Cu uptake in a cyanobacterium: Fate of selected photochemical reactions

Cu uptake in the diazotrophic cyanobacteriumNostoc calcicola Bréb. was accompanied by inhibitions in the in vivo activities of photosystem (PS) II, PS I,¹⁴CO₂-fixation, and decline in the ATP pool. Cyanobacterial cells, while saturated for Cu uptake within 1 h at 40 M Cu, showed more than 50% inhibition of PS II and 95.4% of¹⁴CO₂ fixation compared with only 15.5% decrease in the PS I activity. The total extractable ATP content also declined by 32.2% within 1 h. In a subsequent follow-up study lasting 72 h, PS II activity and¹⁴CO₂ fixation showed complete inhibition, in contrast to 34.4% of PS I activity and 4.2% of ATP still remaining unaffected. The results have been discussed in the light of multiple effects of Cu during and subsequent to its uptake by the cyanobacterium.     

Nitrogen and phosphorus uptake by the Brazilian kelp Laminaria abyssalis (Phaeophyta) in culture

The uptake of ammonium, nitrate and phosphate by laboratory-grown young sporophytes of Laminaria abyssalis was measured in a perturbed system (batch mode) at 18 °C and 35 ± 5 µE m^–2 s^–1 photon flux density. Uptake of all appeared to follow saturation-type nutrient uptake kinetics. The NO _inf3 ^sup– (K _s = 14.0 µM, V _max = 5.0 µmol h^–1 g^–1 dry wt) and NH _inf4 ^sup+ (K _s = 4.6 µM, V _max= 2.0 µmol h^–1 g^–1 dry wt) were taken up simultaneously, although NH _inf4 ^sup+ was taken up more rapidly. Values of K ₃ and V _max for phosphate were, respectively, 2.21 µM and 0.83 µmol h^–1 g^–1 dry wt. Nitrate and phosphate were both consumed in similar rates (V _max /Ks 0.37) at low concentrations. NH _inf4 ^sup+ , thus, might be a more efficient form of N fertilizer if artificial enrichment of seawater is used.     

The physiological basis of increased biomass partitioning to roots upon nitrogen deprivation in young clonal tea (Camellia sinensis (L.) O. Kuntz)

Deprivation of nitrogen (N) increases assimilate partitioning towards roots at the expense of that to shoots. This study was done to determine the physiological basis of increased root growth of tea (sCammellia sinensis L.) under N shortage. Nine-month-old clonal tea (clone TRI2025) was grown in quartz sand under naturally lit glasshouse conditions. Three levels of N (0, 3.75 and 7.5 mM N) were incorporated in to the nutrient solution and applied daily. Plant growth, photosynthesis, root respiration and plant N contents were measured at 10-day intervals over a 45-day period. Root dry weight showed a sharp increase during the first 15 days after the plants were transferred to 0 mM N, whereas no such increase was shown in plants transferred to 7.5 mM N. In contrast, shoot dry weight increased at 7.5 mM N and was significantly greater than at 0 mM N, where no increase was observed. Due to the above changes, root weight ratio increased and leaf weight ratio decreased during the first 15 days of N deprivation. Leaf photosynthetic rates did not vary between N levels during the initial 15-day period. Thereafter, photosynthetic rates were greater at 7.5 mM and 3.75 mM N than at 0 mM N. Root respiration rate decreased at 0 mM N, whereas it increased at 3.75 and 7.5 mM N, probably because of the greater respiratory cost for nitrate uptake. Root respiratory costs associated with maintenance (R _m) and nitrate uptake (R _u) were calculated to investigate whether the sharp increase of root growth observed upon nitrogen deprivation was solely due to the reduced respiratory costs for nitrate uptake. The estimated values for R _m and R _u were 3.241 × 10^–4 mol CO₂ g^–1 (root dry matter) s^–1 and 0.64 mol CO₂ (mol N)^–1, respectively. Calculations showed that decreased respiratory costs for nitrate uptake could not solely account for the significant increase of root biomass upon N deprivation. Therefore, it is concluded that a significant shift in assimilate partitioning towards roots occurs immediately following N deprivation in tea.     

Abiotic metal stress enhances diosgenin yield in Dioscorea bulbifera L. cultures

Lower concentrations of CuSO₄ (25–75 M) in the MS medium supplemented with 0.1 mg l^–1 IAA+5.0 mg l^–1 Kn+500 mg l^–1 CH+10 mg l^–1 Cyst hyd enhanced the growth of regenerants of Dioscorea bulbifera L. CuSO₄ (75 M) induced an appreciable diosgenin yield in the regenerants compared to those obtained on media without Cu. The presence of Cu thus seems to stimulate diosgenin production. The regenerants also differentiated bulbils on lower concentrations of Cu. At CuSO₄ (100 M), however, cultures showed poor growth as well as a low diosgenin yield. Increased proline and protein contents were recorded in cultures grown on Cu-enriched media.     

Hg2+ uptake in a cyanobacterium

The uptake of Hg²⁺ and its regulation in the cyanobacteriumNostoc calcicola Bréb. was studied. Hg²⁺ uptake pattern consisted of two distinct phases: (a) rapid binding of the cation to the negatively charged cell surface (first 10 min) and (b) its subsequent metabolism-dependent intracellular import, at least up to 40 min (saturating concentration 1.5 M Hg²⁺, K_m=1.0M Hg²⁺ and V_max 0.21 nmol Hg²⁺ mg^–1 protein min^–1). Hg²⁺ influx, to a major extent, depended on photosynthetically generated energy, and the supply of exogenous ATP (10 M) or DCMU (5 M) suggested the vital role of PS II-mediated energy to support the process. The significant lowering in Hg²⁺ uptake rate as well as total cellular Hg²⁺ in the presence ofp-chloromercuribenzoate (pCMB), azide (NaN₃), N,N-dicyclohexycarbodiimide (DCCD), and thiol (mercaptoethanol) indicated the role of membrane potential,-SH groups, and ATP hydrolysis in regulating Hg²⁺ transport. While Cu²⁺ antagonized Hg²⁺ intake, Ni²⁺ showed synergism.     

Microcalorimetric study of magnesium-adenosine triphosphate ternary complex

Using an original microcalorimetric method, the existence of the Mg₂ATP ternary chelate has been studied. The thermodynamic parameters of this complex are H=7.2±0.5 kJ mole^–1 andK=49±9 M^–1. These values are compared with those previously obtained for binary chelate Mg ATP^2–. A possible regulation role of Mg₂ATP is discussed.     

Role of energy in oxidative phosphorylation

This article reviews the current status of information regarding the role of energy in the process of oxidative phosphorylation by mitochondria. The available data suggest that in submitochondrial particles (SMP) energy is utilized for the binding of ADP and P_i and for the release of ATP bound at the catalytic sites of F₁-ATPase. The process of ATP synthesis on the surface of F₁ from F₁-bound ADP and P_i appears to be associated with negligible free energy change. The rate of energy production by the respiratory chain modulates the kinetics of ATP synthesis between a lowK _m (for ADP and P_i)-lowV _max mode and a highK _m -highV _max mode. TheK _m extremes for ADP are 2–3 µM and 120–150 µM, andV _max for ATP synthesis at high rates of energy production by bovine-heart SMP is about 440 s^–1 (mole F₁)^–1 at 30°C, which corresponds to 11 µmol ATP (min · mg of protein)^–1. The interaction of dicyclohexylcarbodiimide (DCCD) or oligomycin at the proteolipid (subunitc) of the membrane sector (F₀) of the ATP synthase complex alters the mode of ATP binding at the catalytic sites of F₁, probably to one of lower affinity. It has been suggested that protonic energy might be conveyed to the catalytic sites of F₁ in an analogous manner, i.e., via conformation changes in the ATP synthase complex initiated by proton-induced alterations in the structure of the DCCD-binding proteolipid. Finally, the relationship between the steady-state membrane potential () and the rates of electron transfer and ATP synthesis has been discussed. It has been shown, in agreement with the delocalized chemiosmotic mechanism, that under appropriate conditions is exquisitely sensitive to changes in the rates of energy production and consumption.     

Nitrate deposition in northern hardwood forests and the nitrogen metabolism of Acer saccharum marsh

It is generally assumed that plant assimilation constitutes the major sink for anthropogenic Nitrate NO ₃ ^– deposited in temperate forests because plant growth is usually limited by nitrogen (N) availability. Nevertheless, plants are known to vary widely in their capacity for NO ₃ ^– uptake and assimilation, and few studies have directly measured these parameters for overstory trees. Using a combination of field and greenhouse experiments, we studied the N nutrition of Acer saccharum Marsh. in four northern hardwood forests receiving experimental NO ₃ ^– additions equivalent to 30 kg N ha^–1 year^–1. We measured leaf and fine-root nitrate reductase activity (NRA) of overstory trees using an in vivo assay and used ¹⁵N to determine the kinetic parameters of NO ₃ ^– uptake by excised fine roots. In two greenhouse experiments, we measured leaf and root NRA in A. saccharum seedlings fertilized with 0–3.5 g NO ₃ ^– –N m^–2 and determined the kinetic parameters of NO ₃ ^– and NH ₄ ⁺ uptake in excised roots of seedlings. In both overstory trees and seedlings, rates of leaf and fine root NRA were substantially lower than previously reported rates for most woody plants and showed no response to NO ₃ ^– fertilization (range = non-detectable to 33 nmol NO ₂ ^– g^–1 h^–1). Maximal rates of NO ₃ ^– uptake in overstory trees also were low, ranging from 0.2 to 1.0 mol g^–1 h^–1. In seedlings, the mean V _max for NO ₃ ^– uptake in fine roots (1 mol g^–1 h^–1) was approximately 30 times lower than the V _max for NH ₄ ⁺ uptake (33 mol g^–1 h^–1). Our results suggest that A. saccharum satisfies its N demand through rapid NH ₄ ⁺ uptake and may have a limited capacity to serve as a direct sink for atmospheric additions of NO ₃ ^– .     

Ammonium uptake by Chondrus crispus Stackhouse (Gigartinales,Rhodophyta) in culture

Cultivated Chondrus crispus was used in N-NH₄ uptake experiments in the laboratory. An elevation of temperature increased the apparent rate of uptake, especially up to 11 °C. Uptake in the dark was found to be 83 % of that in the light. The apparent uptake decreased with increasing internal N pool; rates were 26.5, 22.2 and 20.2 µg N g dry wt^–1 min^–1 for internal N pools of 2.7, 3.5 and 4.6%, respectively. Apparent uptake increased with the substrate N concentration. The resulting curve has two components: an active uptake and a diffusion component at high (> 5000 µg N L^–1) external N levels. Ks and V _max were calculated by deducting the diffusion component from the uptake curve: these were of 497 µg N L ^–1 and 14.4 µg N g dry wt^–1 min^–1. respectively, and reflect a low substrate affinity. This could be the result of 10 years of continuous culture of C. crispus. Uptake was similarly followed in the culture tanks and showed comparable results; nighttime would be the most appropriate time to supply nutrients.     

Assessment of Hg2+ toxicity to a N2-fixing cyanobacterium in long- and short-term experiments

Toxicological responses of the filamentous N₂-fixing cyanobacteriumNostoc calcicola Bréb. towards Hg²⁺ were studied to enumerate the decisive lethal events. In low-dose, long-term experiments (0.05–0.25 m Hg²⁺, 10 days), photoautotrophic growth was severely inhibited with concurrent loss of photosynthetic pigments (phycocyanin>chlorophyll >carotenoids) and nucleic acids. The termination of growth after a day 4 exposure to 0.25 m Hg²⁺ has been attributed to the complete inhibition ofin vivo photosynthetic activity in the cyanobacterium (O₂ evolution>¹⁴CO₂ incorporation). The elevated Hg²⁺ concentrations irreversibly damaged the cell membrance as observed under light microscopy, and as indicated by the leakage of intracellular electrolytes and phycocyanin. In high-dose, short-term experiments (0.5–20.0 m Hg²⁺, up to 6 h), thein vivo activities of selected enzymes (glutamine synthetase > nitrate reductase > nitrogenase) were less inhibited by Hg²⁺ than the uptake of nutrient ions (NH ₄ ⁺ >NO ₃ ^– >PO ₄ ^3– ).     

Mechanisms of cytoplasmic calcium signalling in cerebellar bergmann glial cells

Mechanisms underlying intracellular calcium signals in Bergmann glial cells evoked by various neurotransmitters were investigated in experiments on cerebellar slices acutely isolated from 30-day-old mice. [Ca²⁺] _in values were measured by means of a Ca²⁺-sensitive fluorescent probe fura-2. Extracellular application of ATP (10–100 µM), histamine (10–100 µM), or noradrenaline (or adrenaline, 0.1–10.0 µM) caused a temporary increase in cytoplasmic Ca²⁺ concentrations. The effect persisted in Ca²⁺-free extracellular solution and was blocked with thapsigargin (500 nM) or a specific blocker of the inositol-1,4,5-trisphosphate-sensitive intracellular channels heparin. Based on the pharmacological analysis, we postulate the involvement of P₂ purinoreceptors, ₁-adrenoreceptors, and H₁ histamine receptors in an agonist-activated increase in [Ca²⁺] _in in Bergmann glia. Thus, ATP, monoamines, or histamine induce calcium signal generation in Bergmann glial cells via activation of Ca²⁺ release from the inositol-1,4,5-trisphosphate-sensitive internal stores.Neirofiziologiya/Neurophysiology, Vol. 26, No. 6, pp. 417–419, November–December, 1994.     

Comparison of plant uptake and plant toxicity of various ions in wheat

The effects of varying solution concentrations of manganese (Mn), zinc (Zn), copper (Cu), boron (B), iron (Fe), gallium (Ga) and lanthanum (La) on plant chemical concentrations, plant uptake and plant toxicity were determined in wheat (Triticum aestivum L.) grown in a low ionic strength (2.7×10^–3 M solution culture). Increasing the solution concentration of Mn, Zn, Cu, B, Fe, Ga and La increased plant concentrations of that ion. Asymptotic maximum plant concentrations were reached for Zn (10 mg kg DM^–1 in the roots), Ga (2 mg kg DM^–1 in the tops and 18 mg kg DM^–1 in the roots) and La (0.4 mg kg DM^–1 in the tops and 4 mg kg DM^–1 in the roots). Plant ion concentrations were, on average, 3 times higher in the roots than the tops for Mn and Zn, 7 times for Cu, 9 times for Fe, 12 times for Ga and 15 times for La. In contrast, B concentrations were higher in the tops than the roots by, on average, 2 times. The estimated toxicity threshold (plant concentration at which a rapid decrease in yield occurred) in the tops was 0.4 mg g DM^–1 for B, 2 for Zn, 0.075 for Cu and 0.09 for La and in the roots 0.2 mg g DM^–1 for B, 5 for Zn, 0.3 for Cu and 3 for La. Plant uptake rates of the ions (as estimated by the slope of the relationship between solution ion concentrations and plant ion concentrations) was in the order B 250 mg kg DM^–1 M ^–1). Plant toxicity was estimated as the reciprocal of the plant concentration that reduced yield by 50% (change in relative yield per mg ion kg DM^–1). The plant toxicity of the ions tested was in the order Mn相似文献   

Ulva rigida (Ulvales,Chlorophyta) tank culture as biofilters for dissolved inorganic nitrogen from fishpond effluents

Ulva rigida was cultivated in 7501 tanks at different densities with direct and continuous inflow (at 2, 4, 8 and 12 volumes d^–1) of the effluents from a commercial marine fishpond (40 metric tonnes, Tm, of Sparus aurata, water exchange rate of 16 m³ Tm^–1) in order to assess the maximum and optimum dissolved inorganic nitrogen (DIN) uptake rate and the annual stability of the Ulva tank biofiltering system. Maximum yields (40 g DW m^–2 d^–1) were obtained at a density of 2.5 g FW 1^–1 and at a DIN inflow rate of 1.7 g DIN m^–2 d^–1. Maximum DIN uptake rates were obtained during summer (2.2 g DIN M^–2 d^–1), and minimum in winter (1.1 g DIN m^–2 d^–1) with a yearly average DIN uptake rate of 1.77 g DIN m^–2 d^–1 At yearly average DIN removal efficiency (2.0 g DIN m^–2 d^–1, if winter period is excluded), 153 m² of Ulva tank surface would be needed to recover 100% of the DIN produced by 1 Tm of fish.Abbreviations DIN= dissolved inorganic nitrogen (NH _inf4 ^sup+ + NO _inf3 ^sup– + NO _inf2 ^sup– ); - FW= fresh weight; - DW= dry weight; - PFD= photon flux density; - V= DIN uptake rate     

Outdoor pond cultivation of the subtropical marine red algaGracilaria tenuistipitata in brackish water in Sweden. Growth,nutrient uptake,co-cultivation with rainbow trout and epiphyte control

Pond cultivation of the subtropical, euryhaline macroscopic red algaGracilaria tenuisipitata var.liui Zhanget Xia was carried out in brackish seawater (6–7) in the Gryt archipelago on the east coast of Sweden, using four outdoor tanks of 30–40 m³. Growth rate and nutrient uptake in batch culture were measured with the aim of estimating the water purification capacity ofG. tenuisipitata in outdoor conditions. Its ability to withstand epiphytic infections was also studied. An average growth rate of 4 biomass increase per day was recorded during two seasons with a maximum growth rate of 9 d^–1. The initial biomass was usually 1 kgFW m^–3 (FW, fresh weight). The nutrient uptake capacity was on average ca. 1 g N_i kgFW^–1 d^–1 and 0.08 g P_i kgFW^–1 d^–1 and the uptake rates for NH₄ ⁺-N were higher than those for NO₃ ^–-N. Both the growth rate and the nutrient uptake rate were highest at the highest water temperature. Co-cultivation with rainbow trout (Oncorhynchus mykiss) was tested: with trout fodder as the only nutrient inputG. tenuistipitata could grow and maintain low levels of N_i and P_i with optimum efficiency at a trout: alga ratio of 1:1 (w:w). Epiphytic growth of filamentous green and brown algae was limited, probably as a result of the high pH values caused by inorganic carbon uptake byG. tenuistipitata. The growth ofEnteromorpha intestinalis, the only significant epiphyte, was completely inhibited and the majority of plants died by a few days treatment with 100 µg 1^–1 Cu²⁺, a concentration that did not severely affectG. tenuistipitata. We conclude thatG. tenuistipitata can be cultivated in outdoor ponds in southern Sweden during 5–6 months of the year using aerated or unaerated batch cultures and that wastewater from trout cultivation may be used as a nutrient source, resulting in purification with respect to N and P.     

The effects of nitrite on chloride regulation in the crayfish Pacifastacus leniusculus Dana (Crustacea: Decapoda)

Summary Nitrite in the external freshwater medium was found to be toxic to Pacifastacus leniusculus Dana (48 h LC₅₀0.7 mM NO ₂ ^– ). It produced significant changes in haemolymph ionic concentration and acid-base status. Exposure to 1.0 mM NO ₂ ^– resulted in a rapid, active accumulation of nitrite in the haemolymph (to 25 mM NO ₂ ^– after 24 h) and caused the partial inhibition of Cl^– uptake. Some reduction in Cl^– efflux rate was seen. In 1.0 mM NO ₂ ^– a rapid depletion of haemolymph [Cl^–] was observed (50 mM decrease in 27 h). Nitrite competitively inhibited active Cl^– uptake (K_m increased from 0.42 to 1.22 mM; K_i=0.45 mM). To achieve Cl^– balance in this medium, depleted crayfish would require a two-fold increase in external [Cl^–]. A lesser decrease in haemolymph [Na⁺] was found while osmotic pressure was relatively unaffected. Haemolymph [HCO ₃ ^– ] showed a significant increase and was accompanied, unexpectedly, by an acidosis. Possible sources of the excess HCO ₃ ^– , perhaps by inhibition of normal Cl^–/HCO ₃ ^– branchial exchange or release from CaCO₃ stores, are discussed. Haemolymph clearance of NO ₂ ^– was slower than uptake as was the restoration of [Cl^–] on recovery in nitrite-free medium.Abbreviations AFWM artificial freshwater medium - BOD biochemical oxygen demand - J _out ^Cl chloride efflux - J _in ^Cl chloride influx - J _in ^Cl chloride influx - J _net ^base net base flux - J _net ^base net base flux - J _in(p) ^Cl passive chloride influx - J _out efflux - LC ₅₀ median lethal concentration - NEDE N-1-Naphthylethylenediamine - SEM standard error of mean - TEP transepithelial potential difference - V _in ^Cl active chloride uptake     

The effect of carbon dioxide on the growth kinetics of fructose-limited chemostat cultures of Acetobacterium woodii DSM 1030

The growth of the anaerobic acetogenic bacterium Acetobacterium woodii DSM 1030 was investigated in fructose-limited chemostat cultures. A defined medium was developed which contained fructose, mineral salts, cysteine · HCl and Ca pantothenate (1 mg · 1^–1) supplied in a vitamin supplement. Growth at high dilution rates was dependent on the presence of CO₂ in the gas phase. The ^max was found to be 0.16 h^–1 and the fructose maintenance requirement was 0.1 to 0.13 mmol fructose · (g dry wt)^–1 · h^–1. A growth yield of 61 g dry wt · (mol fructose)^–1, corrected for the cell maintenance requirement and for incorporation of fructose carbon into cell biomass, was determined from the fructose consumption. A corresponding growth yield of 69 g dry wt · (mol fructose)^–1 was calculated from the acetate production assuming that fructose fermentation was homoacetogenic. A Y_ATP of 12.2 to 13.8 g dry wt · (mol ATP)^–1 was calculated from these growth yields using a value of 5 mol ATP · (mol fructose)^–1 as an estimate of the amount of ATP synthesised from fructose fermentation. The addition of yeast extract (0.5 g · 1^–1) to the medium did not influence the ^max or cell yield. After prolonged growth under fructose-limited conditions the requirement of the culture for CO₂ in the gas phase was reduced.Abbreviations YE yeast extract - IC inorganic carbon - D fermenter dilution rate : h^–1 - M_X maintenance requirement for X: mmol X · (g dry wt)^–1 · h^–1 - X may be fructose (Fruct), fructose consumed in energy metabolism (Fruct [E]), acetate (Ac) - ATP CO₂, NH _inf4 ^sup+ or Pi - qX specific rate of utilisation or consumption of X: mmol X · (g dry wt)^–1 · h^–1 - V fermenter volume: litre - rC · Cell, fermenter cell carbon production: mmol C · h^–1 - Y_X yield of cells on X: g dry wt · (mol X)^–1 - Y _infx ^supmax the yield corrected for cell maintenance: g dry wt · (mol X)^–1 - S_ATP stoichiometry of ATP synthesis from fructose: mol ATP · (mol frucose)^–1 - x cell concentration: g dry wt · 1^–1 - specific growth rate : h^–1 - ^max maximum specific growth rate: h^–1     

pH modulation of the kinetics of rabbit jejunal,brush-border folate transport

Summary In jejunal brush-border membrane vesicles, an outwardly directed OH^– gradient (in>out) stimulates DIDS-sensitive, saturable folate (F) uptake (Schron, C.M. 1985.J. Clin. Invest. 76:2030–2033), suggesting carrier-mediated folate: OH^– exchange (or phenomenologically indistinguishable H⁺: folate cotransport). In the present study, the precise role of pH in the transport process was elucidated by examining F uptake at varying pH. For pH gradients of identical magnitude, F uptake (0.1 M) was greater at lower (pH_int/pH_ext: 5.5/4.5) compared with higher (6.5/5.5) pH ranges. In the absence of a pH gradient, internal Ftrans stimulated DIDS-sensitive³H-folate uptake only at pH6.0. Since stepwise increments ininternal pH (4.57.5; pH_ext=4.5) stimulated F uptake, an inhibitory effect of higherinternal pH was excluded. In contrast, with increasing external pH (4.356.5; pH_int=7.8), a 50-fold decrement in F uptake was observed (H⁺ K _m =12.8±1.2 M). Hill plots of these data suggest involvement of at least one H⁺ (OH^–) at low pH (monovalent F^– predominates) and at least 2 H⁺ (OH^–) at high pH (divalent F^–2 predominates). Since an inside-negative electrical potential did not affect F uptake at either pH_ext 4.55 or 5.8, transport of F^– and F^–2 is electroneutral. Kinetic parameters for F^– and F^–2 were calculated from uptake data at pH_ext 4.55 and 5.0. Comparison of predictedvs. experimentally determined kinetic parameters at pH_ext5.8 (K _m =1.33vs. 1.70 M;V _max=123.8vs. 58.0 pmol/mg prot min) suggest that increasing external pH lowers theV _max, but does not affect theK _m for carrier-mediated F transport. These data are consistent with similarK _i ^' s for sulfasalazine (competitive inhibitor) at pH_ext 5.35 and 5.8 (64.7 and 58.5 M, respectively). In summary, the jejunal F carrier mediates electroneutral transport of mono- and divalent F and is sensitive to external pH with a H⁺ K _m (or OH^– lC₅₀) corresponding to pH 4.89. External pH effects theV _max, but not theK _m for carriermediated F uptake suggesting a reaction mechanism involving a ternary complex between the outward-facing conformation of the carrier and the transported ions (F and either OH^– or H⁺),rather than competitive binding that is mutually exclusive.     

Mercury (Hg2+) and zinc (Zn2+): Two divalent cations with different actions on voltage-activated calcium channel currents

Summary 1. We examined the actions of mercury (Hg²⁺) and zinc (Zn²⁺) on voltage-activated calcium channel currents of cultured rat dorsal root ganglion (DRG) neurons, using the whole-cell patch clamp technique.2. Micromolar concentrations of both cations reduced voltage-activated calcium channel currents. Calcium channel currents elicited by voltage jumps from a holding potential of –80 to 0 mV (mainly L- and N-currents) were reduced by Hg²⁺ and Zn²⁺. The threshold concentration for Hg²⁺ effects was 0.1 µM and that for Zn²⁺ was 10µM. Voltage-activated calcium channel currents were abolished (>80%) with 5µM Hg²⁺ or 200µM Zn²⁺. The peak calcium current was reduced to 50% (IC₅₀) by 1.1µM Hg²⁺ or 69µM Zn²⁺. While Zn²⁺ was much more effective in reducing the T-type calcium channel current—activated by jumping from –80 to –35 mV—Hg²⁺ showed some increased effectiveness in reducing this current.3. The effects of both cations occurred rapidly and a steady state was reached within 1–3 min. While the action of Zn²⁺ was not dependent on an open channel state, Hg²⁺ effects depended partially on channel activation.4. While both metal cations reduced the calcium channel currents over the whole voltage range, some charge screening effects were detected with Hg²⁺ and with higher concentrations (>100µM) of Zn²⁺.5. As Zn²⁺ in the concentration range used had no influence on resting membrane currents, Hg²⁺ caused a clear inward current at concentrations µM.6. In the present study we discuss whether the actions of both metals on voltage-activated calcium channel currents are mediated through the same binding site and how they may be related to their neurotoxic effects.     

Effect of light intensity and ammonium-N on carotenogenesis ofTrentepohlia odorata andDunaliella bardawil

AxenicTrentepohlia odorata was cultured at three different NH₄Cl levels (3.5 × 10^–2, 3.5 × 10^–3, 3.5 × 10^–4 M) and three different light intensities (48, 76, 122 µmol m^–2 s^–1). Chloride had no effect on growth over this range of concentration. High light intensity and high NH₄Cl concentration enhanced the specific growth rate. The carotenoid content increased under a combination of high light intensity and low N concentration. WhenD. bardawil was exposed to the same combination of growth conditions, there was an increase in its carotenoid content. The light saturation and the light inhibition constants (K _s andK _i, respectively) for growth, and the saturation constant (K _m) for NH₄Cl were determined. TheK _s andK _i values were higher inT. odorata (66.7 and> 122 mol m^–2 s^–1, respectively) than inD. bardawil (5.1 and 14.7 µmol m^–2 s^–1, respectively). TheK _m value determined at 122 µmol m^–2 s^–1, however, was lower inT. odorata (0.048 µM) than inD. bardawil (0.062 µM).Author for correspondence     

Spectroscopic and binding studies of azide to type-2-copper-depleted ascorbate oxidase from zucchini

Summary Binding of azide to type-2-copper-depleted (T2D) zucchini ascorbate oxidase, containing reduced type-3 Cu centers, and met-T2D ascorbate oxidase, containing oxidized type-3 Cu centers, has been studied spectroscopically. In both cases titration with azide in 0.1 M phosphate pH 6.8 produces a broad near-ultraviolet band with maximum at 455 nm (e 2500 M^–1 cm^–1, with respect to the met-T2D enzyme) and shoulder at 390 nm (e 1700 M^–1 cm^–1), that are assigned to(azide)Cu(II) ligand-to-metal charge transfer (LMCT) transitions. This is accompanied by a reduction of absorbance at 330 nm in the met-T2D) enzyme adduct (e –1400 M^–1 cm^–1). A broad circular dichroic band of negative sign between 370–480 nm corresponds to the LMCT absorption band. Analysis of the titration data indicates that one azide ion binds independently to each of the binuclear T3 Cu couples with low affinity (K = 50 M^–1). The ESR signal of the T1 Cu observed in frozen solutions of the T2D enzyme is also perturbed by the addition of azide. The analogies in the azide-binding characteristics between ascorbate oxidase and laccase are discussed.