Effects of membrane channel-forming polypeptides on mitochondrial oxidative phosphorylation. A comparison of alamethicin, gramicidin A, melittin and tetraacetyl melittin

Transmembrane channel-forming polypeptides can function as uncouplers of mitochondrial oxidative phosphorylation. The observed effects are dependent on the phosphate ion (Pi) concentration in the medium. At low Pi (2.5 mM) the order of uncoupling efficiencies is gramicidin A much greater than alamethicin greater than tetraacetyl melittin greater than melittin. The remarkably high activity of gramicidin A suggests insertion of preformed channel dimers into the membrane. It is also suggested that lipid phase association of peptides is necessary in the other cases. At Pi = 100 mM inhibitory effects are observed for alamethicin and tetraacetyl melittin. Less pronounced inhibition is seen for melittin, while no such effect is noted for gramicidin A. The site of inhibition is shown to be complex IV, and the differences in the behavior of the peptides are rationalized in terms of channel structures.

Effects of membrane channel-forming polypeptides on mitochondrial oxidative phosphorylation. A comparison of alamethicin, gramicidin A, melittin and tetraacetyl melittin.

Transmembrane channel-forming polypeptides can function as uncouplers of mitochondrial oxidative phosphorylation. The observed effects are dependent on the phosphate ion (Pi) concentration in the medium. At low Pi (2.5 mM) the order of uncoupling efficiencies is gramicidin A much greater than alamethicin greater than tetraacetyl melittin greater than melittin. The remarkably high activity of gramicidin A suggests insertion of preformed channel dimers into the membrane. It is also suggested that lipid phase association of peptides is necessary in the other cases. At Pi = 100 mM inhibitory effects are observed for alamethicin and tetraacetyl melittin. Less pronounced inhibition is seen for melittin, while no such effect is noted for gramicidin A. The site of inhibition is shown to be complex IV, and the differences in the behavior of the peptides are rationalized in terms of channel structures.

Zervamicins, a structurally characterised peptide model for membrane ion channels

Voltage dependent membrane channels are formed by the zervamicins, a group of α-aminoisobutyric acid containing peptides. The role of polar residues like Thr, Gln and Hyp in promoting helical bundle formation is established by dramatically reduced channel lifetimes for a synthetic apolar analog. Crystal structures of Leu1-zervamicin reveal association of bent helices. Polar contacts between convex faces result in an ‘hour glass’ like arrangement of an aqueous channel with a central constriction. The structure suggests that gating mechanisms may involve movement of the Gln11 carboxamide group. Gln3 may play a role in modulating the size of the channel mouth.

Synthesis of neoglycopeptides and analyses of their biodistributionin vivo to identify tissue specific uptake and novel putative membrane lectins

Complex typeN-linked oligosaccharides derived from fetuin, fibrinogen and thyroglobulin were coupled to acetyltyrosine affording a series of neoglycopeptides with retention of terminal structures and the beta-anomeric configuration of their reducing endN-acetylglycosamine residue. The neoglycopeptides thus synthesized could be labelled to high specific activities with125I in the aromatic side chain of tyrosine. Analysis of the fate of these neoglycopeptides in conjunction with inhibition with asialofetuin and oligosaccharides of defined structure in micein vivo revealed the uptake of galactosylated biantennary compound by kidneys, in addition to the known itinerary of triantennary galactosylated complex oligosaccharide from fetuin to liver and the galactosylated biantennary chain with fucosylation in the core to bone marrows. On the other hand, the agalacto, aglucosamino biantennary chains with and without fucosylation in the core region are taken up by submaxillary glands while the conserved trimannosyl core with fucose is primarily concentrated in stomach tissue. These studies thus define new routes for the uptake of complexN-linked glycans and also subserve to identify lectins presumably involved in their recognition.

Functional role of the Mso1p-Sec1p complex in membrane fusion regulation

Sec1/Munc18 (SM) protein family members are evolutionary conserved proteins. They perform an essential, albeit poorly understood function in SNARE complex formation in membrane fusion. In addition to the SNARE complex components, only a few SM protein binding proteins are known. Typically, their binding modes to SM proteins and their contribution to the membrane fusion regulation is poorly characterised. We identified Mso1p as a novel Sec1p interacting partner. It was shown that Mso1p and Sec1p interact at sites of polarised secretion and that this localisation is dependent on the Rab GTPase Sec4p and its GEF Sec2p. Using targeted mutagenesis and N- and C-terminal deletants, it was discovered that the interaction between an N-terminal peptide of Mso1p and the putative Syntaxin N-peptide binding area in Sec1p domain 1 is important for membrane fusion regulation. The yeast Syntaxin homologues Sso1p and Sso2p lack the N-terminal peptide. Our results show that in addition to binding to the putative N-peptide binding area in Sec1p, Mso1p can interact with Sso1p and Sso2p. This result suggests that Mso1p can mimic the N-peptide binding to facilitate membrane fusion. In addition to Mso1p, a novel role in membrane fusion regulation was revealed for the Sec1p C-terminal tail, which is missing in its mammalian homologues. Deletion of the Sec1p-tail results in temperature sensitive growth and reduced sporulation. Using in vivo and in vitro experiments, it was shown that the Sec1p-tail mediates SNARE complex binding and assembly. These results propose a regulatory role for the Sec1p-tail in SNARE complex formation. Furthermore, two novel interaction partners for Mso1p, the Rab GTPase Sec4p and plasma membrane phospholipids, were identified. The Sec4p link was identified using Bimolecular Fluorescence Complementation assays with Mso1p and the non-SNARE binding Sec1p(1-657). The assay revealed that Mso1p can target Sec1p(1-657) to sites of secretion. This effect is mediated via the Mso1p C-terminus, which previously has been genetically linked to Sec4p. These results and in vitro binding experiments suggest that Mso1p acts in cooperation with the GTP-bound form of Sec4p on vesicle-like structures prior to membrane fusion. Mso1p shares homology with the PIP2 binding domain of the mammalian Munc18 binding Mint proteins. It was shown both in vivo and in vitro that Mso1p is a phospholipid inserting protein and that this insertion is mediated by the conserved Mso1p amino terminus. In vivo, the Mso1p phospholipid binding is needed for sporulation and Mso1p-Sec1p localisation at the sites of secretion at the plasma membrane. The results reveal a novel layer of membrane fusion regulation in exocytosis and propose a coordinating role for Mso1p in connection with membrane lipids, Sec1p, Sec4p and SNARE complexes in this process.

Wave propagation in a micropolar fluid contained in a visco-elastic membrane

The aim of the paper is to investigate the propagation of a pulse in a micropolar fluid contained in a visco-elastic membrane. It was undertaken with a view to study how closely we can approximate the flow of blood in arteries by the above model. We find that for large Reynolds number, the effect of micropolarity is hardly perceptible, whereas for small Reynolds numbers it is of considerable importance.

Frequency-Dependent Viscosity of Membrane Solutions

It is shown that dilute suspensions of membranes have strongly frequency-dependent viscosities. This behaviour should be seen in a variety of measurements such as capillary flow, mechanical impedance and ultrasound damping.

Experimental approach for studying structural changes in axonal membrane upon nerve excitation

The Hodgkin and Huxley (HH) model of action potential has become a central paradigm of neuroscience. Despite its ability to predict action potentials with remarkable accuracy, it fails to explain several biophysical findings related to the initiation and propagation of the nerve impulse. The isentropic heat release and optical phenomena demonstrated by various experiments suggest that action potential is accompanied by a transient phase change in the axonal membrane. In this study a method was developed for preparing a giant axon from the crayfish abdominal cord for studying the molecular mechanisms of action potential simultaneously by electrophysiological and optical methods. Also an alternative setup using a single-cell culture of an Aplysia sensory neuron is presented. In addition to the description of the method, the preliminary results on the effect of phloretin, a dipole potential lowering compound, on the excitability of a crayfish giant axon are presented.

PVA-SSA-HPA Mixed-Matrix-Membrane Electrolytes for DMFCs

Stabilized forms of heteropolyacids (HPAs), namely phosphomolybdic acid (PMA), phosphotungstic acid (PTA), and silicotungstic acid (STA), are incorporated into poly (vinyl alcohol) (PVA) cross-linked with sulfosuccinic acid (SSA) to form mixed-matrix membranes for application in direct methanol fuel cells (DMFCs). Bridging SSA between PVA molecules not only strengthens the network but also facilitates proton conduction in HPAs. The mixed-matrix membranes are characterized for their mechanical stability, sorption capability, ion-exchange capacity, and wetting in conjunction with their proton conductivity, methanol permeability, and DMFC performance. Methanol-release kinetics is studied ex situ by volume-localized NMR spectroscopy (employing point-resolved spectroscopy'') with the results clearly demonstrating that the incorporation of certain inorganic fillers in PVA-SSA viz., STA and PTA, retards the methanol-release kinetics under osmotic drag compared to Nafion, although PVA-SSA itself exhibits a still lower methanol permeability. The methanol crossover rate for PVA-SSA-HPA-bridged-mixed-matrix membranes decreases dramatically with increasing current density rendering higher DMFC performance in relation to a DMFC using a pristine PVA-SSA membrane. A peak power density of 150 mW/cm(2) at a load current density of 500 mA/cm(2) is achieved for the DMFC using a PVA-SSA-STA-bridged-mixed-matrix-membrane electrolyte. (C) 2010 The Electrochemical Society. [DOI: 10.1149/1.3465653] All rights reserved.

On the admittance of lipid bilayer membranes: Part I. Membrane-permeable ions

The modular formalism of Rangarajan [J. Electroanal. Chem., 55 (1974) 297] has been applied to the admittance of lipid bilayer membranes. The method leads to equations which clearly show the interrelations between the various partial processes involved in ion transport, and which allow examination of model assumptions without the need for a complete rederivation of the membrane admittance. Explicit expressions are given for both the continuum and single jump models. The former includes the ionic displacement component, important mostly at high frequencies.

On the admittance of lipid bilayer membranes: Part I. Membrane-permeable ions

The modular formalism of Rangarajan [J. Electroanal. Chem., 55 (1974) 297] has been applied to the admittance of lipid bilayer membranes. The method leads to equations which clearly show the interrelations between the various partial processes involved in ion transport, and which allow examination of model assumptions without the need for a complete rederivation of the membrane admittance. Explicit expressions are given for both the continuum and single jump models. The former includes the ionic displacement component, important mostly at high frequencies.

Stabilization of the bio-membrane by small molecules: interaction of trehalose with the phospholipid bilayer

Anhydrobiotic organisms undergo periods of acute dehydration during their life cycle. It is of interest to understand how the biomembrane remains intact through such stress. A disaccharide, trehalose, which is metabolised during anhydrobiosis is found to prevent disruption of model membrane systems. Molecular modelling techniques are used to investigate the possible mode of interaction of trehalose with a model monolayer. The objective is to maximise hydrogen bonding between the two systems. A phospholipid matrix consisting of 1,2-dimyristoyl-sn-glycero-3-phosphatidylcholine (DMPC) is chosen to represent the monolayer. The crystal structure of DMPC reveals that there are two distinct conformers designated as A and B. An expansion of the monolayer, coplanar with its surface, results in the trehalose molecule being accommodated in a pocket formed by four B conformers. One glucose ring of the sugar rests on the hydrophobic patch provided by the choline methyls of an A conformer. Five hydrogen bonds are formed involving the phosphate oxygens of three of the surrounding B conformers. The model will be discussed with reference to relevant experimental data on the interaction.