Protein Sulfhydryl Groups Research Articles

Вплив лазерного опромінення на ксантиноксидазну активність та генерацію супероксидного радикала в цитозольній фракції печінки щурів-пухлиноносіїв

The effect of tumor growth in the body and laser irradiation on the enzymatic activity of xanthine oxidase, in particular its D- and O-forms, and also the rate of generation of the superoxide radical (O2–) and the level of protein sulfhydryl groups in the liver rat cytosolic fraction has been investigated. It has been found that in the cytosolic fraction of rats with transplanted Guerin’s carcinoma decreases the enzymatic activity of the D-form of xanthine oxidase with a simultaneous increase in its O-form during the period of intensive (14 days, which corresponds to the logarithmic phase of on cogenesis) and the period of final tumor growth (21 days, which corresponds to the stationary phase of oncogenesis). The increase in the enzymatic activity of the O-form of xanthine oxidase was accompanied by an increase the rate of superoxide radical generation and a decrease in the level of protein SH-groups in the liver cytosolic fraction of tumor-bearing rats. Daily directed action of laser irradiation on the area of growth of Guerin’s carcinoma leads to less destructive changes in the liver. Thus, there is an increase in the enzymatic activity of the D-form of xanthine oxidase, a decrease the rate of superoxide radical formation and an increase the content of protein SH-groups in the cytosolic fraction of the liver of experimental animals compared with non-irradiated tumor-bearing rats.

A water-soluble and incubate-free fluorescent environment-sensitive probe for ultrafast visualization of protein thiols within living cells

The amount of protein thiols play a crucial role in maintaining the cellular redox homeostasis and have significant implications to indicate a series of diseases. Therefore, it is necessary to develop an ideal probe for protein thiol detection in a simple and readily implementable method. Consequently, a water-soluble and incubate-free fluorescent environment-sensitive probe DMTs-OCC was synthesized using 7-diethylamincoumarin as the fluorophore and 4-(5-Methanesulfonyl- [1,2,3,4]tetrazol-1-yl)-phenol (MSTP) as a thiol receptor reagent. The blue-shift emission spectra of probe DMTs-OCC was observed by ultrafast binding to protein sulfhydryl groups from the excited intramolecular charge transfer (ICT) to the twisted intramolecular charge transfer (TICT) conversion process in aqueous solution. The experimental results showed that probe DMTs-OCC exhibited an excellent selectivity to protein thiols and biocompatibility in aqueous solution, as well as terrific cell membrane permeability which enabled the successful visualization of BSA protein thiol in living cells. Moreover, no excess probe was cleaned and no incubation time was needed in cell experiments. Therefore, it could provide a new method to the construction of fluorescent probes for protein thiols labelling and visualization.

Endocan and advanced oxidation protein products in adult population with hypertension.

Hypertension is closely related to oxidative stress and inflammation. Endocan is a new inflammation marker whose role is not completely elucidated in hypertension. The aim of this study was to explore the association between endocan and several oxidative stress markers [i.e., advanced oxidation protein products (AOPP), total protein sulfhydryl (SH-) groups and prooxidant-antioxidant balance (PAB)] in adult population with hypertension. A total of 90 patients with hypertension were compared with 44 controls. Blood pressure, anthropometric and biochemical parameters were measured. Associations of clinical data with hypertension were tested with univariable and multivariable logistic ordinal regression analysis. Endocan and AOPP were significantly higher in hypertensive patients than in the controls (p=0.006 and p=0.046, respectively). In the multivariable logistic regression analysis, AOPP and endocan kept their independent positive associations with hypertension. As AOPP rose by 1 μmol/L and endocan rose by 1 pg/mL, the probability for hypertension presence rose by 4.2% and 32.2%, respectively and 39.9% of variation in hypertension could be explained with the Model. The area under the Receiver Operating Characteristic curve [(AUC) for AOPP=0.638 (0.550-0.719), p=0.01 and for endocan=0.679 (0.593-0.757), p<0.001] demonstrated sufficient clinical accuracy towards hypertension. On the contrary, the Model showed very good clinical accuracy [AUC= 0.825 (0.749-0.900), p<0.001]. Endocan and AOPP are independently correlated with hypertension in adult population and these tested markers together could be reliable parameters to discriminate patients with hypertension from normotensive ones.

Composition and foam properties of whole wheat dough liquor as affected by xylanase and glucose oxidase

Xylanase (XYL) and glucose oxidase (GOX) have been applied to improve the crumb structure of whole wheat buns. However, their effects on the constituents dissolved in the dough liquor (DL), which supposedly contributed to gas cell stabilization and crumb structure, have not been elucidated. In this study, we aimed to unravel the effects of these enzymes on the protein and arabinoxylan contents, DL yields, bulk viscosity, as well as foaming properties. DL was first separated through ultracentrifugation from whole wheat dough prior treated by XYL and GOX. The results indicated that XYL and GOX treatment increased the DL yields. Moreover, while XYL increased the water extractable arabinoxylan content, GOX increased the protein level in the DL. The free sulfhydryl groups of soluble proteins declined significantly in the presence of GOX. Moreover, GOX might have catalyzed the formation of large protein aggregates and protein-polysaccharide complexes. Furthermore, while the foaming capacity of DL increased only in the presence of XYL, the foam stability was increased by both XYL and GOX, especially under heat treatment. The present research might help to understand the mechanism of crumb structure improvement of whole wheat buns by XYL/GOX treatment from the view of their effects on the DL.

Effects of oxidative modification by 13-hydroperoxyoctadecadienoic acid on the structure and functional properties of rice protein

13-hydroperoxyoctadecadienoic acid (13-HPODE) was selected as lipid peroxidation primary products during rice ageing to investigate the effects of lipid hydroperoxide modification on the structural characteristics and functional properties of rice protein. Incubation of rice protein with 13-HPODE resulted in protein carbonylation and loss of protein sulfhydryl groups in a concentration-dependent manner. FT-IR evaluated the effects of 13-HPODE on rice protein, and the results indicated increasing concentration of 13-HPODE resulted in continuous loss of α-helix, β-turn and random coil structure, and increase in β-sheet content and amino acid side chains. The content of soluble protein aggregates gradually increased, accompanied by protein crosslink via disulfide bonds. 13-HPODE had a negative impact on the solubility, water holding capacity, foaming capacity, and foam stability of rice protein. 13-HPODE could promote oil holding capacity of rice protein. With the concentration of 13-HPODE below 0.1 mmol/L, emulsifying activity index (EAI) and emulsifying stability index (ESI) of rice protein gradually increased, which due to gradual increase in rice protein surface hydrophobicity and flexibility. However, at relatively high concentration of 13-HPODE (above 1 mmol/L), the adverse effect in EAI and ESI was observed, which might be attributed to a decrease in surface hydrophobicity and protein aggregates formation.

The role of açaí ( Euterpe oleracea Mart. 1824) as a chemoprotective agent in the evaluation of antioxidant defence, oxidative damage and histology of juvenile shrimp Litopenaeus vannamei (BOONE, 1931) exposed to ammonia

This study investigated the effects of the use of the inclusion of açaí on the diet of shrimp Litopenaeus vannamei on antioxidant and histopathological responses after exposure to ammonia. The shrimps were fed two experimental diets: control and with 10% of açaí inclusion (W/W), for 35 days. Afterwards, the organisms were exposed at four concentrations of ammonia (0.01-control; 0.26; 0.48 and 0.91 mg NH3-N L−1) for 96 hr. The total antioxidant capacity (ACAP) of the gills decreased significantly in both diets when exposed to ammonia, whereas in the muscle, the açaí promoted an increase in ACAP. Concomitantly, lipid peroxidation levels increased in the gills and decreased in muscle. After exposure to ammonia, glutathione-S-transferase activity increased in hepatopancreas in shrimps fed with açaí facilitating the detoxification of lipid peroxidation by-products, and the concentration of protein sulfhydryl groups decreased in the gills and muscle of the shrimp of the control diet, evidencing protein damage, an unobserved response in shrimps that received the açaí diet. Histopathological changes decreased in açaí-fed shrimps about the control diet after exposure to ammonia. It is concluded that açaí mitigated ammonia-induced histopathological changes, improved the antioxidant defence system (gills and muscle) and attenuated the lipid damage in the muscle.

Oxidative Modifications in Advanced Atherosclerotic Plaques: A Focus on In Situ Protein Sulfhydryl Group Oxidation.

Although oxidative stress has been long associated with the genesis and progression of the atherosclerotic plaque, scanty data on its in situ effects on protein sulfhydryl group modifications are available. Within the arterial wall, protein sulfhydryls and low-molecular-weight (LMW) thiols are involved in the cell regulation of both Reactive Oxygen Species (ROS) and Reactive Nitrogen Species (RNS) levels and are a target for several posttranslational oxidative modifications that take place inside the atherosclerotic plaque, probably contributing to both atherogenesis and atherosclerotic plaque progression towards complicated lesions. Advanced carotid plaques are characterized by very high intraplaque GSH levels, due to cell lysis during apoptotic and/or necrotic events, probably responsible for the altered equilibrium among protein sulfhydryls and LMW thiols. Some lines of evidence show that the prooxidant environment present in atherosclerotic tissue could modify filtered proteins also by protein-SH group oxidation, and demonstrate that particularly albumin, once filtered, represents a harmful source of homocysteine and cysteinylglycine inside the plaque. The oxidative modification of protein sulfhydryls, with particular emphasis to protein thiolation by LMW thiols and its association with atherosclerosis, is the main topic of this review.

Effects of short-term fermentation with lactic acid bacteria on egg white: Characterization, rheological and foaming activities

Lactic acid fermentation is a green technology to modify functionalities of food ingredients, including proteins. In this study, the commercial lactic acid bacteria (Streptococcus thermophiles and Lactobacillus bulgaricus) were used to ferment egg white for 0 h, 3 h, 6 h, and 9 h, respectively. The microbial composition significantly changed during the process, and the abundance of B. Lactobacillus peaked at 6 h. The free sulfhydryl groups of egg white proteins (EWP) gradually decreased, and the surface hydrophobicity increased from 28.7 to 52.0 after 6 h. The rheological experiments illustrated that fermented samples exhibited a lower apparent viscosity, and the sample at 6 h possessed the highest storage modulus (G'). Correspondingly, the foaming ability of egg white significantly improved from 72.3% to 93.0% after 6 h of fermentation. This study suggested that moderate lactic acid fermentation endowed egg white with stronger foaming activities, and thus had a potential to be applied in improving texture quality of egg products.

The Intensity of Free Radical Processes on Rat Liver Mitochondria under Moderate Hypothermia of Various Duration

Moderate artificial hypothermia is widely used in clinical practice to protect organs from the effects of ischemia (reperfusion), trauma, and hypoxia. However, a decrease in the body temperature of homeothermic animals induces oxidative stress, the severity of which may depend on the time of exposure to the cold factor. Since mitochondria play a key role in the generation of reactive oxygen species, we studied the dependence of the intensity of free radical processes in rat liver mitochondria on the duration of moderate hypothermia (30°C). It turned out that short-term (30 min) hypothermia activates the processes of lipid peroxidation (POL), while the concentration of lipid hydroperoxides, Schiff bases, and malondialdehyde significantly increases. Prolongation of hypothermia to 1 h reduces the content of many lipid peroxidation products, and their normalization is observed with 3-h hypothermia. Short-term hypothermia and its prolongation up to 1 h is accompanied by oxidative destruction of mitochondrial proteins, which is manifested in a decrease in the content of sulfhydryl groups in them and an increase in carbonyl groups. At the same time, 3-h hypothermia contributes to the normalization of the studied markers of protein oxidative modification. The dynamics of changes in the levels of sulfhydryl and carbonyl groups in the mitochondrial matrix proteins is more pronounced in comparison with membrane proteins. The study of the spectral characteristics of mitochondrial membrane proteins showed a decrease in the intensity of their fluorescence at the initial stages of hypothermia. Tryptophan residues localized at the periphery make the main contribution to it. Prolongation of hypothermia up to 3 h contributes to restoration of fluorescence parameters to the control level. The data obtained in the analysis of the second derivative fluorescence spectra indicate changes in the spatial configuration of membrane proteins.

Malondialdehyde and Uric Acid as Predictors of Adverse Outcome in Patients with Chronic Heart Failure.

In chronic heart failure (HF), some parameters of oxidative stress are correlated with disease severity. The aim of this study was to evaluate the importance of oxidative stress biomarkers in prognostic risk stratification (death and combined endpoint: heart transplantation or death). In 774 patients, aged 48-59 years, with chronic HF with reduced ejection fraction (median: 24.0 (20-29)%), parameters such as total antioxidant capacity, total oxidant status, oxidative stress index, and concentration of uric acid (UA), bilirubin, protein sulfhydryl groups (PSH), and malondialdehyde (MDA) were measured. The parameters were assessed as predictive biomarkers of mortality and combined endpoint in a 1-year follow-up. The multivariate Cox regression analysis was adjusted for other important clinical and laboratory prognostic markers. Among all the oxidative stress markers examined in multivariate analysis, only MDA and UA were found to be independent predictors of death and combined endpoint. Higher serum MDA concentration increased the risk of death by 103.0% (HR = 2.103; 95% CI (1.330-3.325)) and of combined endpoint occurrence by 100% (HR = 2.000; 95% CI (1.366-2.928)) per μmol/L. Baseline levels of MDA in the 4th quartile were associated with an increased risk of death with a relative risk (RR) of 3.64 (95% CI (1.917 to 6.926), p < 0.001) and RR of 2.71 (95% CI (1.551 to 4.739), p < 0.001) for the occurrence of combined endpoint as compared to levels of MDA in the 1st quartile. Higher serum UA concentration increased the risk of death by 2.1% (HR = 1.021; 95% CI (1.005-1.038), p < 0.001) and increased combined endpoint occurrence by 1.4% (HR = 1.014; 95% CI (1.005-1.028), p < 0.001), for every 10 μmol/L. Baseline levels of UA in the 4th quartile were associated with an increased risk for death with a RR of 3.21 (95% CI (1.734 to 5.931)) and RR of 2.73 (95% CI (1.560 to 4.766)) for the occurrence of combined endpoint as compared to the levels of UA in the 1st quartile. In patients with chronic HF, increased MDA and UA concentrations were independently related to poor prognosis in a 1-year follow-up.

1892P - Enzalutamide (ENZA) and apalutamide (APA) In vitro chemical reactivity studies and activity in a mouse drug allergy model (MDAM)

BackgroundENZA and APA are androgen receptor inhibitors approved for treatment of castration-resistant prostate cancer (CRPC) and nonmetastatic CRPC, respectively. Their chemical structures differ with ENZA having 2-cyanophenyl and dimethyl moieties and APA 2-cyanopyridyl and cyclobutyl moieties. In the SPARTAN clinical trial, APA treatment was reported to be associated with an increased incidence of skin rash compared with the placebo group, which was not observed with ENZA in the PROSPER trial in a similar patient population; therefore, we examined their in vitro chemical reactivity and hypersensitivity potential in an MDAM. MethodsTo assess possible reactivity of cyanopyridine, in vitro studies examining the chemical stability in buffer (+/- glutathione) and covalent binding to proteins (eg, bovine serum albumin [BSA]) were evaluated. For the MDAM, C57BL/6 mice were injected subcutaneously with APA, ENZA, or RD162 (Analogue 1), daily for 3 days at 25, 50, or 100mg/kg/day. Brachial lymph nodes were harvested on day 6 and lymph node cellularity was analyzed by flow cytometry. ResultsThe 2-cyanopyridine moiety of APA was found to be chemically reactive with the thiol nucleophile glutathione, resulting in rearranged thiazoline products. Radiolabeled APA, but not ENZA, was shown to react with mouse and human plasma proteins. Thiol nucleophiles decreased the extent of covalent binding to the model protein BSA while amine and alcohol nucleophiles had no effect, suggesting a reaction with cysteine sulfhydryl groups of proteins. In the MDAM, dose-dependent higher lymph node cellularity was observed for the APA groups (compared with the vehicle control), suggesting immune activation. ENZA and Analogue 1 demonstrated substantially less covalent binding activity and no change in lymph node cellularity (compared with vehicle) in the MDAM. ConclusionsThese data support the hypothesis that the 2-cyanopyridine moiety present in APA, but absent in ENZA or Analogue 1, may react with cysteine residues in proteins to form haptens that may trigger an immune response leading to the increased incidence of skin rash clinically observed in patients treated with APA, but not with ENZA. Editorial acknowledgementMedical writing and editorial assistance funded by Pfizer Inc. was provided by Ira Mills, PhD, and Michele Salernitano from Ashfield Healthcare Communications. Legal entity responsible for the studyPfizer Inc. and Astellas Pharma, Inc. FundingPfizer Inc. and Astellas Pharma, Inc. DisclosureM. Guha: Shareholder / Stockholder / Stock options, Employee: Pfizer Inc.. C. Ji: Shareholder / Stockholder / Stock options, Employee: Pfizer Inc.. X. Zhu: Shareholder / Stockholder / Stock options, Employee: Pfizer Inc.. J. Whritenour: Shareholder / Stockholder / Stock options, Employee: Pfizer Inc.. M. Hemkens: Shareholder / Stockholder / Stock options, Employee: Pfizer Inc.. S. Tse: Shareholder / Stockholder / Stock options, Employee: Pfizer Inc.. G.S. Walker: Shareholder / Stockholder / Stock options, Employee: Pfizer Inc.. E. Evans: Shareholder / Stockholder / Stock options, Employee: Pfizer Inc.. N.K. Khan: Shareholder / Stockholder / Stock options, Employee: Pfizer Inc.. M.B. Finkelstein: Shareholder / Stockholder / Stock options, Employee: Pfizer Inc.. E. Callegari: Shareholder / Stockholder / Stock options, Employee: Pfizer Inc.. R..S. Obach: Shareholder / Stockholder / Stock options, Employee: Pfizer Inc..

Comparison of Oxidative Stress Parameters in Heart Failure Patients Depending on Ischaemic or Nonischaemic Aetiology.

Background Abnormalities in the oxidative and antioxidant states causing oxidative stress were both found in heart failure (HF) of various aetiologies and atherosclerosis. Aim of Study The goals of the study were as follows: comparison of oxidative stress parameters (OSP) in ischaemic cardiomyopathy (ICM) (n = 479) and nonischaemic cardiomyopathy (nICM) (n = 295) patients; assessment of the relationships of OSP with functional capacity (NYHA class), maximal oxygen consumption (max.O2), left ventricle ejection fraction (LVEF), and NT-proBNP concentration; and determination of the mutual relations of OSP in subgroups of patients with ICM and n-ICM. Methods Serum concentrations of total antioxidant capacity (TAC), total oxidant status (TOS), uric acid (UA), bilirubin, albumin, protein sulfhydryl groups (PSH), and malondialdehyde (MDA) were measured. The oxidative stress index (OSI) and MDA/PSH ratio were calculated. Results Higher concentrations of TAC (1.14 vs 1.11 mmol/l; p < 0.001) and MDA (1.80 vs 1.70 μmol/l; p < 0.05) and higher MDA/PSH ratios (0.435 vs 0.358; p < 0,001) were observed in ICM than in nICM patients. Simultaneously, lower values of the OSI index (4.27 vs 4.6; p < 0, 05), PSH (4.10 vs 4.75 μmol/g of protein; p < 0,001), and bilirubin (12.70 vs 15.40 μmol/l; p < 0,001) concentrations were indicated in ICM patients. There were no differences in TOS, UA, and albumin between the examined groups. The NYHA class and VO2max correlate with MDA, bilirubin, and albumin in both groups, while with UA only in the ICM group. Correlations between the NYHA class, VO2max, and PSH were indicated in nICM. The association of LVEF with UA, bilirubin, and albumin has been demonstrated in the ICM group. The study showed negative correlations between TAC, MDA, and PSH and positive between TAC and MDA in both groups. In ICM patients, MDA positively correlated with UA. A negative correlation between PSH and concentrations of UA and bilirubin was expressed only in the nICM group. Conclusion The obtained results confirm the relationship between the severity of HF and oxidative stress. The mechanisms of oxidative stress and antioxidant defence are partially different in the ICM and the nICM patients.

Is oxidative stress associated with disease severity, pulmonary function and metabolic syndrome in chronic obstructive pulmonary disease?

ObjectiveTo investigate associations between oxidant/antioxidant biomarkers with the disease severity, pulmonary function and diagnosis of metabolic syndrome (MetS) in patients with COPD. MethodsSeventy-four subjects were included, 39 with COPD (age 69±7 years; female 41%) and 35 for control group (age 69±7 years; female 43%). They were diagnosed with MetS and allocated in one of 4 subgroups: COPD and control, with and without MetS, respectively. Advanced oxidation protein products (AOPP), paraoxonase-1, catalase activity, sulfhydryl group and total lipid hydroperoxide were assayed. Pulmonary function was performed with a plethysmograph. ResultsCOPD severity (GOLD≥3) and pulmonary function were associated with sulfhydryl group and AOPP (p≤.03 for all). The prevalence of MetS was associated with AOPP in COPD (p=.04). Individuals with COPD and MetS showed higher AOPP compared to COPD without MetS (p<.0001). ConclusionCOPD severity, worse pulmonary function and presence of metabolic syndrome are associated with oxidative stress in individuals with COPD.

¿Está el estrés oxidativo asociado a la gravedad de la enfermedad, a la función pulmonar y al síndrome metabólico en la enfermedad pulmonar obstructiva crónica?

ObjectiveTo investigate associations between oxidant/antioxidant biomarkers with the disease severity, pulmonary function and diagnosis of metabolic syndrome (MetS) in patients with COPD. MethodsSeventy-four subjects were included, 39 with COPD (age 69±7 years; female 41%) and 35 for control group (age 69±7 years; female 43%). They were diagnosed with MetS and allocated in one of 4 subgroups: COPD and control, with and without MetS, respectively. Advanced oxidation protein products (AOPP), paraoxonase-1, catalase activity, sulfhydryl group and total lipid hydroperoxide were assayed. Pulmonary function was performed with a plethysmograph. ResultsCOPD severity (GOLD≥3) and pulmonary function were associated with sulfhydryl group and AOPP (P≤.03 for all). The prevalence of MetS was associated with AOPP in COPD (P=.04). Individuals with COPD and MetS showed higher AOPP compared to COPD without MetS (P<.0001). ConclusionCOPD severity, worse pulmonary function and presence of metabolic syndrome are associated with oxidative stress in individuals with COPD.

Thallium(I) treatment induces nucleolar stress to stop protein synthesis and cell growth

Thallium is considered as an emergent contaminant owing to its potential use in the superconductor alloys. The monovalent thallium, Tl(I), is highly toxic to the animals as it can affect numerous metabolic processes. Here we observed that Tl(I) decreased protein synthesis and phosphorylated eukaryotic initiation factor 2α. Although Tl(I) has been shown to interact with the sulfhydryl groups of proteins and cause the accumulation of reactive oxygen species, it did not activate endoplasmic reticulum stress. Notably, the level of 60S ribosomal subunit showed significant under-accumulation after the Tl(I) treatment. Given that Tl(I) shares similarities with potassium in terms of the ionic charge and atomic radius, we proposed that Tl(I) occupies certain K+-binding sites and inactivates the ribosomal function. However, we observed neither activation of ribophagy nor acceleration of the proteasomal degradation of 60S subunits. On the contrary, the ribosome synthesis pathway was severely blocked, i.e., the impairment of rRNA processing, deformed nucleoli, and accumulation of 60S subunits in the nucleus were observed. Although p53 remained inactivated, the decreased c-Myc and increased p21 levels indicated the activation of nucleolar stress. Therefore, we proposed that Tl(I) interfered the ribosome synthesis, thus resulting in cell growth inhibition and lethality.

Effects of cold atmospheric plasma on squid proteases and gel properties of protein concentrate from squid (Argentinus ilex) mantle

The effect of cold atmospheric plasma (CAP) on protein concentrate from squid (Argentinus ilex) mantle has been evaluated in terms of gel properties, protease inhibition, texture profile, color attributes, and water holding capacity. Different exposure time (15, 60, 120, 180, 240 and 300 s) at 60 kV have been employed. Our results indicated that protease activity decreases with increasing treatment time. The highest reduction (p < 0.05) in protease activity (64%) was observed after 240 s of CAP treatment. Texture profile analysis, color properties and water holding capacity of the treated squid gel revealed a significant increase. Protein carbonyl and sulfhydryl group contents findings' showed a significant increase in carbonyl content (about three times of the original content), while the total sulfhydryl group decreased (up to about 40%) in the crude extract. Microstructure and SDS-PAGE analysis revealed a high degree of protein aggregation in the squid gel treated with CAP.

Oxidative Stress of Carbon Nanotubes on Proteins Is Mediated by Metals Originating from the Catalyst Remains.

Nanomaterials introduced into biological systems are immediately coated by proteins in vivo. They induce oxidative stress on adsorbed proteins and hence alter the protein structures, which determines the fate pathways and biological impacts of nanomaterials. Carbon nanotubes (CNTs) have been suggested to cause protein oxidation. In this work, we discovered that CNTs induce oxidative stress on proteins in cooperation with coexisting metals originating from catalyst remains. Protein sulfhydryl groups were readily oxidized by the coexistence of CNTs and metals. Numerical simulations of the reaction demonstrated that the metals effectively mediate electron transfer between the CNTs and protein sulfhydryl groups. Thus, the coexistence of CNTs and metals, even in low concentrations, generates oxidative stress on proteins with high reaction rates. Metal catalysts used for CNT growth, in turn, catalyze the oxidation reaction of proteins. The proposed protein oxidation mechanism will advance the fundamental understanding of the biological safety and toxicity of nanomaterials synthesized using metal catalysts.

Crystallographic identification of spontaneous oxidation intermediates and products of protein sulfhydryl groups.

In the absence of protective reducing agents, Cys residues in purified proteins can be oxidized spontaneously by oxygen in the air, as frequently observed in protein crystal structures. However, the formation of an O-bridge via dehydration mechanism between a peroxidized Cys side chain and a primary amine of Lys side chain in proteins has not yet been reported. When an electron density feature was observed for an extra group or an extra atom between side chains of Cys-245 and Lys-158 in the crystal structure of histidinol phosphate phosphatase, mass spectrometric analysis was carried out for its chemical identification. That analysis led to a conclusion that this extra density corresponded to a methylene group. It was then proposed that these two residues were able to absorb CO2 and reduced it to CH2 spontaneously. Further examination of other protein structures in the PDB showed that the formation of this cross-linking species was a widespread phenomenon. This claim is examined in this study using methods recently developed for quantification of electrons around nucleus as the means for direct chemical identification. It is found that an O-bridge is actually formed between Cys and Lys side chains, instead of a CH2 -bridge.

Acetaminophen hepatotoxicity: A mitochondrial perspective.

Acetaminophen (APAP) is a highly effective analgesic, which is safe at therapeutic doses. However, an overdose can cause hepatotoxicity and even liver failure. APAP toxicity is currently the most common cause of acute liver failure in the United States. Decades of research on mechanisms of liver injury have established the role of mitochondria as central players in APAP-induced hepatocyte necrosis and this chapter examines the various facets of the organelle's involvement in the process of injury as well as in resolution of damage. The injury process is initiated by formation of a reactive metabolite, which binds to sulfhydryl groups of cellular proteins including mitochondrial proteins. This inhibits the electron transport chain and leads to formation of reactive oxygen species, which induce the activation of redox-sensitive members of the MAP kinase family ultimately causing activation of c-Jun N terminal kinase, JNK. Translocation of JNK to the mitochondria then amplifies mitochondrial dysfunction, ultimately resulting in mitochondrial permeability transition and release of mitochondrial intermembrane proteins, which trigger nuclear DNA fragmentation. Together, these events result in hepatocyte necrosis, while adaptive mechanisms such as mitophagy remove damaged mitochondria and minimize the extent of the injury. This oscillation between recovery and necrosis is predominant in cells at the edge of the necrotic area in the liver, where induction of mitochondrial biogenesis is important for liver regeneration. All these aspects of mitochondria in APAP hepatotoxicity, as well as their relevance to humans with APAP overdose and development of therapeutic approaches will be examined in detail in this chapter.

Protein structure and sulfhydryl group changes affected by protein gel properties: process of thermal-induced gel formation of myofibrillar protein

ABSTRACT The study investigated the change in protein structure upon thermo-induced gelation of myofibrillar protein (MP, 2%) under different temperatures (30–80°C) at 0.6 M NaCl and pH 6.2. The results showed that heating from 45°C led to changes in physicochemical properties and intermolecular forces involved during MP gelation, which are related to each other. During the heating process, the free sulfhydryl content slowly increased and reached the highest value at 45°C (10.23 ± 0.13 μmol/g protein). When the temperature exceeded 50°C, Ellman’s titration and SDS-PAGE results indicated that MP gel began to form with the formation of disulfide bonds, which contributed to stabilize the gel structure. Gel strength of the heat-induced MP gel formed at 60°C was significantly higher than that at other temperatures (P < .05), reaching 1.12 ± 0.02 N. Strictly speaking, myofibrillar protein gelatinized after 60°C, and water-holding capacity of the MP gel decreased from 60-80°C, and reached a relatively low value at 80°C. Furthermore, Raman spectra showed that the α-helix content decreased with increasing temperature, β-sheets and random coil contents increased, and the content of the twist-twist-trans conformation increased steadily.