Thiolation and nitrosation of cysteines in biological fluids and cells

Abstract

Thiols (RSH) are potent nucleophilic agents, the rates of which depend on the pKa of the sulfhydryl. Unlike compounds having other nucleophile moieties (-OH or -NH(2)), RSH are involved in reactions, such as conjugations, redox and exchange reactions. Although protein SH groups (PSH) react like non-protein thiols (NPSH), the biochemistry of proteins is much more complex for reasons such as steric hindrance, charge distribution and accessibility of PSH to the solvent (protein conformation). The reaction rates and types of end-products of PSH vary a lot from protein to protein. The biological problem is even more complex because in all compartments and tissues, there may be specific competition between thiols (namely between GSH and PSH), regulated by the properties of antioxidant enzymes. Moreover, PSH are divided biologically into essential and non-essential and their respective influence in the various biological systems is unknown. It follows that during phenomena eliciting a prompt thiol response (oxidative stress), the antioxidant PSH response and reaction mechanisms vary considerably from case to case. For example, in spite of a relatively low pKa that should guarantee good antioxidant capacity, PSH of albumin has much less propensity to form adducts with conjugating agents than NPSH; moreover, the structural characteristics of the protein prevent albumin from forming protein disulfides when exposed to oxidants (whereas protein-thiol mixed disulfides are formed in relative abundance). On the other hand, proteins with a relatively high reactivity, such rat hemoglobin, have much greater antioxidant capacity than GSH, but although human hemoglobin has a pKa similar to GSH, for structural reasons it has less antioxidant capacity than GSH. When essential PSH are involved in S-thiolation and S-nitrosation reactions, a similar change in biological activity is observed. S-thiolated proteins are a recurrent phenomenon in oxidative stress elicited by reactive oxygen species (ROS). This event may be mediated by disulfides, that exchange with PSH, or by the protein intermediate sulfenic acid that reacts with thiols to form protein-mixed disulfides. During nitrosative stress elicited by reactive nitrogen species (RNS), depending on the oxygen concentration of the system, nitrosation reactions of thiols may also be accompanied by protein S-thiolation. In this review we discuss a number of cell processes and biochemical modifications of enzymes that indicate that S-thiolation and S-nitrosation may occur simultaneously in the same protein in the presence of appropriate interactions between ROS and RNS.

Accumulation of Free ADP-ribose from Mitochondria Mediates Oxidative Stress-induced Gating of TRPM2 Cation Channels

“…1), including exposure to MNNG, which in addition to inducing nitrosative stress has been shown to produce markedly diminished NAD levels in HEK cells over a time course in which we observed sustained TRPM2 gating (25). Furthermore, disulfide stress induced by the disulfide-inducing reagent diamide did not initiate TRPM2-dependent calcium transients (data not shown), indicating that the pathway leading to TRPM2 activation most likely involves oxidative/nitrosative modification of one or more protein thiol group(s) (2,26,27), as opposed to a more general effect on cellular redox balance. These observations are difficult to reconcile with a proposed model of TRPM2 activation involving accumulation of NAD because of generalized adenine nucleotide oxidation and its agonistic binding to the NUDT9-H domain (14), and suggested to us that oxidative/nitrosative stress-mediated TRPM2 activation must involve an alternative mechanism.…”

Factors Affecting S-Homocysteinylation of LDL Apoprotein B

“…Plasma S-thiolated proteins have been detected in healthy humans, in patients with cardiovascular diseases, and in several cell types after oxidant exposure (20 ). Although increased protein S-thiolation is considered an in vivo marker of oxidative injury caused by noxious agents (21,22 ), the importance of this index in clinical studies has not been well documented.…”

Widespread sulfenic acid formation in tissues in response to hydrogen peroxide

“…There is a growing awareness that reversible cysteine-targeted modifications of proteins, particularly S-thiolation and Snitrosylation, provide an additional mechanism by which proteins can be posttranslationally regulated (14). There is already some evidence that protein sulfenic acid formation is also important in regulation of protein function (13,15), but the full extent of these regulatory mechanisms is yet to be determined.…”

“…When H 2 O 2 is elevated, reactive protein thiols within the cell can become oxidized. A number of protein thiol oxidation states can be formed, including the S-thiolated and the S-nitrosylated form (14), which obviously require the presence of other chemical species.…”