S-thiolation of cytoplasmic cardiac creatine kinase in heart cells treated with diamide

Collison, Mark W.; Thomas, James A.

doi:10.1016/0167-4889(87)90112-1

Cited by 53 publications

(21 citation statements)

References 24 publications

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“…The amount of S-thiolation induced by the thiol-selective diamide was considerably greater than that induced by reperfusion, indicating that only a small proportion of proteins are S-thiolated during the ischemia/ reperfusion protocol. Our findings agree with studies that have used myocytes or cell lines with 35 S-labeled GSH to show that S-glutathiolation occurs following treatment with chemical oxidants, which simulates the oxidative component of ischemia and reperfusion (19). Protein S-thiolation during reperfusion is expected because free radicals are known to be produced at this time (22).…”

Section: S-thiolation During Cardiac Oxidant Stress and Functionalsupporting

confidence: 81%

“…Many studies of protein S-thiolation during oxidant stress have used cultured cells with a radiolabeled thiol pool in which the Sthiolated proteins are detected by autoradiography of nonreducing SDS-PAGE gels (19). The investigation of specific proteins that are known to be S-thiolated has been achieved by non-reducing isoelectric-focusing gel electrophoresis coupled with Western blotting (8).…”

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confidence: 99%

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Detection, Quantitation, Purification, and Identification of Cardiac Proteins S-Thiolated during Ischemia and Reperfusion

Eaton¹,

Byers²,

Leeds³

et al. 2002

Journal of Biological Chemistry

167

133

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We have developed methods that allow detection, quantitation, purification, and identification of cardiac proteins S-thiolated during ischemia and reperfusion. Cysteine was biotinylated and loaded into isolated rat hearts. During oxidative stress, biotin-cysteine forms a disulfide bond with reactive protein cysteines, and these can be detected by probing Western blots with streptavidin-horseradish peroxidase. S-Thiolated proteins were purified using streptavidin-agarose. Thus, we demonstrated that reperfusion and diamide treatment increased S-thiolation of a number of cardiac proteins by 3-and 10-fold, respectively. Dithiothreitol treatment of homogenates fully abolished the signals detected. Fractionation studies indicated that the modified proteins are located within the cytosol, membrane, and myofilament/cytoskeletal compartments of the cardiac cells. This shows that biotin-cysteine gains rapid and efficient intracellular access and acts as a probe for reactive protein cysteines in all cellular locations. Using Western blotting of affinity-purified proteins we identified actin, glyceraldehyde-3-phosphate dehydrogenase, HSP27, protein-tyrosine phosphatase 1B, protein kinase C␣, and the small G-protein ras as substrates for S-thiolation during reperfusion of the ischemic rat heart. MALDI-TOF mass fingerprint analysis of tryptic peptides independently confirmed actin and glyceraldehyde-3-phosphate dehydrogenase S-thiolation during reperfusion. This approach has also shown that triosephosphate isomerase, aconitate hydratase, M-protein, nucleoside diphosphate kinase B, and myoglobin are S-thiolated during post-ischemic reperfusion.

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Section: S-thiolation During Cardiac Oxidant Stress and Functionalsupporting

confidence: 81%

mentioning

confidence: 99%

Detection, Quantitation, Purification, and Identification of Cardiac Proteins S-Thiolated during Ischemia and Reperfusion

Eaton¹,

Byers²,

Leeds³

et al. 2002

Journal of Biological Chemistry

167

133

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“…Early data demonstrating protein S-glutathiolation were largely derived from the exposure of cells to toxic levels of oxidants or activation of an inflammatory response from neutrophils (141), macrophages (961), and monocytes (764). Such studies readily demonstrated a number of S-glutathiolated proteins that spanned a broad range of cellular functions including metabolism (glyceraldehyde-3-phosphate dehydrogenase and creatine kinase) (164,764), the cytoskeleton (actin) (141), and antioxidant protection (SOD and glutathione transferase) (180,812) to name a few. For a more complete list of S-glutathiolated proteins, readers are directed to an excellent review (490).…”

Section: Targets For Rns In Cellular Signalingmentioning

confidence: 99%

Role of Oxidative Modifications in Atherosclerosis

Stocker¹,

Keaney²

2004

Physiological Reviews

2,258

1,756

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This review focuses on the role of oxidative processes in atherosclerosis and its resultant cardiovascular events. There is now a consensus that atherosclerosis represents a state of heightened oxidative stress characterized by lipid and protein oxidation in the vascular wall. The oxidative modification hypothesis of atherosclerosis predicts that low-density lipoprotein (LDL) oxidation is an early event in atherosclerosis and that oxidized LDL contributes to atherogenesis. In support of this hypothesis, oxidized LDL can support foam cell formation in vitro, the lipid in human lesions is substantially oxidized, there is evidence for the presence of oxidized LDL in vivo, oxidized LDL has a number of potentially proatherogenic activities, and several structurally unrelated antioxidants inhibit atherosclerosis in animals. An emerging consensus also underscores the importance in vascular disease of oxidative events in addition to LDL oxidation. These include the production of reactive oxygen and nitrogen species by vascular cells, as well as oxidative modifications contributing to important clinical manifestations of coronary artery disease such as endothelial dysfunction and plaque disruption. Despite these abundant data however, fundamental problems remain with implicating oxidative modification as a (requisite) pathophysiologically important cause for atherosclerosis. These include the poor performance of antioxidant strategies in limiting either atherosclerosis or cardiovascular events from atherosclerosis, and observations in animals that suggest dissociation between atherosclerosis and lipoprotein oxidation. Indeed, it remains to be established that oxidative events are a cause rather than an injurious response to atherogenesis. In this context, inflammation needs to be considered as a primary process of atherosclerosis, and oxidative stress as a secondary event. To address this issue, we have proposed an “oxidative response to inflammation” model as a means of reconciling the response-to-injury and oxidative modification hypotheses of atherosclerosis.

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“…The range of proteins that can be modified by S-thiolation has been determined in studies with human endothelial cells and murine macrophages (40,41). A few proteins that are S-thiolated in response to oxidants, including carbonic anhydrase III (4), actin (3), creatine kinase (8), glycogen phosphorylase b (37) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (39,42), have been positively identified in mammalian cells. The physiological relevance of the modification of these proteins is unclear.…”

mentioning

confidence: 99%

Differential Protein S-Thiolation of Glyceraldehyde-3-Phosphate Dehydrogenase Isoenzymes Influences Sensitivity to Oxidative Stress

Grant¹,

Quinn

Dawes³

1999

Molecular and Cellular Biology

145

126

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The irreversible oxidation of cysteine residues can be prevented by protein S-thiolation, in which protein -SH groups form mixed disulfides with low-molecular-weight thiols such as glutathione. We report here the identification of glyceraldehyde-3-phosphate dehydrogenase as the major target of protein S-thiolation following treatment with hydrogen peroxide in the yeast Saccharomyces cerevisiae. Our studies reveal that this process is tightly regulated, since, surprisingly, despite a high degree of sequence homology (98% similarity and 96% identity), the Tdh3 but not the Tdh2 isoenzyme was S-thiolated. The glyceraldehyde-3-phosphate dehydrogenase enzyme activity of both the Tdh2 and Tdh3 isoenzymes was decreased following exposure to H 2 O 2 , but only Tdh3 activity was restored within a 2-h recovery period. This indicates that the inhibition of the S-thiolated Tdh3 polypeptide was readily reversible. Moreover, mutants lacking TDH3 were sensitive to a challenge with a lethal dose of H 2 O 2 , indicating that the S-thiolated Tdh3 polypeptide is required for survival during conditions of oxidative stress. In contrast, a requirement for the nonthiolated Tdh2 polypeptide was found during exposure to continuous low levels of oxidants, conditions where the Tdh3 polypeptide would be S-thiolated and hence inactivated. We propose a model in which both enzymes are required during conditions of oxidative stress but play complementary roles depending on their ability to undergo S-thiolation.Sulfhydryl groups (-SH) play a remarkably broad range of roles in the cell, since the redox status of cysteine residues is involved in both the structure and function of numerous enzymes, receptors, and transcription factors. However, cysteine residues are among the most easily oxidized residues in proteins, and the oxidation of -SH groups is one of the earliest observable events during reactive oxygen species (ROS)-mediated damage (6, 12). The oxidation of -SH groups can result in the formation of protein disulfides (S-S) through two protein cysteines or mixed disulfides between protein-SH groups and a number of low-molecular-weight thiols (protein S-thiolation). While the role of disulfide bond formation in protein folding has been well characterized (see, for example, reference 23), its role during conditions of oxidative stress is unclear. In contrast, protein S-thiolation has been proposed to serve an antioxidant function by preventing the irreversible oxidation of cysteine residues to higher oxidation states (e.g., sulfenic acid) following exposure to ROS (10, 47).Modification of proteins by S-thiolation does not require an enzymatic activity and has been proposed to occur either by the reaction of partially oxidized protein sulfhydryls (thiyl radical or sulfenic acid intermediates) with thiols such as cysteine or glutathione (GSH) or by thiol-disulfide exchange reactions with the oxidized disulfide form of GSH (GSSG) (47). To provide a defense against conditions of oxidative stress, this modification must be reversible. Dethiolatio...

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S-thiolation of cytoplasmic cardiac creatine kinase in heart cells treated with diamide

Cited by 53 publications

References 24 publications

Detection, Quantitation, Purification, and Identification of Cardiac Proteins S-Thiolated during Ischemia and Reperfusion

Detection, Quantitation, Purification, and Identification of Cardiac Proteins S-Thiolated during Ischemia and Reperfusion

Role of Oxidative Modifications in Atherosclerosis

Differential Protein S-Thiolation of Glyceraldehyde-3-Phosphate Dehydrogenase Isoenzymes Influences Sensitivity to Oxidative Stress

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