Presence Of Negative Charge Research Articles

Scanning capacitance microscopy imaging of threading dislocations in GaN films grown on (0001) sapphire by metalorganic chemical vapor deposition

A combination of atomic force microscopy and scanning capacitance microscopy was used to investigate the relationship between the surface morphology and the near-surface electrical properties of GaN films grown on c-axis sapphire substrates by metalorganic chemical vapor deposition. Local regions surrounding the surface termination of threading dislocations displayed a reduced change in capacitance with applied voltage relative to regions that contained no dislocations. Capacitance–voltage characteristics obtained from these regions indicated the presence of negative charge in the vicinity of dislocations.

E.coli PhoE porin has an opposite voltage-dependence to the homologous OmpF.

We used patch clamp analysis to compare the electrophysiological behavior of two related porins from Escherichia coli, the anion-specific PhoE and the cation-selective OmpF. Outer membrane fractions were obtained from strains expressing just one of these porin types, and the channels were reconstituted into liposomes without prior purification. We show that the orientation of the reconstituted channels is not random and is the same for both PhoE and OmpF. Like cation-selective porins, PhoE shows fast and slow gating to closed levels of various amplitudes, testifying that the channels visit multiple functional states and behave as cooperative entities. The voltage-dependence of PhoE closure is asymmetric, but strikingly, occurs at voltages of inverse polarity from those promoting closures of OmpC and OmpF. Both slow kinetics and inverse voltage-dependence are removed when 70 amino acids from the N-terminal of OmpF are introduced into the homologous region of PhoE. This novel observation regarding the voltage-dependence of the two channel types, along with published results on PhoE and OmpF mutants, allows us to propose a molecular mechanism for voltage sensing and sensor charge movements in bacterial porins. It also offers new cues on the possible physiological relevance in bacteria of this common form of channel modulation.

On the molecular origin of the protein catalysis of the primary process in bacteriorhodopsin photosynthesis: Retinal photoisomerization

Abstract

Assessment of oxide charge density and centroid from Fowler-Nordheim derivative characteristics in MOS structures after uniform gate stress

A new method for the extraction of the oxide charge density and distribution centroid based on the exploitation of the Fowler plot derivative characteristics is proposed. The comparison with the DiMaria method confirms the overall consistency of the new approach. The presence of negative charge within the oxide is shown to be responsible for an increase in the apparent Fowler barrier height after uniform gate stress.

Effect of acidic phospholipids on sphingosine kinase.

Sphingosine-1-phosphate (SPP) is a unique sphingolipid metabolite involved in cell growth regulation and signal transduction. SPP is formed from sphingosine in cells by the action of sphingosine kinase, an enzyme whose activity can be stimulated by growth factors. Little is known of the mechanisms by which sphingosine kinase is regulated. We found that acidic phospholipids, particularly phosphatidylserine, induced a dose-dependent increase in sphingosine kinase activity due to an increase in the apparent Vmax of the enzyme. Other acidic phospholipids, such as phosphatidylinositol, phosphatidic acid, phosphatidylinositol bisphosphate, and cardiolipin stimulated sphingosine kinase activity to a lesser extent than phosphatidylserine, whereas neutral phospholipids had no effect. Diacylglycerol, a structurally similar molecule which differs from phosphatidic acid in the absence of the phosphate group, failed to induce any changes in sphingosine kinase activity. Our results suggest that the presence of negative charges on the lipid molecules is important for the potentiation of sphingosine kinase activity, but the effect does not directly correlate with the number of negative charges. These results also support the notion that the polar group confers specificity in the stimulation of sphingosine kinase by acidic glycerophospholipids. The presence of a fatty acid chain in position 2 of the glycerol backbone was not critical since lysophosphatidylserine also stimulated sphingosine kinase, although it was somewhat less potent. Dioleoylphosphatidylserine was the most potent species, including a fourfold stimulation, whereas distearoyl phosphatidylserine was completely inactive. Thus, the degree of saturation of the fatty acid chain of the phospholipids may also play a role in the activation of sphingosine kinase.

Structural determinants in the interaction of Shaker inactivating peptide and a Ca(2+)-activated K+ channel.

Shaker B inactivating peptide (BP) binds to its receptor in maxi KCa channels obstructing the flow of ions through them. The interaction between KCa channels and BP mutants, with different net charge and hydrophobicity, revealed several structural features of the KCa channel internal mouth. Increasing BP net positive charge or decreasing the internal milieu ionic strength increased the affinity and rate of association, while increasing hydrophobicity augmented blocking times and had limited or no effect on on-rates. These results uncover (a) the presence of negative charges in or near the BP receptor and (b) the existence of a hydrophobic contact surface in the internal channel vestibule that is a structural constituent of the BP receptor in maxi KCa channels.

Surface potentials near the mouth of the large-conductance K+ channel from Chara australis: a new method of testing for diffusion-limited ion flow.

The kinetics of single K+ channels were derived for patch-clamp recordings of membrane patches excised from cytoplasmic drops from the plant, Chara australis R. Br. Specifically, the "tilt effect" model of MacKinnon, Latorre and Miller (1989. Biochemistry 28:8092-8099) has been used to measure the electrostatic potential (surface PD) and fixed charge at the entrances of the channel. The surface PD is derived from the difference between the trans-pore potential difference (PD) and that between the two bulk phases. The trans-pore PD is probed using three voltage-dependent properties of the channel. These are (1) the association and dissociation rates of Ca2+ binding to the channel, from both the cytoplasmic and vacuolar solutions. These were determined from the mean blocked and unblocked durations of the channel in the presence of either 20 mmol liter-1 vacuolar or 1 mmol liter-1 cytoplasmic Ca2+; (2) the closing rate of the channel's intrinsic gating process. This was determined from the mean channel open time in the absence of vacuolar Ca2+ at membrane PDs more negative than -100 mV; and (3) the effect of Mg2+ on channel conductance when added to solutions initially containing 3 mmol liter-1 KCl. The voltage dependence of properties 1 and 2 shifts along the voltage axis according to the ionic strength of the bathing media, consistent with the presence of negative charge in the channel vestibules. Furthermore, the magnitude of this shift depends on the current in a manner consistent with diffusion-limited ion flow in the channel (i.e., the rate of ion diffusion in the external electrolyte limits the channel conductance). Mg2+ on either side of the membrane alters channel conductance in a voltage-dependent way. A novel feature of the Mg2+ effect is that it reverses, from a block to an enhancement, when the membrane PD is more negative than -70 mV. This reversal only appears in solutions of low ionic strength. The attenuating effect is due to voltage-dependent binding of Mg2+ within the pore, which presumably plugs the channel. The enhancing effect is due to screening by Mg2+ of surface potentials arising from diffusion-limited flow of K+. All experimental approaches give a consistent picture of K+ permeation in which the surface charge and convergence permeability of the cytoplasmic vestibule are the major factors in determining channel conductance. The cytoplasmic vestibule has a charge density of -0.035 C/m2 which is similar to that found for maxi K channels in rat muscle.(ABSTRACT TRUNCATED AT 400 WORDS)

Asymmetric electrostatic effects on the gating of rat brain sodium channels in planar lipid membranes

The effects of ionic strength (10-1,000 mM) on the gating of batrachotoxin-activated rat brain sodium channels were studied in neutral and in negatively charged lipid bilayers. In neutral bilayers, increasing the ionic strength of the extracellular solution, shifted the voltage dependence of the open probability (gating curve) of the sodium channel to more positive membrane potentials. On the other hand, increasing the intracellular ionic strength shifted the gating curve to more negative membrane potentials. Ionic strength shifted the voltage dependence of both opening and closing rate constants of the channel in analogous ways to its effects on gating curves. The voltage sensitivities of the rate constants were not affected by ionic strength. The effects of ionic strength on the gating of sodium channels reconstituted in negatively charged bilayers were qualitatively the same as in neutral bilayers. However, important quantitative differences were noticed: in low ionic strength conditions (10-150 mM), the presence of negative charges on the membrane surface induced an extra voltage shift on the gating curve of sodium channels in relation to neutral bilayers. It is concluded that: (a) asymmetric negative surface charge densities in the extracellular (1e-/533A2) and intracellular (1e-/1,231A2) sides of the sodium channel could explain the voltage shifts caused by ionic strength on the gating curve of the channel in neutral bilayers. These surface charges create negative electric fields in both the extracellular and intracellular sides of the channel. Said electric fields interfere with gating charge movements that occur during the opening and closing of sodium channels; (b) the voltage shifts caused by ionic strength on the gating curve of sodium channels can be accounted by voltage shifts in both the opening and closing rate constants; (c) net negative surface charges on the channel's molecule do not affect the intrinsic gating properties of sodium channels but are essential in determining the relative position of the channel's gating curve; (d) provided the ionic strength is below 150 mM, the gating machinery of the sodium channel molecule is able to sense the electric field created by surface changes on the lipid membrane. I propose that during the opening and closing of sodium channels, the gating charges involved in this process are asymmetrically displaced in relation to the plane of the bilayer. Simple electrostatic calculations suggest that gating charge movements are influenced by membrane electrostatic potentials at distances of 48 and 28 A away from the plane of the membrane in the extracellular sides of the channel, respectively.

Fluorine-enhanced nitridation of silicon at low temperatures in a microwave plasma

A microwave plasma discharge at very low temperatures (200–250 °C) has been used for the growth of thin silicon oxynitride films suitable for gate dielectric applications. The addition of CHF3 as a source of fluorine enhances the growth rate. A x-ray photoelectron spectroscopy study indicates the incorporation of fluorine (F/Si≂0.2) in the film. Electrical properties of the grown layers have been evaluated by the characterization of metal-insulator-semiconductor capacitors. Results have indicated the presence of negative charges in the insulator. The estimated charge density is lowest for the fluorinated film. The conduction mechanism in the films at room temperature appears to be Frenkel–Poole type.

The Amino Acid Sequence of Zinc-Carboxypeptidase from Streptomyces griseus

The amino acid sequence of a zinc-carboxypeptidase from S. griseus (Cpase SG) was determined by automated Edman degradation and carboxypeptidase digestion of the S-carboxymethylated protein and by sequence analyses of peptides produced by cyanogen bromide cleavage and by lysyl endopeptidase digestion of the S-carboxymethylated protein. This enzyme is characterized by a uniquely broad substrate specificity which combines the specificities of mammalian Cpase A and Cpase B (J. Biochem. 86, 683-694, 1979). Cpase SG consists of 328 amino acid residues. The amino acid sequence of Cpase SG is partially similar to those of bovine Cpase A and Cpase B (sequence identity, 28-29%). In the sequence of Cpase SG, residues that are functionally important in mammalian Cpase A and Cpase B were all found at the corresponding positions. Residue 255 (according to the numbering system for bovine Cpase A), which, in the other Cpases, contributes to the difference in specificity between Cpase A (Ile-255) and Cpase B (Asp-255), was Asp. However, residue 254 was Ile, in contrast to Ser or Thr in all of the forms of Cpase A and Cpase B examined to date. The increase in hydrophobicity caused by the change at position 254 and the presence of negative charge at position 255 is probably one of the reasons for the broad substrate specificity of Cpase SG.

Activation of porcine pancreatic phospholipase A2 by the presence of negative charges at the lipid-water interface.

The effect of surface charge on the porcine pancreatic phospholipase A2 catalyzed hydrolysis of organized substrates was examined through initial rate enzyme kinetic measurements. Two long-chain phospholipid substrates, phosphatidylglycerol (PG) and phosphatidylcholine (PC), were solubilized in seven detergents differing in polar head-group charge. The neutral or zwitterionic detergents selected were Triton X-100, Zwittergent 314, lauryl maltoside, hexadecylphosphocholine (C16PN), and 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate. The negatively and positively charged detergents used were cholate and CTAB, respectively. In general, the negatively charged phospholipid PG was hydrolyzed much more rapidly than the neutral (zwitterionic) phospholipid PC. The rate of hydrolysis of PG was rapid when solubilized in all the neutral detergents and in cholate but was essentially zero in the positively charged CTAB. Conversely, hydrolysis of PC was negligible when solubilized in neutral detergents, except C16PN, and was maximal in the negatively charged detergent, cholate. The rate of hydrolysis of PC solubilized in a neutral detergent became significant only when a negative surface charge was introduced by addition of SDS. Taken together, these kinetic measurements indicate that the surface charge on the lipid aggregates is an important factor in the rate of hydrolysis of phospholipid substrates and the highest activity is observed when the net surface charge is negative. Fluorescence and electron spin resonance (ESR) spectroscopic data provide additional support for this conclusion. The fluorescence emission spectrum of the single tryptophan of phospholipase A2 is a sensitive monitor of interfacial complex formation and shows that interaction of the protein with detergent micelles is strongly dependent on the presence of a negatively charged amphiphile.(ABSTRACT TRUNCATED AT 250 WORDS)

Modification of the outer surface of Erwinia carotovora ssp. atroseptica by the phytoalexin rishitin

The phytoalexin rishitin (500 μg/mL) modified the wall of Erwinia carotovora ssp. atroseptica by removing an outer layer and causing holes to appear through which the cell contents were extruded. A lower concentration of rishitin (300 μg/mL) produced effects on the cytoplasm visible by electron microscopy but no obvious differences in membrane integrity, although sections stained with the periodic acid – thiosemicarbazide – silver proteinate reaction showed a reduction in the intensity with which the carbohydrate layer in the outer membrane was stained which was similar to that found at the higher concentration (500 μg/mL). Labelling rishitin-treated cells with cationized ferritin showed the presence of negative charges on the outer surface of all cells treated with 500 μg rishitin/mL which were absent on nontreated cells and present in only a few cells treated with 300 μg/mL. After incubation of E. carotovora ssp. atroseptica with rishitin, cells bound to fluorescein isothiocyanate labelled lectins from Ricinus communis, Solanum tuberosum, and Triticum vulgaris and rhodamine-labelled avidin; cells of E. carotovora spp. atroseptica treated with ethanol were not labelled. Rishitin-treated cells did not bind to lectins from Arachis hypogaea, Dolichos biflorus, Glycine max, Limulus polyphemus, or Ulex europaeus. These results suggest that the phytoalexin rishitin can modify existing sites or expose previously blocked sites which may be involved in host–pathogen recognition, and may therefore indicate a reinforcement of other recognition processes in vivo.

Instability of MOSFET's due to redistribution of Oxide charges

It has been reported that electrical instability of MOSFET's occurs with the encapsulation of silicon nitride when these devices are stressed. In order to determine the mechanisms responsible, MOSFET's with and without this capsulation were stressed under combinations of biases which were different from those previously utilized. It is observed that another form of instability is caused by the drain-gate-substrate interaction such that when an abnormally high bias is applied to the drain, oxide charges are redistributed in the direction parallel to the oxide-semiconductor interface. Measurements show that after the stress, net positive charge is accumulated above the channel adjacent to the drain while net negative charge is accumulated in the drain-gate overlap region. The presence of negative charge is apparently associated with hydrogen in the plasma silicon nitride. The positive charge is present with or without the nitride layer and our experiments point out another source of instability other than hot-carrier trapping in devices even without a SiN cap for a drain bias higher than some critical value. It is concluded that this instability is the same as that manifested by the well-known walk-out effect in gated diodes.

Separation of aldehydes and ketones by chromatography on dowex-50 in the ethylenediamine form

A technique is developed that separates a number of carbonyl-containing compounds on the basis of chemical interaction with ethylenediamine in a column of polystyrene resin (AG-50W-X2) in the ethylenediamine form eluted with 10 m m ethylenediamine-phosphate, pH 8.0, at 25°C. Aldehydes are retained more strongly than ketones on this column. Relative elution volumes for several representative compounds are acetaldehyde > glyceraldehyde > 2,5-anhydromannose (chitose) > cyclohexanone > dihydroxyacetone > glyceraldehyde 3-phosphate > pyruvate > dihydroxyacetone phosphate ≥ glucose, chitose 6-phosphate, and glucose 1-phosphate. Within a given class of compounds the extent of hydration does not appear to affect how well a substance is retained, whereas the presence of negative charge(s) appears to decrease retention. In the absence of anionic functional groups, the retention of carbonyl compounds corresponds roughly to what is known about the relative stability of imines.

The interaction of atebrin with phospholipid vesicles

The interaction of atebrin with phosphatidylcholine and phosphatidylcholine-phosphatidic acid vesicles has been followed by equilibrium dialysis, and by photometric, fluorimetric and NMR techniques. The presence of negative charges in the phospholipids enhances the binding of atebrin. The absorbance and NMR spectral changes and fluorescence quenching occurring with phosphatidic acid are attributed to dimerization of the dye interacting electrostatically with negative groups. The dissociation constant of the binding of the dye to phosphatidylcholine vesicles was 1.4 mM; those of binding to the negative sites of phosphatidic acid were approx. 150 and 3 μM. The dye is probably located at the interphase with the acridine ring interacting with the anionic groups of phosphatidic acid and the tail freely floating in the aqueous phase. The results are discussed also in view of the use of atebrin as a probe of the energized state in natural membranes and of the suggestion that atebrin may be used as a transmembrane pH indicator in liposomes or natural membranes.