Low Negative Charge Density Research Articles

Molecular fingerprint in hyaluronan-peptide interactions: Cooperation of electrostatics and side-chain specificity in enhanced arginine effect.

Hyaluronan is a sugar polymer found on the extracellular side, close to the plasma membrane. It is the scaffold of the extracellular matrix due to its high solubility, despite its low negative charge density and unusual length. Hyaluronan is also a known ligand of several glycan-binding proteins. We can improve our understanding of these interactions by characterizing hyaluronan affinity to model amino acid sequences of reduced complexity. Extracted features and fingerprints allow for identifying new potential binding partners and expand the current understanding of hyaluronan's biological role. Here, we use a combination of NMR and molecular dynamics simulations (MD) to study hyaluronan binding to short oligopeptides. We focus on arginine as a known critical amino acid for this interaction and compare it to lysine and glycine to highlight its distinctive features. We can test the binding using 1H NMR chemical shift perturbation (CSP) by titrating hyaluronan with a peptide. CSP not only estimates binding strength but also pinpoints the binding motif involved. Our findings are further explained by MD, which provide an atomistic fingerprint of the hyaluronan-peptide complex. Interestingly, we could not find any NOE signal confirming the CSP observed interactions. The combination of positive CSP with no visible NOEs indicates the presence of a dynamic binding mode in agreement with our MD simulations. This agreement with simulations was only achieved using the prosECCo force field for accounting for electronic polarizability. Overall, we find that arginine, a positively charged amino acid, binds the strongest to the negative hyaluronan, followed by the positive lysine. The hyaluronan's carboxyl and amide groups are the driving force for interacting with the peptides. We prove the importance of the electrostatics interaction but also a specific sidechain affinity that favors arginine.

Influence of the negative charge density of metalate nanosheets on their bottom-up synthesis

Low negative charge density is required for the bottom-up synthesis of metalate nanosheets by an aqueous solution process.

A Fluorescent Probe for Glycosaminoglycans Applied to the Detection of Dermatan Sulfate by a Mix-and-Read Assay.

Glycosaminoglycans are complex biomolecules of great biological and medical importance. The quantification of glycosaminoglycans, in particular in complex matrices, is challenging due to their inherent structural heterogeneity. Heparin Red, a polycationic, fluorescent perylene diimide derivative, has recently emerged as a commercial probe for the convenient detection of heparins by a mix-and-read fluorescence assay. The probe also detects glycosaminoglycans with a lower negative charge density than heparin, although with lower sensitivity. We describe here the synthesis and characterization of a structurally related molecular probe with a higher positive charge of +10 (vs. +8 of Heparin Red). The superior performance of this probe is exemplified by the quantification of low dermatan sulfate concentrations in an aqueous matrix (quantification limit 1 ng/mL) and the detection of dermatan sulfate in blood plasma in a clinically relevant concentration range. The potential applications of this probe include monitoring the blood levels of dermatan sulfate after administration as an antithrombotic drug in the absence of heparin and other glycosaminoglycans.

Parallel Polyadenine Duplex Formation at Low pH Facilitates DNA Conjugation onto Gold Nanoparticles.

DNA-functionalized gold nanoparticles (AuNPs) have been extensively used in sensing, drug delivery, and materials science. A key step is to attach DNA to AuNPs, forming a stable and functional conjugate. Although the traditional salt-aging method takes a full day or longer, a recent low-pH method allows DNA conjugation in a few minutes. The effect of low pH was attributed to the protonation of adenine (A) and cytosine (C), resulting in an overall lower negative charge density on DNA. In this work, the effect of DNA conformation at low pH is studied. Using circular dichroism (CD) spectroscopy, the parallel poly-A duplex (A-motif) is detected when a poly-A segment is linked to a random DNA, a design typically used for DNA conjugation. A DNA staining dye, thiazole orange, is identified for detecting such A-motifs. The A-motif structure is ideal for DNA conjugation because it exposes the thiol group to directly react with the gold surface while minimizing nonspecific DNA base adsorption. For nonthiolated DNA, the optimal procedure is to incubate DNA and AuNPs followed by lowering the pH. The i-motif formed by poly-C DNA at low pH is less favorable to the conjugation reaction because of its unique way of folding. The stability of poly-A and poly-G DNA at low pH is examined. An excellent stability of poly-A DNA is confirmed, but poly-G has lower stability. This study provides new fundamental insights into a practically useful technique of conjugating DNA to AuNPs.

Parkinson’s Protein α-Synuclein Binds Efficiently and with a Novel Conformation to Two Natural Membrane Mimics

Binding of human α-Synuclein, a protein associated with Parkinson’s disease, to natural membranes is thought to be crucial in relation to its pathological and physiological function. Here the binding of αS to small unilamellar vesicles mimicking the inner mitochondrial and the neuronal plasma membrane is studied in situ by continuous wave and pulsed electron paramagnetic resonance. Local binding information of αS spin labeled by MTSL at positions 56 and 69 respectively shows that also helix 2 (residues 50–100) binds firmly to both membranes. By double electron-electron resonance (DEER) on the mutant spin labeled at positions 27 and 56 (αS 27/56) a new conformation on the membrane is found with a distance of 3.6 nm/ 3.7 nm between residues 27 and 56. In view of the low negative charge density of these membranes, the strong interaction is surprising, emphasizing that function and pathology of αS could involve synaptic vesicles and mitochondria.

Interparticle forces in spherical monodispersed silica dispersions: Effects of branched polyethylenimine and molecular weight

The effect of branched polyethylenimine (PEI) of molecular weight (Mw) 600, 1800 and 70,000 on the surface forces interacting between ‘uniform size’ spherical silica particles in water was investigated via the yield stress and zeta potential techniques. This silica has a point of zero charge at pH ∼ 2.0. All PEIs caused the zeta potential–pH curve and the high pH zero zeta potential to shift to a higher pH and the extent of the shift increases with increasing PEI concentration and is not affected by PEI Mw. PEI adsorption on silica is low or negligible at pH less than 3.5 and this is due to a very low negative charge density. Adsorption of PEI beyond 3.5 caused a maximum zeta potential to occur at pH between 4 and 6. The maximum yield stress located at the point zero zeta potential is many times larger than that with no added PEI. It ranged from 20 to 42 times for low Mw PEI and as high as 68 times for Mw 70,000. At low surface coverages, the force responsible for the high yield stress is charged patch–bridging attraction. At complete surface coverage, particle bridging via hydrogen bond and unlike charged attraction between monomeric, dimeric and tetrameric silicate ions with the adsorbed PEI layers of the interacting particles was responsible.

Natural lipid extracts and biomembrane-mimicking lipid compositions are disposed to form nonlamellar phases, and they release DNA from lipoplexes most efficiently

A viewpoint now emerging is that a critical factor in lipid-mediated transfection (lipofection) is the structural evolution of lipoplexes upon interacting and mixing with cellular lipids. Here we report our finding that lipid mixtures mimicking biomembrane lipid compositions are superior to pure anionic liposomes in their ability to release DNA from lipoplexes (cationic lipid/DNA complexes), even though they have a much lower negative charge density (and thus lower capacity to neutralize the positive charge of the lipoplex lipids). Flow fluorometry revealed that the portion of DNA released after a 30-min incubation of the cationic O-ethylphosphatidylcholine lipoplexes with the anionic phosphatidylserine or phosphatidylglycerol was 19% and 37%, respectively, whereas a mixture mimicking biomembranes (MM: phosphatidylcholine/phosphatidylethanolamine/phosphatidylserine /cholesterol 45:20:20:15 w/w) and polar lipid extract from bovine liver released 62% and 74%, respectively, of the DNA content. A possible reason for this superior power in releasing DNA by the natural lipid mixtures was suggested by structural experiments: while pure anionic lipids typically form lamellae, the natural lipid mixtures exhibited a surprising predilection to form nonlamellar phases. Thus, the MM mixture arranged into lamellar arrays at physiological temperature, but began to convert to the hexagonal phase at a slightly higher temperature, ∼ 40–45 °C. A propensity to form nonlamellar phases (hexagonal, cubic, micellar) at close to physiological temperatures was also found with the lipid extracts from natural tissues (from bovine liver, brain, and heart). This result reveals that electrostatic interactions are only one of the factors involved in lipid-mediated DNA delivery. The tendency of lipid bilayers to form nonlamellar phases has been described in terms of bilayer “frustration” which imposes a nonzero intrinsic curvature of the two opposing monolayers. Because the stored curvature elastic energy in a “frustrated” bilayer seems to be comparable to the binding energy between cationic lipid and DNA, the balance between these two energies could play a significant role in the lipoplex–membrane interactions and DNA release energetics.

Modeling the selective partitioning of cations into negatively charged nanopores in water

Partitioning and transport of water and small solutes into and through nanopores are important to a variety of chemical and biological processes and applications. Here we study water structure in negatively charged model cylindrical [carbon nanotube (CNT)-like] nanopores, as well as the partitioning of positive ions of increasing size (Na+, K+, and Cs+) into the pore interior using extensive molecular dynamics simulations. Despite the simplicity of the simulation system-containing a short CNT-like nanopore in water carrying a uniformly distributed charge of qpore=-ne surrounded by n (=0,...,8) cations, making the overall system charge neutral-the results provide new and useful insights on both the pore hydration and ion partitioning. For n=0, that is, for a neutral nanopore, water molecules partition into the pore and form single-file hydrogen-bonded wire spanning the pore length. With increasing n, water molecules enter the pore from both ends with preferred orientations, resulting in a mutual repulsion between oriented water molecules at the pore center and creating a cavity-like low density region at the center. For low negative charge densities on the pore, the driving force for partitioning of positive ions into the pore is weak, and no partitioning is observed. Increasing the pore charge gradually leads to partitioning of positive ions into the pore. Interestingly, over a range of intermediate negative charge densities, nanopores display both thermodynamic as well as kinetic selectivity toward partitioning of the larger K+ and Cs+ ions into their interior over the smaller Na+ ions. Specifically, the driving force is in the order K+>Cs+>Na+, and K+ and Cs+ ions enter the pore much more rapidly than Na+ ions. At higher charge densities, the driving force for partitioning increases for all cations-it is highest for K+ ions-and becomes similar for Na+ and Cs+ ions. The variation of thermodynamic driving force and the average partitioning time with the pore charge density together suggest the presence of free energy barriers in the partitioning process. We discuss the role of ion hydration in the bulk and in the pore interior as well as of the pore hydration in determining the barrier heights for ion partitioning and the observed thermodynamic and kinetic selectivities.

Surface charge on loose nanofiltration membranes

The main objective of this work was to study the effect of salt concentration and pH on the surface charge of loose nanofiltration (NF) membranes through streaming potential measurements. A Dow pilot unit equipped with a plate-and-frame module allowed the simultaneous test of two commercial NF membranes: Celgard N30F and Celgard NF-PES-10. Permeation of 0.2–18 mol/m3 NaCl solutions was carried out at constant flow rate and ambient temperature, 2.5–12.5 bar, whereas pH ranged from 5 to 7. The ion exchange capacity (IEC) of the membranes was assessed by potentiometric titration. The streaming potential developed in NaCl solutions (0.2–5 mol/m3; pH = 5–7) flowing along a thin rectangular slit formed by two parallel identical membranes was measured by an EKA electrokinetic analyser, which automatically calculated the zeta potential through the Helmholtz-Smoluchowsky equation and corrected it for the membrane surface conductance. The membrane surface charge was determined through the Gouy–Chapmann equation. Both membranes bear low negative charge densities, the absolute magnitude of which decreased for decreasing pH, as the membrane isoelectric point was approached. The absolute magnitude of the membrane surface charge increased for increasing concentrations, due to the progressive adsorption of anions onto the membrane surface, though it was impossible to glimmer a Freundlich adsorption isotherm, due to the scattering of streaming potential data at concentrations above 5 mol/m3. The ion exchange capacities of the membranes permselective layers were in the range of weak ion exchange membranes. However, IEC data were one order of magnitude higher than the maximum charge densities assessed by electrokinetic measurements.

Water-soluble single-walled carbon nanotubes films: Preparation, characterization and applications as electrochemical sensing films

Water-soluble single-walled carbon nanotubes (SWNTs) were prepared via noncovalent functionalization by Congo red through a physical grinding treatment. Based on the unique property of strong rebundling when dried, water-soluble SWNTs were firmly immobilized on the surface of a glassy carbon electrode by a simple casting method. The prepared films were characterized by scanning electron microscopy (SEM), cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). The results showed that water-soluble SWNTs formed uniform films with porous network structures of nanosizes on the electrode surface, which were stable in neutral and acidic solutions but were unstable in basic media. In addition, the structural properties and the negative charge density of these films can be conveniently controlled by choosing proper solvents during the washing procedure, providing a simple approach for adjusting their properties to specific applications. For instance, the films prepared from water-soluble SWNTs by N, N′-dimethyl formamide (DMF) washing (SWNTs–CR DMF) had looser structures and lower negative charge density than those by water washing (SWNTs–CR water), which allowed the entry and the succedent electrochemical reactions of both positively and negatively charged species inside the films. The potential applications of these films in electroanalytical chemistry were examined. The enhanced response of dopamine (DA) and the separation of DA oxidation potential from those of uric acid (UA) and ascorbic acid (AA) at these films demonstrated that the water-soluble SWNTs were the ideal materials for constructing SWNTs-based electrochemical sensing films.

Acidic Glycosaminoglycans in Marine Teleost Larvae: Evidence for a Relationship Between Composition and Negative Charge Density in Elopomorph Leptocephali

Glycosaminoglycan (GAG) composition was determined in larvae of 10 species of true eels (subdivision Elopomorpha; order Anguilliformes; families Chlopsidae, Congridae, Moringuidae, Muraenidae and Ophichthidae), two species from the subdivision Clupeomorpha (order Clupeiformes; families Engraulidae and Clupeidae) and one species from the subdivision Euteleostei (order Aulopiformes; family Synodontidae). Using a combination of specific GAG-degrading enzymes, cellulose acetate electrophoresis with known GAG standards and relative degree of staining with Alcian Blue, the predominant GAG in anguilliform leptocephali was identified as undersulfated chondroitin sulfate (uCS); the only exception was Bathycongrus macrurus (Congridae), where heparan sulfate was tentatively identified together with uCS. In representatives of the Clupeomorpha and Euteleostei, however, the predominant GAG was chondroitin sulfate; uCS was not detected. These results, combined with those from previous studies, provide data on GAG composition in 21 species from three of the four currently recognized elopomorph orders (Albuliformes, Elopiformes and Anguilliformes). The combined data suggest that although several different body matrix GAGs have evolved in different groups of leptocephali, selection has favored GAGs (mainly uCS and keratan sulfate) with relatively low average negative charge densities (average of about one negative charge per disaccharide repeat of the acidic GAG chain). The presence of normally sulfated CS (average per disaccharide negative charge density of 2) as the major GAG in the three species of non-elopomorph larvae further suggests that some aspect of the unique developmental pattern in elopomorph leptocephali may place constraints on the maximum negative charge density permitted in the body matrix GAGs. The presence of GAGs with low negative charge density is interpreted as an additional elopomorph synapomorphy, offering further support for the view that the Elopomorpha represents a monophyletic group of fishes. Possible scenarios for the evolution of GAGs in leptocephali are presented.

Interaction of Methylene Blue with Reduced Charge Montmorillonite

Interaction of methylene blue (MB) with reduced charge Li−montmorillonites (RCM) in the aqueous suspensions was investigated using visible absorption spectroscopy. Dye cation agglomeration and protonation at the clay surface depended very sensitively on layer charge density. With increasing layer charge reduction, the content of agglomerates of MB cations decreased in favor of monomers and the protonated form of MB. Lower negative charge density on the clay basal surface induces a greater distance between neighboring MB cations sorbed at the clay surfaces, which suppresses dye agglomeration. Since each form of MB absorbs visible light at a different wavelength, different layer charge densities induce different colors of the resulting clay−dye suspension. Therefore, visible spectra of MB−smectite suspensions may be a simple but extremely sensitive method for the detection of layer charge density of smectites.

Polymers as Soil Conditioners Under Consecutive Irrigations and Rainfall

AbstractLow water infiltration caused by crust formation during rain or sprinkler irrigation is a significant problem in some arid and semiarid regions. Polymers may be applied in irrigation water through a sprinkler system, but must be applied directly to the soil under rainfall conditions. The objectives of this rainfall simulator study were to: (i) determine the effect of drying of crusted vermiculitic soil on the subsequent crust properties and infiltration rate (IR) values, (ii) determine the effect of polymers applied at low concentration in irrigation water of two qualities on the IR under consecutive water applications, and (iii) determine the effectiveness of polymer application to the soil as would be required under rainfall conditions. Two cationic polysaccharide guar derivatives having a higher (HCCP) and a lower (LCCP) charge density and a polyacrylamide (PAM) with a low negative charge density were used in the study. Applications of polymers with the sprinkler water maintain IR in the order HCCP > LCCP > PAM > untreated. Except for PAM, the polymer applications were relatively ineffective in subsequent sprinkler applications with plain water applied with impact energy. The beneficial effects were preserved under water application without impact energy. Spraying concentrated polymer solutions on the soil surface was not effective in preventing crust formation by following rain events except for the case when LCCP was sprayed on in a CaCl2 solution. The results are explained on the basis of polymer adsorption and penetration into the soil surface layer and aggregates.

Mechanics of chromatin template activation. Physical evidence for destabilization of nucleoproteins by polyanions.

Thermal denaturation profiles were used to quantitate the destabilizing activity of five classes of polyanions in an effort to explain previously reported differences in activity among apparently similar polymers. This physical assay for polyanion action on chromatin is free of the biochemical complications associated with template assay systems. Most polyanions tested caused a thermal destabilization of the normal nucleohistone complexes of rat thymus chromatin and exposed free or weakly complexed DNA. Polyphosphates dissociated chromatin even when their net charge was as low as 6, and activity per unit of weight increased with chain length only for polymers containing less than about 20 phosphate units; pyrophosphate was inactive. Two polymers which had no influence on chromatin had a low negative charge density. Some, such as polycytidlate and denatured DNA, possessed a high charge but caused only minor changes in the shape of denaturation profiles and exposed little or no free DNA. The highly active nucleic acids, ribosomal RNA and polyguanylate, were distinguishable from inactive nucleic acids by having more secondary structure and potentially less steric hindrance for anionic interaction with chromatin. There was evidence that some polyanions dissociate certain histones before others. Although data obtained show that some histones may be redistributed from chromatin onto added double-stranded DNA in a medium containing urea, this does not appear to interfere significantly with the interpretation of typical thermal denaturation profiles of chromatin.

Negative Surface Charges of Unground Kaolinites

The negative surface charges of some kaolinites from different origins were determined, paying attention to the distribution of the charges on basal and edge surfaces. The relations between the charge density and the particle morphology or crystallinity were considered.The negative charges on the basal and edge surfaces were assessed from the C. E. C. values at pH 4 and from the difference in the C. E. C. values at pH 10 and pH 4, respectively. The crystallinity was evaluated from the ratio of the two peak intensities (I(021)/(I(060)) of X-ray diffraction. The particle shape was observed electron microscopically.The results obtained are summarized as follows;1) The kaolinites of nine species studied could be classified into two groups by the magnitude of the negative charge density.2) Four kaolinites which showed high negative charge densities were poorly stacked to the direction of c-axis. The particles were rather thin shape owing presumably to high degree of isomorphous substitution. The negative charge densities on the basal surfaces, particularly in fine particles, were larger than those on edge surfaces.3) The remaining five kaolinites having low negative chargede nsities were well stacked and the particles were rather thick. The negative charge densities on the basal surfaces were almost same as those on edge-faces.

STUDIES OF THE DISSOCIATION OF OXIDE SURFACES AT THE LIQUID–SOLID INTERFACE

The dissociation of surface hydroxyl groups of crystalline SiO2, ZrO2, and ThO2, in aqueous suspensions, has been studied as a function of pHs at different ionic strengths of KNO3. The surface groups of quartz dissociate as weak acids, while those of ZrO2 and ThO2 dissociate amphoterically. A reversible double layer is formed at the oxide–solution interface, and H+ and OH− function as the potential-determining ions. Quantitative data have been obtained on (a) surface charge densities, (b) zero point of charge, (c) differential capacities of the double layer on quartz and ZrO2, and (d) the effect of surface charge density and ionic strength on the interfacial tension. The differential capacities indicate specific adsorption of NO3−on ZrO2 and ThO2 while NO3− has zero affinity for quartz surfaces. At low negative charge densities, solvated K+ are adsorbed on quartz and ZrO2 through coulombic interactions while at pHs > 10 specific adsorption of K+ predominates. ThO2 appears to exhibit a greater tendency for the specific adsorption of K+.