Annuario della Società Botanica Italiana 1932

A sandy culture experiment was conducted to investigate the effects of exogenous CaCl₂ on the indole alkaloid accumulation in Catharanthus roseus under salt stress. One-month seedlings of C. roseus were treated with the different concentrations of NaCl (0, 50, and 100 mmol l^? 1) and 7.5 mmol l^? 1 CaCl₂. The plant samples were analyzed after 7 days of the treatments. The NaCl-stressed plants showed decrease of fresh and dry weight and increase of malondialdehyde (MDA) content compared to control. Tryptophan decarboxylase (TDC) activity increased significantly under 50 mmol l^? 1 NaCl without CaCl₂ addition, 50 mmol l^? 1 NaCl with 7.5 mmol l^? 1 CaCl₂, and CaCl₂ treatment without NaCl addition. There was a significant increase in peroxidase activity under NaCl stress compared to control. The vindoline, catharanthine, vincristine, and vinblastine contents increased under salt stress (especially with 50 mmol l^? 1 NaCl treatment with or without CaCl₂). Addition of CaCl₂ to NaCl-stressed plants increased biomass, TDC activity, vindoline, and catharanthine contents and lowered MDA and vincirstine contents compared to the plants without CaCl₂. The plants treated with CaCl₂ alone showed higher TDC activity, vindoline, catharanthine, and vinblastine content when compared to control. The results showed that exogenous CaCl₂ could promote the indole alkaloid metabolism under salt stress.     

The function of 33-kDa protein in the photosynthetic oxygen-evolution system studied by reconstitution experiments

Treatment of Photosystem II particles with 1.2 M CaCl₂ released three proteins of 33, 24 and 18 kDa of the photosynthetic oxygen evolution system, but left Mn bound to the particles as demonstrated by Ono and Inoue (Ono, T. and Inoue, Y. (1983) FEBS Lett. 164, 252–260). Oxygen-evolution activity of the CaCl₂-treated particles was very low in a medium containing 10 mM NaCl as a salt, but could be restored by the 33-kDa protein. When the particles were incubated in 10 mM NaCl at 0°C, two of the four Mn atoms per oxygen-evolution system were released with concomitant loss of oxygen-evolution activity. The 33-kDa protein suppressed the release of Mn and the inactivation during the incubation. These findings from reconstitution experiments suggest that the 33-kDa protein acts to preserve Mn atoms in the oxygen-evolution system. The 33-kDa protein could be partially substituted by 100 or 150 mM Cl⁻ for the preservation of the Mn and oxygen-evolution activity. The Mn in Photosystem II particles enhanced rebinding of the 33-kDa protein to the particles.     

Calcium chloride and gibberellic acid protect linseed (Linum usitatissimum L.) from NaCl stress by inducing antioxidative defence system and osmoprotectant accumulation

Salinity stress affects many metabolic facets of plants and induces anatomical and morphological changes resulting in reduced growth and productivity. To overcome the damaging effects of salinity, different strategies of the application of nutrients with plant hormones are being adopted. The present study was carried out with an aim to find out whether application of calcium chloride (CaCl₂) and gibberellic acid (GA₃) could alleviate the detrimental effects of salinity stress on plant metabolism. Fifteen days old plants were supplied with (1) 0 mM NaCl + 0 mg CaCl₂ kg^?1 sand + 0 M GA₃ (control, T0); (2) 0 mM NaCl + 10 mg CaCl₂ kg^?1 sand + 0 M GA₃ (T1); (3) 0 mM NaCl + 0 mg CaCl₂ kg^?1 sand + 10^?6 M GA₃ (T2); (4) 150 mM NaCl + 0 mg CaCl₂ kg^?1 sand + 0 M GA₃ (T3); (5) 150 mM NaCl + 10 mg CaCl₂ kg^?1 sand + 0 M GA₃ (T4); (6) 150 mM NaCl + 0 mg CaCl₂ kg^?1 sand + 10^?6 M GA₃ (T5); (7) 150 mM NaCl + 10 mg CaCl₂ kg^?1 sand + 10^?6 M GA₃ (T6). To assess the response of the crop to NaCl, CaCl₂ and GA₃, plants were uprooted randomly at 60 days after sowing. The presence of NaCl in the growth medium decreased all the growth and physio-biochemical parameters, except electrolyte leakage, proline (Pro) and glycine betaine (GB) content, thiobarbituric acid reactive substances (TBARS), H₂O₂ content, activities of superoxide dismutase (SOD) and catalase (CAT) and leaf Na content, which exhibited an increase of 37.6, 29.3, 366.9, 107.5, 59.1, 17.1, 28.4 and 255.2%, respectively, compared to the control plants. However, application of CaCl₂ in combination with GA₃ appears to confer greater osmoprotection by the additive role with NaCl in Pro and GB accumulation. Although the activities of antioxidant enzymes (SOD, CAT and POX) were increased by salt stress, the combined application of CaCl₂ and GA₃ to salt-stressed plants further enhanced the activities of these enzymes by 25.1, 6.7 and 47.8%, respectively, compared to plants grown with NaCl alone. The present study showed that application of CaCl₂ and GA₃ alone as well as in combination mitigated the adverse effect of salinity, but combined application of these treatments proved more effective in alleviating the adverse effects of NaCl stress.     

Effect of varying NaCl doses on flavonoid production in suspension cells of Ginkgo biloba: relationship to chlorophyll fluorescence,ion homeostasis,antioxidant system and ultrastructure

Ginkgo suspension cells were used to investigate the mechanism that governs the shift between primary and secondary metabolism under NaCl elicitation. The production of three flavonol glycosides, chlorophyll fluorescence, ion content, the antioxidant system, and the cellular ultrastructure in the presence of NaCl doses from 5 to 175 mM were examined. At low salt doses (5–50 mM), cell growth and flavonol glycosides accumulation were stimulated without damaging cell structure or inducing oxidative stress by maintaining high K⁺ and chlorophyll content. At moderate salt doses (75–125 mM), the cells could withstand the salt stress without an impact on survival by changing internal cellular structure, maintaining high levels of K⁺ and Ca²⁺ and increasing anti-oxidative enzyme activities rather than flavonol glycosides to counteract the inhibition of the photosystem II, the accumulation of Na⁺ and hydrogen peroxide (H₂O₂) in the cells. This allowed cells to divert their metabolism from growth to defense-related pathways and tolerate NaCl stress. At higher salinity (150–175 mM), the cellular structure was damaged, and the high Na⁺ and low K⁺ content led to osmotic stress, and therefore, the stimulation of peroxidase (POD) and catalase (CAT) was not enough to cope with high H₂O₂ accumulation. The high production of flavonol glycosides may be a response of elicitation stimulation to serious damage at 175 mM NaCl. In conclusion, the use of 175 mM NaCl may be desirable for the induction of flavonol glycoside production in Ginkgo suspension cells.     

Metabolic threshold and sulfide-buffering in diffusion controlled marine sediments impacted by continuous organic enrichment

The effects of organic enrichment on sediment biogeochemistry was studied in diffusion controlled sediment mesocosms, where labile organic matter (OM) (fish feed) pulses were added once a week to the sediment surface. Two types of sediments, differing mainly in content of reactive Fe, were used. The aim of this experiment was two-fold, (1) to evaluate the importance of Fe-driven sulfide buffering for sulfide accumulation in surface enriched sediments, and (2) to estimate the diagenetic capacity for degradation of labile OM near the sediment surface. The simulated OM loading rate of 375 mmol C m^?2 day^?1 led to a 5–6 times increase in CO₂-production and a 4–5 times increase in O₂-uptake. Sulfate reduction estimated by radiotracer experiments and CO₂-release was 105–131 mmol m^?2 day^?1, but accumulation of porewater sulfide was low in both sediment types. Instead 99% of sulfide was oxidized with O₂ at the sediment water interface in the low Fe treatment, whereas 46% of produced sulfide precipitated as Fe-S compound in the high Fe treatment resulting in significantly lower O₂-uptake. Furthermore, the accumulation of up to 30% of added OM by the end of the experiment indicated a saturation of the heterotrophic microbial communities in the upper enriched surface layer. These results suggest a maximum diagenetic capacity for OM degradation in the range of ~25 μmol C cm^?3 day^?1 or 260 mmol m^?2 day^?1 for the present sediment types.     

Microdistribution of major to trace elements between roots of Halimione portulacoides and host sediments (Tagus estuary marsh,Portugal)

Aim

This study presents a micrometre-scale map of the elemental distribution within roots and surrounding sediment of Halimione portulacoides of a contaminated salt marsh in the Tagus estuary.

Methods

Microprobe particle induced X-ray emission analysis was performed in sediment slices containing roots with tubular rhizoconcretions attached to host sediments.

Results

Strong concentration gradients were found particularly in the inner part of rhizoconcretions adjacent to the root wall. Local enrichment was observed in sediment interstices with Fe precipitates and other associated elements. A maximum of 55 % of Fe was measured near the concretion–root interface, with a decrease to <5 % in the host sediment. Maximum concentrations of P (3 %), As (1,200 μg g^?1) and Zn (3,000 μg g^?1) were registered in concretions, one order of magnitude above the values of the host sediment. The elemental concentration profiles across roots showed that the epidermis was an efficient selective barrier to the entrance of elements. Fe and As were retained in the epidermis. The highest Cu and Zn concentrations were also observed in the epidermis. However, the concentrations of Mn, Cu and Zn increased in the inner root.

Conclusions

As and Fe were mostly retained in the concretion, whereas P, Mn, Cu and Zn were mobilised by the root.     

Inducing salt tolerance in maize (Zea mays L.) through seed priming with chloride salts: Growth and ion transport at early growth stages

The study was conducted to determine whether salt tolerance could be induced in maize at germination stage by soaking of seeds for 8 h in distilled water or in 200 meq·L⁻¹ of NaCl, KCl, CaCl₂·2H₂O. Both primed and un-primed seeds were subjected for 14 days to 0, 100 or 200 mol·m⁻³ NaCl under controlled conditions. Although all priming agents were effective in alleviating adverse effects of salt stress on maize at germination stage, CaCl₂·2H₂O proved to be more effective since the seeds primed with this salt had significantly higher final germination, rate of germination and fresh and dry weights of plumules and radicles than those treated with other salts or distilled water. Concentration of Na⁺, K⁺ and Ca²⁺ increased significantly in all parts of germinating seeds of maize seeds primed with NaCl, KCl, or CaCl₂·2H₂O, respectively. In addition, seeds primed with CaCl₂·2H₂O were the highest in Cl⁻ accumulation in all parts of the germinating seeds, followed by seeds treated with NaCl and KCl. Most of the Ca²⁺ was retained in seeds and mesocotyl, because of which, transport of this ion to plumules and radicles was low.     

A novel cold-active and salt-tolerant α-amylase from marine bacterium Zunongwangia profunda: molecular cloning,heterologous expression and biochemical characterization

A novel gene (amyZ) encoding a cold-active and salt-tolerant α-amylase (AmyZ) was cloned from marine bacterium Zunongwangia profunda (MCCC 1A01486) and the protein was expressed in Escherichia coli. The gene has a length of 1785 bp and encodes an α-amylase of 594 amino acids with an estimated molecular mass of 66 kDa by SDS-PAGE. The enzyme belongs to glycoside hydrolase family 13 and shows the highest identity (25 %) to the characterized α-amylase TVA II from thermoactinomyces vulgaris R-47. The recombinant α-amylase showed the maximum activity at 35 °C and pH 7.0, and retained about 39 % activity at 0 °C. AmyZ displayed extreme salt tolerance, with the highest activity at 1.5 M NaCl and 93 % activity even at 4 M NaCl. The catalytic efficiency (k _cat/K _m) of AmyZ increased from 115.51 (with 0 M NaCl) to 143.30 ml mg^?1 s^?1 (with 1.5 M NaCl) at 35 °C and pH 7.0, using soluble starch as substrate. Besides, the thermostability of the enzyme was significantly improved in the presence of 1.5 M NaCl or 1 mM CaCl₂. AmyZ is one of the very few α-amylases that tolerate both high salinity and low temperatures, making it a potential candidate for research in basic and applied biology.     

Interactive effects of Ca2+ and NaCl salinity on the growth of two tomato genotypes differing in Ca2+ use efficiency

Two tomato (Lycopersicon esculentum Mill.) lines differing in Ca²⁺ use efficiency (Ca²⁺ use efficient line 113 and Ca²⁺ use inefficient line 67) were subjected to salinity treatments in two separate experiments to determine whether they differed in salt tolerance. In experiment I, three NaCl and two CaCl₂ treatments were imposed. The Na⁺ concentrations were 1.1, 100 and 150 mM and the Ca²⁺ concentrations were either 1.51 or 10 mM. In experiment II, one NaCl and three Ca²⁺ treatments (as CaCl₂ or CaSO₄) were imposed. The treatments consisted of 150 mM NaCl at either 1.51 mM CaCl₂, 10 mM CaCl₂, or 10 mM CaSO₄. Response to treatments was determined by analysis of growth parameters (shoot and root dry weights, plant height, and root length). Shoot and root dry weight, and root length were depressed as salinity increased in plants lacking additional Ca²⁺. No significant differences in salt tolerance were detected between the two tomato lines after 24 d of salinity treatment. An important finding of this study was that root growth and length appeared to be more sensitive to the effect of CaCI₂ treatment alone and to the effects of CaCl₂ × NaCl treatments. This suggests that over the long term, both root growth and root length may be more sensitive indicators of salinity effects than shoots. Supplemental CaCl₂ had no ameliorative effect on NaCl stress in shoot growth. The inability of Ca²⁺ to counter Cl⁻ entry or toxicity may account for the lack of amelioration. Additional Ca²⁺ as CaSO₄ improved shoot growth of plants exposed to 150 mM NaCl. In contrast, root growth and length were improved by 10 mM Ca²⁺ as either CaCl₂ or CaSO₄.     

Thermophilic and halophilic β-agarase from a halophilic archaeon Halococcus sp. 197A

An agar-degrading archaeon Halococcus sp. 197A was isolated from a solar salt sample. The agarase was purified by hydrophobic column chromatography using a column of TOYOPEARL Phenyl-650 M. The molecular mass of the purified enzyme, designated as Aga-HC, was ~55 kDa on both SDS-PAGE and gel-filtration chromatography. Aga-HC released degradation products in the order of neoagarohexose, neoagarotetraose and small quantity of neoagarobiose, indicating that Aga-HC was a β-type agarase. Aga-HC showed a salt requirement for both stability and activity, being active from 0.3 M NaCl, with maximal activity at 3.5 M NaCl. KCl supported similar activities as NaCl up to 3.5 M, and LiCl up to 2.5 M. These monovalent salts could not be substituted by 3.5 M divalent cations, CaCl₂ or MgCl₂. The optimal pH was 6.0. Aga-HC was thermophilic, with optimum temperature of 70 °C. Aga-HC retained approximately 90 % of the initial activity after incubation for 1 hour at 65–80 °C, and retained 50 % activity after 1 hour at 95 °C. In the presence of additional 10 mM CaCl₂, approximately 17 % remaining activity was detected after 30 min at 100 °C. This is the first report on agarase purified from Archaea.     

Salt impacts on organic carbon and nitrogen leaching from senesced vegetation

Senesced vegetation is exposed to a wide range of salt concentrations in surface waters resulting from human activities which include deicing salts and irrigation water chemistry. Both dissolved organic carbon (DOC) and salt concentrations are rising in northern hemisphere watersheds, yet there has been little investigation of sodium as a potential mechanism for DOC increases. The objective of this study was to investigate the impact of solution sodicity and salinity on DOC and dissolved organic nitrogen (DON) leaching from five types of senesced and cut vegetation. Vegetation was soaked for 24 h in a series of sodium chloride (NaCl)–calcium chloride (CaCl₂) solutions with sodium adsorption ratios (SAR) of 2, 10, or 30 and electrical conductivities of 0.1 dS m^?1 through 3.0 dS m^?1. Vegetation was also soaked in a sodium bicarbonate (NaHCO₃) solution at SAR = 30 and stream water from local watersheds with a range of sodicity and salinity. The mass of both DOC and DON released increased as SAR increased in the NaCl solutions, but the total salinity had inconsistent effects on DOC and DON release. NaHCO₃ leached similar amounts of DOC and DON as NaCl. The SAR of the stream water solutions was able to explain 88 % of the variability in DOC leached from vegetation (p < 0.05). The results indicated that sodicity, quantified by SAR, had a significant impact on DOC and DON leaching from senesced vegetation and could be a potential mechanism to explain the observed increases in surface water DOC.     

Neutrophilic, nitrate-dependent, Fe(II) oxidation by a Dechloromonas species

A species of Dechloromonas, strain UWNR4, was isolated from a nitrate-reducing, enrichment culture obtained from Wisconsin River (USA) sediments. This strain was characterized for anaerobic oxidation of both aqueous and chelated Fe(II) coupled to nitrate reduction at circumneutral pH. Dechloromonas sp. UWNR4 was incubated in anoxic batch reactors in a defined medium containing 4.5–5 mM NO₃ ^?, 6 mM Fe²⁺ and 1–1.8 mM acetate. Strain UWNR4 efficiently oxidized Fe²⁺ with 90 % oxidation of Fe²⁺ after 3 days of incubation. However, oxidation of Fe²⁺ resulted in Fe(III)-hydroxide-encrusted cells and loss of metabolic activity, suggested by inability of the cells to utilize further additions of acetate. In similar experiments with chelated iron (Fe(II)-EDTA), encrusted cells were not produced and further additions of acetate and Fe(II)-EDTA could be oxidized. Although members of the genus Dechloromonas are primarily known as perchlorate and nitrate reducers, our findings suggest that some species could be members of microbial communities influencing iron redox cycling in anoxic, freshwater sediments. Our work using Fe(II)-EDTA also demonstrates that Fe(II) oxidation was microbially catalyzed rather than a result of abiotic oxidation by biogenic NO₂ ^?.     

Salinity effects on greenhouse gas emissions from wetland soils are contingent upon hydrologic setting: a microcosm experiment

Coastal forested wetlands provide important ecosystem services such as carbon sequestration, nutrient retention, and flood protection, but they are also important sources of greenhouse gas emissions. Human appropriation of surface water and extensive ditching and draining of coastal plain landscapes are interacting with rising sea levels to increase the frequency and magnitude of saltwater incursion into formerly freshwater coastal wetlands. Both hydrologic change and saltwater incursion are expected to alter carbon and nutrient cycling in coastal forested wetlands. We performed a full factorial experiment in which we exposed intact soil cores from a coastal forested wetland to experimental marine salt treatments and two hydrologic treatments. We measured the resulting treatment effects on the emissions of carbon dioxide (CO₂), methane (CH₄), and nitrous oxide (N₂O) over 112 days. Salinity effects were compared across four treatments to isolate the effects of increases in ionic strength from the impact of adding a terminal electron acceptor (SO₄^2?). We compared control treatments (DI addition), to artificial saltwater (ASW, target salinity of 5 parts per thousand) and to two treatments that added sulfate alone (SO₄^2?, at the concentration found in 5 ppt saltwater) and saltwater with the sulfate removed (ASW-SO₄^2?, with the 5 ppt target salinity maintained by adding additional NaCl). We found that all salt treatments suppressed CO₂ production, in both drought and flooded treatments. Contrary to our expectations, CH₄ fluxes from our flooded cores increased between 300 and 1200% relative to controls in the ASW and ASW-SO₄^2? treatments respectively. In the drought treatments, we saw virtually no CH₄ release from any core, while artificial seawater with sulfate increased N₂O fluxes by 160% above DI control. In contrast, salt and sulfate decreased N₂O fluxes by 72% in our flooded treatments. Our results indicate that salinization of forested wetlands of the coastal plain may have important climate feedbacks resulting from enhanced greenhouse gas emissions and that the magnitude and direction of these emissions are contingent upon wetland hydrology.     

Improving NaCl resistance of red-osier dogwood: role of CaCl2 and CaSO4

The influence of Ca²⁺ salts on the resistance of red-osier dogwood (Cornus sericea) seedlings to salinity was investigated. Red-osier dogwood seedlings were exposed to 5 and 10 mM of CaCl₂ or CaSO₄ in the presence or absence of 50 mM NaCl for 40 days in a controlled environment. Seedlings exposed to CaCl₂ and CaSO₄ recovered from NaCl-induced transpiration reduction after 20 days at a concentration of 10 mM and after 30 days at a concentration of 5 mM; while in absence of additional Ca²⁺, the seedlings recovered only after 40 days. Addition of 10 mM Ca²⁺ to NaCl treatment also limited the accumulation of proline in leaf tissues and caused an increase in leaf and lateral shoot K⁺ content. These results suggest that 10 mM Ca²⁺ could alleviate, at least in part, the osmotic effect of NaCl on red-osier dogwood via control of stomatal closure. On the other hand, ion analysis showed that Ca²⁺ addition was able to reduce the NaCl-induced Na⁺ concentration only in stem tissues suggesting that Ca²⁺ had only a limited effect on the ionic stress. The present study also showed an unexpected NaCl-induced increase in Ca²⁺ content of leaves, lateral shoots and stems that was not observed in our previous hydroponics experiments and seems to be more characteristic of plants growing on sandy soils.     

Oxygen Dependency of Neutrophilic Fe(II) Oxidation by Leptothrix Differs from Abiotic Reaction

Neutrophilic Fe(II) oxidizing microorganisms are found in many natural environments. It has been hypothesized that, at low oxygen concentrations, microbial iron oxidation is favored over abiotic oxidation. Here, we compare the kinetics of abiotic Fe(II) oxidation to oxidation in the presence of the bacterium Leptothrix cholodnii Appels isolated from a wetland sediment. Rates of Fe(II) oxidation were determined in batch experiments at 20°C, pH 7 and oxygen concentrations between 3 and 120 μmol/l. The reaction progress in experiments with and without cells exhibited two distinct phases. During the initial phase, the oxygen dependency of microbial Fe(II) oxidation followed a Michaelis-Menten rate expression (K_M = 24.5 ± 10 μmol O₂/l, v_max = 1.8 ± 0.2 μmol Fe(II)/(l min) for 10⁸ cells/ml). In contrast, abiotic rates increased linearly with increasing oxygen concentrations. At similar oxygen concentrations, initial Fe(II) oxidation rates were faster in the experiments with bacteria. During the second phase, the accumulated iron oxides catalyzed further oxidative iron precipitation in both abiotic and microbial reaction systems. That is, abiotic oxidation also dominated the reaction progress in the presence of bacteria. In fact, in some experiments with bacteria, iron oxidation during the second phase proceeded slower than in the absence of bacteria, possibly due to an inhibitory effect of extracellular polymeric substances on the growth of Fe(III) oxides. Thus, our results suggest that the competitive advantage of microbial iron oxidation in low oxygen environments may be limited by the autocatalytic nature of abiotic Fe(III) oxide precipitation, unless the accumulation of Fe(III) oxides is prevented, for example, through a close coupling of Fe(II) oxidation and Fe(III) reduction.     

Water potential in soil and Atriplex nummularia (phytoremediator halophyte) under drought and salt stresses

Atriplex nummularia is a halophyte widely employed to recover saline soils and was used as a model to evaluate the water potentials in the soil-plant system under drought and salt stresses. Potted plants grown under 70 and 37% of field capacity irrigated with solutions of NaCl and of a mixture of NaCl, KCl, MgCl₂ and CaCl₂ reproducing six electrical conductivity (EC): 0, 5, 10, 20, 30, and 40 dS m^?1. After 100 days, total water (Ψ_{w, plant}) and osmotic (Ψ_{o, plant}) potentials at predawn and midday and Ψ_{o, soil}, matric potential (Ψ_{m, soil}) and Ψ_{w, soil} were determined. The type of ion in the irrigation water did not influence the soil potential, but was altered by EC. The soil Ψ_o component was the largest contributor to Ψ_{w, soil}. Atriplex is surviving ECs close to 40 dS m^?1 due to the decrease in the Ψ_w. The plants reached a Ψ_w of approximately ?8 MPa. The water potentials determined for different moisture levels, EC levels and salt types showed huge importance for the management of this species in semiarid regions and can be used to recover salt affected soils.     

Warfarin modulates the nitrite reductase activity of ferrous human serum heme–albumin

Human serum heme–albumin (HSA–heme–Fe) displays reactivity and spectroscopic properties similar to those of heme proteins. Here, the nitrite reductase activity of ferrous HSA–heme–Fe [HSA–heme–Fe(II)] is reported. The value of the second-order rate constant for the reduction of $ {\text{NO}}_{2}^{ - } $ to NO and the concomitant formation of nitrosylated HSA–heme–Fe(II) (i.e., k _on) is 1.3 M^?1 s^?1 at pH 7.4 and 20 °C. Values of k _on increase by about one order of magnitude for each pH unit decrease between pH 6.5 to 8.2, indicating that the reaction requires one proton. Warfarin inhibits the HSA–heme–Fe(II) reductase activity, highlighting the allosteric linkage between the heme binding site [also named the fatty acid (FA) binding site 1; FA1] and the drug-binding cleft FA2. The dissociation equilibrium constant for warfarin binding to HSA–heme–Fe(II) is (3.1 ± 0.4) × 10^?4 M at pH 7.4 and 20 °C. These results: (1) represent the first evidence for the $ {\text{NO}}_{2}^{ - } $ reductase activity of HSA–heme–Fe(II), (2) highlight the role of drugs (e.g., warfarin) in modulating HSA(–heme–Fe) functions, and (3) strongly support the view that HSA acts not only as a heme carrier but also displays transient heme-based reactivity.     

Identification of novel halotolerant bacillopeptidase F-like proteinases from a moderately halophilic bacterium, Virgibacillus sp. SK37

Aims: Virgibacillus sp. SK37 isolated from Thai fish sauce produced numerous NaCl‐activated subtilisin‐like proteinases. Our objectives were to purify, characterize and identify these extracellular proteinases. Methods and Results: Three major subtilisin‐like enzymes including 19, 34 and 44 kDa were partially purified and showed maximum activity at pH 8, 55–60°C, 25–30% NaCl and 70–100 mmol l^?1 CaCl₂. Enzymes showed stability at 0–30% NaCl and <20 mmol l^?1 CaCl₂ and were completely inhibited by phenylmethanesulphonyl fluoride but not by ethylenediaminetetraacetic acid. The isoelectric points of 19‐, 34‐ and 44‐kDa proteinases were at 3·6, 5·2 and 3·8, respectively, based on 2D electrophoresis. Peptide mass fingerprint and de novo peptide homology analysis of tryptic peptides using MALDI‐TOF and LC–MS/MS, respectively, suggested that all three enzymes were novel and homologous to bacillopeptidase F. Conclusions: The three major proteinases are a member of bacillopeptidase F‐like enzymes exhibiting thermophilic and halotolerant characteristics with high stability at 30% NaCl. Significance and Impact of the Study: This is the first report on bacillopeptidase F‐like proteinases in genus Virgibacillus with a distinct halotolerant feature. They showed potential to be a processing aid for food and biotechnological applications, particularly in high salt condition.     

Alleviation of NaCl toxicity in the cyanobacterium <Emphasis Type="Italic">Synechococcus</Emphasis> sp. PCC 7942 by exogenous calcium supplementation

Salinity (NaCl) is one of the major problems associated with irrigated agricultural lands, especially rice fields. Being the common inhabitants of rice fields, cyanobacteria frequently experience high concentration of NaCl which in turn causes cellular damage. Therefore, mitigation of NaCl stress in cyanobacteria, plant growth-promoting microorganisms, is of utmost importance. The present study was designed to investigate the role of calcium in the alleviation of NaCl stress-induced cellular in Synechococcus sp. PCC 7942. The cyanobacterium was subjected to sub-lethal concentration of NaCl (800 mM) with and without the supplementation of calcium (1 mM CaCl₂) for 8 days. The results showed a drastic reduction in growth due to excess NaCl, but supplementation of CaCl₂ reduced the salt stress damage and partially restored growth. Application of calcium increased pigment contents, photosynthetic efficiency, antioxidative enzyme activity, osmolyte contents and reduced the intracellular sodium ion concentration, MDA content, electrolyte leakage and free oxygen radical generation. Furthermore, proteins involved in photosynthesis, respiration, ATP synthesis and protein synthesis along with two hypothetical proteins were also observed to be upregulated in the cyanobacterium in presence of calcium. Furthermore, proteins related to oxidative stress defence, nitrogen metabolism, carbohydrate metabolism, fatty acid metabolism and secondary metabolism were found to be upregulated by several fold. Therefore, our study suggests that calcium suppresses salt toxicity in Synechococcus sp. PCC 7942 by restricting the entry of Na⁺ into the cell, increasing osmolyte production and upregulating defence-related proteins.     

Synthesis,DNA Binding,and Photonuclease Activity of New Tetraaza Macrocyclic Constrained Isoxazole Rings as Subunit in Metal Complexes

The DNA-binding and photonuclease activity of newly synthesized tetra-azamacrocyclic ligand L (C₃₂H₃₂N₈O₄) and its complexes of type [MLCl₂] and [ML]Cl₂ (where M = Co(II), Fe(II) and Cu(II); L = N,N′-[3-(4-{5-[(2-amino-ethylamino)-methyl]-isoxazol-3yl}-phenyl)-isoxazol-5-yl methyl-ethane-1,2-diamine] are specified. An octahedral geometry has been proposed for Fe(II) and Co(II) complexes, while the Cu(II) complex has a square planar environment. The absorption spectral results indicate that the complexes bind with the base pairs of DNA, with an intrinsic binding constant K_b of Fe(II), Co(II), and Cu(II) complexes found to be 3.2 × 10⁴ M^?1, 5.3 × 10⁴ M^?1, and 4.2 × 10⁴ M^?1, respectively, in 5 mM Tris-HCl/50 mM NaCl buffer at pH 7.2. The large enhancement in the relative viscosity of DNA on binding to the complexes supports the proposed DNA binding modes. The viscosity and thermal denaturation studies sustain the effective intercalation with DNA. The DNA photocleavage studies demonstrated that compounds exhibit significant photonuclease activity by a concentration dependent on singlet oxygen mediated mechanism.