Amino Acids (2007) 32: 543–551 DOI 10.1007/s00726-006-0430-y Printed in The Netherlands Sulfenic acid in human serum albumin Review Article S. Carballal 1;2;3 , B. Alvarez 1;3 , L. Turell 1;3 , H. Botti 2;3 , B. A. Freeman 4 , and R. Radi 2;3 1 Laboratorio de Enzimologı ´a, Facultad de Ciencias, Universidad de la Rep ublica, Montevideo, Uruguay 2 Departamento de Bioquı ´mica, Facultad de Medicina, Universidad de la Rep ublica, Montevideo, Uruguay 3 Center for Free Radical and Biomedical Research, Universidad de la Rep ublica, Montevideo, Uruguay 4 Department of Pharmacology, University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania, USA Received June 10, 2006 Accepted June 30, 2006 Published online October 24, 2006; # Springer-Verlag 2006 Summary. Sulfenic acid (RSOH) is a central intermediate in both the reversible and irreversible redox modulation by reactive species of an increasing number of proteins involved in signal transduction and en- zymatic pathways. In this paper we focus on human serum albumin (HSA), the most abundant plasma protein, proposed to serve antiox- idant functions in the vascular compartment. Sulfenic acid in HSA has been previously detected using different methods after oxidation of its single free thiol Cys34 through one- or two-electron mechanisms. Since recent evidence suggests that sulfenic acid in HSA is stabilized within the protein environment, this derivative represents an appropri- ate model to examine protein sulfenic acid biochemistry, structure and reactivity. Sulfenic acid in HSA could be involved in mixed disufide formation, supporting a role of HSA-Cys34 as an important redox reg- ulator in extracellular compartments. Keywords: Thiol – Human serum albumin – Sulfenic acid – Peroxyni- trite – Hydrogen peroxide – Free radicals Abbreviations: HSA, human serum albumin; HSA-SH, the thiol of HSA; BSA, bovine serum albumin; SH=HSA, amount of thiol per albumin molecule; RSOH, sulfenic acid; NBD-Cl, 7-chloro-4-nitrobenzo-2-oxa- 1,3-diazol; DTNB, 5,5 0 -dithio-bis(2-nitrobenzoic acid); dimedone, 5,5- dimethyl-1,3-cyclohexanedione; GSH, glutathione; GSSG, glutathione disulfide. Introduction The overproduction of reactive oxygen species including nitrogen oxides under conditions of oxidative stress may damage various biomolecules leading to cell and tissue toxicity. In addition, there is increasing evidence that these reactive mediators play a central role in signal trans- duction under physiological and physiopathological con- ditions, leading to new proposals on the definition of oxi- dative stress in terms of disruption of redox signaling and control (Hansen et al., 2006a). Thiols are preferential targets of reactive species. With one- and two-electron reduction potentials E 0 (RS  =RSH) of 0.92 V and E 0 (RSSR=2RSH) of 0.24 V for cysteine (Surdhar and Armstrong, 1986; Keire et al., 1992), thiols can be oxidized by a wide spectrum of radical and non- radical species, usually at rates several orders of magnitude faster than other amino acids. At the high concentrations that thiols can achieve in different cell compartments, thiols can effectively compete with other targets. Last, several thiol oxidation products can be efficiently repaired. In the last few years, the reactions of reactive species with thiols, far from constituting simple scavenging pathways, are being recognized to participate in modulatory and sig- naling mechanisms, coupling changes in the redox state of the milieu to biochemical and cellular processes (Schafer and Buettner, 2001). Among the different posttranslational modifications generated by thiol oxidation, sulfenic acid is receiving increased interest since it has been identified in a growing list of proteins, where it serves catalytic or regula- tory functions. In this paper we will focus on the formation of sulfenic acid in human serum albumin and revise its properties and possible biological function. Thiol oxidation states and sulfenic acid formation Oxidation of low molecular weight or protein thiols typ- ically results in the formation of disulfide bonds (RSSR).