scholarly journals Selective, Modular Probes for Thioredoxins Enabled by Rational Tuning of a Unique Disulfide Structure Motif

2021 ◽  
Author(s):  
Jan Felber ◽  
Lukas Zeisel ◽  
Lena Poczka ◽  
Karoline Scholzen ◽  
Sander Busker ◽  
...  
Keyword(s):  
Redox Homeostasis ◽  
Selective Reduction ◽  
Effector Proteins ◽  
Exchange Reactions ◽  
Redox Systems ◽  
Disulfide Reduction ◽  
Redox Active ◽  
Design Paradigm ◽  
Fluorogenic Probes ◽  
Ca 50

<p>Specialised cellular networks of oxidoreductases coordinate the dithiol/disulfide-exchange reactions that control metabolism, protein regulation, and redox homeostasis. For probes to be selective for redox enzymes and effector proteins (nM to µM concentrations), they must also be able to resist nonspecific triggering by the ca. 50 mM background of non-catalytic cellular monothiols. However, no such selective reduction-sensing systems have yet been established. Here, we used rational structural design to independently vary thermodynamic and kinetic aspects of disulfide stability, creating a series of unusual disulfide reduction trigger units designed for stability to monothiols. We integrated the motifs into modular series of fluorogenic probes that release and activate an arbitrary chemical cargo upon reduction, and compared their performance to that of the literature-known disulfides. The probes were comprehensively screened for biological stability and selectivity against a range of redox effector proteins and enzymes. This design process delivered the first disulfide probes with excellent stability to monothiols, yet high selectivity for the key redox-active protein effector, thioredoxin. We anticipate that further applications of these novel disulfide triggers will deliver unique probes targeting cellular thioredoxins. We also anticipate that further tuning following this design paradigm will deliver redox probes for other important dithiol-manifold redox proteins, that will be useful in revealing the hitherto hidden dynamics of endogenous cellular redox systems.</p>

scholarly journals Selective, Modular Probes for Thioredoxins Enabled by Rational Tuning of a Unique Disulfide Structure Motif

2021 ◽  
Author(s):  
Jan Felber ◽  
Lukas Zeisel ◽  
Lena Poczka ◽  
Karoline Scholzen ◽  
Sander Busker ◽  
...  
Keyword(s):  
Redox Homeostasis ◽  
Selective Reduction ◽  
Effector Proteins ◽  
Exchange Reactions ◽  
Redox Systems ◽  
Disulfide Reduction ◽  
Redox Active ◽  
Design Paradigm ◽  
Fluorogenic Probes ◽  
Ca 50

<p>Specialised cellular networks of oxidoreductases coordinate the dithiol/disulfide-exchange reactions that control metabolism, protein regulation, and redox homeostasis. For probes to be selective for redox enzymes and effector proteins (nM to µM concentrations), they must also be able to resist nonspecific triggering by the ca. 50 mM background of non-catalytic cellular monothiols. However, no such selective reduction-sensing systems have yet been established. Here, we used rational structural design to independently vary thermodynamic and kinetic aspects of disulfide stability, creating a series of unusual disulfide reduction trigger units designed for stability to monothiols. We integrated the motifs into modular series of fluorogenic probes that release and activate an arbitrary chemical cargo upon reduction, and compared their performance to that of the literature-known disulfides. The probes were comprehensively screened for biological stability and selectivity against a range of redox effector proteins and enzymes. This design process delivered the first disulfide probes with excellent stability to monothiols, yet high selectivity for the key redox-active protein effector, thioredoxin. We anticipate that further applications of these novel disulfide triggers will deliver unique probes targeting cellular thioredoxins. We also anticipate that further tuning following this design paradigm will deliver redox probes for other important dithiol-manifold redox proteins, that will be useful in revealing the hitherto hidden dynamics of endogenous cellular redox systems.</p>

scholarly journals Endogenous formaldehyde scavenges cellular glutathione resulting in cytotoxic redox disruption

2020 ◽  
Author(s):  
Carla Umansky ◽  
Agustín Morellato ◽  
Marco Scheidegger ◽  
Matthias Rieckher ◽  
Manuela R. Martinefski ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Dna Repair ◽  
Fanconi Anemia ◽  
Redox Homeostasis ◽  
Thiol Group ◽  
Cellular Damage ◽  
Repair Pathway ◽  
Cellular Redox ◽  
Redox Active ◽  
Development And Aging

AbstractFormaldehyde (FA) is a ubiquitous endogenous and environmental metabolite that is thought to exert cytotoxicity through DNA and DNA-protein crosslinking. We show here that FA can cause cellular damage beyond genotoxicity by triggering oxidative stress, which is prevented by the enzyme alcohol dehydrogenase 5 (ADH5/GSNOR). Mechanistically, we determine that endogenous FA reacts with the redox-active thiol group of glutathione (GSH) forming S-hydroxymethyl-GSH, which is metabolized by ADH5 yielding reduced GSH thus preventing redox disruption. We identify the ADH5-ortholog gene in Caenorhabditis elegans and show that oxidative stress also underlies FA toxicity in nematodes. Moreover, we show that endogenous GSH can protect cells lacking the Fanconi Anemia DNA repair pathway from FA, which might have broad implications for Fanconi Anemia patients and for healthy BRCA2-mutation carriers. We thus establish a highly conserved mechanism through which endogenous FA disrupts the GSH-regulated cellular redox homeostasis that is critical during development and aging.

scholarly journals Understanding Cellular Redox Homeostasis: A Challenge for Precision Medicine

2021 ◽  
Vol 23 (1) ◽  
pp. 106
Author(s):  
Verena Tretter ◽  
Beatrix Hochreiter ◽  
Marie Louise Zach ◽  
Katharina Krenn ◽  
Klaus Ulrich Klein
Keyword(s):  
Oxidative Stress ◽  
Precision Medicine ◽  
Redox Homeostasis ◽  
Reactive Species ◽  
Scientific Field ◽  
Redox Systems ◽  
Cellular Redox ◽  
Redox Biology ◽  
Living Organisms ◽  
Large Repertoire

Living organisms use a large repertoire of anabolic and catabolic reactions to maintain their physiological body functions, many of which include oxidation and reduction of substrates. The scientific field of redox biology tries to understand how redox homeostasis is regulated and maintained and which mechanisms are derailed in diverse pathological developments of diseases, where oxidative or reductive stress is an issue. The term “oxidative stress” is defined as an imbalance between the generation of oxidants and the local antioxidative defense. Key mediators of oxidative stress are reactive species derived from oxygen, nitrogen, and sulfur that are signal factors at physiological concentrations but can damage cellular macromolecules when they accumulate. However, therapeutical targeting of oxidative stress in disease has proven more difficult than previously expected. Major reasons for this are the very delicate cellular redox systems that differ in the subcellular compartments with regard to their concentrations and depending on the physiological or pathological status of cells and organelles (i.e., circadian rhythm, cell cycle, metabolic need, disease stadium). As reactive species are used as signaling molecules, non-targeted broad-spectrum antioxidants in many cases will fail their therapeutic aim. Precision medicine is called to remedy the situation.

scholarly journals Redox regulation as a platform for search and design of new type drugs. Search of gastroprotectors among sunstituted coumarins

2014 ◽  
Vol 12 (4) ◽  
pp. 22-42
Author(s):  
Edgar Andreevich Parfenov ◽  
Vladimir Alexandrovich Trapkov ◽  
Petr Dmitriyevich Shabanov
Keyword(s):  
Redox Regulation ◽  
Redox Homeostasis ◽  
Complex Compounds ◽  
Active Metabolites ◽  
Defence Mechanisms ◽  
Redox Active ◽  
Acute Ethanol ◽  
New Type ◽  
Important Position ◽  
Effective Drugs

Redox homeostasis controls most or all processes of normal and pathological physiology. Important position in the defence mechanisms are redox active metabolites As a robust platform for the development of new effective drugs is not screened, but directed the design and search for low molecular weight metabolites natural redox active compounds. This statement confirms the synthesis and study of new gastroprotektors of complex compounds of copper and zinc with coumarin ligands. On the model of acute ethanol rat gastric ulcer shown that preprocessing all tested complex compounds in equimolar doses of 0.15 mmol/kg leads to a considerable reduction in the size of the damage to the wall of the stomach, compared to control, and sucralfat (for 72,5-87,9 %, sucralfat - 52,3%).

The Inhibition of Photosynthetic Electron Transport by DBMIB and its Restoration by p-Phenylenediamines; Studied by Means of Prompt and Delayed Chlorophyll Fluorescence of Green Algae

1974 ◽  
Vol 29 (11-12) ◽  
pp. 725-732 ◽  
Author(s):  
Robert Bauer ◽  
Mathijs J. G. Wijnands
Keyword(s):  
Chlorophyll Fluorescence ◽  
Electron Transport ◽  
Photosystem Ii ◽  
Photosynthetic Electron Transport ◽  
Primary Electron ◽  
Delayed Fluorescence ◽  
Fluorescence Induction ◽  
Chlorophyll Fluorescence Induction ◽  
Exchange Reactions ◽  
Redox Systems

Abstract The effect of the plastohydroquinone antagonist dibromothym oquinone (DBMIB) on photosynthetic electron transport reactions was studied in the presence and absence of p-phenylene-diamines by means of measurements of prompt and delayed chlorophyll fluorescence induction of the green alga Scenedesm us obliquus. Prompt and delayed chlorophyll fluorescence induction phenomena are valid indicators for the native presence of and cooperation between the two photosynthetic light reactions. Their kinetics reflect the balancing of electron exchange reactions in the chain of coupled redox-systems between the two photosystems upon sudden illumination. From distinct alterations of the short-term (sec) light induced changes in the yield of prom pt and delayed chlorophyll fluorescence it is concluded that DBMIB inhibits the photosynthetic electron transport in the chain of redox-systems between the two light reactions. There is evidence to show that upon illumination of DBMIB treated cells only the reduction of primary electron ac­ceptor pools of photosystem II (i. e. Q and PQ) is still possible. After their reduction the further electron transport through photosystem II is blocked. The addition of p-phenylenediamines to DBM IB-treated cells abolishes the typical DBMIB-affected prom pt and delayed fluorescence inhibition curves and the normal induction curves re­ appear qualitatively in all their important features. From these measurements it is suggested that the redox properties of p-phenylenediamines allow an electron transport bypass of the DBMIB inhibition site which results in a fully restored photosynthetic electron transport from water to NADP.

scholarly journals Redox Systems in Botrytis cinerea: Impact on Development and Virulence

2014 ◽  
Vol 27 (8) ◽  
pp. 858-874 ◽  
Author(s):  
Anne Viefhues ◽  
Jens Heller ◽  
Nora Temme ◽  
Paul Tudzynski
Keyword(s):  
Botrytis Cinerea ◽  
Redox Homeostasis ◽  
Thioredoxin Reductase ◽  
Redox System ◽  
Redox Status ◽  
Redox Systems ◽  
Gene Encoding ◽  
Cellular Redox ◽  
Thioredoxin System ◽  
The Impact

The thioredoxin system is of great importance for maintenance of cellular redox homeostasis. Here, we show that it has a severe influence on virulence of Botrytis cinerea, demonstrating that redox processes are important for host-pathogen interactions in this necrotrophic plant pathogen. The thioredoxin system is composed of two enzymes, the thioredoxin and the thioredoxin reductase. We identified two genes encoding for thioredoxins (bctrx1, bctrx2) and one gene encoding for a thioredoxin reductase (bctrr1) in the genome of B. cinerea. Knockout mutants of bctrx1 and bctrr1 were severely impaired in virulence and more sensitive to oxidative stress. Additionally, Δbctrr1 showed enhanced H2O2 production and retarded growth. To investigate the impact of the second major cellular redox system, glutathione, we generated deletion mutants for two glutathione reductase genes. The effects were only marginal; deletion of bcglr1 resulted in reduced germination and, correspondingly, to retarded infection as well as reduced growth on minimal medium, whereas bcglr2 deletion had no distinctive phenotype. In summary, we showed that the balanced redox status maintained by the thioredoxin system is essential for development and pathogenesis of B. cinerea, whereas the second major cellular redox system, the glutathione system, seems to have only minor impact on these processes.

Redox signals as a language of interorganellar communication in plant cells

2013 ◽  
Vol 8 (12) ◽  
pp. 1153-1163 ◽  
Author(s):  
Tomasz Kopczewski ◽  
Elżbieta Kuźniak
Keyword(s):  
Redox Regulation ◽  
Biological Effects ◽  
Physiological Responses ◽  
Plant Cells ◽  
Enzymatic Antioxidants ◽  
Oxygen Species ◽  
Redox Systems ◽  
Disulfide Exchange ◽  
Redox Active

AbstractPlants are redox systems and redox-active compounds control and regulate all aspects of their life. Recent studies have shown that changes in reactive oxygen species (ROS) concentration mediated by enzymatic and non-enzymatic antioxidants are transferred into redox signals used by plants to activate various physiological responses. An overview of the main antioxidants and redox signaling in plant cells is presented. In this review, the biological effects of ROS and related redox signals are discussed in the context of acclimation to changing environmental conditions. Special attention is paid to the role of thiol/disulfide exchange via thioredoxins (Trxs), glutaredoxins (Grxs) and peroxiredoxins (Prxs) in the redox regulatory network. In plants, chloroplasts and mitochondria occupying a chloroplasts and mitochondria play key roles in cellular metabolism as well as in redox regulation and signaling. The integrated redox functions of these organelles are discussed with emphasis on the importance of the chloroplast and mitochondrion to the nucleus retrograde signaling in acclimatory and stress response.

scholarly journals Functional Studies of Multiple Thioredoxins from Mycobacterium tuberculosis

2008 ◽  
Vol 190 (21) ◽  
pp. 7087-7095 ◽  
Author(s):  
Mohd Akif ◽  
Garima Khare ◽  
Anil K. Tyagi ◽  
Shekhar C. Mande ◽  
Abhijit A. Sardesai
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Redox Homeostasis ◽  
Biologically Active ◽  
Dependent Manner ◽  
Cytoplasmic Protein ◽  
Redox Potentials ◽  
Exchange Reactions ◽  
E Coli ◽  
Thioredoxin System ◽  
Protein Reduction

ABSTRACT Cytoplasmic protein reduction via generalized thiol/disulfide exchange reactions and maintenance of cellular redox homeostasis is mediated by the thioredoxin superfamily of proteins. Here, we describe the characterization of the thioredoxin system from Mycobacterium tuberculosis, whose genome bears the potential to encode three putative thioredoxins from the open reading frames designated trxAMtb , trxBMtb , and trxCMtb . We show that all three thioredoxins, overproduced in Escherichia coli, are able to reduce insulin, a model substrate, in the presence of dithiothreitol. However, we observe that thioredoxin reductase is not capable of reducing TrxA Mtb in an NADPH-dependent manner, indicating that only TrxB Mtb and TrxC Mtb are the biologically active disulfide reductases. The absence of detectable mRNA transcripts of trxAMtb observed when M. tuberculosis strain H37Rv was cultivated under different growth conditions suggests that trxAMtb expression may be cryptic. The measured redox potentials of TrxB Mtb and TrxC Mtb (−262 ± 2 mV and −269 ± 2 mV, respectively) render these proteins somewhat more oxidizing than E. coli thioredoxin 1 (TrxA). In E. coli strains lacking components of cytoplasmic protein reduction pathways, heterologous expression of the mycobacterial thioredoxins was able to effectively substitute for their function.

Thioredoxin inhibitor PX-12 induces mitochondria-mediated apoptosis in acute lymphoblastic leukemia cells

2020 ◽  
Vol 401 (2) ◽  
pp. 273-283
Author(s):  
Vanessa Ehrenfeld ◽  
Simone Fulda
Keyword(s):  
Cell Death ◽  
Acute Lymphoblastic Leukemia ◽  
Caspase 3 ◽  
Therapeutic Potential ◽  
Lymphoblastic Leukemia ◽  
Redox Homeostasis ◽  
Dependent Manner ◽  
Redox Active ◽  
Ros Scavenger ◽  
Pre Treatment

AbstractImbalances in redox homeostasis have been described to be involved in the development, progression and relapse of leukemia. As the thioredoxin (Trx) system, one of the major cellular antioxidant networks, has been implicated in acute lymphoblastic leukemia (ALL), we investigated the therapeutic potential of Trx inhibition in ALL. Here, we show that the Trx inhibitor PX-12 reduced cell viability and induced cell death in a dose- and time-dependent manner in different ALL cell lines. This antileukemic activity was accompanied by an increase in reactive oxygen species (ROS) levels and enhanced PRDX3 dimerization. Pre-treatment with the thiol-containing ROS scavenger N-acetylcysteine (NAC), but not with non-thiol-containing scavengers α-tocopherol (α-Toc) or Mn(III)tetrakis(4-benzoic acid) porphyrin chloride (MnTBAP), significantly rescued PX-12-induced cell death. Furthermore, PX-12 triggered activation of BAK. Importantly, knockdown of BAK reduced PX-12-stimulated ROS production and cell death. Similarly, silencing of NOXA provided significant protection from PX-12-mediated cell death. The relevance of mitochondria-mediated, caspase-dependent apoptosis was further supported by data showing that PX-12 triggered cleavage of caspase-3 and that addition of the broad-range caspase inhibitor carbobenzoxy-valyl-alanyl-aspartyl-[O-methyl]-fluoromethylketone (zVAD.fmk) potently blocked cell death upon PX-12 treatment. This study provides novel insights into the mechanisms of PX-12-induced cell death in ALL and further highlights the therapeutic potential of redox-active compounds in ALL.

scholarly journals Structural determinants of persulfide-sensing specificity in a dithiol-based transcriptional regulator

2020 ◽  
Author(s):  
Daiana A. Capdevila ◽  
Brenna J. C. Walsh ◽  
Yifan Zhang ◽  
Christopher Dietrich ◽  
Giovanni Gonzalez-Gutierrez ◽  
...  
Keyword(s):  
Redox Homeostasis ◽  
Bacterial Pathogen ◽  
Structural Determinants ◽  
Sulfur Species ◽  
Antibiotic Resistant ◽  
Cellular Processes ◽  
Redox Active ◽  
Bacterial Infectious Disease ◽  
Potential Applications ◽  
Crystallographic Structures

AbstractCysteine thiol-based transcriptional regulators orchestrate coordinated regulation of redox homeostasis and other cellular processes by “sensing” or detecting a specific redox-active molecule, which in turn activates the transcription of a specific detoxification pathway. The extent to which these sensors are truly specific in cells for a singular class of reactive small molecule stressors, e.g., reactive oxygen or sulfur species, is largely unknown. Here we report novel structural and mechanistic insights into a thiol-based transcriptional repressor SqrR, that reacts exclusively with organic and inorganic oxidized sulfur species, e.g., persulfides, to yield a unique tetrasulfide bridge that allosterically inhibits DNA operator-promoter binding. Evaluation of five crystallographic structures of SqrR in various derivatized states, coupled with the results of a mass spectrometry-based kinetic profiling strategy, suggest that persulfide selectivity is determined by structural frustration of the disulfide form. This energetic roadblock effectively decreases the reactivity toward major oxidants to kinetically favor formation of the tetrasulfide product. These findings lead to the identification of an uncharacterized repressor from the increasingly antibiotic-resistant bacterial pathogen, Acinetobacter baumannii, as a persulfide sensor, illustrating the predictive power of this work and potential applications to bacterial infectious disease.

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