Binding and regulatory properties of phosphofructokinase from swine kidney

The influence of fructose 2,6-bisphosphate on the activation of purified swine kidney phosphofructokinase as a function of the concentration of fructose 6P, ATP and citrate was investigated. The purified enzyme was nearly completely inhibited in the presence of 2 mM ATP. The addition of 20 nM fructose 2,6-P2 reversed the inhibition and restored more than 80% of the activity. In the absence of fructose 2,6-P2 the reaction showed a sigmoidal dependence on fructose-6-phosphate. The addition of 10 nM fructose 2,6-bisphosphate decreased the K0.5 for fructose 6-phosphate from 3 mM to 0.4 mM in the presence of 1.5 mM ATP. These results clearly show that fructose 2,6-bisphosphate increases the affinity of the enzyme for fructose 6-phosphate and decreases the inhibitory effect of ATP. The extent of inhibition by citrate was also significantly decreased in the presence of fructose 2,6-phosphate. The influence of various effectors of phosphofructokinase on the binding of ATP and fructose 6-P to the enzyme was examined in gel filtration studies. It was found that kidney phosphofructokinase binds 5.6 moles of fructose 6-P per mole of enzyme, which corresponds to about one site per subunit of tetrameric enzyme. The KD for fructose 6-P was 13 microM and in the presence of 0.5 mM ATP it increased to 27 microM. The addition of 0.3 mM citrate also increased the KD for fructose 6-P to about 40 microM. AMP, 10 microM, decreased the KD to 5 microM and the addition of fructose 2,6-phosphate decreased the KD for fructose 6-P to 0.9 microM. The addition of these compounds did not effect the maximal amount of fructose 6-P bound to the enzyme, which indicated that the binding site for these compounds might be near, but was not identical to the fructose 6-P binding site. The enzyme bound a maximum of about 12.5 moles of ATP per mole, which corresponds to 3 moles per subunit. The KD of the site with the highest affinity for ATP was 4 microM, and it increased to 15 microM in the presence of fructose 2,6-bisphosphate. The addition of 50 microM fructose 1,6-bisphosphate increased the KD for ATP to 5.9 microM. AMP increased the KD to 5.9 microM whereas 0.3 mM citrate decreased the KD for ATP to about 2 microM.(ABSTRACT TRUNCATED AT 400 WORDS).

Algal Photosynthetic Dynamics in Urban Lakes under Stress Conditions

Urban lakes form vital ecosystems supporting livelihood with social, economic and aesthetic benefits that are essential for quality life. This depends on the biotic and abiotic components in an ecosystem. The structure of an ecosystem forms a decisive factor in sustaining its functional abilities which include nutrient cycling, oxygen production, etc. A community assemblage of primary producers (algae) plays a crucial role in maintaining the balance as they form the base of energy pyramid in the ecosystem. Algae assimilate carbon in the environment via photosynthetic activities and releases oxygen for the next level of biotic elements in an ecosystem. Besides these, algal cells rich in protein serve as food and feed, used as manure and for production of biofuels. Understanding algal photosynthetic dynamics helps in assessing the level of dissolved oxygen (DO), food (fish, etc.), waste assimilation, etc. Algal chlorophyll content, algal biomass, primary productivity and algal photosynthetic quotient are some of the parameters that help in assessing the status of urban lakes. Chlorophyll content gives a measure of the growth, spread and quantity of algae. Unplanned rapid urbanization in Bangalore in recent times has resulted in either disappearance of lake ecosystems or deteriorated the lake water quality impairing the ecological processes. This paper computes algal growth, community structure, primary productivity and composition for three major lakes (T G Halli, Bellandur and Varthur lakes) under contrast levels of anthropogenic influences.

Role of degree of saturation in denitrification of unsaturated sand specimens

Nitrate contamination of groundwater arises from anthropogenic activities, such as, fertilizer and animal manure applications and infiltration of wastewater/leachates. During migration of wastewater and leachates, the vadose zone (zone residing above the groundwater table), is considered to facilitate microbial denitrification. Particle voids in vadose zone are deficient in dissolved oxygen as the voids are partially filled by water and the remainder by air. Discontinuities in liquid phase would also restrict oxygen diffusion and therefore facilitate denitrification in the vadose/unsaturated soil zone. The degree of saturation of soil specimen (S (r)) quantifies the relative volume of voids filled with air and water. Unsaturated specimens have S (r) values ranging between 0 and 100 %. Earlier studies from naturally occurring nitrate losses in groundwater aquifers in Mulbagal town, Kolar District, Karnataka, showed that the sub-surface soils composed of residually derived sandy soil; hence, natural sand was chosen in the laboratory denitrification experiments. With a view to understand the role of vadose zone in denitrification process, experiments are performed with unsaturated sand specimens (S (r) = 73-90 %) whose pore water was spiked with nitrate and ethanol solutions. Experimental results revealed 73 % S (r) specimen facilitates nitrate reduction to 45 mg/L in relatively short durations of 5.5-7.5 h using the available natural organic matter (0.41 % on mass basis of sand); consequently, ethanol addition did not impact rate of denitrification. However, at higher S (r) values of 81 and 90 %, extraneous ethanol addition (C/N = 0.5-3) was needed to accelerate the denitrification rates.

Humanized antibody neutralizing 2009 pandemic H1N1 virus

The 2009 pandemic H1N1 S-OIV (swine origin influenza A virus) caused noticeable morbidity and mortality worldwide. In addition to vaccine and antiviral drug therapy, the use of influenza virus neutralizing monoclonal antibodies (MAbs) for treatment purposes is a viable alternative. We previously reported the isolation of a high affinity, potently neutralizing murine MAb MA2077 against 2009 pandemic H1N1 virus. We describe here the humanization of MA2077 and its expression in a mammalian cell line. Six complementarity-determining regions (CDRs) of MA2077 were grafted onto the human germline variable regions; along with six and eight back mutations in the framework of heavy and light chains, respectively, pertaining to the vernier zone and interchain packing residues to promote favorable CDR conformation and facilitate antigen binding. The full length humanized antibody, 2077Hu2, expressed in CHO-K1 cells, showed high affinity to hemagglutinin protein (K-D = 0.75 +/- 0.32 nM) and potent neutralization of pandemic H1N1 virus (IC50 = 0.17 mu g/mL), with marginally higher IC50 as compared to MA2077 (0.08 mu g/mL). In addition, 2077Hu2 also retained the epitope specificity for the ``Sa'' antigenic site on pandemic HA. To the best of our knowledge, this is the first report of a humanized neutralizing antibody against pandemic H1N1 virus.

Impact of compost, vermicompost and biochar on soil fertility, maize yield and soil erosion in Northern Vietnam: A three year mesocosm experiment

Compost, vermicompost and biochar amendments are thought to improve soil quality and plant yield. However, little is known about their long-term impact on crop yield and the environment in tropical agro-ecosystems. In this study we investigated the effect of organic amendments (buffalo manure, compost and verrnicompost) and biochar (applied alone or with vermicompost) on plant yield, soil fertility, soil erosion and water dynamics in a degraded Acrisol in Vietnam. Maize growth and yield, as well as weed growth, were examined for three years in terrestrial mesocosms under natural rainfall. Maize yield and growth showed high inter-annual variability depending on the organic amendment. Vermicompost improved maize growth and yield but its effect was rather small and was only significant when water availability was limited (year 2). This suggests that vermicompost could be a promising substrate for improving the resistance of agrosystems to water stress. When the vermicompost biochar mixture was applied, further growth and yield improvements were recorded in some cases. When applied alone, biochar had a positive influence on maize yield and growth, thus confirming its interest for improving long-term soil productivity. All organic amendments reduced water runoff, soil detachment and NH4+ and NO3- transfer to water. These effects were more significant with vermicompost than with buffalo manure and compost, highlighting that the beneficial influence of vermicompost is not limited to its influence on plant yield. In addition, this study showed for the first time that the combination of vermicompost and biochar may not only improve plant productivity but also reduce the negative impact of agriculture on water quality. (C) 2015 Elsevier B.V. All rights reserved.