Alpha-selenoconotoxins, a new class of potent alpha7 neuronal nicotinic receptor antagonists

Disulfide bonds are important structural motifs that play an essential role in maintaining the conformational stability of many bioactive peptides. Of particular importance are the conotoxins, which selectively target a wide range of ion channels that are implicated in numerous disease states. Despite the enormous potential of conotoxins as therapeutics, their multiple disulfide bond frameworks are inherently unstable under reducing conditions. Reduction or scrambling by thiol-containing molecules such as glutathione or serum albumin in intracellular or extracellular environments such as blood plasma can decrease their effectiveness as drugs. To address this issue, we describe a new class of selenoconotoxins where cysteine residues are replaced by selenocysteine to form isosteric and nonreducible diselenide bonds. Three isoforms of alpha-conotoxin ImI were synthesized by t-butoxycarbonyl chemistry with systematic replacement of one ([Sec(2,8)]ImI or [Sec(3,12)]ImI), or both ([Sec(2,3,8,12)]ImI) disulfide bonds with a diselenide bond. Each analogue demonstrated remarkable stability to reduction or scrambling under a range of chemical and biological reducing conditions. Three-dimensional structural characterization by NMR and CD spectroscopy indicates conformational preferences that are very similar to those of native ImI, suggesting fully isomorphic structures. Additionally, full bioactivity was retained at the alpha7 nicotinic acetylcholine receptor, with each selenoanalogue exhibiting a dose-response curve that overlaps with wild-type ImI, thus further supporting an isomorphic structure. These results demonstrate that selenoconotoxins can be used as highly stable scaffolds for the design of new drugs.     

Effect of sulfite treatment on total antioxidant capacity, total oxidant status, lipid hydroperoxide, and total free sulfydryl groups contents in normal and sulfite oxidase-deficient rat plasma

Sulfites, which are commonly used as preservatives, are continuously formed in the body during the metabolism of sulfur-containing amino acids. Sulfite oxidase (SOX) is an essential enzyme in the pathway of the oxidative degradation of sulfite to sulfate protecting cells from sulfite toxicity. This article investigated the effect of sulfite on total antioxidant capacity (TAC), total oxidant status, lipid hydroperoxide (LOOH), and total free sulfydryl groups (-SH) levels in normal and SOX-deficient male albino rat plasma. For this purpose, rats were divided into four groups: control, sulfite-treated, SOX-deficient, and sulfite-treated SOX-deficient groups. SOX deficiency was established by feeding rats a low molybdenum diet and adding to their drinking water 200 ppm tungsten. Sulfite (70 mg/kg) was administered to the animals via their drinking water. SOX deficiency together with sulfite treatment caused a significant increase in the plasma LOOH and total oxidant status levels. -SH content of rat plasma significantly decreased by both sulfite treatment and SOX deficiency compared to the control. There was also a significant decrease in plasma TAC level by sulfite treatment. In conclusion, sulfite treatment affects the antioxidant/oxidant balance of the plasma cells of the rats toward oxidants in SOX-deficient groups.     

Investigation of ash deposition in a pilot-scale fluidized bed combustor co-firing biomass with lignite

This study presents the results from investigation of ash deposition characteristics of a high ash and sulfur content lignite co-fired with three types of biomass (olive residue, 49 wt%; hazelnut shell, 42 wt%; and cotton residue, 41 wt%) in 0.3 MW(t) Middle East Technical University (METU) Atmospheric Bubbling Fluidized Bed Combustion (ABFBC) Test Rig. Deposit samples were collected on an air-cooled probe at a temperature of 500 degrees C. Samples were analyzed by SEM/EDX and XRD methods. The results reveal that co-firing lignite with olive residue, hazelnut shell and cotton residue show low deposition rates. High concentrations of silicon, calcium, sulfur, iron, and aluminum were found in deposit samples. No chlorine was detected in deposits. Calcium sulfate and potassium sulfate were detected as major and minor components of the deposits, respectively. High sulfur and alumina-silicate content of lignite resulted in formation of alkali sulfates instead of alkali chlorides. Therefore, fuel blends under consideration can be denoted to have low-fouling propensity.     

The Modified Sequential Treatment Regimen Containing Levofloxacin for Helicobacter pylori Eradication in Turkey

Background:  Eradication rates of Helicobacter pylori have declined to unacceptable levels in recent years. New and effective treatment options are warranted both as a first and second line treatment.
Aim:  To test an effectiveness of modified sequential therapy with levofloxacin for H. pylori eradication in Turkey.
Material and Methods:  Helicobacter pylori infected dyspeptic patients were included to the study. Subjects were treated with modified sequential therapy consisting of rabeprazole 20 mg b.i.d. and amoxicillin 1 g b.i.d., for 7 days followed by rabeprazole 20 mg b.i.d, levofloxacin 500 mg q.d. and metronidazole 500 mg b.i.d for the remaining 7 days.
Results:  Sixty-three treatment naive patients and 37 previous treatment failures were enrolled to the study (59 F, 41 M, age: 21–80 years). There was five drop out. Helicobacter pylori eradication was achieved in 80 patients, intention-to-treat (ITT): 80% (95% CI: 71–87%) and per-protocol (PP): 84.2% (95% CI: 75–91%), totally. In treatment naive patients ITT and PP eradication rates were 82.5% (95% CI: 71–91%), and 86.7% (95% CI: 75–94%), respectively. As a second line treatment eradication was successful in ITT 75.7%.(95% CI: 59–88%), and PP 80% (95% CI: 63–92%).Mild side effects were reported by 8 patients (8.4%).
Conclusions:  Sequential therapy using "rabeprazole and amoxicillin 7 days followed by rabeprazole, metronidazole and levofloxacin for 7 days" is a new regimen with acceptable eradication rates in naïve patients in Turkey. Further modifications in the dose or duration of this new sequential therapy might increase its effectiveness as both first and second line treatment.     

Polar lipid fatty acids,LPS-hydroxy fatty acids,and respiratory quinones of three <Emphasis Type="Italic">Geobacter</Emphasis> strains,and variation with electron acceptor

The polar lipid fatty acids, lipopolysaccharide hydroxy-fatty acids, and respiratory quinones of Geobacter metallireducens str. GS-15, Geobacter sulfurreducens str. PCA, and Geobacter bemidjiensis str. Bem are reported. Also, the lipids of G. metallireducens were compared when grown with Fe³⁺ or nitrate as electron acceptors and G. sulfurreducens with Fe³⁺ or fumarate. In all experiments, the most abundant polar lipid fatty acids were 14:0, i15:0, 16:1ω7c, 16:1ω5c, and 16:0; lipopolysaccharide hydroxy-fatty acids were dominated by 3oh16:0, 3oh14:0, 9oh16:0, and 10oh16:0; and menaquinone-8 was the most abundant respiratory quinone. Some variation in lipid profiles with strain were observed, but not with electron acceptor. D. C. White: Deceased.     

A flexible docking scheme to explore the binding selectivity of PDZ domains

Modeling of protein binding site flexibility in molecular docking is still a challenging problem due to the large conformational space that needs sampling. Here, we propose a flexible receptor docking scheme: A dihedral restrained replica exchange molecular dynamics (REMD), where we incorporate the normal modes obtained by the Elastic Network Model (ENM) as dihedral restraints to speed up the search towards correct binding site conformations. To our knowledge, this is the first approach that uses ENM modes to bias REMD simulations towards binding induced fluctuations in docking studies. In our docking scheme, we first obtain the deformed structures of the unbound protein as initial conformations by moving along the binding fluctuation mode, and perform REMD using the ENM modes as dihedral restraints. Then, we generate an ensemble of multiple receptor conformations (MRCs) by clustering the lowest replica trajectory. Using ROSETTA LIGAND , we dock ligands to the clustered conformations to predict the binding pose and affinity. We apply this method to postsynaptic density‐95/Dlg/ZO‐1 (PDZ) domains; whose dynamics govern their binding specificity. Our approach produces the lowest energy bound complexes with an average ligand root mean square deviation of 0.36 Å. We further test our method on (i) homologs and (ii) mutant structures of PDZ where mutations alter the binding selectivity. In both cases, our approach succeeds to predict the correct pose and the affinity of binding peptides. Overall, with this approach, we generate an ensemble of MRCs that leads to predict the binding poses and specificities of a protein complex accurately.     

Differential protein expression in the metal-reducing bacterium Geobacter sulfurreducens strain PCA grown with fumarate or ferric citrate

Geobacter sulfurreducens, generally considered to be a strict anaerobe, is a predominant microbe in subsurface environments, where it utilizes available metals as electron acceptors. To better understand the metabolic processes involved in the metal-reduction capability of this microbe, the proteins expressed by cells grown anaerobically with either fumarate or ferric citrate as electron acceptor were compared. Proteins were separated by 2-DE under denaturing or nondenaturing conditions, and proteins varying in abundance with a high level of statistical significance (p<0.0001) were identified by peptide mass analysis. Denaturing 2-DE revealed significant differences in the relative abundance of the membrane proteins OmpA and peptidoglycan-associated lipoprotein, several metabolic enzymes, and, in addition, superoxide dismutase and rubredoxin oxidoreductase. Nondenaturing 2-DE revealed elevated catalase in cells grown with ferric citrate. These results suggest that, in addition to adjustments in membrane transport and specific metabolic pathways in response to these two different electron acceptors, distinct differences exist in the oxidative environment within the cell when fumarate or soluble ferric citrate is provided as electron acceptor. Although an anaerobe, G. sulfurreducens appears to have alternate mechanisms for dealing with reactive oxygen species in response to increased intracellular soluble iron.     

Metabolic model for acetate uptake by a mixed culture of phosphate- and glycogen-accumulating organisms under anaerobic conditions

This paper proposes a new metabolic model for acetate uptake by a mixed culture of phosphate- and glycogen-accumulating organisms (PAOs and GAOs) under anaerobic conditions. The model uses variable overall stoichiometry based on the assumption that PAOs may have the ability of using the glyoxylate pathway to produce the required reducing power for polyhydroxyalkonate (PHA) synthesis. The proposed model was tested and verified by experimental results. A sequencing batch reactor system was operated for enhanced biological phosphorus removal (EBPR) with acetate as the sole carbon source at different influent acetate/phosphate ratios. The resulting experimental data supported the validity of the proposed model, indicating the presence of GAOs for all tested HAc/P ratios, especially under P-limiting conditions. Strong agreement is observed between experimental values and model predictions for all model components, namely, PHB production, PHA composition, glycogen utilization, and P release.     

Microorganisms associated with uranium bioremediation in a high-salinity subsurface sediment

Although stimulation of dissimilatory metal reduction to promote the reductive precipitation of uranium has been shown to successfully remove uranium from some aquifer sediments, the organisms in the family Geobacteraceae that have been found to be associated with metal reduction in previous studies are not known to grow at the high salinities found in some uranium-contaminated groundwaters. Studies with a highly saline uranium-contaminated aquifer sediment demonstrated that the addition of acetate could stimulate the removal of U(VI) from the groundwater. This removal was associated with an enrichment in microorganisms most closely related to Pseudomonas and Desulfosporosinus species.