Impact of oxygen supply on rtPA expression in <Emphasis Type="Italic">Escherichia coli</Emphasis> BL21 (DE3): ammonia effects

A phytase with high activity at neutral pH and typical water temperatures (∼25°C) could effectively hydrolyze phytate in aquaculture. In this study, a phytase-producing strain, Pedobacter nyackensis MJ11 CGMCC 2503, was isolated from glacier soil, and the relevant gene, PhyP, was cloned using degenerate PCR and thermal asymmetric interlaced PCR. To our knowledge, this is the first report of detection of phytase activity and cloning of phytase gene from Pedobacter. PhyP belongs to beta-propeller phytase family and shares very low identity (∼28.5%) with Bacillus subtilis phytase. The purified recombinant enzyme (r-PhyP) from Escherichia coli displayed high specific activity for sodium phytate of 24.4 U mg⁻¹. The optimum pH was 7.0, and the optimum temperature was 45°C. The K _m, V _max, and k _cat values were 1.28 mM, 71.9 μmol min⁻¹ mg⁻¹, and 45.1 s⁻¹, respectively. Compared with Bacillus phytases, r-PhyP had higher relative activity at 25°C (r-PhyP (>50%), B. subtilis phytase (<8%)) and hydrolyzed phytate from soybean with greater efficacy at neutral pH. These characteristics suggest that r-PhyP might be a good candidate for an aquatic feed additive in the aquaculture industry.     

Characterization of an ecto-ATPase activity in Fonsecaea pedrosoi

In this work, we characterized an ecto-ATPase activity in intact mycelial forms of Fonsecaea pedrosoi, the primary causative agent of chromoblastomycosis. In the presence of 1 mM EDTA, fungal cells hydrolyzed adenosine-5′-triphosphate (ATP) at a rate of 84.6 ± 11.3 nmol Pi h⁻¹ mg⁻¹ mycelial dry weight. The ecto-ATPase activity was increased at about five times (498.3 ± 27.6 nmol Pi h⁻¹ mg⁻¹) in the presence of 5 mM MgCl₂, with values of V _max and apparent K _m for Mg-ATP²⁻corresponding to 541.9 ± 48.6 nmol Pi h⁻¹ mg⁻¹ cellular dry weight and 1.9 ± 0.2 mM, respectively. The Mg²⁺-stimulated ecto-ATPase activity was insensitive to inhibitors of intracellular ATPases such as vanadate (P-ATPases), bafilomycin A₁ (V-ATPases)_, and oligomycin (F-ATPases). Inhibitors of acid phosphatases (molybdate, vanadate, and fluoride) or alkaline phosphatases (levamizole) had no effect on the ecto-ATPase activity. The surface of the Mg²⁺-stimulated ATPase in F. pedrosoi was confirmed by assays in which 4,4′-diisothiocyanostylbene-2,2′-disulfonic acid (DIDS), a membrane impermeant inhibitor, and suramin, an inhibitor of ecto-ATPase and antagonist of P₂ purinoreceptors. Based on the differential expression of ecto-ATPases in the different morphological stages of F. pedrosoi, the putative role of this enzyme in fungal biology is discussed.     

Nucleotide-Dependent Isomerization of Glutamate Dehydrogenase in Relation to Total RNA Contents of Peanut

The physiological function of glutamate dehydrogenase (GDH) was investigated by treating germinating peanut (Arachis hypogaea L.) seeds with nucleoside triphosphate (NTP) solutions in order to alter the isoenzyme distribution patterns. The free nucleosides and nucleotides of the GTP-treated peanut were the highest [8.7 μmol g⁻¹(f.m.)], and they decreased through the ATP-treated peanut [5.8 μmol g⁻¹(f.m.)], and CTP-treated peanut [5.5 μmol g⁻¹(f.m.)], to the UTP-treated peanut [4.1 μmol g⁻¹(f.m.)]. The combination of 4 NTPs induced 20 % higher content of Pi [173 nmol g⁻¹(f.m.)] than in the control, but the combined ATP+UTP treatment induced the lowest (93.0 nmol g⁻¹(f.m.)] Pi. The 4 NTP treatment also induced the highest number of GDH isoenzymes (28) followed by the purine NTP treatments (15 to 20), but the pyrimidine NTP treatments and the combined purine + pyrimidine NTP treatments induced the lowest numbers (<15) of isoenzymes. The deamination/amination ratios were generally higher in the UTP (0.11), and CTP (0.06) treated peanuts than in the GTP (0.04), and ATP (0.07) treated peanuts. There were mutual relationships between higher numbers of GDH isoenzymes present in the GTP-, and ATP-treated peanuts and higher RNA (236.5 and 239.4 μg g⁻¹, respectively) contents on one hand, and between the lower numbers of isoenzymes in the CTP-, and UTP-treated peanuts and lower RNA (162.0 and 152.5 μg g⁻¹, respectively) contents. The recurrent relationships of the effects of the NTP treatments of peanut were UTP > ATP > CTP > GTP. This revised version was published online in July 2006 with corrections to the Cover Date.     

Expression,purification and characterization of pectate lyase A from <Emphasis Type="Italic">Aspergillus nidulans</Emphasis> in <Emphasis Type="Italic">Escherichia coli</Emphasis>

Pectate lyase A (PelA) of Aspergillus nidulans was successfully expressed in Escherichia coli and effectively purified using a Ni²⁺-nitrilotriacetate-agarose column. Enzyme activity of the recombinant PelA could reach 360 U ml⁻¹ medium. The expressed PelA exhibited its optimum level of activity over the range of pH 7.5–10 at 50°C. Mn²⁺, Ca²⁺, Fe²⁺, Mg²⁺ and Fe³⁺ ions stimulated the pectate lyase activity, but Cu²⁺ and Zn²⁺ inhibited it. The recombinant PelA had a V _max of 77 μmol min⁻¹ mg⁻¹ and an apparent K _m of 0.50 mg ml⁻¹ for polygalacturonic acid. Low-esterified pectin was the optimum substrate for the PelA, whereas higher-esterified pectin was hardly cleaved by it. PelA efficiently macerated mung bean hypocotyls and potato tuber tissues into single cells.     

Production of red pigments byMonascus purpureus in solid-state culture

To maximize and sustain the productivity ofMonascus pigments, various environmental and nutritional parameters, such as the initial moisture content, pH, inoculum size, sample size, and nutrient supplement, that influence pigment production were evaluated in solid-state cultures as follows: initial moisture content, 50%; pH, 6.0; inoculum size 1 x 10⁴ spore cells (grams of dry solid substrate)⁻¹; sample size, 300 g. All supplementary nutrients (carbon, nitrogen, and mineral sources) added has inhibitory effects on the cell growth and red pigment production. In open tray culture the maximum biomass yield and specific productivity of red pigments were 223 mg DCW (grams of initial dry substrate)⁻¹ and, 47.6 OD₅₀₀ (DCW grams)⁻¹ h⁻¹, respectively.     

The effect of co-cultivation and selection parameters on <Emphasis Type="Italic">Agrobacterium</Emphasis>-mediated transformation of Chinese soybean varieties

In the present study, an efficient Agrobacterium-mediated gene transformation system was developed for soybean [Glycine max (L.) Merrill] based on the examinations of several factors affecting plant transformation efficiency. Increased transformation efficiencies were obtained when the soybean cotyledonary node were inoculated with the Agrobacterium inoculum added with 0.02% (v/v) surfactant (Silwet L-77). The applications of Silwet L-77 (0.02%) during infection and l-cysteine (600 mg l⁻¹) during co-cultivation resulted in more significantly improved transformation efficiency than each of the two factors alone. The optimized temperature for infected explant co-cultivation was 22°C. Regenerated transgenic shoots were selected and produced more efficiently with the modified selection scheme (initiation on shoot induction medium without hygromycin for 7 days, with 3 mg l⁻¹ hygromycin for 10 days, 5 mg l⁻¹ hygromycin for another 10 days, and elongation on shoot elongation medium with 8 mg l⁻¹ hygromycin). Using the optimized system, we obtained 145 morphologically normal and fertile independent transgenic plants in five important Chinese soybean varieties. The transformation efficacies ranged from 3.8 to 11.7%. Stable integration, expression and inheritance of the transgenes were confirmed by molecular and genetic analysis. T₁ plants were analyzed and transmission of transgenes to the T₁ generation in a Mendelian fashion was verified. This optimized transformation system should be employed for efficient Agrobacterium-mediated soybean gene transformation.     

Supplementation requirements of brewery’s spent grain hydrolysate for biomass and xylitol production by Debaryomyces hansenii CCMI 941

The effect of nutrient supplementation of brewery’s spent grain (BSG) hydrolysates was evaluated with respect to biomass and xylitol production by Debaryomyces hansenii. For optimal biomass production, supplementation of full-strength BSG hydrolysates required only phosphate (0.5 g l⁻¹ KH₂PO₄), leading to a biomass yield and productivity of 0.60 g g⁻¹ monosaccharides and 0.55 g l⁻¹ h⁻¹, respectively. Under the conditions studied, no metabolic products other than CO₂ and biomass were identified. For xylitol production, fourfold and sixfold concentrated hydrolysate-based media were used to assess the supplementation effects. The type of nutrient supplementation modulated the ratio of total polyols/total extracellular metabolites as well as the xylitol/arabitol ratio. While the former varied from 0.8 to 1, the xylitol/arabitol ratio reached a maximum value of 2.6 for yeast extract (YE)-supplemented hydrolysates. The increase in xylitol productivity and yield was related to the increase of the percentage of consumed xylose induced by supplementation. The best xylitol yield and productivity were found for YE supplementation corresponding to 0.55 g g⁻¹ and 0.36 g l⁻¹ h⁻¹, respectively. In sixfold concentrated hydrolysates, providing that the hydrolysate was supplemented, the levels of xylitol produced were similar or higher than those for arabitol. Xylitol yield exhibited a further increase in the sixfold hydrolysate supplemented with trace elements, vitamins and minerals to 0.65 g g⁻¹, albeit the xylitol productivity was somewhat lower. The effect of using activated charcoal detoxification in non-supplemented versus supplemented sixfold hydrolysates was also studied. Detoxification did not improve polyols formation, suggesting that the hemicellulose-derived inhibitor levels present in concentrated BSG hydrolysates are well tolerated by D. hansenii.     

Production of inulinase by solid-state fermentation: effect of process parameters on production and preliminary characterization of enzyme preparations

This work was aimed at producing inulinase by solid-state fermentation of sugarcane bagasse, using factorial design to identify the effect of corn steep liquor (CSL) and soybean bran concentration, particle size of bagasse and size of inoculum. Maximum inulinase activity achieved was 250 U per g of dry substrate (gds) at 20% (w/w) of CSL, 5% (w/w) of soybean bran, 1 × 10¹⁰ cells mL⁻¹ and particle size of bagasse in the range 9/32 mesh. The use of soybean bran decreased the time to reach maximum activity from 96 to 24 h and the maximum productivity achieved was 8.87 U gds⁻¹ h⁻¹. The maximum activity was obtained at pH 5.0 and 55.0°C. Within the investigated range, the enzyme extract was more thermostable at 50.0°C, showing a D-value of 123.1 h and deactivation energy of 343.9 kJ gmol⁻¹. The extract showed highest stability from pH 4.5 to 4.8. Apparent K _m and V _max are 7.1 mM and 17.79 M min⁻¹, respectively.     

Hydrolysis of organic phosphates by corn and soybean roots

Because of the importance of organic phosphates as sources of P for plants, this work was performed to study the hydrolysis of nine organic phosphates by sterile, intact corn (Zea mays L.) and soybean (Glycine max L.) roots. Results showed that the rates of hydrolysis ofp-nitrophenyl phosphate (PNP) in buffered solutions by roots of three varieties of corn and three varieties of soybean ranged from 13 to 22 μmol PO₄−P g⁻¹ root h⁻¹ and from 2.1 to 2.2 μmol PO₄−P 0.1 g⁻¹ root h⁻¹, respectively. The average rate of hydrolysis of PNP in nonbuffered solutions was 2- to 3-fold lower for corn roots and 6- to 10-fold lower for soybean roots as compared with those obtained with buffered solutions. The orthophosphate released from hydrolysis of organic P compounds in buffered solutions during a 48-h incubation of corn roots showed that the maximum rate of hydrolysis of PNP was 4 to 6 times greater than the commonly used substrates: α- and β-glycerophosphates, phenolphthalein diphosphate, and glucose-6-phosphate. The rates of hydrolysis of glucose-6-phosphate and glucose-1-phosphate were similar and about 6- to 12-fold lower than that of PNP. Phosphoethanolamine and phosphocholine were hydrolyzed slightly, ando-carboxyphenyl phosphate was not hydrolyzed. The rates of hydrolysis of organic P compounds in nonbuffered solutions by corn and soybean roots were 1 to 3 and 1 to 10 times lower than those in buffered solutions, respectively. The trends in rates of hydrolysis by soybean roots of buffered organic P substrates were similar to those observed with corn roots, with the exception of glucose-1-phosphate and phosphoethanolamine.     

Competition in tree row agroforestry systems. 1. Distribution and dynamics of fine root length and biomass

Complementarity in the distribution of tree and crop root systems is important to minimise competition for resources whilst maximising resource use in agroforestry systems. A field study was conducted on a kaolinitic Oxisol in the sub-humid highlands of western Kenya to compare the distribution and dynamics of root length and biomass of a 3-year-old Grevillea robusta A. Cunn. ex R. Br. (grevillea) tree row and a 3-year-old Senna spectabilis DC. (senna) hedgerow grown with Zea mays L. (maize). Tree roots were sampled to a 300 cm depth and 525 cm distance from the tree rows, both before and after maize cropping. Maize roots were sampled at two distances from the tree rows (75–150 cm and 450–525 cm) to a maximum depth of 180 cm, at three developmental stages. The mean root length density (L_rv) of the trees in the upper 15 cm was 0.55 cm cm⁻³ for grevillea and 1.44 cm cm⁻³ for senna, at the start of the cropping season. The L_rv of senna decreased at every depth during the cropping season, whereas the L_rv of grevillea only decreased in the crop rooting zone. The fine root length of the trees decreased by about 35% for grevillea and 65% for senna, because of maize competition, manual weeding, seasonal senescence or pruning regime (senna). At anthesis, the L_rv of maize in the upper 15 cm was between 0.8 and 1.5 cm cm⁻³. Maize root length decreased with greater proximity to the tree rows, potentially reducing its ability to compete for soil resources. However, the specific root length (m g⁻¹) of maize was about twice that of the trees, so may have had a competitive uptake advantage even when tree root length was greater. Differences in maize fine root length and biomass suggest that competition for soil resources and hence fine root length may have been more important for maize grown with senna than grevillea. This revised version was published online in June 2006 with corrections to the Cover Date.     

Nutrient and water addition effects on day- and night-time conductance and transpiration in a C3 desert annual

Recent research has shown that many C₃ plant species have significant stomatal opening and transpire water at night even in desert habitats. Day-time stomatal regulation is expected to maximize carbon gain and prevent runaway cavitation, but little is known about the effect of soil resource availability on night-time stomatal conductance (g) and transpiration (E). Water (low and high) and nutrients (low and high) were applied factorially during the growing season to naturally occurring seedlings of the annual Helianthus anomalus. Plant height and biomass were greatest in the treatment where both water and nutrients were added, confirming resource limitations in this habitat. Plants from all treatments showed significant night-time g (~0.07 mol m⁻² s⁻¹) and E (~1.5 mol m⁻² s⁻¹). In July, water and nutrient additions had few effects on day- or night-time gas exchange. In August, however, plants in the nutrient addition treatments had lower day-time photosynthesis, g and E, paralleled by lower night-time g and E. Lower predawn water potentials and higher integrated photosynthetic water-use efficiency suggests that the nutrient addition indirectly induced a mild water stress. Thus, soil resources can affect night-time g and E in a manner parallel to day-time, although additional factors may also be involved.     

Differential expression of thermophilic phosphatases in the wild type and auxotrophic mutant strains of <Emphasis Type="Italic">Thermoactinomyces vulgaris</Emphasis>

In the wild type strain (stock no. 1227) of Thermoactinomyces vulgaris, as reported earlier [Sinha and Singh (1980) Biochem. J. 190, 457–460], all phosphatase isoenzymes (three alkaline — AlpI, AlpII and AlpIII, and one acidic — Acp) are present. However, the auxotrophic mutants, the strains 1286 (thi ⁻), 1279 (nic ⁻, ura ⁻) and 1278 (thi ⁻, ura ⁻) exhibited two alkaline phosphatase isoenzymes (AlpII and AlpIII), but AlpI was lacking. In the strain 1261 (nic ⁻, thi ⁻), only AlpIII was expressed, and AlpI and AlpII isoenzymes were missing. The results suggest that the strains, which require either thiamine (1286 and 1278) or nicotinamide (1279) for their growth, were AlpI ⁻ mutants; and the strain (1261), which requires both thiamine and nicotinamide for its growth, was AlpI ⁻/AlpII ⁻ double mutant. There was no direct correlation between uracil auxotrophy and the expression of phosphatases. The uniform expression of AlpIII and Acp in all the strains, irrespective of their nutrient requirements, suggest that these constitutive phosphatases are species-specific. The specific activities of the thermophilic acid and alkaline phosphatases were maximum in the wild type strain (1227) of T. vulgaris. The next in phosphatase activity was the strain 1279 (an AlpI ⁻ mutant), followed by their decrease, in order, in the strains 1286 and 1278 (which were also AlpI ⁻ mutants); while least activity of these enzymes was observed in the obligate thermophile strain 1261 (AlpI ⁻/AlpII ⁻ double mutant).     

Enhanced iturin A production by <Emphasis Type="Italic">Bacillus subtilis</Emphasis> and its effect on suppression of the plant pathogen <Emphasis Type="Italic">Rhizoctonia solani</Emphasis>

The behavior of Streptomyces peucetius var. caesius N47 was studied in a glucose limited chemostat with a complex cultivation medium. The steady-state study yielded the characteristic constants μ _max over 0.10 h⁻¹, Y _XS 0.536 g g⁻¹, and m_S 0.54 mg g⁻¹ h⁻¹. The product of secondary metabolism, ɛ-rhodomycinone, was produced with characteristics Y _PX 12.99 mg g⁻¹ and m _P 1.20 mg g⁻¹ h⁻¹. Significant correlations were found for phosphate and glucose consumption with biomass and ɛ-rhodomycinone production. Metabolic flux analysis was conducted to estimate intracellular fluxes at different dilution rates. TCA, PPP, and shikimate pathway fluxes exhibited bigger values with production than with growth. Environmental perturbation experiments with temperature, airflow, and pH changes on a steady-state chemostat implied that an elevation of pH could be the most effective way to shift the cells from growing to producing, as the pH change induced the biggest transient increase to the calculated ɛ-rhodomycinone flux.     

Biofiltering efficiency, uptake and assimilation rates of Ulva clathrata (Roth) J. Agardh (Clorophyceae) cultivated in shrimp aquaculture waste water

The growth, biofiltering efficiency and uptake rates of Ulva clathrata were studied in a series of outdoor tanks, receiving waste water directly from a shrimp (Litopenaeus vannamei) aquaculture pond, under constant aeration and two different water regimes: (1) continuous flow, with 1 volume exchange a day (VE day^-1) and (2) static regime, with 1 VE after 4 days. Water temperature, salinity, pH, dissolved inorganic nitrogen (DIN), phosphate (PO₄), chlorophyll-a (chl-a), total suspended solids (TSS), macroalgal biomass (fresh weight) and tissue nutrient assimilation were monitored over 12 days. Ulva clathrata was highly efficient in removing the main inorganic nutrients from effluent water, stripping 70–82% of the total ammonium nitrogen (TAN) and 50% PO₄ within 15 h. Reductions in control tanks were much lower (Tukey HSD, P < 0.05). After 3 days, the mean uptake rates by the seaweed biomass under continuous flow were 3.09 mg DIN g DW day⁻¹ (383 mg DIN m⁻² day⁻¹) and 0.13 mg PO₄ g DW day⁻¹ (99 mg PO₄ m⁻² day⁻¹), being significantly higher than in the static regime (Tukey HSD, P < 0.05). The chl-a decreased in seaweed tanks, suggesting that U. clathrata inhibited phytoplankton growth. Correlations between the cumulative values of DIN removed from the water and total nitrogen assimilated into the seaweed biomass (r = 0.7 and 0.8, P < 0.05), suggest that nutrient removal by U. clathrata dominated over other processes such as phytoplankton and bacterial assimilation, ammonia volatilization and nutrient precipitation.     

Improvement of artemisinin production by chitosan in hairy root cultures of <Emphasis Type="Italic">Artemisia annua</Emphasis> L.

Artemisinin production by hairy roots of Artemisia annua L. was increased 6-fold to 1.8 μg mg⁻¹ dry wt over 6 days by adding 150 mg chitosan l⁻¹. The increase was dose-dependent. Similar treatment of hairy roots with methyl jasmonate (0.2 mM) or yeast extract (2 mg ml⁻¹) increased artemisinin production to 1.5 and 0.9 μg mg⁻¹ dry wt, respectively.     

A novel highly acidic β-mannanase from the acidophilic fungus Bispora sp. MEY-1: gene cloning and overexpression in Pichia pastoris

Using degenerate polymerase chain reaction (PCR) and thermal asymmetric interlaced PCR, a 1,347-bp full-length complementary DNA fragment encompassing the gene man5A, which encodes a 429-amino acid β-mannanase with a calculated mass of 46.8 kDa, was cloned from acidophilic Bispora sp. MEY-1. The deduced amino acid sequence (catalytic domain) displayed highest identity (54.1%) with the Emericella nidulans endo-β-1,4-d-mannanase, a member of the glycoside hydrolase family 5. Recombinant MAN5A was overexpressed in Pichia pastoris, and its activity in the culture medium reached 500 U ml⁻¹. The enzyme was acidophilic, with highest activity at pH 1.0–1.5, lower than any known mannanases, and optimal temperature for activity was 65°C. MAN5A had good pH adaptability, excellent thermal and pH stability, and high resistance to both pepsin and trypsin. The specific activity, K _m, and V _max for locust bean gum substrate was 3,373 U mg⁻¹, 1.56 mg ml⁻¹, and 6,587.6 μmol min⁻¹ mg⁻¹, respectively. The enzymatic activity was not significantly affected by ions such as Ca²⁺, Cr³⁺, Co²⁺, Zn²⁺, Na⁺, K⁺, and Mg²⁺ and enhanced by Ni²⁺, Fe³⁺, Mn²⁺ and Ag⁺. These favorable properties make MAN5A a potential candidate for use in various industrial applications.     

Purification and Characterization of the Glutathione-S-transferases from the Northern Quahog Mercinaria mercinaria

Glutathione-S-transferase (GST) was isolated from the northern hardshell clam Mercinaria mercinaria (quahog) using a two-step procedure involving ammonium sulfate precipitation and affinity chromatography. Kinetic analysis of the purified enzyme using 1-chloro-2,4-dinitrobenzene as substrate revealed a specific activity of 38.0 μmol min⁻¹ mg⁻¹, while V _max and K _m values were estimated as 48.0 μmol min⁻¹ mg⁻¹ and 0.24 mM, respectively. Electrophoretic analysis of GST indicated multiple forms of the dimeric enzyme in quahogs with subunit molecular masses of 22, 24, 25, and 27 kDa. Isoelectric focusing analysis resulted in pI values for three isoenzymes of 5.1, 4.9, and 4.6. The acidic pI values obtained indicated that quahog GST belongs to the π class. Inhibition of quahog GST by tetrapyrroles was similar to that of GST from oyster and rat liver. Quantitative comparison of tetrapyrrole inhibition patterns of quahog GST with those of oyster and rat liver GST indicated lower inhibition rates by three of the four tetrapyrroles tested (bilirubin, biliverdin, and chlorophillyin), suggesting that quahog GST could differ structurally or functionally from oyster and rat liver GSTs. Received March 17, 1998; accepted August 18, 1998.     

Characterisation of acidic sites of Pseudomonas biomass capable of binding protons and cadmium and removal of cadmium via biosorption

Summary The ability of Pseudomonas aeruginosa to accumulate Cd(II) ions from wastewater industries was experimentally investigated and mathematically modelled. From the potentiometric titration and non-ideal competitive analysis (NICA) model, it was found that the biomass contains three acidic sites. The values of proton binding (pK _i =1.66±3.26×10⁻³, 1.92±1.63×10⁻⁴ and 2.16±3.79×10⁻⁴) and binding constant of cadmium metal ions (pK _M1=1.99±2.45×10⁻³ and pK _M2=1.67±4.08×10⁻³) on the whole surface of biomass showed that protonated functional groups and biosorption of Cd(II) ions could be attributed to a monodentate binding to one acidic site, mainly the carboxylic group. From the isothermal sorption experimental data and Langmuir model, it was also found that the value of Langmuir equilibrium (pK _f) constant is 2.04±2.1×10⁻⁵ suggesting that the carboxyl group is the main active binding site. In addition, results showed that the maximum cadmium capacity (q _max) and affinity of biomass towards cadmium metal ions (b) at pH 5.1 and 20 min were 96.5±0.06 mg/g and 3.40×10⁻³± 2.10×10⁻³, respectively. Finally, interfering metal ions such as Pb(II), Cu(II), Cr(III), Zn(II), Fe(II), Mn(II), Ca(II) and Mg(II) inhibited Cd(II) uptake. Comparing the biosorption of Cd(II) by various Pseudomonas isolates from contaminated environment samples (soil and sewage treatment plant) showed that maximum capacities and equilibrium times were different, indicating that there was a discrepancy in the chemical composition between biomasses of different strains.     

Purification and characterization of dihydroorotase fromPseudomonas putida

Dihydroorotase was purified to homogeneity fromPseudomonas putida. The relative molecular mass of the native enzyme was 82 kDa and the enzyme consisted of two identical subunits with a relative molecular mass of 41 kDa. The enzyme only hydrolyzed dihydro-l-orotate and its methyl ester, and the reactions were reversible. The apparentK _m andV _max values for dihydro-l-orotate hydrolysis (at pH 7.4) were 0.081 mM and 18 μmol min⁻¹ mg⁻¹, respectively; and those forN-carbamoyl-dl-aspartate (at pH 6.0) were 2.2 mM and 68 μmol min⁻¹ mg⁻¹, respectively. The enzyme was inhibited by metal ion chelators and activated by Zn²⁺. However, excessive Zn²⁺ was inhibitory. The enzyme was inhibited by sulfhydryl reagents, and competitively inhibited byN-carbamoylamino acids such asN-carbamoylglycine, with aK _i value of 2.7 mM. The enzyme was also inhibited noncompetitively by pyrimidine-metabolism intermediates such as dihydrouracil and orotate, with aK _i value of 3.4 and 0.75 mM, respectively, suggesting that the enzyme activity is regulated by pyrimidine-metabolism intermediates and that dihydroorotase plays a role in the control of pyrimidine biosynthesis.     

A coupled model of stomatal conductance and photosynthesis for winter wheat

The model couples stomatal conductance (g _s) and net photosynthetic rate (P _N) describing not only part of the curve up to and including saturation irradiance (I _max), but also the range above the saturation irradiance. Maximum stomatal conductance (g _smax) and I _max can be calculated by the coupled model. For winter wheat (Triticum aestivum) the fitted results showed that maximum P _N (P _max) at 600 μmol mol⁻¹ was more than at 350 μmol mol⁻¹ under the same leaf temperature, which can not be explained by the stomatal closure at high CO₂ concentration because g _smax at 600 μmol mol⁻¹ was less than at 350 μmol mol⁻¹. The irradiance-response curves for winter wheat had similar tendency, e.g. at 25 °C and 350 μmol mol⁻¹ both P _N and g _s almost synchronously reached the maximum values at about 1 600 μmol m⁻² s⁻¹. At 25 °C and 600 μmol mol⁻¹ the I _max corresponding to P _max and g _smax was 2 080 and 1 575 μmol m⁻² s⁻¹, respectively.