Review ArticleNonequilibrium thermodynamics of thiol/disulfide redox systems: A perspective on redox systems biology
Introduction
Cysteine (Cys) and methionine (Met) are the only amino acids in proteins which contain elements undergoing reversible oxidation under biologic conditions. These elements are aptly described as “sulfur switches” because the reversible oxidations provide a means to control a broad range of activity and structure of proteins [1]. The sulfur atoms of both Cys and Met can undergo multiple oxidations, but the reversible oxidation of thiols to disulfides has been studied most extensively and serves as the basis for the current review. Changes in oxidation/reduction (redox) state of thiol/disulfide couples affect protein conformation, enzyme activity, transporter activity, ligand binding to receptors, protein–protein interactions, protein–DNA interactions, protein trafficking, and protein degradation [2], [3], [4], [5], [6].
Considerable knowledge exists concerning specific redox–dependent components in redox signaling, transcriptional regulation, cell proliferation, apoptosis, hormonal signaling, and other fundamental cell functions [7], [8], [9], [10]. These redox–dependent components exist in redox signaling pathways [11]: the biological significance of these signaling pathways [12], the multiplicity of such pathways [13], and the sensitivity of these pathways to disruption by oxidative stress [14] have led to a refinement in the definition of oxidative stress as “an imbalance in prooxidants and antioxidants with associated disruption of redox circuitry and macromolecular damage” [15], [16]. This definition accommodates the multiple redox–dependent pathways (circuits) that exist and the heterogeneity in responses which can occur due to imbalances of oxidative and reductive processes [15], [16], [17]. At the same time, this definition suggests a need for integrated network maps to guide the search for redox–sensitive sites which can serve as targets for therapeutic interventions to prevent and treat oxidative stress. There has been limited progress in developing such maps and integrating the multiple redox pathways and compartments into common models.
The developing discipline of “systems biology” offers novel approaches for understanding the complex regulation of interacting redox networks. Established modeling formalisms, such as biochemical systems theory [18] and metabolic control analysis [19], which were once limited to metabolic pathways have been expanded as signal transduction and gene regulation regulatory networks have demonstrated similar control motifs [20]. Emerging principles in systems biology include the necessity of control points within a network (often referred as nodes) which serve as regulators of information or substrate flow. The realization that large biological networks can effectively be organized into functional modules [21] linked together by control nodes has allowed researchers to tease apart the spaghetti diagrams into tractable pieces for validation and testing. The pace of scientific discovery and emphasis on cataloging and databasing [22], [23], [24] have increased the ability of descriptive kinetic modeling at every organizational level within the cell. Most importantly, the availability of high-throughput technologies for rapid, large-scale data acquisition has allowed novel data analysis tools to be developed for inferring network structures and extracting relationships between components. These computational and experimental tools can provide complementary approaches to traditional reductionist strategies for determining the interconnectivity between multiple redox couples that function as control nodes and their respective molecular targets.
The present review focuses on the redox states, stability, and control mechanisms of the central thiol/disulfide redox couples (thioredoxins, GSH/GSSG, and Cys/CySS) in mammalian cells with the intent to promote development of redox systems biology. These couples have previously been described as “redox control nodes” within electron–conducting circuits [15], [25]. These redox control nodes appear to regulate the redox state of different sets of target proteins [26]. The purpose of the present review is to summarize the current knowledge of steady-state redox potentials of these redox control nodes and provide a framework for development of kinetic models which can describe the relatively stable nonequilibrium steady-state values of these redox couples and their functions in redox biology. A long-term goal of such an approach is to describe the steady-state redox potentials of individual redox couples as a function of all of the interacting electron transfer reactions within and between the cellular compartments.
The first section summarizes studies of the nonequilibrium redox states of the major thiol/disulfide redox couples in the subcellular and extracellular compartments. The second section contains an overview of some of the evolving principles which can be used to formulate models for redox systems biology. The third describes elements of kinetic models needed to account for nonequilibrium steady states of sulfur switches, using the GSH/GSSG couple as an example. This example illustrates the need for rate information for each step of biosynthesis, degradation/metabolism, transport, oxidation, and reduction, to describe the steady-state relationships. The fourth section provides a framework to extend this approach to a more comprehensive systems biology description of redox signaling and control. The last section briefly considers the potential utility of such models for predictive health and development of new strategies to protect against disease.
Section snippets
Major thiol/disulfide couples are not at equilibrium in biologic systems
The redox potential (electromotive force, Eh) is a measure of the tendency of a chemical species to accept or donate electrons. This tendency is quantitatively expressed in millivolts relative to the standard hydrogen electrode reaction (H2/2H+ + 2e−). The Eh for an oxidation/reduction couple (e.g., GSH/GSSG) is dependent on the inherent tendency of the chemical species to accept/donate electrons (Eo) and the concentrations of the respective acceptors and donors, defined by the Nernst equation
Formulation of models for thiol/disulfide redox systems biology
Most proteins contain at least one Cys or Met which is subject to oxidation, and all aspects of life depend on redox reactions. Consequently, there is considerable complexity to development of redox systems biology. Certain features are emerging which suggest that this complexity can be simplified by a set of rules or principles governing the biological redox reactions, which can be considered part of a biologic “redox code” [84], [85], [86]. For initial model development, these principles can
Kinetic models and potential mechanisms for control of nonequilibrium steady states
Electron transfer rates between specific components within biologic systems can vary considerably, but the overall rates are ultimately dictated by the rate of transfer to O2, the terminal electron acceptor for aerobic organisms. The maximal rate of O2 consumption by an athletically trained human is about 4 L/min, which is equivalent to about 2.5 mM min− 1 averaged throughout the body. This average equates to about 10 nmol/mg protein per minute at the cellular level, which is about 40% of
Perspectives for redox systems biology
Fig. 4, which is focused on the GSH/GSSG couple, delineates only one aspect of the overall cellular redox system including Trx couples and Cys/CySS. The description can be expanded to include the Trx and Cys/CySS couples by incorporating relevant fluxes and distributions across cellular compartments such as shown in Fig. 5. These redox control nodes, in turn, provide the set points by which different sulfur switches can operate independently. Because systems biology is driven to explore the
Acknowledgments
Research by the authors on which this review was based was supported by NIH Grants ES011195, ES009047, and ES012929, and by support from the Whitaker Foundation.
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