Impact of calcium on salivary α-amylase activity, starch paste apparent viscosity and thickness perception

Thickness perception of starch-thickened products during eating has been linked to starch viscosity and salivary amylase activity. Calcium is an essential cofactor for α-amylase and there is anecdotal evidence that adding extra calcium affects amylase activity in processes like mashing of beer. The aims of this paper were to (1) investigate the role of salivary calcium on α-amylase activity and (2) to measure the effect of calcium concentration on apparent viscosity and thickness perception when interacting with salivary α-amylase in starch-based samples. α-Amylase activity in saliva samples from 28 people was assessed using a typical starch pasting cycle (up to 95 °C). The activity of the enzyme (as measured by the change in starch apparent viscosity) was maintained by the presence of calcium, probably by protecting the enzyme from heat denaturation. Enhancement of α-amylase activity by calcium at 37 °C was also observed although to a smaller extent. Sensory analysis showed a general trend of decreased thickness perception in the presence of calcium, but the result was only significant for one pair of samples, suggesting a limited impact of calcium enhanced enzyme activity on perceived thickness

In vitro effects of zinc on the cytokine production from peripheral blood mononuclear cells in patients with zinc allergy.

Metals, such as nickel, cobalt, chromium and zinc, are ubiquitous in the environment. Systemic reactions, including hand dermatitis and generalized eczematous reactions, can be caused by the dietary ingestion of metals. In this study, we aimed to determine whether the cytokine production from peripheral blood mononuclear cells (PBMCs) obtained from zinc allergy patients can be used as a sensitive marker to investigate zinc-allergic contact dermatitis. The diagnosis of sensitivity to metal was made based on the results of a metal patch test. The PBMCs were stimulated with various concentrations (5-100 μM) of zinc sulfate (ZnSO4) for 24 h. The culture supernatants were collected and analyzed using ELISA for measurement of the cytokine production. The levels of IFN-γ, TNF-α, IL-1β, IL-5, IL-13 and MIF were significantly higher in the zinc-allergic patients (n = 5) than in the healthy controls (n = 5) at 100 μM of ZnSO4 stimulation. Although, patch testing is considered as standard test to diagnose metal allergy but false-positive and -negative reactions may limit its use in conditions of existing dermatitis. Therefore, this study suggest that in support of patch testing the determination of cytokine production using PBMCs cultures would be helpful for making an early diagnosis of such conditions

Predicting zinc binding at the proteome level

BACKGROUND: Metalloproteins are proteins capable of binding one or more metal ions, which may be required for their biological function, for regulation of their activities or for structural purposes. Metal-binding properties remain difficult to predict as well as to investigate experimentally at the whole-proteome level. Consequently, the current knowledge about metalloproteins is only partial. RESULTS: The present work reports on the development of a machine learning method for the prediction of the zinc-binding state of pairs of nearby amino-acids, using predictors based on support vector machines. The predictor was trained using chains containing zinc-binding sites and non-metalloproteins in order to provide positive and negative examples. Results based on strong non-redundancy tests prove that (1) zinc-binding residues can be predicted and (2) modelling the correlation between the binding state of nearby residues significantly improves performance. The trained predictor was then applied to the human proteome. The present results were in good agreement with the outcomes of previous, highly manually curated, efforts for the identification of human zinc-binding proteins. Some unprecedented zinc-binding sites could be identified, and were further validated through structural modelling. The software implementing the predictor is freely available at: CONCLUSION: The proposed approach constitutes a highly automated tool for the identification of metalloproteins, which provides results of comparable quality with respect to highly manually refined predictions. The ability to model correlations between pairwise residues allows it to obtain a significant improvement over standard 1D based approaches. In addition, the method permits the identification of unprecedented metal sites, providing important hints for the work of experimentalists

Transcriptional regulation of the cinnamyl alcohol dehydrogenase gene from sweetpotato in response to plant developmental stage and environmental stress

Cinnamyl alcohol dehydrogenase (CAD) is a key enzyme in the biosynthesis of lignin. We have isolated full length of a cDNA encoding CAD (IbCAD1) that was previously identified as the most abundant gene in an EST library of sweetpotato suspension cells. Phylogenetic analysis revealed that IbCAD1 belongs to the family of defense-related CADs. High levels of IbCAD1 mRNA were found in the roots of sweetpotato, but not in the leaves and petioles. The IbCAD1 gene transcripts were highly induced by cold, wounding, and reactive oxygen species. Analyses of transcriptional regulation of the IbCAD1 gene in transgenic tobacco plants carrying the IbCAD1 promoter–GUS revealed that IbCAD1 promoter expression was strong in the roots, but barely detectable in the cotyledons. IbCAD1 promoter activity increased with increasing root age, and strong promoter expression was observed in the lateral root emergence sites and in root tips. Weak GUS expression was observed in lignified tissues of vascular system of mature leaves and stems. IbCAD1 promoter activity was strongly induced in response to the biotic and abiotic stresses, with the strongest inducer being wounding, and was also induced by salicylic acid (SA) and jasmonic acid (JA) as well as by abscisic acid (ABA) and 6-benzylaminopurine. Taken together, our data suggest that IbCAD1 can be involved in JA- and SA-mediated wounding response and ABA-mediated cold response, respectively. The IbCAD1 gene may play a role in the resistance mechanism to biotic and abiotic stresses as well as in tissue-specific developmental lignification

Caenorhabditis elegans Battling Starvation Stress: Low Levels of Ethanol Prolong Lifespan in L1 Larvae

The nematode Caenorhabditis elegans arrests development at the first larval stage if food is not present upon hatching. Larvae in this stage provide an excellent model for studying stress responses during development. We found that supplementing starved larvae with ethanol markedly extends their lifespan within this L1 diapause. The effects of ethanol-induced lifespan extension can be observed when the ethanol is added to the medium at any time between 0 and 10 days after hatching. The lowest ethanol concentration that extended lifespan was 1 mM (0.005%); higher concentrations to 68 mM (0.4%) did not result in increased survival. In spite of their extended survival, larvae did not progress to the L2 stage. Supplementing starved cultures with n-propanol and n-butanol also extended lifespan, but methanol and isopropanol had no measurable effect. Mass spectrometry analysis of nematode fatty acids and amino acids revealed that L1 larvae can incorporate atoms from ethanol into both types of molecules. Based on these data, we suggest that ethanol supplementation may extend the lifespan of L1 larvae by either serving as a carbon and energy source and/or by inducing a stress response

Clioquinol Inhibits Zinc-Triggered Caspase Activation in the Hippocampal CA1 Region of a Global Ischemic Gerbil Model

Background: Excessive release of chelatable zinc from excitatory synaptic vesicles is involved in the pathogenesis of selective neuronal cell death following transient forebrain ischemia. The present study was designed to examine the neuroprotective effect of a membrane-permeable zinc chelator, clioquinol (CQ), in the CA1 region of the gerbil hippocampus after transient global ischemia. Methodology/Principal Findings: The common carotid arteries were occluded bilaterally, and CQ (10 mg/kg, i.p.) was injected into gerbils once a day. The zinc chelating effect of CQ was examined with TSQ fluorescence and autometallography. Neuronal death, the expression levels of caspases and apoptosis inducing factor (AIF) were evaluated using TUNEL, in situ hybridization and Western blotting, respectively. We were able to show for the first time that CQ treatment attenuates the ischemia-induced zinc accumulation in the CA1 pyramidal neurons, accompanied by less neuronal loss in the CA1 field of the hippocampus after ischemia. Furthermore, the expression levels of caspase-3,-9, and AIF were significantly decreased in the hippocampus of CQ-treated gerbils. Conclusions/Significance: The present study indicates that the neuroprotective effect of CQ is related to downregulation o

Minimal Functional Sites Allow a Classification of Zinc Sites in Proteins

Zinc is indispensable to all forms of life as it is an essential component of many different proteins involved in a wide range of biological processes. Not differently from other metals, zinc in proteins can play different roles that depend on the features of the metal-binding site. In this work, we describe zinc sites in proteins with known structure by means of three-dimensional templates that can be automatically extracted from PDB files and consist of the protein structure around the metal, including the zinc ligands and the residues in close spatial proximity to the ligands. This definition is devised to intrinsically capture the features of the local protein environment that can affect metal function, and corresponds to what we call a minimal functional site (MFS). We used MFSs to classify all zinc sites whose structures are available in the PDB and combined this classification with functional annotation as available in the literature. We classified 77% of zinc sites into ten clusters, each grouping zinc sites with structures that are highly similar, and an additional 16% into seven pseudo-clusters, each grouping zinc sites with structures that are only broadly similar. Sites where zinc plays a structural role are predominant in eight clusters and in two pseudo-clusters, while sites where zinc plays a catalytic role are predominant in two clusters and in five pseudo-clusters. We also analyzed the amino acid composition of the coordination sphere of zinc as a function of its role in the protein, highlighting trends and exceptions. In a period when the number of known zinc proteins is expected to grow further with the increasing awareness of the cellular mechanisms of zinc homeostasis, this classification represents a valuable basis for structure-function studies of zinc proteins, with broad applications in biochemistry, molecular pharmacology and de novo protein design

Loss of Pluripotency in Human Embryonic Stem Cells Directly Correlates with an Increase in Nuclear Zinc

The pluripotency of human embryonic stem cells (hESCs) is important to investigations of early development and to cell replacement therapy, but the mechanism behind pluripotency is incompletely understood. Zinc has been shown to play a key role in differentiation of non-pluripotent cell types, but here its role in hESCs is directly examined. By mapping the distribution of metals in hESCs at high resolution by x-ray fluorescence microprobe (XFM) and by analyzing subcellular metal content, we have found evidence that loss of pluripotency is directly correlated with an increase in nuclear zinc. Zinc elevation not only redefines our understanding of the mechanisms that support pluripotency, but also may act as a biomarker and an intervention point for stem cell differentiation

Angiotensin II Requires Zinc and Downregulation of the Zinc Transporters ZnT3 and ZnT10 to Induce Senescence of Vascular Smooth Muscle Cells

Senescence, a hallmark of mammalian aging, is associated with the onset and progression of cardiovascular disease. Angiotensin II (Ang II) signaling and zinc homeostasis dysfunction are increased with age and are linked to cardiovascular disease, but the relationship among these processes has not been investigated. We used a model of cellular senescence induced by Ang II in vascular smooth muscle cells (VSMCs) to explore the role of zinc in vascular dysfunction. We found that Ang II-induced senescence is a zinc-dependent pathway mediated by the downregulation of the zinc transporters ZnT3 and ZnT10, which work to reduce cytosolic zinc. Zinc mimics Ang II by increasing reactive oxygen species (ROS), activating NADPH oxidase activity and Akt, and by downregulating ZnT3 and ZnT10 and inducing senescence. Zinc increases Ang II-induced senescence, while the zinc chelator TPEN, as well as overexpression of ZnT3 or ZnT10, decreases ROS and prevents senescence. Using HEK293 cells, we found that ZnT10 localizes in recycling endosomes and transports zinc into vesicles to prevent zinc toxicity. Zinc and ZnT3/ZnT10 downregulation induces senescence by decreasing the expression of catalase. Consistently, ZnT3 and ZnT10 downregulation by siRNA increases ROS while downregulation of catalase by siRNA induces senescence. Zinc, siZnT3 and siZnT10 downregulate catalase by a post-transcriptional mechanism mediated by decreased phosphorylation of ERK1/2. These data demonstrate that zinc homeostasis dysfunction by decreased expression of ZnT3 or ZnT10 promotes senescence and that Ang II-induced senescence is a zinc and ROS-dependent process. Our studies suggest that zinc might also affect other ROS-dependent processes induced by Ang II, such as hypertrophy and migration of smooth muscle cells

Interaction Between Marginal Zinc and High Fat Supply on Lipid Metabolism and Growth of Weanling Rats

The impact of a moderate Zn deficiency on growth and plasma and liver lipids was investigated in two 4-week experiments with male weanling rats fed fat-enriched diets. Semisynthetic, approximately isocaloric diets containing 3% soybean oil were supplemented with either 7 or 100 mg Zn/kg diet and with 22% beef tallow (BT) or sunflower oil (SF). In Experiment 1, which compared the dietary fat level and the fat source in a factorial design of treatments, all diets were fed ad libitum to 6 × 8 animals, whereas intake of the high-Zn BT and SF diets was restricted in Experiment 2 (5 × 6 rats) to the level of intake of the respective low-Zn diets. The low-Zn SF diet consistently depressed food intake and final live weights of the animals to a greater extent than the other low-Zn diets, while intake and growth were comparable among the animals fed the high-Zn diets. The marginal Zn deficit per se did not alter plasma triglyceride and cholesterol concentrations nor hepatic concentrations of triglyceride, cholesterol and phospholipids. The fatty acid pattern of liver phospholipids did not indicate that chain elongation and desaturation of fatty acids was impaired by a lack of zinc. It was concluded that dietary energy and fat intake, and fat source have a greater effect on plasma and liver lipids than a moderate Zn deficiency. Marginally Zn-deficient diets enriched with sunflower oil as a major energy source cause a greater growth retardation than diets rich in carbohydrates or beef tallow