Alkaline phosphatase as a reporter of sigma(S) levels and rpoS polymorphisms in different E-coli strains

sigma(S) is responsible for the transcriptional regulation of genes related to protection against stresses and bacterial survival and it accumulates in the cell under conditions of stress, such as nutrient limitation. An increase in the levels of sigma(S) causes a reduction in the expression of genes that are transcribed by RNA polymerase associated with the principal sigma factor, sigma(70). phoA, that encodes alkaline phosphatase (AP) is expressed under phosphate shortage conditions, and is also repressed by sigma(S). Here we show that in a Pi-limited chemostat, accumulation of rpoS mutations is proportional to the intrinsic level of sigma(S) in the cells. Acquisition of mutations in rpoS relieves repression of the PHO genes. We also devised a non-destructive method based on the rpoS effect on AP that differentiates between rpo(S+) and rpoS mutants, as well as between high and low-sigma(S) producers. Using this method, we provide evidence that sigma(S) contributes to the repression of AP under conditions of Pi excess and that AP variation among different strains is at least partly due to intrinsic variation in sigma(S) levels. Consequently, a simple and non-destructive AP assay can be employed to differentiate between strains expressing different levels of sigma(S) on agar plates.

Crystal structures and thermal behavior of alkaline earth tricyanomethanides

The alkaline earth tricyanomethanides Mg(tcm)(2) center dot 2H(2)O, Ca(tcm)(2), Sr(tcm)(2) - H2O and Ba(tcm)(2) center dot 2H(2)O were prepared from aqueous solutions of the respective chlorides and silver tricyanomethanide. Their IR spectra and thermal behavior are described. The crystal structures of Ca(tcm)(2) and Ba(tcm)(2) center dot 2H(2)O were determined by single crystal X-ray diffraction. The structure of Ca(tcm)(2) is of the type found for several transition metal tricyanomethanides [1], containing two independent interpenetrating networks. Ba(tcm)(2) center dot 2H(2)O has a unique crystal structure corresponding to a three-dimensional coordination polymer with nine fold coordinated Ba atoms connected by water molecules and tricyanomethanide anions.

Mafic dykes at the southwestern margin of Eastern Ghats belt: Evidence of rifting and collision

The southwestern margin of the Eastern Ghats Belt characteristically exposes mafic dykes intruding massif-type charnockites. Dykes of olivine basalt of alkaline composition have characteristic trace element signatures comparable with Ocean Island Basalt (OIB). Most importantly strong positive Nb anomaly and low values of Zr/Nb ratio are consistent with OIB source of the mafic dykes. K-Ar isotopic data indicate two cooling ages at 740 and 530 Ma. The Pan-African thermal event could be related to reactivation of major shear zones and represented by leuco-granite vein along minor shear bands. And 740 Ma cooling age may indicate the low grade metamorphic imprints, noted in some of the dykes. Although no intrusion age could be determined from the present dataset, it could be constrained by some age data of the host charnockite gneiss and Alkaline rocks of the adjacent Prakasam Province. Assuming an intrusion age of similar to 1.3 Ga, Sr-Nd isotopic composition of the dykes indicate that they preserved time-integrated LREE enrichment. In view of the chemical signatures of OIB source, the mafic dykes could as well be related to continental rifting, around 1.3 Ga, which may have been initiated by intra-plate volcanism.

Microgranitic Enclaves as Products of Self-mixing Events: a Study of Open-system Processes in the Maua Granite, Sao Paulo, Brazil, Based on in situ Isotopic and Trace Elements in Plagioclase

Felsic microgranular enclaves with structures indicating that they interacted in a plastic state with their chemically similar host granite are abundant in the Maua Pluton, SE Brazil. Larger plagioclase xenocrysts are in textural disequilibrium with the enclave groundmass and show complex zoning patterns with partially resorbed An-rich cores (locally with patchy textures) surrounded by more sodic rims. In situ laser ablation-(multi-collector) inductively coupled plasma mass spectrometry trace element and Sr isotopic analyses performed on the plagioclase xenocrysts indicate open-system crystallization; however, no evidence of derivation from more primitive basic melts is observed. The An-rich cores have more radiogenic initial Sr isotopic ratios that decrease towards the outermost part of the rims, which are in isotopic equilibrium with the matrix plagioclase. These profiles may have been produced by either (1) diffusional re-equilibration after rim crystallization from the enclave-forming magma, as indicated by relatively short calculated residence times, or (2) episodic contamination with a decrease of the contaminant ratio proportional to the extent to which the country rocks were isolated by the crystallization front. Profiles of trace elements with high diffusion coefficients would require unrealistically long residence times, and can be modeled in terms of fractional crystallization. A combination of trace element and Sr isotope data suggests that the felsic microgranular enclaves from the Maua Pluton are the products of interaction between end-member magmas that had similar compositions, thus recording `self-mixing` events.

Zr- AND Ba-RICH MINERALS FROM THE PONTE NOVA ALKALINE MAFIC-ULTRAMAFIC MASSIF, SOUTHEASTERN BRAZIL: INDICATION OF AN ENRICHED MANTLE SOURCE

Zirconium- and Ba-rich minerals are found in gabbroic rocks from the Ponte Nova alkaline mafic-ultramafic massif in southeastern Brazil. The unusual mineralogical assemblage includes zirconolite, baddeleyite, Ba-rich alkali feldspar, and Ba- and Ti-rich biotite. Zirconolite of the Ponte Nova massif has higher levels of Zr (up to 1.172 apfu) than those registered in other terrestrial rocks and a prominent enrichment in the light rare-earth elements. Baddeleyite contains small quantities of Hf, Ti, and Fe. The Ba-rich alkali feldspar and Ba- and Ti-rich biotite contain up to 9.25 and 7.35 wt% BaO, respectively, and the biotite contains up to 12.01 wt% TiO(2). In the different intrusions of the Ponte Nova massif, such an unusual assemblage typifies the residual magma after the crystallization of clinopyroxene and olivine from previously enriched basanitic parental magma. The different trends of enrichments in REE and Th + U found for zirconolite of the intrusions of the Ponte Nova massif provide a better understanding of the variable degrees of enrichment of incompatible elements of the distinct gabbroic bodies. A lithospheric mantle source enriched in incompatible elements by the metasomatic action of volatile-rich fluids and with the presence of phlogopite or amphibole (or both) and other minor accessory phases could explain the presence of the Zr- and Ba-rich minerals in this gabbroic massif.

Alkaline magmatism in the Amambay area, NE Paraguay: The Cerro Sarambi complex

The Early Cretaceous alkaline magmatism in the northeastern region of Paraguay (Amambay Province) is represented by stocks, plugs, dikes, and dike swarms emplaced into Carboniferous to Triassic-Jurassic sediments and Precambrian rocks. This magmatism is tectonically related to the Ponta Pora Arch, a NE-trending structural feature, and has the Cerro Sarambi and Cerro Chiriguelo carbonatite complexes as its most significant expressions. Other alkaline occurrences found in the area are the Cerro Guazu and the small bodies of Cerro Apua, Arroyo Gasory, Cerro Jhu, Cerro Tayay, and Cerro Teyu. The alkaline rocks comprise ultramafic-mafic, syenitic, and carbonatitic petrographic associations in addition to lithologies of variable composition and texture occurring as dikes; fenites are described in both carbonatite complexes. Alkali feldspar and clinopyroxene, ranging from diopside to aegirine, are the most abundant minerals, with feldspathoids (nepheline, analcime), biotite, and subordinate Ti-rich garnet; minor constituents are Fe-Ti oxides and cancrinite as the main alteration product from nepheline. Chemically, the Amambay silicate rocks are potassic to highly potassic and have miaskitic affinity, with the non-cumulate intrusive types concentrated mainly in the saturated to undersaturated areas in silica syenitic fields. Fine-grained rocks are also of syenitic affiliation or represent more mafic varieties. The carbonatitic rocks consist dominantly of calciocarbonatites. Variation diagrams plotting major and trace elements vs. SiO(2) concentration for the Cerro Sarambi rocks show positive correlations for Al(2)O(3), K(2)O, and Rb, and negative ones for TiO(2), MgO, Fe(2)O(3), CaO, P(2)O(5), and Sr, indicating that fractional crystallization played an important role in the formation of the complex. Incompatible elements normalized to primitive mantle display positive spikes for Rb, La, Pb, Sr, and Sm, and negative for Nb-Ta, P, and Ti, as these negative anomalies are considerably more pronounced in the carbonatites. Chondrite-normalized REE patterns point to the high concentration of these elements and to the strong LRE/HRE fractionation. The Amambay rocks are highly enriched in radiogenic Sr and have T(DM) model ages that vary from 1.6 to 1.1 Ga. suggesting a mantle source enriched in incompatible elements by metasomatic events in Paleo-Mesoproterozoic times. Data are consistent with the derivation of the Cerro Sarambi rocks from a parental magma of lamprophyric (minette) composition and suggest an origin by liquid immiscibility processes for the carbonatites. (C) 2011 Elsevier Ltd. All rights reserved.

Fabrics of pre- and syntectonic granite plutons and chronology of shear zones in the Eastern Borborema Province, NE Brazil

We used the fabrics of two granite plutons and U/Pb (SHRIMP) zircon ages to constrain the tectonic evolution of the E-trending Patos shear zone (Borborema Province, NE Brazil). The pre-tectonic Teixeira batholith consists of an amphibole leucogranite locally with aegirine-augite. Zircons from a syenogranite yielded crystallization ages of 591 +/- 5 Ma. The batholith fabrics were determined by anisotropy of magnetic susceptibility (AMS) and mineral shape preferred orientation. The fabrics support pre-transcurrent batholith emplacement, as evidenced by: (i) magmatic/magnetic fabrics in low susceptibility (<0.35 mSI) leucogranites highly discordant to the regional host rock structure, and (ii) concordant magnetic fabrics restricted to high susceptibility (>1 mSI) corridors connected to shear zones branching off from Patos. One of these satellite shear zones controlled the syntectonic emplacement of the Serra Redonda pluton, which yields a crystallization age of 576 +/- 3 Ma. This late shearing event marks the peak regional deformation that, south of Patos, was coupled to crustal shortening nearly perpendicular to the shear belt. The chronology of the deformational events indicates that the major shear zones of the eastern Borborema are late structures active after the crustal blocks amalgamated. (C) 2007 Elsevier Ltd. All rights reserved.

Characterization of a beta-1,3-glucanase active in the alkaline midgut of Spodoptera frugiperda larvae and its relation to beta-glucan-binding proteins

Spodoptera frugiperda beta-1,3-glucanase (SLam) was purified from larval midgut. It has a molecular mass of 37.5 kDa, an alkaline optimum pH of 9.0, is active against beta-1,3-glucan (laminarin), but cannot hydrolyze yeast beta-1,3-1,6-glucan or other polysaccharides. The enzyme is an endoglucanase with low processivity (0.4), and is not inhibited by high concentrations of substrate. In contrast to other digestive beta-1,3-glucanases from insects, SLam is unable to lyse Saccharomyces cerevisae cells. The cDNA encoding SLam was cloned and sequenced, showing that the protein belongs to glycosyl hydrolase family 16 as other insect glucanases and glucan-binding proteins. Multiple sequence alignment of beta-1,3-glucanases and beta-glucan-binding protein supports the assumption that the beta-1,3-glucanase gene duplicated in the ancestor of mollusks and arthropods. One copy originated the derived beta-1,3-glucanases by the loss of an extended N-terminal region and the beta-glucan-binding proteins by the loss of the catalytic residues. SLam homology modeling suggests that E228 may affect the ionization of the catalytic residues, thus displacing the enzyme pH optimum. SLam antiserum reacts with a single protein in the insect midgut. Immunocytolocalization shows that the enzyme is present in secretory vesicles and glycocalyx from columnar cells. (C) 2010 Elsevier Ltd. All rights reserved.

Bismuth Modified Gold Microelectrode for Pb(II) Determination in Wine Using Alkaline Medium

Electrodeposition of bismuth on gold microelectrodes for determination of Pb(II) by square wave anodic stripping voltammetry (SWASV) was accomplished by an in situ procedure in alkaline solution. A linear calibration plot for Pb(II) in the concentration range 40 to 6700 nmol L(-1) (r=0.998) was obtained, the detection limit was found to be 12.5 nmol L(-1) (S/N = 3) and the relative standard deviation in Solutions containing 1 mu mol L(-1) Pb(II) was 4% (n = 12). The analytical performance of the proposed sensor wits tested by measuring the Pb(II) concentration in a wine sample. The result Was in good agreement with the one obtained by GFAAS.

Ex Situ Scanning Electrochemical Microscopy (SECM) Investigation of Bismuth- and Bismuth/Lead Alloy Film-Modified Gold Electrodes in Alkaline Medium

Scanning electrochemical microscopy (SECM) in feedback mode was employed to characterise the reactivity and microscopic peculiarities of bismuth and bismuth/lead alloys plated onto gold disk substrates in 0.1 molL(-1) NaOH solutions. Methyl viologen was used as redox mediator, while a platinum microelectrode was employed as the SECM tip. The metal films were electrodeposited ex situ from NaOH solutions containing either bismuth ions only or both bismuth and lead ions. Approach curves and SECM images indicated that the metal films were conductive and locally reactive with oxygen to provide Bi(3+) and Pb(2+) ions. The occurrence of the latter chemical reactions was verified by local anodic stripping voltammetry (ASV) at the substrate solution interface by using a mercury-coated platinum SECM tip. The latter types of measurements allowed also verifying that lead was not uniformly distributed onto the bismuth film electrode substrate. These findings were confirmed by scanning electron microscopy images. The surface heterogeneity produced during the metal deposition process, however, did not affect the analytical performance of the bismuth coated gold electrode in anodic stripping voltammetry for the determination of lead in alkaline media, even in aerated aqueous solutions. Under the latter conditions, stripping peak currents proportional to lead concentration with a satisfactory reproducibility (within 5% RSD) were obtained.

Synthesis, characterization and spectroscopic investigation of new tetrakis(acetylacetonato)thulate(III) complexes containing alkaline metals as countercations

In this work a series of tetrakis complexes C[Tm(acac)(4)] where C(+) = Li(+) Na(+) and K(+) countercations and acac = acetylacetonate ligand were synthesized and characterized for photoluminescence investigation The relevant aspect is that these complexes are water-free in the first coordination sphere The emission spectra of the tetrakis Tm(3+)-complexes present narrow bands characteristic of the (1)G(4)->(3)H(6) (479 nm) (1)G(4)->(3)F(4) (650 nm) and (1)G(4) ->(3)H(5) (779 nm) transitions of the Tm(3+) ion with the blue emission color at 479 nm as the most prominent one The lifetime values (tau) of the emitting (1)G(4) level of the C[Tm(acac)(4)] complexes were 344 360 and 400 ns for the Li(+) Na(+) and K(+) countercations respectively showing an increasing linear behavior versus the ionic radius of the alkaline ion An efficient intramolecular energy transfer process from the triplet state (T) of the ligands to the emitting (1)G(4) state of the Tm(3+) ion is observed This fact together with the absence of water molecules in first coordination sphere allows these tetrakis Tm(3+)-complexes to act as efficient blue light conversion molecular devices (c) 2010 Elsevier B V All rights reserved

Pt monolayer electrocatalysts for O-2 reduction: PdCo/C substrate-induced activity in alkaline media

We measured the activity of electrocatalysts, comprising Pt monolayers deposited on PdCo/C substrates with several Pd/Co atomic ratios, in the oxygen reduction reaction in alkaline solutions. The PdCo/C substrates have a core-shell structure wherein the Pd atoms are segregated at the particle`s surface. The electrochemical measurements were carried out using an ultrathin film rotating disk-ring electrode. Electrocatalytic activity for the O-2 reduction evaluated from the Tafel plots or mass activities was higher for Pt monolayers on PdCo/C compared to Pt/C for all atomic Pd/Co ratios we used. We ascribed the enhanced activity of these Pt monolayers to a lowering of the bond strength of oxygenated intermediates on Pt atoms facilitated by changes in the 5d-band reactivity of Pt. Density functional theory calculations also revealed a decline in the strength of PtOH adsorption due to electronic interaction between the Pt and Pd atoms. We demonstrated that very active O-2 reduction electrocatalysts can be devised containing only a monolayer Pt and a very small amount of Pd alloyed with Co in the substrate.

Molecular masses and sedimentation coefficients of extracellular hemoglobin of Glossoscolex paulistus: Alkaline oligomeric dissociation

The giant extracellular hemoglobin of Glossoscolex paulistus (HbGp) has a molecular mass (M) of 3600 +/- 100 kDa and a standard sedimentation coefficient (s(20.w)(0)) of 58 S. estimated by analytical ultracentrifugation (AUC). In the present work, further AUC studies were developed for HbGp, at pH 10.0, which favors oligomeric dissociation into lower M species. The HbGp oligomer is formed by globin chains a, b, c and d plus the linker chains. The pure monomeric fraction, subunit d, and HbGp at pH 10.0, in the presence of beta-mercaptoethanol, were also studied. Our results indicate that for samples of pure subunit d, besides the monomeric species with s(20.w)(0) of 2.0 S, formation of dimer of subunit d is observed with s(20.w)(0) of around 2.9 S. For the whole HbGp at pH 10.0 contributions from monomers, trimers and linkers are observed. No contribution from 58 S species was observed for the sample of oxy-HbGp at pH 10.0, showing its complete dissociation. For cyanomet-HbGp form a contribution of 17% is observed for the un-dissociated oligomer, consistent with data from other techniques that show the cyanomet-form is more stable as compared to oxy-HbGp. Masses of HbGp subunits, especially trimer abc and monomeric chains a, b, c and d, were also estimated from sedimentation equilibrium data, and are in agreement with the results from MALDI-TOF-MS. (C) 2010 Elsevier B.V. All rights reserved.

Culture of osteogenic cells from human alveolar bone: A useful source of alkaline phosphatase

The aim of this study was to obtain membrane-bound alkaline phosphatase from osteoblastic-like cells of human alveolar bone. Cells were obtained by enzymatic digestion and maintained in primary culture in osteogenic medium until subconfluence. First passage cells were cultured in the same medium and at 7, 14, and 21 days, total protein content, collagen content, and alkaline phosphatase activity were evaluated. Bone-like nodule formation was evaluated at 21 days. Cells in primary culture at day 14 were washed with Tris-HCl buffer, and used to extract the membrane-bound alkaline phosphatase. Cells expressed osteoblastic phenotype. The apparent optimum pH for PNPP hydrolysis by the enzyme was pH 10.0. This enzyme also hydrolyzes ATP, ADP, fructose-1-phosphate, fructose-6-phosphate, pyrophosphate and beta-glycerophosphate. PNPPase activity was reduced by typical inhibitors of alkaline phosphatase. SDS-PAGE of membrane fraction showed a single band with activity of similar to 120 kDa that could be solubilized by phospholipase C or Polidocanol. (c) 2007 International Federation for Cell Biology. Published by Elsevier Ltd. All rights reserved.

Allosteric modulation of pyrophosphatase activity of rat osseous plate alkaline phosphatase by magnesium ions

Pyrophosphatase activity of rat osseous plate alkaline phosphatase was studied at different concentrations of calcium and magnesium ions. with the aim of characterizing the modulation of enzyme activity by these metals. In the absence of metal ions, the enzyme hydrolysed pyrophosphate following Michaelian kinetics with a specific activity of 36.7 U/mg and K-0.5 = 88 mu M. In the presence of low concentrations (0.1 mM) of magnesium (or calcium) ions, the enzyme also exhibited Michaclian kinetics for the hydrolysis of pyrophosphate, but a significant increase in specific activity (123 U/mg) was observed. K-m values remained almost unchanged. Quite different behavior occurred in the presence of 2 mM magnesium (or calcium) ions. In addition to low-affinity sites (K-0.5 = 40 and 90 mu M, for magnesium and calcium, respectively), high-affinity sites were also observed with K-0.5 values 100-fold lower. The high-affinity sites observed in the presence of calcium ions represented about 10% of those observed for magnesium ions. This was correlated with the fact that only magnesium ions triggered conformational changes yielding a fully active enzyme. These results suggested that the enzyme could hydrolyse pyrophosphate, even at physiological concentrations (4 mu M), since magnesium concentrations are high enough to trigger conformational changes increasing the enzyme activity. A model, suggesting the involvement of magnesium ions in the hydrolysis of pyrophosphate by rat osseous plate alkaline phosphatase is proposed. (C) 1998 Published by Elsevier B.V. Ltd. All rights reserved.