S-Nitrosation of Glutathione Transferase P1-1 Is Controlled by the Conformation of a Dynamic Active Site Helix

Balchin, David; Wallace, Louise; Dirr, Heini W.

doi:10.1074/jbc.m113.462671

Cited by 11 publications

(23 citation statements)

References 48 publications

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“…Conserved cysteine nitrosation is known to negatively regulate many metabolic enzymes including cytosolic glyceraldehyde-3-phosphate dehydrogenase 36 , glutathione transferase P1 48 , and protein disulfide isomerase 49 . In these cases, modified cysteines are either active site residues or are located in catalytic domains.…”

Section: Discussionmentioning

confidence: 99%

S-Nitrosation of Conserved Cysteines Modulates Activity and Stability of S-Nitrosoglutathione Reductase (GSNOR)

et al. 2016

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The free radical nitric oxide (NO•) regulates diverse physiological processes from vasodilation in humans to gas exchange in plants. S-nitrosoglutathione (GSNO) is considered a principal nitroso reservoir due to its chemical stability. GSNO accumulation is attenuated by GSNO reductase (GSNOR), a cysteine-rich cytosolic enzyme. Regulation of protein nitrosation is not well understood since NO•-dependent events proceed without discernible changes in GSNOR expression. Because GSNORs contain evolutionarily-conserved cysteines that could serve as nitrosation sites, we examined the effects of treating plant (Arabidopsis thaliana), mammalian (human), and yeast (Saccharomyces cerevisiae) GSNORs with nitrosating agents in vitro. Enzyme activity was sensitive to nitroso donors, while the reducing agent dithiothreitol (DTT) restored activity, suggesting catalytic impairment was due to S-nitrosation. Protein nitrosation was confirmed by mass spectrometry, by which mono-, di-, and tri-nitrosation were observed, and these signals were sensitive to DTT. GSNOR mutants in specific non-zinc coordinating cysteines were less sensitive to catalytic inhibition by nitroso donors and exhibited reduced nitrosation signals by mass spectrometry. Nitrosation also coincided with decreased tryptophan fluorescence, increased thermal aggregation propensity, and increased polydispersity—properties reflected by differential solvent accessibility of amino acids important for dimerization and the shape of the substrate and coenzyme binding pockets as assessed by hydrogen-deuterium exchange mass spectrometry. Collectively, these data suggest a mechanism for NO• signal transduction in which GSNOR nitrosation and inhibition transiently permit GSNO accumulation.

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Section: Discussionmentioning

confidence: 99%

S-Nitrosation of Conserved Cysteines Modulates Activity and Stability of S-Nitrosoglutathione Reductase (GSNOR)

et al. 2016

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“…These results confirm that the intrinsic Prx1 fluorescence can be used to follow S-nitrosation of Prx1 and suggest that treatment of Prx1C83SC173S with GSNO leads to Prx1C83SC173S nitrosation, which is reversed (denitrosated) by GSH addition. Changes in the intrinsic fluorescence of proteins upon their modification by oxidants and nitrosating agents have been extensively used to perform kinetic investigations, such as the kinetics of Prx1 oxidation by peroxides [44][45][46] and of glutathione S-transferase S-nitrosation by GSNO [47]. Therefore, we started by treating the mutant Prx1C83SC173S (5 μM) with GSNO (400 μM) and examined the changes in its intrinsic fluorescence.…”

Section: Kinetics and Products Of The Reaction Of Prx1 With Gsnomentioning

confidence: 99%

The Peroxidatic Thiol of Peroxiredoxin 1 is Nitrosated by Nitrosoglutathione but Coordinates to the Dinitrosyl Iron Complex of Glutathione

et al. 2020

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Protein S-nitrosation is an important consequence of NO●·metabolism with implications in physiology and pathology. The mechanisms responsible for S-nitrosation in vivo remain debatable and kinetic data on protein S-nitrosation by different agents are limited. 2-Cys peroxiredoxins, in particular Prx1 and Prx2, were detected as being S-nitrosated in multiple mammalian cells under a variety of conditions. Here, we investigated the kinetics of Prx1 S-nitrosation by nitrosoglutathione (GSNO), a recognized biological nitrosating agent, and by the dinitrosyl-iron complex of glutathione (DNIC-GS; [Fe(NO)2(GS)2]−), a hypothetical nitrosating agent. Kinetics studies following the intrinsic fluorescence of Prx1 and its mutants (C83SC173S and C52S) were complemented by product analysis; all experiments were performed at pH 7.4 and 25 ℃. The results show GSNO-mediated nitrosation of Prx1 peroxidatic residue ( k + N O C y s 52 = 15.4 ± 0.4 M−1. s−1) and of Prx1 Cys83 residue ( k + N O C y s 83 = 1.7 ± 0.4 M−1. s−1). The reaction of nitrosated Prx1 with GSH was also monitored and provided a second-order rate constant for Prx1Cys52NO denitrosation of k − N O C y s 52 = 14.4 ± 0.3 M−1. s−1. In contrast, the reaction of DNIC-GS with Prx1 did not nitrosate the enzyme but formed DNIC-Prx1 complexes. The peroxidatic Prx1 Cys was identified as the residue that more rapidly replaces the GS ligand from DNIC-GS ( k D N I C C y s 52 = 7.0 ± 0.4 M−1. s−1) to produce DNIC-Prx1 ([Fe(NO)2(GS)(Cys52-Prx1)]−). Altogether, the data showed that in addition to S-nitrosation, the Prx1 peroxidatic residue can replace the GS ligand from DNIC-GS, forming stable DNIC-Prx1, and both modifications disrupt important redox switches.

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“…Among of them, Tyr8 functions as a critical player for stabilization of GSH [32,33]. On the other hand, the H-site residues of C-terminal domain are relatively flexible compared with that of others that is associated with capability of adaptation to a wider spectrum of substrates [34,35].…”

Section: Discussionmentioning

confidence: 99%

“…In general, the Gsite is highly specific for GSH binding and contains eleven highly conserved amino acids: Tyr7, Arg13, Trp38, Lys46, Gln53, Leu54, Pro55, Gln66, Ser67, Glu98 and Asp99 [32,33]. The H-site is the binding site for electrophilic substrates and has eight conserved residues: Tyr7, Phe8, Val10, Arg13, Val104, Tyr108, Asn204, and Gly205 [34]. Notably, an obvious diversity of conserved residues derived from G-site and H-site is detected between AwGST1 and AwGST2.…”

Section: Discussionmentioning

confidence: 99%

“…Positively selected residues of the GSTs are located close to essential catalytic residues, it represents a mechanism for substrate diversity of the GSTs family [36,37]. The active sites of GSTs require specific amino acid residues and accurate orientation in protein structures, thereby resulting in greater functional and structural constraints [18,32,34,37,38]. In sigmaclass GSTs, Tyr residue responds for stabilization of GSH, this stabilization residue play an important role in formation of ball-andsocket structure of cytosolic GSTs [32,33].…”

Section: Discussionmentioning

confidence: 99%

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Isolation and characterization of two glutathione S-transferases from freshwater bivalve Anodonta woodiana : Chronic effects of pentachlorophenol on gene expression profiles

Liu

Shang

et al. 2017

Fish & Shellfish Immunology

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Glutathione S-transferases (GST) play a prominent role in protecting cells against oxidative stress. Our previous study showed that the reactive oxygen species (ROS) generated from pentachlorophenol (PCP) could cause an acute impact on freshwater bivalve Anodonta Woodiana, but its chronic toxicity remain unclear. In order to investigate the chronic effect of PCP, clams A. Woodiana were randomly grouped into PCP treated group in which animals were administrated with 13.9 μg/L concentrations of PCP, and control group those with similar volume dimethyl sulfoxide. In addition, two complete GST sequences were isolated from A. Woodianaa and respectively named AwGST1 and AwGST2. The full-length cDNA of AwGST1 was consisted of a 5' untranslated region (UTR) of 132 bp, a 3' UTR of 80 bp and an open reading frame (ORF) of 609 bp encoding a polypeptide of 203 amino acids. The full-length cDNA of AwGST2 contained a 5' UTR of 57 bp, a 3' UTR of 291 bp and an ORF of 678 bp encoding a polypeptide of 226 amino acids. The constitutive expression levels of AwGST1 and AwGST2 were examined in different tissues including foot, mantle, adductor muscle, heart, hepatopancreas, hemocytes and gill. Administration of PCP could result in a significant increase of AwGST1 and AwGST2 expression in the hepatopancreas, gill and hemocytes. In the hepatopancreas, AwGST1 mRNA levels of PCP treated group increased more than 28.73% at day 1, then 70.37% (P < 0.05) at day 3, reach to 6.64 times (P < 0.01) at day 15 in contrasted with that of control group. AwGST2 increased more 18.18%, 82.88% (P < 0.05) and 2.43 times (P < 0.01) at day 1, 3 and 15, respectively. In the gill, AwGST1 expression showed a significant up-regulation in the PCP treated group during experiment observed compared with that of control group, mRNA level of AwGST2 increased more than 1.44 times (P < 0.05). In addition, expressions of AwGST1 and AwGST2 were significantly induced after PCP treatment in the hemocytes. These results indicated that up-regulations of AwGST1 and AwGST2 expression in bivalve A. woodiana are contribute to against oxidative stress derived from PCP treatment during experiment observed.

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S-Nitrosation of Glutathione Transferase P1-1 Is Controlled by the Conformation of a Dynamic Active Site Helix

Cited by 11 publications

References 48 publications

S-Nitrosation of Conserved Cysteines Modulates Activity and Stability of S-Nitrosoglutathione Reductase (GSNOR)

S-Nitrosation of Conserved Cysteines Modulates Activity and Stability of S-Nitrosoglutathione Reductase (GSNOR)

The Peroxidatic Thiol of Peroxiredoxin 1 is Nitrosated by Nitrosoglutathione but Coordinates to the Dinitrosyl Iron Complex of Glutathione

Isolation and characterization of two glutathione S-transferases from freshwater bivalve Anodonta woodiana : Chronic effects of pentachlorophenol on gene expression profiles

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