Fluorescent alamethicin fragments A study of membrane activity and aqueous phase aggregation

The linear polypeptide antibiotic alamethicin is known to form channels in artificial lipid membranes. Synthetic 13- and 17-residue alamethicin fragments, labelled with a fluorescent dansyl group at the N-terminus, have been shown to translocate divalent cations across phospholipid membranes and to uncouple oxidative phosphorylation in rat liver mitochondria, in a manner analogous to the parent peptides. From studies of the aqueous phase aggregation behavior of the peptides, as well as their interaction with rat liver mitochondria, it is concluded that the interaction of the peptides with membranes is a complex process, probably involving both aqueous and membrane phase aggregation.

Design and flight testing of a bio-inspired plume tracking algorithm for unmanned serial vehicles

There is an increased interest on the use of UAVs for environmental research such as tracking bush fires, volcanic eruptions, chemical accidents or pollution sources. The aim of this paper is to describe the theory and results of a bio-inspired plume tracking algorithm. A method for generating sparse plumes in a virtual environment was also developed. Results indicated the ability of the algorithms to track plumes in 2D and 3D. The system has been tested with hardware in the loop (HIL) simulations and in flight using a CO2 gas sensor mounted to a multi-rotor UAV. The UAV is controlled by the plume tracking algorithm running on the ground control station (GCS).

Weakening of the Gram-negative bacterial outer membrane - A tool for increasing microbiological safety

Gram-negative bacteria are harmful in various surroundings. In the food industy their metabolites are potential cause of spoilage and this group also includes many severe or potential pathogens, such as Salmonella. Due to their ability to produce biofilms Gram-negative bacteria also cause problems in many industrial processes as well as in clinical surroundings. Control of Gram-negative bacteria is hampered by the outer membrane (OM) in the outermost layer of the cells. This layer is an intrinsic barrier for many hydrophobic agents and macromolecules. Permeabilizers are compounds that weaken OM and can thus increase the activity of antimicrobials by facililating entry of hydrophobic compounds and macromolecules into the cell where they can reach their target sites and inhibit or destroy cellular functions. The work described in this thesis shows that lactic acid acts as a permeabilizer and destabilizes the OM of Gram-negative bacteria. In addition, organic acids present in berriers, i.e. malic, sorbic and benzoic acid, were shown to weaken the OM of Gram-negative bacteria. Organic acids can poteniate the antimicrobial activity of other compounds. Microbial colonic degradation products of plant-derived phenolic compounds (3,4-dihydroxyphenylacetic acid, 3-hydroxyphenylacetic acid, 3,4-dihydroxyphenylpropionic acid, 4-hydroxyphenylpropionic acid, 3-phenylpropionic acid and 3-hydroxyphenylpropionic acid) efficiently destabilized OM of Salmonella. The studies increase our understanding of the mechanism of action of the classical chelator, ethylenediaminetetra-acetic acid (EDTA). In addition, the results indicate that the biocidic activity of benzalkonium chloride against Pseudomonas can be increased by combined use with polyethylenimine (PEI). In addition to PEI, several other potential permeabilizers, such as succimer, were shown to destabilize the OM of Gram-negative bacteria. Furthermore, combination of the results obtained from various permeability assays (e.g. uptake of a hydrophobic probe, sensitization to hydrophobic antibiotics and detergents, release of lipopolysaccharide (LPS) and LPS-specific fatty acids) with atomic force microscopy (AFM) image results increases our knowledge of the action of permeabilizers.

Immunologic studies on nucleic acids and their components. I. An analysis of the specificity of anti-deoxyadenylate antibodies by a membrane-binding technique

Anti-deoxyadenylate antibodies were produced in rabbits by injecting a conjugate of deoxyadenosine 5′-phosphate with bovine serum albumin. The antisera, as analyzed by double diffusion in agar and the quantitative precipitin reaction, showed hapten-specific antibodies. The specific interaction between [3H]deoxyadenylate and antiserum was studied by a sensitive nitrocellulose membrane-binding assay. The specificity of the antibodies was analyzed by measuring the effectiveness of other nucleotides or derivatives to inhibit the hapten-antibody binding. The requirements for recognition by the antibody sites were studied by using a series of naturally occurring nucleic acid components as well as some synthetic derivatives as inhibitors. The antibodies were found to show a high degree of specificity for the whole nucleotide, the base, sugar and phosphate playing almost equally important roles. There was cross reactivity with other mononucleotides, although of a low order. The antibodies were able to react with DNA and tRNA.

Paramagnetic probes in membrane biophysics

The use of paramagnetic probes in membrane research is reviewed. Electron paramagnetic resonance studies on model and biological membranes doped with covalent and non-covalent spin-labels have been discussed with special emphasis on the methodology and the type of information obtainable on several important phenomena like membrane fluidity, lipid flip-flop, lateral diffusion of lipids, lipid phase separation, lipid bilayer phase transitions, lipid-protein interactions and membrane permeability. Nuclear magnetic resonance spectroscopy has also been effectively used to study the conformations of cation mediators across membranes and to analyse in detail the transmembrane ionic motions at the mechanistic level.

Vanadate-stimulated NADH oxidation in plasma membrane

The rate of NADH oxidation with oxygen as the acceptor is very low in mouse liver plasma membrane and erythrocyte membrane. When vanadate is added, this rate is stimulated 10- to 20-fold. The absorption spectrum of vanadate does not change with the disappearance of NADH. The reaction is inhibited by superoxide dismutase, and there is no activity under an argon atmosphere. This indicates that oxygen is the electron acceptor and the reaction is mediated by superoxide. The vanadate stimulation is not limited to plasma membrane. Golgi apparatus and endoplasmic reticulum show similar increase in NADH oxidase activity when vanadate is added. The endomembranes have significant vanadate-stimulated activity with both NADH and NADPH. The vanadate-stimulated NADH oxidase in plasma membrane is inhibited by compounds, which inhibit NADH dehydrogenase activity: catechols, anthracycline drugs and manganese. This activity is stimulated by high phosphate and sulfate anion concentrations.

Studies on bicuculline binding sites on neuronal membrane using fluorescent antibody technique: Comparative binding of gaba and bicuculline

Highly purified fluorescent labelled anti-bicuculline antibodies were used to mark bicuculline binding sites in cerebral cortex of monkey brain. Specific binding of bicuculline could be demonstrated in the synaptosomal fraction, when bicuculline was added both Image and Image . Addition of γ-aminobutyric acid (GABA) to the bicucullinised membrane led to a decrease in fluorescence indicating same receptor loci and establishing GABA-bicuculline antagonism at a molecular level.

Modified design procedure for a sparger reactor sequence

In an earlier communication[l] we have indicated a general graphical design procedure for a sequence of sparger reactors in which a second order liquid phase reaction proceeds in a stagewise fashion. The prediction of the reactant concentration in each stage and hence the conversion depended on a search procedure initiated along a straight line representing the mass balance equation at the given stage and drawn from the known feed stage located on the abscissa in a E-IU diagram for the given system.

Liquid phase hydrogenation of aniline in a trickle bed reactor

A rate equation is developed for the liquid phase hydrogenation of aniline over cylindrical catalyst pellets of 30% nickel deposited on clay in a trickle bed reactor. The equation takes into account external and internal diffusional limitations, and describes the experimental data adequately. The hydrogenation reaction is first order with respect to hydrogen and zero order with respect to aniline. Effectiveness factors are in the range 0.003-0.03. Apparent activation energy of the reaction is 12.7 kcal/mol and true activation energy is 39.6 kcal/mol.

Separation of sulfur-containing components of transfer ribonucleic acid on Bio-Gel P-2 and Sephadex G-10 columns

S-Labeled nucleosides of E. coli tRNA and some of the derivatives of thionucleosides were separated on Bio-Gel P-2 and Sephadex G-10 columns employing buffers of low salt concentration and high pH.

Lymphatic vessels in health and disease: Role of the VEGF-C/VEGFR-3 pathway and the transcription factor FOXC2

The circulatory system consists of two vessel types, which act in concert but significantly differ from each other in several structural and functional aspects as well as in mechanisms governing their development. The blood vasculature transports oxygen, nutrients and cells to tissues whereas the lymphatic vessels collect extravasated fluid, macromolecules and cells of the immune system and return them back to the blood circulation. Understanding the molecular mechanisms behind the developmental and functional regulation of the lymphatic system long lagged behind that of the blood vasculature. Identification of several markers specific for the lymphatic endothelium, and the discovery of key factors controlling the development and function of the lymphatic vessels have greatly facilitated research in lymphatic biology over the past few years. Recognition of the crucial importance of lymphatic vessels in certain pathological conditions, most importantly in tumor metastasis, lymphedema and inflammation, has increased interest in this vessel type, for so long overshadowed by its blood vascular cousin. VEGF-C (Vascular Endothelial Growth Factor C) and its receptor VEGFR-3 are essential for the development and maintenance of embryonic lymphatic vasculature. Furthermore, VEGF-C has been shown to be upregulated in many tumors and its expression found to positively correlate with lymphatic metastasis. Mutations in the transcription factor FOXC2 result in lymphedema-distichiasis (LD), which suggests a role for FOXC2 in the regulation of lymphatic development or function. This study was undertaken to obtain more information about the role of the VEGF-C/VEGFR-3 pathway and FOXC2 in regulating lymphatic development, growth, function and survival in physiological as well as in pathological conditions. We found that the silk-like carboxyterminal propeptide is not necessary for the lymphangiogenic activity of VEGF-C, but enhances it, and that the aminoterminal propeptide mediates binding of VEGF-C to the neuropilin-2 coreceptor, which we suggest to be involved in VEGF-C signalling via VEGFR-3. Furthermore, we found that overexpression of VEGF-C increases tumor lymphangiogenesis and intralymphatic tumor growth, both of which could be inhibited by a soluble form of VEGFR-3. These results suggest that blocking VEGFR-3 signalling could be used for prevention of lymphatic tumor metastasis. This might prove to be a safe treatment method for human cancer patients, since inhibition of VEGFR-3 activity had no effect on the normal lymphatic vasculature in adult mice, though it did lead to regression of lymphatic vessels in the postnatal period. Interestingly, in contrast to VEGF-C, which induces lymphangiogenesis already during embryonic development, we found that the related VEGF-D promotes lymphatic vessel growth only after birth. These results suggest, that the lymphatic vasculature undergoes postnatal maturation, which renders it independent of ligand induced VEGFR-3 signalling for survival but responsive to VEGF-D for growth. Finally, we show that FOXC2 is necessary for the later stages of lymphatic development by regulating the morphogenesis of lymphatic valves, as well as interactions of the lymphatic endothelium with vascular mural cells, in which it cooperates with VEGFR-3. Furthermore, our study indicates that the absence of lymphatic valves, abnormal association of lymphatic capillaries with mural cells and an increased amount of basement membrane underlie the pathogenesis of LD. These findings have given new insight into the mechanisms of normal lymphatic development, as well as into the pathogenesis of diseases involving the lymphatic vasculature. They also reveal new therapeutic targets for the prevention and treatment of tumor metastasis and lymphatic vascular failure in certain forms of lymphedema. Several interesting questions were posed that still need to be addressed. Most importantly, the mechanism of VEGF-C promoted tumor metastasis and the molecular nature of the postnatal lymphatic vessel maturation remain to be elucidated.