Negative Membrane Surface Charge Research Articles

Nanofiltration membrane functionalization with enhanced hardness cation removal using a mono-component interlayer

Current drinking water treatment demands new nanofiltration (NF) membranes to achieve efficient separation performance of both divalent hardness cations and new contaminants (ECs). In this work, we utilized sodium alginate as an interlayer to fabricate modified NF membrane with enhanced separation capacity. The interlayer was formed on substrate before interfacial polymerization reaction, controlling the irregular mass transfer of piperazine monomers and forming a crosslinked network at the bottom of the active layer to enhance the separation performance. Alginate modification method was proved achieving excellent separation of hardness cations by the surface potential shift and sharpened pore size distribution, while exhibiting high rejection for SO42− (≥99.00 %) by maintaining a negative membrane surface charge. The modified membrane provided efficient hardness removal (90.79 % for Ca2+, 95.94 % for Mg2+) and enhanced anionic ECs rejection (95.24 % for PFOA, 98.44 % for PFOS), while maintaining operational stability, thus providing drinking water treatment potential for groundwater with high total dissolved solids.

Transport of trace organic compounds through novel forward osmosis membranes: Role of membrane properties and the draw solution

Forward osmosis (FO) offers to be a very promising technology for the removal of trace organic compounds (TrOCs) from contaminated wastewater, and with the recent developments in FO membranes, the effect of both a higher water flux and reverse salt flux on the rejection of TrOCs needs to be explored. In this study two novel thin-film composite (TFC) membranes with greater water permeability and selectivity than the benchmark cellulose tri-acetate (CTA) membrane were compared at bench-scale in terms of TrOCs permeability. By probing the solute-membrane interactions that dictate the transport of TrOCs through the two membranes in the absence and presence of a draw solution, several conclusions were drawn. Firstly, steric hindrance is the main TrOCs transport -limiting mechanism through TFC membranes unless the negative membrane surface charge is significant, in which case, electrostatic interactions can dominate over steric hindrance. Secondly, the increase in ionic strength induced by the draw solution in the vicinity of and perhaps inside the membrane seems to favour the rejection of TrOCs by “shrinking” the membrane pores or by “shielding” the negative surface charge. Lastly, during FO operation, solute concentration polarisation becomes detrimental when working at high water fluxes, whereas the reverse solute flux has no direct impact on the transport of TrOCs through the membrane.

Relating Silica Scaling in Reverse Osmosis to Membrane Surface Properties

We investigated the relationship between membrane surface properties and silica scaling in reverse osmosis (RO). The effects of membrane hydrophilicity, free energy for heterogeneous nucleation, and surface charge on silica scaling were examined by comparing thin-film composite polyamide membranes grafted with a variety of polymers. Results show that the rate of silica scaling was independent of both membrane hydrophilicity and free energy for heterogeneous nucleation. In contrast, membrane surface charge demonstrated a strong correlation with the extent of silica scaling (R2 > 0.95, p < 0.001). Positively charged membranes significantly facilitated silica scaling, whereas a more negative membrane surface charge led to reduced scaling. This observation suggests that deposition of negatively charged silica species on the membrane surface plays a critical role in silica scale formation. Our findings provide fundamental insights into the mechanisms governing silica scaling in reverse osmosis and highlight the potential of membrane surface modification as a strategy to reduce silica scaling.

The impacts of operational conditions on phenol removal by nanofiltration membranes

The main purpose of this study was to investigate the effectiveness of various nanofiltration (NF) membranes in rejecting phenol from waters. The impacts of operational conditions and water chemistry on phenol rejections in model solutions were studied. Single-solute phenol model solutions were prepared in distilled and deionized water. All membrane tests were conducted using a lab-scale cross-flow flat-sheet configuration test unit. For all the tested NF membranes, the main factor affecting phenol rejections was solution pH. Increasing solution pH from 7.0 to 10.5 significantly increased phenol rejections. Both the enhanced negative membrane surface charge at higher pH and the dominance of negatively charged phenolate species led to higher phenol rejections at pH 10.5 compared to neutral pH. No significant changes in flux values were observed at neutral pH and pH of 10.5. There was not a correlation among salt and phenol rejections by the tested polyamide NF membranes. The initial phenol concentrations (50 and 200 mg/l) did not affect permeate flux and phenol rejections. Phenol rejections increased by increasing the pressure from 100 to 200 psi. The results overall indicated that although both size exclusion and electrostatic repulsion mechanisms play role in phenol rejections by NF membranes, electrostatic repulsion mechanism contributes more to rejections than size exclusion mechanism. Supporting the electrostatic repulsion mechanism through pH increase in NF applications may result in both enhanced phenol rejections and prevention of phenol adsorption and further diffusion inside the membranes. On the other hand, the pH adjustments should be within the range of manufacturer specifications since increased membrane degradation could occur at higher pH values in polyamide membranes.

Osmosensing and osmosignaling in Corynebacterium glutamicum

The Gram-positive soil bacterium Corynebacterium glutamicum is used in microbial biotechnology for the large-scale production of amino acids, e.g., L: -glutamate and L: -lysine. We have studied the response of this organism to hyperosmotic challenge at the level of both transcription and protein activity. Two systems responding to hyperosmotic stress in C. glutamicum are reviewed here, the two component system MtrAB and the glycine-betaine uptake system BetP. The osmosensory two-component system consists of the membrane-bound histidine kinase MtrB and the soluble response regulator MtrA. MtrB was shown to perceive a so far unknown physical stimulus related to hyperosmotic stress via the cytoplasmically oriented phosphorylation domain, and to transduce the signal to the DNA via MtrA. The secondary active transporter BetP takes up betaine in cotransport with two Na(+) ions. BetP responds to hyperosmotic stress by increased transcription mediated via MtrAB signaling, and by instant activation of transport. In the mechanism of BetP activation, the C-terminal, regulatory domain of BetP, the cytoplasmic concentration of K(+), and negative membrane surface charges are involved. The molecular mechanism of the activation process is discussed in relation to the recently published X-ray structure of BetP.

Influence of Membrane Surface Charge and Post-Translational Modifications to Myelin Basic Protein on Its Ability To Tether the Fyn-SH3 Domain to a Membrane in Vitro

Myelin basic protein (MBP) is a highly post-translationally modified, multifunctional structural component of central nervous system myelin, adhering to phospholipid membranes and assembling cytoskeletal proteins, and has previously been shown to bind SH3 domains in vitro and tether them to a membrane surface [Polverini, E., et al. (2008) Biochemistry 47, 267-282]. Since molecular modeling shows that the Fyn-SH3 domain has a negative surface charge density even after binding the MBP ligand, we have investigated the influence of negative membrane surface charge and the effects of post-translational modifications to MBP on the interaction of the Fyn-SH3 domain with membrane-associated MBP. Using a sedimentation assay with multilamellar vesicles consisting of neutral phosphatidylcholine (PC) and negatively charged phosphatidylinositol (PI), we demonstrate that increasing the negative surface charge of the membrane by increasing the proportion of PI reduces the amount of Fyn-SH3 domain that binds to membrane-associated MBP, due to electrostatic repulsion. When one of the phosphoinositides, PI(4)P or PI(4,5)P(2) was substituted for PI in equal proportion, none of the Fyn-SH3 domain bound to MBP under the conditions that were used. Post-translational modifications of MBP which reduced its net positive charge, i.e., phosphorylation or arginine deimination, increased the degree of repulsion of Fyn-SH3 from the membrane surface, an effect further modulated by the lipid charge. This study suggests that changes in membrane negative surface charge due to protein or lipid modifications, which could occur during cell signaling, can regulate the binding of the Fyn-SH3 domain to membrane-associated MBP and thus could regulate the activity of Fyn at the oligodendrocyte membrane surface.

Anticancer &amp;#945;-Helical Peptides and Structure / Function Relationships Underpinning Their Interactions with Tumour Cell Membranes

Cancer is a major cause of premature death and there is an urgent need for new anticancer agents with novel mechanisms of action. Here we review recent studies on a group of peptides that show much promise in this regard, exemplified by arthropod cecropins and amphibian magainins and aureins. These molecules are alpha-helical defence peptides, which show potent anticancer activity (alpha-ACPs) in addition to their established roles as antimicrobial factors and modulators of innate immune systems. Generally, alpha-ACPs exhibit selectivity for cancer and microbial cells primarily due to their elevated levels of negative membrane surface charge as compared to non-cancerous eukaryotic cells. The anticancer activity of alpha-ACPs normally occurs at micromolar levels but is not accompanied by significant levels of haemolysis or toxicity to other mammalian cells. Structure/function studies have established that architectural features of alpha-ACPs such as amphiphilicty levels and hydrophobic arc size are of major importance to the ability of these peptides to invade cancer cell membranes. In the vast majority of cases the mechanisms underlying such killing involves disruption of mitochondrial membrane integrity and/or that of the plasma membrane of the target tumour cells. Moreover, these mechanisms do not appear to proceed via receptor-mediated routes but are thought to be effected in most cases by the carpet/toroidal pore model and variants. Usually, these membrane interactions lead to loss of membrane integrity and cell death utilising apoptic and necrotic pathways. It is concluded that that alpha-ACPs are major contenders in the search for new anticancer drugs, underlined by the fact that a number of these peptides have been patented in this capacity.

A Study of the Regional Effects of α-Synuclein on the Organization and Stability of Phospholipid Bilayers

Associations between the protein alpha-synuclein (alpha-syn) and presynaptic vesicles have been implicated in synaptic plasticity and neurotransmitter release and may also affect how the protein aggregates into fibrils found in Lewy bodies, the cellular inclusions associated with neurodegenerative diseases. This work investigated how alpha-syn interacts with model phospholipid membranes and examined what effect protein binding has upon the physical properties of lipid bilayers. Wide line 2H and 31P NMR spectra of phospholipid vesicles revealed that alpha-syn associates with membranes containing lipids with anionic headgroups and can disrupt the integrity of the lipid bilayer, but the protein has little effect on membranes of zwitterionic phosphatidylcholine. A peptide, alpha-syn(10-48), which corresponds to the lysine-rich N-terminal region of alpha-syn, was found to associate with lipid headgroups with a preference for a negative membrane surface charge. Another peptide, alpha-syn(120-140), which corresponds to the glutamate-rich C-terminal region, also associates weakly with lipid headgroups but with a slightly higher affinity for membranes with no net surface charge than for negatively charged membrane surfaces. Binding of alpha-syn(10-48) and alpha-syn(120-140) to the lipid vesicles did not disrupt the lamellar structure of the membranes, but both peptides appeared to induce the lateral segregation of the lipids into clusters of acidic lipid-enriched and acidic lipid-deficient domains. From these findings, it is speculated that the N-terminal and C-terminal domains of full-length alpha-syn might act in concert to organize the membrane components during normal protein function and perhaps play a role in presynaptic vesicle synthesis, maintenance, and fusion.

Involvement of membrane surface charge in thermal stability of the rat ovarian LH/hCG receptor

Analysis of fluorescence of membrane-bound 8-anilino-1-naphthalene sulfonate and monodansylcadaverine probes revealed that a negative membrane surface charge derived from free fatty acids (FFA) resulted in destabilization of structure–functional properties of the rat ovarian LH/hCG receptor. Removal of FFA from rat luteal and porcine ovarian granulosa cells by BSA increased gonadotropin responsiveness of cells in cAMP formation.

Altered gating of HERG potassium channels by cobalt and lanthanum.

Activation of the rapid, delayed rectifier K current (IKr) is important for normal repolarization of cardiac action potentials, especially in mammalian ventricular muscle. The study of this current has been greatly aided by the discovery that the human ether-a-go-go-related gene (HERG) encodes the pore-forming alpha subunits of these channels. As for other voltage-activated K+ channels, divalent and trivalent cations affect the gating of HERG channels by screening negative membrane surface charges or by direct interaction with the channel gating mechanism. Previous studies have reported that IKr of myocytes, and HERG channels heterologously expressed in Xenopus oocytes, are reduced by external Co2+ and La3+. We have reinvestigated the "blocking" effect of Co2+ and La3+ on HERG channels expressed in Xenopus oocytes. At concentrations previously reported to block IKr or HERG current (IHERG), Co2+ (10 mM) and La3+ (10 microM) had only small effects on the magnitude of fully activated IHERG. The apparent block results from altered kinetics and voltage dependence of gating, similar to the effects of Ca2+ on HERG channels. Under control conditions, the half-points for voltage-dependent activation and inactivation of HERG were -35+/-2.1 and -76.3+/-1.7 mV, respectively. Co2+ and La3+ accelerated the rate of deactivation, decreased the rate of current activation, and shifted the half-point of the HERG channel activation curve by +53 and +65 mV, respectively. Co2+ shifted the voltage dependence of inactivation by + 14 mV, whereas La3+ had no effect. Co2+ also slowed the onset of IHERG inactivation and accelerated the rate of recovery from inactivation. These results indicate that reduction of IHERG by Co2+ (10 mM) and La3+ (10 microM) during depolarizing pulses is caused by a positive shift in the voltage dependence of activation, and does not result from pore block.

Thermodynamics of the membrane insertion process of the M13 procoat protein, a lipid bilayer traversing protein containing a leader sequence.

For the first time, the standard free energy change, delta Gzero, of a membrane-inserting protein with a leader sequence has been determined experimentally, using M13 procoat protein as an example. The partition coefficient for the distribution of the procoat protein between the aqueous phase and the membrane phase of preformed lipid vesicles yielded a value of gamma = 6.5 x 10(5) M-1, corresponding to a delta Gzero of -10.4 kcal/mol, based on measurements of the fluorescence energy transfer between the intrinsic tryptophan of the protein and a suitably labeled lipid membrane of POPC. For comparison, the partition coefficient of the M13 coat protein between the aqueous and the POPC lipid bilayer phase was determined to be distinctly lower: gamma = 1 x 10(5) M-1 (delta Gzero = -9.3 kcal/mol). Proteinase K digestion experiments have been performed, showing that 20% of the procoat protein bound to lipid vesicles spontaneously integrate in a transbilayer form, whereas 80% remain inserted in the interfacial membrane region. By taking together these results, an upper limit for the free energy change of the transmembrane insertion of procoat protein was estimated to be -14.8 kcal/mol. In order to distinguish further the contribution arising from insertion of the procoat protein into the membrane interfacial region from that due to transmembrane insertion, the partition coefficient of the mutant procoat protein OM30R [which contains a positively charged amino acid in its mature hydrophobic segment (exchange of a Val to an Arg residue at position 30)] was determined, yielding gamma = 0.3 x 10(5) M-1 (delta Gzero = -8.6 kcal/mol). Previously reported in vivo experiments have shown that the OM30R mutant protein is not translocated across Escherichia coli membranes but only binds to the inner surface. The results presented here indicate that although the insertion of the procoat protein into the interfacial region of the lipid bilayer contributes the major part to delta Gzero, it is the final energy gain of the interaction of the hydrophobic portions of the folded pre-protein with the lipid chains which drives the transmembrane insertion of the M13 procoat protein. Neither the leader sequence nor the mature coat protein alone yields this free energy gain. For the different proteins investigated here, spontaneous membrane insertion occurs only for fluid lipid bilayers, but not for membranes in the crystalline lipid phase. Furthermore, by using lipid bilayers with negative membrane surface charges, it was shown that both procoat and coat proteins are electrostatically attracted to the surface of the lipid membrane, though only to a small extent, with apparent partition coefficients of the same order of magnitude as for the phosphatidylcholine lipid membrane.

Structural basis of potent antiperoxidation activity of the triterpene celastrol in mitochondria: Effect of negative membrane surface charge on lipid peroxidation

The structural basis of the potent inhibitory effect of the triterpene celastrol on lipid peroxidation of rat liver mitochondria initiated by adenosine 5′-diphosphate (ADP) and Fe 2+ was studied in comparison with the effects of its analogs, pristimerin and acetylcelastrol. The dienone-phenol moiety and the anionic carboxyl group of celastrol were concluded to be important for their antiperoxidative action: The former moiety directly scavenges radicals and the latter donates the membrane with a negative surface charge, making it more resistant to peroxidation. Celastrol is suggested to inhibit the peroxidation of the outer and inner mitochondrial membrane by direct radical scavenging, and also to prevent the attack of oxygen radicals on the inner membrane by increasing its negative surface charge.

Influence of lipid lateral distribution on the surface charge response of the phosphatidylcholine headgroup as detected using 2H nuclear magnetic resonance

The effect of lipid lateral distribution on the surface charge response of the phosphatidylcholine headgroup, in bilayers composed of binary mixtures of 1-palmitoyl-2-oleoyl- sn-glycero-3-phosphocholine (POPC) and 1,2-dimyristoyl- sn-glycero-3-phosphate (DMPA), was investigated by monitoring the deuterium nuclear magnetic resonance ( 2H-NMR) spectrum of choline-deuterated phosphatidylcholine as a function of temperature and DMPA concentration. Addition of DMPA at temperatures corresponding to fully liquid-crystalline membranes caused a progressive increase (decrease) in the 2H-NMR quadrupole splitting from POPC- α- d 2 (POPC- β- d 2), in agreement with the known response of phosphatidylcholine to negative membrane surface charge (Seelig, J., Macdonald, P.M. and Scherer, P.G. (1987) Biochemistry 26, 7535–7541). Lateral phase separation of DMPA-rich domains was induced in these mixtures by lowering the temperature in the range from 60°C to −15°C, and was accompanied by a reversal of the original effects of DMPA on the quadrupole splitting. Analysis of the 2H-NMR spectral response allows one to generate a temperature/composition phase diagram for the POPC/DMPA system. We conclude that 2H-NMR of headgroup-deuterated phosphatidylcholine can be employed to sense and to quantify inhomogeneities in the lateral distribution of charged membrane components.

Interaction of hemoglobin derivatives with liposomes. Membrane cholesterol protects against the changes of hemoglobin.

Hemoglobin (Hb) was incubated with small unilamellar vesicles of different lipid compositions in physiological or low pH-low ionic strength media, and the alterations in the protein and bilayer structure were analyzed by measuring (i) the sedimentation properties and elution pattern of the vesicles upon gel filtration, (ii) the quenching effect of Hb on the fluorescence of membrane-embedded probes, (iii) the intrinsic fluorescence of Hb, and (iv) the Soret spectra of Hb. The results indicate complex formation between Hb and the membrane, followed by intercalation of the protein into the bilayer. These processes can lead to secondary alterations, including aggregation, peroxidative decomposition of unsaturated fatty acids, unfolding of Hb, oxidation of the heme iron, displacement of the heme relative to globin, and oxidative or nonoxidative deconjugation of the porphyrin ring. Complex formation and intercalation of Hb into the bilayer are primarily due to hydrophobic interaction between the protein and the membrane, whereas displacement of the heme and its nonoxidative deconjugation are elicited by ionic interaction between the heme and negative membrane surface charges. These charges, on the other hand, inhibit the oxidative processes in unsaturated lipid vesicles. The rate and extent of alterations both in the protein and in the membrane show inverse correlation with the stability of the heme-globin linkage. The liquid-crystalline phase state promotes the penetration of Hb into the bilayer, whereas displacement of the heme is most expressed in the case of saturated bilayers in the gel state. Cholesterol inhibits both the hydrophobic and ionic interactions between the protein and bilayer, most remarkably the partition of the heme into negatively charged saturated bilayers. It is suggested that the effects of cholesterol are mediated through changes in membrane fluidity.

The effect of polyphosphoinositides and phosphatidic acid on the phosphatidylinositol transfer protein from bovine brain: a kinetic study

The phosphatidylinositol transfer protein from bovine brain (PI-TP) has lipid transfer characteristics which make it well suited to maintain phosphatidylinositol (PI) levels in intracellular membranes (Van Paridon, P.A., Gadella, Jr., T.W.J., Somerharju, P.J. and Wirtz, K.W.A. (1987) Biochim. Biophys. Acta 903, 68–77). Using a continuous fluorimetric transfer assay we have investigated in what way phosphatidylinositol 4-phosphate (PIP), phosphatidylinositol 4,5-bisphosphate (PIP 2) and phosphatidic acid (PA) affect the transfer activity of this protein in model systems. The effects were analysed by application of a kinetic model which yielded the association constant ( K) and dissociation rate constant ( k −) for the PI-TP/vesicle complex. Incorporation of PA, PIP and PIP 2 into the phosphatidylcholine-containing vesicles increased the association constant solely by diminishing the dissociation rate constant. This effect could be completely accounted for by changes in the membrane surface charge density. In contrast to the inhibitory effect of PA, the inhibition caused by PIP 2 was completely abolished by the addition of neomycin, in agreement with the observed preferential binding of this polyamine antibiotic to PIP 2. A rise in pH from 5.5 to 8 drastically reduced the association constant for vesicles containing 16 mol% PA (e.g., from 38 to 2 mM −1), without affecting the V max. This effect could be mainly attributed to an increase in the negative charge on PI-TP (isoelectric point 5.5), resulting in an enhanced repulsion. Increasing the negative membrane surface charge at pH 7.4 had the opposite effect. This is interpreted to indicate that the membrane interaction site on PI-TP must be positively charged, overcoming the repulsive forces between PI-TP and the vesicle. Addition of PIP 2 micelles as a third component in the transfer assay strongly inhibited PI-TP transfer activity. The extent of inhibition suggests a very high affinity of PI-TP for this lipid.

Induced hydrogen ion transport in lipid membranes as origin of toxic effect of pentachlorophenol in an alga

Pentachlorophenol (PCP) decreases the rate of carbon assimilation in the alga Selenastrum capricornutum. In parallel with the reduction of carbon assimilation in this alga there is a decrease of electrical resistance of lipid membranes and development of negative membrane surface charge. The experimental results suggest that PCP toxicity to algae is due to adsorption of negatively charged PCP ions at the membrane surface that act as carriers of hydrogen ion across the membrane. This protonophoretic action of PCP causes the decrease of membrane electrical resistance and the dissipation of hydrogen ion electrochemical potential gradients across cellular and subcellular membranes, which reduces the ability of algae to assimilate carbon.