L, d_P = d_L, cytb₆f predominantly binds to anionic lipids—several phosphatidylglycerol (PG) molecules and one sulfoquinovosyldiacylglycerol (SQDG) molecule. Upon the induction of the transition to State 2, when d_P increases and induces a positive HMM (d_P > d_L), the neutral, non-bilayer-forming lipid monogalactosyldiacylglycerol (MGDG) replaces some of the bound PGs. Upon the induction of the transition to State 1, when d_P decreases and induces a negative HMM (d_P < d_L), PGs and SQDG detach from their binding sites, and two neutral, bilayer-forming lipids such as digalactosyldiacylglycerol (DGDG) occupy two sites. Additionally, this research uncovers two lipid-mediated signaling pathways from Chla to the center of flexibility, the Phe/Tyr124^fg-loop-suIV residue—one of which involves β-carotene. This study identifies two novel types of lipid raft-like nanodomains that are devoid of typical components, such as sphingomyelin and cholesterol. These findings firmly validate the HMM model and underscore the STs as the first recognized functional process that fully utilizes the unique and evolutionarily conserved composition of just four thylakoid lipid classes. This research contributes to our understanding of membrane dynamics in general and STs in particular. It introduces a novel and simple approach for reversible protein reorganization driven purely by biophysical mechanisms, with promising implications for various membrane-based applications. Full article

Fluorescent cholesterol probes are indispensable tools for studying membrane structure, dynamics, and trafficking. To better understand the structure–function relationship of fluorescent cholesteryl probes, we developed a series of five new modular naphthalimide-containing cholesteryl probes (CND15–CND19). These probes incorporate an L-lysine linker between the cholesterol moiety and the fluorophore, along with a series of distinct head groups. We conducted extensive biophysical characterizations of these probes, including the determination of their solvatochromic properties and lipid partitioning behavior using giant unilamellar vesicles. Molecular dynamics simulations were employed to identify key molecular interactions of these probes within model lipid membranes. Furthermore, live-cell imaging in 3T3 fibroblasts demonstrated the potential applications of these analogs in live-cell imaging, measuring cellular membrane dynamics and studying cholesterol-related processes. The results of this study underscore the critical role of the linker and head group in designing fluorescent cholesterol-mimicking probes. These findings provide valuable insights into optimizing probe designs for future cholesterol and membrane biology research. Full article

Biological materials are distinguished by their hierarchical structures in which the organization of the basic building blocks is precisely controlled on many discrete length scales. This biophysical organization, i.e., the structure, along with the biochemical attributes, dictates their properties and function. This article is a review, and also a tutorial, that describes the use of small-angle X-ray scattering (SAXS) for determining the structures at the nanometer and sub-micron length scales in three distinct classes of scattering patterns that arise from fibrous structures, lamellae, and solutions. Fibrous structures are discussed using results from collagen, bone, hair, feathers, and silk. The use of SAXS to study the lamellar structures is illustrated using the results from myelin and membranes. SAXS in solutions is discussed by highlighting the results from multidomain proteins such as monoclonal antibodies and facile structures in intrinsically disordered proteins and protein condensates. The goal is to describe the different methods for analyzing the distinct classes of scattering patterns arising from 1- and 2-D ordered structures and from 3D structures in solutions and to illustrate how the structure imparts unique functions and properties to the biological materials. An understanding of the hierarchical structures in biology is expected to be useful in medical diagnosis and serve as a guide for fabricating functional biomaterials by mimicking these structures. Full article

The perivitellin-2 (PV2) from snails is an unusual neuro and enterotoxin comprising a pore-forming domain of the Membrane Attack Complex and Perforin Family (MACPF) linked to a lectin. While both domains have membrane binding capabilities, PV2’s mechanism of action remains unclear. We studied the apple snail Pomacea maculata PV2’s (PmPV2’s) interaction with lipid membranes using various biophysical and cell biology approaches. In vitro studies showed that PmPV2 toxicity decreased when cholesterol (Chol) was diminished from enterocyte cell membranes. Chol enhanced PmPV2 association with phosphatidylcholine membranes but did not induce pore formation. In contrast, using rat brain lipid models, rich in glycolipids, PmPV2 exhibited high affinity and induced vesicle permeabilization. Negative stain electron microscopy and atomic force microscopy confirmed the formation of pore-like structures in brain lipid vesicles. Our findings suggest that Chol is a necessary lipid component and point to PmPV2–glycolipid interactions as potential activators critical to triggering PmPV2’s pore-forming activity, providing insights into this novel toxin’s mechanism. Full article

Archaeal membranes exhibit remarkable stability under extreme environmental conditions, a feature attributed to their unique lipid composition. While it is widely accepted that tetraether lipids confer structural integrity by forming monolayers, the role of bilayer-forming diether lipids in membrane stability remains unclear. Here, we demonstrate that incorporating diethers into archaeal-like lipid assemblies enhances membrane organization and adaptability under thermal stress. Using neutron diffraction, we show that membranes composed of mixed diethers and tetraethers exhibit greater structural order and stability compared to pure lipid systems. Contrary to expectations, monolayer-forming tetraethers alone display increased variability in lamellar spacing under fluctuating temperature and humidity, whereas mixed lipid membranes maintain a consistent architecture. Furthermore, neutron-scattering length density profiles reveal an unexpected density feature at the bilayer midplane, challenging conventional models of archaeal monolayer organization. These findings suggest that molecular diversity of lipid molecules, rather than tetraether dominance, plays a critical role in membrane auto-assembly, stability, and adaptability. Our results provide new insights into archaeal membrane adaptation strategies, with implications for the development of bioinspired, robust synthetic membranes for industrial and biomedical applications. Full article