Modification Of Thiol Groups Research Articles

Supersulfides to Regulate Electrophilic Stress

Environmental electrophiles modify thiol groups of proteins in organs, disrupting cellular functions carried out by the modified proteins and increasing the risk of various diseases. The transcription factor NF-E2-related factor 2 (Nrf2) plays a crucial role in detoxifying electrophiles by forming glutathione adducts and subsequently excreting them into extracellular spaces. Supersulfides such as cysteine persulfides (CysSSH) produced by cystathionine γ-lyase (CSE) capture environmental electrophiles through sulfur adduct formation. However, the Nrf2 and CSE contributions to blocking environmental electrophile-mediated toxicity have yet to be evaluated. Therefore, we assessed the individual and combined roles of Nrf2 and CSE in suppressing toxicity induced by environmental electrophiles using Nrf2 knockout (KO), CSE KO, and Nrf2/CSE double KO (DKO) mice. Our findings indicate that CSE/Nrf2 DKO mice are more sensitive to environmental electrophiles compared to their single KO counterparts, highlighting the distinct mechanisms through which both pathways mitigate the toxic effects of reactive electrophiles. Moreover, diverse metabolites produced by symbiotic gut bacteria in the human body are known to exert various effects on host organ functions beyond the intestinal tract. We observed reduced blood supersulfide levels in mice lacking gut microflora compared to normal mice. Furthermore, we identified intestinal bacteria belonging to the families Ruminococcaceae and Lachnospiraceae as high CysSSH-producing bacteria. This suggests that the gut microbiota serves as a source of in vivo supersulfide molecules. These findings suggest that supersulfide derived from gut bacteria may act protectively against environmental electrophilic exposure in the host.

Transient Receptor Potential channels (TRP) in GtoPdb v.2023.3

Transient Receptor Potential channels (TRP) in GtoPdb v.2023.3

Transient Receptor Potential channels (TRP) in GtoPdb v.2023.2

Transient Receptor Potential channels (TRP) in GtoPdb v.2023.2

P-085 Reactive molecules drive sperm capacitation via protein posttranslational modifications

Abstract Study question Are S-sulfenylation and S-nitrosylation, two essential posttranslational modifications (PTMs), increased during sperm capacitation? Which proteins are subjected to S-sulfenylation? Summary answer S-sulfenylation and S-nitrosylation increase during sperm capacitation. S-sulfenylation affects certain proteins, including A-kinase anchor protein 13, Fibrilline-1, and Spermatogenesis-associated protein 7. What is known already There is a significant increase in reactive oxygen species (ROS) and reactive nitrogen species (RNS) production within sperm capacitation. In contrast to tyrosine phosphorylation, a well-established mark of sperm capacitation, ROS, and RNS lead to PTMs themselves and modify thiol groups of cysteines. Despite these facts, the dynamics of ROS/RNS-derived PTMs during sperm capacitation remain unknown. Therefore, we consider both S-nitrosylation and S-sulfenylation of sperm proteins as novel modifications associated with the capacitation. Study design, size, duration C57BL6 mouse sperm were isolated from the caudae epididymae of 12-20-week-old males (n = 5). The study compared non-capacitated and capacitated spermatozoa using proteomic profiling. Participants/materials, setting, methods Sperm proteins were extracted into RIPA buffer. The protein lysate was incubated with 5mM dimedone, a hapten that specifically labels S-sulfenylated proteins, and immunogenically detected with an anti-dimedone antibody (Merk, Germany). Alternatively, S-nitrosylated proteins were detected by the anti-nitrosocysteine antibody (Merk, Germany). S-sulfenylated and S-nitrosylated proteins were visualized by western blotting. S-sulfenylated proteins were simultaneously identified by mass spectrometry. Main results and the role of chance We demonstrated an increase in S-sulfenylation and S-nitrosylation along with sperm capacitation. In addition, both S-sulfenylation and S-nitrosylation are positively correlated with tyrosine phosphorylation. Using mass spectrometry, we identified S-sulfenylated proteins, mostly belonging to the cytoskeleton: A-kinase anchor protein 13, Fibrilline-1, Spermatogenesis-associated protein 7, and Rab-like protein 2A. Limitations, reasons for caution The limitation of this study is the instability of PTMs. ROS/RNS-derived PTMs hypothetically arise and cease during protein sample preparation. The experiment was limited to one animal model. Further experiments should be performed on other mammalian models. Wider implications of the findings Studying novel sperm PTMs and their mechanism of action may improve the diagnosis of male infertility. Further, it could bring a novel approach to modulate the fertilizing ability of sperm used for in vitro fertilization. Trial registration number 260 536

Transient Receptor Potential channels (TRP) in GtoPdb v.2023.1

Transient Receptor Potential channels (TRP) in GtoPdb v.2023.1

Cover Feature: Magnetic Relaxation and Modification of Thiol Groups on Co‐Mg Ferrite Nanoparticles for Theranostics (ChemNanoMat 10/2022)

Magnetic nanoparticles (MNPs) have been actively researched for biomedical applications. This work reports the magnetic and thermal properties of various types of MNPs and confirms a significant hyperthermia effect of Mn-Zn ferrite nanoparticles in human breast cancer cells. Doping with Co ions with larger anisotropy is a possible approach to obtain higher magnetization. However, Co-ferrite particles exhibit hysteresis in their magnetization curves, hindering the maintenance of the superparamagnetic behavior. In contrast, Mg-ferrite micro-order clusters have remarkable heat generation properties for hyperthermia treatment in external magnetic fields. Thus, new properties can be expected from the synergistic effect of Co and Mg ions. Co1-xMgxFe2O4 (x = 0.0, 0.2, 0.4, 0.6, 0.8, 1.0) nanoparticles embedded in amorphous SiO2 with particle sizes of approximately 4.5 nm were prepared using an original wet chemical method. Spin-echo magnetic resonance imaging measurements were performed for Co1-xMgxFe2O4 nanoparticles using a 0.3-T MRI system. The particles exhibited a significant T2 shortening effect compared with that of agarose, depending on the composition. All particles exhibited effective relaxivity, R2, and a significant contrast was observed in the phantom image. More information can be found in the Research Article by Yuko Ichiyanagi et al.

Abstract 6184: Kras Co-mutations lead to resistance to mutant TP53 targeted therapy in mouse models of spontaneous non-small cell lung cancer

Abstract Background: Lung cancer results in more than 131,000 deaths per year with estimated 236,000 new cases in 2021 in the United States. About 85% of lung cancers are non-small cell lung cancer (NSCLC). TP53 is an important tumor-suppressor and about 50% of human lung NSCLCs contain a mutant TP53. Targeting of mutant p53 is very limited due to the lack of drugs. PRIMA-1 is a first in class anti-mutant TP53 molecule with the ability to induce apoptosis by restoring the wild-type function of TP53 through covalently binding and modifying thiol groups in the central domain of the mutated protein. Recently, the FDA granted breakthrough therapy designation to the PRIMA-1 methylated analog, APR-246 in combination with azacytidine for the treatment of patients with myelodysplastic syndromes and a TP53 mutation. The major setback of targeted therapy today continues to be resistance. Herein, we report our findings related to resistance to PRIMA-1 targeted therapy in spontaneous P53 mutant driven NSCLC. Methods: We have previously reported the generation of a line of transgenic mice with a human mutant p53(273H) expressed in a lung-specific manner. Human mutant TP53 expression was regulated under the transcriptional control of the human surfactant protein C (SP-C) promoter. After the age of 12 months, the mice developed spontaneous lung adenocarcinomas. We treated SPC-p53-273H lung tumor mice via intraperitoneal injection (i.p) with PRIMA-1 at a dose of 100 mg/kg in 0.2 mL PBS every other day for two weeks. The tumors were identified using micro-computed tomography (micro-CT) system and pre-and post-treatment diameters were estimated based on the CT image. K-ras status was analyzed by PCR-based Sanger sequencing. Results: Of the 10 lung tumor-bearing mice treated with PRIMA-1, micro-CT derived sum of the largest diameters indicated that six mice (60%) experienced tumor volume reductions whereas four mice experienced tumor volume increase. Sequencing analysis showed five tumors contained K-ras mutations (two p.G12D, one p.G12F, one p.G12V and one p.Q61L). On average, lung tumor diameter had a 31% reduction post-treatment of PRIMA-1 for the lung tumors containing wild-type K-ras gene. Whereas it increased 18% on the average post-treatment of PRIMA-1 for the lung tumors containing a mutant K-ras gene. There is a significant difference statistically (p = 0.016) in the response to the treatment of PRIMA-1 between the mutant K-ras cohort and the wild-type K-ras cohort. Conclusion: PRIMA 1 therapy results in antitumor activity against P53 mutant lung tumors. However, the presence of K-ras co-mutation is a significant resistance factor. Combination of APR-246 and K-ras inhibitor therapy would be needed to overcome this resistance. Citation Format: David T. Costa, Shaylyn Grier, Melanie Perez, Li Gao, Miguel A. Villalona-Calero, Wenrui Duan. Kras Co-mutations lead to resistance to mutant TP53 targeted therapy in mouse models of spontaneous non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6184.

Magnetic Relaxation and Modification of Thiol Groups on Co‐Mg Ferrite Nanoparticles for Theranostics

AbstractCo1‐xMgxFe2O4(x=0.0, 0.2, 0.4, 0.6, 0.8, 1.0) nanoparticles embedded in amorphous SiO2with particle sizes of approximately 4.5 nm were prepared using an original wet chemical method. We performed spin‐echo magnetic resonance imaging measurements for Co1‐xMgxFe2O4nanoparticles using a 0.3‐T MRI system. The particles exhibited a significantT2shortening effect compared with that of agarose, depending on the composition. All particles exhibited effective relaxivity,R2, and a significant contrast was observed in the phantom image. Furthermore, thiol group was modified to enable particles to bind specifically to maleimide proteins. These particles are expected to be potential theranostic agents.

Transient Receptor Potential channels (TRP) in GtoPdb v.2022.1

Transient Receptor Potential channels (TRP) in GtoPdb v.2022.1

Methods to Analyze the Redox Reactivity of Plant Proteins.

Proteins can be covalently modified by a broad range of highly reactive chemicals and redox mechanisms. Reversible redox-mediated post-translational modifications of sensitive cysteine thiol groups in proteins impact protein characteristics such as interaction behavior and activity state. Evaluating the response of proteins to redox perturbation or reactive chemical species is critical for understanding the underlying mechanisms involved and their contribution to plant stress physiology. Here we provide a detailed workflow that includes procedures for (i) purification, processing, and analysis of protein samples with redox agents, (ii) determining redox-modulated monomer to oligomer transitions using size exclusion chromatography, and (iii) activity assays for monitoring the impact of redox agents on purified enzymes and in crude extracts from plants subjected to oxidative stress. We exemplified how to apply several of the methods discussed for analyzing redox-sensing metallopeptidases, such as thimet oligopeptidases. We anticipate that these protocols should find broad applications in monitoring biochemical properties of other classes of redox-sensitive plant proteins.

Synthesis of novel thiol-modified lysozyme coated magnetic nanoparticles for the high selective adsorption of Hg(II)

As a common heavy metal ion in wastewater, Hg(II) has high toxicity even at low concentration. In this study, a novel magnetic nano-adsorbent (FS@PTL-SH) was synthesized by modifying thiol groups on phase-transited lysozyme (PTL) coated magnetic nanoparticles. FS@PTL-SH exhibited superior adsorption performance for Hg(II) in a wide pH range. The maximum adsorption capacity of FS@PTL-SH fitted by Langmuir model could reach 1001.21 mg/g at 298 K, and 90% of the equilibrium uptake could be achieved within 5 min. The interference of inorganic ions and organic complexing agents was limited even at high concentrations. High adsorption selectivity of Hg(II) was observed in the mixed solution containing various heavy metal ions. Moreover, FS@PTL-SH could still maintain above 90% of the adsorption capacity after five adsorption–desorption cycles. For the mechanisms, the adsorption of FS@PTL-SH for Hg(II) was mainly relied on the strong complexation of the surface functional groups, especially thiol groups.

A new electrochemical aptasensor based on gold/nitrogen-doped carbon nano-onions for the detection of Staphylococcus aureus

Bacterial diseases are serious problems in the world today. The Staphylococcus aureus (S. aureus) bacterium is one of the most invasive and resistant bacteria that infects humans, making it a common cause of hospital infections. Herein, a very sensitive aptasensor for S. aureus was constructed using nitrogen-doped carbon nano-onions, gold nanoparticles (AuNPs) and thiol-terminated, aptamer-modified, screen-printed carbon electrodes. The physicochemical characteristics of the nanoparticles and the analytical parameters of the aptasensor were determined by several methods: energy dispersive X-ray spectroscopy, scanning and transmission electron microscopy, cyclic voltammetry and electrochemical impedance spectroscopy. The aptamer was self-assembled on the electrode surface through the covalent modification of thiol groups with AuNPs. The analytical parameters of this sensor were improved by immobilizing a carbon layer on the electrode surface. The developed aptasensor can measure S. aureus quickly with a limit of detection of 3 CFU/mL in the linear range of 10–108 CFU/mL. The aptasensor for S. aureus has good repeatability, reproducibility, and long-term stability and can measure S. aureus bacteria in human serum samples.

Transient Receptor Potential channels (TRP) in GtoPdb v.2021.3

Transient Receptor Potential channels (TRP) in GtoPdb v.2021.3

Study of Effect of Nitrosamine Stress in Progression of Leprosy

Objective: In several countries, including India, leprosy is an older disease & till now continues to be an important health issue. Leprosy is a chronic granulomatous disease, the contributing agent for this is mycobacterium leprae. Nitric oxide (NO) plays a significant role in immunity to fight against bacteria. But increased NO can harm host tissue by causing altered structure of thiol-containing compounds, which is a significant event that influences the pathogenesis of leprosy. Considering this, the study was planned to determine the type of relationship between NO, nitrothiols, and thiols as disease advances.
 Methods: 50 newly diagnosed leprosy patients & 50 healthy controls were included in the study. In the leprosy group,16 were Paucibacillary (PB), and 34 were Multibacillary (MB) type, leprosy patients.
 Results: Serum NO and serum nitrothiol significantly increased (p<0.01) in leprosy patients than controls. Further, Serum NO and serum nitrothiol significantly increased (p<0.01) in MB leprosy patients than PB leprosy patients. On the other hand, we found a significantly decreased (p<0.01) level of total thiols in leprosy patients compared with controls. Serum thiols significantly decreased (p<0.01) in MB leprosy patients than PB leprosy patients. Among Both leprosy patients, the negative correlation of NO with thiol and nitrothiol with thiols was observed.
 Conclusion: Thus, in leprosy, increased nitric oxide causes modification of thiol groups of proteins and impaired their activity which may be responsible for the severity of disease; hence therapy aimed to reduce nitrosative modification of proteins leprosy.

Persulfidation is the mechanism underlying sulfide-signaling of autophagy

ABSTRACT In this commentary, we highlight the findings described in a recent paper regarding the mechanism of H2S regulation of macroautophagy/autophagy in mammalian cells and discuss the similarities/divergencies with plant cells. The main outcome is that the posttranslational modification of thiol groups of cysteine residues to form persulfides is a conserved molecular mechanism.

Hydrogen sulfide signaling in plant adaptations to adverse conditions: molecular mechanisms.

Hydrogen sulfide (H2S) is a signaling molecule that regulates critical processes and allows plants to adapt to adverse conditions. The molecular mechanism underlying H2S action relies on its chemical reactivity, and the most-well characterized mechanism is persulfidation, which involves the modification of protein thiol groups, resulting in the formation of persulfide groups. This modification causes a change of protein function, altering catalytic activity or intracellular location and inducing important physiological effects. H2S cannot react directly with thiols but instead can react with oxidized cysteine residues; therefore, H2O2 signaling through sulfenylation is required for persulfidation. A comparative study performed in this review reveals 82% identity between sulfenylome and persulfidome. With regard to abscisic acid (ABA) signaling, widespread evidence shows an interconnection between H2S and ABA in the plant response to environmental stress. Proteomic analyses have revealed persulfidation of several proteins involved in the ABA signaling network and have shown that persulfidation is triggered in response to ABA. In guard cells, a complex interaction of H2S and ABA signaling has also been described, and the persulfidation of specific signaling components seems to be the underlying mechanism.

Ginsenoside derivatives inhibit advanced glycation end-product formation and glucose–fructose mediated protein glycation in vitro via a specific structure–activity relationship

Ginseng (Panax ginseng and red ginseng) extract has been reported to inhibit the formation of advanced glycation end-products (AGEs); however, the potential inhibitory activity of its major constituents (ginsenosides) against AGE formation is still unknown. In the present study, we investigated the inhibitory effect of ginsenoside derivatives on AGE formation. Herein, we assessed the activity of 22 ginsenosides, most of which significantly inhibited fluorescent AGE formation. Notably, ginsenoside Rh2, ginsenoside Rh1, and compound K exhibited the most potent AGE inhibitory potential with IC50 values of 3.38, 8.42, and 10.85 µM, respectively. The structure– activity relationship revealed that the presence of sugar moieties, hydroxyl groups, and their linkages, and the stereostructure of the ginsenoside skeleton played an important role in the inhibition of AGE formation. Furthermore, the inhibitory activity of the most active ginsenoside Rh2 on fructose-glucose-mediated protein glycation and oxidation of bovine serum albumin (BSA) was explored. Rh2 (0.1–12.5 µM) inhibited the formation of fluorescent AGE and non-fluorescent AGE, as well as the level of fructosamine and prevented protein oxidation by decreasing protein carbonyl formation and protein thiol group modification. Rh2 also suppressed the formation of the β-cross amyloid structure of BSA. Ginsenosides might be promising new anti-glycation agents for the prevention of diabetic complications via inhibition of AGE formation and oxidation-dependent protein damage.

Alcohol Binge Drinking Selectively Stimulates Protein S-Glutathionylation in Aorta and Liver of ApoE -/- Mice.

Background: Binge drinking has become the most common and deadly pattern of excessive alcohol use in the United States, especially among younger adults. It is closely related to the increased risk of cardiovascular disease. Oxidative stress as a result of ethanol metabolism is the primary pathogenic factor for alcohol-induced end organ injury, but the role of protein S-glutathionylation—a reversible oxidative modification of protein cysteine thiol groups that mediates cellular actions by oxidants—in binge drinking-associated cardiovascular disease has not been explored. The present study defines the effect of alcohol binge drinking on the formation of protein S-glutathionylation in a mouse model of atherosclerosis.Methods and Results: To mimic the weekend binge drinking pattern in humans, ApoE deficient (ApoE−/−) mice on the Lieber-DeCarli liquid diet received ethanol or isocaloric maltose (as a control) gavages (5 g/kg/day, 2 consecutive days/week) for 6 weeks. The primary alcohol-targeted organs (liver, brain), and cardiovascular system (heart, aorta, lung) of these two groups of the mice were determined by measuring the protein S-glutathionylation levels and its regulatory enzymes including [Glutaredoxin1(Grx1), glutathione reductase (GR), glutathione-S-transferase Pi (GST-π)], as well as by assessing aortic endothelial function and liver lipid levels. Our results showed that binge drinking selectively stimulated protein S-glutathionylation in aorta, liver, and brain, which coincided with altered glutathionylation regulatory enzyme expression that is downregulated Grx1 and upregulated GST-π in aorta, massive upregulation of GST-π in liver, and no changes in Grx1 and GST-π in brain. Functionally, binge drinking induced aortic endothelial cell function, as reflected by increased aortic permeability and reduced flow-mediated vasodilation.Conclusions: This study is the first to provide in vivo evidence for differential effects of binge drinking on formation of protein S-glutathionylation and its enzymatic regulation system in major alcohol-target organs and cardiovascular system. The selective induction of protein S-glutathionylation in aorta and liver is associated with aortic endothelial dysfunction and fatty liver, which may be a potential redox mechanism for the increased risk of vascular disease in human binge-drinkers.

Synchronous Construction of Hierarchical Porosity and Thiol Functionalization in COFs for Selective Extraction of Cationic Dyes in Water Samples.

Pore size and functionalization are two critical factors for covalent organic frameworks (COFs) as effective adsorbents. However, due to the low crystallinity of COFs, it is a grand challenge to accomplish pore diameter adjustment and functionalization at the same time. In this work, we developed a simple and ingenious strategy, cutting off linkage, to synchronously construct hierarchical porosity and modify thiol groups in COFs under mild conditions. The hybrid COFs containing disulfide bonds were designed and synthesized, and then the disulfide bonds were cleaved by glutathione, resulting in the formation of thiol groups as well as the increase in pore size caused by skeleton defects. The pore diameter of thiol-functionalized hierarchical porous COFs (denoted as HP-TpEDA-SH) was concentrated at 2.6 and 3.5 nm. Thanks to the electrostatic attraction of thiol groups to cationic dyes and the higher number of available adsorption sites, the maximum extraction amounts of methylene blue (MB), malachite green (MG), and crystal violet (CV) by HP-TpEDA-SH were 2.6, 2.1, and 3.3 times those of microporous COFs under optimal extraction conditions, respectively. The proposed analytical method (solid-phase extraction-high-performance liquid chromatography/ultraviolet (SPE-HPLC/UV)) with HP-TpEDA-SH as the adsorbent showed low detection limits of 1.3, 0.13, and 0.12 μg·L-1 for MB, MG, and CV, respectively. The recoveries of three spiked water samples ranged from 81.5 to 113.8%, with relative standard deviations (RSDs) less than 9.7%. This work not only opened a new avenue for the preparation of functionalized hierarchical porous COFs but also established an effective method for detecting trace cationic dyes in fishery water.

Cysteamine assay for the evaluation of bioactive electrophiles

Covalent modifications of thiol and amine groups may control the function of proteins involved in the regulatory and signaling pathways of the cell. In this study, we developed a simple cysteamine assay which can be used to study the reactivity of electrophilic compounds towards primary amine and thiol groups in an aqueous environment. The detection principle is based on the electrochemical, photometrical and mass spectrometric analyses of cysteamine (2-aminoethanethiol) as the molecular probe. This technique is useful for studying the reaction kinetics of electrophiles with thiol (SH) and amino (NH2) groups. The decrease in analytical responses of cysteamine was monitored to evaluate the reactivity of three electrophilic activators of the Nrf2 pathway, which mediates the cellular stress response. The SH-reactivity under cell-free conditions of the tested electrophiles decreased in the following order: 4-hydroxy-2-nonenal ≥ nitro-oleic acid > sulforaphane. However, as shown in RAW264.7 cells, the tested compounds activated Nrf2-dependent gene expression in the opposite order: sulforaphane > nitro-oleic acid ≥ 4-hydroxy-2-nonenal. Although other factors in addition to chemical reactivity play a role in biological systems, we conclude that this cysteamine assay is a useful tool for screening potentially bioactive electrophiles and for studying their reactivity at a molecular level.