Antimicrobial peptides are part of the immune defense system involved in protection against pathogenic agents, such as bacteria and fungi. Most of them are cationic and amphipathic, an essential feature that confers them special affinity for their main target: the lipid matrix of the cell membrane of microorganisms. Their activity as biocides depends on perturbation of the membrane barrier, either by causing the opening of pores or by inducing membrane disintegration. Lipid bilayers have been widely used as biomimetic system of the complex cell membrane and were essential in providing mechanistic detail on the mode of action of antimicrobial peptides. In particular, giant unilamellar vesicles (GUVs) are cell-sized lipid vesicles that can be individually followed under a microscope, providing a direct way to correlate peptide binding with membrane permeabilization and domain formation. Here, an overview on the contributions of optical microscopy to reveal the mode of action of several membrane-active peptides is given and discussed. A complete characterization of the mechanism of action of the antimicrobial peptide gomesin from the Brazilian spider Acanthoscurria gomesiana developed by our group in recent works is presented, combining optical microscopy observation of GUVs and complementary techniques on large unilamellar vesicles, such as isothermal titration calorimetry, light scattering, and a leakage assay of an entrapped fluorescent dye.
Apoptosis is a form of programmed cell death that plays an important role in key biological processes like development of organisms, the correct functioning of the immune system, and the maintenance of the cellular homeostasis. Dysregulation in the apoptotic pathway leads to diseases like cancer or neurodegenerative disorders. The proteins of the B-cell lymphoma 2 (Bcl-2) family are key regulators of mitochondrial outer membrane permeabilization (MOMP) during apoptosis, which is a critical step in the cell's commitment to death. However, their mechanism of action is still under intense investigation. Here, we discuss how microscopy approaches applied to model membranes are used to understand the intrinsic apoptotic pathway involving MOMP. We describe how model membranes mimicking the outer mitochondrial membrane (LUVs, GUVs, SLB) are used to understand the mechanism of Bcl-2-mediated apoptosis using state-of-the-art techniques like atomic force microscopy and fluorescence correlation spectroscopy. These studies have revealed interesting features like the role of membrane in altering the affinity of Bcl-2 proteins, Bax and Bak proapoptotic activity, mechanistic differences between pro- and antiapoptotic members, and the critical helices involved in pore formation by Bax.
There has been a great effort to study lipid lateral organization in biomembranes in the past decades, in order to unravel the structural basis and functional significance of membrane lipid domains. However, in both respects fundamental doubts still persist, and recent results have widened this topic well beyond lipid rafts. In particular, the detection of sphingolipid-enriched gel domains in the yeast Saccharomyces cerevisiae plasma membrane, which are not the prototypical liquid ordered, sterol-enriched lipid rafts, will be described. The critical role of ordered lipid domains will be demonstrated with biophysical studies of membrane lipid organization in living cells and in model systems, concerning mammalian and fungal membranes. Membrane interactions with different types of bioactive molecules will be briefly presented, including endogenous molecules such as the hormone epinephrine or membrane proteins, as well as drugs, including anticancer and antitubercular compounds. Strategies to tackle the complexity of living cell membranes will be discussed, in an attempt to reach a compromise between lipid lamellar phases in artificial or reconstituted systems and observations in living cells. The development of new and improved biomimetic systems might provide answers to some of the open questions in the lipid domains field. Therefore, new lipid bilayer membrane models containing lipid domains stably formed on a conducting support (gold), where powerful surface and electrochemical techniques can be employed, will also be presented. The redox behavior of the catecholamine hormone epinephrine studied in such system showed that the lipid bilayer has a crucial role for the hormone chemical stability.
Lipids are integral part of cell membranes and play a vital role in biological processes such as membrane fusion, exocytosis, cell signaling, and disease progression. Uniquely defined chemical structures of lipids as well as their assembly in the aqueous environment are often known to dictate their biological activity. Some of these features of lipids have been exploited to develop carriers for drugs, bioactive molecules, and nucleic acids. Lipid-based nanocarriers (specifically liposomes) have claimed their status in the field of cancer nanomedicine. Liposomes, the longest studied nanocarriers (at least since four decades), are currently used in the clinics for delivery of drugs to treat certain types of cancers though not at their full potential. Further development in this field by utilizing site-specific targeting molecules and/or efforts to generate tunable liposomes can be predicted to enhance their suitability in the clinical settings. In general, the chemical and biophysical properties of lipid molecules primarily dictate the development of tunable (stimuli-sensitive) liposomes. Research activities toward stimuli-responsive liposomes can be broadly classified into internal (pH, redox, and enzyme) or external (heat, light and/or magnetic field, and ultrasound) stimulus. In this review, we will discuss the current status and latest developments in the field of stimuli-sensitive liposomes. The topics covered in this review will include (i) a brief summary of liposomes used in the clinics, stimuli-sensitive liposomes; (ii) triggering events by inclusion of coagents in the liposomes; (iii) triggering based on properties of lipids to yield heat, light, pH, redox, and enzyme-activatable liposomes; and (iv) limitations and future directions for the development of clinically viable stimuli-responsive liposomes.
Lipids are integral part of cell membranes and play a vital role in biological processes such as membrane fusion, exocytosis, cell signaling, and disease progression. Uniquely defined chemical structures of lipids as well as their assembly in the aqueous environment are often known to dictate their biological activity. Some of these features of lipids have been exploited to develop carriers for drugs, bioactive molecules, and nucleic acids. Lipid-based nanocarriers (specifically liposomes) have claimed their status in the field of cancer nanomedicine. Liposomes, the longest studied nanocarriers (at least since four decades), are currently used in the clinics for delivery of drugs to treat certain types of cancers though not at their full potential. Further development in this field by utilizing site-specific targeting molecules and/or efforts to generate tunable liposomes can be predicted to enhance their suitability in the clinical settings. In general, the chemical and biophysical properties of lipid molecules primarily dictate the development of tunable (stimuli-sensitive) liposomes. Research activities toward stimuli-responsive liposomes can be broadly classified into internal (pH, redox, and enzyme) or external (heat, light and/or magnetic field, and ultrasound) stimulus. In this review, we will discuss the current status and latest developments in the field of stimuli-sensitive liposomes. The topics covered in this review will include (i) a brief summary of liposomes used in the clinics, stimuli-sensitive liposomes; (ii) triggering events by inclusion of coagents in the liposomes; (iii) triggering based on properties of lipids to yield heat, light, pH, redox, and enzyme-activatable liposomes; and (iv) limitations and future directions for the development of clinically viable stimuli-responsive liposomes.
In this study, we discuss applications of the electrochemical impedance (EI) response of tethered bilayer membranes (tBLMs) in protein (peptide)/phospholipid membrane interaction studies. Because of highly asymmetric geometry, tBLMs exhibit unique EI response, which cannot be modeled by simple equivalent circuits consisting of capacitors and resistors. Special functions describe the characteristic features of EI spectra. They take into account the structural and the dielectric properties of the tBLMs. The analysis of the EI response provides the theoretical background for the utility of tBLMs as bioanalytical sensors for the membrane-damaging agents, such as pore-forming toxins. We demonstrate that the magnitude and frequency of the minimum of the negative of the phase as well as the modulus of impedance are the parameters indicative of the extent of the membrane damage and may be used to estimate the defect density in bilayers. The precision of such estimates is highly dependent on the knowledge of the physical properties of the submembrane reservoir separating phospholipid bilayer and solid surface. Clustering of the defects affects the EI response in a unique way, which may be used for the qualitative analysis of protein–membrane interactions.
Diverse cellular functions are supported by membrane protein assemblies associated with the cell membrane. Although considered to be protein-mediated, membrane components are now being recognized as critical in modulating and sometime dictating function. This chapter discusses the effect of the lipid bilayer, in particular its composition on membrane protein organization. Computational methods have been successful in quantifying transmembrane protein association and general features of dimerization profiles are explored. Understanding the molecular basis of the interactions has lead to the recognition of the lipophobic effects. These nonspecific effects include those that arise from membrane perturbations and lipid chain packing and have been shown to modulate the energetics as well as the structural characteristics of membrane protein dimerization. In addition, specific interactions arising from direct protein-lipid interactions and protein-cholesterol interactions have been suggested to influence membrane protein association. We summarize here a few examples highlighting the role of the lipid bilayer on membrane protein organization.
An intriguing question in cell biology is "how do cells regulate their shape?" It is commonly believed that the observed cellular morphologies are a result of the complex interaction among the lipid molecules (constituting the cell membrane), and with a number of other macromolecules, such as proteins. It is also believed that the common biophysical processes essential for the functioning of a cell also play an important role in cellular morphogenesis. At the cellular scale-where typical dimensions are in the order of micrometers-the effects arising from the molecular scale can either be modeled as equilibrium or non-equilibrium processes. In this chapter, we discuss the dynamically triangulated Monte Carlo technique to model and simulate membrane morphologies at the cellular scale, which in turn can be used to investigate several questions related to shape regulation in cells. In particular, we focus on two specific problems within the framework of isotropic and anisotropic elasticity theories: namely, (i) the origin of complex, physiologically relevant, membrane shapes due to the interaction of the membrane with curvature remodeling proteins, and (ii) the genesis of steady state cellular shapes due to the action of non-equilibrium forces that are generated by the fission and fusion of transport vesicles and by the binding and unbinding of proteins from the parent membrane.
All antimicrobial agents are inherently membrane active and the complex interactions involved can lead to considerable conformational changes in the agents, while also inducing structural rearrangements of membrane lipids. Such structural modifications can trigger series of events enabling the agent to affect the structural integrity of the microbial membrane or translocate to the interior of the microbial cell. Therapeutic use of such agents requires a detailed understanding of the interaction of such antimicrobial agents with bacterial membranes. It is of interest to note that despite the existence of diversity in chemical compositions of many naturally occurring antimicrobial agents, the possible number of secondary structure conformations that they possess, either in solution or in membrane environment, is very small in number. In spite of considerable effort put in to probe the relationship between secondary structure and mode of antimicrobial action over the past two decades through experiments and simulations, a detailed understanding of the same is yet to be achieved. Furthermore, recent experimental and simulation results suggest that built-in well-defined secondary conformations such as a-helix or beta-sheet may not be the essential feature of potent antimicrobial agents, but rather the ability of these agents to acquire amphiphilic conformations, involving the spatial separation of charged and hydrophobic moieties, near the bacterial membrane. In this chapter, we review different antimicrobial agents that have been the focus of various studies with special emphasis on computer simulations and their role in understanding the interactions of biomimetic antimicrobial polymers, based on methacrylate copolymers, with bacterial membranes.
Lyotropic liquid crystalline nanostructured particles, such as cubosomes, have grown in popularity as drug delivery systems in the last few years. These systems require steric stabilizers to maintain colloidal stability in an aqueous medium, with Pluronic®F127, a block copolymer, being the most commonly employed stabilizer. However, in recent years, alternative, more effective stabilizers, as well as rationally designed systems with opportunities for further biofunctionalization have been reported. The purpose of this chapter is to collate and collectively interpret studies in the field of steric stabilization of this important emerging class of nanoparticles for drug and medical imaging agent delivery.
Membrane microvesiculation is a common process in cells. Membrane constituents undergo lateral redistribution coupled to the change in local membrane curvature. Thin necks that are formed in this process can be torn by mechanical stress and membrane-enclosed fragments that contain various biologically active molecules become more or less free to move with fluids. Released vesicles are small in size (micrometer down to tens of nanometers). They interact with distant cells and thereby present an intercellular communication system which plays important physiological role in organisms. Micro and nanovesicles (NVs) can be isolated from body fluids. It was found that the concentration of NVs is increased in isolates from blood of patients with different diseases (e.g., cancer, inflammation, infection, thromboembolic diseases) indicating an increased vesiculability of blood cells. Here, we present some mechanisms of microvesiculation of biological membranes and suggest a possible mechanism for suppression of microvesiculation by a mediated attractive interaction between membranes.
There has been a great effort to study lipid lateral organization in biomembranes in the past decades, in order to unravel the structural basis and functional significance of membrane lipid domains. However, in both respects fundamental doubts still persist, and recent results have widened this topic well beyond lipid rafts. In particular, the detection of sphingolipid-enriched gel domains in the yeast Saccharomyces cerevisiae plasma membrane, which are not the prototypical liquid ordered, sterol-enriched lipid rafts, will be described.The critical role of ordered lipid domains will be demonstrated with biophysical studies of membrane lipid organization in living cells and in model systems, concerning mammalian and fungal membranes. Membrane interactions with different types of bioactive molecules will be briefly presented, including endogenous molecules such as the hormone epinephrine or membrane proteins, as well as drugs, including anticancer and antitubercular compounds. Strategies to tackle the complexity of living cell membranes will be discussed, in an attempt to reach a compromise between lipid lamellar phases in artificial or reconstituted systems and observations in living cells.The development of new and improved biomimetic systems might provide answers to some of the open questions in the lipid domains field. Therefore, new lipid bilayer membrane models containing lipid domains stably formed on a conducting support (gold), where powerful surface and electrochemical techniques can be employed, will also be presented. The redox behavior of the catecholamine hormone epinephrine studied in such system showed that the lipid bilayer has a crucial role for the hormone chemical stability.
Hydrophobically modified chitosan was prepared using the hydrophobic compound derived from natural resource material such as cashew nut shell liquid. Chitosan was modified using 3-pentadecyl cyclohexane carbaldehyde to different extents (2, 3, and 5 mol%). Solution properties of hydrophobically modified chitosan were studied by rheology and light scattering. These indicated the aggregation behavior above the critical association concentration. Further, it was concluded that above the critical association concentration, the dynamics of the network formed due to the associations slowed down significantly.
Advances in Planar Lipid Bilayers and Liposomes cover a broad range of topics, including main arrangements of the reconstituted system, namely planar lipid bilayers as well as spherical liposomes. The invited authors present the latest results of their own research groups in this exciting, multidisciplinary field.Incorporates contributions from newcomers and established and experienced researchersExplores the planar lipid bilayer systems and spherical liposomes from theoretical and experimental perspectivesServes as an indispensable source of information for new scientists
