ICAT (Isotope‐Coded Affinity Tag) Approach to Redox Proteomics: Identification and Quantification of Oxidant‐Sensitive Protein Thiols

Mahadevan, Sethuraman; McComb, Mark E.; Huang, Hongjuan; Huang, Sequin; Heibeck, Tyler H.; Costello, Catherine E.; Cohen, Richard A.

doi:10.1002/0471973122.ch10

Cited by 2 publications

(4 citation statements)

References 26 publications

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“…The presence of a reactive thiol (-SH) group in proteins and amino thiols such as GSH not only predisposes these molecules to oxidative modification but also confers potent antioxidant properties. 12 The assessment of total or reduced, oxidized, and protein bound thiols could both provide an estimate of the antioxidant potential, and could also provide insight into the redox-status. 12 T-SH status in menopause has predominantly been investigated in whole blood and plasma.…”

Section: Discussionmentioning

confidence: 99%

“…12 The assessment of total or reduced, oxidized, and protein bound thiols could both provide an estimate of the antioxidant potential, and could also provide insight into the redox-status. 12 T-SH status in menopause has predominantly been investigated in whole blood and plasma. 4,5 The total amount of thiol containing molecules or the amount of specific thiols has been reported to be lower in plasma or erythrocyte lysate of women with menopause as comparing the corresponding values in women receiving hormone replacement therapy.…”

Section: Discussionmentioning

confidence: 99%

“…Proteins containing Cys thiol groups are particularly susceptible to oxidation by ROS. 12 Glutathione (GSH) can be reversibly bound to protein thiol groups by a mechanism called Sglutathionylation and leading to the formation of Sglutathionylated proteins. 13 Malondialdehyde (MDA) is a physiological ketoaldehyde produced by peroxidative decomposition of unsaturated lipids as a byproduct of arachidonic acid metabolism.…”

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confidence: 99%

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Effects of estrogens on oxidative protein damage in plasma and tissues in ovariectomised rats

Topçuoğlu¹,

Uzun²,

Balcı³

et al. 2009

CIM

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Purpose: To assess estrogen-related changes in the redox status of the brain and liver proteins as well as the systemic oxidative stress in ovarectomised (OVX) rats Methods: Twelve-week-old, sexually mature female Sprague–Dawley rats (200-250g) were randomly divided into four groups: The following treatment combinations were administrated daily to all in 0.05 ml 96% ethanol solution by gastric gavage. (1) Sham operation (2) OVX rats (3) OVX rats [0.02 mg/kg/day of 17?-estradiol (E2) and 0.01 mg/kg/day of norethisterone acetate] (4) OVX rats [E2 (0.01 mg/kg/day) and drospirenon (0.02 mg/kg/day)]. Estrogen levels were determined using routine clinical-chemistry methods. We also measured protein oxidation parameters such as protein carbonyl (PCO), total thiol (T-SH) and the other oxidative stress markers malondialdehyde (MDA) and glutathione (GSH). Results: Ovariectomy resulted in abnormal elevation of plasma and tissue oxidative stress markers and changes in redox status of the proteins in tissue dependent manner. Supplementation of various estrogens combinations partially alleviated these abnormalities and restored redox homeostasis of proteins after the ovariectomy. Among the studied protein oxidation parameters, plasma and tissue PCO levels of the OVX rats were higher than those of the control groups (P < 0.01). Hormone replacement therapies (HRT) caused a decrease in PCO and MDA in both plasma and tissue of the OVX rats (P < 0.01). HRT in OVX rats decreased plasma MDA and increased liver and brain GSH (P < 0.01). Liver MDA levels of the Drospirenon-treated rats were lower than in the norethisterone acetate group (P < 0.01). On the other hand, Drospirenon increases brain GSH s more effectively than norethisterone acetate (P < 0.01). After bilateral oopherectomy, plasma and tissue T-SH levels decreased in the OVX group compared with control (P < 0.01). Norethisterone acetate increased plasma T-SH more effectively than Drospirenon (P < 0.05) Conclusions: The study showed the extent of oxidative protein damage (OPD) in this model of estrogen deficiency. The protective effect of estrogens against tissue specific OPD suggests that estrogens play an important role within the antioxidant defense systems in plasma, liver and brain. The exact molecular mechanisms leading to these findings are not yet completely known. Meanwhile, hormone replacement therapy for the prevention of OPD in a tissue specific manner may be required.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Effects of estrogens on oxidative protein damage in plasma and tissues in ovariectomised rats

Topçuoğlu¹,

Uzun²,

Balcı³

et al. 2009

CIM

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“…There is considerable interest in being able to monitor the oxidation status of protein thiols in response to disease and toxicant exposure, and several methods have been presented in the literature. Thorough reviews of currently available methods based on both electrophoresis (35) and mass spectrometry (36) have been published recently. All have advantages and limitations.…”

Section: Discussionmentioning

confidence: 99%

Measurement of Protein Sulfhydryls in Response to Cellular Oxidative Stress Using Gel Electrophoresis and Multiplexed Fluorescent Imaging Analysis

Spiess

Morin

Jewell

et al. 2008

Chem. Res. Toxicol.

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The significance of free radicals in biology has been established by numerous investigations spanning a period of over 40 years. Whereas there are many intracellular targets for these radical species, the importance of cysteine thiol posttranslational modification has received considerable attention. The current studies present a highly sensitive method for measurement of the posttranslational modification of protein thiols. This method is based on labeling of proteins with monofunctional maleimide dyes followed by 2D gel electrophoresis to separate proteins and multiplexed fluorescent imaging analysis. The method correctly interrogates the thiol/disulfide ratio present in commercially available proteins. Exposure of pulmonary airway epithelial cells to high concentrations of menadione or t-butyl hydroperoxide resulted in the modification of cysteines in more than 141 proteins of which 60 were subsequently identified by MALDI-TOF/TOF MS. Although some proteins were modified similarly by these two oxidants, several showed detectably different maleimide ratios in response to these two agents. Proteins that were modified by one or both oxidants include those involved in transcription, protein synthesis and folding, and cell death/growth. In conclusion, these studies provide a novel procedure for measuring the redox status of cysteine thiols on individual proteins with a clearly demonstrated applicability to interactions of chemicals with pulmonary epithelial cells.

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ICAT (Isotope‐Coded Affinity Tag) Approach to Redox Proteomics: Identification and Quantification of Oxidant‐Sensitive Protein Thiols

Cited by 2 publications

References 26 publications

Effects of estrogens on oxidative protein damage in plasma and tissues in ovariectomised rats

Effects of estrogens on oxidative protein damage in plasma and tissues in ovariectomised rats

Measurement of Protein Sulfhydryls in Response to Cellular Oxidative Stress Using Gel Electrophoresis and Multiplexed Fluorescent Imaging Analysis

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