共查询到20条相似文献,搜索用时 31 毫秒
1.
Beatriz Garzón Javier Gayarre Severine Gharbi Beatriz Díez-Dacal Francisco J. Sánchez-Gómez John F. Timms Dolores Pérez-Sala 《Chemico-biological interactions》2010,183(1):212-221
The cyclopentenone prostaglandin (cyPG) PGA1 displays potent anti-proliferative and anti-inflammatory effects. Therefore, PGA1 derivatives are being studied as therapeutic agents. One major mechanism for cyPG action is the modification of protein cysteine residues, the nature of the modified proteins being highly dependent on the structure of the cyPG. Biotinylated cyPGs may aid in the proteomic identification of cyPG targets of therapeutic interest. However, for the identified targets to be relevant it is critical to assess whether biotinylated cyPGs retain the desired biological activity. Here we have explored the anti-inflammatory, anti-proliferative and cell stress-inducing effects of a biotinylated analog of PGA1 (PGA1-biotinamide, PGA1-B), to establish its validity to identify cyPG–protein interactions of potential therapeutic interest. PGA1 and PGA1-B displayed similar effects on cell viability, Hsp70 and heme oxygenase-1 induction and pro-inflammatory gene inhibition. Remarkably, PGA1-B did not activate PPAR. Therefore, this biotinylated analog can be useful to identify PPAR-independent effects of cyPGs. Protein modification and subcellular distribution of PGA1-B targets were cell-type-dependent. Through proteomic and biochemical approaches we have identified a novel set of PGA1-B targets including proteins involved in stress response, protein synthesis, cytoskeletal regulation and carbohydrate metabolism. Moreover, the modification of several of the targets identified could be reproduced in vitro. These results unveil novel interactions of PGA1 that will contribute to delineate the mechanisms for the anti-proliferative and metabolic actions of this cyPG. 相似文献
2.
Pérez-Sala D 《Chemico-biological interactions》2011,192(1-2):96-100
Electrophilic eicosanoids are reactive mediators that arise by non-enzymatic transformations of arachidonic acid or of its products and display varied biological actions. Various electrophilic eicosanoids have shown anti-proliferative and anti-inflammatory effects, which have elicited a great interest in their study as potential therapeutic agents. A key feature of these compounds is their ability to covalently modify proteins, thus altering their structure and function. The modification of several components of the NF-κB pathway contributes to the anti-inflammatory effects of electrophilic eicosanoids, whereas addition to redox-sensitive proteins plays a key role in the antioxidant response. However, electrophilic eicosanoids may also have a dark side, and accumulating evidence points towards their involvement in neurotoxicity and/or neurodegeneration. The ability of some electrophilic eicosanoids to induce protein oligomerization or aggregation through various mechanisms may contribute to these effects. Biochemical and proteomic studies have led to the identification of numerous protein targets for modification by electrophilic eicosanoids, the number of which continues to expand, revealing novel potential functions for these compounds and providing a basis for their pleiotropic effects. The ample number of targets identified, together with the non-enzymatic nature of the modification argue against the potential specificity or regulation of electrophilic eicosanoid action. However, protein modification displays selectivity depending on structural features of the proteins and of the electrophilic compounds as well as on context factors such as cell type and GSH availability. Understanding the factors which control the extent and selectivity of protein modification by electrophilic eicosanoids is therefore essential to elucidate their pathophysiological roles and therapeutic potential in specific settings. 相似文献
3.
Chouchani ET James AM Fearnley IM Lilley KS Murphy MP 《Current opinion in chemical biology》2011,15(1):120-128
Protein cysteine residues are central to redox signaling and to protection against oxidative damage through their interactions with reactive oxygen and nitrogen species, and electrophiles. Although there is considerable evidence for a functional role for cysteine modifications, the identity and physiological significance of most protein thiol alterations are unknown. One way to identify candidate proteins involved in these processes is to utilize the proteomic methodologies that have been developed in recent years for the identification of proteins that undergo cysteine modification in response to redox signals or oxidative damage. These tools have proven effective in uncovering novel protein targets of redox modification and are important first steps that allow for a better understanding of how reactive molecules may contribute to signaling and damage. Here, we discuss a number of these approaches and their application to the identification of a variety of cysteine-centered redox modifications. 相似文献
4.
The process of lipid peroxidation is widespread in biology and is mediated through both enzymatic and non-enzymatic pathways. A significant proportion of the oxidized lipid products are electrophilic in nature, the RLS (reactive lipid species), and react with cellular nucleophiles such as the amino acids cysteine, lysine and histidine. Cell signalling by electrophiles appears to be limited to the modification of cysteine residues in proteins, whereas non-specific toxic effects involve modification of other nucleophiles. RLS have been found to participate in several physiological pathways including resolution of inflammation, cell death and induction of cellular antioxidants through the modification of specific signalling proteins. The covalent modification of proteins endows some unique features to this signalling mechanism which we have termed the 'covalent advantage'. For example, covalent modification of signalling proteins allows for the accumulation of a signal over time. The activation of cell signalling pathways by electrophiles is hierarchical and depends on a complex interaction of factors such as the intrinsic chemical reactivity of the electrophile, the intracellular domain to which it is exposed and steric factors. This introduces the concept of electrophilic signalling domains in which the production of the lipid electrophile is in close proximity to the thiol-containing signalling protein. In addition, we propose that the role of glutathione and associated enzymes is to insulate the signalling domain from uncontrolled electrophilic stress. The persistence of the signal is in turn regulated by the proteasomal pathway which may itself be subject to redox regulation by RLS. Cell death mediated by RLS is associated with bioenergetic dysfunction, and the damaged proteins are probably removed by the lysosome-autophagy pathway. 相似文献
5.
Cyclopentenone prostaglandins (cyPG) with antiinflammatory and antiproliferative properties have been envisaged as leads for the development of therapeutic agents. Because cyPG effects are mediated in part by the formation of covalent adducts with critical signaling proteins, it is important to assess the specificity of this interaction. By using biotinylated derivatives of 15-deoxy-Delta(12,14)-PGJ(2) (15d-PGJ(2)-B) and PGA(1) (PGA(1)-B) we herein provide novel evidence for the differential selectivity of protein modification by distinct cyPG. The marked quantitative and qualitative differences in the binding of 15d-PGJ(2)-B and PGA(1)-B to cellular proteins were related to a differential reactivity in the presence of glutathione (GSH), both in vitro and in intact cells. Therefore GSH levels may influence not only the intensity but also the specificity of cyPG action. 相似文献
6.
Sánchez-Gómez FJ Gayarre J Avellano MI Pérez-Sala D 《Archives of biochemistry and biophysics》2007,457(2):150-159
Glutathione-S-transferases (GST) catalyze the conjugation of electrophilic compounds to glutathione, thus playing a key role in cell survival and tumor chemoresistance. Cyclopentenone prostaglandins (cyPG) are electrophilic eicosanoids that display potent antiproliferative properties, through multiple mechanisms not completely elucidated. Here we show that the cyPG 15-deoxy-Delta(12,14)-PGJ2 (15d-PGJ2) binds to GSTP1-1 covalently, as demonstrated by mass spectrometry and by the use of biotinylated 15d-PGJ2. Moreover, cyPG inactivate GSTP1-1 irreversibly. The presence of the cyclopentenone moiety is important for these effects. Covalent interactions also occur in cells, in which 15d-PGJ2 binds to endogenous GSTP1-1, irreversibly reduces GST free-thiol content and inhibits GST activity. Protein delivery of GSTP1-1 improves cell survival upon serum deprivation whereas 15d-PGJ2-treated GSTP1-1 displays a reduced protective effect. These results show the first evidence for the formation of stable adducts between cyPG and GSTP1-1 and may offer new perspectives for the development of irreversible GST inhibitors as anticancer agents. 相似文献
7.
Oeste CL Díez-Dacal B Bray F García de Lacoba M de la Torre BG Andreu D Ruiz-Sánchez AJ Pérez-Inestrosa E García-Domínguez CA Rojas JM Pérez-Sala D 《PloS one》2011,6(1):e15866
Ras proteins are crucial players in differentiation and oncogenesis and constitute important drug targets. The localization and activity of Ras proteins are highly dependent on posttranslational modifications at their C-termini. In addition to an isoprenylated cysteine, H-Ras, but not other Ras proteins, possesses two cysteine residues (C181 and C184) in the C-terminal hypervariable domain that act as palmitoylation sites in cells. Cyclopentenone prostaglandins (cyPG) are reactive lipidic mediators that covalently bind to H-Ras and activate H-Ras dependent pathways. Dienone cyPG, such as 15-deoxy-Δ(12,14)-PGJ(2) (15d-PGJ(2)) and Δ(12)-PGJ(2) selectively bind to the H-Ras hypervariable domain. Here we show that these cyPG bind simultaneously C181 and C184 of H-Ras, thus potentially altering the conformational tendencies of the hypervariable domain. Based on these results, we have explored the capacity of several bifunctional cysteine reactive small molecules to bind to the hypervariable domain of H-Ras proteins. Interestingly, phenylarsine oxide (PAO), a widely used tyrosine phosphatase inhibitor, and dibromobimane, a cross-linking agent used for cysteine mapping, effectively bind H-Ras hypervariable domain. The interaction of PAO with H-Ras takes place in vitro and in cells and blocks modification of H-Ras by 15d-PGJ(2). Moreover, PAO treatment selectively alters H-Ras membrane partition and the pattern of H-Ras activation in cells, from the plasma membrane to endomembranes. These results identify H-Ras as a novel target for PAO. More importantly, these observations reveal that small molecules or reactive intermediates interacting with spatially vicinal cysteines induce intramolecular cross-linking of H-Ras C-terminus potentially contributing to the modulation of Ras-dependent pathways. 相似文献
8.
Lia S. Nakao Robert A. Everley Stefano M. Marino Sze M. Lo Luiz E. de Souza Steven P. Gygi Vadim N. Gladyshev 《The Journal of biological chemistry》2015,290(9):5685-5695
Thioredoxin (Trx)-fold proteins are protagonists of numerous cellular pathways that are subject to thiol-based redox control. The best characterized regulator of thiols in proteins is Trx1 itself, which together with thioredoxin reductase 1 (TR1) and peroxiredoxins (Prxs) comprises a key redox regulatory system in mammalian cells. However, there are numerous other Trx-like proteins, whose functions and redox interactors are unknown. It is also unclear if the principles of Trx1-based redox control apply to these proteins. Here, we employed a proteomic strategy to four Trx-like proteins containing CXXC motifs, namely Trx1, Rdx12, Trx-like protein 1 (Txnl1) and nucleoredoxin 1 (Nrx1), whose cellular targets were trapped in vivo using mutant Trx-like proteins, under conditions of low endogenous expression of these proteins. Prxs were detected as key redox targets of Trx1, but this approach also supported the detection of TR1, which is the Trx1 reductant, as well as mitochondrial intermembrane proteins AIF and Mia40. In addition, glutathione peroxidase 4 was found to be a Rdx12 redox target. In contrast, no redox targets of Txnl1 and Nrx1 could be detected, suggesting that their CXXC motifs do not engage in mixed disulfides with cellular proteins. For some Trx-like proteins, the method allowed distinguishing redox and non-redox interactions. Parallel, comparative analyses of multiple thiol oxidoreductases revealed differences in the functions of their CXXC motifs, providing important insights into thiol-based redox control of cellular processes. 相似文献
9.
The redoxome: Proteomic analysis of cellular redox networks 总被引:1,自引:0,他引:1
Redox-regulated proteins play fundamentally important roles not only during the defense of organisms against oxidative stress conditions but also as targets of cellular signaling events. This realization has spurred the development of proteomic techniques geared towards characterizing the redoxome; proteins with highly reactive cysteine residues, whose thiol oxidation state controls the function of the proteins, and by extension, the pathways they are part of. We will here summarize the most recent advances made in the field of redox proteomic analysis, aimed to elucidate the cellular redox networks that appear to control prokaryotic and eukaryotic organisms. 相似文献
10.
Michelet L Zaffagnini M Vanacker H Le Maréchal P Marchand C Schroda M Lemaire SD Decottignies P 《The Journal of biological chemistry》2008,283(31):21571-21578
Glutathionylation is the major form of S-thiolation in cells. This reversible redox post-translational modification consists of the formation of a mixed disulfide between a free thiol on a protein and a molecule of glutathione. This recently described modification, which is considered to occur under oxidative stress, can protect cysteine residues from irreversible oxidation, and alter positively or negatively the activity of diverse proteins. This modification and its targets have been mainly studied in non-photosynthetic organisms so far. We report here the first proteomic approach performed in vivo on photosynthetically competent cells, using the eukaryotic unicellular green alga Chlamydomonas reinhardtii with radiolabeled [(35)S]cysteine to label the glutathione pool and diamide as oxidant. This method allowed the identification of 25 targets, mainly chloroplastic, involved in various metabolic processes. Several targets are related to photosynthesis, such as the Calvin cycle enzymes phosphoglycerate kinase and ribose-5-phosphate isomerase. A number of targets, such as chaperones and peroxiredoxins, are related to stress responses. The glutathionylation of HSP70B, chloroplastic 2-Cys peroxiredoxin and isocitrate lyase was confirmed in vitro on purified proteins and the targeted residues were identified. 相似文献
11.
Tsaytler PA C O'Flaherty M Sakharov DV Krijgsveld J Egmond MR 《Journal of proteome research》2008,7(9):3868-3878
Oxidative stress induced in tumor cells undergoing photodynamic treatment (PDT) leads to extensive modification of many proteins in these cells. Protein oxidation mainly gives rise to formation of carbonyls and oxidized thiols. The immediate targets of PDT-induced protein oxidation in A431 tumor cells have been identified using a proteomic approach involving selective biotinylation, affinity purification and mass spectrometric identification of modified proteins. In all, 314 proteins were shown to undergo PDT-mediated oxidative modifications. While abundant structural proteins and chaperones represented a significant fraction of the carbonylated proteins, labeling of proteins containing oxidized thiols allowed identification of many proteins at low abundance and those involved in signaling and redox homeostasis. On the basis of the identification of these proteins, several likely mechanisms of PDT-induced triggering of apoptosis were put forward. This may not only lead to a further understanding of the complex network of cellular responses to oxidative stress, but it may also help in detailed targeting of photodynamic treatment applied to cancer. 相似文献
12.
Glutathione oxidation and protein glutathionylation are considered hallmarks of oxidative stress in cells because they reflect thiol redox status in proteins. Our aims were to analyze the redox status of thiols and to identify mixed disulfides and targets of redox signaling in pancreas in experimental acute pancreatitis as a model of acute inflammation associated with glutathione depletion. Glutathione depletion in pancreas in acute pancreatitis is not associated with any increase in oxidized glutathione levels or protein glutathionylation. Cystine and homocystine levels as well as protein cysteinylation and γ-glutamyl cysteinylation markedly rose in pancreas after induction of pancreatitis. Protein cysteinylation was undetectable in pancreas under basal conditions. Targets of disulfide stress were identified by Western blotting, diagonal electrophoresis, and proteomic methods. Cysteinylated albumin was detected. Redox-sensitive PP2A and tyrosine protein phosphatase activities diminished in pancreatitis and this loss was abrogated by N-acetylcysteine. According to our findings, disulfide stress may be considered a specific type of oxidative stress in acute inflammation associated with protein cysteinylation and γ-glutamylcysteinylation and oxidation of the pair cysteine/cystine, but without glutathione oxidation or changes in protein glutathionylation. Two types of targets of disulfide stress were identified: redox buffers, such as ribonuclease inhibitor or albumin, and redox-signaling thiols, which include thioredoxin 1, APE1/Ref1, Keap1, tyrosine and serine/threonine phosphatases, and protein disulfide isomerase. These targets exhibit great relevance in DNA repair, cell proliferation, apoptosis, endoplasmic reticulum stress, and inflammatory response. Disulfide stress would be a specific mechanism of redox signaling independent of glutathione redox status involved in inflammation. 相似文献
13.
14.
Nitric oxide (NO), a gaseous free radical that is synthesized in organisms by nitric oxide synthases, participates in a critical
fashion in the regulation of diverse physiological functions such as vascular and neuronal signal transduction, host defense,
and cell death regulation. Two major pathways of NO signaling involve production of the second messenger guanosine 3′,5′-cyclic
monophosphate (cGMP) and posttranslational modification (PTM) of redox-sensitive cysteine thiols of proteins. We recently
clarified the physiological formation of 8-nitroguanosine 3′,5′-cyclic monophosphate (8-nitro-cGMP) as the first demonstration,
since the discovery of cGMP more than 40 years ago, of a new second messenger derived from cGMP in mammals. 8-Nitro-cGMP is
electrophilic and reacts efficiently with sulfhydryls of proteins to produce a novel PTM via cGMP adduction, a process that
we named protein S-guanylation. 8-Nitro-cGMP may regulate electrophilic signaling on the basis of its electrophilicity through induction of
S-guanylation of redox sensor proteins. Examples include S-guanylation of the redox sensor protein Kelch-like ECH-associated protein 1 (Keap1), which leads to activation of NF-E2-related
factor 2 (Nrf2)-dependent expression of antioxidant and cytoprotective genes. This S-guanylation-mediated activation of an antioxidant adaptive response may play an important role in cytoprotection during bacterial
infections and oxidative stress. Identification of new redox-sensitive proteins as targets for S-guanylation may help development of novel therapeutics for oxidative stress- and inflammation-related disorders and vascular
diseases as well as understanding of cellular protection against oxidative stress. 相似文献
15.
16.
Kamel Chibani Jérémy Couturier Benjamin Selles Jean-Pierre Jacquot Nicolas Rouhier 《Photosynthesis research》2010,104(1):75-99
The post-translational modification consisting in the formation/reduction of disulfide bonds has been the subject of intense
research in plants since the discovery in the 1970s that many chloroplastic enzymes are regulated by light through dithiol–disulfide
exchange reactions catalyzed by oxidoreductases called thioredoxins (Trxs). Further biochemical and proteomic studies have
considerably increased the number of target enzymes and processes regulated by these mechanisms in many sub-cellular compartments.
Recently, glutathionylation, a modification consisting in the reversible formation of a glutathione adduct on cysteine residues,
was proposed as an alternative redox regulation mechanism. Glutaredoxins (Grxs), proteins related to Trxs, are efficient catalysts
for deglutathionylation, the opposite reaction. Hence, the Trxs- and Grxs-dependent pathways might constitute complementary
and not only redundant regulatory processes. This article focuses on these two multigenic families and associated protein
partners in poplar and on their involvement in the regulation of some major chloroplastic processes such as stress response,
carbohydrate and heme/chlorophyll metabolism. 相似文献
17.
Giulia Gorini Tania Gamberi Tania Fiaschi Michele Mannelli 《Free radical research》2019,53(2):126-138
The imbalance between the reactive oxygen (ROS) and nitrogen (RNS) species production and their handling by the antioxidant machinery (low molecular weight antioxidant molecules and antioxidant enzymes), also known as oxidative stress, is a condition caused by physiological and pathological processes. Moreover, oxidative stress may be due to an overproduction of free radicals during physical exercise. Excess of radical species leads to the modification of molecules, such as proteins – the most susceptible to oxidative modification – lipids and DNA. With regard to the oxidation of proteins, carbonylation is an oxidative modification that has been widely described. Several studies have detected changes in the total amount of protein carbonyls following different types of physical exercise, but only few of these identified the specific amino acidic residues targets of such oxidation. In this respect, proteomic approaches allow to identify the proteins susceptible to carbonylation and in many cases, it is also possible to identify the specific protein carbonylation sites. This review focuses on the role of protein oxidation, and specifically carbonyl formation, for plasma and skeletal muscle proteins, following different types of physical exercise performed at different intensities. Furthermore, we focused on the proteomic strategies used to identify the specific protein targets of carbonylation. Overall, our analysis suggests that regular physical activity promotes a protection against protein carbonylation, due to the activation of the antioxidant defence or of the turnover of protein carbonyls. However, we can conclude that from the comprehensive bibliography analysed, there is no clearly defined specific physiological role about this post-translational modification of proteins. 相似文献
18.
Lipid peroxidation involves a cascade of reactions in which production of free radicals occurs selectively in the lipid components of cellular membranes. Polyunsaturated fatty acids easily undergo lipid peroxidation chain reactions, which, in turn, lead to the formation of highly reactive electrophilic aldehydes. Among these, the most abundant aldehydes are 4-hydroxy-2-nonenal (HNE) and malondialdehyde, while acrolein is the most reactive. Proteins are susceptible to posttranslational modifications caused by aldehydes binding covalently to specific amino acid residues, in a process called Michael adduction, and these types of protein adducts, if not efficiently removed, may be, and generally are, dangerous for cellular homeostasis. In the present review, we focused the discussion on the selective proteins that are identified, by redox proteomics, as selective targets of HNE modification during the progression and pathogenesis of Alzheimer disease (AD). By comparing results obtained at different stages of the AD, it may be possible to identify key biochemical pathways involved and ideally identify therapeutic targets to prevent, delay, or treat AD. 相似文献
19.
随着抗生素药物的开发与利用,细菌在对抗药物过程中逐渐发展出各种不同的耐药机制.近年来高通量的蛋白质组学技术已逐渐用于细菌耐药性机理研究,但主要集中在对细菌外膜蛋白的作用进行分析.本文采用2-D native/SDS PAGE方法从蛋白质复合物角度分析接近生理条件的胞浆蛋白在头孢曲松耐药性中的作用.结果发现8个耐药性相关蛋白质,通过对蛋白质功能分析,揭示了细菌通过调整能量代谢相关蛋白产生耐药性的新机制.进一步对相关菌株的次抑菌浓度和生存率分析,提示MalP和SucC等关键蛋白质可作为设计和开发新型抗菌分子的作用靶点. 相似文献
20.
Lind C Gerdes R Hamnell Y Schuppe-Koistinen I von Löwenhielm HB Holmgren A Cotgreave IA 《Archives of biochemistry and biophysics》2002,406(2):229-240
Redox modification of proteins is proposed to play a central role in regulating cellular function. However, high-throughput techniques for the analysis of the redox status of individual proteins in complex mixtures are lacking. The aim was thus to develop a suitable technique to rapidly identify proteins undergoing oxidation of critical thiols by S-glutathionylation. The method is based on the specific reduction of mixed disulfides by glutaredoxin, their reaction with N-ethylmaleimide-biotin, affinity purification of tagged proteins, and identification by proteomic analysis. The method unequivocally identified 43 mostly novel cellular protein substrates for S-glutathionylation. These include protein chaperones, cytoskeletal proteins, cell cycle regulators, and enzymes of intermediate metabolism. Comparisons of the patterns of S-glutathionylated proteins extracted from cells undergoing diamide-induced oxidative stress and during constitutive metabolism reveal both common protein substrates and substrates failing to undergo enhanced S-glutathionylation during oxidative stress. The ability to chemically tag, select, and identify S-glutathionylated proteins, particularly during constitutive metabolism, will greatly enhance efforts to establish posttranslational redox modification of cellular proteins as an important biochemical control mechanism in coordinating cellular function. 相似文献