Evidence for the complex relationship between free amino acid and sugar concentrations and acrylamide-forming potential in potato

Free amino acids and reducing sugars participate in the Maillard reaction during high-temperature cooking and processing. This results not only in the formation of colour, aroma and flavour compounds, but also undesirable contaminants, including acrylamide, which forms when the amino acid that participates in the reaction is asparagine. In this study, tubers of 13 varieties of potato (Solanum tuberosum), which had been produced in a field trial in 2010 and sampled immediately after harvest or after storage for 6 months, were analysed to show the relationship between the concentrations of free asparagine, other free amino acids, sugars and acrylamide-forming potential. The varieties comprised five that are normally used for crisping, seven that are used for French fry production and one that is used for boiling. Acrylamide formation was measured in heated flour, and correlated with glucose and fructose concentration. In French fry varieties, which contain higher concentrations of sugars, acrylamide formation also correlated with free asparagine concentration, demonstrating the complex relationship between precursor concentration and acrylamide-forming potential in potato. Storage of the potatoes for 6 months at 9°C had a significant, variety-dependent impact on sugar and amino acid concentrations and acrylamide-forming potential.

IntFOLD: an integrated server for modelling protein structures and functions from amino acid sequences

IntFOLD is an independent web server that integrates our leading methods for structure and function prediction. The server provides a simple unified interface that aims to make complex protein modelling data more accessible to life scientists. The server web interface is designed to be intuitive and integrates a complex set of quantitative data, so that 3D modelling results can be viewed on a single page and interpreted by non-expert modellers at a glance. The only required input to the server is an amino acid sequence for the target protein. Here we describe major performance and user interface updates to the server, which comprises an integrated pipeline of methods for: tertiary structure prediction, global and local 3D model quality assessment, disorder prediction, structural domain prediction, function prediction and modelling of protein-ligand interactions. The server has been independently validated during numerous CASP (Critical Assessment of Techniques for Protein Structure Prediction) experiments, as well as being continuously evaluated by the CAMEO (Continuous Automated Model Evaluation) project. The IntFOLD server is available at: http://www.reading.ac.uk/bioinf/IntFOLD/

Critical role for an acidic amino acid region in platelet signaling by the HemITAM (hemi-immunoreceptor tyrosine-based activation motif) containing receptor CLEC-2 (C-type lectin receptor-2)

CLEC-2 is a member of new family of C-type lectin receptors characterized by a cytosolic YXXL downstream of three acidic amino acids in a sequence known as a hemITAM (hemi-immunoreceptor tyrosine-based activation motif). Dimerization of two phosphorylated CLEC-2 molecules leads to recruitment of the tyrosine kinase Syk via its tandem SH2 domains and initiation of a downstream signaling cascade. Using Syk-deficient and Zap-70-deficient cell lines we show that hemITAM signaling is restricted to Syk and that the upstream triacidic amino acid sequence is required for signaling. Using surface plasmon resonance and phosphorylation studies, we demonstrate that the triacidic amino acids are required for phosphorylation of the YXXL. These results further emphasize the distinct nature of the proximal events in signaling by hemITAM relative to ITAM receptors.

Peptide based hydrogels for cancer drug release: modulation of stiffness, drug release and proteolytic stability of hydrogels by incorporating D-amino acid residue(s)

Synthetic tripeptide based noncytotoxic hydrogelators have been discovered for releasing an anticancer drug at physiological pH and temparature. Interestingly, gel stiffness, drug release capacity and proteolytic stability of these hydrogels have been successfully modulated by incorporating D-amino acid residues, indicating their potential use for drug delivery in the future.

Legumes regulate Rhizobium bacteroid development and persistence by the supply of branched-chain amino acids

One of the largest contributions to biologically available nitrogen comes from the reduction of N-2 to ammonia by rhizobia in symbiosis with legumes. Plants supply dicarboxylic acids as a carbon source to bacteroids, and in return they receive ammonia. However, metabolic exchange must be more complex, because effective N-2 fixation by Rhizobium leguminosarum bv viciae bacteroids requires either one of two broad-specificity amino acid ABC transporters (Aap and Bra). It was proposed that amino acids cycle between plant and bacteroids, but the model was unconstrained because of the broad solute specificity of Aap and Bra. Here, we constrain the specificity of Bra and ectopically express heterologous transporters to demonstrate that branched-chain amino acid (LIV) transport is essential for effective N-2 fixation. This dependence of bacteroids on the plant for LIV is not due to their known down-regulation of glutamate synthesis, because ectopic expression of glutamate dehydrogenase did not rescue effective N-2 fixation. Instead, the effect is specific to LIV and is accompanied by a major reduction in transcription and activity of LIV biosynthetic enzymes. Bacteroids become symbiotic auxotrophs for LIV and depend on the plant for their supply. Bacteroids with aap bra null mutations are reduced in number, smaller, and have a lower DNA content than wild type. Plants control LIV supply to bacteroids, regulating their development and persistence. This makes it a critical control point for regulation of symbiosis. MICROBIOLOGY

Characterization of a gamma-aminobutyric acid transport system of rhizobium leguminosarum bv. viciae 3841

Spontaneous mutants of Rhizobium leguminosarum bv. viciae 3841 were isolated that grow faster than the wild type on gamma-aminobutyric acid (GABA) as the sole carbon and nitrogen source. These strains (RU1736 and RU1816) have frameshift mutations (gtsR101 and gtsR102, respectively) in a GntR-type regulator (GtsR) that result in a high rate of constitutive GABA transport. Tn5 mutagenesis and quantitative reverse transcription-PCR showed that GstR regulates expression of a large operon (pRL100242 to pRL100252) on the Sym plasmid that is required for GABA uptake. An ABC transport system, GtsABCD (for GABA transport system) (pRL100248-51), of the spermidine/putrescine family is part of this operon. GtsA is a periplasmic binding protein, GtsB and GtsC are integral membrane proteins, and GtsD is an ATP-binding subunit. Expression of gtsABCD from a lacZ promoter confirmed that it alone is responsible for high rates of GABA transport, enabling rapid growth of strain 3841 on GABA. Gts transports open-chain compounds with four or five carbon atoms with carboxyl and amino groups at, or close to, opposite termini. However, aromatic compounds with similar spacing between carboxyl and amino groups are excellent inhibitors of GABA uptake so they may also be transported. In addition to the ABC transporter, the operon contains two putative mono-oxygenases, a putative hydrolase, a putative aldehyde dehydrogenase, and a succinate semialdehyde dehydrogenase. This suggests the operon may be involved in the transport and breakdown of a more complex precursor to GABA. Gts is not expressed in pea bacteroids, and gtsB mutants are unaltered in their symbiotic phenotype, suggesting that Bra is the only GABA transport system available for amino acid cycling.

Major advances in fundamental dairy cattle nutrition

Fundamental nutrition seeks to describe the complex biochemical reactions involved in assimilation and processing of nutrients by various tissues and organs, and to quantify nutrient movement (flux) through those processes. Over the last 25 yr, considerable progress has been made in increasing our understanding of metabolism in dairy cattle. Major advances have been made at all levels of biological organization, including the whole animal, organ systems, tissues, cells, and molecules. At the whole-animal level, progress has been made in delineating metabolism during late pregnancy and the transition to lactation, as well as in whole-body use of energy-yielding substrates and amino acids for growth in young calves. An explosion of research using multicatheterization techniques has led to better quantitative descriptions of nutrient use by tissues of the portal-drained viscera (digestive tract, pancreas, and associated adipose tissues) and liver. Isolated tissue preparations have provided important information on the interrelationships among glucose, fatty acid, and amino acid metabolism in liver, adipose tissue, and mammary gland, as well as the regulation of these pathways during different physiological states. Finally, the last 25 yr has witnessed the birth of "molecular biology" approaches to understanding fundamental nutrition. Although measurements of mRNA abundance for proteins of interest already have provided new insights into regulation of metabolism, the next 25 yr will likely see remarkable advances as these techniques continue to be applied to problems of dairy cattle biology. Integration of the "omics" technologies (functional genomics, proteomics, and metabolomics) with measurements of tissue metabolism obtained by other methods is a particularly exciting prospect for the future. The result should be improved animal health and well being, more efficient dairy production, and better models to predict nutritional requirements and provide rations to meet those requirements.

Effects of host plant drought stress on the performance of the bird cherry-oat aphid, Rhopalosiphum padi (L.): a mechanistic analysis

1. The growth (increase in height and leaf number) of four grass species was reduced by a -0.5 MPa drought stress, but the performance of an associated herbivore, Rhopalosiphum padi (L.), was not affected consistently. The intrinsic rate of increase of R. padi was reduced by drought stress on three grass species, including Dactylis glomerata (L.), but was unaffected on Arrhenatherum elatius (L.). Therefore, there is no general relationship in the effect of plant drought on an insect herbivore, even among closely related host plant species. 2. Drought stress increased the quality of plant phloem sap, as indicated by increased sieve element osmotic pressure and essential amino acid concentrations. Thus, diet quality could not account for the reduced performance of R. padi under drought stress. The concentration of essential amino acids in the phloem of well-watered A. elatius was, however, lower than that of well-watered D. glomerata, correlating with the decreased performance of aphids on well-watered A. elatius. 3. There were no differences in aphid feeding duration between watering treatments or plant species but sap ingestion rates were reduced significantly under drought stress. 4. Using the measure of dietary amino acid concentrations and the estimate of sap ingestion, the essential amino acid flux through aphids was calculated. Compared with the flux through aphids feeding on well-watered D. glomerata, there was a reduction in aphids feeding on drought-stressed D. glomerata and drought-stressed A. elatius due to lower sap ingestion rates. The flux through aphids on well-watered A. elatius was also reduced due to low phloem essential amino acid concentrations. Thus, the performance of an aphid is correlated with the availability and accessibility of essential amino acids.

Metabolic changes of rhizobia in legume nodules

Bacteria have evolved a wide variety of metabolic strategies to cope with varied environments. Some are specialists and only able to survive in restricted environments; others are generalists and able to cope with diverse environmental conditions. Rhizolbia (e.g. Rhizobium, Sinorhizobium, Bradyrhizobium, Mesorhizobium and Azorhizobium species) can survive and compete for nutrients in soil and the plant rhizosphere but can also form a beneficial symbiosis with legumes in a highly specialized plant cell environment. Inside the legume-root nodule, the bacteria (bacteroids) reduce dinitrogen to ammonium, which is secreted to the plant in exchange for a carbon and energy source. A new and challenging aspect of nodule physiology is that nitrogen fixation requires the cycling of amino acids between the bacteroid and plant. This review aims to summarize the metabolic plasticity of rhizobia and the importance of amino acid cycling.

Metabolism of Rhizobium bacteroids

Nitrogen fixation within legume nodules results from a complex metabolic exchange between bacteria of the family Rhizobiaciae and the plant host. Carbon is supplied to the differentiated bacterial cells, termed bacteroids, in the form of dicarboxylic acids to fuel nitrogen fixation. In exchange, fixed nitrogen is transferred to the plant. Both the bacteroid and the plant-derived peribacteroid membrane tightly regulate the exchange of metabolites. In the bacteroid oxidation of dicarboxylic acids via the TCA cycle occurs in an oxygen-limited environment. This restricts the TCA cycle at key points, such as the 2-oxoglutarate dehydrogenase complex, and requires that inputs of carbon and reductant are balanced with outputs from the TCA cycle. This may be achieved by metabolism through accessory pathways that can remove intermediates, reductant, or ATP from the cycle. These include synthesis of the carbon polymers PHB and glycogen and bypass pathways such as the recently identified 2-oxoglutarate decarboxylase reaction in soybean bacteroids. Recent labeling data have shown that bacteroids synthesize and secrete amino acids, which has led to controversy over the role of amino acids in nodule metabolism. Here we review bacteroid carbon metabolism in detail, evaluate the labeling studies that relate to amino acid metabolism by bacteroids, and place the work in context with the genome sequences of Mesorhizobium loti and Sinorhizobium meliloti. We also consider a wider range of metabolic pathways that are probably of great importance to rhizobia in the rhizosphere, during nodule initiation, infection thread development, and bacteroid development.

1,3-Dipolar cycloadditions to unsymmetrical ketone-derived chiral stabilized azomethine ylides: strategies for the synthesis of highly substituted amino acids

We report herein, the first generation of unsymmetrical ketone-derived chiral stabilized azomethine ylides. Intrairiolecular and intermolecular cycloaddition strategies have been utilized to synthesize both an enantiornerically pure bicyclic proline derivative and an enantionierically pure beta-hydroxy-alpha-amino acid.

m-aminobenzoic acid inserted beta-turn in acyclic tripeptides: a peptidomimetic design

X-ray diffraction studies show that peptides Boc-Leu-Aib-m-ABA-OMe (I) (Aib, alpha-aminoisobutyric acid; m-ABA, meta-aminobenzoic acid) and Boc-Phe-Aib-m-ABA-OMe, (II) adopt a type-II beta-turn conformation, solely stabilized by co-operative steric interactions amongst the amino acid residues. This type of U-turn without any intramolecular hydrogen bonding is generally referred to as an open turn. Although there are some examples of constrained cyclic peptides in which o-substituted benzenes have been inserted to mimic the turn region of the neurotrophin, a nerve growth factor, peptides I and II present novel two examples where m-aminobenzoic acid has been incorporated in the beta-turn of acyclic tripeptides. The result also demonstrates the first crystallographic evidence of a beta-turn structure containing an inserted m-aminobenzoic acid, which can be considered as a rigid gamma-aminobutyric acid with an all-trans extended configuration. (c) 2005 Elsevier Ltd. All rights reserved.

Investigations on the effect of amino acids on acrylamide, pyrazines, and Michael addition products in model systems

Acrylamide and pyrazine formation, as influenced by the incorporation of different amino acids, was investigated in sealed low-moisture asparagine-glucose model systems. Added amino acids, with the exception of glycine and cysteine and at an equimolar concentration to asparagine, increased the rate of acrylamide formation. The strong correlation between the unsubstituted pyrazine and acrylamide suggests the promotion of the formation of Maillard reaction intermediates, and in particular glyoxal, as the determining mode of-action. At increased amino acid concentrations, diverse effects were observed. The initial rates of acrylamide formation remained high for valine, alanine, phenylalanine, tryptophan, glutamine, and Ieucine, while a significant mitigating effect, as evident from the acrylamide yields after 60 min of heating at 160 degrees C, was observed for proline, tryptophan, glycine, and cysteine. The secondary amine containing amino acids, proline and tryptophan, had the most profound mitigating effect on acrylamide after 60 min of heating. The relative importance of the competing effect of added amino acids for alpha-dicarbonyls and acrylamide-amino, acid alkylation reactions is discussed and accompanied by data on the relative formation rates of selected amino acid-AA adducts.

Adverse gastrointestinal effects of arginine and related amino acids

Oral supplements of arginine and citrulline increase local nitric oxide (NO production in the small intestine and this may be harmful under certain circumstances. Gastrointestinal toxicity was therefore reviewed with respect to the intestinal physiology of arginine, citrulline, ornithine, and cystine (which shares the same transporter) and the many clinical trials of supplements of the dibasic amino acids or N-acetylcysteine (NAC. The human intestinal dibasic amino acid transport system has high affinity and low capacity. L-Arginine (but not lysine, ornithine, or D-arginine) induces water and electrolyte secretion that is mediated by NO, which acts as an absorbagogue at low levels and as a secretagogue at high levels. The action of many laxatives is NO mediated and there are reports of diarrhea following oral administration of arginine or ornithine ihine. The clinical data cover a wide span of arginine intakes f rom 3 g/d to > 100 g/d, but the standard of reporting adverse effects (e.g. nausea, vomiting, and diarrhea) was variable. Single doses of 3-6 g rarely provoked side effects and healthy athletes appeared to be more susceptible than diabetic patients to gastrointestinal symptoms at individual doses >9 g. This may relate to an effect of disease on gastrointestinal motility and pharmacokinetics. Most side effects of arginine and NAC occurred at single doses of >9 g in adults >140 mg/kg) often when part of a daily regime of similar to>30 g/d (>174 mmol/d). In the case of arginine, this compares with the laxative threshold of the nonabsorbed disaccharide alcohol, lactitol (74 g or 194 mmol). Adverse effects seemed dependent on the dosage regime and disappeared if divided doses were ingested (unlike lactitol). Large single doses of poorly absorbed amino acids seem to provoke diarrhea. More research is needed to refine dosage strategies that reduce this phenomenon. It is suggested that dipeptide forms of arginine may meet this criterion.