Protein phosphatase type 2B (calcineurin)-mediated, FK506-sensitive regulation of intracellular ions in yeast is an important determinant for adaptation to high salt stress conditions.

To assess the physiological function of Ca(2+)-dependent protein phosphatase (PP2B) in the yeast Saccharomyces cerevisiae, the phenotypes of PP2B-deficient mutants were investigated. Although PP2B was dispensable for growth under normal conditions, the mutations did, however, cause growth inhibition under certain stress circumstances. The growth of the mutants was inhibited by NaCl and LiCl, but not by KCl, CaCl2, MgCl2 or nonspecific osmotic stresses. Upon shift to high NaCl medium, intracellular Na+ levels of both wild type yeast and the mutants initially increased at a comparable rate. However, internal Na+ in wild type cells started to decline more rapidly than the mutant cells during cultivation in high NaCl medium, indicating that PP2B is important in maintaining a gradient across the membrane. The protection against salt stress was achieved, at least in part, by the stimulation of Na+ export. The maintenance of a high level of internal K+ in high NaCl medium was also PP2B-dependent. In the presence of the immunosuppressant FK506, the growth behaviour and intracellular Na+ and K+ of wild type cells in high NaCl medium became very similar to those of the PP2B-deficient mutant in a manner dependent on the presence of the FK506 binding protein.     

Effect of the preparation method on Na+-H+ exchange and ion permeabilities in rat renal brush-border membranes

The delta pH-dependent quenching of Acridine orange was used to characterize Na+-H+ exchange and K+ and H+ conductances in brush-border membrane vesicles isolated by precipitation with either CaCl2 or MgCl2 from rat kidney cortex. A transmembrane pH difference of 2.5 units (inside acidic) was imposed and the initial rate of its dissipation was followed after injecting a puls of tetramethylammonium gluconate (control) or sodium or potassium gluconate. In membranes isolated by CaCl2, the Na+-H+ exchange was partially electroneutral (45% to 77% of the total exchange) and the rest was due to electrically coupled Na+ and H+ movements through conductive pathways in the membranes. In membranes prepared by MgCl2, the rate of total Na+-H+ exchange was about twice as high as that in membranes obtained by CaCl2 precipitation. However, total and electroneutral exchanges were equal indicating negligible electrically coupled Na+ and H+ movements in these membranes. K0.5 for Na+ in all preparations was in the same range, being in average 30 mM. Amiloride was a competitive inhibitor of Na+-H+ exchange in membranes obtained with both preparations; Ki values ranged between 0.1 and 0.58 mM. The rates of delta pH-dissipation with K+ gradients (+/- valinomycin) were by 50% to 150% higher in membranes prepared with CaCl2 than in membranes isolated with MgCl2 indicating much higher H+ and K+ conductances in membranes obtained with CaCl2. Therefore, the rate of Na+-H+ exchange as well as the conductances for various ions in the isolated brush-border membranes depend on membrane preparation.     

植物单盐毒害和离子拮抗机理研究Ⅰ.单盐和混合盐对植物原生质体膜某些特性、超氧物歧化酶和过氧化氢酶活性的影响

单盐(KCl, CaCl_2或MgCl_2)和混合盐(KC_1+CaCl_2或KCl+MgCl_2)对植物原生质体完整率、存活率和膜透性等均有明显影响。K~+、Ca~(2+)或Mg~(2+)等单种阳离子明显降低原生质体膜完整率和存活率而增加其物质渗漏量,其中以单价阳离子K~+的影响为甚。上述单种阳离子还明显降低小麦幼叶超氧物歧化酶(SOD)和过氧化氢酶活性。只有由单价和二价阳离子组成的平衡混合盐才能使原生质体维持较高的完整率、存活率和较正常的膜透性.并能使细胞维持较高的SOD和过氧化氢酶活性。 认为单盐毒害机理可能是首先引起细胞膜发生不正常的膜相变或细胞累积较多的有害氧自由基,引起膜脂发生过氧化或脱酯化而破坏膜结构。在离子平衡混合盐作用下,膜系才能维持正常液晶相,具有较高活性的SOD和过氧化氢酶等生物保护性酶系是离子拮抗作用之原因。     

An ionometric study of Na+ and K+ fluxes across the nystatin-modified human erythrocyte membrane]

The application of ion-selective electrodes is discussed for the kinetic determination of K+ and Na+ concentrations in the system, containing human red blood cells modified by nystatin. A series of mixed solutions was worked out, according to which the Na(+)-glass and the K(+)-thick membrane valinomycin electrodes were calibrated. The human erythrocytes were washed for 3 times with the basic solution (in mol per liter: 0.141 NaCl, 0.004 KCl, 0.002 CaCl2, 0.003 MgCl2, 0.01 glucose), and then were resuspended in it. The suspension was kept in a shaking bath at 37 degrees C. The modification of the cell membranes was performed by the introduction of different amounts of the antibiotic nystatin into the probe. Under these conditions the concentration of Na+ decreased, while K+ concentration increased. The values of concentration were registered ionometrically. In an hour and a half the stationary lines were obtained. Being based on the values of the stationary cation concentrations and the final concentrations, registered after the complete lysis of erythrocytes promoted by saponin, the ratio of cation fluxes across the modified membrane to the flux across the nonmodified membrane was calculated in accordance with the Hodgkin-Katz equation.     

Capacity of the outer membrane of a gram-negative marine bacterium in the presence of cations to prevent lysis by Triton X-100.

Cells of marine pseudomonad B-16 (ATCC 19855) washed with a solution containing 0.3 M NaCl, 50 mM MgCl2, and 10 mM KCl (complete salts) could be protected from lysis in a hypotonic environment if the suspending medium contained either 20 mM Mg2+, 40 mM Na+, or 300 mM K+. When the outer double-track layer (the outer membrane) of the cell envelope was removed to yield mureinoplasts, the Mg2+, Na+ or K+, requirements to prevent lysis were raised to 80, 210, and 400 mM, respectively. In the presence of 0.1% Triton X-100, 220, 320, and 360 mM Mg2+, Na+ or K+, respectively, prevented lysis of the normal cells. Mureinoplasts and protoplasts, however, lysed instantly in the presence of the detergent at all concentrations of Mg2+, Na+, or K+ tested up to 1.2 M. Thus, the structure of the outer membrane appears to be maintained by appropriate concentrations of Mg2+ or Na+ in a form preventing the penetration of Triton X-100 and thereby protecting the cytoplasmic membrane from dissolution by the detergent. K+ was effective in this capacity with cells washed with complete salts solution but not with cells washed with a solution of NaCl, suggesting that bound Mg2+ was required in the cell wall membrane for K+ to be effective in preventing lysis by the detergent. At high concentrations (1 M) K+ and Mg2+, but not Na+, appeared to destabilize the structure of the outer membrane in the presence of Triton X-100.     

Significance of regional difference in ion concentrations in lizard, Hemidactylus flaviviridis (Rüppell): assessment of ionic influence on sperm motility in vitro

A gradual increase in the concentration of Ca2+ from anterior to the posterior region was observed when mono- and divalent cations were estimated in different segments of the epididymis in wall lizard. Na+ and K+ levels increased from anterior to middle segment but declined significantly in the posterior segment. However, no significant difference in the levels of Mg2+ was observed in various segments. To study the influence of mono- and divalent cations on sperm motility in vitro, the spermatozoa from posterior region of the epididymis were incubated in medium with varying concentrations of Na+, K+, Ca2+ and Mg2+. Spermatozoa were non-motile when suspended in Na+-free medium. Addition of NaCl induced the acquisition of sperm motility in a concentration-dependent manner. Further, amiloride, a Na+-influx blocker, markedly reduced the Na+-induced forward progressive motility. Unlike Na+, the presence of K+ or Ca2+ in the incubation medium reduced the motility of spermatozoa even at very low concentrations. The inhibitory effect of Ca2+ decreased when nifedipine, a Ca2+-influx blocker, was added to the medium. Mg2+ at high concentrations only was able to reduce the forward progressive motility.     

Modeling the acid–base properties of glutathione in different ionic media, with particular reference to natural waters and biological fluids

The acid-base properties of γ-L-glutamyl-L-cysteinyl-glycine (glutathione, GSH) were determined by potentiometry (ISE-H(+), glass electrode) in pure NaI((aq)) and in NaCl((aq))/MgCl(2(aq)), and NaCl((aq))/CaCl(2(aq)) mixtures, at T = 298.15 K and different ionic strengths (up to I(c) ~ 5.0 mol L(-1)). In addition, the activity coefficients of glutathione were also determined by the distribution method at the same temperature in various ionic media (LiCl((aq)), NaCl((aq)), KCl((aq)), CsCl((aq)), MgCl(2(aq)), CaCl(2(aq)), NaI((aq))). The results obtained were also used to calculate the Specific ion Interaction Theory (SIT) and Pitzer coefficients for the dependence on medium and ionic strength of glutathione species, as well as the formation constants of weak Mg(j)H( i )(GSH)((i+2j-3)) and Ca(j)H(i)(GSH)((i+2j-3)) complexes. Direct calorimetric titrations were also carried out in pure NaCl((aq)) and in NaCl((aq))/CaCl(2(aq)) mixtures at different ionic strengths (0.25 ≤ I (c )/mol L(-1) ≤ 5.0) in order to determine the enthalpy changes for the protonation and complex formation equilibria in these media at T = 298.15 K. Results obtained are useful for the definition of glutathione speciation in any aqueous media containing the main cations of natural waters and biological fluids, such as Na(+), K(+), Mg(2+), and Ca(2+). Finally, this kind of systematic studies, where a series of ionic media (e.g., all alkali metal chlorides) is taken into account in the determination of various thermodynamic parameters, is useful for the definition of some trends in the thermodynamic behavior of glutathione in aqueous solution.     

Induction of the acrosome reaction in goat spermatozoa in simple physiological salt solution

The present study was carried out to determine whether K+, Mg2+, PO4(3-), and HCO3- in a medium are necessary for inducing the acrosome reaction in ejaculated goat spermatozoa. Washed goat spermatozoa were resuspended in K-1 medium, containing NaCl, KCl, CaCl2, MgCl2, NaH2PO4, and NaHCO3; in K-2 medium, containing NaCl, CaCl2, NaH2PO4, and NaHCO3; in K-3 medium, containing NaCl, CaCl2, and NaHCO3; and in K-4 medium, containing only NaCl and CaCl2, followed by preincubation in a sealed glass tube at 39.5 degrees C for 1, 2, or 3 h. The sperm acrosome reaction was evaluated by the trypan blue-Giemsa method and hamster test. The results were essentially the same in all cases. Following preincubation for 1 h, however, the percentage of acrosome-reacted spermatozoa, the proportion of zona-free hamster eggs penetrated by spermatozoa, and the average number of spermatozoa in the vitellus of these penetrated eggs were low; all values indicated a significant increase with preincubation for 2 and 3 h. The presence of K+, Mg2+, PO4(3-), and HCO3- in the medium thus does not appear necessary for inducing the acrosome reaction in goat spermatozoa, since they can undergo the reaction during preincubation in a simple physiological salt solution containing only NaCl and CaCl2 when preincubated in sealed glass tubes at 39.5 degrees C.     

In vitro chemotactic responses of Brugia pahangi infective larvae to sodium ions

In vitro chemotactic responses of infective third-stage larvae (L3) of Brugia pahangi to NaCl, Na2HPO4, KCl, K2HPO4, MgCl2 and CaCl2 were assessed. Compared to deionized water as a control, 200?mm NaCl and 100?mm Na2HPO4 significantly attracted L3 (P?相似文献   

Transport ATPases in the erythrocytes of rats acclimatized to intermittent altitude hypoxia

The activity of Na+/K+- and Ca2+-ATPase and some allosteric properties of Na+/K+-ATPase were studied in whole erythrocytes and their membrane preparations (ghosts) from rats exposed to intermittent altitude hypoxia (10 and 24 exposures, 8 h/day in an altitude chamber, stepwise up to an altitude of 7,000 m). Ca2+-ATPase activity was increased both in whole erythrocytes and ghosts after the first phase of acclimatization (10 exposures). In a standard incubation medium (containing 3 mmol.l-1 MgCl2 ), Na+/K+-ATPase activity in the ghosts was also increased after the initial phase of acclimatization whereas in whole erythrocytes Na+/K+-ATPase was only decreased in the regression phase. At high MgCl2 concentrations (12 mmol.l-1) changes of Na+/K+-ATPase activity both in whole erythrocytes and in the ghosts followed similar time course with a pronounced increase in the first phase of acclimatization (10 exposures) followed by an abrupt drop (24 exposures) and then by a gradual normalization in the regression phase. Sensitivity of the enzyme to mounting MgCl2 concentrations was increased in the ghosts at the end of acclimatization and was decreased in whole erythrocytes during acclimatization and especially in the regression phase. It has been suggested that chronic altitude hypoxia leads to the alteration of cooperative interaction of the Na+/K+-ATPase subunits in the erythrocyte membrane and accumulation of some factor in the cells inhibiting this enzyme.     

Substituting manganese for magnesium alters certain reaction properties of the (Na+ + K+)-ATPase

MnCl2 was partially effective as a substitute for MgCl2 in activating the K+- dependent phosphatase reaction catalyzed by a purified (Na+ + K+)-ATPase enzyme preparation from canine kidney medulla, the maximal velocity attainable being one-fourth that with MgCl2. Estimates of the concentration of free Mn2+ available when the reaction was half-maximally stimulated lie in the range of the single high-affinity divalent cation site previously identified (Grisham, C.M. and Mildvan, A.S. (1974) J. Biol. Chem. 249, 3187--3197). MnCl2 competed with MgCl2 as activator of the phosphatase reaction, again consistent with action through a single site. However, with MnCl2 appreciable ouabain-inhibitable phosphatase activity occurred in the absence of added KCl, and the apparent affinities for K+ as activator of the reaction and for Na+ as inhibitor were both decreased. For the (Na+ + K+)-ATPase reaction substituting MnCl2 for MgCl2 was also partially effective, but no stimulation in the absence of added KCl, in either the absence or presence of NaCl, was detectable. Moreover, the apparent affinity for K+ was increased by the substitution, although that for Na+ was decreased as in the phosphatase reaction. Substituting MnCl2 also altered the sensitivity to inhibitors. For both reactions the inhibition by ouabain and by vanadate was increased, as was binding of [48V] -vanadate to the enzyme; furthermore, binding in the presence of MnCl2 was, unlike that with MgCl2, insensitive to KCl and NaCl. Inhibition of the phosphatase reaction by ATP was decreased with 1 mM but not 10 mM KCl. Finally, inhibition of the (Na+ + K+)-ATPase reaction by Triton X-100 was increased, but that by dimethylsulfoxide decreased after such substitution. These findings are considered in terms of Mn2+ at the divalent cation site being a better selector than Mg2+ of the E2 conformational states of the enzyme, states also selected by K+ and by dimethylsulfoxide and reactive with ouabain and vanadate; the E1 conformational states, by contrast, are those selected by Na+ and ATP, and also by Triton X-100.     

Effects of medium amino acids on ouabain-sensitive 86Rb+ -uptake and membrane-potential dependent [3H]tetraphenylphosphonium accumulation in Friend erythroleukemia cells

The effects of amino acids present in minimal essential medium were investigated on 86Rb+ -fluxes and on the membrane-potential dependent accumulation of the lipophilic cation [3H]tetraphenylphosphonium (TTP+) in logarithmically growing Friend erythroleukemia cells. The ouabain-sensitive 86Rb+ -uptake measured as well in complete growth medium as in Earle's balanced salt solution (EBSS) with amino acid composition present in growth medium, was 3 to 4-fold increased in comparison to the 86Rb+-uptake measured in pure EBSS only. The Na+,K+,2Cl- -cotransport measured as piretanide-sensitive 86Rb+-uptake was reduced in the presence of amino acids. Stimulation of the ouabain-sensitive 86Rb+ -uptake could be brought about by the addition of alanine alone or of the sodium ionophore monensin. In spite of the activation of the Na+,K+ -pump the membrane-potential dependent accumulation of [3H]TPP+ was about 40 per cent reduced in the presence of medium amino acids indicating a decreased membrane potential under these conditions. On the other hand, monensin which induces an electrically silent Na+ -influx via Na+/H+ -exchange was shown to hyperpolarize the membrane on the basis of [3H]TPP+-accumulation. These results suggest that the intensive uptake of neutral amino acids by Na+-cotransport in rapidly growing cells may be responsible for both stimulation of the Na+,K+ -pump and decrease in the transmembrane potential.     

Effect of cyclic nucleotides on calcium dependent K+ channels, in human erythrocytes

K+ efflux has been analyzed in human erythrocytes incubated in a K+ free medium containing ouabain, bumetanide, CaCl2, and the Ca2+ ionophore A23187. In these conditions, a K+ efflux, which is exponentially dependent on the concentration of A23187 present in the medium, has been observed. This flux is almost completely abolished by either quinine or EGTA, so that, the above K+ efflux has been considered Ca2+ dependent. The effects of cAMP, and cGMP, have been tested on this flux. Ca2+ dependent K+ efflux decreases in presence of millimolar concentrations of cAMP in the medium. The addition of methyl-isobutyl-xanthine to the incubation medium containing cAMP enhances the inhibitory effect of this compound. cGMP also inhibits the Ca2+ dependent K+ efflux. Our results suggest that cyclic nucleotides may modulate the activation of Ca2+ dependent K+ channels in human erythrocytes.     

Kinetic studies on the (Na+ + K+)-dependent ATPase evidence for coexisting sites for Na+, K+ and Mg2+

Na+-ATPase activity of a dog kidney (Na+ + K+)-ATPase enzyme preparation was inhibited by a high concentration of NaCl (100 mM) in the presence of 30 microM ATP and 50 microM MgCl2, but stimulated by 100 mM NaCl in the presence of 30 microM ATP and 3 mM MgCl2. The K0.5 for the effect of MgCl2 was near 0.5 mM. Treatment of the enzyme with the organic mercurial thimerosal had little effect on Na+ -ATPase activity with 10 mM NaCl but lessened inhibition by 100 mM NaCl in the presence of 50 microM MgCl2. Similar thimerosal treatment reduced (Na+ + K+)-ATPase activity by half but did not appreciably affect the K0.5 for activation by either Na+ or K+, although it reduced inhibition by high Na+ concentrations. These data are interpreted in terms of two classes of extracellularly-available low-affinity sites for Na+: Na+-discharge sites at which Na+-binding can drive E2-P back to E1-P, thereby inhibiting Na+-ATPase activity, and sites activating E2-P hydrolysis and thereby stimulating Na+-ATPase activity, corresponding to the K+-acceptance sites. Since these two classes of sites cannot be identical, the data favor co-existing Na+-discharge and K+-acceptance sites. Mg2+ may stimulate Na+-ATPase activity by favoring E2-P over E1-P, through occupying intracellular sites distinct from the phosphorylation site or Na+-acceptance sites, perhaps at a coexisting low-affinity substrate site. Among other effects, thimerosal treatment appears to stimulate the Na+-ATPase reaction and lessen Na+-inhibition of the (Na+ + K+)-ATPase reaction by increasing the efficacy of Na+ in activating E2-P hydrolysis.     

Calcium inhibition of the ATPase and phosphatase activities of (Na+ + K+)-ATPase

In experiments performed at 37 degrees C, Ca2+ reversibly inhibits the Na+-and (Na+ + K+)-ATPase activities and the K+-dependent phosphatase activity of (Na+ + K+)-ATPase. With 3 mM ATP, the Na+-ATPase was less sensitive to CaCl2 than the (Na+ + K+)-ATPase activity. With 0.02 mM ATP, the Na+-ATPase and the (Na+ + K+)-ATPase activities were similarly inhibited by CaCl2. The K0.5 for Ca2+ as (Na+ + K+)-ATPase inhibitor depended on the total MgCl2 and ATP concentrations. This Ca2+ inhibition could be a consequence of Ca2+-Mg2+ competition, Ca . ATP-Mg . ATP competition or a combination of both mechanisms. In the presence of Na+ and Mg2+, Ca2+ inhibited the K+-dependent dephosphorylation of the phosphoenzyme formed from ATP, had no effect on the dephosphorylation in the absence of K+ and inhibited the rephosphorylation of the enzyme. In addition, the steady-state levels of phosphoenzyme were reduced in the presence both of NaCl and of NaCl plus KCl. With 3 mM ATP, Ca2+ alone sustained no more than 2% of the (Na+ + K+)-ATPase activity and about 23% of the Na+-ATPase activity observed with Mg2+ and no Ca2+. With 0.003 mM ATP, Ca2+ was able to maintain about 40% of the (Na+ + K+)-ATPase activity and 27% of the Na+-ATPase activity seen in the presence of Mg2+ alone. However, the E2(K)-E1K conformational change did not seem to be affected. Ca2+ inhibition of the K+-dependent rho-nitrophenylphosphatase activity of the (Na+ + K+)-ATPase followed competition kinetics between Ca2+ and Mg2+. In the presence of 10 mM NaCl and 0.75 mM KCl, the fractional inhibition of the K+-dependent rho-nitrophenylphosphatase activity as a function of Ca2+ concentration was the same with and without ATP, suggesting that Ca2+ indeed plays the important role in this process. In the absence of Mg2+, Ca2+ was unable to sustain any detectable ouabain-sensitive phosphatase activity, either with rho-nitrophenylphosphate or with acetyl phosphate as substrate.     

Reversible induction of actin rods in mouse C3H-2K cells by incubation in salt buffers and by treatment with non-ionic detergents

Incubating conditions which induced actin paracrystal-like intracellular structures (actin rods) were investigated by using several cell lines. We have found that an incubation of cells of a mouse fibroblastic cell line, C3H-2K, in an isotonic solution of NaCl containing 1 mM MgCl2, 1 mM CaCl2 and 10 mM MES, pH 6.5, induced disintegration of stress fibers and formation of actin rods in the cytoplasm. Actin rods were induced also by incubating in salt buffers in which Na+ of the above solution was substituted by most cations except K+ or Rb+. When the actin rod-forming cells were transferred back to DMEM containing 10% FBS, actin rods disappeared and stress fibers subsequently re-formed within 1 h at 37 degrees C. Although the induction was observed in NaCl buffer at a wide range of pH values (5.5-10), the optimal pH was 6.5. Formation of actin rods is dependent upon cellular metabolism, as it was inhibited at 4 degrees C, or by metabolic inhibitors. Incubation in NaCl buffer induced actin rods in HeLa, L, NRK, BALB/c 3T3 and Swiss 3T3 cells, but not in CEF or MEF cells. A decrease in cell volume was observed parallel with the induction of actin rods, except for CEF and MEF cells. Alterations in intracellular concentrations of Na, K or Ca were not correlated with the induction, however. Actin rods were also induced in C3H-2K cells by a brief treatment with non-ionic detergents. Tween 80 at concentrations as low as 0.003% was effective for the induction, but did not increase the passive membrane transport of p-nitrophenylphosphate. In contrast to the induction by NaCl buffer, treatment with Tween 80 induced numerous tiny actin rods at 4 degrees C, which became larger when further incubated at 37 degrees C. Double immunofluorescence staining with anti-actin antibody and anti-vinculin antibody showed that vinculin plaques remained at least in an early stage of the actin rod formation. We discuss the mechanism for the induction of actin rods based upon the present findings.     

Electrical characteristics in an excitable element of lipid membrane.

Electrical characteristics in a membrane constructed from a porous filter adsorbed with a lipid analogue, dioleoyl phosphate (DOPH), were investigated in a situation interposed between 100 mM NaCl + 3 mM CaCl2 and 100 mM KCl. Calcium ions affected significantly the membrane characteristics. The membrane potential was negative on the KCl side, which implies the higher permeability to K+ than Na+; this tendency was increased by a tiny amount of Ca2+. While the membrane showed a low electrical resistance of several k omega . cm2 under K+/Na+ gradient, it showed several M omega . cm2 by Ca2+. The surface structure of the membrane exhibited many voids in the low-resistance state, but the surface was covered by oil droplets in the high-resistance state. Oscillations of the membrane potential appeared spontaneously with application of the electrical current from the KCl side to the NaCl + CaCl2 side. The frequency was increased with the electrical current. All these results were explained comprehensively using an electrochemical kinetic model taking account of the Ca2+ binding effect, where DOPH assemblies make a phase transition between oil droplets due to Ca2+ and multi-bilayers with excess K+. The oscillation arises from coupling of the phase transition to accumulation and release of K+ or Ca2+. This membrane can be used as an excitable element regulated by Ca2+ in neuro-computer devices.     

Mutant selection of <Emphasis Type="Italic">Hahella chejuensis</Emphasis> KCTC 2396 and statistical optimization of medium components for prodigiosin yield-up

Prodigiosin is a natural red pigment with algicidal activity against Cochlodinium polykrikoides, a major harmful red-tide microalga. To increase the yield of prodigiosin, a mutant of Hahella chejuenesis KCTC 2396, assigned M3349, was developed by an antibiotic mutagenesis using chloramphenicol. When cultured in Sucrose-based Marine Broth medium (SMB), M3349 could produce prodigiosin at 1.628+/-0.06 g/L, while wild type producing at 0.658+/-0.12 g/L under the same conditions. To increase the yield of prodigiosin production by M3349, significant medium components were determined using a two-level Plackett-Burman statistical design technique. Among fourteen components included in SMB medium, NaCl, Na2SiO3, MgCl2, H3BO3, Na2HPO4, Na2SO4, and CaCl2 were determined to be important for prodigiosin production. The medium formulation was finally optimized using a Box-Behnken design as follows: sucrose 10.0, peptone 8.0, yeast extract 2.0, NaCl 10.0, Na2SO4 12.0, CaCl2 1.8, MgCl2 0.7 g/L; and H3BO3 22.0, Na2HPO4 20.0, Na2SiO3 8.0 mg/L. The predicted maximum yield of prodigiosin in the optimized medium was 2.43 g/L by the Box-Behnken design, while the practical production was 2.60+/-0.176 g/L, which was 3.9 times higher than wild type with SMB Medium (0.658 g/L).     

Electrical perception of the 'death message' in Chara: characterization of K+ -induced depolarization

When the nodal end of an internodal cell of Chara corallina was subjected to a pulse treatment with artificial cell sap, a depolarization lasted in artificial pond water. This depolarization could also be induced by pulse treatment with KCl solution, indicating that K+ in the artificial cell sap is responsible (K+ -induced depolarization). The depolarization was prolonged in the presence of 2 mM KCl and the prolonged depolarization was terminated by supplementing with either CaCl2, MgCl2 or NaCl. These results supported the hypothesis that K+ released from the killed cell is responsible for generation of the wound-induced membrane depolarization.     

Use of a D-optimal design with constrains to quantify the effects of the mixture of sodium,potassium, calcium and magnesium chloride salts on the growth parameters of <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>

The combined effect of NaCl, KCl, CaCl(2), and MgCl(2) on the water activity (a (w)) and the growth parameters of Saccharomyces cerevisiae was studied by means of a D-optimal mixture design with constrains (total salt concentrations < or = 9.0%, w/v). The a (w) was linearly related to the concentrations of the diverse salts; its decrease, by similar concentrations of salts, followed the order NaCl > CaCl(2) > KCl > MgCl(2), regardless of the reference concentrations used (total absence of salts or 5% NaCl). The equations that expressed the maximum specific growth (mu (max)), lag phase duration (lambda), and maximum population reached (N (max)) showed that the values of these parameters depended on linear effects and two-way interactions of the studied chloride salts. The mu (max) decreased as NaCl and CaCl(2) increased (regardless of the presence or not of previous NaCl); however, in the presence of a 5% NaCl, a further addition of KCl and MgCl(2) markedly increased mu (max). The lambda was mainly affected by MgCl(2) and the interactions NaCl x CaCl(2) and CaCl(2) x MgCl(2). The further addition of NaCl and CaCl(2) to a 5% NaCl medium increased the lag phase while KCl and MgCl(2) had negligible or slightly negative effect, respectively. N (max) was mainly affected by MgCl(2) and its interactions with NaCl, KCl, and CaCl(2); MgCl(2) stimulated N (max) in the presence of 5% NaCl while KCl, NaCl, and CaCl(2) had a progressive decreasing effect. These results can be of interest for the fermentation and preservation of vegetable products, and foods in general, in which this yeast could be present.