The Three-dimensional Solution Structure of NaD1, a New Floral Defensin from Nicotiana alata and its Application to a Homology Model of the Crop Defense Protein alfAFP

NMR spectroscopy and simulated annealing calculations have been used to determine the three-dimensional structure of NaD1, a novel antifungal and insecticidal protein isolated from the flowers of Nicotiana alata. NaD1 is a basic, cysteine-rich protein of 47 residues and is the first example of a plant defensin from flowers to be characterized structurally. Its three-dimensional structure consists of an a-helix and a triple-stranded anti-parallel beta-sheet that are stabilized by four intramolecular disulfide bonds. NaD1 features all the characteristics of the cysteine-stabilized up motif that has been described for a variety of proteins of differing functions ranging from antibacterial insect defensins and ion channel-perturbing scorpion toxins to an elicitor of the sweet taste response. The protein is biologically active against insect pests, which makes it a potential candidate for use in crop protection. NaD1 shares 31% sequence identity with alfAFP, an antifungal protein from alfalfa that confers resistance to a fungal pathogen in transgenic potatoes. The structure of NaD1 was used to obtain a homology model of alfAFP, since NaD1 has the highest level of sequence identity with alfAFP of any structurally characterized antifungal defensin. The structures of NaD1 and alfAFP were used in conjunction with structure - activity data for the radish defensin Rs-AFP2 to provide an insight into structure-function relationships. In particular, a putative effector site was identified in the structure of NaD1 and in the corresponding homology model of alfAFP. (C) 2002 Elsevier Science Ltd. All rights reserved.

The JK three-product cyclone - performance and potential applications

In spite of their wide application in comminution circuits, hydrocyclones have at least one significant disadvantage in that their operation inherently tends to return the fine denser liberated minerals to the grinding mill. This results in unnecessary overgrinding which adds to the milling cost and can adversely affect the efficiency of downstream processes. In an attempt to solve this problem, a three-product cyclone has been developed at the Julius Kruttschnitt Mineral Research Centre (JKMRC) to generate a second overflow in which the fine dense liberated minerals can be selectively concentrated for further treatment. In this paper, the design and operation of the three-product cyclone are described. The influence of the length of the second vortex finder on the performance of a 150-mm unit treating a mixture of magnetite and silica is investigated. Conventional cyclone tests were also conducted under similar conditions. Using the operational performance data of the three-product and conventional cyclones, it is shown that by optimising the length of the second vortex finder, the amount of fine dense mineral particles that reports to the three-product cyclone underflow can be reduced. In addition, the three-product cyclone can be used to generate middlings stream that may be more suitable for flash flotation than the conventional cyclone underflow, or alternatively, could be classified with a microscreen to separate the valuables from the gangue. At the same time, a fines stream having similar properties to those of the conventional overflow can be obtained. Hence, if the middlings stream was used as feed for flash flotation or microscreening, the fines stream could be used in lieu of the conventional overflow without compromising the feed requirements for the conventional flotation circuit. Some of the other potential applications of the new cyclone are described. (C) 2003 Elsevier Science B.V. All rights reserved.

Fatty acid binding protein is a major determinant of hepatic pharmacokinetics of palmitate and its metabolites

Disposition kinetics of [H-3] palmitate and its low-molecular-weight metabolites in perfused rat livers were studied using the multiple-indicator dilution technique, a selective assay for [H-3] palmitate and its low-molecular-weight metabolites, and several physiologically based pharmacokinetic models. The level of liver fatty acid binding protein (L-FABP), other intrahepatic binding proteins (microsomal protein, albumin, and glutathione S-transferase) and the outflow profiles of [H-3] palmitate and metabolites were measured in four experimentalgroups of rats: 1) males; 2) clofibrate-treated males; 3) females; and 4) pregnant females. A slow-diffusion/bound model was found to better describe the hepatic disposition of unchanged [H-3] palmitate than other pharmacokinetic models. The L-FABP levels followed the order: pregnant female > clofibrate-treated male > female > male. Levels of other intrahepatic proteins did not differ significantly. The hepatic extraction ratio and mean transit time for unchanged palmitate, as well as the production of low-molecular-weight metabolites of palmitate and their retention in the liver, increased with increasing L-FABP levels. Palmitate metabolic clearance, permeability-surface area product, retention of palmitate by the liver, and cytoplasmic diffusion constant for unchanged [H-3] palmitate also increased with increasing L-FABP levels. It is concluded that the variability in hepatic pharmacokinetics of unchanged [H-3] palmitate and its low-molecular-weight metabolites in perfused rat livers is related to levels of L-FABP and not those of other intrahepatic proteins.

Heat resistance of Bacillus spores when adhered to stainless steel and its relationship to spore hydrophobicity

Twenty-one strains of Bacillus (10 B. stearothermophilus, 3 B. cereus, and 8 B. licheniformis strains) were assayed for spore surface hydrophobicity on the basis of three measures: contact angle measurement (CAM), microbial adhesion to hydrocarbons (MATH), and hydrophobic interaction chromatography (HIC). On the basis of the spore surface characteristics obtained from these assays, along with data on the heat resistance of these spores in water, eight strains of Bacillus (three B. stearothermophilus, three B. cereus, and two B. licheniformis strains) either suspended in water or adhering to stainless steel were exposed to sublethal heat treatments at 90 to 110degreesC to determine heat resistance (D-value). Significant increases in heat resistance (ranging from 3 to 400%) were observed for the eight strains adhering to stainless steel. No significant correlation was found between these heat resistance increases and spore surface characteristics as determined by the three hydrophobicity assays. There was a significant positive correlation between the hydrophobicity data obtained by the MATH assay and those obtained by the HIC assay, but these data did not correlate with those obtained by the CAM assay.

Nitrate ammonification and its relationship to the accumulation of ammonium in a Vertisol subsoil

High concentrations of ammonium ( up to 270 kg N/ha) have been observed in a Vertisol soil below 1 m depth near Warra in south-east Queensland. This study examined the possibility that increased water movement into the subsoil after the removal of native vegetation, and a subsequent increase in periods of waterlogging, could have triggered nitrate ammonification and be responsible for the production of ammonium. Two incubation experiments were conducted to test this hypothesis. The first involved the incubation of repacked cores that had been amended with 30 mg N/kg of 5 atom% N-15 nitrate under low oxygen conditions for a period of 360 days. Over this time period the N-15 enrichment of the exchangeable ammonium fraction was monitored in order to detect any reduction of nitrate to ammonium. The second experiment involved the incubation of soil amended with 30 mg N/ kg of 5 atom% N-15 nitrate under waterlogged and low oxygen conditions for 75 days. During this period the redox potential of the soil was monitored using a field test to determine if reducing conditions would develop in this soil over a period of waterlogging, combined with the monitoring of any nitrate reduction to ammonium. The results of these experiments indicated that a small amount of nitrate ammonification (< 0.1 mg N/ kg) could be observed in the Warra subsoil, but that unless the rate of reduction were to significantly increase with time, this could not account for the accumulation of ammonium observed in the field. The environmental conditions that would make either dissimilatory or abiotic nitrate ammonification favourable were not observed to develop. Consequently, it has been concluded that the observed nitrate ammonification occurred via an assimilatory pathway. Due to the low rate of microbial activity in this subsoil it is considered unlikely that this process was responsible for the subsoil ammonium accumulation at Warra.

Subsoil nitrogen mineralisation and its potential to contribute to NH4 accumulation in a Vertosol

High concentrations of NH4+ (up to 270 kg N/ha) have been observed in a Vertosol below 1 m depth in south-east Queensland. This study examined the possibility that mineralisation associated with the removal of native vegetation (Acacia harpophylla) for cropping was responsible for the production of NH4+. Particularly, the potential contribution of decomposing root material and/or dissolved organic nitrogen (DON) leached into the subsoil after clearing was investigated. The amount of N that was contained within native vegetation root material was determined from an area of native vegetation adjacent to the cleared site containing elevated NH4+ concentrations. In addition, the amount of NH4+ that could be mineralised in the native vegetation soil was determined by monitoring NH4+ concentrations over 360 days in intact cores, and by conducting waterlogged incubations. To determine the rate at which a source of DON leached into the subsoil would mineralise, soil was amended with glutamic acid at a rate of 250 mg N/kg and placed under waterlogged incubation. The possibility that the acidic pH of the subsoil, or the lack of a significant subsoil microbial population, was inhibiting mineralisation was also examined by increasing soil pH from 4.4 to 7.0, and inoculating the subsoil with surface soil microorganisms during waterlogged incubations. Low concentrations of N, approximately 90 kg N/ha between 1.2 and 3 m, were found in the native vegetation root material. In addition, no net N mineralisation was observed in either the extended incubation of intact cores or in the control samples of the waterlogged incubations. Net N mineralisation was also not detected when the subsoil was amended with a source of organic N. Results indicate that this lack of mineralisation is largely due to pH inhibition of the microbial population. It is concluded that the mineralisation of either in situ organic material, or DON transported to the subsoil during leaching events, is unlikely to have significantly contributed to the subsoil NH4 accumulation at the study site.

Biodegradation and ecotoxicity evaluation of a bionolle and starch blend and its degradation products in compost

A polymer based on a blend of starch and Bionolle(TM) has been prepared and tested for biodegradation in compost. The polymer was completely mineralised to carbon dioxide in 45 days. The potential toxicity of the polymer was tested against the earthworm Eisenia fetida using a modification of the American Standard for Testing Materials E1976-97. The earthworms were exposed to 30 g of the polymer for 28 days and changes in weight recorded. In addition, the polymer was firstly degraded by the compost and the worms exposed to the breakdown products for 28 days. Differences in weight were also recorded. In each case the production of juveniles was noted and all earthworms were examined for pathology. The results obtained were processed statistically using a t-test. The number of juveniles, produced from the breakdown products, was highly significant (P < 0.001) when compared to the earthworms added to the intact polymer. There was a definitely significant difference (P < 0.01, t = 3.25) in change in weight between the earthworms that were exposed to the polymer directly and those that were exposed to the breakdown products. There was no indication of any pathology of any earthworms. The polymer is considered safe for this species. (C) 2002 Elsevier Science Ltd. All rights reserved.

Biased EPR entanglement and its application to teleportation

We consider pure continuous variable entanglement with non-equal correlations between orthogonal quadratures. We introduce a simple protocol which equates these correlations and in the process transforms the entanglement onto a state with the minimum allowed number of photons. As an example we show that our protocol transforms, through unitary local operations, a single squeezed beam split on a beam splitter into the same entanglement that is produced when two squeezed beams are mixed orthogonally. We demonstrate that this technique can in principle facilitate perfect teleportation utilizing only one squeezed beam.