Stream Of Nitrogen Gas Research Articles

The Structure of a Full-length Response Regulator from Mycobacterium tuberculosis in a Stabilized Three-dimensional Domain-swapped, Activated State

The full-length, two-domain response regulator RegX3 from Mycobacterium tuberculosis is a dimer stabilized by three-dimensional domain swapping. Dimerization is known to occur in the OmpR/PhoB subfamily of response regulators upon activation but has previously only been structurally characterized for isolated receiver domains. The RegX3 dimer has a bipartite intermolecular interface, which buries 2357 A(2) per monomer. The two parts of the interface are between the two receiver domains (dimerization interface) and between a composite receiver domain and the effector domain of the second molecule (interdomain interface). The structure provides support for the importance of threonine and tyrosine residues in the signal transduction mechanism. These residues occur in an active-like conformation stabilized by lanthanum ions. In solution, RegX3 exists as both a monomer and a dimer in a concentration-dependent equilibrium. The dimer in solution differs from the active form observed in the crystal, resembling instead the model of the inactive full-length response regulator PhoB.

Ternary compound B4CN4 prepared by direct nitridation of B4C

Abstract Borocarbide (B4C) can be directly nitrided to a stoichiometric ternary compound B4CN4 by heating in a stream of nitrogen gas at temperatures in a range of 1600–1900 °C. The reaction was diffusion-controlled with an activation energy of 242 kJ/mol. The resulting B4CN4 powder had the turbostratic layered boron nitride (t-BN) structure, which was changed into a three-dimensionally ordered layered structure (h-BN) with the lattice parameters of a = 2.506(1) A and c = 6.710(9) A on heating at 2250 °C in a closed nitrogen atmosphere. The a parameter is very close to that of h-BN, but the c parameter is larger than that of h-BN, rather close to that of graphite. The compound was reversibly decomposed to B4C + 2N2 on heating in Ar atmosphere at 2100 °C.

Retinoschisin (RS1), the Protein Encoded by the X-linked Retinoschisis Gene, Is Anchored to the Surface of Retinal Photoreceptor and Bipolar Cells through Its Interactions with a Na/K ATPase-SARM1 Complex

Retinoschisin or RS1 is a discoidin domain-containing protein encoded by the gene responsible for X-linked retinoschisis (XLRS), an early onset macular degeneration characterized by a splitting of the retina. Retinoschisin, expressed and secreted from photoreceptors and bipolar cells as a homo-octameric complex, associates with the surface of these cells where it serves to maintain the cellular organization of the retina and the photoreceptor-bipolar synaptic structure. To gain insight into the role of retinoschisin in retinal cell adhesion and the pathogenesis of XLRS, we have investigated membrane components in retinal extracts that interact with retinoschisin. Unlike the discoidin domain-containing blood coagulation proteins Factor V and Factor VIII, retinoschisin did not bind to phospholipids or retinal lipids reconstituted into unilamellar vesicles or immobilized on microtiter plates. Instead, co-immunoprecipitation studies together with mass spectrometric-based proteomics and Western blotting showed that retinoschisin is associated with a complex consisting of Na/K ATPase (alpha3, beta2 isoforms) and the sterile alpha and TIR motif-containing protein SARM1. Double labeling studies for immunofluorescence microscopy confirmed the co-localization of retinoschisin with Na/K ATPase and SARM1 in photoreceptors and bipolar cells of retina tissue. We conclude that retinoschisin binds to Na/K ATPase on photoreceptor and bipolar cells. This interaction may be part of a novel SARM1-mediated cell signaling pathway required for the maintenance of retinal cell organization and photoreceptor-bipolar synaptic structure.

Chemical composition of the essential oils of flowers of Rosa banksiae from China

Rosa banksiae is a climber, growing to about 20 feet tall, and is very long lived. The flowers of Rosa banksiae have a lovely fragrance, similar to violets. It has no serious disease or insect problems. Rosa banksiae is an important and widespread woody plant in southwest China. It is reported that the flowers and leaves of this plant show free radical scavenging activity [1, 2]. Doctors of traditional Chinese medicine believe that the radical and leaves of this plant can stop pain and bleeding. However, it is noteworthy that the flowers of Rosa banksiae Ait.f. have a distinctive aroma. So far, there is no published scientific information in the literature on the chemical constituents of the volatiles of flowers of Rosa banksiae Ait.f. The objective of the present study is to provide qualitative and quantitative information on the volatiles present in flowers of Rosa banksiae Ait.f. from China. The flowers of Rosa banksiae Ait.f. were collected during flowering (April, 2006) in Enshi, Hubei Province, China, as identified by Prof. Kaigong Lu (School of Biological Science & Technology, Hubei Institute for Nationalities), and placed in the shade to dry. Two hundred grams of dried flowers was placed in a three-neck flask. Distilled water (1500 mL) was added and the contents distilled for 5h in an oil bath. The distillate was extracted with anhydrous ether (AR grade). The extracted solution was combined, dried over anhydrous sodium sulfate (AR grade) overnight, and concentrated by passing a slow stream of oxygen-free nitrogen gas (99.99%) to it without ether. A bright reddish-yellow essential oil (0.2612 g) was obtained and stored at 4°C until analysis. The yield was 0.13%. In total, 46 compounds were identified in flowers of Rosa banksiae Ait.f. from China (Table 1). These identified components constitute 94.15% of all identified peak areas using GC-FID with a DB-5 column. Of the total number of components identified, there are one acetal, 12 fatty hydrocarbons, three aromatic aldehydes, three carboxylic acids, five alcohols, two arenes, 17 terpenoids, and three phenols. As the main compounds (concentration higher than 1.0%), the following were found: octane (4.73%), phenylethyl alcohol (5.78%), 2-bornanone (26.34%), cis-verbenol (2.68%), borneol (3.78%), dodecane (41.01%), 6-methyldodecane (1.00%), elemicin (3.01%), and α-cadinol (1.06%). Among all the detected compounds in the flowers, we focus on the 17 terpenoids, which are responsible for the flowery odor.

Three-dimensional micro-XRF under cryogenic conditions: a pilot experiment for spatially resolved trace analysis in biological specimens

Three-dimensional micro-XRF is a recently developed microprobe which facilitates three-dimensional resolved chemical analyses with a resolution of around 20 mum. Arbitrary sites or sections of samples can be investigated without the need to section specimens physically. In this paper we demonstrate the use of the microprobe in combination with a cold nitrogen gas stream for the cryogenic fixation of specimens. A 3D micro-XRF setup at the new microfocus beamline at BESSY II was equipped with a nitrogen cryogenic stream. The distribution of Ca, Fe, Zn and Cu across virtual cross sections of a water-rich sample, the root of common duckweed, could be investigated without further sample preparation. This paper demonstrates the capabilities of 3D micro-XRF under cryogenic conditions for investigations of biological specimens.

Formation and Healing of Micrometer-Sized Channel Networks on Highly Mobile Au(111) Surfaces

Practical protocols are presented to reproducibly prepare micrometer-sized Au(111) substrates. Au(111) terraces of micrometer dimensions and atomic smoothness were crystallized by flame-annealing vacuum-deposited gold films on glass and on millimetric amorphous gold shots. Gold films and shots that were slowly cooled in a moderately applied stream of nitrogen gas exhibited large and stable crystal surfaces with Au(111) morphologies. Similarly, flame-annealed gold samples cooled with another protocol--in much rougher streams of nitrogen gas--produced morphologically unstable and highly mobile Au(111) layers. Within the first hour after preparation, however, rapid microscale restructuring in the layers produced complex morphologies of hexagonal channel networks and islands that were predominantly triangular. These channeled gold layers fused slowly in the following hours, with velocities of 0.01-0.2 A/s, as quantified by digital image correlation (DIC). Atomically smooth, stable, and predominantly triangular Au(111) terraces on the scale of micrometers were observed approximately 24 h after the sample preparations.

Suppression of endothelial or lipoprotein lipase in THP-1 macrophages attenuates proinflammatory cytokine secretion

LPL and endothelial lipase (EL) are associated with macrophages in human atherosclerotic lesions, and overexpression of LPL in mouse macrophages is associated with a greater extent of atherosclerosis. To investigate potential mechanisms by which macrophage-derived lipase expression may mediate proatherogenic effects, we used lentivirus-mediated RNA interference to suppress the expression of either LPL or EL within THP-1 macrophages. After suppression of either LPL or EL, significant decreases in the concentration of interleukin-1beta, interleukin-6, monocyte chemoattractant protein-1, and tumor necrosis factor-alpha were observed. Incubation of THP-1 macrophages with either mildly or extensively oxidized LDL consistently decreased cytokine expression, which was additive to that contributed by lipase suppression. Decreased lipase expression was also associated with an altered lipid composition, with reduced percentages of cholesterol (unesterified and esterified), triglycerides, and lysophosphatidylcholine. Microarray data indicated a decreased expression of proinflammatory genes, growth factors, and antiapoptotic genes. By contrast, there was an increased expression of lipoprotein receptors (scavenger receptor 1, low density lipoprotein receptor, scavenger receptor class B type I, and CD36). Thus, we conclude that the suppression of either LPL or EL decreases proinflammatory cytokine expression and influences the lipid composition of THP-1 macrophages. These results provide further insight into the specific metabolic and potential pathological roles of LPL and EL in human macrophages.

The removal of dichloromethane from atmospheric pressure nitrogen gas streams using plasma-assisted catalysis

A non-thermal, atmospheric pressure plasma utilising a dielectric packed bed was used to study the destruction of dichloromethane, CH2Cl2, (DCM) in gas streams of nitrogen with the presence of catalysts. The effect of plasma-assisted catalysis was investigated in two configurations; one where the catalyst was incorporated into the packed bed itself and the other where the catalyst was downstream of the plasma. The combination of plasma and a catalyst allowed improved destruction of DCM. γ-Al2O3 in a one-stage reactor configuration was the most successful in terms of the total destruction of DCM as well as providing the best result in the two-stage reactor configuration compared with the eight catalysts chosen which included alumina, TiO2 and various zeolites. Products as detected by FTIR were CO, CO2, HCN, HCl and H2O with the by-product of N2O. The nature of the catalyst plays a vital role with respect to the effectiveness of DCM destruction and the selectivity of the end-products.

Evidence for Specificity in Lipid-Rhodopsin Interactions

The interaction of bovine rhodopsin with poly- and monounsaturated lipids was studied by (1)H MAS NMR with magnetization transfer from rhodopsin to lipid. Experiments were conducted on bovine rod outer segment (ROS) disks and on recombinant membranes containing lipids with polyunsaturated, docosahexaenoyl (DHA) chains. Poly- and monounsaturated lipids interact specifically with different sites on the rhodopsin surface. Rates of magnetization transfer from protein to DHA are lipid headgroup-dependent and increased in the sequence PC < PS < PE. Boundary lipids are in fast exchange with the lipid matrix on a time scale of milliseconds or shorter. All rhodopsin photointermediates transferred magnetization preferentially to DHA-containing lipids, but highest rates were observed for Meta-III rhodopsin. The experiments show clearly that the surface of rhodopsin has sites for specific interaction with lipids. Current theories of lipid-protein interaction do not account for such surface heterogeneity.

Crystal structure of the catalytic domain of human DUSP5, a dual specificity MAP kinase protein phosphatase

Crystal structure of the catalytic domain of human DUSP5, a dual specificity MAP kinase protein phosphatase

Positional specificity of lysosomal phospholipase A2

Lysosomal phospholipase A(2) (Lpla2) is highly expressed in alveolar macrophages and may mediate the phospholipid metabolism of surfactant. Studies on the properties of this phospholipase are consistent with the presence of both phospholipase A(1) and phospholipase A(2) activities. These activities were studied through the production of O-acyl compounds, produced by the transacylase activity of Lpla2. Liposomes containing POPC and N-acetylsphingosine (NAS) were incubated with the soluble fraction obtained from MDCK cells stably transfected with the mouse Lpla2 gene. Two 1-O-acyl-NASs, 1-O-palmitoyl-NAS and 1-O-oleoyl-NAS, were produced by Lpla2. The formation rate of 1-O-oleoyl-NAS was 2.5-fold that of 1-O-palmitoyl-NAS. When 1-oleoyl-2-palmitoyl-sn-glycero-3-phosphocholine (OPPC) was used, the formation rate of 1-O-oleoyl-NAS was 5-fold higher than that of 1-O-palmitoyl-NAS. Thus, Lpla2 can act on acyl groups at both sn-1 and sn-2 positions of POPC and OPPC. When 1-palmitoyl-2-unsaturated acyl-sn-glycero-3-phosphocholines were used as acyl donors, the transacylation of the acyl group from the sn-2 position to NAS was preferred to that of the palmitoyl group from the sn-1 position. An exception was observed for 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (PAPC), for which the formation rate of 1-O-palmitoyl-NAS from PAPC was 4-fold greater than that of 1-O-arachidonoyl-NAS. Thus, Lpla2 has broad positional specificity for the sn-1 and sn-2 acyl groups in phosphatidylcholine and phosphatidylethanolamine.

Contribution of Sensory Neurons to Sex Difference in the Development of Stress-Induced Gastric Mucosal Injury in Mice

Sensory neurons play a critical role in reducing stress-induced gastric mucosal injury by releasing calcitonin gene-related peptide (CGRP) through an increase in gastric mucosal levels of prostacyclin (PGI(2)). Because estrogen enhances nerve growth factor-mediated CGRP production in sensory neurons, we hypothesized that stress-induced gastric mucosal injury occurs less in females than in males. Gastric ulcer index, gastric myeloperoxidase activity, and gastric tissue levels of CGRP and 6-keto-PGF(1alpha), a stable metabolite of PGI(2), were determined in male and female wild-type (CGRP(+/+)) mice and CGRP knockout (CGRP(-/-)) mice subjected to water-immersion restraint stress. In CGRP(+/+) mice, ulcer index and myeloperoxidase activities were lower and gastric tissue levels of CGRP and 6-keto-PGF(1alpha) were higher in female mice than in male mice, but there were no such sex differences in CGRP(-/-)mice. Sex differences in CGRP(+/+) mice were eliminated by pretreatment with SB366791 (500 microg/kg intraperitoneally), a vanilloid receptor antagonist, and by ovariectomy. Reversal of sex differences by ovariectomy was not observed in female CGRP(+/+) mice with estradiol replacement (1 mg . kg(-1). wk(-1) for 3 weeks). Levels of CGRP messenger RNA in dorsal root ganglion neurons isolated from female CGRP(+/+) mice were decreased by ovariectomy, and these decreases were reversed by estradiol replacement. Estrogen-mediated increases in CGRP levels in sensory neurons might contribute to reduce stress-induced gastric mucosal injury by attenuating inflammatory responses. This might at least partly explain the sex difference observed in the development of stress-induced gastric mucosal injury in mice.

Direct VUV photolysis of chlorinated methanes and their mixtures in a nitrogen stream

The gas-phase decomposition of CCl 4, CHCl 3 and CH 2Cl 2 and their binary mixtures was studied in a flow-type reactor in a nitrogen gas stream, using a low-pressure mercury vapour lamp covered with a high-purity silica quartz sleeve. The 184.9 nm vacuum-ultraviolet (VUV) light emitted is able to rupture the C–Cl bond in these target substances. For H-containing compounds, the decomposition takes place not only by direct photolysis, but also by H abstraction by Cl formed during the direct photolysis of the target substances. The relative contributions of direct photolysis and Cl-sensitized reactions to the decomposition were estimated at different initial concentrations. The addition of CCl 4 to CHCl 3 or CH 2Cl 2 increased their decomposition rates via increase of the Cl concentration, whereas the addition of CH 2Cl 2 to CHCl 3 decreased its degradation rate, suggesting that CH 2Cl 2 acts as a Cl radical scavenger. The variation of the product distribution confirms the effect of the composition of the irradiated gas mixtures on the relative contributions of Cl-sensitized reactions and direct photolysis.

Structurally Altered Peptides Reveal an Important Role for N-terminal Heptad Repeat Binding and Stability in the Inhibitory Action of HIV-1 Peptide DP178

Human immunodeficiency virus 1 gp41 folds into a six-helix bundle whereby three C-terminal heptad repeat regions pack in an anti-parallel manner against the coiled-coil formed by three N-terminal heptad repeats (NHR). Peptides that inhibit bundle formation contributed significantly to the understanding of the entry mechanism of the virus. DP178, which partially overlaps C-terminal heptad repeats, prevents bundle formation through an undefined mechanism; additionally it has been suggested to bind other ENV regions and arrest fusion in an unknown manner. We used two structurally altered DP178 peptides; in each, two sequential amino acids were substituted into their d configuration, d-SQ in the hydrophilic N-terminal region and d-LW in the hydrophobic C-terminal. Importantly, we generated an elongated NHR peptide, N54, obtaining the full N-helix docking site for DP178. Interestingly, d-LW retained wild type fusion inhibitory activity, whereas d-SQ exhibited significantly reduced activity. In correlation with the inhibitory data, CD spectroscopy and fluorescence studies revealed that all the DP178 peptides interact with N54, albeit with different stabilities of the bundles. We conclude that strong binding of DP178 N-terminal region to the endogenous NHR, without significant contribution of the C-terminal sequence of DP178 to core formation, is vital for DP178 inhibition. The finding that d-amino acid incorporation in the C terminus did not affect activity or membrane binding as revealed by surface plasmon resonance correlates with an additional membrane binding site, or membrane anchoring role, for the C terminus, which works synergistically with the N terminus to inhibit fusion.

Ss-LrpB from Sulfolobus solfataricus Condenses about 100 Base Pairs of Its Own Operator DNA into Globular Nucleoprotein Complexes

Ss-LrpB from the hyperthermoacidophilic crenarchaeote Sulfolobus solfataricus P2 is a member of the Lrp-like family of Bacterial/Archaeal transcription regulators that binds its own control region at three regularly spaced and partially conserved 15-bp-long imperfect palindromes. We have used atomic force microscopy to analyze the architecture of Ss-LrpB.DNA complexes with a different stoichiometry formed with the wild type operator and with an operator mutant. Binding of dimeric Ss-LrpB to all three target sites is accompanied by the formation of globular complexes, in which the protein induces strong DNA deformations. Furthermore, DNA contour length foreshortening of these complexes indicates DNA wrapping, with about 100 bp being condensed. The average bending angle is 260 degrees . The establishment of protein-protein contacts between Ss-LrpB dimers in these globular complexes will contribute to the cooperativity of the binding. The profound remodeling of the control region is expected to have a strong impact on gene expression and might constitute the key element in the autoregulatory process.

An analysis of the shielding gas flow from a coaxial conical nozzle during high power CO2 laser welding

An experimental and theoretical study on the role of the nitrogen gas stream, exiting from a conventional conical nozzle tip during a laser welding process, has been carried out. A mathematical model has been used, based on the Navier–Stokes equations which express fundamental conservation laws of mass, momentum and energy for a compressible fluid. Numerical simulations of the gas stream colliding onto a plane surface have been performed showing the effects of variations of inlet gas pressure, nozzle exit diameter and standoff distance on the density and Mach number contours, axis pressure of the gas jet and plate pressure produced on the workpiece surface. Laser welding experiments have been performed on carbon and stainless steel specimens, by varying the process parameters in the same range as in the simulations and keeping constant the incident power and the travel speed. Two different gas stream regimes were found, namely sonic and subsonic, which were experimentally verified to produce cutting and welding conditions, respectively.Weld performances have been evaluated in terms of bead width, penetration depth and melted area. Nozzle standoff distance was found to have a negligible influence, while the exit diameter and the flow rate significantly affect the weld results. The numerical predictions allowed an explanation of the experimental results yielding useful suggestions for enhancing the weld quality, acting simply on the shielding gas parameters.

Nontoxic crystal protein from Bacillus thuringiensis demonstrates a remarkable structural similarity to β‐pore‐forming toxins

Nontoxic crystal protein from <i>Bacillus thuringiensis</i> demonstrates a remarkable structural similarity to β‐pore‐forming toxins

Regulation of bile acid biosynthesis by hepatocyte nuclear factor 4α

Hepatocyte nuclear factor 4alpha (HNF4alpha) regulates many genes that are preferentially expressed in liver. Mice lacking hepatic expression of HNF4alpha (HNF4alphaDeltaL) exhibited markedly increased levels of serum bile acids (BAs) compared with HNF4alpha-floxed (HNF4alphaF/F) mice. The expression of genes involved in the hydroxylation and side chain beta-oxidation of cholesterol, including oxysterol 7alpha-hydroxylase, sterol 12alpha-hydroxylase (CYP8B1), and sterol carrier protein x, was markedly decreased in HNF4alphaDeltaL mice. Cholesterol 7alpha-hydroxylase mRNA and protein were diminished only during the dark cycle in HNF4alphaDeltaL mice, whereas expression in the light cycle was not different between HNF4alphaDeltaL and HNF4alphaF/F mice. Because CYP8B1 expression was reduced in HNF4alphaDeltaL mice, it was studied in more detail. In agreement with the mRNA levels, CYP8B1 enzyme activity was absent in HNF4alphaDeltaL mice. An HNF4alpha binding site was found in the mouse Cyp8b1 promoter that was able to direct HNF4alpha-dependent transcription. Surprisingly, cholic acid-derived BAs, produced as a result of CYP8B1 activity, were still observed in the serum and gallbladder of these mice. These studies reveal that HNF4alpha plays a central role in BA homeostasis by regulation of genes involved in BA biosynthesis, including hydroxylation and side chain beta-oxidation of cholesterol in vivo.

Analysis of Solids, Liquids, and Biological Tissues Using Solids Probe Introduction at Atmospheric Pressure on Commercial LC/MS Instruments

Direct analysis of samples using atmospheric pressure ionization (API) provides a more rapid method for analysis of volatile and semivolatile compounds than vacuum solids probe methods and can be accomplished on commercial API mass spectrometers. With only a simple modification to either an electrospray (ESI) or atmospheric pressure chemical ionization (APCI) source, solid as well as liquid samples can be analyzed in seconds. The method acts as a fast solids/liquid probe introduction as well as an alternative to the new direct analysis in real time (DART) and desorption electrospray ionization (DESI) methods for many compound types. Vaporization of materials occurs in the hot nitrogen gas stream flowing from an ESI or APCI probe. Ionization of the thermally induced vapors occurs by corona discharge under standard APCI conditions. Accurate mass and mass-selected fragmentation are demonstrated as is the ability to obtain ions from biological tissue, currency, and other objects placed in the path of the hot nitrogen stream.

Crystal Structure of the Yersinia enterocolitica Type III Secretion Chaperone SycT

Several Gram-negative pathogens deploy type III secretion systems (TTSSs) as molecular syringes to inject effector proteins into host cells. Prior to secretion, some of these effectors are accompanied by specific type III secretion chaperones. The Yersinia enterocolitica TTSS chaperone SycT escorts the effector YopT, a cysteine protease that inactivates the small GTPase RhoA of targeted host cells. We solved the crystal structure of SycT at 2.5 angstroms resolution. Despite limited sequence similarity among TTSS chaperones, the SycT structure revealed a global fold similar to that exhibited by other structurally solved TTSS chaperones. The dimerization domain of SycT, however, differed from that of all other known TTSS chaperone structures. Thus, the dimerization domain of TTSS chaperones does not likely serve as a general recognition pattern for downstream processing of effector/chaperone complexes. Yersinia Yop effectors are bound to their specific Syc chaperones close to the Yop N termini, distinct from their catalytic domains. Here, we showed that the catalytically inactive YopT(C139S) is reduced in its ability to bind SycT, suggesting an ancillary interaction between YopT and SycT. This interaction could maintain the protease inactive prior to secretion or could influence the secretion competence and folding of YopT.