A theoretical study on the reaction mechanism of O<Subscript>2</Subscript> with C<Subscript>4</Subscript>H<Subscript>9</Subscript>• radical

Ab initio calculations have been performed using the complete basis set model (CBS-QB3) to study the reaction mechanism of butane radical (C₄H₉•) with oxygen (O₂). On the calculated potential energy surface, the addition of O₂ to C₄H₉• forms three intermediates barrierlessly, which can undergo subsequent isomerization or decomposition reaction leading to various products: HOO• + C₄H₈, C₂H₅• + CH₂CHOOH, OH• + C₃H₇CHO, OH• + cycle-C₄H₈O, CH₃• + CH₃CHCHOOH, CH₂OOH• + C₃H₆. Five pathways are supposed in this study. After taking into account the reaction barrier and enthalpy, the most possible reaction pathway is C₄H₉• + O₂ → IM1 → TS5 → IM3 → TS6 → IM4 → TS7 → OH• + cycle-C₄H₈O.     

Thiosulfate stimulates growth and alleviates silver and copper toxicity in tomato root cultures

The influence of supplemented thiosulfate (S₂O₃ ²⁻) as well as a complex of either Ag⁺ or Cu²⁺ with S₂O₃ ²⁻ in the culture medium on proliferating root cultures of tomato (Solanum lycopersicum) was investigated. The presence of 10–300 μM sodium thiosulfate (Na₂S₂O₃) in half-strength Murashige and Skoog (MS) basal salt medium promoted root elongation and proliferation of lateral roots. Growth was enhanced by 1–2 μM AgNO₃, but was completely arrested at 5 μM AgNO₃; moreover, growth inhibition was elicited by dissolved silver (Ag⁺) and by silver in silver precipitate particles. Root elongation was also inhibited by 50 μM CuSO₄ supplemented to the basal medium. Roots subjected to either AgNO₃ or CuSO₄ growth inhibiting treatments were unable to recover following transfer to medium lacking either Ag⁺ or Cu²⁺. When the basal medium was supplemented with either silver or copper in the form of silver thiosulfate complex or copper thiosulfate complex, root cultures continued to elongate and proliferate, thus either completely alleviating or diminishing the inhibitory effects of Ag⁺ and Cu²⁺, respectively. It was concluded that tomato roots sensed and responded to S₂O₃ ²⁻, hence root proliferation could be promoted by adding Na₂S₂O₃ to the medium. Moreover, a complex of Ag⁺ with S₂O₃ ²⁻ detoxified dissolved Ag⁺ and prevented the generation of toxic silver particle precipitates. Consequently, silver thiosulfate was superior to AgNO₃ in enhancing root culture. Finally, a complex of Cu²⁺ with S₂O₃ ²⁻ ligand reduced toxicity of Cu²⁺ to root cultures of tomato.     

Reversible sporicidal action of cuprous ions

At 10 mM, Cu⁺ was highly protective against killing of spores of Bacillus megaterium ATCC 19213 by H₂O₂, while at higher concentrations, from 15–100 mM, killing was augmented. In contrast, Cu²⁺, Fe²⁺, Fe³⁺, Co²⁺ or Co³⁺ ions acted only protectively. Cu⁺ itself was sporicidal in the absence of H₂O₂ or ascorbate, and its sporicidal action did not depend on generation of highly reactive oxygen species. It appeared that killing involved either inhibition of germination or copper toxicity to germinated cells in that Cu⁺-inactivated spores did not germinate readily but chemical decoating of the cells prior to plating on a solid medium resulted in reversal of the sporicidal effect. Received 12 July 1996/ Accepted in revised form 03 November 1996     

Adhesion forces in the self-recognition of oligosaccharide epitopes of the proteoglycan aggregation factor of the marine sponge <Emphasis Type="Italic">Microciona prolifera</Emphasis>

Cell aggregation in the marine sponge Microciona prolifera is mediated by a multimillion molecular-mass aggregation factor, termed MAF. Earlier investigations revealed that the cell aggregation activity of MAF depends on two functional domains: (i) a Ca²⁺-independent cell-binding domain and (ii) a Ca²⁺-dependent proteoglycan self-interaction domain. Structural analysis of involved carbohydrate fragments of the proteoglycan in the self-association established a sulfated disaccharide β-d-GlcpNAc3S-(1→3)-α-l-Fucp and a pyruvated trisaccharide β-d-Galp4,6(R)Pyr-(1→4)-β-d-GlcpNAc-(1→3)-α-l-Fucp. Recent UV, SPR, and TEM studies, using BSA conjugates and gold nanoparticles of the synthetic sulfated disaccharide, clearly demonstrated self-recognition on the disaccharide level in the presence of Ca²⁺-ions. To determine binding forces of the carbohydrate–carbohydrate interactions for both synthetic MAF oligosaccharides, atomic force microscopy (AFM) studies were carried out. It turned out that, in the presence of Ca²⁺-ions, the force required to separate the tip and sample coated with a self-assembling monolayer of thiol-spacer-containing β-d-GlcpNAc-(1→3)-α-l-Fucp-(1→O)(CH₂)₃S(CH₂)₆S- was found to be quantized in integer multiples of 30 ± 6 pN. No binding was observed between the two monolayers in the absence of Ca²⁺-ions. Cd²⁺-ions could partially induce the self-interaction. In contrast, similar AFM experiments with thiol-spacer-containing β-d-Galp4,6(R)Pyr-(1→4)-β-d-GlcpNAc-(1→3)-α-l-Fucp-(1→O)(CH₂)₃S(CH₂)₆S- did not show a binding in the presence of Ca²⁺-ions. Also TEM experiments of gold nanoparticles coated with the pyruvated trisaccharide could not make visible aggregation in the presence of Ca²⁺-ions. It is suggested that the self-interaction between the sulfated disaccharide fragments is stronger than that between the pyruvated trisaccharide.     

Carbonylation and other metal-catalysed C?C coupling reactions used in industry

The formation of new carbon-carbon bonds is probably the most fundamental reaction in organic chemistry. Many of the most selective and energy efficient reactions today involve the use of metals and their complexes as promoters. Key routes to C−C bond formation involve carbonylation (using CO) and coupling reactions (of olefins, dienes, and arenes) following an activation step. The largest carbonylation reactions in volume terms are the hydrogenation of CO, to make long chain hydrocarbons for diesel fuel (over supported Fe, Co, Ru or Rh metal catalysts; the Fischer-Tropsch reaction) or, over a Cu−ZnO catalyst, to make methanol. Many of the important carbonylations are homogeneously catalysed, using soluble complexes of the late transition metals, such as Co, Rh, Ir, or Pd. The carbonylation of methanol to acetic acid, MeOH+CO→MeCOOH was originally developed using a Co/I⁻ catalyst; improvements in conditions (better energy use and selectivity), led by mechanistic considerations resulted in, first the Rh/I⁻ (Monsanto) and more recently, the Ir/I⁻ (BP, Cativa) processes. The hydroformylation of olefins,e.g., MeCH=CH₂+H₂+CO→MeCH₂CH₂CHO+MeCH(CHO)CH₃ was also originally developed using a soluble cobalt catalyst, but again the use of rhodium catalysts (bearing phosphine ligands) has largely superseded the older processes. Even newer technologies, for example involvingsupported homogeneous catalysts, now promise still cleaner and more selective processes. Direct coupling reactions include Friedel-Crafts type processes, for example, C₆H₆+CH₂=CHR→C₆H₅−C₂H₄R Zeolites are now the strong acid catalysts employed heterogeneously rather than the previously used AlCl₃ or HF; the new catalysts are much more environmentally friendly (?greener?) as they do not involve halides which cause corrosion and undesirable side reactions. Valuable highly selective reactions include the Cr-catalysed oligomerization of ethylene to 1-hexene or 1-octene, and of butadiene to cyclo-octadiene or cyclo-dodecatriene, over Ni catalysts. Palladium catalysed coupling and carbonylation reactions have made key steps much more environmentally acceptable in the syntheses of the important anti-inflammatory pharmaceuticalsibuprofen andnaproxen.

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Riassunto   Carbonilazione ed altri accoppiamenti C−C catalizzati da metalli impiegati nell'industria. La formazione di nuovi legami carbonio-carbonio è forse la reazione più fondamentale in chimica organica. Molte delle reazioni più selettive ed energeticamente efficienti implicano oggi l'impiego di metalli e loro complessi come promotori. Le vie chiave per la formazione di legami C−C sono la carbonilazione (mediante CO) e le reazioni di accoppiamento (di olefine, dieni ed areni) precedute da uno stadio di attivazione. Le maggiori reazioni di carbonilazione in termini di volume sono l'idrogenazione del CO per la produzione di idrocarburi a lunga catena destinati all'impiego come combustibili Diesel (su catalizzatiri metallici supportati a base di Fe, Co, Ru o Rh; reazione di Fischer-Tropsch) o di metanolo su catalizzatore Cu−ZnO. Molte tra le più importanti carbonilazioni avvengono su catalizzatori in fase omogenea costituiti da complessi solubili di metalli alla fine di una serie di transizione come Co, Rh, Ir e Pd. La carbonilazione del metanolo ad acido acetico MeOH+CO→MeCOOH venne originariamente sviluppata usando un catalizzatore Co/I⁻; il miglioramento delle condizioni (miglior uso dell'energia e maggior selettività) sulla base di considerazioni meccanicistiche condusse in un primo tempo al catalizzatore Rh/I⁻ (processo Monsanto) e più recentemente al catalizzatore Ir/I⁻ (processo BP, Cativa). L'idroformilazione delle olefine, per esempio: MeCH=CH₂+H₂+CO→MeCH₂CH₂CHO+MeCH(CHO)CH₃ venne anch'essa sviluppata originariamente usando un complesso solubile del cobalto, ma di nuovo l'uso dei catalizzatori a base di rodio (contenenti leganti fosfinici) ha largamente superato i processi più vecchi. Anche le nuove technologie basate su catalizzatori omogenei supportati promettono ora processi ancora più puliti e selettivi. Le reazioni di accoppiamento diretto comprendono i processi di tipo Friedel-Crafts, per esempio: C₆H₆+CH₂=CHR→C₆H₅−C₂H₄R. Le zeoliti rappresentano ora i catalizzatori fortemente acidi impiegati in fase etrogenea al posto di AlCl₃ od HF usati in precedenza; i nuovi catalizzatori sono molto più compatibili ambientalmente (?più verdi?) in quanto non contengono alogenuri che possono provocare corrosioni ed altre reazioni secondarie non desiderate. Pregevoli reazioni ad alta selettività comprendono l'oligomerizzazione dell'etilene ad 1-esene od 1-ottene catalizzata dal cromo o del butadiene a cicloottadiene o ciclododecatriene catalizzata dal nichel. Attraverso reazioni di accoppiamento e carbonilazione catalizzate dal palladio sono stati realizzati passaggi chiave molto più accettabili ambientalmente nella sintesi degli importanti composti farmaceuticiibuprofen enaproxen.

     

Characteristics of non-specific permeability and H+-ATPase inhibition induced in the plasma membrane of Nitella flexilis by excessive Cu2+

Effects of Cu²⁺ on a non-specific conductance and H⁺-ATPase activity in the plasma membrane of the freshwater alga Nitella flexilis L. Agardh was studied using a conventional microelectrode voltage-clamp technique. We show that a Cu²⁺-induced increase in the non-specific conductance is related to the formation of pores in the plasma membrane. Pore formation is the result of unidentified chemical reactions, since the Q₁₀ for the rate of increase of conductance over time was about 3. Various oxidants and antioxidants (10 mmol/l H₂O₂, 10 mmol/l ascorbate, 100 μg/ml superoxide dismutase, and 100 μg/ml catalase) did not alter Cu²⁺-induced changes in the plasma membrane conductance, suggesting that the effect of Cu²⁺ was unrelated to peroxidation of plasma-membrane lipids. In contrast, organic and inorganic Ca²⁺-channel antagonists (nifedipine, Zn²⁺, Cd²⁺, Fe²⁺, Ni²⁺) inhibited the Cu²⁺-induced non-specific conductance increase. This suggests that changes in Ca²⁺ influx underlie this effect of Cu²⁺. Decreasing the pH or the ionic strength of external solutions also inhibited the Cu²⁺-induced plasma-membrane conductance increase. Copper was also found to inhibit plasma-membrane H⁺-ATPase activity with half-maximal inhibition occurring at about 5–20 μmol/l and full inhibition at about 100–300 μmol/l. The Hill coefficient of Cu²⁺ inhibition of the H⁺-ATPase was close to two. Received: 8 December 1999 / Accepted: 16 August 2000     

Energy transduction efficiencies in nitrogenous oxide respirations of Azospirillum brasilense Sp7

For Azospirillum brasilense Sp7, the energy transformation efficiencies were measured in anaerobic respirations with either nitrate, nitrite or nitrous oxide as respiratory electron acceptors by determining the maximal molar growth yields and the H⁺-translocations using the oxidant pulse method. In continuous cultures grown with malate limiting, the maximal molar growth yields (Y _s ^max -values) were essentially the same with O₂ or N₂O but were 1/3 and 2/3 lower with NO ₂ ^- or NO ₃ ^- , respectively, as respiratory electron acceptors. Both the maximal molar growth yields and the maintenance energy coefficients were surprisingly high when Azospirillum was grown with nitrite as the sole electron acceptor and source for N-assimilation. Growth under N₂-fixing conditions drastically reduced the Y _s ^max -values in the N₂O and O₂-respiring cells. In the H⁺-translocation measurements, the /oxidant ratios were 5.6 for O₂→H₂O, 2.5–2.8 for NO ₃ ^- →NO ₂ ^- , 2.2 for NO ₂ ^- →N₂O and 3.1 for N₂O→N₂ respirations when the cells were preincubated with valinomycin and K⁺. All the values were enhanced when the experiments were performed with valinomycin plus methyltriphenylphosphonium (=TPMP⁺) cation. The uncoupler carbonyl cyanide-m-chlorophenyl-hydrazone diminished the H⁺-excretion indicating that this translocation was due to vectorial flow across the membrane. In the absence of any ionophore, nitrate and nitrite respirations were accompanied by a H⁺-uptake . Any significant H⁺-translocation could not be detected in N₂O- and O₂-respirations under these conditions. It is concluded that nitrate reduction proceeds inside the cytoplasmic membrane, whereas nitrite is reduced extramembraneously. The data are not conclusive for the location of nitrous oxide reductase. The maximal molar growth yield determinations and the absence of any H⁺-uptake in untreated cells indicate a cytoplasmic orientation of the enzyme similar to the terminal cytochrome oxidase of respiration. The low H⁺-extrusion values for N₂O-respiration compared to O₂-respiration in cells treated with valinomycin plus TPMP⁺ are, however, not in accord with such an interpretation.     

Theoretical study of crown ethers with incorporated azobenzene moiety

A series of crown ethers containing the azobenzene moiety incorporated into crowns of various sizes [Cr(O₆)_, Cr(O₇) and Cr(O₈)] and their corresponding alkali metal cation (Li⁺, Na⁺, K⁺, Rb⁺) complexes have been studied theoretically. The density functional theory (DFT) method was employed to elucidate the stereochemical structural natures and thermodynamic properties of all of the target molecules at the B3LYP/6-31 G(d) and LANL2DZ level for the cation Rb⁺. The fully optimized geometries had real frequencies, thus indicating their minimum-energy status. In addition, the bond lengths between the metal cation and oxygen atoms, atomic torsion angles and thermodynamic energies for complexes were studied. Natural bond orbital (NBO) analysis was used to explore the origin of the internal forces and the intermolecular interactions for the metal complexes. The calculated results show that the most significant interaction is that between the lone pair electrons of electron-donating oxygens in the cis-forms of azobenzene crown ethers (cis-ACEs) and the LP* (1-center valence antibond lone pair) orbitals of the alkali-metal cations (Li⁺, Na⁺, K⁺ and Rb⁺). The electronic spectra for the cis-ACEs [cis-Cr(O₆), cis-Cr(O₇) and cis-Cr(O₈)] are obtained by the time-dependent density functional theory (TDDFT) at the B3LYP/6-31 G(d) level. The spectra of the cis-isomers show broad π → π* (S₀ → S₂) absorption bands at 310–340 nm but weaker n → π* (S₀ → S₁) bands at 480–490 nm. The calculated results are in good agreement with the experimental results.     

Kinetic studies on the reactions of Fusarium galactose oxidase with five different substrates in the presence of dioxygen

 Reactions (25  °C) of galactose oxidase, GOase_ox from Fusarium NRRL 2903 with five different primary-alcohol-containing substrates RCH₂OH:- D-galactose (I) and 2-deoxy-d-galactose (II) (monosaccharides); methyl-β-d-galactopyranoside (III) (glycoside);d-raffinose (IV) (trisaccharide); and dihydroxyacetone (V) have been studied in the presence of O₂. The GOase_ox state has a tyrosyl radical coordinated at a square-pyramidal Cu^II active site, and is a two-equivalent oxidant. Reactant concentrations were [GOase_ox] (0.8–10 μM), RCH₂OH (1.0–6.0 mM), and O₂ (0.14–0.29 mM), with I=0.100 M (NaCl). The reactions, monitored at 450 nm by stopped-flow spectrophotometry, terminated with depletion of the O₂. Each trace was fitted to the competing reactions GOase_ox+RCH₂ OH → GOase_redH₂+RCHO (k ₁), and GOase_redH₂+O₂→ GOase_ox+H₂O₂ (k ₂), with GOase_redH₂ written as the doubly protonated two-electron-reduced Cu^I product. It was necessary to avoid auto-redox interconversion of GOase_ox and GOase_semi . Information obtained at pH 7.5 indicates a 5 : 95 (ox : semi) "native" mix equilibration complete in ∼3 h. At pH >7.5, rate constants 10^–4 k ₁ / M^–1 s^–1 for the reactions of GOase_ox with (I) (1.19), (II) (1.07), (III) (1.29), (IV) (1.81), (V) (2.94) were determined. On decreasing the pH to 5.5, k ₁ values decreased by factors of up to a half, and acid dissociation pK _as in the range 6.6–6.9 were obtained. UV-Vis spectrophotometric studies on GOase_ox gave an independently determined pK _a of 6.7. No corresponding reactions of the Tyr495Phe variant were observed, and there are no similar UV-Vis absorbance changes for this variant. The pK _a is therefore assigned to protonation of Tyr-495 which is a ligand to the Cu. The rate constant k ₂ (1.01×10⁷ M^–1 s^–1) is independent of pH in the range 5.5–9.0 investigated, suggesting that H⁺ (or H-atoms) for the O₂ → H₂O₂ change are provided by the active site of GOase_red . The Cu^I of GOase_red is less extensively complexed, and a coordination number of three is likely. Received: 4 February 1997 / Accepted: 16 May 1997     

The reaction mechanism of nitrosothiols with copper(I)

Copper and other transition metal ions and their complexes are catalysts for the decomposition of nitrosothiols. In this way they catalyze the biological functions of nitrosothiols. The kinetics and mechanism of the reaction of two nitrosothiols, S-nitrosothiolactic acid and S-nitrosoglutathione (GSNO), with copper(I) are reported. The kinetics of the reaction of Cu(MeCN) _n ⁺ (n=0–3) with the nitrosothiols were studied. The results indicate that Cu⁺ _aq is the active species in the GSNO system, with k(Cu⁺ _aq+GSNO)=(9.4 ±2.0)×10⁷ dm³ mol⁻¹ s⁻¹. The results also indicate that the Cu(MeCN) _n ⁺ (n=0–3) complexes react with S-nitrosothiolactic acid. Transient species are formed in these processes. The results suggest that these species contain copper(I) and thiol. The results shed light on the catalytic role of copper complexes in the decomposition of S-nitrosothiols. Received 10 April 1999 / Accepted 17 December 1999     

An assessment of thermodynamic reaction constants for simulating aqueous environmental monomethylmercury speciation

ABSTRACT

Monomethylmercury (CH₃Hg ⁺) is both the most ecologically significant and the least well characterized species of mercury in environmental settings. Our understanding of the environmental speciation behavior of this compound is limited both as the result of lesser available laboratory data (when compared to inorganic mercury) as well as the uncertainties associated with our understanding of the properties of environmental ligands. A careful examination and synthesis of data reported in the technical literature led to the following findings: (1) a 25°C, zero ionic strength bicarbonate ion complexation constant estimate is remarkably close to an earlier reported value at 0.4 M: CH₃Hg⁺ + HCo₃^-?CH₃HgHCO₃,log₁₀K = 2.6 (±0.22, 1 SD), (2) three 25°C zero ionic strength reaction constants reported by DeRobertis et al.(1998) were confirmed to within ~±0.1 log₁₀K units: CH₃Hg ⁺+ OH^-?CH₃HgOH, log₁₀K = 9.47; 2CH₃Hg ⁺ + H₂O?(CH₃Hg)₂OH ⁺ + H⁺, log₁₀K =?2.15; CH₃Hg ⁺+ Cl^-?CH₃HgCl, log₁₀K = 5.45, (3) “best estimate” literature complexation constants corrected to zero ionic strength include: CH₃Hg ⁺ + F^-?CH₃HgF, log₁₀K = 1.75 (20°C corr. Schwart-zenbach and Schellenberg, 1965); CH₃Hg ⁺ + Br^-?CH₃HgBr, log₁₀K = 6.87 (20°C corr. Schwartzenbach and Schellenberg, 1965); CH₃Hg ⁺ +1^-?CH₃HgI, log₁₀K = 8.85 (20°C corr. Schwartzenbach and Schellenberg, 1965); and CH₃Hg ⁺+ SO₄^2-?CH₃HgSO₄^-,log₁₀K = 2.64 (25°C, DeRobertis et al., 1998), (4) literature reported values for simulating monomethylmercury complexation with the carbonate ion may be too low: CH₃Hg ⁺+ CO₃^2-?CH₃HgCO₃^-, log₁₀K = 6.1 (Rabenstein et al., 1976; Erni, 1981), and (5) ‘‘best estimate’’ constants for simulating methyl mercury complexation with reduced environmental sulfur species include: CH₃Hg ⁺ + S^2-?CH₃HgS ^-, log₁₀K = 21.1; CH³Hg ⁺+ SH ^-? CH3HgSH, log₁₀K = 14.5 (H ⁺ + SH^-?CH₂S, log₁₀K = 6.88; Dyrssen and Wedborg, 1991); CH₃Hg ⁺ + RS^-?CH₃HgSR, log₁₀K = 16.5 (H + + RS^-?RSH, log₁₀K = 9.96; Qian et al., 2002); and CH₃Hg ⁺+ CH₃HgS^{1 -}?(CH₃Hg)₂S, log₁₀K = 16.32 (Schwartzenbach and Schellenberg, 1965; Rabenstein et al., 1978; and Erni, 1981).     

Investigation of the redox interaction between Mn-bicarbonate complexes and reaction centers from Rhodobacter sphaeroides R-26, Chromatium minutissimum, and Chloroflexus aurantiacus

The change in the dark reduction rate of photooxidized reaction centers (RC) of type II from three anoxygenic bacteria (Rhodobacter sphaeroides R-26, Chromatium minutissimum, and Chloroflexus aurantiacus) having different redox potentials of the P⁺/P pair and availability of RC for exogenous electron donors was investigated upon the addition of Mn²⁺ and HCO₃⁻. It was found that the dark reduction of P₈₇₀⁺ from Rb. sphaeroides R-26 is considerably accelerated upon the combined addition of 0.5 mM MnCl₂ and 30–75 mM NaHCO₃ (as a result of formation of “low-potential” complexes [Mn(HCO₃)₂]), while MnCl₂ and NaHCO₃ added separately had no such effect. The effect is not observed either in RC from Cf. aurantiacus (probably due to the low oxidation potential of the primary electron donor, P₈₆₅, which results in thermodynamic difficulties of the redox interaction between P₈₆₅⁺ and Mn²⁺) or in RC from Ch. minutissimum (apparently due to the presence of the RC-bound cytochrome preventing the direct interaction between P₈₇₀⁺ and Mn²⁺). The absence of acceleration of the dark reduction of P₈₇₀⁺ in the RC of Rb. sphaeroides R-26 when Mn²⁺ and HCO₃⁻ were replaced by Mg²⁺ or Ca²⁺ and by formate, oxalate, or acetate, respectively, reveals the specificity of the Mn²⁺-bicarbonate complexes for the redox interaction with P⁺. The results of this work might be considered as experimental evidence for the hypothesis of the participation of Mn²⁺ complexes in the evolutionary origin of the inorganic core of the water oxidizing complex of photosystem II.     

Stimulation and oxidative catalytic inactivation of thermolysin by copper•Cys-Gly-His-Lys

[Cu²⁺•Cys-Gly-His-Lys] stimulates thermolysin (TLN) activity at low concentration (below 10 μM) and inhibits the enzyme at higher concentration, with binding affinities of 2.0 and 4.9 μM, respectively. The metal-free Cys-Gly-His-Lys peptide also stimulates TLN activity, with an apparent binding affinity of 2.2 μM. Coordination of copper through deprotonated imine nitrogens, the histidyl nitrogen, and the free N-terminal amino group is consistent with the characteristic absorption spectrum of a Cu²⁺–amino-terminal copper and nickel binding motif (λ _max ∼ 525 nm). The lack of thiol coordination is suggested by both the absence of a thiol to Cu²⁺ charge transfer band and electrochemical studies, since the electrode potential (vs. Ag/AgCl) 0.84 V (ΔE = 92 mV) for the Cu^3+/2+ redox couple obtained for [Cu²⁺•Cys-Gly-His-Lys] was found to be in close agreement with that of a related complex [Cu²⁺•Lys-Gly-His-Lys]⁺ (0.84 V, ΔE = 114 mV). The N-terminal cysteine appears to be available as a zinc-anchoring residue and plays a critical functional role since the [Cu²⁺•Lys-Gly-His-Lys]⁺ homologue exhibits neither stimulation nor inhibition of TLN. Under oxidizing conditions (ascorbate/O₂) the catalyst is shown to mediate the complete irreversible inactivation of TLN at concentrations where enzyme activity would otherwise be stimulated. The observed rate constant for inactivation of TLN activity was determined as k _obs = 7.7 × 10⁻² min⁻¹, yielding a second-order rate constant of (7.7 ± 0.9) × 10⁴ M⁻¹ min⁻¹. Copper peptide mediated generation of reactive oxygen species that subsequently modify active-site residues is the most likely pathway for inactivation of TLN rather than cleavage of the peptide backbone.     

The behaviors of metal ions in the CdTe quantum dots–H2O2 chemiluminescence reaction and its sensing application

The behaviors of 15 kinds of metal ions in the thiol‐capped CdTe quantum dots (QDs)–H₂O₂ chemiluminescence (CL) reaction were investigated in detail. The results showed that Ag⁺, Cu²⁺ and Hg²⁺ could inhibit CdTe QDs and H₂O₂ CL reaction. A novel CL method for the selective determination of Ag⁺, Cu²⁺ and Hg²⁺ was developed, based on their inhibition of the reaction of CdTe QDs and H₂O₂. Under the optimal conditions, good linear relationships were realized between the CL intensity and the logarithm of concentrations of Ag⁺, Cu²⁺ and Hg²⁺. The linear ranges were from 2.0 × 10^?6 to 5.0 × 10^?8 mol L^?1 for Ag⁺, from 5.0 × 10^?6 to 7.0 × 10^?8 mol L^?1 for Cu²⁺ and from 2.0 × 10^?5 to 1.0 × 10^?7 mol L^?1 for Hg²⁺, respectively. The limits of detection (S/N = 3) were 3.0 × 10^?8, 4.0 × 10^?8 and 6.7 × 10^?8 mol L^?1 for Ag⁺, Cu²⁺ and Hg²⁺, respectively. A possible mechanism for the inhibition of CdTe QDs and H₂O₂ CL reaction was also discussed. Copyright © 2009 John Wiley & Sons, Ltd.     

Convergence of Ca<Superscript>2+</Superscript> signaling pathways in adipocytes. The role of <Emphasis Type="Italic">L</Emphasis>-arginine and protein kinase G in generation of transient and periodic Ca<Superscript>2+</Superscript> signals

Experiments on cultured mouse adipocytes (9 days in vitro) using fluorescent microscopy have shown that activation of α₁- and α₂-adrenoceptors by norepinephrine (NE) or α₂-adrenoreceptors by L-arginine evokes transient Ca²⁺ signals, while activation of m₃-cholinoreceptors by acetylcholine (ACh) or betaine causes sustained or damped Ca²⁺ oscillations. The presence in the incubation medium of L-arginine at a low concentration (100–200 μM) is necessary for a vigorous manifestation of these effects, apparently due to transition of protein kinase G (PKG) and phosphodiesterase V into an active state. In the presence of 1–10 mM L-arginine, the amplitude of the Ca²⁺ transient response to NE increases and signal duration decreases. ACh and NE upon a sequential addition mutually potentiate their effects. Using an inhibitory analysis we show that the observed modes are related to the operation of a signaling pathway with the participation of phosphatidylinositol 3-kinase (PI3K), protein kinase B (PKB), endothelial NO synthase (eNOS), cytoplasmic guanylate cyclase (sGC), protein kinase G (PKG), ADP-ribosyl cyclase (CD38), and the ryanodine receptor (RyR). The formation of several loops of positive feedbacks (PF) and negative feedbacks (NF) in the signaling system is possible: (i) short PF loops due to Ca²⁺-induced Ca²⁺ release (CICR) from internal stores through the inositol trisphosphate receptor (IP₃R) and RyR participating in the transient signal formation; (ii) long PF loop Ca²⁺ → eNOS → sGC → PKG → CD38 → RyR → Ca²⁺, which can provide necessary conditions for calcium oscillations arising from short PF loops (CICR); (iii) several NF loops based on PKG-mediated inhibition of IP₃R and activation of Ca²⁺-ATPases of sarco(endo)plasmic reticulum and of the plasma membrane providing a shutdown of signaling by the pathway phospholipase C → IP₃R → Ca²⁺ and limiting Ca²⁺ rise caused by the pathway PI3K → PKB → eNOS → sGC → PKG → CD38 → RyR → Ca²⁺. Convergence of signaling pathways that involve α₁-, α₂-, and m₃-receptors and then Gβγ-subunits of G_q and G_q proteins acting on PI3Kγ can provide activation of cytoplasmic PKG, which plays a key role in producing transient responses, in activation of Ca²⁺ removal and generation of [Ca²⁺]_i oscillations. PKG inhibition (implemented here by KT5823 application) in the presence of any agonist results in rupture of NF loops controlling Ca²⁺ transporting systems activity that leads to uncontrolled [Ca²⁺]_i rise and cell death.     

Salinity and Copper-Induced Oxidative Damage and Changes in the Antioxidative Defence Systems of Anabaena doliolum

Summary This study provides first-hand information on the salinity and copper-induced oxidative damage and its protection in Anabaena doliolum by the antioxidant defence system. Oxidative damage measured in terms of lipid peroxidation, electrolyte leakage and H₂O₂ production was induced by different concentrations of NaCl and Cu²⁺. A greater electrolyte leakage by NaCl than Cu²⁺ supported the hypothesis of salinity being more injurious than copper. To explore the survival strategies of A. doliolum under NaCl and Cu stress, enzymatic antioxidant activities e.g. superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), and glutathione reductase (GR) and nonenzymatic antioxidant contents such as glutathione reduced (GSH), ascorbate, α-tocopherol, and carotenoid were measured. A general induction in SOD and APX activities as well as ascorbate and α-tocopherol contents was found under NaCl and Cu²⁺ stress. In contrast to this, an appreciable decline in GR activity, GSH pool and carotenoid content under Cu²⁺ and an increase under NaCl stress were observed. CAT activity was completely inhibited at high doses of NaCl but stimulated following Cu²⁺ treatment. The above results suggest the involvement of APX and CAT in the scavenging of H₂O₂ under Cu²⁺ stress. In contrast to this, only APX was involved in H₂O₂ scavenging under salt stress. Our postulate of Cu²⁺-mediated antagonism of salt stress can be explained by a conceivable reversion of Na⁺-induced disturbance of cellular homeostasis by redox active Cu²⁺.     

Acetyl-CoA decarbonylase/synthase complex from Archaeoglobus fulgidus

The acetyl-CoA decarbonylase/synthase (ACDS) multienzyme complex catalyzes the reversible cleavage and synthesis of acetyl-CoA in methanogens. This report of the enzyme complex in Archaeoglobus fulgidus demonstrates the existence of a functional ACDS complex in an organism that is not a methanogen. The A. fulgidus enzyme complex contained five subunits of 89, 72, 50, 49.5, and 18.5 kDa, and it catalyzed the overall synthesis of acetyl-CoA according to the following reaction: w CO₂ + 2 Fd_red(Fe²⁺) + 2 H⁺ + CH₃– H₄SPt + CoA ⇌ acetyl-CoA + H₄SPt + 2 Fd_ox(Fe³⁺) + H₂O where Fd is ferredoxin, and CH₃–H₄SPt and H₄SPt denote N ⁵-methyl-tetrahydrosarcinapterin and tetrahydrosarcinapterin, respectively. Received: 27 October 1997 / Accepted: 29 January 1998     

Horizontal gene transfer of stress resistance genes through plasmid transport

The horizontal gene transfer of plasmid-determined stress tolerance was achieved under lab conditions. Bacterial isolates, Enterobacter cloacae (DGE50) and Escherichia coli (DGE57) were used throughout the study. Samples were collected from contaminated marine water and soil to isolate bacterial strains having tolerance against heavy metals and antimicrobial agents. We have demonstrated plasmid transfer, from Amp⁺Cu⁺Zn⁻ strain (DGE50) to Amp⁻Cu⁻Zn⁺ strain (DGE57), producing Amp⁺Cu⁺Zn⁺ transconjugants (DGE_TC50→57) and Amp⁺Cu⁻Zn⁺ transformants (DGE_TF50→57). DGE57 did not carry any plasmid, therefore, it can be speculated that zinc tolerance gene in DGE57 is located on chromosome. DGE50 was found to carry three plasmids, out of which two were transferred through conjugation into DGE57, and only one was transferred through transformation. Plasmid transferred through transformation was one out of the two transferred through conjugation. Through the results of transformation it was revealed that the genes of copper and ampicillin tolerance in DGE50 were located on separate plasmids, since only ampicillin tolerance genes were transferred through transformation as a result of one plasmid transfer. By showing transfer of plasmids under lab conditions and monitoring retention of respective phenotype via conjugation and transformation, it is very well demonstrated how multiple stress tolerant strains are generated in nature.     

Measurements of the gas temperature in an oxygen plasma by spectroscopy of the O2( b 1Σ g+ ) → O2( X 3Σ g? ) transition

A method for measuring the gas temperature in an oxygen plasma by spectroscopy of the electronic transition from the O₂(b ¹Σ _g ⁺ , v = 0) metastable state of molecular oxygen into the O₂(X ³Σ _g ⁻ , v = 0) ground state is considered in detail. The method is verified experimentally for the plasma of dc glow discharge in pure oxygen. It is shown that the gas temperature can be determined by analyzing high-resolution spectra of the P branch of this transition, no matter whether its fine structure (^P P and ^P Q branches) is resolved or masked, provided that the rotational structure of the spectrum is resolved. The feasibility of the method proposed in 1999 by P. Maco and P. Veis for determining the gas temperature from the ratio between the intensity maxima of the R and P branches of the O₂(b ¹Σ _g ⁺ , v = 0) → O₂(X ³Σ _g ⁻ , v = 0) transition in a poorly resolved spectrum was studied experimentally. It is shown that, in order to use this method, it is necessary to know the spectrograph instrumental function. The effect of the spatial inhomogeneity of the temperature and concentration of O₂(b ¹Σ _g ⁺ ) molecules on the accuracy of integral (over the plasma volume) measurements of the gas temperature is investigated using spatially resolved spectroscopy of the O₂(b ¹Σ _g ⁺ , v = 0) → O₂(X ³Σ _g ⁻ , v = 0) transition. It is shown that precise measurements of the temperature require that the optical measurement system be thoroughly adjusted in order for the temperature and concentration of the emitting particles to vary insignificantly over the optically selected volume. Original Russian Text ? S.M. Zyryanov, D.V. Lopaev, 2007, published in Fizika Plazmy, 2007, Vol. 33, No. 6, pp. 563–574.     

Phenoxyl-copper(II) complexes: models for the active site of galactose oxidase

 The reaction of the macrocycles 1,4,7-tris (3,5-di-tert-butyl-2-hydroxy-benzyl)-1,4,7-triazacyclononane, L¹H₃, or 1,4,7-tris(3-tert-butyl-5-methoxy-2-hydroxy-benzyl)-1,4,7-triazacyclononane, L²H₃, with Cu(ClO₄)₂·6H₂O in methanol (in the presence of Et₃N) affords the green complexes [Cu^II(L¹H)] (1), [Cu^II(L²H)]·CH₃OH (2) and (in the presence of HClO₄) [Cu^II(L¹H₂)](ClO₄) (3) and [Cu^II(L²H₂)] (ClO₄) (4). The Cu^II ions in these complexes are five-coordinate (square-base pyramidal), and each contains a dangling, uncoordinated pendent arm (phenol). Complexes 1 and 2 contain two equatorially coordinated phenolato ligands, whereas in 3 and 4 one of these is protonated, affording a coordinated phenol. Electrochemically, these complexes can be oxidized by one electron, generating the phenoxyl-copper(II) species [Cu^II(L¹H)]^+ ·, [Cu(L²H)]^+ ·, [Cu^II(L¹H₂)]^2+ ·, and [Cu^II(L²H₂)]^2+ ·, all of which are EPR-silent. These species are excellent models for the active form of the enzyme galactose oxidase (GO). Their spectroscopic features (UV-VIS, resonance Raman) are very similar to those reported for GO and unambiguously show that the complexes are phenoxyl-copper(II) rather than phenolato-copper(III) species. Received: 10 February 1997 / Accepted: 7 April 1997