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1.
Kim HT  Ko HJ  Kim N  Kim D  Lee D  Choi IG  Woo HC  Kim MD  Kim KH 《Biotechnology letters》2012,34(6):1087-1092
A gene, alg7D, from Saccharophagus degradans, coding for a putative alginate lyase belonging to the family of polysaccharide lyase-7, was overexpressed in Escherichia coli. The properties of the recombinant Alg7D were characterized. The enzyme endolytically depolymerized alginate by β-elimination into oligo-alginates with degrees of polymerization of 2–5. Its activity was maximal at 50°C and pH 7 and was slightly increased in the presence of Na+. The K M , V max , k cat , and k cat /K M values were: 3 mg ml−1, 6.2 U mg−1, 1.9 × 10−2 s−1, and 6.3 × 10−3 mg−1 ml s−1, respectively.  相似文献   

2.
A chemically defined medium for mycelial growth and exopolysaccharide (EPS) production by submerged culture of Phellinus igniarius was investigated. The mainly defined medium compositions were optimized by using orthogonal matrix method. The optimal defined medium (per liter) was 40.0 g glucose, 4.0 g. glutamic acid, 4.0 g (NH4)2SO4, and initial pH 6.0. Under the optimal medium, the maximal mycelial biomass and EPS production were 12.33 ± 0.89 and 1.21 ± 0.08 g l−1 at 192 h in shake flask, while the maximal mycelial biomass and EPS production reached 13.86 ± 0.52 and 1.92 ± 0.07 g l−1 at 168 h in 3 l fermenter, respectively. The molecular weights (g mol−1) of four fractions isolated from EPS by gel permeation were about 6.4 × 106, 3.3 × 105, 2.7 × 105 and 2.9 × 103. This study should be widely applied to other secondary metabolites production from higher fungus in a chemically defined medium and quantitative regulation of the metabolic flux in polysaccharide biosynthesis.  相似文献   

3.
The activity of Na+/H+ exchanger to remove toxic Na+ is important for growth of organisms under high salinity. In this study, the halotolerant cyanobacterium Aphanothece halophytica was shown to possess Na+/H+ exchange activity since exogenously added Na+ could dissipate a pre-formed pH gradient, and decrease extracellular pH. Kinetic analysis yielded apparent K m (Na+) and V max of 20.7 ± 3.1 mM and 3,333 ± 370 nmol H+ min−1 mg−1, respectively. For cells grown under salt-stress condition, the apparent K m (Na+) and V max was 18.3 ± 3.5 mM and 3,703 ± 350 nmol H+ min−1 mg−1, respectively. Three cations with decreasing efficiency namely Li+, Ca2+, and K+ were also able to dissipate pH gradient. Only marginal exchange activity was observed for Mg2+. The exchange activity was strongly inhibited by Na+-gradient dissipators, monensin, and sodium ionophore as well as by CCCP, a protonophore. A. halophytica showed high Na+/H+ exchange activity at neutral and alkaline pH up to pH 10. Cells grown at pH 7.6 under high salinity exhibited higher Na+/H+ exchange activity than those grown under low salinity during 15 days of growth suggesting a role of Na+/H+ exchanger for salt tolerance in A. halophytica. Cells grown at alkaline pH of 9.0 also exhibited a progressive increase of Na+/H+ exchange activity during 15 days of growth.  相似文献   

4.
An alkaline protease from marine Engyodontium album was characterized for its physicochemical properties towards evaluation of its suitability for potential industrial applications. Molecular mass of the enzyme by matrix-assisted laser desorption ionization-mass spectrometry (MALDI-MS) analysis was calculated as 28.6 kDa. Isoelectric focusing yielded pI of 3–4. Enzyme inhibition by phenylmethylsulfonyl fluoride (PMSF) and aprotinin confirmed the serine protease nature of the enzyme. K m, V max, and K cat of the enzyme were 4.727 × 10−2 mg/ml, 394.68 U, and 4.2175 × 10−2 s−1, respectively. Enzyme was noted to be active over a broad range of pH (6–12) and temperature (15–65°C), with maximum activity at pH 11 and 60°C. CaCl2 (1 mM), starch (1%), and sucrose (1%) imparted thermal stability at 65°C. Hg2+, Cu2+, Fe3+, Zn2+, Cd+, and Al3+ inhibited enzyme activity, while 1 mM Co2+ enhanced enzyme activity. Reducing agents enhanced enzyme activity at lower concentrations. The enzyme showed considerable storage stability, and retained its activity in the presence of hydrocarbons, natural oils, surfactants, and most of the organic solvents tested. Results indicate that the marine protease holds potential for use in the detergent industry and for varied applications.  相似文献   

5.
Dey K  Roy P 《Biotechnology letters》2011,33(6):1101-1105
A Bacillus sp., capable of degrading chloroform, was immobilized in calcium alginate. The beads in 20 g alginate l−1 (about 2 × 108 cells/bead) could be re-used nine times for degradation of chloroform at 40 μM. The immobilized cells had a higher range of tolerance (pH 6.5–9 and 20–41°C) than free cells (pH 7–8.5 and 28–32°C). At 5 g alginate l−1, leakage of the cells from the beads was 0.51 mg dry wt ml−1. This species is the first reported Bacillus that can degrade chloroform as the sole carbon source.  相似文献   

6.
Maltose and yeast extract were the most favourable carbon and nitrogen sources for exopolysaccharide production by submerged culture of Shiraia bambusicola WZ-003, and initial maltose and yeast extract concentrations were at 30 and 3 g l−1, respectively. Plant oils could increase the mycelial growth and exopolysaccharide production in tested concentration. K+ and Mg2+ could enhance the mycelial growth and exopolysaccharide biosynthesis. The optimal cultivation temperature and initial pH were found to be 26°C and 6.0, respectively. Exopolysaccharide concentration reached 0.53 g l−1 in 15-l fermenter under optimal nutritional conditions.  相似文献   

7.
A gene from Withania somnifera (winter cherry), encoding a highly stable chloroplastic Cu/Zn superoxide dismutase (SOD), was cloned and expressed in Escherichia coli. The recombinant enzyme (specific activity of ~4,200 U mg−1) was purified and characterized. It retained ~90 and ~70% residual activities after 1 h at 80 and 95°C, respectively. At 95°C, thermal inactivation rate constant (K d) of the enzyme was 2.46 × 10−3 min−1 and half-life of heat inactivation was 4.68 h. The enzyme was stable against a broad pH range (2.5–11.0). It also showed a high degree of resistance to detergent, ethanol and protease digestion. This recombinant Cu/Zn SOD could therefore have useful applications.  相似文献   

8.
We investigated the effects of limiting (1.96 × 10−9 mol l−1 total Cu, corresponding to pCu 14.8; where pCu = −log [Cu2+]) and toxic Cu concentrations up to 8.0 × 10−5 mol l−1 total Cu (equivalent to pCu 9.5) on growth rates and photosynthetic activity of exponentially grown Phaeocystis cordata, using batch and semi-continuous cultures. With pulse amplitude modulated (PAM) fluorometry, we determined the photochemical response of P. cordata to the various Cu levels, and showed contrasting results for the batch and semi-continuous cultures. Although maximum photosystem II (PSII) quantum yield (ΦM) was optimal and constant in the semi-continuous P. cordata, the batch cultures showed a significant decrease in ΦM with culture age (0–72 h). The EC50 for the batch cultures was higher (2.0 × 10−10 mol l−1, pCu9.7), than that for the semi-continuous cultures (6.3 × 10−11 mol l−1, pCu10.2). The semi-continuous cultures exhibited a systematic and linear decrease in ΦM as Cu levels increased (for [Cu2+] < 1.0 × 10−12 mol l−1, pCu12.0), however, no effect of high Cu was observed on their operational PSII quantum yield (Φ′M). Similarly, semi-continuous cultures exhibited a significant decrease in ΦM, but not in Φ′M, because of low-Cu levels. Thus, Cu toxicity and Cu limitation damage the PSII reaction centers, but not the processes downstream of PSII. Quenching mechanisms (NPQ and Q n) were lower under high Cu relative to the controls, suggesting that toxic Cu impairs photo-protective mechanisms. PAM fluorometry is a sensitive tool for detecting minor physiological variations. However, culturing techniques (batch vs. semi-continuous) and sampling time might account for literature discrepancies on the effects of Cu on PSII. Semi-continuous culturing might be the most adequate technique to investigate Cu effects on PSII photochemistry.  相似文献   

9.
Withania somnifera is an important medicinal plant that contains withanolides and withaferins, both bioactive compounds. We have tested the effects of macroelements and nitrogen source in W. somnifera cell suspension cultures with the aim of optimizing the production of biomass and withanolide A. The effects of the macroelements NH4NO3, KNO3, CaCl2, MgSO4 and KH2PO4 at concentrations of 0.0, 0.5, 1.0, 1.5 and 2.0× strength and of the nitrogen source [NH4 +/NO3 (mM/mM) ratio of: 0.00/18.80, 7.19/18.80, 14.38/18.80, 21.57/18.80, 28.75/18.80, 14.38/0.00, 14.38/9.40, 14.38/18.80, 14.38/28.20, and 14.38/37.60 (mM)] in Murashige and Skoog medium were tested for biomass and withanolide A production. The highest accumulation of biomass [147.81 g l−1 fresh weight (FW) and 14.02 g l−1 (dry weight (DW)] was recorded in the medium containing a 0.5× concentration of NH4NO3, and the highest production of withanolide A content was recorded in the medium with 2.0× KNO3 (4.36 mg g−1 DW). The NH4 +/NO3 ratio also influenced cell growth and withanolide A production, with both parameters being larger when the NO3 concentration was higher than that of NH4 +. Maximum biomass growth (110.45 g l−1 FW and 9.29 g l−1 DW) was achieved at an NH4 +/NO3 ratio of 7.19/18.80, while withanolide A production was greatest (3.96 mg g−1 DW) when the NH4 +/NO3 ratio was 14.38/37.60 mM.  相似文献   

10.
This article reports rate constants for thiol–thioester exchange (k ex), and for acid-mediated (k a), base-mediated (k b), and pH-independent (k w) hydrolysis of S-methyl thioacetate and S-phenyl 5-dimethylamino-5-oxo-thiopentanoate—model alkyl and aryl thioalkanoates, respectively—in water. Reactions such as thiol–thioester exchange or aminolysis could have generated molecular complexity on early Earth, but for thioesters to have played important roles in the origin of life, constructive reactions would have needed to compete effectively with hydrolysis under prebiotic conditions. Knowledge of the kinetics of competition between exchange and hydrolysis is also useful in the optimization of systems where exchange is used in applications such as self-assembly or reversible binding. For the alkyl thioester S-methyl thioacetate, which has been synthesized in simulated prebiotic hydrothermal vents, k a = 1.5 × 10−5 M−1 s−1, k b = 1.6 × 10−1 M−1 s−1, and k w = 3.6 × 10−8 s−1. At pH 7 and 23°C, the half-life for hydrolysis is 155 days. The second-order rate constant for thiol–thioester exchange between S-methyl thioacetate and 2-sulfonatoethanethiolate is k ex = 1.7 M−1 s−1. At pH 7 and 23°C, with [R″S(H)] = 1 mM, the half-life of the exchange reaction is 38 h. These results confirm that conditions (pH, temperature, pK a of the thiol) exist where prebiotically relevant thioesters can survive hydrolysis in water for long periods of time and rates of thiol–thioester exchange exceed those of hydrolysis by several orders of magnitude.  相似文献   

11.
Copper (II) complex of formulation [Cu–Phen–Tyr](H2O)](ClO4) (Phen = 1,10-phenanthroline, l-Tyr = l-tyrosine), has been prepared, and their induced DNA oxidative cleavage activity studied. The complex binds to DNA by intercalation, as deduced from the absorption and fluorescence spectral data. Scatchard plots constructed from the absorption titration data gave binding constant 2.44 × 104 M−1 of base pairs. Extensive hypochromism, broadening, and red shifts in the absorption spectra were observed. Upon binding to DNA, the fluorescence from the DNA–ethidium bromide system was efficiently quenched by the copper (II) complex. Stern–Volmer quenching constant 0.61 × 103 M−1 obtained from the linear quenching plots. [Cu–Phen–Tyr] complex efficiently cleave the supercoiled DNA to its nicked circular form with gallic acid as biological reductant at appropriate complex concentration. The gallic acid as reductant could observably improve copper (II) complex to DNA damage. The pseudo-Michaelis–Menten kinetic parameters (k cat, K M) were calculated to be 1.32 h−1 and 5.46 × 10−5 M for [Cu–Phen–Tyr] complex. Mechanistic studies reveal the involvement of superoxide anions and hydroxyl radical (HO·) as the reactive species under an aerobic medium.  相似文献   

12.
Interfacial tension has been determined for phosphatidylcholine (PC)–decanoic acid (DA) and PC–decylamine (DE) membranes. PC (lecithin), DA and DE were used in the experiments; the interfacial tension values of the pure components are 1.62 × 10−3, −2.38 × 10−2 and −3.88 × 10−2 N/m (hypothetical values for DA and DE), respectively. The 1:1 complexes were formed during formation of PC–DA and PC–DE membranes. The following parameters describing the complexes were determined: the surface concentrations of the lipid membranes formed from these complexes, A3 - 1 A_{3}^{ - 1} ; the interfacial tensions of such membranes, γ 3; and the stability constants of these complexes, K.  相似文献   

13.
This study reports on the optimization of protoplast yield from two important tropical agarophytes Gracilaria dura and Gracilaria verrucosa using different cell-wall-degrading enzymes obtained from commercial sources. The conditions for achieving the highest protoplast yield was investigated by optimizing key parameters such as enzyme combinations and their concentrations, duration of enzyme treatment, enzyme pH, mannitol concentration, and temperature. The significance of each key parameter was also further validated using the statistical central composite design. The enzyme composition with 4% cellulase Onozuka R-10, 2% macerozyme R-10, 0.5% pectolyase, and 100 U agarase, 0.4 M mannitol in seawater (30‰) adjusted to pH 7.5 produced the highest protoplast yields of 3.7 ± 0.7 × 106 cells g−1 fresh wt for G. dura and 1.2 ± 0.78 × 106 cells g−1 fresh wt for G. verrucosa when incubated at 25°C for 4–6 h duration. The young growing tips maximally released the protoplasts having a size of 7–15 μm in G. dura and 15–25 μm in G. verrucosa, mostly from epidermal and upper cortical regions. A few large-size protoplasts of 25–35 μm, presumably from cortical region, were also observed in G. verrucosa.  相似文献   

14.
The increasing emergence of antibiotic-resistant bacteria has produced a growing interest among scientists in bacteriophages as alternative antimicrobial agents. This article reports a lytic phage against an antibiotic-resistant strain of Pseudomonas aeruginosa. Phage BVPaP-3 is a member of the Podoviridae family and morphologically similar to the T7-like phage gh-1. The phage has a hexagonal head of 58–59 nm in diameter and a short tail of 10 × 8 nm. It is stable at a wide range of pH (6–10) and temperatures (4–40°C). Its optimal growth temperature is 37°C and the adsorption rate constant is 1.19 × 10−9. Latent and eclipse periods are 20 and 15 min, respectively, and the burst size is 44 after 35 min at 37°C. The phage has a DNA size of 41.31 kb and a proteome of 11 proteins. The major protein is 33 kDa in size.  相似文献   

15.
An endoglucanase (1, 4-β-d glucan glucanohydrolase, EC 3.2.1.4) which was catalytically more active and exhibited higher affinity towards barley β-glucan, xyloglucan and lichenin as compared to carboxymethylcellulose (CMC) was purified from Aspergillus terreus strain AN1 following ion-exchange and hydrophobic interaction chromatography and gel filtration. The purified enzyme (40-fold) that apparently lacked a cellulose-binding domain showed a specific activity of 60 μmol mg−1 protein−1 against CMC. The purified enzyme had a molecular weight of 78 and 80 KDa as indicated by sodium dodecyl sulphate–polyacrylamide gel electrophoresis and gel filtration, respectively, and a pI of 3.5. The enzyme was optimally active at temperature 60°C and pH 4.0, and was stable over a broad range of pH (3.0–5.0) at 50°C. The endoglucanase activity was positively modulated in the presence of Cu2+, Mg2+, Ca2+, Na+, DTT and mercaptoethanol. Endoglucanase exhibited maximal turn over number (K cat) and catalytic efficiency (K cat/km) of 19.11 × 105 min−1 and 29.7 × 105 mM−1 min−1 against barley β-glucan as substrate, respectively. Hydrolysis of CMC and barley β-glucan liberated cellobiose, cellotriose, cellotetraose and detectable amount of glucose. The hydrolysis of xyloglucan, however, apparently yielded positional isomers of cellobiose, cellotriose and cellotetraose as well as larger oligosaccharides.  相似文献   

16.
Surface inoculation dose–response and time–response bioassays and detached fruit bioassays were conducted with a novel South African isolate of the Cryptophlebia leucotreta granulovirus (CrleGV-SA) against Thaumatotibia leucotreta (Meyrick) (Lepidoptera: Noctuidae) neonate larvae. LC50 and LC90 values were estimated to be 4.095 × 103 and 1.185 × 105 OBs ml−1, respectively. LT50 and LT90 values were estimated to be 4 days 22 h and 7 days 8 h, respectively, categorising the virus as a fast or type 2 granulovirus. There was a conspicuous difference in behaviour between larvae on inoculated diet and untreated diet, resulting in a significant reduction in penetration of diet. Bioassays on detached Navel oranges revealed LC50 and LC90 values of 9.310 × 107 and 1.515 × 109 OBs ml−1, when using data on numbers of larvae per fruit rather than on numbers of infested fruit. Field trials will be conducted.  相似文献   

17.
Simultaneous measurements of chlorophyll (Chl) fluorescence and CO2 assimilation (A) in Vicia faba leaves were taken during the first weeks of growth to evaluate the protective effect of 24-epibrassinolide (EBR) against damage caused by the application of the herbicide terbutryn (Terb) at pre-emergence. V. faba seeds were incubated for 24 h in EBR solutions (2 × 10−6 or 2 × 10−5 mM) and immediately sown. Terb was applied at recommended doses (1.47 or 1.96 kg ha−1) at pre-emergence. The highest dose of Terb strongly decreased CO2 assimilation, the maximum quantum yield of PSII photochemistry in the dark-adapted state (F V/F M), the nonphotochemical quenching (NPQ), and the effective quantum yield (ΔF/FM) during the first 3–4 weeks after plant emergence. Moreover, Terb increased the basal quantum yield of nonphotochemical processes (F 0/F M), the degree of reaction center closure (1 − q p), and the fraction of light absorbed in PSII antennae that was dissipated via thermal energy dissipation in the antennae (1 − FV/FM). The herbicide also significantly reduced plant growth at the end of the experiment as well as plant length, dry weight, and number of leaves. The application of EBR to V. faba seeds before sowing strongly diminished the effect of Terb on fluorescence parameters and CO2 assimilation, which recovered 13 days after plant emergence and showed values similar to those of control plants. The protective effect of EBR on CO2 assimilation was detected at a photosynthetic photon flux density (PFD) of 650 μmol m−2 s−1 and the effect on ΔF/FM and photosynthetic electron transport (J) was detected under actinic lightings up to 1750 μmol m−2 s−1. The highest dose of EBR also counteracted the decrease in plant growth caused by Terb, and plants registered the same growth values as controls.  相似文献   

18.
The effects of temperature, irradiance, and daylength on Sargassum horneri growth were examined at the germling and adult stages to discern their physiological differences. Temperature–irradiance (10, 15, 20, 25, 30°C × 20, 40, 80 μmol photons m−2s−1) and daylength (8, 12, 16, 24 h) experiments were carried out. The germlings and blades of S. horneri grew over a wide range of temperatures (10–25°C), irradiances (20–80 μmol photons m−2s−1), and daylengths (8–24 h). At the optimal growth conditions, the relative growth rates (RGR) of the germlings were 21% day−1 (25°C, 20 μmol photons m−2s−1) and 13% day−1 (8 h daylength). In contrast, the RGRs of the blade weights were 4% day−1 (15°C, 20 μmol photons m−2s−1) and 5% day−1 (12 h daylength). Negative growth rates were found at 20 μmol photons m−2s−1 of 20°C and 25°C treatments after 12 days. This phenomenon coincides with the necrosis of S. horneri blades in field populations. In conclusion, we found physiological differences between S. horneri germlings and adults with respect to daylength and temperature optima. The growth of S. horneri germlings could be enhanced at 25°C, 20 μmol photons m−2s−1, and 8 h daylength for construction of Sargassum beds and restoration of barren areas.  相似文献   

19.
The auxin formation in a submerged culture of the xylotrophic basidiomycete Lentinus edodes (Berk.) Sing (Lentinula edodes (Berk.) Pegler) (shiitake) is studied. Biologically active substances of an indole nature are identified, “the effect of small doses” of which lies in not only the stimulation of growth of the mycelium (indole-3-acetic acid, 2 × 10−7–2 × 10−4 g/l), but also in the induction of tryptophan-independent paths of auxin biosynthesis. The above-mentioned path is realized in the presence of exogenous indole (1 × 10−3–1 × 10−4 g/l), as well as while inducing the biosynthesis of indole-3-acetic acid by its microadditives (1 × 10−5−1 × 10−8 g/l), and is accompanied by the formation of anthranilic acid (up to 1.5 mg/l). Induction of the generative development stage of shiitake by indole derivatives is revealed. It was found that among the studied compounds only indoleacetamide at a concentration of an order of ×10−4 g/l in the culture fluid of L. edodes had a pronounced stimulatory effect on the formation of shiitake’s brown mycelial film.  相似文献   

20.
Summary The use of Phanerochaete chrysosporium biomass for the removal of Reactofix Golden Yellow from aqueous solution and eight textile dyes (four azo and four anthraquinone) from a synthetic effluent (0.6 g/l) at different pH, temperature and biomass concentrations was studied. Adsorption was maximum at pH 2.0 and 40 °C using 2.45 g mycelial biomass. The rate constant of adsorption was 1.95×10−1/min for Reactofix Golden Yellow and 1.64×10−1/min for synthetic effluent. In both cases, the equilibrium data fitted well in the Langmuir but not the Freundlich model of adsorption, and the adsorption was biphasic. Adsorption decreased the COD of Reactofix Golden Yellow and synthetic effluent by 54 and 57%, respectively. Desorption (80–84%) of dyes from P. chrysosporium mycelial surface occurred as the pH increased from 2 to 10.  相似文献   

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