<i>S</i>-Guanylation Proteomics for Redox-Based Mitochondrial Signaling

Rahaman, Md. Mizanur; Sawa, Tomohiro; Ahtesham, Ahmed Khandaker; Khan, Shahzada; Inoue, Hirofumi; Irie, Atsuhi; Fujii, Shigemoto; Akaike, Takaaki

doi:10.1089/ars.2012.4606

Cited by 28 publications

(43 citation statements)

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“…31) S-Guanylation at Cys434 of Keap1 may facilitate dissociation of the protein from Nrf2, which would lead to activation of Nrf2-dependent gene expression. 31) Other targets of protein S-guanylation identified so far include mitochondrial heat shock protein-60, 32) small GTPase H-Ras, 9) SNAP25 in the SNARE complex, 33) and tau protein. 34) We found that 8-nitro-cGMP causes protein S-guanylation of H-Ras in the mouse heart during myocardial infarction and in rat cardiac fibroblasts in culture.…”

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confidence: 99%

Synthesis and Characterization of 8-Nitroguanosine 3′,5′-Cyclic Monophosphorothioate Rp-Isomer as a Potent Inhibitor of Protein Kinase G1α

Ahmed

Zhang

Ono

et al. 2017

Biological & Pharmaceutical Bulletin

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Guanosine 3′,5′-cyclic monophosphate (cGMP)-dependent protein kinases (PKG) are kinases regulating diverse physiological functions including vascular smooth muscle relaxation, neuronal synaptic plasticity, and platelet activities. Certain PKG inhibitors, such as Rp-diastereomers of derivatives of guanosine 3′,5′-cyclic monophosphorothioate (Rp-cGMPS), have been designed and used to study PKG-regulated cell signaling. 8-Nitroguanosine 3′,5′-cyclic monophosphate (8-nitro-cGMP) is an endogenous cGMP derivative formed as a result of excess production of reactive oxygen species and nitric oxide. 8-Nitro-cGMP causes persistent activation of PKG1α through covalent attachment of cGMP moieties to cysteine residues of the enzyme (i.e., the process called protein S-guanylation). In this study, we synthesized a nitrated analogue of Rp-cGMPS, 8-nitroguanosine 3′,5′-cyclic monophosphorothioate Rp-isomer (Rp-8-nitro-cGMPS), and investigated its effects on PKG1α activity. We synthesized Rp-8-nitro-cGMPS by reacting Rp-8-bromoguanosine 3′,5′-cyclic monophosphorothioate (Rp-8-bromo-cGMPS) with sodium nitrite. Rp-8-Nitro-cGMPS reacted with the thiol compounds cysteine and glutathione to form Rp-8-thioalkoxy-cGMPS adducts to a similar extent as did 8-nitro-cGMP. As an important finding, a protein S-guanylation-like modification was clearly observed, by using Western blotting, in the reaction between recombinant PKG1α and Rp-8-nitro-cGMPS. Rp-8-Nitro-cGMPS inhibited PKG1α activity with an inhibitory constant of 22 µM in a competitive manner. An organ bath assay with mouse aorta demonstrated that Rp-8-nitro-cGMPS inhibited vascular relaxation induced by acetylcholine or 8-bromo-cGMP more than Rp-8-bromo-cGMPS did. These findings suggest that Rp-8-nitro-cGMPS inhibits PKG through induction of an S-guanylation-like modification by attaching the Rp-cGMPS moiety to the enzyme. Additional study is warranted to explore the potential application of Rp-8-nitro-cGMPS to biochemical and therapeutic research involving PKG1α activation.

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Section: )mentioning

confidence: 99%

Synthesis and Characterization of 8-Nitroguanosine 3′,5′-Cyclic Monophosphorothioate Rp-Isomer as a Potent Inhibitor of Protein Kinase G1α

Ahmed

Zhang

Ono

et al. 2017

Biological & Pharmaceutical Bulletin

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“…Anti-8-thioalkoxy-cGMP antibodies that recognize the Sguanylation moiety have been developed to detect protein S-guanylation by means of immunocytochemistry/immunohistochemistry [8,17], Western blotting [3,[7][8][9]17], immunoaffinity purification coupled with mass spectrometry [3,7,9], and enzyme-linked immunosorbent assay (ELISA) [40]. S-Guanylation structure is stable against treatment with conventional denaturing agents such as urea and dithiothreitol.…”

Section: Analysis Of Protein S-guanylationmentioning

confidence: 99%

“…Thus, S-guanylated proteins can be subjected for tryptic and other related enzyme digestions after reduction-alkylation processes [41]. Subsequently, peptides bearing S-guanylated moieties can be analyzed by LC-MS/MS to identify the sites of modifications without any types of derivatization like biotin labeling [9]. S-Guanylation has been registered as ''cGMP (C)'' in the Matrix Science, thus, one can search S-guanylation in the peptides with Mascot MS/MS ion searches of the National Center for Biotechnology Information-nonredundant (NCBI nr) database [9].…”

Section: Analysis Of Protein S-guanylationmentioning

confidence: 99%

“…This S-guanylation proteomics comprised two approaches: (i) direct protein digestion, followed by immunoaffinity capture of S-guanylated peptides that were subjected to LC-MS/MS, and (ii) 2D polyacrylamide gel electrophoresis separation of S-guanylated proteins that were visualized by Western blotting, and corresponding S-guanylation spots were extracted and subjected to in-gel digestion, followed by LC-MS/MS [9].…”

Section: Analysis Of Protein S-guanylationmentioning

confidence: 99%

“…Accumulating evidence has suggested the occurrence of S-guanylation on proteins that critically involved in the regulation of cellular responses to oxidative, metabolic or environmental stress [4][5][6]. Examples in- clude protein S-guanylation of Kelch-like ECH-associated protein 1 (Keap1) [3,7], H-Ras [8] and mitochondrial heat shock proteins [9]. 8-Nitro-cGMP signaling via protein S-guanylation may thus have evolved to convey adaptive cellular responses to stress in general [5,6].…”

Section: Introductionmentioning

confidence: 99%

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Formation, signaling functions, and metabolisms of nitrated cyclic nucleotide

et al. 2013

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8-Nitro-cGMP: A Novel Protein-Reactive cNMP and Its Emerging Roles in Autophagy

Arimoto

Takahashi

2017

Handbook of Experimental Pharmacology

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Nitric oxide (NO) raises the intracellular 3',5'-cyclic guanosine monophosphate (cGMP) level through the activation of soluble guanylate cyclase and, in the presence of reactive oxygen species (ROS), reacts with biomolecules to produce nitrated cGMP derivatives. 8-Nitro-cGMP was the first endogenous cGMP derivative discovered in mammalian cells (2007) and was later found in plant cells. Among the six nitrogen atoms in this molecule, the one in the nitro group (NO) comes from NO. This chapter asserts that this newly found cGMP is undoubtedly one of the major physiological cNMPs. Multiple studies suggest that its intracellular abundance might exceed that of unmodified cGMP. The characteristic chemical feature of 8-nitro-cGMP is its ability to modify proteinous cysteine residues via a stable sulfide bond. In this posttranslational modification, the nitro group is detached from the guanine base. This modification, termed "protein S-guanylation," is known to regulate the physiological functions of several important proteins. Furthermore, 8-nitro-cGMP participates in the regulation of autophagy. For example, in antibacterial autophagy (xenophagy), S-guanylation accumulates around invading bacterial cells and functions as a "tag" for subsequent clearance of the organism via ubiquitin modifications. This finding suggests the existence of a system for recognizing the cGMP structure on proteins. Autophagy induction by 8-nitro-cGMP is mechanistically distinct from the well-described starvation-induced autophagy and is independent of the action of mTOR, the master regulator of canonical autophagy.

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S-Guanylation Proteomics for Redox-Based Mitochondrial Signaling

Abstract: Our S-guanylation proteomic method determined that mitochondrial HSPs may be novel targets for redox modification via protein S-guanylation that participates in mPTP regulation and mitochondrial redox signaling.

Cited by 28 publications

References 50 publications

Synthesis and Characterization of 8-Nitroguanosine 3′,5′-Cyclic Monophosphorothioate Rp-Isomer as a Potent Inhibitor of Protein Kinase G1α

Synthesis and Characterization of 8-Nitroguanosine 3′,5′-Cyclic Monophosphorothioate Rp-Isomer as a Potent Inhibitor of Protein Kinase G1α

Formation, signaling functions, and metabolisms of nitrated cyclic nucleotide

8-Nitro-cGMP: A Novel Protein-Reactive cNMP and Its Emerging Roles in Autophagy

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