Regulatory Control or Oxidative Damage? Proteomic Approaches to Interrogate the Role of Cysteine Oxidation Status in Biological Processes

Abstract

Oxidation is a double-edged sword for cellular processes and its role in normal physiology, cancer and aging remains only partially understood. Although oxidative stress may disrupt biological function, oxidation-reduction (redox) reactions in a cell are often tightly regulated and play essential physiological roles. Cysteines lie at the interface between these extremes since the chemical properties that make specific thiols exquisitely redox-sensitive also predispose them to oxidative damage by reactive oxygen or nitrogen species during stress. Thus, these modifications can be either under reversible redox regulatory control or, alternatively, a result of reversible or irreversible oxidative damage. In either case, it has become increasingly important to assess the redox status of protein thiols since these modifications often impact such processes as catalytic activity, conformational alterations, or metal binding. To better understand the redox changes that accompany protein cysteine residues in complex biological systems, new experimental approaches have been developed to identify and characterize specific thiol modifications and/or changes in their overall redox status. In this review, we describe the recent technologies in redox proteomics that have pushed the boundaries for detecting and quantifying redox cysteine modifications in a cellular context. While there is no one-size-fits-all analytical solution, we highlight the rationale, strengths, and limitations of each technology in order to effectively apply them to specific biological questions. Several technological limitations still remain unsolved, however these approaches and future developments play an important role toward understanding the interplay between oxidative stress and redox signaling in health and disease. Molecular & Cellular Proteomics 11: 10.1074/mcp.R111.013037, 1-14, 2012. CYSTEINE: AN UNCOMMONLY REACTIVE AMINO ACIDThe nucleophilic sulfur atom allows cysteines to undergo a broad range of chemical modifications. These modifications include redox reactions, lipid acylation, and metal binding motifs that are important for protein structure, localization, regulation, and catalysis. Metal binding and oxidation play a role in protein structure through iron-sulfur (Fe-S) clusters, zinc fingers (ZF) 1 , and disulfide bonding, among others. Catalytic cysteines are essential to the function of numerous enzymes such as the E1 and E2 ligases of ubiquitin and ubiquitin-like proteins; the HECT domain of ubiquitin E3 ligases; SENP family sumo proteases; the tyrosine phosphatases protein phosphatase 1b (PTP1b) and PTEN; and many others including antioxidants in the thioredoxin, glutaredoxin, and peroxiredoxin families.Multiple thiol chemistries can converge to regulate the function of individual cysteines in a biological context. An example of the interconnection between the catalytic and redox properties of a cysteine is found in the cysteine-dependent aspartate-directed protease family of caspases. Essential to apoptosis, caspases are cysteine pr...

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