Network formation of lipid membranes: Triggering structural transitions by chain melting

Phospholipids when dispersed in excess water generally form vesicular membrane structures. Cryo-transmission and freeze-fracture electron microscopy are combined here with calorimetry and viscometry to demonstrate the reversible conversion of phosphatidylglycerol aqueous vesicle suspensions to a three-dimensional structure that consists of extended bilayer networks. Thermodynamic analysis indicates that the structural transitions arise from two effects: (i) the enhanced membrane elasticity accompanying the lipid state fluctuations on chain melting and (ii) solvent-associated interactions (including electrostatics) that favor a change in membrane curvature. The material properties of the hydrogels and their reversible formation offer the possibility of future applications, for example in drug delivery, the design of structural switches, or for understanding vesicle fusion or fission processes.

Amphipols: Polymers that keep membrane proteins soluble in aqueous solutions

Amphipols are a new class of surfactants that make it possible to handle membrane proteins in detergent-free aqueous solution as though they were soluble proteins. The strongly hydrophilic backbone of these polymers is grafted with hydrophobic chains, making them amphiphilic. Amphipols are able to stabilize in aqueous solution under their native state four well-characterized integral membrane proteins: (i) bacteriorhodopsin, (ii) a bacterial photosynthetic reaction center, (iii) cytochrome b6f, and (iv) matrix porin.

Interaction between like-charged colloidal spheres in electrolyte solutions

How colloidal particles interact with each other is one of the key issues that determines our ability to interpret experimental results for phase transitions in colloidal dispersions and our ability to apply colloid science to various industrial processes. The long-accepted theories for answering this question have been challenged by results from recent experiments. Herein we show from Monte-Carlo simulations that there is a short-range attractive force between identical macroions in electrolyte solutions containing divalent counterions. Complementing some recent and related results by others, we present strong evidence of attraction between a pair of spherical macroions in the presence of added salt ions for the conditions where the interacting macroion pair is not affected by any other macroions that may be in the solution. This attractive force follows from the internal-energy contribution of counterion mediation. Contrary to conventional expectations, for charged macroions in an electrolyte solution, the entropic force is repulsive at most solution conditions because of localization of small ions in the vicinity of macroions. Both Derjaguin–Landau–Verwey–Overbeek theory and Sogami–Ise theory fail to describe the attractive interactions found in our simulations; the former predicts only repulsive interaction and the latter predicts a long-range attraction that is too weak and occurs at macroion separations that are too large. Our simulations provide fundamental “data” toward an improved theory for the potential of mean force as required for optimum design of new materials including those containing nanoparticles.

On the acidity and reactivity of HNO in aqueous solution and biological systems

The gas phase and aqueous thermochemistry and reactivity of nitroxyl (nitrosyl hydride, HNO) were elucidated with multiconfigurational self-consistent field and hybrid density functional theory calculations and continuum solvation methods. The pKa of HNO is predicted to be 7.2 ± 1.0, considerably different from the value of 4.7 reported from pulse radiolysis experiments. The ground-state triplet nature of NO− affects the rates of acid-base chemistry of the HNO/NO− couple. HNO is highly reactive toward dimerization and addition of soft nucleophiles but is predicted to undergo negligible hydration (Keq = 6.9 × 10−5). HNO is predicted to exist as a discrete species in solution and is a viable participant in the chemical biology of nitric oxide and derivatives.

Designing surfaces that kill bacteria on contact

Poly(4-vinyl-N-alkylpyridinium bromide) was covalently attached to glass slides to create a surface that kills airborne bacteria on contact. The antibacterial properties were assessed by spraying aqueous suspensions of bacterial cells on the surface, followed by air drying and counting the number of cells remaining viable (i.e., capable of growing colonies). Amino glass slides were acylated with acryloyl chloride, copolymerized with 4-vinylpyridine, and N-alkylated with different alkyl bromides (from propyl to hexadecyl). The resultant surfaces, depending on the alkyl group, were able to kill up to 94 ± 4% of Staphylococcus aureus cells sprayed on them. A surface alternatively created by attaching poly(4-vinylpyridine) to a glass slide and alkylating it with hexyl bromide killed 94 ± 3% of the deposited S. aureus cells. On surfaces modified with N-hexylated poly(4-vinylpyridine), the numbers of viable cells of another Gram-positive bacterium, Staphylococcus epidermidis, as well as of the Gram-negative bacteria Pseudomonas aeruginosa and Escherichia coli, dropped more than 100-fold compared with the original amino glass. In contrast, the number of viable bacterial cells did not decline significantly after spraying on such common materials as ceramics, plastics, metals, and wood.

Oxidative Burst and Hypoosmotic Stress in Tobacco Cell Suspensions

Oxidative burst constitutes an early response in plant defense reactions toward pathogens, but active oxygen production may also be induced by other stimuli. The oxidative response of suspension-cultured tobacco (Nicotiana tabacum cv Xanthi) cells to hypoosmotic and mechanical stresses was characterized. The oxidase involved in the hypoosmotic stress response showed similarities by its NADPH dependence and its inhibition by iodonium diphenyl with the neutrophil NADPH oxidase. Activation of the oxidative response by hypoosmotic stress needed protein phosphorylation and anion effluxes, as well as opening of Ca2+ channels. Inhibition of the oxidative response impaired Cl− efflux, K+ efflux, and extracellular alkalinization, suggesting that the oxidative burst may play a role in ionic flux regulation. Active oxygen species also induced the cross-linking of a cell wall protein, homologous to a soybean (Glycine max L.) extensin, that may act as part of cell volume and turgor regulation through modification of the physical properties of the cell wall.

Increased liposome extravasation in selected tissues: effect of substance P.

We have used a pharmacologic mediator to open intercellular connections in selected vessels to allow liposomes to escape from the blood stream and to extravasate into tissues that have appropriate receptors. We have examined the effects of substance P (SP), a peptide known to increase vascular permeability in selected tissues, such as trachea, esophagus, and urinary bladder in rats. We used quantitative fluorescence analysis of tissues to measure two fluorescent markers, one attached to the lipid (rhodamine-phosphatidylethanolamine) and another, doxorubicin (an anti-tumor drug), encapsulated within the aqueous interior. We have also examined the deposition of liposomes microscopically by the use of encapsulated colloidal gold and silver enhancement. Analysis of the biochemical and morphological observations indicate the following: (i) Injection of SP produces a striking increase in both liposome labels, but only in tissues that possess receptors for SP in postcapillary venules; (ii) liposome material in these tissues has extravasated and is found extracellularly near a variety of cells beyond the endothelial layer over the first few hours; (iii) 24 h following injection of liposomes and SP, liposome material is found in these tissues, localized intracellularly in both endothelial cells and macrophages. We propose that appropriate application of tissue-specific mediators can result in liposome extravasation deep within tissues that normally do not take up significant amounts of liposomes from the blood. Such liposomes are able to carry a variety of pharmacological agents that can be released locally within selected target tissues for therapeutic purposes.

Phase separation in aqueous solutions of lens gamma-crystallins: special role of gamma s.

We have studied liquid-liquid phase separation in aqueous ternary solutions of calf lens gamma-crystallin proteins. Specifically, we have examined two ternary systems containing gamma s--namely, gamma IVa with gamma s in water and gamma II with gamma s in water. For each system, the phase-separation temperatures (Tph (phi)) alpha as a function of the overall protein volume fraction phi at various fixed compositions alpha (the "cloud-point curves") were measured. For the gamma IVa, gamma s, and water ternary solution, a binodal curve composed of pairs of coexisting points, (phi I, alpha 1) and (phi II, alpha II), at a fixed temperature (20 degrees C) was also determined. We observe that on the cloud-point curve the critical point is at a higher volume fraction than the maximum phase-separation temperature point. We also find that typically the difference in composition between the coexisting phases is at least as significant as the difference in volume fraction. We show that the asymmetric shape of the cloud-point curve is a consequence of this significant composition difference. Our observation that the phase-separation temperature of the mixtures in the high volume fraction region is strongly suppressed suggests that gamma s-crystallin may play an important role in maintaining the transparency of the lens.

Electrostatic force microscope for probing surface charges in aqueous solutions.

A scanning force microscope was converted to an electrostatic force microscope by charging the usually neutral cantilever with phospholipids. The electrostatic force microscope was used to study surface electrostatic charges of samples in aqueous solutions. Lysozymes, DEAE-Sephadex beads, 3-propyltriethoxysilane-treated glass and mica were imaged in water or phosphate buffer with electrostatic force microscopy. The adhesion force measured when a charged probe and oppositely charged specimen interacted was up to 500 times greater than when a bare probe was used. This dramatic increase in measured adhesion force can be attributed to the energy required to break the salt bridges formed between the charged probe and the specimen. The use of phospholipids to functionalize the cantilever tip allows the incorporation of other biomolecules and ligands that can be used as biologically specific tips (e.g., receptors, drugs) for the study of intermolecular interactions.