Yersiniabactin metal binding characterization and removal of nickel from industrial wastewater

Yersiniabactin (Ybt) is a metal‐binding natural product that has been re‐purposed for water treatment. The early focus of this study was the characterization of metal binding breadth attributed to Ybt. Using LC‐MS analysis of water samples exposed to aqueous and surface‐localized Ybt, quantitative assessment of binding was completed with metals that included Pd²⁺, Mg²⁺, and Zn²⁺. In total, Ybt showed affinity for 10 metals. Next, Ybt‐modified XAD‐16N resin (Ybt‐XAD) was utilized to quantify the affinity for metal removal, showing a rank order of Fe³⁺ > Ga³⁺ > Ni²⁺ > Cu²⁺ > Cr²⁺≈Zn²⁺ > Co²⁺ > Pd²⁺ > Mg²⁺ > Al³⁺, and in the applied treatment of wastewater from a local precious metal plating company, showing selective removal of nickel from the aqueous effluent. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1548–1554, 2017     

Metal ion binding to anticoagulation factor II from the venom of <Emphasis Type="Italic">Agkistrodon acutus</Emphasis>: stabilization of the structure and regulation of the binding affinity to activated coagulation factor X

Anticoagulation factor II (ACF II) isolated from the venom of Agkistrodon acutus is an activated coagulation factor X (FXa)-binding protein with both anticoagulant and hypotensive activities. The thermodynamics of the binding of alkaline earth metal ions to ACF II and their effects on the stability of ACF II and the binding of ACF II to FXa were investigated by isothermal titration calorimetry, fluorescence, differential scanning calorimetry, and surface plasmon resonance. The binding of ACF II to FXa does not have an absolute requirement for Ca²⁺. Mg²⁺, Sr²⁺, and Ba²⁺ can induce the binding of ACF II to FXa. The radii of the cations bound in ACF II crucially affect the binding affinity of ACF II for cations and the structural stability of ACF II against guanidine hydrochloride and thermal denaturation, whereas the radii of cations bound in FXa markedly affect the binding affinity between ACF II and FXa. The binding affinities of ACF II for cations and the capacities of metal-induced stabilization of ACF II follow the same trend: Ca²⁺ > Sr²⁺ > Ba²⁺. The metal-induced binding affinities of ACF II for FXa follow the trend Mg²⁺ > Ca²⁺ > Sr²⁺ > Ba²⁺. Although Mg²⁺ shows significantly low binding affinity with ACF II, Mg²⁺ is the most effective to induce the binding of ACF II with FXa. Our observations suggest that in blood the bindings of Ca²⁺ in two sites of ACF II increase the structural stability of ACF II, but these bindings are not essential for the binding of ACF II with FXa, and that the binding of Mg²⁺ and Ca²⁺ to FXa may be essential for the recognition between FXa and ACF II. Like Ca²⁺, the abundant Mg²⁺ in blood also plays an important role in the anticoagulation of ACF II.     

Removing heavy metals from synthetic effluents using “kamikaze” Saccharomyces cerevisiae cells

One key step of the bioremediation processes designed to clean up heavy metal contaminated environments is growing resistant cells that accumulate the heavy metals to ensure better removal through a combination of biosorption and continuous metabolic uptake after physical adsorption. Saccharomyces cerevisiae cells can easily act as cation biosorbents, but isolation of mutants that are both hyperaccumulating and tolerant to heavy metals proved extremely difficult. Instead, mutants that are hypersensitive to heavy metals due to increased and continuous uptake from the environment were considered, aiming to use such mutants to reduce the heavy metal content of contaminated waters. In this study, the heavy metal hypersensitive yeast strain pmr1∆ was investigated for the ability to remove Mn²⁺, Cu²⁺, Co²⁺, or Cd²⁺ from synthetic effluents. Due to increased metal accumulation, the mutant strain was more efficient than the wild-type in removing Mn²⁺, Cu²⁺, or Co²⁺ from synthetic effluents containing 1–2 mM cations, with a selectivity $ {\text{Mn}}^{{{\text{2}} + }} > {\text{Co}}^{{{\text{2}} + }} ~ > {\text{Cu}}^{{{\text{2}} + }} $ {\text{Mn}}^{{{\text{2}} + }} > {\text{Co}}^{{{\text{2}} + }} ~ > {\text{Cu}}^{{{\text{2}} + }} and also in removing Mn²⁺ and Cd²⁺ from synthetic effluents containing 20–50 μM cations, with a selectivity Mn²⁺ > Cd²⁺.     

Biotreatment of acidic zinc- and copper-containing wastewater using ethanol-fed sulfidogenic anaerobic baffled reactor

The treatment of acidic (pH 6.5–3), sulfate- (2–3 g/L), Zn- and Cu- (total metal 0–500 mg/L) containing wastewater was studied in a four-stage anaerobic baffled reactor (ABR) at 35 °C for 250 days. Ethanol was supplemented (COD/SO₄ ²⁻ = 0.67) as carbon and electron source for sulfate reducing bacteria. Sulfate reduction, COD oxidation and metal precipitation efficiencies were 70–92, 80–94 and >99%, respectively. The alkalinity produced from sulfidogenic ethanol oxidation increased the wastewater pH from 3.0 to 7.0–8.0. The electron flow from organic oxidation to sulfate averaged 87%. Decreasing feed pH to 3 and increasing total metal concentrations to 500 mg/L did not adversely affect the performance of ABR and sufficient alkalinity was produced to increase the effluent pH to neutral values. More than 99% of metals were precipitated in the form of metal-sulfides. Accumulation of precipitated metals in the first compartment allowed metal recovery without disturbing reactor performance seriously.     

Effect of lactoferrin on oxidative features of ceruloplasmin

In our previous report we first described a complex between lactoferrin (Lf) and ceruloplasmin (Cp) with K _d ~ 1.8 μM. The presence of this complex in colostrum that never contains more than 0.3 μM Cp questions the reliability of K _d value. We carefully studied Lf binding to Cp and investigated the enzymatic activity of the latter in the presence of Lf, which allowed obtaining a new value for K _d of Cp–Lf complex. Lf interacting with Cp changes its oxidizing activity with various substrates, such as Fe²⁺, o-dianisidine (o-DA), p-phenylenediamine (p-PD) and dihydroxyphenylalanine (DOPA). The presence of at least two binding sites for Lf in Cp molecule is deduced from comparison of substrates’ oxidation kinetics with and without Lf. When Lf binds to the first site affinity of Cp to Fe²⁺ and to o-DA increases, but it decreases towards DOPA and remains unchanged towards p-PD. Oxidation rate of Fe²⁺ grows, while that of o-DA, p-PD and DOPA goes down. Subsequent Lf binding to the second center has no effect on iron oxidation, hampers DOPA and o-DA oxidation, and reduces affinity towards p-PD. Scatchard plot for Lf sorbing to Cp-Sepharose allowed estimating K _d for Lf binding to high-affinity (~13.4 nM) and low-affinity (~211 nM) sites. The observed effect of Lf on ferroxidase activity of Cp is likely to have physiological implications.     

Synthesis and characterization of glucosyl-curcuminoids as Fe3+ suppliers in the treatment of iron deficiency

The Fe³⁺ chelating ability of some curcumin glucosyl derivatives (Glc-H; Glc-OH; Glc-OCH₃) is tested by means of UV and NMR study. The pK _a values of the ligands and the overall stability constants of Fe³⁺ and Ga³⁺ complexes are evaluated from UV spectra. The only metal binding site of the ligand is the β-diketo moiety in the keto-enolic form; the glucosyl moiety does not interact with metal ion but it contributes to the stability of metal/ligand 1:2 complexes by means of hydrophilic interactions. These glucosyl derivatives are able to bind Fe³⁺ in a wide pH rage, forming complex species thermodynamically more stable than those of other ligands commonly used in the treatment of iron deficiency. In addition they demonstrate to have a poor affinity for competitive biological metal ions such as Ca²⁺. All ligands and their iron complexes have a good lypophilicity (log P > −0.7) suggesting an efficient gastrointestinal absorption in view of their possible use as iron supplements in oral therapy. The ligand molecules are also tested for their antioxidant properties in “ex vivo” biological system.     

Biochemical characterization of Helicobacter pylori α-1,4 fucosyltransferase: metal ion requirement, donor substrate specificity and organic solvent stability

The effect of metal ions on the activity, the donor substrate specificity, and the stability in organic solvents of Helicobacter pylori α-1,4 fucosyltransferase were studied. The recombinant enzyme was expressed as soluble form in E. coli strain AD494 and purified in a one step affinity chromatography. Its activity was highest in cacodylate buffer at pH 6.5 in the presence of 20 mM Mn²⁺ ions at 37°C. Mn²⁺ ions could be substituted by other metal ions. In all cases, Mn²⁺ ions proofed to be the most effective (Mn²⁺ > Co²⁺ > Ca²⁺ > Mg²⁺ > Cu²⁺ > Ni²⁺ > EDTA). The enzyme shows substrate specificity for Type I disaccharide (1) with a K _M of 114 μM. In addition, the H. pylori α-1,4 fucosyltransferase efficiently transfers GDP-activated l-fucose derivatives to Galβ1-3GlcNAc-OR (1). Interestingly, the presence of organic solvents such as DMSO and methanol up to 20% in the reaction medium does not affect significantly the enzyme activity. However, at the same concentration of dioxane, activity is totally abolished.     

65Zn2+ transport by isolated gill epithelial cells of the American lobster, Homarus americanus

Gills are the first site of impact by metal ions in contaminated waters. Work on whole gill cells and metal uptake has not been reported before in crustaceans. In this study, gill filaments of the American lobster, Homarus americanus, were dissociated in physiological saline and separated into several cell types on a 30, 40, 50, and 80% sucrose gradient. Cells from each sucrose solution were separately resuspended in physiological saline and incubated in ⁶⁵Zn²⁺ in order to assess the nature of metal uptake by each cell type. Characteristics of zinc accumulation by each kind of cell were investigated in the presence and absence of 10 mM calcium, variable NaCl concentrations and pH values, and 100 μM verapamil, nifedipine, and the calcium ionophore A23187. ⁶⁵Zn²⁺ influxes were hyperbolic functions of zinc concentration (1–1,000 μM) and followed Michaelis–Menten kinetics. Calcium reduced both apparent zinc binding affinity (K _m) and maximal transport velocity (J _max) for 30% sucrose cells, but doubled the apparent maximal transport velocity for 80% sucrose cells. Results suggest that calcium, sodium, and protons enter gill epithelial cells by an endogenous broad-specificity cation channel and trans-stimulate metal uptake by a plasma membrane carrier system. Differences in zinc transport observed between gill epithelial cell types appear related to apparent affinity differences of the transporters in each kind of cell. Low affinity cells from 30% sucrose were inhibited by calcium, while high affinity cells from 80% sucrose were stimulated. ⁶⁵Zn²⁺ transport was also studied by isolated, intact, gill filament tips. These intact gill fragments generally displayed the same transport properties as did cells from 80% sucrose and provided support for metal uptake processes being an apical phenomenon. A working model for zinc transport by lobster gill cells is presented.     

Efficient Zn and Pb uptake by filamentous fungus Paecilomyces marquandii with engagement of metal hydrocarbonates precipitation

Zinc and lead biosorption by living non-growing filamentous fungus Paecilomyces marquandii was examined for its potential application in heavy metals elimination from contaminated areas. Metal uptake by the studied fungus was pH dependent and reached the level of 308 mg of Zn²⁺ g⁻¹ and 505 mg of Pb²⁺ g⁻¹ at pH of 7.5 caused by microprecipitation in slightly alkaline environment. All other metal studies were cultivated with unregulated pH yielding the maximum of 186.2 mg of Zn²⁺ g⁻¹ and 305.8 mg of Pb²⁺ g⁻¹. Interestingly, zinc binding by mycelium increased intensively after 15 h of incubation, whereas the lead concentration in biomass extended gradually and proportionally to the initial concentration and the time of contact. The study showed that thermal pretreatment of mycelium led to a decline in metal uptake, especially in the case of zinc. The mycelium slightly digested by the cell wall lytic enzyme complex, could adsorb lead twice as well after 2 h of exposure whereas zinc loading did not differ from the metal uptake by mycelia without any digestion procedure. The release of potassium ions from the mycelium, concomitant with lead uptake was observed suggesting ion exchange participation in lead binding. Energy-dispersive X-ray analysis, X-ray diffraction and FTIR spectroscopy revealed the presence of both metals hydrocarbonates on the mycelium surface. Additionally, the contribution of carboxyl and amide groups, originating from the mycelium, in metal binding was confirmed by FTIR analysis.The obtained results suggest that the effective metals uptake by P. marquandii was due to a combined mechanism with a dominant role of metabolism dependent microprecipitation.     

Combination of chemometrically assisted voltammetry, calorimetry, and circular dichroism as a new method for the study of bioinorganic substances: application to selenocystine metal complexes

Selenium-containing compounds play an important role in antioxidant defense systems, binding to toxic metals, preventing their uptake into cells, and thus protecting cells from metal-induced formation of reactive oxygen species. Here, we present a proposal for a relatively new method as a complement to the more usual methods used in selenium studies. A systematic study of the metal-binding properties of selenocystine (SeCyst) in the presence of divalent metal cations (Cd, Co, Hg, Ni, and Zn) is reported. Isothermal titration calorimetry provides thermodynamic parameters of the systems. Titrations produced curves that could be fit reasonably well to the one set of sites model. The data clearly demonstrate that one M²⁺ binds one SeCyst molecule, and the stable M(SeCyst) complex is formed under these conditions. The order of the SeCyst binding constant for the metal ions is Hg²⁺ > Cd²⁺ ~ Zn²⁺ > Ni²⁺> Co²⁺. Cadmium ion was selected as a modulator for the behavior of SeCyst in the presence of a nonessential metal, and zinc was selected for the case of an essential element. These interactions of SeCyst with Cd²⁺ and Zn²⁺, either individually or combined, were studied in aqueous buffered solutions at physiological pH by differential pulse polarography and circular dichroism spectroscopy. Furthermore, recently developed chemometric tools were applied to differential pulse polarography data obtained in mixtures of SeCyst and glutathione in the presence of Cd²⁺ at physiological pH.     

Luminescent sensor for Cd2+, Hg2+ and Ag+ in water based on a sulphur‐containing receptor: quantitative binding–softness relationship

A water‐soluble, high‐output fluorescent sensor, based on a lumazine ligand with a thiophene substituent for Cd²⁺, Hg²⁺ and Ag⁺ metal ions, is reported. The sensor displays fluorescence enhancement upon Cd²⁺ binding (log  β = 2.79 ± 0.08) and fluorescence quenching by chelating with Ag⁺ and Hg²⁺ (log β = 4.31 ± 0.15 and 5.42 ± 0.1, respectively). The mechanism of quenching is static and occurs by formation of a ground‐state non‐fluorescent complex followed by rapid intersystem crossing. The value of the Stern–Volmer quenching rate constant (k_q) by Ag⁺ ions is close to 6.71 × 10¹² mol/L/s at 298 K. The thermodynamic parameters (ΔG, ΔH and ΔS) were also evaluated and indicated that the complexation process is spontaneous, exothermic and entropically favourable. The quantitative linear relationship between the softness values of Klopman (σ_K) or Ahrland (σ_A) and the experimental binding constants (β) being in the order of Hg²⁺ > Ag⁺ > Cd²⁺ suggests that soft–soft interactions are the key for the observed sensitivity and selectivity in the presence of other metal ions, such as: Pb²⁺, Ni²⁺, Mn²⁺, Cu²⁺, Co²⁺, Zn²⁺ and Mg²⁺ ions. Copyright © 2008 John Wiley & Sons, Ltd.     

Characterization of a metal resistant Pseudomonas sp. isolated from uranium mine for its potential in heavy metal (Ni2+, Co2+, Cu2+, and Cd2+) sequestration

Heavy metal sequestration by a multimetal resistant Pseudomonas strain isolated from a uranium mine was characterized for its potential application in metal bioremediation. 16S rRNA gene analysis revealed phylogenetic relatedness of this isolate to Pseudomonas fluorescens. Metal uptake by this bacterium was monophasic, fast saturating, concentration and pH dependent with maximum loading of 1048 nmol Ni²⁺ followed by 845 nmol Co²⁺, 828 nmol Cu²⁺ and 700 nmol Cd²⁺ mg^?1 dry wt. Preferential metal deposition in cell envelope was confirmed by TEM and cell fractionation. FTIR spectroscopy and EDX analysis revealed a major role of carboxyl and phosphoryl groups along with a possible ion exchange mechanism in cation binding. Binary system demonstrated selective metal binding affinity in the order of Cu²⁺ > Ni²⁺ > Co²⁺ > Cd²⁺. A comparison with similar metal uptake reports considering live bacteria strongly indicated the superiority of this strain in metal sequestration, which could be useful for developing efficient metal removal system.     

Metal dependent hydrolysis of β‐casein by sIgA antibodies from human milk

We present the first evidence that electrophoretically and immunologically homogeneous sIgAs purified from milk of healthy human mothers by chromatography on Protein A‐Sepharose and FPLC gel filtration contain intrinsically bound metal ions (Ca > Mg ≥ Al > Fe ≈ Zn ≥ Ni ≥ Cu ≥ Mn), the removal of which by a dialysis against ethylenediamine tetraacetic acid (EDTA) leads to a significant decrease in the β‐casein‐hydrolyzing activity of these antibodies (Abs). An affinity chromatography of total sIgAs on benzamidine‐Sepharose interacting with canonical serine proteases separates a small metalloprotease sIgA fraction (6.8 ± 2.4%) from the main part of these Abs with a serine protease‐like β‐casein‐hydrolyzing activity. The relative activity of this metalloprotease sIgA fraction containing intrinsically bound metal ions increases ～1.2–1.9‐fold after addition of external metal ions (Mg²⁺ > Fe²⁺ > Cu²⁺ ≥ Ca²⁺ ≥ Mn²⁺) but decreases by 85 ± 7% after the removal of the intrinsically bound metals. The metalloprotease sIgA fraction free of intrinsic metal ions demonstrates a high β‐casein‐hydrolyzing activity in the presence of individual external metal ions (Fe²⁺ > Ca²⁺ > Co²⁺ ≥ Ni²⁺) and especially several combinations of metals: Co²⁺ + Ca²⁺ < Mg²⁺ + Ca²⁺ < Ca²⁺ + Zn²⁺ < Fe²⁺ + Zn²⁺ < Fe²⁺ + Co²⁺ < Fe²⁺ + Ca²⁺. The patterns of hydrolysis of a 22‐mer oligopeptide corresponding to one of sIgA‐dependent specific cleavage sites in β‐casein depend significantly on the metal used. Metal‐dependent sIgAs demonstrate an extreme diversity in their affinity for casein‐Sepharose and chelating Sepharose, and interact with Sepharoses bearing immobilized monoclonal mouse IgGs against λ‐ and κ‐type light chains of human Abs. Possible ways of the production of metalloprotease abzymes (Abz) by human immune system are discussed. Copyright © 2010 John Wiley & Sons, Ltd.     

Cu,Zn-superoxide dismutase is differently regulated by cadmium and lead in roots of soybean seedlings

The cadmium (Cd²⁺) and lead (Pb²⁺)-induced changes in Cu,Zn-SOD gene expression on the level of mRNA accumulation and enzyme activity were analyzed in roots of soybean (Glycine max) seedlings. The Cd²⁺ caused the induction of copper–zinc superoxide dismutase (Cu,Zn-SOD) mRNA accumulation, at each analyzed metal concentration (5–25 mg/l), whereas in Pb²⁺-treated roots this effect was observed only at the medium metal concentrations (50–100 mg/l of Pb²⁺). The analysis of Cu,Zn-SOD activity proved an increase in enzyme activity during Cd²⁺/Pb²⁺ stresses, however in Pb²⁺-treated plants the activity of enzyme was not correlated with respective mRNAs level. Presented data suggest that different metals may act on various level of Cu,Zn-SOD expression in plants exposed to heavy metals stress.     

Interaction of Pb and Cd with gum kondagogu (Cochlospermum gossypium): A natural carbohydrate polymer with biosorbent properties

Gum kondagogu (Cochlospermum gossypium), an exudates tree gum from India was explored for its potential to decontaminate toxic metals (Pb²⁺ and Cd²⁺). Optimum biosorption of metals were determined by investigating the contact time, pH, initial concentration of metal ions and biosorbent dose at 25 ± 2 °C. The maximum metal biosorption capacity for gum kondagogu was observed for Pb²⁺ (48.52 mg g⁻¹) and Cd²⁺ (47.48 mg g⁻¹) as calculated by Langmuir isotherm model. Kinetic studies showed that the biosorption rates could be described by pseudo-second-order expression. The metal interactions with biopolymer were assessed by FT-IR, SEM–EDXA and XPS analysis. Results based on these techniques suggest that mechanism of metal binding by the biopolymer involves micro-precipitation, ion-exchange and metal complexation.     

Gating of aquaporins by heavy metals in <Emphasis Type="Italic">Allium cepa</Emphasis> L. epidermal cells

Changes in the water permeability, aquaporin (AQP) activity, of leaf cells were investigated in response to different heavy metals (Zn²⁺, Pb²⁺, Cd²⁺, Hg²⁺). The cell pressure probe experiments were performed on onion epidermal cells as a model system. Heavy metal solutions at different concentrations (0.05 μM–2 mM) were used in our experiments. We showed that the investigated metal ions can be arranged in order of decreasing toxicity (expressed as a decrease in water permeability) as follows: Hg>Cd>Pb>Zn. Our results showed that β-mercaptoethanol treatment (10 mM solution) partially reverses the effect of AQP gating. The magnitude of this reverse differed depending on the metal and its concentration. The time course studies of the process showed that the gating of AQPs occurred within the first 10 min after the application of a metal. We also showed that after 20–40 min from the onset of metal treatment, the water flow through AQPs stabilized and remained constant. We observed that irrespective of the metal applied, the effect of AQP gating can be recorded within the first 10 min after the administration of metal ions. More generally, our results indicate that the toxic effects of investigated metal ions on the cellular level may involve AQP gating.     

Kinetic and thermodynamic properties of wall bound invertase in heat tolerant and susceptible cultivars of wheat

We have identified two types of invertases, one bound ionically and the other covalently to the particulate fraction in grains of heat tolerant C 306 and heat susceptible WH 542 cultivars of wheat (Triticum aestivum L.). The cell walls contained a high level of invertase activity, of which 79.2–72.8% was extractable by 2 M NaCl and 14.9–21.1% by 0.5% EDTA in C 306 and WH 542, respectively. The NaCl-released invertase constituted the predominant fraction. Using 5–100 mM sucrose and pH range of 4.0–7.0, the apparent Michaelis constant (K _m, enzyme substrate affinity measure) of enzyme ranged from 5.73 to 16.06 mM for C 306 and from 6.08 to 19.86 mM for WH 542. The V _max (maximum catalytic rate) values at these pH were higher in C 306 (0.63–11.04 μg sucrose hydrolysed min⁻¹) than WH 542 (0.51–8.73 μg sucrose hydrolysed min⁻¹). By employing photo-oxidation and by studying the effect of pH on K _m and V _max, the involvement of histidine and α-carboxyl groups at the active site of the enzyme was indicated. The two cultivars also showed differential response in terms of thermodynamic properties of the enzyme i.e. energy of activation (E _a), enthalpy change (ΔH) and entropy change (ΔS). NaCl-released invertase showed differential response to metal ions in two cultivars suggesting their distinctive nature. Mn²⁺, Cu²⁺, Hg²⁺, Mg²⁺, Zn²⁺ and Cd²⁺ were strong inhibitors in WH 542 as compared to C 306 while K⁺, Ca²⁺ were stimulators in both the cultivars. Overall the results suggest that genetic differences exist in wall bound invertase properties of wheat grains as evident in its altered kinetic behaviour.     

Factors modulating alkaline phosphatase activity in the diazotrophic rice-field cyanobacterium, Anabaena oryzae

Summary Alkaline phosphatases (APase), both phosphomonoesterase (PMEase) and phosphodiesterase (PDEase) were studied in the cyanobacterium Anabaena oryzae for their specific requirements of temperature, pH, micro- and macronutrients and their activities in the presence of salinity and heavy metal stress. The alkaline phosphatases (PMEase and PDEase) are quite stable enzymes and require a narrow range of pH (pH 10–10.2) and temperature (35–40 °C) for their optimal activity.A pH of 10, 10.2 and 10.2 supported optimal activity of cellular PMEase, cellular PDEase and extracellular PMEase, respectively, whereas temperatures of 35, 38 and 40 °C were required for their optimal activity. The requirement for Ca²⁺ and Mg²⁺ as macronutrients and the significance of the micronutrients Zn²⁺, Co²⁺, Fe²⁺, Mn²⁺ and Cu²⁺ in APase activity in the cyanobacterium suggests nutritional regulation of enzyme activity in A. oryzae. The metals Pb²⁺, Cr⁶⁺ and Ni²⁺ severely inhibited APase activity, whereas the NaCl stress had a dual role, which was concentration dependent. NaCl stress at lower concentrations (≤20 mM) caused an increase in cellular PMEase activity while its higher concentration (>20 mM) favoured release of the extracellular PMEase. The decrease in cellular activity and an increase in extracellular activity suggest that the higher concentrations of salt stimulate the release of the enzyme.The data suggest that the cyanobacterium A. oryzae possess a potential application as biofertilizer in high salinity and alkaline (Ca²⁺-rich) soils because of its ability to release PO₄³⁻ enzymatically under these conditions.     

Copper and Zinc Mediated Oligomerisation of Aβ Peptides

The accumulation of senile plaques composed primarily of aggregated amyloid β-peptide (Aβ), is the major characteristic of Alzheimer’s disease. Many studies correlate plaque accumulation and the presence of metal ions, particularly copper and zinc. The metal binding sites of the amyloid Aβ peptide of Alzheimer’s disease are located in the N-terminal region of the full-length peptide. In this work, the interactions with metals of a model peptide comprising the first 16 amino acid residues of the amyloid Aβ peptide, Aβ(1–16), were studied. The effect of Cu²⁺ and Zn²⁺ binding to Aβ(1–16) on peptide structure and oligomerisation are reported. The results of ESI-MS, gel filtration chromatography and NMR spectroscopy demonstrated formation of oligomeric complexes of the peptide in the presence of the metal ions and revealed the stoichiometry of Cu²⁺ and Zn²⁺ binding to Aβ(1–16), with Cu²⁺ showing a higher affinity for binding the peptide than Zn²⁺.     

Ion Permeation and Block of P2X2 Purinoceptors: Single Channel Recordings

P2X₂ purinoceptors are cation-selective channels activated by ATP and its analogues. Using single channel measurements we studied the channel's selectivity for the alkali metal ions and organic monovalent cations NMDG⁺, Tris⁺, TMA⁺, and TEA⁺. The selectivity sequence for currents carried by alkali metal ions is: K⁺ > Rb⁺ > Cs⁺ > Na⁺ > Li⁺, which is Eisenman sequence IV. This is different from the mobility sequence of the ions in free solution suggesting there is weak interaction between the ions and the channel interior. The relative conductance for alkali ions increases linearly in relation to the Stokes radius. The organic ions NMDG⁺, Tris⁺, TMA⁺ and TEA⁺ were virtually impermeant. The divalent ions (Mn²⁺, Mg²⁺, Ca²⁺ and Ba²⁺) induced a fast block visible as a reduction in amplitude of the unitary currents. Using a single-site binding model, the divalent ions exhibited an equilibrium affinity sequence of Mn²⁺ > Mg²⁺ > Ca²⁺ > Ba²⁺. Received: 3 May 1999/Revised: 23 August 1999