Change In Negative Charge Research Articles

Transforms of Cell Surface Glycoproteins Charge Influences Tumor Cell Metastasis via Atypically Inhibiting Epithelial-Mesenchymal Transition Including Matrix Metalloproteinases and Cell Junctions.

Cell communication and signal transduction rely heavily on the charge on the cell surface. The cell surface is negatively charged, with glycoproteins on the cell membrane providing a large percentage of the charge. Sialic acid is found on the outermost side of glycan chains and contributes to glycoprotein's negative charge. Sialic acid is highly expressed in tumor cells and plays an important role in tumor metastasis and immune escape by interacting with extracellular ligands. However, the specific effect of negative charge changes on glycoproteins is still poorly understood. In this study, we used 9-azido sialic acid (9Az-Sia) to create artificial epitopes on glycoproteins via metabolic glycan labeling, and we attached charged groups such as amino and carboxyl to 9Az-Sia via a click reaction with dibenzocyclooctyne (DBCO). The charge of glycoproteins was changed by metabolic glycan labeling and click modification. The results showed that the migration and invasion ability of the MDA-MB-231 cell labeled with 9Az-Sia was significantly reduced after the modification with amino groups rather than carboxyl groups. Epithelial-mesenchymal transition (EMT) is the biological process of metastatic tumor cells, with an increasing ability of tumor cells to migrate and invade. In particular, the expression of adhesion molecules increased in the amine-linked group, whereas the expression of matrix metalloproteinases (MMPs) increased significantly, which is not identical to EMT characteristics. In vivo experiments have demonstrated that the loss of negative charge on glycoproteins has an inhibitory effect on tumors. In conclusion, modifying the positive charge on the surface of glycoproteins can inhibit tumor cell metastasis and has great potential for tumor therapy.

Role of charged residues of the “electrostatic loop” of hSOD1 in promotion of aggregation: Implications for the mechanism of ALS-associated mutations under amyloidogenic conditions

Protein misfolding and amyloid formation are hallmarks of numerous diseases, including amyotrophic lateral sclerosis (ALS), in which hSOD1 aggregation is involved in pathogenesis. We used two point mutations in the electrostatic loop, G138E and T137R, to analyze charge distribution under destabilizing circumstances to gain more about how ALS-linked mutations affect SOD1 protein stability or net repulsive charge. We show that protein charge is important in the ALS disease process using bioinformatics and experiments. The MD simulation findings demonstrate that the mutant protein differs significantly from WT SOD1, which is consistent with the experimental evidence. The specific activity of the wild type was 1.61 and 1.48 times higher than that of the G138E and T137R mutants, respectively. Under amyloid induction conditions, the intensity of intrinsic and ANS fluorescence in both mutants reduced. Increasing the content of β-sheet structures in mutants can be attributed to aggregation propensity, which was confirmed using CD polarimetry and FTIR spectroscopy. Our findings show that two ALS-related mutations promote the formation of amyloid-like aggregates at near physiological pH under destabilizing conditions, which were detected using spectroscopic probes such as Congo red and ThT fluorescence, and also further confirmation of amyloid-like species by TEM. Overall, our results provide evidence supporting the notion that negative charge changes combined with other destabilizing factors play an important role in increasing protein aggregation by reducing repulsive negative charges.

Changes in Negative Charge at the Luminal Mouth of the Pore Alter Ion Handling and Gating in the Cardiac Ryanodine-Receptor

We have tested the hypothesis that a high density of negative charge at the luminal mouth of the RyR2 pore plays a pivotal role in the high cation conductance and limited selectivity observed in this channel by introducing into each monomer a double point mutation to neutralize acidic residues in this region of the mouse RyR2 channel. The resultant channel, ED4832AA, is capable of functioning as a calcium-release channel in situ. Consistent with our hypothesis, the ED4832AA mutation altered the ion handling characteristics of single RyR2 channels. The mutant channel retains the ability to discriminate between cations and anions but cation conductance is altered significantly. Unitary K + conductance is reduced at low levels of activity but increases dramatically as activity is raised and shows little sign of saturation. ED4832AA no longer discriminates between divalent and monovalent cations. In addition, the gating characteristics of single RyR2 channels are altered markedly by residue neutralization. Open probability in the ED4832AA channel is substantially higher than that of the wild-type channel. Moreover, at holding potentials in excess of ±50 mV several subconductance states become apparent in ED4832AA and are more prevalent at very high holding potentials. These observations are discussed within the structural framework provided by a previously developed model of the RyR2 pore. Our data indicates that neutralization of acidic residues in the luminal mouth of the pore produces wide-ranging changes in the electric field in the pore, the interaction energies of permeant ions in the pore and the stability of the selectivity filter region of the pore, which together contribute to the observed changes ion handling and gating.

High-pressure optical studies of Y2O3:Eu3+ nanoparticles

The fluorescence spectra of Y2O3:Eu3+ nanoparticles have been measured under the pressure of up to 78 kbar at room temperature. In this pressure range, a red-shift of 0.02(1) nm/kbar−1 is noticed for the 0–2 line (5D0→7F2 transition). This shift is explained by the change of negative charge of the surrounding ligands. Compatibility between measured and calculated values for the 0–2 line position was obtained. The luminescence decay curves of the 5D0→7F2 transition were studied up to 78 kbar and were found to behave exponentially for all pressures studied. The fluorescence lifetime τ for the 0–2 line (5D0→7F2 transition) slowly decreased with pressure. The pressure effect on τ for the 0–2 line (5D0→7F2 transition) was explained by a model which considers the pressure effect on the line position, inter-ionic distance, ion volume and polarizability, molecular volume and polarizability, molecular refractive index and the refractive index medium n med of the surrounding hydrostatic medium. The fluorescence lifetime calculated by the present model is in close correspondence with the experimental values.

Ab Initio Fragment Molecular Orbital Study of Molecular Interactions between Liganded Retinoid X Receptor and Its Coactivator: Roles of Helix 12 in the Coactivator Binding Mechanism

On the basis of the fragment molecular orbital method we addressed molecular interactions of liganded retinoid X receptor (RXR) with steroid receptor co-activating factor-1 (SRC1) coactivator to examine the contribution of helix 12 (H12), which contains the core of the transcriptional activation function 2 activating domain, to the coactivator binding of RXR. The interaction between H12 and SRC1 was proved to be the main cause for the stabilization of the coactivator binding. In particular, highly conserved charged (Glu453) and hydrophobic (Phe450) residues in H12 were found to have stronger electrostatic and dispersion interactions with SRC1 than the other charged and hydrophobic residues in H12, respectively. In addition, the charge transfer (CT) from RXR to SRC1 was found to occur mainly by the changes in charges of H12 residues. Large positive and negative charge changes were observed especially for Glu453 and for Lys631 and Ile632 in SRC1, respectively, indicating that Glu453 is an electron donor for Lys631 and Ile632 in this CT. Taken together, our findings quantitatively demonstrated that H12 and its highly conserved residues significantly contribute to the coactivator binding not only by the Coulomb and dispersion interactions but also by the CT described with the quantum-mechanical framework.

Analysis of Oligosaccharide Negative Charge by Anion‐Exchange Chromatography

This unit presents the analysis of negative charge on labeled N- or O-linked oligosaccharides. These protocols may be used in the initial screening of oligosaccharides to detect negative charge, for analytical or preparative separation of oligosaccharides based on their negative charge, or to analyze the type of negative charge found on the oligosaccharides. The basic Protocol describes the use of anion-exchange (QAE-Sephadex) chromatography with stepwise elution for estimating the number of negative charges on an oligosaccharide sample derived from glycosidase treatment of a glycoprotein. In an Alternate Protocol, gradient elution is used for the preparative separation of oligosaccharides based on negative charge. A Support Protocol describes a method for measuring loss of or change in negative charge after treatment of the oligosaccharide sample with mild acid and/or phosphatases.

Relationship between Electron Transport across the Plasmalemma and a pH Decrease in the Bulk Medium

The hypothesis is tested that acidification of the bulk medium during transplasmalemma electron transport to ferricyanide is due solely to a requirement for charge balance. According to this hypothesis, reduction of the trivalent anion, ferricyanide, to the tetravalent anion, ferrocyanide, results in a charge difference that is balanced by protons. A coulometric device is used that rapidly and efficiently reoxidizes ferrocyanide to ferricyanide, thus maintaining a constant charge in the bulk medium. Oat (Avena sativa L. cv Garry) mesophyll protoplasts are chosen as experimental material to facilitate ferricyanide reduction and the concomitant ferrocyanide reoxidation by the coulometric device. The kinetics of ferricyanide reduction and proton excretion by protoplasts are similar to those of other cell types and tissues. Rates of net proton excretion are identical regardless of whether the ferrocyanide is simultaneously reoxidized. We conclude that acidification may occur during transplasmalemma electron transport when there is no change in negative charge of the bulk medium.

Factors influencing interaction of phosphorylated and dephosphorylated myosin with actin

The influence of various factors on the interaction of phosphorylated and dephosphorylated myosin with action was examined. It was found that the difference between the values of specific activity of the two myosin forms of actin-stimulated Mg 2+-ATPase is affected by changes in KCl, MgATP and acting concentration. The effect of increased pH on the differences in the rate of ATP hydrolysis by actomyosin containing phosphorylated myosin as compared with that of the dephosphorylated one, observed in the presence of EGTA, is abolished by addition of Ca 2+. Tropomyosin strongly inhibits the actin-stimulated Mg 2+-ATPase of phosphorylated myosin (by about 60%). The tropomyosin-troponin complex and native tropomyosin lowered the rate of ATP hydrolysis by actomyosin containing both phosphorylated and dephosphorylated myosin by about of 60% of the value obtained in the absence of those proteins. These results indicate that the change of negative charge on the myosin head due to phosphorylation and dephosphorylation of myosin light chains modulates the actin-myosin interaction at different steps of the ATP hydrolysis cycle. Phosphorylation of myosin seems to be a factor decreasing the rate of ATP hydrolysis by actomyosin under physiological conditions.

Gallium induces reduction of negative charge of the cell membrane and redifferentiation of cancer cells.

In order to obtain an insight into the mechanism of experimentally induced redifferentiation of cancer cells, changes in negative charge of the cell membrane were examined by cytopherometry following in vitro exposure of the cancer cells to non-radioactive gallium. Reduction of the negative charge was observed at 24 hr and later. The gallium ions must have neutralized the negative charge of the cancer cell membrane to the normal range. This event may contribute in some way to cancer cell redifferentiation.

Ionic strength of soil solution and its effect on charge properties of some New Zealand soils

SummarySelected horizons from six New Zealand soils under permanent natural vegetation, four of which form a development sequence, were chosen to provide variations in organic matter, phyllosilicate clay mineral and short‐range ordered aluminosilicate (allophane) contents. The ionic strength of the soil solution (μ) extracted from the soils of the development sequence was low, being always less than 0.005. For all horizons of the six soils, the negative charge changes linearly with μ1/2 within the ionic strength range of 0.001 to 1. The change was negligible for the least weathered and leached soil, indicating that it contained mainly permanently charged colloids. As soil development increased and/or the components carrying variable charge increased in amount, the change in charge with ionic strength increased. A regression equation showed a strong relationship between change in negative charge with ionic strength and organic carbon and oxalate‐extractable A1 (R2=0.976). This equation was tested by predicting observed changes in charge with ionic strength for a second selection of soils. The change in positive charge with ionic strength was correlated with oxalate‐extractable A1 (r2=0.914). The results are discussed in relation to measurement of soil charge and iron movement in soils.