Thermodynamic analysis of regulation in metabolic networks using constraint-based modeling

<p>Abstract</p> <p>Background</p> <p><it>Geobacter sulfurreducens </it>is a member of the <it>Geobacter </it>species, which are capable of oxidation of organic waste coupled to the reduction of heavy metals and electrode with applications in bioremediation and bioenergy generation. While the metabolism of this organism has been studied through the development of a stoichiometry based genome-scale metabolic model, the associated regulatory network has not yet been well studied. In this manuscript, we report on the implementation of a thermodynamics based metabolic flux model for <it>Geobacter sulfurreducens</it>. We use this updated model to identify reactions that are subject to regulatory control in the metabolic network of <it>G. sulfurreducens </it>using thermodynamic variability analysis.</p> <p>Findings</p> <p>As a first step, we have validated the regulatory sites and bottleneck reactions predicted by the thermodynamic flux analysis in <it>E. coli </it>by evaluating the expression ranges of the corresponding genes. We then identified ten reactions in the metabolic network of <it>G. sulfurreducens </it>that are predicted to be candidates for regulation. We then compared the free energy ranges for these reactions with the corresponding gene expression fold changes under conditions of different environmental and genetic perturbations and show that the model predictions of regulation are consistent with data. In addition, we also identify reactions that operate close to equilibrium and show that the experimentally determined exchange coefficient (a measure of reversibility) is significant for these reactions.</p> <p>Conclusions</p> <p>Application of the thermodynamic constraints resulted in identification of potential bottleneck reactions not only from the central metabolism but also from the nucleotide and amino acid subsystems, thereby showing the highly coupled nature of the thermodynamic constraints. In addition, thermodynamic variability analysis serves as a valuable tool in estimating the ranges of Δ_rG' of every reaction in the model leading to the prediction of regulatory sites in the metabolic network, thereby characterizing the regulatory network in both a model organism such as <it>E. coli </it>as well as a non model organism such as <it>G. sulfurreducens</it>.</p

A scalable algorithm to explore the Gibbs energy landscape of genome-scale metabolic networks

The integration of various types of genomic data into predictive models of biological networks is one of the main challenges currently faced by computational biology. Constraint-based models in particular play a key role in the attempt to obtain a quantitative understanding of cellular metabolism at genome scale. In essence, their goal is to frame the metabolic capabilities of an organism based on minimal assumptions that describe the steady states of the underlying reaction network via suitable stoichiometric constraints, specifically mass balance and energy balance (i.e. thermodynamic feasibility). The implementation of these requirements to generate viable configurations of reaction fluxes and/or to test given flux profiles for thermodynamic feasibility can however prove to be computationally intensive. We propose here a fast and scalable stoichiometry-based method to explore the Gibbs energy landscape of a biochemical network at steady state. The method is applied to the problem of reconstructing the Gibbs energy landscape underlying metabolic activity in the human red blood cell, and to that of identifying and removing thermodynamically infeasible reaction cycles in the Escherichia coli metabolic network (iAF1260). In the former case, we produce consistent predictions for chemical potentials (or log-concentrations) of intracellular metabolites; in the latter, we identify a restricted set of loops (23 in total) in the periplasmic and cytoplasmic core as the origin of thermodynamic infeasibility in a large sample ($10^6$) of flux configurations generated randomly and compatibly with the prior information available on reaction reversibility.Comment: 11 pages, 6 figures, 1 table; for associated supporting material see http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.100256

Fixation, transient landscape and diffusion's dilemma in stochastic evolutionary game dynamics

Agent-based stochastic models for finite populations have recently received much attention in the game theory of evolutionary dynamics. Both the ultimate fixation and the pre-fixation transient behavior are important to a full understanding of the dynamics. In this paper, we study the transient dynamics of the well-mixed Moran process through constructing a landscape function. It is shown that the landscape playing a central theoretical "device" that integrates several lines of inquiries: the stable behavior of the replicator dynamics, the long-time fixation, and continuous diffusion approximation associated with asymptotically large population. Several issues relating to the transient dynamics are discussed: (i) multiple time scales phenomenon associated with intra- and inter-attractoral dynamics; (ii) discontinuous transition in stochastically stationary process akin to Maxwell construction in equilibrium statistical physics; and (iii) the dilemma diffusion approximation facing as a continuous approximation of the discrete evolutionary dynamics. It is found that rare events with exponentially small probabilities, corresponding to the uphill movements and barrier crossing in the landscape with multiple wells that are made possible by strong nonlinear dynamics, plays an important role in understanding the origin of the complexity in evolutionary, nonlinear biological systems.Comment: 34 pages, 4 figure

Severe zinc depletion of escherichia coli: roles for high affinity zinc binding by ZinT, zinc transport and zinc-independent proteins

Zinc ions play indispensable roles in biological chemistry. However, bacteria have an impressive ability to acquire Zn2+ from the environment, making it exceptionally difficult to achieve Zn2+ deficiency, and so a comprehensive understanding of the importance of Zn2+ has not been attained. Reduction of the Zn2+ content of Escherichia coli growth medium to 60 nM or less is reported here for the first time, without recourse to chelators of poor specificity. Cells grown in Zn2+-deficient medium had a reduced growth rate and contained up to five times less cellular Zn2+. To understand global responses to Zn2+ deficiency, microarray analysis was conducted of cells grown under Zn2+-replete and Zn2+-depleted conditions in chemostat cultures. Nine genes were up-regulated more than 2-fold (p<0.05) in cells from Zn2+-deficient chemostats, including zinT (yodA). zinT is shown to be regulated by Zur ( zinc uptake regulator). A mutant lacking zinT displayed a growth defect and a 3-fold lowered cellular Zn2+ level under Zn2+ limitation. The purified ZinT protein possessed a single, high affinity metal-binding site that can accommodate Zn2+ or Cd2+. A further up-regulated gene, ykgM, is believed to encode a non-Zn2+ finger-containing paralogue of the Zn2+ finger ribosomal protein L31. The gene encoding the periplasmic Zn2+- binding protein znuA showed increased expression. During both batch and chemostat growth, cells "found" more Zn2+ than was originally added to the culture, presumably because of leaching from the culture vessel. Zn2+ elimination is shown to be a more precise method of depleting Zn2+ than by using the chelator N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine

Detailed Enzyme Kinetics in Terms of Biochemical Species: Study of Citrate Synthase

The compulsory-ordered ternary catalytic mechanism for two-substrate two-product enzymes is analyzed to account for binding of inhibitors to each of the four enzyme states and to maintain the relationship between the kinetic constants and the reaction equilibrium constant. The developed quasi-steady flux expression is applied to the analysis of data from citrate synthase to determine and parameterize a kinetic scheme in terms of biochemical species, in which the effects of pH, ionic strength, and cation binding to biochemical species are explicitly accounted for in the analysis of the data. This analysis provides a mechanistic model that is consistent with the data that have been used support competing hypotheses regarding the catalytic mechanism of this enzyme

Roles of the creatine kinase system and myoglobin in maintaining energetic state in the working heart

<p>Abstract</p> <p>Background</p> <p>The heart is capable of maintaining contractile function despite a transient decrease in blood flow and increase in cardiac ATP demand during systole. This study analyzes a previously developed model of cardiac energetics and oxygen transport to understand the roles of the creatine kinase system and myoglobin in maintaining the ATP hydrolysis potential during beat-to-beat transient changes in blood flow and ATP hydrolysis rate.</p> <p>Results</p> <p>The theoretical investigation demonstrates that elimination of myoglobin only slightly increases the predicted range of oscillation of cardiac oxygenation level during beat-to-beat transients in blood flow and ATP utilization. In silico elimination of myoglobin has almost no impact on the cytoplasmic ATP hydrolysis potential (Δ<it>G</it>_ATPase). In contrast, disabling the creatine kinase system results in considerable oscillations of cytoplasmic ADP and ATP levels and seriously deteriorates the stability of Δ<it>G</it>_ATPasein the beating heart.</p> <p>Conclusion</p> <p>The CK system stabilizes Δ<it>G</it>_ATPaseby both buffering ATP and ADP concentrations and enhancing the feedback signal of inorganic phosphate in regulating mitochondrial oxidative phosphorylation.</p

Modelling the impact of changes in the extracellular environment on the cytosolic free NAD+/NADH ratio during cell culture.

Cancer cells depend on glucose metabolism via glycolysis as a primary energy source, despite the presence of oxygen and fully functioning mitochondria, in order to promote growth, proliferation and longevity. Glycolysis relies upon NAD+ to accept electrons in the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) reaction, linking the redox state of the cytosolic NAD+ pool to glycolytic rate. The free cytosolic NAD+/NADH ratio is involved in over 700 oxidoreductive enzymatic reactions and as such, the NAD+/NADH ratio is regarded as a metabolic readout of overall cellular redox state. Many experimental techniques that monitor or measure total NAD+ and NADH are unable to distinguish between protein-bound and unbound forms. Yet total NAD+/NADH measurements yield little information, since it is the free forms of NAD+ and NADH that determine the kinetic and thermodynamic influence of redox potential on glycolytic rate. Indirect estimations of free NAD+/NADH are based on the lactate/pyruvate (L/P) ratio at chemical equilibrium, but these measurements are often undermined by high lability. To elucidate the sensitivity of the free NAD+/NADH ratio to changes in extracellular substrate, an in silico model of hepatocarcinoma glycolysis was constructed and validated against in vitro data. Model simulations reveal that over experimentally relevant concentrations, changes in extracellular glucose and lactate concentration during routine cancer cell culture can lead to significant deviations in the NAD+/NADH ratio. Based on the principles of chemical equilibrium, the model provides a platform from which experimentally challenging situations may be examined, suggesting that extracellular substrates play an important role in cellular redox and bioenergetic homeostasis

Different Prey Resources Suggest Little Competition Between Non-Native Frogs and Insectivorous Birds Despite Isotopic Niche Overlap

Non-native amphibians often compete with native amphibians in their introduced range, but their competitive effects on other vertebrates are less well known. The Puerto Rican coqui frog (Eleutherodactylus coqui) has colonized the island of Hawaii, and has been hypothesized to compete with insectivorous birds and bats. To address if the coqui could compete with these vertebrates, we used stable isotope analyses to compare the trophic position and isotopic niche overlap between the coqui, three insectivorous bird species, and the Hawaiian hoary bat. Coquis shared similar trophic position to Hawaii amakihi, Japanese white-eye, and red-billed leiothrix. Coquis were about 3 ‰ less enriched in δ15N than the Hawaiian hoary bat, suggesting the bats feed at a higher trophic level than coquis. Analyses of potential diet sources between coquis and each of the three bird species indicate that there was more dietary overlap between bird species than any of the birds and the coqui. Results suggest that Acari, Amphipoda, and Blattodea made up \u3e90% of coqui diet, while Araneae made up only 2% of coqui diet, but approximately 25% of amakihi and white-eye diet. The three bird species shared similar proportions of Lepidoptera larvae, which were ~25% of their diet. Results suggest that coquis share few food resources with insectivorous birds, but occupy a similar trophic position, which could indicate weak competition. However, resource competition may not be the only way coquis impact insectivorous birds, and future research should examine whether coqui invasions are associated with changes in bird abundance

Amphibians of Santa Teresa, Brazil: The Hotspot Further Evaluated

A checklist of the amphibians of Santa Teresa municipality, in southeastern Brazil is presented based on fieldwork, examination of specimens in collections, and a literature review. This new amphibian list of Santa Teresa includes 108 species, of which 106 (~98%) belong to Anura and two (~2%) to Gymnophiona. Hylidae was the most represented family with 47 species (43%). Compared to the previous amphibian lists for Santa Teresa, 14 species were added, 17 previously reported species were removed, and 13 species were re-identified based on recent taxonomic rearrangements. Of the 14 species added, 11 (79%) were first recorded during our fieldwork and specimen examination. It is also the first list of caecilians for Santa Teresa. This list suggests that Santa Teresa has 0.16 species per km2 (i.e., 108 species/683 km2), one of the highest densities of amphibian species in the world at a regional scale. This richness represents 78% of the 136 anurans from Espírito Santo state and 10% of the 1,080 amphibians from Brazil. We highlight the need for long-term monitoring to understand population trends and develop effective conservation plans to safeguard this remarkable amphibian richness

Learning in the Palaver Hut: The ‘Africa Study Visit’ as teaching tool.

noThe aim of this article is to assess the experiential learning environment of the African Study Visit (ASV). It presents a theoretically grounded analysis of the ASV. Although field visits are not a new phenomenon within Higher Education, they seem, but with few exceptions, to be considered as an add-on teaching method. By drawing from the experiential learning literature, we demonstrate that there are sound pedagogical reasons for incorporating field visits like the ASV into the curriculum as stand-alone components. Thus, the original contribution of this article is to place the ASV within the experiential learning literature such that the theoretical, practical and conceptual benefits for students are understood. Its significance is that this article offers a set of practices from an experiential learning perspective that can be used for deepening the levels of comprehension of political issues in Africa for international studies students