Influence of sugars on endoglucanase and β-xylanase activities of aBacillus strain

Summary ABacillus sp. screened from termite infested soils produced significant amount of endoglucanase and xylanase enzymes when grown on a lignocellulosic substrate, rice husk. Biosynthesis of these enzymes was significantly enhanced by the addition of 0.2% cellobiose or glucose for endoglucanase and xylose for -xylanase activities. In the actual hydrolyses, glucose and cellobiose at low concentrations acted as activitors of endoglucanase activity whereas cellobiose and xylose acted as inhibitors of -xylanase activity.     

Evolutionary relationships of eukaryotic kingdoms

The evolutionary relationships of four eukaryotic kingdoms—Animalia, Plantae, Fungi, and Protista—remain unclear. In particular, statistical support for the closeness of animals to fungi rather than to plants is lacking, and a preferred branching order of these and other eukaryotic lineages is still controversial even though molecular sequences from diverse eukaryotic taxa have been analyzed. We report a statistical analysis of 214 sequences of nuclear small-subunit ribosomal RNA (srRNA) gene undertaken to clarify these evolutionary relationships. We have considered the variability of substitution rates and the nonindependence of nucleotide substitution across sites in the srRNA gene in testing alternative hypotheses regarding the branching patterns of eukaryote phylogeny. We find that the rates of evolution among sites in the srRNA sequences vary substantially and are approximately gamma distributed with size and shape parameter equal to 0.76. Our results suggest that (1) the animals and true fungi are indeed closer to each other than to any other crown group in the eukaryote tree, (2) red algae are the closest relatives of animals, true fungi, and green plants, and (3) the heterokonts and alveolates probably evolved prior to the divergence of red algae and animal-fungus-green-plant lineages. Furthermore, our analyses indicate that the branching order of the eukaryotic lineages that diverged prior to the evolution of alveolates may be generally difficult to resolve with the srRNA sequence data.     

Predictions for oxygen supply control to enhance population stability of engineered production strains

The simultaneous growth and product formation in a microbial culture is an important feature of several laboratory, industrial, and environmental bioprocesses. Metabolic burden associated with product formation in these bioprocesses may lead to growth advantage of a nonproducing mutant leading to a loss of the producing population over time. A simple population dynamics model demonstrates the extreme sensitivity of population stability to the engineered productivity of a strain. Here we use flux balance analysis to estimate the effects of the metabolic burden associated with product secretion on optimal growth rates. Comparing the optimal growth rates of the producing and nonproducing strains under a given processing condition allows us to predict the population stability. In order to increase stability of an engineered strain, we determine processing conditions that simultaneously maximize the growth rate of the producing population while minimizing the growth rate of a nonproducing population. Using valine, tryptophan, and lysine production as specific examples, we demonstrate that although an appropriate choice of oxygenation may increase culture longevity more than twofold, total production as governed by economic criterion can be increased by several orders of magnitude. Choice of optimal nutrient and oxygen supply rates to enhance stability is important both for strain screening as well as for culture of engineered strains. Appropriate design of the culture environment can thus be used to enhance the productivity of bioprocesses that use engineered production strains. (c) 1994 John Wiley & Sons, Inc.     

Stoichiometric interpretation of Escherichia coli glucose catabolism under various oxygenation rates.

Metabolic by-product secretion is commonly observed in oxygen-limited cultures. Oxygen limitations occur because of limits in the capacity of the respiratory system or because of the oxygenation limits of the cultivation method used. The latter restriction is of considerable practical importance since it results in a critical cell concentration above which oxygenation is insufficient, leading to by-product secretion. In this study we used a flux balance approach to determine optimal metabolic performance of Escherichia coli under variable oxygen limitations. This method uses linear optimization to find optimal metabolic flux patterns with respect to cell growth. Cell growth was defined as precursor requirements on the basis of a composition analysis. A growth-associated maintenance requirement of 23 mmol of ATP per g of biomass and a non-growth-associated maintenance value of 5.87 mmol at ATP per g (dry weight)-h were incorporated on the basis of a comparison with experimental data. From computations of optimal growth increased oxygen limitations were found to result in the secretion of acetate, formate, and ethanol in that order. Consistent with the experimental data in the literature, by-product secretion rates increased linearly with the growth rate. The computed optimal growth under increasing oxygen limitation revealed four critical growth rates at which changes in the by-product secretion pattern were observed. Concomitant with by-product secretion under oxygen limitations were changes in metabolic pathway utilization. The shifts in metabolism were characterized by changes in the metabolic values (computed as shadow prices) of the various redox carriers. The redox potential was thus identified as a likely trigger that leads to metabolic shifts.2+ ă     

Identification of Phorbol Myristate Acetate Stimulated Kinase in Zea mays

Following DEAE-Sephacel and affinity chromatography a highly enriched lipid stimulated kinase activity could be recovered with a purification fold of 1725. The peak kinase activity fraction eluted with 0.1 mM calcium from phosphatidyl serine affinity chromatography showed a major protein of 70 kD and a minor band of 55 kD molecular weight and showed kinase activity that was stimulated by phorbol myristate acetate in the presence of phosphatidylserine and calcium. The optimum requirement was 2.5 × 10^?6 M, 1.25 × 10^?4 M, 1 × 10^?4 M, and 1.7 × 10^?6 M for phorbol myristate acetate, phosphatidyl serine, oleyl acetyl glycerol and free calcium respectively. The kinase activity was inhibited by H-7 and staurosporine. The binding of [³H]-phorbol myristate acetate was associated with purified fraction as resolved by get electrophoresis and the kinase activity was also precipitated by animal protein kinase C antibodies. The present data give strong evidence for the presence of phorbol myristate acetate stimulated kinase in plants.     

Enzymatic labeling of nucleic acids

Enzymatic labeling of nucleic acids is a fundamental tool in molecular biology with virtually every aspect of nucleic acid hybridization technique involving the use of labeled probes. Different methods for enzymatic labeling of DNA, RNA and oligonucleotide probes are available today. In this review, we will describe both radioactive and nonradioactive labeling methods, yet the choice of system for labeling the probe depends on the application under study.     

Effect of moisture on lipase catalysed esterification of geraniol of palmarosa oil in non-aqueous system

Summary Optimisation of reaction conditions for the esterification of geraniol of palmarosa oil with n-butyric acid using immobilized lipase from Mucor miehei in non-aqueous system was carried out. Palmarosa oil could be easily esterified upto 95% w/w at 40°C in 24 h. Effect of moisture content was studied using Na₂SO₄ and recycling of the immobilized enzyme.     

Parametric sensitivity of stoichiometric flux balance models applied to wild-type Escherichia coli metabolism

The esterification of lauric acid with geraniol catalyzed by the commercially immobilized lipase preparation from Mucor miehei, Lipozyme(R), was studied in well-stirred flasks. The enzyme support was characterized in terms of its internal and external surface area, protein location, and protein content. It was found that the enzyme was mainly located on the external surface of the support, therefore, internal diffusional limitations were not important. It was also shown that the protein content of the support depends on the size of the particle, with smaller particles containing higher amounts of protein per unit weight. Under the conditions studied, the reaction was not under external mass transfer limitations, and the initial reaction rate depended on the size of the support particles. This was mainly due to the different protein contents on the support as a function of particle size and not to internal or external mass transfer limitations. Also, it was found that the inhibition exerted by water was predominantly a physical effect due to its accumulation around the enzyme. It was also found that the reaction was substrate inhibited by lauric acid, but not by geraniol. (c) 1995 John Wiley & Sons, Inc.     

Transmission Studies with Cucumber Green Mottle Mosaic Virus

Cucumber green mottle mosaic virus may spread in bottlegourd under field conditions through soil contaminated with infected plant debris followed by contact. No seed transmission was noticed in bottlegourd (Lagenaria siceraria) or vegetable marrow (Cucurbita pepo) although pollen grains and cotyledons from infected bottlegourd flowers or seeds, respectively, contained negliginle amounts of virus. Cucumber leaf beerles (Raphidopalpa fevicollis) are probable vectors since their regurgitated fluid and excretes contained infective virus particles. No vector fungi were found in the soil around infected bottlegourd plants.