Identification of the tyrosine nitration sites in human endothelial nitric oxide synthase by liquid chromatography-mass spectrometry

The formation of nitric oxide (NO) in biological systems has led to the discovery of a number of post- translational protein modifications that can affect biological conditions such as vasodilation. Studies both from our laboratory and others have shown that beside its effect on cGMP generation from soluble guanylate cylcase, NO can produce protein modifications through both S-nitrosylation of cysteine residues. Previously, we have identified the potential S-nitrosylation sites on endothelial NO synthase (eNOS). Thus, the goal of this study was to further increase our understanding of reactive nitrogen protein modifications of eNOS by identifing tyrosine residues within eNOS that are susceptible to nitration in vitro. To accomplish this, nitration was carried out using tetranitromethane followed by tryptic digest of the protein. The resulting tryptic peptides were analyzed by liquid chromatography/mass spectrometry (LC/MS) and the position of nitrated tyrosines in eNOS were identified. The eNOS sequence contains 30 tyrosine residues and our data indicate that multiple tyrosine residues are capable of being nitrated. We could identify 25 of the 30 residues in our tryptic digests and 19 of these were susceptible to nitration. Interstingly, our data identified four tyrosine residues that can be modified by nitration that are located in the region of eNOS responsible for the binding to heat shock protein 90 (Hsp90), which is responsible for ensuring efficient coupling of eNOS.     

Polydimethylsiloxane-LiNbO3 surface acoustic wave micropump devices for fluid control into microchannels

This paper presents prototypical microfluidic devices made by hybrid microchannels based on piezoelectric LiNbO(3) and polydimethylsiloxane. This system enables withdrawing micropumping by acoustic radiation in microchannels. The withdrawing configuration, integrated on chip, is here quantitatively investigated for the first time, and found to be related to the formation and coalescence dynamics of droplets within the microchannel, primed by surface acoustic waves. The growth dynamics of droplets is governed by the water diffusion on LiNbO(3), determining the advancement of the fluid front. Observed velocities are up to 2.6 mm s(-1) for 30 dBm signals applied to the interdigital transducer, corresponding to tens of nl s(-1), and the micropumping dynamics is described by a model taking into account an acoustic power exponentially decaying upon travelling along the microchannel. This straighforward and flexible micropumping approach is particularly promising for the withdrawing of liquids in lab-on-chip devices performing cycling transport of fluids and biochemical reactions.     

Ligand-mediated decarbonylation as an efficient synthetic method to Re(I) and Re(II) dicarbonyl complexes

A novel synthetic method based on a ligand-mediated decarbonylation reaction of complexes of the common fac-[Re(CO)3]+ core efficiently yields Re(I) and Re(II) dicarbonyl species.     

Conus ventricosus venom peptides profiling by HPLC-MS: a new insight in the intraspecific variation

Conus is a genus of predatory marine gastropods that poison the prey with a complex mixture of compounds active on muscle and nerve cells. An individual cone snail's venom contains a mixture of pharmacological agents, mostly short, structurally constrained peptides. This study is focused on the composition of the venom employed by Conus ventricosus Gmelin, 1791, a worm-hunting cone snail living in the Mediterranean Sea. For this purpose, LC coupled to MS techniques has been successfully used to establish qualitative and quantitative differences in conopeptides from minute amounts of venom ducts. We were able to prove variability in the venom conopeptide complement, possibly related to different trophic habits of the species in the Mediterranean Sea. Moreover, the information-rich MS techniques enabled us to identify two novel C. ventricosus peptides, here named Conotoxin-Vn and -Conotoxin-Vn. On the basis of the structural data collected so far, we suggest that Conotoxin-Vn is a conopeptide belonging to the -family that recognizes calcium channels through a specific pharmacophore. Similarly, molecular modeling data suggest that -Conotoxin-Vn should represent a competitive antagonist of neuronal nicotinic acetylcholine receptors (nAChRs).     

Pressure-induced core packing and interfacial dehydration in nonionic C12E6 micelle in aqueous solution

A spherical micelle of C12E6 is simulated at different pressures, from 0.001 to 3 kbar, by molecular dynamics. On increasing the pressure the alkyl tails of the surfactants pack tightly and stretch. At 3 kbar we observe dynamical slowing down of the oil core of the micelle. At that pressure the core is characterized by a high oil density, rho oil approximately 0.85 g/cm(3), regular density oscillation, and low chain entropy. Pressure affects the interfacial region as well. Dehydration, induced by the collapse of the hydrophilic head groups, is observed in the inner part of the interface. Such dehydration resembles temperature dehydration but differs in details. Our results support the interpretation of recent experiments on micellar solutions at high pressure.     

Synthesis and characterization of epoxy-modified cement slurries--kinetic data at hardened slurries/HCl interfaces

In this study, two new epoxy-modified oilwell cement slurries were synthesized. All characterization measurements were performed and compared in relation to a standard cement slurry, which was synthesized using only cement and water. The XRD patterns, as well as TG/DTG and DSC analyses have suggested that portlandite (Ca(OH)2) and carbonate phases were not present in detectable amounts in the new cement slurries. The isothermal kinetic data of the interaction of HCl with the epoxy-modified cement were obtained using the batch mode from 25 to 55 degrees C. The kinetic data were best fitted to the second-order and the Avrami kinetic models, since differences between experimental and calculated values were lower for these models. The intraparticle diffusion model has pointed out that the new slurries present positive values for the parameter "C," which is related to higher boundary layer resistance to HCl diffusion into the internal parts of the slurries, in relation to the standard slurry. The characterization and the HCl interaction kinetic studies have suggested that the new cement slurries present good potential, from the chemical viewpoint, to be used in oilwell operations.     

Electrokinetics in nanochannels: part II. Mobility dependence on ion density and ionic current measurements

In the first of this two-paper series, a new model was developed for calculating the electric potential field in a long, thin nanochannel with overlapped electric double layers. The model takes into account the dependence of ion mobility on local ion densities and pH. This model is used here to study and demonstrate the effect of ion density and pH on ionic current measurements. A comparison is shown of predictions based on each of three boundary conditions, as studied in Part I. The model developed in Part I is validated by comparing simulations with measurements of ionic current as a function of sodium borate concentration. Results show that predictions based on extended Debye-Hückel theory for ion mobility significantly improve the accuracy of simulations, but that these do not predict exact scaling behavior. A simple bulk conductivity measurement used as input parameter for the simulations, in place of the predicted bulk conductivity (K(0)), guarantees agreement with data in the thin EDL region. Results also indicate that the charge regulation boundary condition, complemented with an adequate bulk electrolyte model, provides better agreement with experimental trends than the specified zeta potential or specified surface net charge boundary conditions. Further, it is shown that currents due to advection (by electroosmotic flow) are in all cases studied less than 25% of the total current in the system.     

Electrokinetics in nanochannels: part I. Electric double layer overlap and channel-to-well equilibrium

In this paper a new model is described for calculating the electric potential field in a long, thin nanochannel with overlapped electric double layers. Electrolyte concentration in the nanochannel is predicted self-consistently via equilibrium between ionic solution in the wells and within the nanochannel. Differently than published models that require detailed iterative numerical solutions of coupled differential equations, the framework presented here is self-consistent and predictions are obtained solving a simple one-dimensional integral. The derivation clearly shows that the electric potential field depends on three new parameters: the ratio of ion density in the channel to ion density in the wells; the ratio of free-charge density to bulk ion density within the channel; and a modified Debye-Hückel thickness, which is the relevant scale for shielding of surface net charge. For completeness, three wall-surface boundary conditions are analyzed: specified zeta-potential; specified surface net charge density; and charge regulation. Predictions of experimentally observable quantities based on the model proposed here, such as depth-averaged electroosmotic flow and net ionic current, are significantly different than results from previous overlapped electric double layer models. In this first paper of a series of two, predictions are presented where channel depth is varied at constant well concentration. Results show that under conditions of electric double layer overlap, electroosmosis contributes only a small fraction of the net ionic current, and that most of the measurable current is due to ionic conduction in conditions of increased counterion density in the nanochannel. In the second of this two-paper series, predictions are presented where well-concentration is varied and the channel depth is held constant, and the model described here is employed to study the dependence of ion mobility on ionic strength, and compare predictions to measurements of ionic current as a function of channel depth and ion density.     

Direct palladium-catalyzed C-3 arylation of free (NH)-indoles with aryl bromides under ligandless conditions

A new method for the efficient, practical, and highly regioselective direct palladium-catalyzed C-3 arylation of free (NH)-indole and its electron-rich 1-unsubstituted derivatives under ligandless conditions is described. The reactions, which are run outside a glovebox without purification of solvent and reagents, involve treatment of free (NH)-indoles with activated, unactivated, and deactivated aryl bromides in refluxing toluene in the presence of K2CO3 as the base and a catalyst system consisting of a combination of Pd(OAc)2 and benzyl(tributyl)ammonium chloride. The experimental results are consistent with a catalytic cycle based on an electrophilic palladation pathway at the 3-position of 1-indolyl potassium salts.