Control of plant mitochondrial respiration

Plant mitochondria are characterised by the presence of both phosphorylating (cytochrome) and non-phosphorylating (alternative) respiratory pathways, the relative activities of which directly affect the efficiency of mitochondrial energy conservation. Different approaches to study the regulation of the partitioning of reducing equivalents between these routes are critically reviewed. Furthermore, an updated view is provided regarding the understanding of plant mitochondrial respiration in terms of metabolic control. We emphasise the extent to which kinetic modelling and 'top-down' metabolic control analysis improve the insight in phenomena related to plant mitochondrial respiration. This is illustrated with an example regarding the affinity of the plant alternative oxidase for oxygen.     

Kinetic modelling of plant metabolic pathways

This paper provides a review of kinetic modelling of plant metabolic pathways as a tool for analysing their control and regulation. An overview of different modelling strategies is presented, starting with those approaches that only require a knowledge of the network stoichiometry; these are referred to as structural. Flux-balance analysis, metabolic flux analysis using isotope labelling, and elementary mode analysis are briefly mentioned as three representative examples. The main focus of this paper, however, is a discussion of kinetic modelling, which requires, in addition to the stoichiometry, a knowledge of the kinetic properties of the constituent pathway enzymes. The different types of kinetic modelling analysis, namely time-course simulation, steady-state analysis, and metabolic control analysis, are explained in some detail. An overview is presented of strategies for obtaining model parameters, as well as software tools available for simulation of such models. The kinetic modelling approach is exemplified with discussion of three models from the general plant physiology literature. With the aid of kinetic modelling it is possible to perform a control analysis of a plant metabolic system, to identify potential targets for biotechnological manipulation, as well as to ascertain the regulatory importance of different enzymes (including isoforms of the same enzyme) in a pathway. Finally, a framework is presented for extending metabolic models to the whole-plant scale by linking biochemical reactions with diffusion and advective flow through the phloem. Future challenges include explicit modelling of subcellular compartments, as well as the integration of kinetic models on the different levels of the cellular and organizational hierarchy.     

The respiratory source of CO2

Abstract Approximately half of the carbon plants fix in photosynthesis is lost in dark respiration. The major pathways for dark respiration and their control are briefly discussed in the context of a growing plant. It is suggested that whole-plant respiration may be largely ADP-limited and that fine control of the respiratory network serves to select the respiratory substrate and to partition carbon between the numerous possible fates within the network. The striking stoichiometry between whole-plant growth and respiration is reviewed, and the relationships between substrate-limited growth and ADP-limited respiration are discussed.     

Experimental and mathematical approaches to modeling plant metabolic networks

To support their sessile and autotrophic lifestyle higher plants have evolved elaborate networks of metabolic pathways. Dynamic changes in these metabolic networks are among the developmental forces underlying the functional differentiation of organs, tissues and specialized cell types. They are also important in the various interactions of a plant with its environment. Further complexity is added by the extensive compartmentation of the various interconnected metabolic pathways in plants. Thus, although being used widely for assessing the control of metabolic flux in microbes, mathematical modeling approaches that require steady-state approximations are of limited utility for understanding complex plant metabolic networks. However, considerable progress has been made when manageable metabolic subsystems were studied. In this article, we will explain in general terms and using simple examples the concepts underlying stoichiometric modeling (metabolic flux analysis and metabolic pathway analysis) and kinetic approaches to modeling (including metabolic control analysis as a special case). Selected studies demonstrating the prospects of these approaches, or combinations of them, for understanding the control of flux through particular plant pathways are discussed. We argue that iterative cycles of (dry) mathematical modeling and (wet) laboratory testing will become increasingly important for simulating the distribution of flux in plant metabolic networks and deriving rational experimental designs for metabolic engineering efforts.     

Structural organization of the mitochondrial respiratory chain

Two models exist of the mitochondrial respiratory chain: the model of a random organization of the individual respiratory enzyme complexes and that of a super-complex assembly formed by stable association between the individual complexes. Recently Sch?gger, using digitonin solubilization and Blue Native PAGE produced new evidence of preferential associations, in particular a Complex I monomer with a Complex III dimer, and suggested a model of the respiratory chain (the respirasome) based on direct electron channelling between complexes. Discrimination between the two models is amenable to kinetic testing using flux control analysis. Experimental evidence obtained in beef heart SMP, according to the extension of the Metabolic Control Theory for pathways with metabolic channelling, showed that enzyme associations involving Complex I and Complex III take place in the respiratory chain while Complex IV seems to be randomly distributed, with cytochrome c behaving as a mobile component. Flux control analysis at anyone of the respiratory complexes involved in aerobic succinate oxidation indicated that Complex II and III are not functionally associated in a stable supercomplex. A critical appraisal of the solid-state model of the mitochondrial respiratory chain requires its reconciliation with previous biophysical and kinetic evidence that CoQ behaves as a homogeneous diffusible pool between all reducing enzyme and all oxidizing enzymes: the hypothesis can be advanced that both models (CoQ pool and supercomplexes) are true, by postulating that supercomplexes physiologically exist in equilibrium with isolated complexes depending on metabolic conditions of the cell.     

A kinetic model describes metabolic response to perturbations and distribution of flux control in the benzenoid network of Petunia hybrida

In recent years there has been much interest in the genetic enhancement of plant metabolism; however, attempts at genetic modification are often unsuccessful due to an incomplete understanding of network dynamics and their regulatory properties. Kinetic modeling of plant metabolic networks can provide predictive information on network control and response to genetic perturbations, which allow estimation of flux at any concentration of intermediate or enzyme in the system. In this research, a kinetic model of the benzenoid network was developed to simulate whole network responses to different concentrations of supplied phenylalanine (Phe) in petunia flowers and capture flux redistributions caused by genetic manipulations. Kinetic parameters were obtained by network decomposition and non‐linear least squares optimization of data from petunia flowers supplied with either 75 or 150 mm ²H₅‐Phe. A single set of kinetic parameters simultaneously accommodated labeling and pool size data obtained for all endogenous and emitted volatiles at the two concentrations of supplied ²H₅‐Phe. The generated kinetic model was validated using flowers from transgenic petunia plants in which benzyl CoA:benzyl alcohol/phenylethanol benzoyltransferase (BPBT) was down‐regulated via RNAi. The determined in vivo kinetic parameters were used for metabolic control analysis, in which flux control coefficients were calculated for fluxes around the key branch point at Phe and revealed that phenylacetaldehyde synthase activity is the primary controlling factor for the phenylacetaldehyde branch of the benzenoid network. In contrast, control of flux through the β‐oxidative and non‐β‐oxidative pathways is highly distributed.     

Potential impacts of climate change on Sub-Saharan African plant priority area selection

The Global Strategy for Plant Conservation (GSPC) aims to protect 50% of the most important areas for plant diversity by 2010. This study selects sets of 1-degree grid cells for 37 sub-Saharan African countries on the basis of a large database of plant species distributions. We use two reserve selection algorithms that attempt to satisfy two of the criteria set by the GSPC. The grid cells selected as important plant cells (IPCs) are compared between algorithms and in terms of country and continental rankings between cells. The conservation value of the selected grid cells are then considered in relation to their future species complement given the predicted climate change in three future periods (2025, 2055, and 2085). This analysis uses predicted climate suitability for individual species from a previous modelling exercise.
We find that a country-by-country conservation approach is suitable for capturing most, but not all, continentally IPCs. The complementarity-based reserve selection algorithms suggest conservation of a similar set of grid cells, suggesting that areas of high plant diversity and rarity may be well protected by a single pattern of conservation activity.
Although climatic conditions are predicted to deteriorate for many species under predicted climate change, the cells selected by the algorithms are less affected by climate change predictions than non-selected cells. For the plant species that maintain areas of climatic suitability in the future, the selected set will include cells with climate that is highly suitable for the species in the future. The selected cells are also predicted to conserve a large proportion of the species richness remaining across the continent under climate change, despite the network of cells being less optimal in terms of future predicted distributions.
Limitations to the modelling are discussed in relation to the policy implications for those implementing the GSPC.     

Effect of regulatory mechanism on hyperbolic reaction network properties

The notion that the regulated and flux-controlling enzyme in a metabolic network need not correspond suggests that the purpose of regulation may not be flux homeostasis under all physiological circumstances. Additionally, the fact that diversity in the function of intact metabolic networks exists suggests that in addition to time constant separation, other kinetic structure/regulatory mechanism patterns exist. In order to compliment and expand prior work on identifying kinetic structure-property relationships in networks, the present work explores in a general way how the control, dynamic, and energetic properties of metabolic networks depend on operating point, kinetic structure, and regulatory mechanism. The basic feature of trade-offs between properties is illustrated and used as a basis for indicating how particular subsets of structure, regulatory mechanism, and operating point emphasize certain properties that can be associated with a physiological function. Examples of scavenging trace metabolites and amphibolite coordination are proposed. Microstructure logic in terms of turnover number distributions as well as a potential mixed polynomial network analysis approach are also discussed.     

A critique of the use of inhibitors to estimate partitioning of electrons between mitochondrial respiratory pathways in plants

The contribution of individual plant mitochondrial respiratory pathways to total respiration is commonly assessed by titration with specific inhibitors of different components in the branched electron transport chain. A pathway's contribution is equal to the activity when the other branch is blocked by an inhibitor multiplied by the degree (0-1.0) to which this activity is engaged when both pathways are operating. According to Bahr and Bonner (1973. J. Biol. Chem. 218: 3441–3445) the plot of the activities of identical titrations, one performed in the absence and the other in the presence of a specific inhibitor of the other branch of the respiratory chain, yields a straight line whose slope indicates the engagement of the titrated pathway during uninhibited respiration. An initial slope of zero may occur if electron flux is diverted between pathways during titrations. However, beyond the breakpoint (representing the point of pathway saturation), a straight line is obtained with a slope representing engagement. This technique assumes that the kinetics of inhibiting a specific component of the respiratory chain are independent of the absolute rate of electron flux through the total pathway. To test this assumption, the activity of respiratory pathways in isolated soybean (Glycine max [L]. Merr. cv. Stevens) mitochondria was titrated with specific inhibitors of the cytochrome and alternative oxidases. Under these conditions, the electron flux through a given pathway was manipulated by poising the rate of succinate oxidation with the succinate dehydrogenase inhibitor malonate. Construction of activity plots in the presence versus absence of malonate failed to result in straight lines for either KCN (when titrating the cytochrome pathway) or salicylhydroxamic acid (when titrating the alternative pathway). Rather, the resultant plots were always curvilinear whenever the activity in the presence of malonate divided by the activity in the absence of malonate was less than 1.0. In no case could the real engagement of the pathway be precisely estimated from the titration data. Titrations of cytochrome pathway activity in isolated potato tuber (Solanum tuberosum L. cv. Sabago and Canabex) mitochondria (which lack the alternative oxidase) showed that as the inhibitor concentration was increased, so did the reduction status of the ubiquinone pool, to a new steady state. The dependence of inhibition kinetics on the rate of flux through the pathway, and the increase in ubiquinone pool reduction upon KCN addition, are explained in terms of the elasticity of component enzymes as outlined in the theory of metabolic control analysis. The implications of this finding for the use of titrations to estimate engagement of plant respiratory pathways are discussed.     

高等植物对环境胁迫的适应与其胁迫信号的转导

高等植物适应环境胁迫有多种水平与尺度的生理与生化方式,但其本质却是分子水平的基因时空表达与调控,它又受到胁迫信号转导途径的多重调控与影响。环境胁迫的主要形式是冷害、干旱、盐碱胁迫与UV-B辐射等,而它们又是影响高等植物生长、发育、繁殖等重要过程的生态因子,同时也是作物高效生产必需重视的因素,对其与植物相互作用的分子机理的认识有重要理论意义与实践意义。从细胞与组织和器官水平获得的分子生物学规律,只有应用到个体,群体,及生态系统中才会更有生命力。如何将这些数据资料成为宝贵的永续资源是21世纪植物系统生物学面临的主要挑战之一。主要从农业生态环境角度阐述环境胁迫信号转导的分子生物学作用方式,新进展资料的整合并建立起它们的可能联系及本领域中存在的相关问题和可能的解决途径,为高效的农业生态可持续发展提供分子生物学方面的理论基础。     

A computational model of a class of gene networks with positive and negative controls

This article proposes a computational framework for modelling the logical behavior of a class of gene networks. We characterize the basic behavior of genes in terms of a state-and-transition structure, and model the individual genes as language-generating automata. We consider positive and negative controls as the interaction mechanisms among the genes, and treat such controls as constraints (also expressed in automata) imposed on the behavior of the gene network. By computing the intersection of the languages generated by the gene models and the constraints, we obtain the complete set of pathways in a gene network. Implications and possible improvement of this work are discussed.     

Something from nothing: bridging the gap between constraint-based and kinetic modelling

Two divergent modelling methodologies have been adopted to increase our understanding of metabolism and its regulation. Constraint-based modelling highlights the optimal path through a stoichiometric network within certain physicochemical constraints. Such an approach requires minimal biological data to make quantitative inferences about network behaviour; however, constraint-based modelling is unable to give an insight into cellular substrate concentrations. In contrast, kinetic modelling aims to characterize fully the mechanics of each enzymatic reaction. This approach suffers because parameterizing mechanistic models is both costly and time-consuming. In this paper, we outline a method for developing a kinetic model for a metabolic network, based solely on the knowledge of reaction stoichiometries. Fluxes through the system, estimated by flux balance analysis, are allowed to vary dynamically according to linlog kinetics. Elasticities are estimated from stoichiometric considerations. When compared to a popular branched model of yeast glycolysis, we observe an excellent agreement between the real and approximate models, despite the absence of (and indeed the requirement for) experimental data for kinetic constants. Moreover, using this particular methodology affords us analytical forms for steady state determination, stability analyses and studies of dynamical behaviour.     

Small-angle scattering: a view on the properties, structures and structural changes of biological macromolecules in solution

A self-contained presentation of the main concepts and methods for interpretation of X-ray and neutron-scattering patterns of biological macromolecules in solution, including a reminder of the basics of X-ray and neutron scattering and a brief overview of relevant aspects of modern instrumentation, is given. For monodisperse solutions the experimental data yield the scattering intensity of the macromolecules, which depends on the contrast between the solvent and the particles as well as on their shape and internal scattering density fluctuations, and the structure factor, which is related to the interactions between macromolecules. After a brief analysis of the information content of the scattering intensity, the two main approaches for modelling the shape and/or structure of macromolecules and the global minimization schemes used in the calculations are presented. The first approach is based, in its more advanced version, on the spherical harmonics approximation and relies on few parameters, whereas the second one uses bead models with thousands of parameters. Extensions of bead modelling can be used to model domain structure and missing parts in high-resolution structures. Methods for computing the scattering patterns from atomic models including the contribution of the hydration shell are discussed and examples are given, which also illustrate that significant differences sometimes exist between crystal and solution structures. These differences are in some cases explainable in terms of rigid-body motions of parts of the structures. Results of two extensive studies--on ribosomes and on the allosteric protein aspartate transcarbamoylase--illustrate the application of the various methods. The unique bridge between equilibrium structures and thermodynamic or kinetic aspects provided by scattering techniques is illustrated by modelling of intermolecular interactions, including crystallization, based on an analysis of the structure factor and recent time-resolved work on assembly and protein folding.     

Power-law models of signal transduction pathways

The mathematical modelling of signal transduction pathways has become a valuable aid to understanding the complex interactions involved in intracellular signalling mechanisms. An important aspect of the mathematical modelling process is the selection of the model type and structure. Until recently, the convention has been to use a standard kinetic model, often with the Michaelis-Menten steady state assumption. However this model form, although valuable, is only one of a number of choices, and the aim of this article is to consider the mathematical structure and essential features of an alternative model form--the power-law model. Specifically, we analyse how power-law models can be applied to increase our understanding of signal transduction pathways when there may be limited prior information. We distinguish between two kinds of power law models: a) Detailed power-law models, as a tool for investigating pathways when the structure of protein-protein interactions is completely known, and; b) Simplified power-law models, for the analysis of systems with incomplete structural information or insufficient quantitative data for generating detailed models. If sufficient data of high quality are available, the advantage of detailed power-law models is that they are more realistic representations of non-homogenous or crowded cellular environments. The advantages of the simplified power-law model formulation are illustrated using some case studies in cell signalling. In particular, the investigation on the effects of signal inhibition and feedback loops and the validation of structural hypotheses are discussed.     

Understanding the regulation of aspartate metabolism using a model based on measured kinetic parameters

The aspartate‐derived amino‐acid pathway from plants is well suited for analysing the function of the allosteric network of interactions in branched pathways. For this purpose, a detailed kinetic model of the system in the plant model Arabidopsis was constructed on the basis of in vitro kinetic measurements. The data, assembled into a mathematical model, reproduce in vivo measurements and also provide non‐intuitive predictions. A crucial result is the identification of allosteric interactions whose function is not to couple demand and supply but to maintain a high independence between fluxes in competing pathways. In addition, the model shows that enzyme isoforms are not functionally redundant, because they contribute unequally to the flux and its regulation. Another result is the identification of the threonine concentration as the most sensitive variable in the system, suggesting a regulatory role for threonine at a higher level of integration.     

Common reeds (Phragmites australis) as sustainable energy source: experimental and modelling analysis of torrefaction and pyrolysis processes

The aim of this study is to apply advanced analytical techniques and kinetic modelling to common reeds (Phragmites australis) to characterize its pyrolysis and torrefaction as possible environmental friendly and sustainable pathways of fuel upgrading. Simultaneous thermogravimetric and differential scanning calorimetry analysis have been carried out on common reeds. The evolved gases during the decomposition process have been analysed by a coupled infrared gas analyser and gas chromatograph/mass spectrometer. Different reed origins (China and Italy) and plant parts (stem and leaves) have been compared. The results have been used to calibrate a torrefaction kinetic model. The model has also been tested simulating a reed torrefaction run occurring in a bench‐scale apparatus, supplementing the chemical analysis with a thermal simulation of the reactor carried out through a finite elements approach. The results show that the proposed modelling approach allows the prediction of the reaction products with a satisfying degree of accuracy. Besides its phytodepuration potential, P. australis has proven to be an interesting natural biomass resource for thermochemical conversion processes and energy production both for its suitability and availability.     

Bridging from molecular simulation to biochemical networks

How can we make the connection between the three-dimensional structures of individual proteins and understanding how complex biological systems involving many proteins work? The modelling and simulation of protein structures can help to answer this question for systems ranging from multimacromolecular complexes to organelles and cells. On one hand, multiscale modelling and simulation techniques are advancing to permit the spatial and temporal properties of large systems to be simulated using atomic-detail structures. On the other hand, the estimation of kinetic parameters for the mathematical modelling of biochemical pathways using protein structure information provides a basis for iterative manipulation of biochemical pathways guided by protein structure. Recent advances include the structural modelling of protein complexes on the genomic level, novel coarse-graining strategies to increase the size of the system and the time span that can be simulated, and comparative molecular field analyses to estimate enzyme kinetic parameters.