共查询到20条相似文献,搜索用时 788 毫秒
1.
Ana C. Ebrecht Matías D. Asención Diez Claudia V. Piattoni Sergio A. GuerreroAlberto A. Iglesias 《Biochimica et Biophysica Acta (BBA)/General Subjects》2015
Background
Giardia lamblia is a pathogen of humans and other vertebrates. The synthesis of glycogen and of structural oligo and polysaccharides critically determine the parasite's capacity for survival and pathogenicity. These characteristics establish that UDP-glucose is a relevant metabolite, as it is a main substrate to initiate varied carbohydrate metabolic routes.Results
Herein, we report the molecular cloning of the gene encoding UDP-glucose pyrophosphorylase from genomic DNA of G. lamblia, followed by its heterologous expression in Escherichia coli. The purified recombinant enzyme was characterized to have a monomeric structure. Glucose-1-phosphate and UTP were preferred substrates, but the enzyme also used galactose-1-phosphate and TTP. The catalytic efficiency to synthesize UDP-galactose was significant. Oxidation by physiological compounds (hydrogen peroxide and nitric oxide) inactivated the enzyme and the process was reverted after reduction by cysteine and thioredoxin. UDP-N-acetyl-glucosamine pyrophosphorylase, the other UTP-related enzyme in the parasite, neither used galactose-1-phosphate nor was affected by redox modification.Conclusions
Our results suggest that in G. lamblia the UDP-glucose pyrophosphorylase is regulated by oxido-reduction mechanism. The enzyme exhibits the ability to synthesize UDP-glucose and UDP-galactose and it plays a key role providing substrates to glycosyl transferases that produce oligo and polysaccharides.General significance
The characterization of the G. lamblia UDP-glucose pyrophosphorylase reinforces the view that in protozoa this enzyme is regulated by a redox mechanism. As well, we propose a new pathway for UDP-galactose production mediated by the promiscuous UDP-glucose pyrophosphorylase of this organism. 相似文献2.
Maria Laura Aon-Bertolino Juan Ignacio Romero Pablo Galeano Mariana Holubiec Maria Sol Badorrey Gustavo Ezequiel Saraceno Eva-Maria Hanschmann Christopher Horst Lillig Francisco Capani 《Biochimica et Biophysica Acta (BBA)/General Subjects》2011
Background
The oxidoreductases of the thioredoxin (Trx) family of proteins play a major role in the cellular response to oxidative stress. Redox imbalance is a major feature of brain damage. For instance, neuronal damage and glial reaction induced by a hypoxic–ischemic episode is highly related to glutamate excitotoxicity, oxidative stress and mitochondrial dysfunction. Most animal models of hypoxia–ischemia in the central nervous system (CNS) use rats to study the mechanisms involved in neuronal cell death, however, no comprehensive study on the localization of the redox proteins in the rat CNS was available.Methods
The aim of this work was to study the distribution of the following proteins of the thioredoxin and glutathione/glutaredoxin (Grx) systems in the rat CNS by immunohistochemistry: Trx1, Trx2, TrxR1, TrxR2, Txnip, Grx1, Grx2, Grx3, Grx5, and γ-GCS, peroxiredoxin 1 (Prx1), Prx2, Prx3, Prx4, Prx5, and Prx6. We have focused on areas most sensitive to a hypoxia–ischemic insult: Cerebellum, striatum, hippocampus, spinal cord, substantia nigra, cortex and retina.Results and conclusions
Previous studies implied that these redox proteins may be distributed in most cell types and regions of the CNS. Here, we have observed several remarkable differences in both abundance and regional distribution that point to a complex interplay and crosstalk between the proteins of this family.General significance
We think that these data might be helpful to reveal new insights into the role of thiol redox pathways in the pathogenesis of hypoxia–ischemia insults and other disorders of the CNS.This article is part of a Special Issue entitled Human and Murine Redox Protein Atlases. 相似文献3.
Carine F. Djuika Sabine Fiedler Martina Schnölzer Cecilia Sanchez Michael Lanzer Marcel Deponte 《Biochimica et Biophysica Acta (BBA)/General Subjects》2013
Background
Peroxiredoxins are important heterogeneous thiol-dependent hydroperoxidases with a variety of isoforms and enzymatic mechanisms. A special subclass of glutaredoxin/glutathione-dependent peroxiredoxins has been discovered in bacteria and eukaryotes during the last decade, but the exact enzymatic mechanisms of these enzymes remain to be unraveled.Methods
We performed a comprehensive analysis of the enzyme kinetics and redox states of one of these glutaredoxin/glutathione-dependent peroxiredoxins, the antioxidant protein from the malaria parasite Plasmodium falciparum, using steady-state kinetic measurements, site-directed mutagenesis, redox mobility shift assays, gel filtration, and mass spectrometry.Results
P. falciparum antioxidant protein requires not only glutaredoxin but also glutathione as a true substrate for the reduction of hydroperoxides. One peroxiredoxin cysteine residue and one glutaredoxin cysteine residue are sufficient for catalysis, however, additional cysteine residues of both proteins result in alternative redox states and conformations in vitro with implications for redox regulation. Our data furthermore point to a glutathione-dependent peroxiredoxin activation and a negative subunit cooperativity.Conclusions
The investigated glutaredoxin/glutathione/peroxiredoxin system provides numerous new insights into the mechanism and redox regulation of peroxiredoxins.General significance
As a member of the special subclass of glutaredoxin/glutathione-dependent peroxiredoxins, the P. falciparum antioxidant protein could become a reference protein for peroxiredoxin catalysis and regulation. 相似文献4.
Satoshi Hara Tatsuya Nojima Kohji Seio Masasuke Yoshida Toru Hisabori 《Biochimica et Biophysica Acta (BBA)/General Subjects》2013
Background
Thiol-mediated redox regulation of proteins plays a key role in many cellular processes.Methods
To understand the redox status of cysteinyl thiol groups of the desired proteins, we developed a new maleimide reagent: a maleimide-conjugated single strand DNA, DNA-maleimide (DNA-Mal).Results
DNA-Mal labelled proteins run as a distinct band on SDS-PAGE, with a discrete 9.32 kDa mobility shift per label regardless of the protein species or electrophoretic conditions.Conclusions
DNA-Mal labels free thiols like standard maleimide reagents, but possesses practical advantages in titration of the number and relative content of free thiols in a protein.General significance
The versatility of DNA molecule enhances the application of DNA-Mal in a broader range of cysteine containing proteins. 相似文献5.
Bruno Manta Marcelo Comini Andrea Medeiros Martín Hugo Madia Trujillo Rafael Radi 《Biochimica et Biophysica Acta (BBA)/General Subjects》2013
Background
Trypanosomatids are early-diverging eukaryotes devoid of the major disulfide reductases – glutathione reductase and thioredoxin reductase – that control thiol-redox homeostasis in most organisms. These protozoans have evolved a unique thiol-redox system centered on trypanothione, a bis-glutathionyl conjugate of spermidine. Notably, the trypanothione system is capable to sustain several cellular functions mediated by thiol-dependent (redox) processes.Scope of review
This review provides a summary of some historical and evolutionary aspects related to the discovery and appearance of trypanothione in trypanosomatids. It also addresses trypanothione's biosynthesis, physicochemical properties and reactivity towards biologically-relevant oxidants as well as its participation as a cofactor for metal binding. In addition, the role of the second most abundant thiol of trypanosomatids, glutathione, is revisited in light of the putative glutathione-dependent activities identified in these organisms.Major conclusions
Based on biochemical and genome data, the occurrence of a thiol-redox system that is strictly dependent on trypanothione appears to be a feature unique to the order Kinetoplastida. The properties of trypanothione, a dithiol, are the basis for its unique reactivity towards a wide diversity of oxidized and/or electrophilic moieties in proteins and low molecular weight compounds from endogenous or exogenous sources. Novel functions have emerged for trypanothione as a potential cofactor in iron metabolism.General significance
The minimalist thiol-redox system, developed by trypanosomatids, is an example of metabolic fitness driven by the remarkable physicochemical properties of a glutathione derivative. From a pharmacological point of view, such specialization is the Achilles' heel of these ancient and deadly parasites. This article is part of a Special Issue entitled Cellular functions of glutathione. 相似文献6.
7.
Rebecca Charles Tamani JayawardhanaPhilip Eaton 《Biochimica et Biophysica Acta (BBA)/General Subjects》2014
Background
The key to understanding the full significance of oxidants in health and disease is the development of tools and methods that allow the study of proteins that sense and transduce changes in cellular redox. Oxidant-reactive deprotonated thiols commonly operate as redox sensors in proteins and a variety of methods have been developed that allow us to monitor their oxidative modification.Scope of the review
This outline review specifically focuses on gel-based methods used to detect, quantify and identify protein thiol oxidative modifications. The techniques we discuss fall into one of two broad categories. Firstly, methods that allow oxidation of thiols in specific proteins or the global cellular pool to be monitored are discussed. These typically utilise thiol-labelling reagents that add a reporter moiety (e.g. affinity tag, fluorophore, chromophore), in which loss of labelling signifies oxidation. Secondly, we outline methods that allow specific thiol oxidation states of proteins (e.g. S-sulfenylation, S-nitrosylation, S-thionylation and interprotein disulfide bond formation) to be investigated.Major conclusions
A variety of different gel-based methods for identifying thiol proteins that are sensitive to oxidative modifications have been developed. These methods can aid the detection and quantification of thiol redox state, as well as identifying the sensor protein.General significance
By understanding how cellular redox is sensed and transduced to a functional effect by protein thiol redox sensors, this will help us better appreciate the role of oxidants in health and disease. This article is part of a Special Issue entitled Current methods to study reactive oxygen species - pros and cons and biophysics of membrane proteins. Guest Editor: Christine Winterbourn. 相似文献8.
Maria Lisa De Benedetto Concetta Rosa Capo Alberto Ferri Cristiana Valle Renato Polimanti Maria Teresa Carrì Luisa Rossi 《Biochimica et Biophysica Acta (BBA)/General Subjects》2014
Background
Glutaredoxin 1 (Grx1), a small protein belonging to the thioredoxin family, is involved in redox-regulation since it catalyzes the reduction of protein disulfides and that of mixed disulfides. It was reported to modulate active copper extrusion from cells, by affecting the function of the pumps ATP7A and B. These are components of the network of protein chaperones involved in the control of the homeostasis of copper, an essential, though harmful, metal. However, the effect of Grx1 on copper levels, copper chaperones and copper-elicited cell toxicity was never investigated.Methods
In order to investigate the effect of Grx1 on copper metabolism, we constitutively overexpressed Grx1 in human neuroblastoma SH-SY5Y cells (SH-Grx1 cells) and assessed a number of copper-related parameters.Results
SH-Grx1 cells show a basal intracellular copper level higher than control cells, accumulate more copper upon CuSO4 treatment, but are more resistant to copper-induced toxicity. Grx1 shows copper-binding properties and copper overload produces a decrease of Grx1 enzyme activity in SH-Grx1 cells. Finally, Grx1 overexpression decreases copper accumulation in mitochondria upon copper overload and modulates the expression of copper transporter 1 (Ctr1).Conclusion
Altogether, these data demonstrate that Grx1 is a major player in copper metabolism in neuronal cells. 相似文献9.
Dmitry S. Bilan Mikhail E. Matlashov Andrey Yu. Gorokhovatsky Carsten Schultz Grigori Enikolopov Vsevolod V. Belousov 《Biochimica et Biophysica Acta (BBA)/General Subjects》2014
Background
The ratio of NAD+/NADH is a key indicator that reflects the overall redox state of the cells. Until recently, there were no methods for real time NAD+/NADH monitoring in living cells. Genetically encoded fluorescent probes for NAD+/NADH are fundamentally new approach for studying the NAD+/NADH dynamics.Methods
We developed a genetically encoded probe for the nicotinamide adenine dinucleotide, NAD(H), redox state changes by inserting circularly permuted YFP into redox sensor T-REX from Thermus aquaticus. We characterized the sensor in vitro using spectrofluorometry and in cultured mammalian cells using confocal fluorescent microscopy.Results
The sensor, named RexYFP, reports changes in the NAD+/NADH ratio in different compartments of living cells. Using RexYFP, we were able to track changes in NAD+/NADH in cytoplasm and mitochondrial matrix of cells under a variety of conditions. The affinity of the probe enables comparison of NAD+/NADH in compartments with low (cytoplasm) and high (mitochondria) NADH concentration. We developed a method of eliminating pH-driven artifacts by normalizing the signal to the signal of the pH sensor with the same chromophore.Conclusion
RexYFP is suitable for detecting the NAD(H) redox state in different cellular compartments.General significance
RexYFP has several advantages over existing NAD+/NADH sensors such as smallest size and optimal affinity for different compartments. Our results show that normalizing the signal of the sensor to the pH changes is a good strategy for overcoming pH-induced artifacts in imaging. 相似文献10.
11.
Tatsuya Ueki Masayuki Uwagaki Sohei Yamamoto Hitoshi Michibata 《Biochimica et Biophysica Acta (BBA)/General Subjects》2014
Background
It is well-understood that ascidians accumulate high levels of vanadium, a reduced form of V(III), in an extremely acidic vacuole in their blood cells. Vanabins are small cysteine-rich proteins that have been identified only from vanadium-rich ascidians. A previous study revealed that Vanabin2 can act as a V(V)-reductase in the glutathione cascade.Methods
AsTrx1, a thioredoxin gene, was cloned from the vanadium-rich ascidian, Ascidia sydneiensis samea, by PCR. AsTrx1 and Vanabin2 were prepared as recombinant proteins, and V(V)-reduction by Vanabin2 was assessed by ESR and ion-exchange column chromatography. Site-directed mutagenesis was performed to examine the direct involvement of cysteine residues. Tissue expression of AsTrx1 was also examined by RT-PCR.Results
When reduced AsTrx1 and Vanabin2 were combined, Vanabin2 adopted an SS/SH intermediate structure while V(V) was reduced to V(IV). The loss of cysteine residues in either Vanabin2 or AsTrx1 caused a significant loss of reductase activity. Vapp and Kapp values for Vanabin2-catalyzed V(V)-reduction in the thioredoxin cascade were 0.066 mol-V(IV)/min/mol-Vanabin2 and 0.19 mM, respectively. The Kapp value was 2.7-fold lower than that observed in the glutathione cascade. The AsTrx1 gene was expressed at a very high level in blood cells, in which Vanabins 1–4 were co-expressed.Conclusions
AsTrx1 may contribute to a significant part of the redox cascade for V(V)-reduction by Vanabin2 in the cytoplasm of vanadocytes, but prevails only at low V(V) concentrations.General significance
This study is the first to report the reduction of V(V) in the thioredoxin cascade. 相似文献12.
José Rodrigo Godoy Maria Funke Waltraud Ackermann Petra Haunhorst Sabrina Oesteritz Francisco Capani Hans-Peter Elsässer Christopher Horst Lillig 《Biochimica et Biophysica Acta (BBA)/General Subjects》2011
Background
Oxidoreductases of the thioredoxin family of proteins have been thoroughly studied in numerous cellular and animal models mimicking human diseases. Despite of their well documented role in various disease conditions, no systematic information on the presence of these proteins is available.Methods
Here, we have systematically analyzed the presence of some of the major constituents of the glutaredoxin (Grx)-, peroxiredoxin (Prx)-, and thioredoxin (Trx)-systems, i.e. Grx1, Grx2, Grx3 (TXNL-2/PICOT), Grx5, nucleoredoxin (Nrx), Prx1, Prx2, Prx3, Prx4, Prx5, Prx6, Trx1, thioredoxin reductase 1 (TrxR1), Trx2, TrxR2, and γ-glutamyl cysteine synthetase (γ-GCS) in various tissues of the mouse using immunohistochemistry.Results
The identification of the Trx family proteins in the central nervous system, sensory organs, digestive system, lymphatic system, reproductive system, urinary system, respiratory system, endocrine system, skin, heart, and muscle revealed a number of significant differences between these proteins with respect to their distribution in these tissues.Conclusion
Our results imply more specific functions and interactions between the proteins of this family than previously assumed.General significance
Crucial functions of Trx family proteins have been demonstrated in various disease conditions. A detailed overview on their distribution in various tissues will be helpful to fully comprehend their potential role and the interactions of these proteins in the most thoroughly studied model for human diseases—the laboratory mouse.This article is part of a Special Issue entitled Human and Murine Redox Protein Atlases. 相似文献13.
Background
Hydrogen peroxide (H2O2) is an important signaling compound that has recently been identified as a new substrate for several members of the aquaporin superfamily in various organisms. Evidence is emerging about the physiological significance of aquaporin-facilitated H2O2 diffusion.Scope of review
This review summarizes current knowledge about aquaporin-facilitated H2O2 diffusion across cellular membranes. It focuses on physicochemical and experimental evidence demonstrating the involvement of aquaporins in the transport of this redox signaling compound and discusses the regulation and structural prerequisites of these channels to transmit this signal. It also provides perspectives about the potential importance of aquaporin-facilitated H2O2 diffusion processes and places this knowledge in the context of the current understanding of transmembrane redox signaling processes.Major conclusions
Specific aquaporin isoforms facilitate the passive diffusion of H2O2 across biological membranes and control H2O2 membrane permeability and signaling in living organisms.General significance
Redox signaling is a very important process regulating the physiology of cells and organisms in a similar way to the well-characterized hormonal and calcium signaling pathways. Efficient transmembrane diffusion of H2O2, a key molecule in the redox signaling network, requires aquaporins and makes these channels important players in this signaling process. Channel-mediated membrane transport allows the fine adjustment of H2O2 levels in the cytoplasm, intracellular organelles, the apoplast, and the extracellular space, which are essential for it to function as a signal molecule. This article is part of a Special Issue entitled Aquaporins. 相似文献14.
Nicola Giangregorio Ferdinando Palmieri Cesare Indiveri 《Biochimica et Biophysica Acta (BBA)/General Subjects》2013
Background
The mitochondrial carnitine/acylcarnitine carrier (CAC) is essential for cell metabolism since it catalyzes the transport of acylcarnitines into mitochondria allowing the β-oxidation of fatty acids. CAC functional and structural properties have been characterized. Cys residues which could form disulfides suggest the involvement of CAC in redox switches.Methods
The effect of GSH and GSSG on the [3H]-carnitine/carnitine antiport catalyzed by the CAC in proteoliposomes has been studied. The Cys residues involved in the redox switch have been identified by site-directed mutagenesis. Glutathionylated CAC has been assessed by glutathionyl-protein specific antibody.Results
GSH led to increase of transport activity of the CAC extracted from liver mitochondria. A similar effect was observed on the recombinant CAC. The presence of glutaredoxin-1 (Grx1) accelerated the GSH activation of the recombinant CAC. The effect was more evident at 37 °C. GSSG led to transport inhibition which was reversed by dithioerythritol (DTE). The effects of GSH and GSSG were studied on CAC Cys-mutants. CAC lacking C136 and C155 was insensitive to both reagents. Mutants containing these two Cys responded as the wild-type. Anti-glutathionyl antibody revealed the formation of glutathionylated CAC.Conclusions
CAC is redox-sensitive and it is regulated by the GSH/GSSG couple. C136 and C155 are responsible for the regulation which occurs through glutathionylation.General significance
CAC is sensitive to the redox state of the cell switching between oxidized and reduced forms in response to variation of GSSG and GSH concentrations. 相似文献15.
Background
The term GSSG/GSH redox potential is frequently used to explain redox regulation and other biological processes.Scope of review
The relevance of the GSSG/GSH redox potential as driving force of biological processes is critically discussed. It is recalled that the concentration ratio of GSSG and GSH reflects little else than a steady state, which overwhelmingly results from fast enzymatic processes utilizing, degrading or regenerating GSH.Major conclusions
A biological GSSG/GSH redox potential, as calculated by the Nernst equation, is a deduced electrochemical parameter based on direct measurements of GSH and GSSG that are often complicated by poorly substantiated assumptions. It is considered irrelevant to the steering of any biological process. GSH-utilizing enzymes depend on the concentration of GSH, not on [GSH]2, as is predicted by the Nernst equation, and are typically not affected by GSSG. Regulatory processes involving oxidants and GSH are considered to make use of mechanistic principles known for thiol peroxidases which catalyze the oxidation of hydroperoxides by GSH by means of an enzyme substitution mechanism involving only bimolecular reaction steps.General significance
The negligibly small rate constants of related spontaneous reactions as compared with enzyme-catalyzed ones underscore the superiority of kinetic parameters over electrochemical or thermodynamic ones for an in-depth understanding of GSH-dependent biological phenomena. At best, the GSSG/GSH potential might be useful as an analytical tool to disclose disturbances in redox metabolism. This article is part of a Special Issue entitled Cellular Functions of Glutathione. 相似文献16.
Stephanie B. Wall M. Ryan SmithKarina Ricart Fen ZhouPraveen K. Vayalil Joo-Yeun OhAimee Landar 《Biochimica et Biophysica Acta (BBA)/General Subjects》2014
Background
Redox signaling is an important emerging mechanism of cellular function. Dysfunctional redox signaling is increasingly implicated in numerous pathologies, including atherosclerosis, diabetes, and cancer. The molecular messengers in this type of signaling are reactive species which can mediate the post-translational modification of specific groups of proteins, thereby effecting functional changes in the modified proteins. Electrophilic compounds comprise one class of reactive species which can participate in redox signaling. Electrophiles modulate cell function via formation of covalent adducts with proteins, particularly cysteine residues.Scope of review
This review will discuss the commonly used methods of detection for electrophile-sensitive proteins, and will highlight the importance of identifying these proteins for studying redox signaling and developing novel therapeutics.Major conclusions
There are several methods which can be used to detect electrophile-sensitive proteins. These include the use of tagged model electrophiles, as well as derivatization of endogenous electrophile–protein adducts.General significance
In order to understand the mechanisms by which electrophiles mediate redox signaling, it is necessary to identify electrophile-sensitive proteins and quantitatively assess adduct formation. Strengths and limitations of these methods will be discussed. This article is part of a Special Issue entitled Current methods to study reactive oxygen species - pros and cons and biophysics of membrane proteins. Guest Editor: Christine Winterbourn. 相似文献17.
Background
Glutathione-dependent catalysis is a metabolic adaptation to chemical challenges encountered by all life forms. In the course of evolution, nature optimized numerous mechanisms to use glutathione as the most versatile nucleophile for the conversion of a plethora of sulfur-, oxygen- or carbon-containing electrophilic substances.Scope of review
This comprehensive review summarizes fundamental principles of glutathione catalysis and compares the structures and mechanisms of glutathione-dependent enzymes, including glutathione reductase, glutaredoxins, glutathione peroxidases, peroxiredoxins, glyoxalases 1 and 2, glutathione transferases and MAPEG. Moreover, open mechanistic questions, evolutionary aspects and the physiological relevance of glutathione catalysis are discussed for each enzyme family.Major conclusions
It is surprising how little is known about many glutathione-dependent enzymes, how often reaction geometries and acid–base catalysts are neglected, and how many mechanistic puzzles remain unsolved despite almost a century of research. On the one hand, several enzyme families with non-related protein folds recognize the glutathione moiety of their substrates. On the other hand, the thioredoxin fold is often used for glutathione catalysis. Ancient as well as recent structural changes of this fold did not only significantly alter the reaction mechanism, but also resulted in completely different protein functions.General significance
Glutathione-dependent enzymes are excellent study objects for structure–function relationships and molecular evolution. Notably, in times of systems biology, the outcome of models on glutathione metabolism and redox regulation is more than questionable as long as fundamental enzyme properties are neither studied nor understood. Furthermore, several of the presented mechanisms could have implications for drug development. This article is part of a Special Issue entitled Cellular functions of glutathione. 相似文献18.
Joshua Heinemann Timothy Hamerly Walid S. Maaty Navid Movahed Joseph D. Steffens Benjamin D. Reeves Jonathan K. Hilmer Jesse Therien Paul A. Grieco John W. Peters Brian Bothner 《Biochimica et Biophysica Acta (BBA)/General Subjects》2014
Background
The current paradigm of intracellular redox chemistry maintains that cells establish a reducing environment maintained by a pool of small molecule and protein thiol to protect against oxidative damage. This strategy is conserved in mesophilic organisms from all domains of life, but has been confounded in thermophilic organisms where evidence suggests that intracellular proteins have abundant disulfides.Methods
Chemical labeling and 2-dimensional gel electrophoresis were used to capture disulfide bonding in the proteome of the model thermophile Sulfolobus solfataricus. The redox poise of the metabolome was characterized using both chemical labeling and untargeted liquid chromatography mass spectrometry. Gene annotation was undertaken using support vector machine based pattern recognition.Results
Proteomic analysis indicated the intracellular protein thiol of S. solfataricus was primarily in the disulfide form. Metabolic characterization revealed a lack of reduced small molecule thiol. Glutathione was found primarily in the oxidized state (GSSG), at relatively low concentration. Combined with genetic analysis, this evidence shows that pathways for synthesis of glutathione do exist in the archaeal domain.Conclusions
In observed thermophilic organisms, thiol abundance and redox poise suggest that this system is not directly utilized for protection against oxidative damage. Instead, a more oxidized intracellular environment promotes disulfide bonding, a critical adaptation for protein thermostability.General significance
Based on the placement of thermophilic archaea close to the last universal common ancestor in rRNA phylogenies, we hypothesize that thiol-based redox systems are derived from metabolic pathways originally tasked with promoting protein stability. 相似文献19.
Radosveta Gencheva Qing Cheng Elias S.J. Arnér 《Biochimica et Biophysica Acta (BBA)/General Subjects》2018,1862(11):2511-2517
Background
Toxoflavin (1,6-dimethylpyrimido[5,4-e][1,2,4]triazine-5,7-dione; xanthothricin) is a well-known natural toxin of the pyrimidinetriazinedione type that redox cycles with oxygen under reducing conditions. In mammalian systems, toxoflavin is an inhibitor of Wnt signaling as well as of SIRT1 and SIRT2 activities, but other molecular targets in mammalian cells have been scarcely studied. Interestingly, in a library of nearly 400,000 compounds (PubChem assay ID 588456), toxoflavin was identified as one out of only 56 potential substrates of the mammalian selenoprotein thioredoxin reductase 1 (TrxR1, TXNRD1). This activity was here examined in further detail.Methods
Kinetic parameters in interactions of toxoflavin with rat or human TrxR isoenzymes were determined and compared with those of juglone (5-Hydroxy-1,4-naphthoquinone; walnut toxin) and 9,10-phenanthrene quinone. Selenocysteine dependence was examined using Sec-to-Cys and Sec-to-Ser substituted variants of recombinant rat TrxR1.Results
Toxoflavin was confirmed as an efficient substrate for TrxR. Rat and human cytosolic TrxR1 supported NADPH-dependent redox cycling coupled to toxoflavin reduction, accompanied by H2O2 production under aerobic conditions. Apparent kinetic parameters for the initial rates of reduction showed that rat TrxR1 displayed higher apparent turnover (kcat?=?1700?±?330?min?1) than human TrxR1 (kcat?=?1100?±?82?min?1) but also a higher Km (Km?=?24?±?4.3?μM for human TrxR1 versus Km?=?54?±?18?μM for rat TrxR1). Human TrxR2 (TXNRD2) was less efficient in reduction of toxoflavin (Km?=?280?±?110?μM and kcat?=?740?±?240?min?1). The activity was absolutely dependent upon selenocysteine (Sec). Toxoflavin was also a subversive substrate indirectly inhibiting reduction of other substrates of TrxR1.Conclusions
Our results identify toxoflavin as an efficient redox cycling substrate of mammalian TrxR enzymes, in a strict Sec-dependent manner.General significance
Тhe interactions of toxoflavin with mammalian TrxR isoenzymes can help to explain parts of the molecular mechanisms giving rise to the well-known toxicity as well as pro-oxidant properties of this toxin. 相似文献20.