Microbial relatives of the seed storage proteins of higher plants: conservation of structure and diversification of function during evolution of the cupin superfamily.

This review summarizes the recent discovery of the cupin superfamily (from the Latin term "cupa," a small barrel) of functionally diverse proteins that initially were limited to several higher plant proteins such as seed storage proteins, germin (an oxalate oxidase), germin-like proteins, and auxin-binding protein. Knowledge of the three-dimensional structure of two vicilins, seed proteins with a characteristic beta-barrel core, led to the identification of a small number of conserved residues and thence to the discovery of several microbial proteins which share these key amino acids. In particular, there is a highly conserved pattern of two histidine-containing motifs with a varied intermotif spacing. This cupin signature is found as a central component of many microbial proteins including certain types of phosphomannose isomerase, polyketide synthase, epimerase, and dioxygenase. In addition, the signature has been identified within the N-terminal effector domain in a subgroup of bacterial AraC transcription factors. As well as these single-domain cupins, this survey has identified other classes of two-domain bicupins including bacterial gentisate 1, 2-dioxygenases and 1-hydroxy-2-naphthoate dioxygenases, fungal oxalate decarboxylases, and legume sucrose-binding proteins. Cupin evolution is discussed from the perspective of the structure-function relationships, using data from the genomes of several prokaryotes, especially Bacillus subtilis. Many of these functions involve aspects of sugar metabolism and cell wall synthesis and are concerned with responses to abiotic stress such as heat, desiccation, or starvation. Particular emphasis is also given to the oxalate-degrading enzymes from microbes, their biological significance, and their value in a range of medical and other applications.

The interrelationships of winter wheat cultivars, crop density and competition of naturally occurring weed flora

The effects of intraspecific and interspecific competition on a wide range of winter wheat cultivars were investigated in two consecutive split plot field experiments. Significant reductions of grain yield at greatly reduced seed rates were observed in the first experiment, whereas increasing crop density up to 380 plants m(-2) in the second experiment failed to produce a significant yield response due to compensation through increased ears and grains per plant at lower crop densities. Appreciable weed suppression and acceptable grain yield can be achieved at crop densities between 150 and 270 plants m(-2). Reductions in final yield due to weed competition occurred in both experiments; 11.7 and 13.6% for the first and second experiment, respectively, with the onset of weed competition occurring from tittering in the first experiment and from stem elongation in the second. The possibility of enhancing crop competitiveness for weed suppression and improved grain yield is discussed.

Toughness & cutting: A new way of determining ductile fracture toughness and strength

A review is given of the mechanics of cutting, ranging from the slicing of thin floppy offcuts (where there is negligible elasticity and no permanent deformation of the offcut) to the machining of ductile metals (where there is severe permanent distortion of the offcut/chip). Materials scientists employ the former conditions to determine the fracture toughness of ‘soft’ solids such as biological materials and foodstuffs. In contrast, traditional analyses of metalcutting are based on plasticity and friction only, and do not incorporate toughness. The machining theories are inadequate in a number of ways but a recent paper has shown that when ductile work of fracture is included many, if not all, of the shortcomings are removed. Support for the new analysis is given by examination of FEM simulations of metalcutting which reveal that a ‘separation criterion’ has to be employed at the tool tip. Some consideration shows that the separation criteria are versions of void-initiation-growth-and-coalescence models employed in ductile fracture mechanics. The new analysis shows that cutting forces for ductile materials depend upon the fracture toughness as well as plasticity and friction, and reveals a simple way of determining both toughness and flow stress from cutting experiments. Examples are given for a wide range of materials including metals, polymers and wood, and comparison is made with the same properties independently determined using conventional testpieces. Because cutting can be steady state, a new way is presented for simultaneously measuring toughness and flow stress at controlled speeds and strain rates.

Fecal water as a non-invasive biomarker in nutritional intervention: comparison of preparation methods and refinement of different endpoints

The assessment of cellular effects by the aqueous phase of human feces (fecal water, FW) is a useful biomarker approach to study cancer risks and protective activities of food. In order to refine and develop the biomarker, different protocols of preparing FW were compared. Fecal waters were prepared by 3 methods: (A) direct centrifugation; (B) extraction of feces in PBS before centrifugation; and (C) centrifugation of lyophilized and reconstituted feces. Genotoxicity was determined in colon cells using the Comet assay. Selected samples were investigated for additional parameters related to carcinogenesis. Two of 7 FWs obtained by methods A and B were similarly genotoxic. Method B, however, yielded higher volumes of FW, allowing sterile filtration for long-term culture experiments. Four of 7 samples were non-genotoxic when prepared according to all 3 methods. FW from lyophilized feces and from fresh samples were equally genotoxic. FWs modulated cytotoxicity, paracellular permeability, and invasion, independent of their genotoxicity. All 3 methods of FW preparation can be used to assess genotoxicity. The higher volumes of FWobtained by preparation method B greatly enhance the perspectives of measuring different types of biological parameters and using these to disclose activities related to cancer development.

PIGOK: linking protein identity to gene ontology and function

Here we introduce a computer database that allows for the rapid retrieval of physicochemical properties, Gene Ontology, and Kyoto Encyclopedia of Genes and Genomes information about a protein or a list of proteins. We applied PIGOK analyzing Schizosaccharomyces pombe proteins displaying differential expression under oxidative stress and identified their biological functions and pathways. The database is available on the Internet at http://pc4-133.ludwig.ucl.ac.uk/pigok.html.

Effects of food structure and ingredient interactions on antioxidant capacity

Foods are complex biological materials, and the lipids within the food are susceptible to lipid oxidation, which is retarded by antioxidants. The precise structure and composition of the food may affect the antioxidant activity quite strongly in some cases. Solubility of the antioxidant in the phases present is one of the main parameters that affects the variation in antioxidant activity with phase composition of food. Polar antioxidants are more effective in oils, whereas non-polar antioxidants are more effective in oil-in-water emulsions. Antioxidant activity has been reported in a range of different media, including oils, emulsions, liposomes, microemulsions, fish and meat muscles, and the antioxidant activity may vary from one medium to another. Synergy between antioxidants may also vary from one medium to another. Interactions with metals and with proteins affect antioxidant activity and these interactions are also dependent on the phases present.

The effect of condensed tannins in fresh sainfoin (Onobrychis viciifolia) on in vivo and in situ digestion in sheep

This study focused on effects of structure, content and biological activity of condensed tannins (CT) in leaves, stems and whole plant of sainfoin (Onobrychis viciifolia) on its in vivo and in situ digestive characteristics in sheep. Sainfoin was studied as fresh forage during the first vegetation cycle at two phenological stages (i.e., end of flowering and green seeds) and during the second vegetation cycle (i.e., start of flowering). The feeding experiment used 12 sheep; with six dosed, through the rumen cannula, with polyethylene glycol (PEG) to neutralise CT effects. Organic matter digestibility (OMD), total tract N disappearance and N balance were measured in sheep fed the whole plant. The residues of dry matter (DM) and N from nylon bags suspended in the rumen were determined on leaves and stems. Intestinal digestibility was measured using other, intestinally fistulated sheep. PEG addition and vegetation cycle increased total tract N digestibility (P<0.001) but PEG affected OMD only at the end of flowering. PEG inactivated the CT and increased urinary N excretion (P<0.05) but this was offset by lower faecal N excretion (P<0.001). Feeding sainfoin can be used to alter the form of excreted N (i.e., urine vs faeces) and thus potentially reduce environmental N pollution without affecting body N retention. Kinetic studies of total N, ammonia N (NH3-N) and volatile fatty acids (VFA) in rumen fluid were made before and 1.5, 3 and 6 h after feeding. Sainfoin CT decreased rumen fluid soluble N (P<0.05) and NH3-N (P<0.01). Ruminal N disappearance (DisN) of leaves or stems was lower in the presence of active CT compared to PEG-inactivated CT (P<0.001) for both vegetation cycles. PEG also increased intestinal digestibility (P<0.05) of leaves and stems. Leaves had lower ruminal DisN, but higher N disappearing from intestine than stems. The biological activity and content of CT in the whole plant decreased as phenological stage increased. Prodelphinidin:procyanidin (PD:PC) ratios of leaves varied with vegetation cycle and phenological stage. The molecular size of CT in the whole plant, as indicated by their mean degree of polymerisation (mDP), was lowest at the start of flowering and coincided with the higher biological activity and content of CT.

Purification and functional characterisation of Rhinocerase, a novel serine protease from the venom of Bitis gabonica rhinoceros

Background: Serine proteases are a major component of viper venoms and are thought to disrupt several distinct elements of the blood coagulation system of envenomed victims. A detailed understanding of the functions of these enzymes is important both for acquiring a fuller understanding of the pathology of envenoming and because these venom proteins have shown potential in treating blood coagulation disorders. Methodology/Principal Findings: In this study a novel, highly abundant serine protease, which we have named rhinocerase, has been isolated and characterised from the venom of Bitis gabonica rhinoceros using liquid phase isoelectric focusing and gel filtration. Like many viper venom serine proteases, this enzyme is glycosylated; the estimated molecular mass of the native enzyme is approximately 36kDa, which reduces to 31kDa after deglycosylation. The partial amino acid sequence shows similarity to other viper venom serine proteases, but is clearly distinct from the sequence of the only other sequenced serine protease from Bitis gabonica. Other viper venom serine proteases have been shown to exert distinct biological effects, and our preliminary functional characterization of rhinocerase suggest it to be multifunctional. It is capable of degrading α and β chains of fibrinogen, dissolving plasma clots and of hydrolysing a kallikrein substrate. Conclusions/Significance: A novel multifunctional viper venom serine protease has been isolated and characterised. The activities of the enzyme are consistent with the known in vivo effects of Bitis gabonica envenoming, including bleeding disorders, clotting disorders and hypotension. This study will form the basis for future research to understand the mechanisms of serine protease action, and examine the potential for rhinocerase to be used clinically to reduce the risk of human haemostatic disorders such as heart attacks and strokes.

Development and validation of a qPCR-based method for quantifying Shiga toxin-encoding and other lambdoid bacteriophages

P>To address whether seasonal variability exists among Shiga toxin-encoding bacteriophage (Stx phage) numbers on a cattle farm, conventional plaque assay was performed on water samples collected over a 17 month period. Distinct seasonal variation in bacteriophage numbers was evident, peaking between June and August. Removal of cattle from the pasture precipitated a reduction in bacteriophage numbers, and during the winter months, no bacteriophage infecting Escherichia coli were detected, a surprising occurrence considering that 1031 tailed-bacteriophages are estimated to populate the globe. To address this discrepancy a culture-independent method based on quantitative PCR was developed. Primers targeting the Q gene and stx genes were designed that accurately and discriminately quantified artificial mixed lambdoid bacteriophage populations. Application of these primer sets to water samples possessing no detectable phages by plaque assay, demonstrated that the number of lambdoid bacteriophage ranged from 4.7 x 104 to 6.5 x 106 ml-1, with one in 103 free lambdoid bacteriophages carrying a Shiga toxin operon (stx). Specific molecular biological tools and discriminatory gene targets have enabled virus populations in the natural environment to be enumerated and similar strategies could replace existing propagation-dependent techniques, which grossly underestimate the abundance of viral entities.

Persistence of anticancer activity in berry extracts after simulated gastrointestinal digestion and colonic fermentation.

Fruit and vegetable consumption is associated at the population level with a protective effect against colorectal cancer. Phenolic compounds, especially abundant in berries, are of interest due to their putative anticancer activity. After consumption, however, phenolic compounds are subject to digestive conditions within the gastrointestinal tract that alter their structures and potentially their function. However, the majority of phenolic compounds are not efficiently absorbed in the small intestine and a substantial portion pass into the colon. We characterized berry extracts (raspberries, strawberries, blackcurrants) produced by in vitro-simulated upper intestinal tract digestion and subsequent fecal fermentation. These extracts and selected individual colonic metabolites were then evaluated for their putative anticancer activities using in vitro models of colorectal cancer, representing the key stages of initiation, promotion and invasion. Over a physiologically-relevant dose range (0-50 µg/ml gallic acid equivalents), the digested and fermented extracts demonstrated significant anti-genotoxic, anti-mutagenic and anti-invasive activity on colonocytes. This work indicates that phenolic compounds from berries undergo considerable structural modifications during their passage through the gastrointestinal tract but their breakdown products and metabolites retain biological activity and can modulate cellular processes associated with colon cancer.

Soils and food security: challenges and opportunities

Soils most obviously contribute to food security in their essential role in crop and fodder production, so affecting the local availability of particular foods. They also have a direct influence on the ability to distribute food, the nutritional value of some foods and, in some societies, the access to certain foods through local processes of allocation and preferences. The inherent fertility of some soils is greater than that of others, so that crop yields vary greatly under semi-natural conditions. Husbandry practices, including the use of manures and fertilisers, have evolved to improve biological, chemical and physical components of soil fertility and thereby increase crop production. The challenge for the future is to sustain soil fertility in ways that increase the yield per unit area while simultaneously avoiding other detrimental environmental consequences. This will require increased effort to develop practices that use inputs such as nutrients, water and energy more efficiently. Opportunities to achieve this include adopting more effective ways to apply water and nutrients, adopting tillage practices that promote water infiltration and increase of organic matter, and breeding to improve the effectiveness of root systems in utilising soil-based resources.

Error, reproducibility and sensitivity: a pipeline for data processing of Agilent oligonucleotide expression arrays

Background: Expression microarrays are increasingly used to obtain large scale transcriptomic information on a wide range of biological samples. Nevertheless, there is still much debate on the best ways to process data, to design experiments and analyse the output. Furthermore, many of the more sophisticated mathematical approaches to data analysis in the literature remain inaccessible to much of the biological research community. In this study we examine ways of extracting and analysing a large data set obtained using the Agilent long oligonucleotide transcriptomics platform, applied to a set of human macrophage and dendritic cell samples. Results: We describe and validate a series of data extraction, transformation and normalisation steps which are implemented via a new R function. Analysis of replicate normalised reference data demonstrate that intrarray variability is small (only around 2 of the mean log signal), while interarray variability from replicate array measurements has a standard deviation (SD) of around 0.5 log(2) units (6 of mean). The common practise of working with ratios of Cy5/Cy3 signal offers little further improvement in terms of reducing error. Comparison to expression data obtained using Arabidopsis samples demonstrates that the large number of genes in each sample showing a low level of transcription reflect the real complexity of the cellular transcriptome. Multidimensional scaling is used to show that the processed data identifies an underlying structure which reflect some of the key biological variables which define the data set. This structure is robust, allowing reliable comparison of samples collected over a number of years and collected by a variety of operators. Conclusions: This study outlines a robust and easily implemented pipeline for extracting, transforming normalising and visualising transcriptomic array data from Agilent expression platform. The analysis is used to obtain quantitative estimates of the SD arising from experimental (non biological) intra- and interarray variability, and for a lower threshold for determining whether an individual gene is expressed. The study provides a reliable basis for further more extensive studies of the systems biology of eukaryotic cells.

Language in genetic syndromes and cognitive modularity

In recent years, research into the impact of genetic abnormalities on cognitive development, including language, has become recognized for its potential to make valuable contributions to our understanding of the brain–behaviour relationships underlying language acquisition as well as to understanding the cognitive architecture of the human mind. The publication of Fodor’s ( 1983 ) book The Modularity of Mind has had a profound impact on the study of language and the cognitive architecture of the human mind. Its central claim is that many of the processes involved in comprehension are undertaken by special brain systems termed ‘modules’. This domain specificity of language or modularity has become a fundamental feature that differentiates competing theories and accounts of language acquisition (Fodor 1983 , 1985 ; Levy 1994 ; Karmiloff-Smith 1998 ). However, although the fact that the adult brain is modularized is hardly disputed, there are different views of how brain regions become specialized for specific functions. A question of some interest to theorists is whether the human brain is modularized from the outset (nativist view) or whether these distinct brain regions develop as a result of biological maturation and environmental input (neuroconstructivist view). One source of insight into these issues has been the study of developmental disorders, and in particular genetic syndromes, such as Williams syndrome (WS) and Down syndrome (DS). Because of their uneven profiles characterized by dissociations of different cognitive skills, these syndromes can help us address theoretically significant questions. Investigations into the linguistic and cognitive profiles of individuals with these genetic abnormalities have been used as evidence to advance theoretical views about innate modularity and the cognitive architecture of the human mind. The present chapter will be organized as follows. To begin, two different theoretical proposals in the modularity debate will be presented. Then studies of linguistic abilities in WS and in DS will be reviewed. Here, the emphasis will be mainly on WS due to the fact that theoretical debates have focused primarily on WS, there is a larger body of literature on WS, and DS subjects have typically been used for the purposes of comparison. Finally, the modularity debate will be revisited in light of the literature review of both WS and DS. Conclusions will be drawn regarding the contribution of these two genetic syndromes to the issue of cognitive modularity, and in particular innate modularity.

Terrestrial exposure of oilfield flowline additives diminish soil structural stability and remediative microbial function

Onshore oil production pipelines are major installations in the petroleum industry, stretching many thousands of kilometres worldwide which also contain flowline additives. The current study focuses on the effect of the flowline additives on soil physico-chemical and biological properties and quantified the impact using resilience and resistance indices. Our findings are the first to highlight deleterious effect of flowline additives by altering some fundamental soil properties, including a complete loss of structural integrity of the impacted soil and a reduced capacity to degrade hydrocarbons mainly due to: (i) phosphonate salts (in scale inhibitor) prevented accumulation of scale in pipelines but also disrupted soil physical structure; (ii) glutaraldehyde (in biocides) which repressed microbial activity in the pipeline and reduced hydrocarbon degradation in soil upon environmental exposure; (iii) the combinatory effects of these two chemicals synergistically caused severe soil structural collapse and disruption of microbial degradation of petroleum hydrocarbons.