Esterases in mycobacteria

Esterases ofMycobacterium phlei (acetic ester acetyl hydrolase E.C.3.1.6 and carboxylic esterhydrolase E.C.3.1.1.1.) obtained after separation on Sephadex G-100 can be temporarily, for a short time interval, activated by adding calcium ions. The activation of esterases isolated from cells was non-repeteable, whereas the temporary activation of esterases from the culture filtrate could be repeated by increasing concentrations of calcium ions. However, the value of activation gradually decreased. Similarly with calcium ions strontium ions were also effective, however, higher concentrations were required and the activation was non-repeatable. Magnesium ions were practically without any effect. Possible mechanisms of the temporary activation of esterases ofMycobacterium phlei are discussed.     

A role of G-proteins and calcium in light-regulated primary leaf formation in Sorghum bicolor

Primary leaf development of Sorghum bicolor is a phytochrome-mediated response. Primary leaves are not produced in Sorghum seedlings even after 10 d of germination if grown in darkness. However, 5 min irradiation with white light or red light given to 5 d etiolated seedlings resulted in the formation of etiolated leaves. This effect of red light was reversed by far-red light. When calcium (3-5 mM) was added exogenously, complete leaf formation was obtained in darkness; however, the kinetics of the response was slower than that seen with light irradiation. This effect was also obtained with potassium ions but magnesium ions had no effect. Light- and calcium-mediated leaf development could be arrested at the stage of leaf emergence or leaf expansion by the addition of inhibitors of G-proteins or by calcium channel blockers suggesting a role of G-proteins and calcium in phytochrome signal transduction during primary leaf development.Key words: Leaf formation, G-proteins, calcium, potassium, Sorghum bicolor.      

Effect of Calcium and Zinc on the Activity and Thermostability of Superoxide Dismutase

The effect of calcium and zinc ions on superoxide dismutase (SOD) from four plant species (Taxus baccata, Pinus sylvestris, Medicago rigidula, and Zea mays) was followed at three temperatures: optimal (20 °C), increased (50 °C), and high, inhibiting temperature (70 – 80 °C). At 20 and 50 °C in vitro added calcium increases SOD activity, but the degree was different for the plants investigated. The effect of zinc ions at the same temperatures varied in the investigated plants from activation to inhibition. An inhibiting effect of high temperature on SOD activity was diminished in the presence of calcium or zinc ions. It was shown that calcium and zinc ions can increase activity and thermostabilize different SOD isoforms.     

Effect of Calcium Ions on Hydrodynamic Properties of Pentameric and Decameric C-Reactive Protein in Solution

The molecular mass and sedimentation coefficient of native C-reactive protein in solution were determined by analytical ultracentrifugation in the presence and absence of calcium ions. Pentameric C-reactive protein was shown to be the major macroscopic form of this protein in solution. The removal of calcium ions from solution caused decompaction of the protein accompanied by changes in its hydrodynamic parameters. The sedimentation coefficient s ⁰ _20,w of pentameric C-reactive protein in solution containing 2 mM Ca²⁺ (6.6S) exceeded that for C-reactive protein in solution containing 2 mM EDTA (6.4S). Analysis of average molecular masses M _w and M _z obtained from sedimentation data demonstrated that the solution of highly purified protein was not homogeneous. As shown by intermolecular crosslinking, the solution also contained the 241-kDa decamer of C-reactive protein (9.5S) as a separate macroscopic form, whose share hardly reached 10% in the presence of 2 mM Ca²⁺ and increased after removal of calcium ions. The decamers were shown to result from intermolecular association of the pentamers.     

Influence of Echinostoma paraensei (Lie and Basch, 1967) infection on the calcium content in Biomphalaria glabrata (Say, 1818)

The calcium content in the hemolymph and shell of Biomphalaria glabrata (Say, 1818) was determined after exposure to different parasite burdens (5 and 50 miracidia) of Echinostoma paraensei (Lie and Basch, 1967). The snails were dissected 1, 2, 3, and 4 weeks after infection to collect the hemolymph and shell. An increase in calcemia was observed in snails infected with both miracidial doses. A significant decrease in the calcium ions in the shell was observed, coinciding with the calcemia peak in the hemolymph. This indicates greater mobilization of calcium between the shell and hemolymph to regulate the calcium content in the body when the snail is exposed to stress conditions, as has also been observed in some other infected snail species. The results obtained indicate that in this model, the calcium metabolism depends on the miracidial dose used.     

Utilization in alginate beads for Cu(II) and Ni(II) adsorption of an exopolysaccharide produced by <Emphasis Type="Italic">Chryseomonas luteola</Emphasis> TEM05

Copper and nickel adsorption onto calcium alginate, sodium alginate with an extracellular polysaccharide (EPS) produced by the activated sludge bacterium Chryseomonas luteola TEM05 and the immobilized C. luteola TEM05 from aqueous solutions were studied. After that, the multi metal ions containing these ions together were prepared and partial competitive adsorptions of these mixtures were also investigated. The metal adsorption of gel beads were carried out at pH 6.0, 25 °C. The maximum adsorption capacities in Langmuir isotherm for calcium alginate, calcium alginate + EPS, calcium alginate + C. luteola TEM05 and calcium alginate + EPS + C. luteola TEM05 were 1.505, 1.989, 1.976, 1.937 mmol/g dry weight for Cu(II) and 0.996, 1.224, 1.078, 1.219 mol/g dry weight for Ni(II), respectively.The competitive biosorption capacities of the carrier for all metal ions were lower than single conditions.     

Further Investigations on the Nature of the Heat Resistance of Thermophilic Bacteria

When cells of Bacillus stearothermophilus, strain NCA 1503, were grown in tryptone starch broth and subsequently transferred to tris buffer, a fraction of the cells: rapidly died in ttie buffer. This fraction increased with increasing content of calcium chloride in the growth medium. The' addition of sodium, potassium or magnesium chloride to the growth medium had no such effect. The rapid dying of the cells in tris buffer was associated with a leakage of organic material and calcium ions from the cells. The results obtained are probably caused by a damage to the osmotic barrier of the cells during their contact with the buffer. Observations: made during the present investigation and a previous one (Ljunger 1970) indicate that the heat resistance of thermophilic bacteria depends on the maintenance of a high intracellular concentration of free calcium ions.     

DNA double-strand breaks in mammalian cells exposed to Á-rays and very heavy ions

The spatial distribution of DNA double-strand breaks (DSB) was assessed after treatment of mammalian cells (V79) with densely ionizing radiation. Cells were exposed to beams of heavy charged particles (calcium ions: 6.9 MeV/u, 2.1⋅10³ keV/μm; uranium ions: 9.0 MeV/u, 1.4⋅10⁴ keV/μm) at the linear accelerator UNILAC of GSI, Darmstadt. DNA was isolated in agarose plugs and subjected to pulsed-field gel electrophoresis under conditions that separated DNA fragments of size 50 kbp to 5 Mbp. The measured fragment distributions were compared to those obtained after γ-irradiation and were analyzed by means of a convolution and a deconvolution technique. In contrast to the finding for γ-radiation, the distributions produced by heavy ions do not correspond to the random breakage model. Their marked overdispersion and the observed excess of short fragments reflect spatial clustering of DSB that extends over large regions of the DNA, up to several mega base pairs (Mbp). At fluences of 0.75 and 1.5/μm², calcium ions produce nearly the same shape of fragment spectrum, merely with a difference in the amount of DNA entering the gel; this suggests that the DNA is fragmented by individual calcium ions. At a fluence of 0.8/μm² uranium ions produce a profile that is shifted to smaller fragment sizes in comparison to the profile obtained at a fluence of 0.4/μm²; this suggests cumulative action of two separate ions in the formation of fragments. These observations are not consistent with the expectation that the uranium ions, with their much larger LET, should be more likely to produce single particle action than the calcium ions. However, a consideration of the greater lateral extension of the tracks of the faster uranium ions explains the observed differences; it suggests that the DNA is closely coiled so that even DNA locations several Mbp apart are usually not separated by less than 0.1 or 0.2 μm. Received: 27 January 1998 / Accepted in revised form: 15 April 1998     

The effect of calcium alginate entrapment on the physiology of Mycobacterium sp. strain E3

The influence of calcium alginate entrapment on the physiology of Mycobacterium sp. E3 is reported. As a model system the NADH-requiring conversion of propene to 1,2-epoxypropane in the presence and absence of glucose as co-substrate was selected. The co-factor-dependent reaction was used as a measure of the physiological status of the resting cells. Initial kinetic experiments established a system free from diffusional limitations. In the presence of glucose there were no differences between the physiology of the free and immobilized cells. The apparent differences observed in the absence of co-substrate were demonstrated to be caused by calcium ions and to a lesser degree alginate; the addition of calcium, alginate or calcium alginate beads containing no cells to the free cells gave similar data to that obtained with immobilized cells. The results presented highlight the high concentrations of calcium to which cells immobilized in calcium alginate beads can be exposed. Correspondence to: M. R. Smith     

Optimization of Bifidobacterium longum growth by use of calcium carbonate-alginate beads

High concentrations of ammonium and sodium ions inhibited Bifidobacterium longum growth more than a high calcium ion concentration. The optimal pH for B. longum growth was determined to be 5.0 due to the lower accumulation of ammonium ion. To reduce the accumulation of ammonium ion and obtain an enhanced growth of B. longum, the pH of the culture containing immobilized calcium carbonate beads was controlled to 5.0 with ammonia water. The concentrations of ammonium, sodium, and calcium ions in the culture were maintained at the desired level. The maximum cell mass increased to 16.8 g/l, 1.23 times higher than cultures without calcium carbonate beads. The number of viable cells in the culture increased to 5.0 × 10¹⁰, 1.67 times more than cultures without calcium carbonate beads.     

INFLUENCE OF IONIC CONDITIONS ON CELL DIFFERENTIATION AND MORPHOGENESIS OF THE CELLULAR SLIME MOLDS

Effects of various ionic conditions on the development of the cellular slime molds D. discoideum and D. mucoroides were studied. A certain concentration of lithium ions (7 mM) promoted differentiation of the stalk cells and conversely inhibited formation of the spores. The presence of calcium and magnesium ions was needed for Li to manifest its specific effect. A high concentration of Ca (100-120 mM) also facilitated differentiation of the stalk cells. On the other hand, fluoride ions stimulated considerable formation of spores at 15 mM. In the absence of divalent cations, sodium ions inhibited morphogenesis and cell differentiation proportionately with its concentration, and complete inhibition was obtained at 20 mM. The inhibitory effect of Na was nullified by addition of small amounts of Ca. Possible mechanisms by which these ions exert their influences on development of this organism were discussed.     

Lanthanide ions are agonists of transient gene expression in rice protoplasts and act in synergy with ABA to increase Em gene expression

Previous work has shown that in rice suspension cells, NaCl at 0.4 M can induce Em gene expression and act synergistically with ABA, possibly by potentiating the ABA response pathway through a rate-limiting intermediate (R.M. Bostock and R.S. Quatrano (1992) Plant Physiol., 98, 1356–1363). Since calcium is an intermediate in ABA regulation of stomatal closure, we tested the effect of calcium changes on ABA-inducible Em gene expression in transiently transformed rice protoplasts. We show that calcium is required for ABA-inducible Em-GUS expression and can act in synergy with ABA. The trivalent ions lanthanum, gadolinium, and aluminum, which are known to interact with calcium- and other signaling pathways, can act at sub-millimolar concentrations to increase GUS reporter gene expression driven by several promoters in transiently transformed rice protoplasts. This effect is not specific for the ABA-inducible Em promoter, but is synergistic with ABA. The lanthanum synergy with ABA does not require calcium. In rice suspension cells, lanthanum alone does not induce Em gene expression, in contrast to transiently transformed protoplasts, yet can act synergistically with ABA to effectively increase the sensitivity to ABA greater than tenfold. Trivalent ions may be a useful tool to study the regulation of gene expression. The possible effects of trivalent ions on ABA signal transduction and gene expression are discussed.     

Degradation of myelin basic protein by calcium-activated neutral protease (CANP) in human brain and inhibition by E-64 analogue

A calcium-activated neutral protease (CANP) was extracted from human brain and partially purified. The activity was measured using alkali-denatured casein (Hammersten) as a substrate. The optimum pH was around 7.0. The activity required the presence of calcium ions, maximum activity was obtained with over 5 mM calcium ions. TheK _m for the casein concentration was about 1.62 mg/ml. The activity of CANP was inhibited by one of the thiol protease inhibitors, E-64 analogue (E-64-a). The rate of inhibition was about 50% at an E-64-a concentration of 10^–5M. This CANP degraded selectively basic protein in myelin proteins and the degradation was inhibited by E-64-a or EGTA. The role of the brain CANP in the process of demyelination was suggested by this study.     

On the Nature of the Heat Stabilizing Effect of Inorganic Ions on Escherichia coli

The growth lag of Escherichia coli at 45°C was reduced by the addition of sodium, potassium, magnesium and calcium ions to the growth medium. A method to quantitatively determine the lag-reducing effect of these ions was developed. The results obtained showed that equivalent amounts of the ions produced the same reduction of the growth lag. According to the results of plasmolysis experiments cells of E. coli suspended in peptone broth were permeable to all four ions. The course of plasmolysis and subsequent deplasmolysis was registered as changes in the cells' ability to scatter light. The heat stability of catalase from E. coli was increased by addition of the four ions. This was observed in experiments with intact cells and with a crude cell-free preparation of catalase. The results of our experiments are most easily explained by assuming a stabilizing effect of the ions tested on the intracellular bacterial proteins.     

The ionic dependence of voltage-activated inward currents in the pharyngeal muscle of Caenorhabditis elegans

The pharynx of Caenorhabditis elegans consists of a syncytium of radially orientated muscle cells that contract synchronously and rhythmically to ingest and crush bacteria and pump them into the intestine of the animal. The action potentials that support this activity are superficially similar to vertebrate cardiac action potentials in appearance with a long, calcium-dependent plateau phase. Although the pharyngeal muscle can generate action potentials in the absence of external calcium ions, action potentials are absent when sodium is removed from the extracellullar solution (Franks et al. 2002). Here we have used whole cell patch clamp recordings from the pharynx and show low voltage-activated inward currents that are present in zero external calcium and reduced in zero external sodium ions. Whilst the lack of effect of zero calcium when sodium ions are present is not surprising in view of the known permeability of voltage-gated calcium channels to sodium ions, the reduction in current in zero sodium when calcium ions are present is harder to explain in terms of a conventional voltage-gated calcium channel. Inward currents were also recorded from egl-19 (n582) which has a loss of function mutation in the pharyngeal L-type calcium channel and these were also markedly reduced in zero external sodium. Despite this apparent dependence on external sodium ions, the current was partially blocked by the divalent cations, cadmium, barium and nickel. Using single-channel recordings we identified a cation channel for which the open-time duration was increased by depolarisation. In inside-out patches, the single-channel conductance was highest in symmetrical sodium solution. Further studies are required to determine the contribution of these channels to the pharyngeal action potential.     

Citric acid fermentation by the mutant strain of theAspergillus niger resistant to manganese ions inhibition

Summary A new mutant strain,Aspergillus niger GS-III, showing resistance to manganese ions inhibition of citric acid fermentation on a sugarcane molasses containing medium was induced fromAspergillus niger KCU 520, a high citric acid-yielding strain. In submerged, surface or continuous cultures in the presence of manganese ions concentration upto 1.5 ppm the mutant strain yielded citric acid about 90 KgM^–3 . The citric acid yield was comparable to that obtained with the parental strain KCU 520 in the absence of manganese ions, but it was atleast 3-fold higher than that obtained by the latter in the presence of manganese ions. The mutant strain immobilized in calcium alginate beads was used in combination with surface-stabilized cultures for about 36-days in a continuous flow horizontal fermenter without any apparent loss in citric acid productivity. These results indicate that the manganese-resistant mutant is stable and may be used in the presence of sufficient manganese ions concentration (1.5 ppm) in the fermentation medium. This capability of the mutant strainA. niger GS-III has been correlated with greatly reduced levels (about one-thirds) of the NADP⁺ -isocitric dehydrogenase, one of the control points for citric acid accumulation.     

Effects of Coexistence of β-Casein on Gelation of κ-Carrageenan

The effects of caseins on the rheological properties of κ-carrageenan-calcium gel was investigated by measuring the gel breaking strength. The existence of β-casein in the system promoted the gelation of κ-carrageenan in the presence of calcium ion. Beta-casein increased the strength of calcium gels of κ-carrageenan with increasing NaCl concentration up to 80 mM and strengthened the κ-carrageenan-calcium gel at neutral pH. The values obtained from the slopes of the logarithmic plots of the gel strength versus concentration were 2.15 for κ-carrageenan gel and 2.27 for a β-casein-κcarrageenan mixture gel, suggesting that β-casein may participate in the gelation of κ-carrageenan through the mediation of calcium ions.     

Investigation of the TEA-resistant outward current in the somatic membrane of perfused nerve cells

Outward currents remaining after addition of 20–50 mM of tetraethylammonium (TEA) ions to the extracellular or intracellular solution, were investigated in perfused isolatedHelix neurons. After this addition, the inactivated inward current carried by potassium ions, the potential-dependent and kinetic characteristics of which differ from those of potassium outward currents suppressed by TEA, is preserved in the membrane. A component dependent on the inward calcium current was found in this TEA-resistant outward current; it was abolished by replacement of the extra-cellular calcium ions by magnesium ions, by blocking of the calcium channels by extracellular cadmium ions, and by their destruction by intracellular fluoride ions. Increasing the intracellular concentration of free calcium ions by perfusing the cell with solutions containing calcium-EGTA buffer potentiated the TEA-resistant component of the outward current, whereas removal of these ions with EGTA weakened it. It is concluded that a system of outward current channels whose activation depends on the presence of calcium ions near the inner surface of the membrane is present in the somatic membrane. It is suggested that to keep these channels capable of being activated, calcium ions must bind with the structures forming their internal opening.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Translated from Neirofiziologiya, Vol. 11, No. 5, pp. 460–468, September–October, 1979.     

Calcium,Calcium Oxalate Crystals,and Leaf Differentiation in the Common Bean (Phaseolus vulgaris L.)*

Young plants of Phaseolus vulgaris were grown in nutrient solutions at different levels of calcium concentration. When the calcium concentration was low more palisade parenchyma and less extended bundle sheath was formed at the adaxial side of minor veins of the leaves as compared to leaves of plants grown with higher calcium supply. The number of calcium oxalate crystals in the bundle sheath extensions was positively correlated to the amount of calcium fed to the plants. The ion induces additional cell divisions in the bundle sheath extensions. A high supply of calcium leads to the formation of a second type of crystal in the bundle sheath.