Redox Enzymes

Laccases are valuable enzymes as an excellent ecological alternative for bioremediation issues because they can oxidize persistent xenobiotic compounds. The production and characterization of extracellular laccases from saprotrophic fungi from disturbed environments have been scarcely explored, even though this could diversify their functional characteristics and expand the conditions in which they carry out their catalysis. Agrocybe pediades, isolated from a disturbed forest, produces an extracellular laccase in liquid culture. The enzyme was purified, identified and characterized. Copper and hexachlorobenzene do not function as inducers for the laccase produced. Partial amino acid sequences were obtained by LC-MS/MS that share similarity with laccases from other fungi. Purified laccase is a monomer with a molecular mass between 55-60 kDa and had an optimum activity at pH 5.0 and the optimum temperature at 45 • C using 2,6-dimethoxyphenol (2,6-DMP) as substrate. The K m and V max also determined with 2,6-DMP were 100 µM and 285 µmol•min -1 •mg -1 , respectively, showing that the laccase of A. pediades has a higher affinity for this substrate than that of other Agaricales. These features could provide a potential catalyst for different toxic substrates and in the future laccase could be used in environmental recovery processes.

ZnO NPs (zinc oxide nanoparticles) has generated significant scientific interest as a novel antibacterial and anticancer agent. Since oxidative stress is a critical determinant of ZnO NPs-induced damage, it is necessary to characterize their underlying mode of action. Different structural and physicochemical properties of ZnO NPs such as particle surface, size, shape, crystal structure, chemical position, and presence of metals can lead to changes in biological activities including ROS (reactive oxygen species) production. However, there are some inconsistencies in the literature on the relation between the physicochemical features of ZnO NPs and their plausible oxidative stress mechanism. Herein, the possible oxidative stress mechanism of ZnO NPs was reviewed. This is worthy of further detailed evaluations in order to improve our understanding of vital NPs characteristics governing their toxicity. Therefore, this study focuses on the different reported oxidative stress paradigms induced by ZnO NPs including ROS generated by NPs, oxidative stress due to the NPs-cell interaction, and role of the particle dissolution in the oxidative damage. Also, this study tries to characterize and understand the multiple pathways involved in oxidative stress induced by ZnO NPs. Knowledge about different cellular signaling cascades stimulated by ZnO NPs lead to the better interpretation of the toxic influences induced by the cellular and acellular parameters. Regarding the potential benefits of toxic effects of ZnO NPs, in-depth evaluation of their toxicity mechanism and various effects of these nanoparticles would facilitate their implementation for biomedical applications.

β-Ionone, an end-ring analogue of β-carotenoid, which is a constituent of vegetables and fruits, has been analyzed for colon cancer chemoprevention and treatment. β-Ionone induced cell growth inhibition and apoptosis in human colon cancer HCT116 cell line. We tested the in vivo chemopreventive efficacy in rat colon carcinogenesis model using aberrant crypt foci (ACF) as endpoint marker. HCT116 cells treated with subtoxic concentrations of β-ionone resulted dose-dependent cell growth suppression with G1-S-phase growth arrest and significant induction of apoptosis. β-Ionone up-regulated expression of retinoid X receptor-α mRNA dose-dependently in HCT116 cells. To evaluate inhibitory properties of β-ionone on colonic ACF, 7-week-old male F344 rats were fed experimental diets containing 0%, 0.1%, or 0.2% β-ionone. After 1 week, rats received s.c. injections of azoxymethane, 15 mg/kg body weight, once weekly for 2 weeks. Rats were continued on respective experimental diets and sacrificed...

Interdisciplinary research has combined the efforts of many scientists and engineers to gain an understanding of biotic and abiotic electrochemical processes, materials properties, biomedical, and engineering approaches for the development of alternative power‐generating and/or energy‐harvesting devices, aiming to solve health‐related issues and to improve the quality of human life. This review intends to recapitulate the principles of biofuel cell development and the progress over the years, thanks to the contribution of cross‐disciplinary researchers that have combined knowledge and innovative ideas to the field. The emergence of biofuel cells, as a response to the demand of electrical power devices that can operate under physiological conditions, are reviewed. Implantable biofuel cells operating inside living organisms have been envisioned for over fifty years, but few reports of implanted devices have existed up until very recently. The very first report of an implanted biofuel ...

Maintenance of the cellular redox balance has vital importance for correcting organism functioning. Methionine sulfoxide reductases (Msrs) are among the key members of the cellular antioxidant defence system. To work properly, methionine sulfoxide reductases need to be reduced by their biological partner, thioredoxin (Trx). This process, according to the available kinetic data, represents the slowest step in the Msrs catalytic cycle. In the present paper, we investigated structural aspects of the intermolecular complex formation between mammalian MsrB1 and Trx. NMR spectroscopy and biocomputing were the two mostly used through the research approaches. The formation of NMR detectable MsrB1/Trx complex was monitored and studied in attempt to understand MsrB1 reduction mechanism. Using NMR data, molecular mechanics, protein docking, and molecular dynamics simulations, it was found that intermediate MsrB1/Trx complex is stabilized by interprotein β-layer. The complex formation accompanied by distortion of disulfide bond within MsrB1 facilitates the reduction of oxidized MsrB1 as it is evidenced by the obtained data.

Ferritins are ubiquitous and conserved proteins endowed with enzymatic ferroxidase activity, that oxidize Fe(II) ions at the dimetal ferroxidase centre to form a mineralized Fe(III) oxide core deposited within the apo-protein shell. Herein, the in vitro formation of a heterodimetal cofactor constituted by Fe and Mn ions has been investigated in human H ferritin (hHFt). Namely, Mn and Fe binding at the hHFt ferroxidase centre and its effects on Fe(II) oxidation have been investigated by UV-Vis ferroxidation kinetics, fluorimetric titrations, multifrequency EPR, and preliminary Mössbauer spectroscopy. Our results show that in hHFt, both Fe(II) and Mn(II) bind the ferroxidase centre forming a Fe-Mn cofactor. Moreover, molecular oxygen seems to favour Mn(II) binding and increases the ferroxidation activity of the Mn-loaded protein. The data suggest that Mn influences the Fe binding and the efficiency of the ferroxidation reaction. The higher efficiency of the Mn-Fe heterometallic centre...

Abstract: Owing to their radical scavenging and UV-filtering properties, ceria nanoparticles (CeO2-NPs) are currently used for various applications, including as catalysts in diesel particulate filters. Because of their ability to filter UV light, CeO2-NPs have garnered significant interest in the medical field and, consequently, are poised for use in various applications. The aim of this work was to investigate the effects of short-term (24 h) and long-term (10 days) CeO2-NP exposure to A549, CaCo2 and HepG2 cell lines. Cytotoxicity assays tested CeO2-NPs over a concentration range of 0.5 μg/mL to 5000 μg/mL, whereas genotoxicity assays tested CeO2-NPs over a concentration range of 0.5 μg/mL to 5000 μg/mL. In vitro assays showed almost no short-term exposure toxicity on any of the tested cell lines. Conversely, long-term CeO2-NP exposure proved toxic for all tested cell lines. NP genotoxicity was detectable even at 24-h exposure. HepG2 was the most sensitive cell line overall; howe...

The inefficiency of cyanide/HCN (CN) binding with heme proteins (under physiological regimes) is demonstrated with an assessment of thermodynamics, kinetics, and inhibition constants. The acute onset of toxicity and CN’s mg/Kg LD50(μM lethal concentration) suggests that the classical hemeFe binding-based inhibition rationale is untenable to account for the toxicity of CN. In vitro mechanistic probing of CN-mediated inhibition of hemeFe reductionist systems was explored as a murburn model for mitochondrial oxidative phosphorylation (mOxPhos). The effect of CN in haloperoxidase catalyzed chlorine moiety transfer to small organics was considered as an analogous probe for phosphate group transfer in mOxPhos. Similarly, inclusion of CN in peroxidase-catalase mediated one-electron oxidation of small organics was used to explore electron transfer outcomes in mOxPhos, leading to water formation. The free energy correlations from a Hammett study and IC50/Hill slopes analyses and comparison w...

Flavin-binding Kelch repeat F-box (FKF1) protein plays important roles in the photoregulation of flowering in Arabidopsis. FKF1 has a light, oxygen, and voltage (LOV) sensing domain binding a flavin mononucleotide (FMN) as a chromophore noncovalently. Photoreaction of the FKF1-LOV polypeptide was studied by low-temperature absorption spectroscopy. Upon blue light irradiation, a ground state, D 450 , is converted to S 390 known as a cysteinyl-flavin adduct intermediate in the photoreaction of phototropin. Below 150 K, bleaching of D 450 was much reduced and a new photoproduct, Z 370 , appeared as well as S 390 formation. The calculated absorption spectrum for Z 370 is very similar to those of flavoproteins in an anion radical state. On the basis of the results that S 390 formation proceeds to Z 370 formation and that Z 370 formed at low temperatures reverts to D 450 upon temperature increase, Z 370 is concluded to be not an intermediate from D 450 to S 390. Z 370 is suggested to be formed from the biradical triplet-excited state after relaxing to the ground state with the FMN anion radical trapped at the low temperature, in which the SH of the cysteine is in the wrong position that is able to produce a radical pair but unable to form the cysteinyl-flavin adduct. The counter SH in the cationic radical state may revert to the ground state by extracting an electron from the unidentified amino acid residue. Interestingly, S 390 that has been thought to be irreversible to D 450 was revealed to revert to D 450 very slowly with a half-life time of 62.5 h in solution at 298 K. The photoreaction mechanism is discussed in reference to the calculated activation energy of the reaction processes.

Background: Titanium dioxide (TiO 2) nanoparticles (NPs) are among the most important and usable photocatalysts. Recently, the biological properties of these NPs, particularly, its anticancer activity, have been considered. Glioblastoma and neuroblastoma are two fatal brain tumors with a high mortality rate in humans, the hope for treatment of which is weak by the common methods. Methods: In this study, the cytotoxicity effects of TiO 2 alone and in combination with ultraviolet A (UVA) irradiation on two different cell lines, neuroblastoma (SH-SY5Y) and glioblastoma U87, were investigated. After administration of 10, 50, 100, and 500 μg/mL TiO 2 , 0.043 and 1.4 mW/cm 2 UVA irradiation, cell viability was investigated after 4, 24, and 48 hours by 3-(4,5-dimethylthiazol-2-yl)-2,5diphenyltetrazolium bromide (MTT) assay. Results: MTT assay and light microscope demonstrated that the effect of TiO 2 NPs varied based on the dose of the substance, the impact time, the cell type, and the amount of radiation. In this study, for NPs alone, both toxicity and non-toxicity of the substance were observed. For NPs in the presence of UV, based on the comparison with its status alone and the difference in the viability assay of the two groups, both the photocatalytic and the coating effect of the NPs were observed. Conclusion: According to the results, different concentrations of TiO 2 can be used for different purposes. Low concentrations of TiO 2 can be used to increase the efficiency of photodynamic therapy and high concentrations of TiO 2 can be used to protect the normal cell. This strategy improves the photodynamic therapy and reduces the harmful effects.

The cytochromes P-450 (P-450s) constitute an extremely large family ('superfamily ') of haemoproteins that catalyse the oxidation of a wide range of physiological and non-physiological compounds. A remarkable feature of the P-450s is the manipulation of the same basic structure and chemistry to achieve an enormous range of functions in organisms as diverse as bacteria and man. Indeed, the P-450s have been described as 'the most versatile biological catalyst known'. Much research is focussed on mammalian P-450s, with their roles in such processes as steroid transformations and the metabolism of carcinogens and other xenobiotics. However, our knowledge of the structure and function of the P-450s has been advanced by analysis of a limited number of its bacterial members, primarily P-450cam from Pseudomonas putida. Four P-450 structures have been solved to date, all of which are from bacterial sources. The aim of this review is to assess current knowledge of the many bacterial P-450s, with emphasis on their diverse biological roles and on the advances in our knowledge of this extremely important enzyme class, which have been made feasible through their study.

Current trends in the field of MXenes emphasize the importance of controlling their surface features for successful application in biotechnological areas. The ability to stabilize the surface properties of MXenes has been demonstrated here through surface charge engineering. It was thus determined how changing the surface charges of two-dimensional (2D) Ti3C2 MXene phase flakes using cationic polymeric poly-L-lysine (PLL) molecules affects the colloidal and biological properties of the resulting hybrid 2D nanomaterial. Electrostatic adsorption of PLL on the surface of delaminated 2D Ti3C2 flakes occurs efficiently, leads to changing an MXene’s negative surface charge toward a positive value, which can also be effectively managed through pH changes. Analysis of bioactive properties revealed additional antibacterial functionality of the developed 2D Ti3C2/PLL MXene flakes concerning Escherichia. coli Gram-negative bacteria cells. A reduction of two orders of magnitude of viable cells ...

Titanium dioxide nanoparticles are widely used in industry to produce a number of products (cosmetics, paints, foods). Tio2 nanoparticles form aggregates and their toxicity varies in function of their size. in this study, aggregate sizes were determined using dynamic light scattering (DlS). The stability of the suspension was evaluated by turbidimetry and the size by transmission electronic microscopy (TeM). in the present study the toxicity of Tio2 nanoparticles was evaluated in vitro on rat kidney proximal tubular cells (nrK-52e). Cells were exposed to Tio2 nanoparticles (20 μg/mL) for 24, 48, 72 and 96 hours. TiO2 nanoparticles induce the production of reactive oxygen species (roS). The relative number of mitosis decreased, while an increase of apoptotic cells was noted. The number of S-phase cells evidenced by BrdU immunoreactivity in cultures was significantly reduced by the exposure to the nanoparticles. These results attest to the toxic effect of the Tio2 nanoparticles on nrK-52e cells.

Silver nanoparticles are increasingly being used in a wide variety of ways that may lead to frequency of exposure for humans and the environment. Thus, it is necessary to understand the biological effect(s) of these nanoparticles. Previously, we showed that AgNPs activated the kynurenine pathway in rat brain independently of oxidative stress in rats. This present study is aimed at evaluating the effect of AgNPs on some selected cytokines, redox parameters, and on the kynurenine level in rats. Male Wistar rats (130-150 g) were divided into 4 groups. Rats were grouped into control, AgNPs only (50 mg/kg bw), coadministration of AgNPs (50 mg/kg bw), and dexamethasone (100 mg/kg bw) and dexamethasone only (100 mg/kg bw). Results indicated that AgNPs did not significantly elevate MDA levels in rat plasma and brain relative to the control group. AgNPs caused a significant alteration in the level of rat brain and plasma total protein concentration. Meanwhile, AgNPs led to an elevation in th...

The article clarifies role of the monooxygenase system in the development of acute pancreatitis. Authors determine cytochrome P-450 possesses unique properties that ensure the maintenance of chemical homeostasis in physiological and pathological states of the internal organism medium. The liver is the main organ supporting physico-chemical homeostasis, detoxification of endogenously formed and exogenously introduced xenobiotics into the body. But there is also extrahepatic metabolism of foreign compounds. The intestine plays an important role in this process. Endogenous intoxication syndrome disrupts metabolism and reduces the functional activity of natural detoxification systems, in particular by inhibiting the liver's monooxygenated detoxification system.

The article clarifies role of the monooxygenase system in the development of acute pancreatitis. Authors determine cytochrome P-450 possesses unique properties that ensure the maintenance of chemical homeostasis in physiological and pathological states of the internal organism medium. The liver is the main organ supporting physico-chemical homeostasis, detoxification of endogenously formed and exogenously introduced xenobiotics into the body. But there is also extrahepatic metabolism of foreign compounds. The intestine plays an important role in this process. Endogenous intoxication syndrome disrupts metabolism and reduces the functional activity of natural detoxification systems, in particular by inhibiting the liver’s monooxygenated detoxification system.

Biosensors are the devices that capture the biological signal and convert it into a detectable electrical signal. It involves the combination of biological entities like DNA, RNA, and proteins/enzymes to the electrochemical transducers in order to detect and observe certain biological analytes like antibody-antigen interaction. Several types of biosensors have been known that have been successfully employed in the fields of environment, biomedical and food industries to detect and remove certain contaminants, weather non-living or living entities. Amperometric, Optical, Surface Plasmon Resonance, enzymatic, DNA, Phage, and bacterial sensors are the common sensors being employed today. These biosensors can be used for the detection of the broad spectrum of biological analytes and have shown greater responses and success in medical laboratories, food bioanalysis, microbial detection etc. Detection of the lower or higher limits of glucose in body, microbial invasion in body and food, heavy metals detection in soil, water and airborne microbes, pesticides in water and soil and various harmful chemicals produced by body, can be easily and timely monitored with high precision using the different types of biosensors with few modifications.

far the highest cytotoxicity, all the treatment regimens alter the characteristic morphology of the cells. Conclusion: Drugs loaded PHBHV NPs exert cytotoxic effects on cancer cells and could be further used for in vivo studies on animal models in order to study their distributions, mechanism of action and clearance.

From Bacillus subtilis cell extracts, ferredoxin-NADP + reductase (FNR) was purified to homogeneity and found to be the yumC gene product by N-terminal amino acid sequencing. YumC is a ~94 kDa homodimeric protein with one molecule of non-covalently bound FAD per subunit. In a diaphorase assay with 2,6-dichlorophenol-indophenol as an electron acceptor, the affinity to NADPH was much higher than to NADH, with K m values of 0.57 vs. >200 µM. K cat values of YumC with NADPH were 22.7 and 35.4 s-1 in diaphorase and in a ferredoxin-dependent NADPH-cytochrome c reduction assay, respectively. The cell extracts contained another diaphorase-active enzyme, the yfkO gene product, but its affinity for ferredoxin was very low. The deduced YumC amino acid sequence has high identity to that of the recently identified Chlorobium tepidum FNR. A genomic database search indicated that there are more than 20 genes encoding proteins that share a high level of amino acid sequence identity with YumC and annotated variously as NADH oxidase, thioredoxin reductase, thioredoxin reductase-like protein, etc. These genes are found notably in Gram-positive bacteria except for Clostridia, and less frequently in archaea and proteobacteria. We propose that YumC and C. tepidum FNR constitute a new group of FNR which should be added to the already established plant type, bacteria type, and mitochondria type FNR groups.

Colorectal cancer is a sporadic, hereditary, or familial based disease in its origin, caused due to diverse set of mutations in large intestinal epithelial cells. Colorectal cancer (CRC) is a common and deadly disease that accounts for the 4 th worldwide highly variable malignancy. For the early detection of CRC, the most common predictive biomarker found endogenously are KRAS and ctDNA/cfDNA along with SEPT9 methylated DNA. Early detection and screening for CRC are necessary and multiple methods can be employed to screen and perform early diagnosis of CRC. Colonoscopy, an invasive method is most prevalent for diagnosing CRC or confirming the positive result as compared to other screening methods whereas several non-invasive techniques such as molecular analysis of breath, urine, blood, and stool can also be performed for early detection. Interestingly, widely used medicines known as non-steroidal anti-inflammatory drugs (NSAIDs) to reduce pain and inflammation have reported chemopreventive impact on gastrointestinal malignancies, especially CRC in several epidemiological and preclinical types of research. NSAID acts by inhibiting two cyclooxygenase enzymes, thereby preventing the synthesis of prostaglandins (PGs) and causing NSAID-induced apoptosis and growth inhibition in CRC cells. This review paper majorly focuses on the diversity of natural and synthetic biomarkers and various techniques for the early detection of CRC. An approach toward current advancement in CRC detection techniques and the role of NSAIDs in CRC chemoprevention has been explored systematically. Several prominent governing mechanisms of the anticancer effects of NSAIDs and their synergistic effect with statins for an effective chemopreventive measure have also been discussed in this review paper.

Colorectal cancer is a sporadic, hereditary, or familial based disease in its origin, caused due to diverse set of mutations in large intestinal epithelial cells. Colorectal cancer (CRC) is a common and deadly disease that accounts for the 4th worldwide highly variable malignancy. For the early detection of CRC, the most common predictive biomarker found endogenously are KRAS and ctDNA/cfDNA along with SEPT9 methylated DNA. Early detection and screening for CRC are necessary and multiple methods can be employed to screen and perform early diagnosis of CRC. Colonoscopy, an invasive method is most prevalent for diagnosing CRC or confirming the positive result as compared to other screening methods whereas several non-invasive techniques such as molecular analysis of breath, urine, blood, and stool can also be performed for early detection. Interestingly, widely used medicines known as non-steroidal anti-inflammatory drugs (NSAIDs) to reduce pain and inflammation have reported chemopre...

Two laccase isoenzymes (LacA and LacB) were isolated from a novel Trichoderma harzianum S7113 isolate employing ammonium sulfate precipitation, Sephadex G100, and DEAE Sepharose ion exchange chromatography. The molecular weights of the purified LacA and LacB laccases were estimated to be 63 and 48 kDa, respectively. The two isoenzymes had their optimum activities at the same temperature (50 °C), but at slightly different pH values (pH 3.0 for LacA and pH 2.5 for LacB). LacA and LacB had the same thermal stability at 40 °C and pH stability at pH 9.0. The two isoenzymes also showed a high level of specific activity toward ABTS, where the K m values of LacA and LacB were 0.100 and 0.065 mM, whereas their V max values were 0.603 and 0.182 µmol min −1 , respectively. LacA and LacB catalytic activity was stimulated by Mg 2+ , Zn 2+ , K + , and Ni 2+ , whereas it was inhibited by Hg 2+ and Pb 2+ , β-mercaptoethanol, EDTA, and SDS, and completely inhibited by sodium azide. Our findings indicate that purified laccase has a promising capacity for bisphenol A (BPA) bioremediation across a broad pH range. This finding opens up new opportunities for the commercialization of this technique in a variety of biotechnology-based applications, particularly for removing endocrine chemicals from the environment.

Data collections provide a basis for solving numerous problems by data mining approach. The advantages of data mining consists in the retrieving of a new knowledge from existing information. The comprehensiveness of the data collection, the structure and quality of the data, and the selection of relevant data sets are extremely important to get correct results. In the crystallographic field, scientists will find several databases dealing with crystal structures of inorganic and organic compounds, or proteins. Usually databases have detailed data evaluation mechanisms integrated in their database production process and offer comprehensive and reliable data sets. The CIF standard enables the scientists to exchange the data. As an example, the Inorganic Crystal Structure Database (ICSD), a source of information for crystallographers, mineralogists, physicists, and chemists will be presented here. The ICSD contains about 120,000 entries (March 2009) of fully determined crystal structures. This chapter gives a detailed description of data collection, the contents of the data fields, data evaluation, and finally search the functionality of the ICSD database.

The interprotein electron transfer (ET) reactions of the cupredoxin amicyanin, which mediates ET from the tryptophan tryptophylquinone (TTQ) cofactor of methylamine dehydrogenase to cytochrome c-551i have been extensively studied. However, it was not possible to perform certain key experiments in that native system. This study examines the ET reaction from reduced amicyanin to an alternative electron acceptor, the diheme protein MauG. It was possible to vary the ΔG° for this ET reaction by simply changing pH to determine the dependence of kET on ΔG°. A P94A mutation of amicyanin significantly altered its oxidation-reduction midpoint potential value. It was not possible to study the ET from reduced P94A amicyanin to cytochrome c-551i in the native system because that reaction was kinetically coupled. However, the reaction from reduced P94A amicyanin to MauG was a true ET reaction and it was possible to determine values of reorganization energy (λ) and electronic coupling for the reac...

The electron transfer (ET) properties of two types of high-valent hemes were studied within the same protein matrix; the bis-Fe(IV) state of MauG and the Compound I state of Y294H MauG. The latter is formed as a consequence of mutation of the tyrosine which forms the distal axial ligand of the six-coordinate heme that allows it to stabilize Fe(IV) in the absence of an external ligand. The rates of the ET reaction of each high-valent species with the type I copper protein, amicyanin, were determined at different temperatures and analysed by ET theory. The reaction with bis-Fe(IV) wild-type (WT) MauG exhibited a reorganization energy (λ) that was 0.39 eV greater than that for the reaction of Compound I Y295H MauG. It is concluded that the delocalization of charge over the two hemes in the bis-Fe(IV) state is responsible for the larger λ, relative to the Compound I state in which the Fe(V) equivalent is isolated on one heme. Although the increase in λ decreases the rate of ET, the delo...

Electron transfer (ET) through and between proteins is a fundamental biological process. The rates and mechanisms of these ET reactions are controlled by the proteins in which the redox centers that donate and accept electrons reside. The protein influences the magnitudes of the ET parameters, the electronic coupling and reorganization energy that are associated with the ET reaction. The protein can regulate the rates of the ET reaction by requiring reaction steps to optimize the system for ET, leading to kinetic mechanisms of gated or coupled ET. Amino acid residues in the segment of the protein through which long range ET occurs can also modulate the ET rate by serving as staging points for hopping mechanisms of ET. Specific examples are presented to illustrate these mechanisms by which proteins control rates of ET reactions.

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The proton-pumping NADH:ubiquinone oxidoreductase, also called complex I, is the first energy-transducing complex of many respiratory chains. It couples the transfer of electrons from NADH to ubiquinone with the translocation of protons across the membrane. One FMN and up to nine ironsulfur (FeS) clusters participate in the redox reaction. So far, complex I has been described mainly by means of EPR-and UV-vis spectroscopy. Here, we report for the first time an infrared spectroscopic characterization of complex I. Electrochemically induced FT-IR difference spectra of complex I from Escherichia coli and of the NADH dehydrogenase fragment of this complex were obtained for critical potential steps. The spectral contributions of the FMN in both preparations were derived from a comparison using model compounds and turned out to be unexpectedly small. Furthermore, the FT-IR difference spectra reveal that the redox transitions of the FMN and of the FeS clusters induce strong reorganizations of the polypeptide backbone. Additional signals in the spectra of complex I reflect contributions induced by the redox transition of the high-potential FeS cluster N2 which is not present in the NADH dehydrogenase fragment. Part of these signals are attributed to the reorganization of protonated/deprotonated Asp or Glu side chains. On the basis of these data we discuss the role of N2 for proton translocation of complex I.

Many health related problems such as cardiovascular diseases are associated with the formation of excessive clot in the blood (thrombus). Approaches in cardiovascular disease treatment are preventing the formation or removing the thrombus. The present thrombolytic agents can be classified as plasminogen activators, fibrinolytic enzyme which directly degrades fibrinogen or fibrin and heparin type which act as thrombin inhibitor. Recently, microbial fibrinolytic enzymes of food origin receive more attention that leads to escalating efforts to explore traditional fermented foods as the natural sources. We have successfully isolated microorganism from Indonesian fermented soybean tofu dregs "Oncom" that secretes fibrinolytic enzyme. The microorganism identified as Stenotrophomonas sp. is unique because most of the reported fibrinolytic microorganism belongs to Bacillus sp. This isolate was found to produce extracellular fibrinolytic enzyme which could degrade fibrinogen and fibrin directly as determined by fibrinogen zymography and fibrin plate methods. More importantly, the 30-kD purified enzymes was found to demonstrate not only fibrin and fibrinogen degradation capabilities, but also acted as thrombin inhibitor as determined using specific substrates for thrombin. This is the first report of a fibrinolytic enzyme that demonstrates additional synergistic activities. This finding accentuates the importance of further development of the enzyme into a powerful agent to treat the thrombus-related disease effectively.

Cobalt oxide (Co 3 O 4) nanoparticles have been extensively used in various industrial and medical applications due to their special optical, magnetic, and electrical activity features. However, there is a lack of information about their toxicity and adverse effects on human health, especially concerning the kidney, which is considered to be a secondary target organ. We investigated the toxic potentials of Co 3 O 4 nanoparticles on NRK-52E kidney epithelial cells by in vitro assays. Co 3 O 4 nanoparticles were taken up by the kidney cells, and caused a decrease in cell viability, by significantly inducing apoptosis/ necrosis at 100 µg/mL. However, no significant DNA damage was observed. Co 3 O 4 nanoparticles induced cellular toxicity in kidney cells. These results should raise concern about the safety of Co 3 O 4 nanoparticles in their various applications. Further studies are needed to elucidate their toxic mechanism.

An electrochemical study was performed on the behavior of Ti 3 C 2 T x MXenes prepared by using either HF (MXene1) or LiF/HCl as etchants (MXene2). The use of two redox probes indicates the presence of a higher negative charge density on MXene2 in comparison to MXene1. The characterization of two nanomaterials shows that titanium and fluoride are present higher by one order of magnitude at the interface of MXene2, compared to MXene1. The high Ti and F content is accompanied by a 82-fold larger (249 μA•cm −2 vs. 5.64 μA•cm −2) anodic peak at the peak potential near 0.4 V (vs. Ag/AgCl). Similarly, the peak current on MXene2 is 317-fold higher for the oxygen reduction at pH 7.0 (at a voltage of −0.84 V) and 215-fold higher for the reduction of H 2 O 2 at −0.89 V, when compared to MXene1. Keywords MXene. Ti 3 C 2 T X. Electrocatalysis. Electrochemistry. H 2 O 2 reduction. Oxygenreduction reactions. Redox stability

Nanomaterials or nanoparticles are commonly used in the cosmetics, medicine, and food industries. Many researchers studied the possible side effects of several nanoparticles including aluminum oxide (Al2O3-nps) and zinc oxide nanoparticles (ZnO-nps). Although, there is limited information available on their direct or side effects, especially on the brain, heart, and lung functions. This study aimed to investigate the neurotoxicity, cardiotoxicity, and lung toxicity induced by Al2O3-nps and ZnO-nps or in combination via studying changes in gene expression, alteration in cytokine production, tumor suppressor protein p53, neurotransmitters, oxidative stress, and the histological and morphological changes. Obtained results showed that Al2O3-nps, ZnO-nps and their combination cause an increase in 8-hydroxy-2´-deoxyguanosine (8-OHdG), cytokines, p53, oxidative stress, creatine kinase, norepinephrine, acetylcholine (ACh), and lipid profile. Moreover, significant changes in the gene express...

Antioxidant nanoparticles have recently gained tremendous attention for their enormous potential in biomedicine. However, discrepant reports of either medical benefits or toxicity, and lack of reproducibility of many studies, generate uncertainties delaying their effective implementation. Herein, the case of cerium oxide is considered, a well‐known catalyst in the petrochemistry industry and one of the first antioxidant nanoparticles proposed for medicine. Like other nanoparticles, it is now described as a promising therapeutic alternative, now as threatening to health. Sources of these discrepancies and how this analysis helps to overcome contradictions found for other nanoparticles are summarized and discussed. For the context of this analysis, what has been reported in the liver is reviewed, where many diseases are related to oxidative stress. Since well‐dispersed nanoparticles passively accumulate in liver, it represents a major testing field for the study of new nanomedicines a...

The manufacture of personalized, real time, specific, and sensitive diagnostic tools continue to be of importance as the need for rapid diagnosis and monitoring of disease persists. In many health conditions like cancer and Covid-19 infection timely diagnosis of diseases can affect chances of survival. The accurate and timely diagnosis of diseases is enhanced by the unique optical, fluorescent, and magnetic properties of nanoscale elements. However, ultra-fine Nano particles unintentionally released into the air through human activities like combustion have been linked to respiratory and cardiovascular diseases. This makes it important for Nano engineered devices to be assessed for safety and toxicity concerns. Toxicity studies on the bulk material of the nanoscale elements used in diagnosis does not suffice as Nano materials due to their small size have unique interactions with biological membranes. Radioactive functionalized nanoparticles have been employed in several diagnostic i...

Ammonia (NH3) is a vital compound in diversified fields, including agriculture, automotive, chemical, food processing, hydrogen production and storage, and biomedical applications. Its extensive industrial use and emission have emerged hazardous to the ecosystem and have raised global public health concerns for monitoring NH3 emissions and implementing proper safety strategies. These facts created emergent demand for translational and sustainable approaches to design efficient, affordable, and high-performance compact NH3 sensors. Commercially available NH3 sensors possess three major bottlenecks: poor selectivity, low concentration detection, and room-temperature operation. State-of-the-art NH3 sensors are scaling up using advanced nano-systems possessing rapid, selective, efficient, and enhanced detection to overcome these challenges. MXene–polymer nanocomposites (MXP-NCs) are emerging as advanced nanomaterials of choice for NH3 sensing owing to their affordability, excellent cond...

The manufacture of personalized, real time, specific, and sensitive diagnostic tools continue to be of importance as the need for rapid diagnosis and monitoring of disease persists. In many health conditions like cancer and Covid-19 infection timely diagnosis of diseases can affect chances of survival. The accurate and timely diagnosis of diseases is enhanced by the unique optical, fluorescent, and magnetic properties of nanoscale elements. However, ultra-fine Nano particles unintentionally released into the air through human activities like combustion have been linked to respiratory and cardiovascular diseases. This makes it important for Nano engineered devices to be assessed for safety and toxicity concerns. Toxicity studies on the bulk material of the nanoscale elements used in diagnosis does not suffice as Nano materials due to their small size have unique interactions with biological membranes. Radioactive functionalized nanoparticles have been employed in several diagnostic i...

far the highest cytotoxicity, all the treatment regimens alter the characteristic morphology of the cells. Conclusion: Drugs loaded PHBHV NPs exert cytotoxic effects on cancer cells and could be further used for in vivo studies on animal models in order to study their distributions, mechanism of action and clearance.

The main manuscript body consists of 5 Figures Supplementary material consists of 6 Figures and 2 videos Highlights  Sensors based on graphene oxide (GO) nanosheets were deposited on five different substrates  Highly oriented pyrolyric graphite, glass slides, silicon wafers, polydimethylsiloxane, and mica were used as substrates  High affinity of GO-modified surfaces enabled immobilization of isolated single cells.  Raman spectroscopy analysis of the chemical composition of individual yeast cells and cell colonies was realized

An electrochemical study was performed on the behavior of Ti 3 C 2 T x MXenes prepared by using either HF (MXene1) or LiF/HCl as etchants (MXene2). The use of two redox probes indicates the presence of a higher negative charge density on MXene2 in comparison to MXene1. The characterization of two nanomaterials shows that titanium and fluoride are present higher by one order of magnitude at the interface of MXene2, compared to MXene1. The high Ti and F content is accompanied by a 82-fold larger (249 μA•cm −2 vs. 5.64 μA•cm −2) anodic peak at the peak potential near 0.4 V (vs. Ag/AgCl). Similarly, the peak current on MXene2 is 317-fold higher for the oxygen reduction at pH 7.0 (at a voltage of −0.84 V) and 215-fold higher for the reduction of H 2 O 2 at −0.89 V, when compared to MXene1. Keywords MXene. Ti 3 C 2 T X. Electrocatalysis. Electrochemistry. H 2 O 2 reduction. Oxygenreduction reactions. Redox stability

Nanotechnology is considered a new technology playing an essential role in several science fields. Titanium dioxide nanoparticles (TiO2 NPs) are extensively utilized inseveral fields like electronics, medicine, agriculture, food additives, paint, and sunscreens. Oral exposure mainly occurs through food products containing TiO2 NPadditives. Therefore, exposure to TiO2 NPs orally may have some adverse effects on exposed mammals and thus might be in potential health risk. The current study aims to study the possibility of liver damage in male rats exposed to a single oral dose (250 mg/kg b.w.) of 25 ± 5 TiO2 NPs. The gene expression of oxidative stress genes (Gpx and Cu-Zn SOD), protein folding gene (Hsp70), apoptosis gene (p53), metal toxicity gene (Mt1) as target genes, and GAPDH as housekeeping genes were determined. The results indicated that significant over-expression of the oxidative stress, apoptosis and protein folding genes was observed in the liver at 7 days post-treatment while there were no significant changes at day 28 as compared with control. Meanwhile, expression of the metal toxicity gene was increased significantly at both 7 and 28 days after exposure.Based on the abovementioned observation, it should be considered to possible health risks associated with the consumption of food products containing high concentrations of TiO2 NP-additives.

Extracorporeal irradiation sterilizes resected tumor bone used as autografts in reconstruction surgery. Therapeutic irradiation is a standard technique in head and neck cancer therapy that aims to preserve organ function. Bone irradiation has a complex, mostly inhibitory, effect on remodeling and regeneration, although the underlying mechanisms are still not fully understood. It remains unclear if extracorporeal irradiation affects the paracrine-like activity of the corresponding autografts. We recently reported that bone-conditioned medium from autogenous bone chips contains a number of factors that might affect cell activity. In the present study, we investigated the effects of extracorporeal irradiation of porcine cortical bone chips on the activity of the corresponding bone-conditioned medium. The effects of bone-conditioned medium on the expressions of transforming growth factor-beta (TGF-β) target genes in oral fibroblasts were assessed. Bone-conditioned medium from bone chips exposed to a total radiation dose up to 120 Gy did not affect expressions of TGF-β target genes, including adrenomedullin, BTB/ POZ domain-containing protein 11, proteoglycan 4, NADPH oxidase 4, and interleukin 11, in oral fibroblasts. In conclusion, bone irradiation does not alter the capability of the corresponding bone-conditioned medium to provoke a robust fibroblastic cell response in vitro.

Light-oxygen-voltage (LOV) domains absorb blue light for mediating various biological responses in all three domains of life. Aureochromes from stramenopile algae represent a subfamily of photoreceptors that differs by its inversed topology with a C-terminal LOV sensor and an N-terminal effector (basic region leucine zipper, bZIP) domain. We crystallized the LOV domain including its flanking helices, A 0 a and Ja, of aureochrome 1a from Phaeodactylum tricornutum in the dark state and solved the structure at 2.8 Å resolution. Both flanking helices contribute to the interface of the native-like dimer. Small-angle X-ray scattering shows light-induced conformational changes limited to the dimeric envelope as well as increased flexibility in the lit state for the flanking helices. These rearrangements are considered to be crucial for the formation of the light-activated dimer. Finally, the LOV domain of the class 2 aureochrome PtAUREO2 was shown to lack a chromophore because of steric hindrance caused by M301.

While cytochrome c (Cyt. c) is a soluble protein involved in mitochondrial electron transfer (ET) reactions between Complex III and Complex IV, cytochrome b5 (Cyt. b5) is a microsomal membrane protein acting as a redox aide for diverse cytochrome P450s and their unique reductase. We found little conservation in the sequence and surface amino acid residues of Cyt. c and b5 proteins among evolutionarily diverse species. Hence, the logic that these proteins mediate ET through affinity binding via specific surface residues is weak. Also, analysis of putative protein-protein interactions in the crystal structures of these proteins and their redox partners did not point to any specific interaction logic. The literature on kinetic and thermodynamic constants of mutants (with altered surface residues) did not provide strong evidence to support the binding-based ET paradigm. Topographically divergent Cyt. b5 from one species has been shown to enhance the activity of CYP450 from another speci...

The inefficiency of cyanide/HCN (CN) binding with heme proteins (under physiological regimes) is demonstrated with an assessment of thermodynamics, kinetics, and inhibition constants. The acute onset of toxicity and CN’s mg/Kg LD50(μM lethal concentration) suggests that the classical hemeFe binding-based inhibition rationale is untenable to account for the toxicity of CN. In vitro mechanistic probing of CN-mediated inhibition of hemeFe reductionist systems was explored as a murburn model for mitochondrial oxidative phosphorylation (mOxPhos). The effect of CN in haloperoxidase catalyzed chlorine moiety transfer to small organics was considered as an analogous probe for phosphate group transfer in mOxPhos. Similarly, inclusion of CN in peroxidase-catalase mediated one-electron oxidation of small organics was used to explore electron transfer outcomes in mOxPhos, leading to water formation. The free energy correlations from a Hammett study and IC50/Hill slopes analyses and comparison w...

Recently, electron transfers and catalyses in a bevy of redox reactions mediated by hemeproteins were explained by murburn concept. The term "murburn" is abstracted from "" or "" and connotes a novel "" interaction paradigm. Quite unlike the genetic regulations and protein-level affinity-based controls that govern order and specificity/selectivity in conventional treatments, murburn concept is based on stochastic/thermodynamic regulatory principles. The novel insight necessitates a "reactivity outside the active-site" perspective, because select redox enzymatic activity is obligatorily mediated via diffusible radical/species. Herein, reactions employing key hemeproteins (as exemplified by CYP2E1) establish direct experimental connection between "additive-influenced redox catalysis" and "unusual dose responses" in reductionist and physiological milieu. Thus, direct and conclusive molecular-level experimental eviden...

Two laccase isoenzymes (LacA and LacB) were isolated from a novel Trichoderma harzianum S7113 isolate employing ammonium sulfate precipitation, Sephadex G100, and DEAE Sepharose ion exchange chromatography. The molecular weights of the purified LacA and LacB laccases were estimated to be 63 and 48 kDa, respectively. The two isoenzymes had their optimum activities at the same temperature (50 °C), but at slightly different pH values (pH 3.0 for LacA and pH 2.5 for LacB). LacA and LacB had the same thermal stability at 40 °C and pH stability at pH 9.0. The two isoenzymes also showed a high level of specific activity toward ABTS, where the K m values of LacA and LacB were 0.100 and 0.065 mM, whereas their V max values were 0.603 and 0.182 µmol min −1 , respectively. LacA and LacB catalytic activity was stimulated by Mg 2+ , Zn 2+ , K + , and Ni 2+ , whereas it was inhibited by Hg 2+ and Pb 2+ , β-mercaptoethanol, EDTA, and SDS, and completely inhibited by sodium azide. Our findings indicate that purified laccase has a promising capacity for bisphenol A (BPA) bioremediation across a broad pH range. This finding opens up new opportunities for the commercialization of this technique in a variety of biotechnology-based applications, particularly for removing endocrine chemicals from the environment.

Reduction of Complex I (NADH:ubiquinone oxidoreductase I) from Escherichia coli by NADH was investigated optically by means of an ultrafast stopped-flow approach. A locally designed microfluidic stopped-flow apparatus with a low volume (0.2 μl) but a long optical path (10 mm) cuvette allowed measurements in the time range from 270 μs to seconds. The data acquisition system collected spectra in the visible range every 50 μs. Analysis of the obtained time-resolved spectral changes upon the reaction of Complex I with NADH revealed three kinetic components with characteristic times of b 270 μs, 0.45-0.9 ms and 3-6 ms, reflecting reduction of different FeS clusters and FMN. The rate of the major (τ = 0.45-0.9 ms) component was slower than predicted by electron transfer theory for the reduction of all FeS clusters in the intraprotein redox chain. This delay of the reaction was explained by retention of NAD + in the catalytic site. The fast optical changes in the time range of 0.27-1.5 ms were not altered significantly in the presence of 10-fold excess of NAD + over NADH. The data obtained on the NuoF E95Q variant of Complex I shows that the single amino acid replacement in the catalytic site caused a strong decrease of NADH binding and/or the hydride transfer from bound NADH to FMN.

MXenes are recently developed 2D layered nanomaterials that provide unique capabilities for bioanalytical applications. These include high metallic conductivity, large surface area, hydrophilicity, high ion transport properties, low diffusion barrier, biocompatibility, and ease of surface functionalization. MXenes are composed of transition metal carbides, nitrides, or carbonitrides and have a general formula M n+1 X n , where M is an early transition metal while X is carbon and/or nitrogen. Due to their unique features, MXenes have attracted significant attention in fields such as clean energy production, electronics, fuel cells, supercapacitors, and catalysis. Their composition and layered structure make MXenes attractive for biosensing applications. The high conductivity allows these materials to be used in the design of electrochemical biosensors and the multilayered configuration makes them an efficient immobilization matrix for the retention of activity of the immobilized biomolecules. These properties are applicable to many biosensing systems and applications. This review describes the progress made on the use and application of MXenes in the development of electrochemical and optical biosensors and highlights future needs and opportunities in this field. In particular, opportunities for developing wearable sensors and systems with integrated biomolecule recognition are highlighted.