NADH Oxidase Research Articles

Assessing the risk factors for myocardial infarction in diet-induced prediabetes: myocardial tissue changes

BackgroundHyperglycaemia is known to result in oxidative stress tissue injury and dysfunction. Interestingly, studies have reported hepatic and renal oxidative stress injury during prediabetes; however, any injury to the myocardium during prediabetes has not been investigated. Hence this study aims to assess changes in the myocardial tissue in an HFHC diet-induced model of prediabetes.MethodsMale Sprague Dawley rats were randomly grouped into non-prediabetes and prediabetes (n = 6 in each group) and consumed a standard rat chow or fed a high-fat-high-carbohydrate diet respectively for a 20-week prediabetes induction period. Post induction, prediabetes was confirmed using the ADA criteria. Aldose reductase, NADH oxidase 1, superoxide dismutase, glutathione peroxide, cardiac troponins were analysed in cardiac tissue homogenate using specific ELISA kits. Lipid peroxidation was estimated by determining the concentration of malondialdehyde in the heart tissue homogenate according to the previously described protocol. Myocardial tissue sections were stained with H&E stain and analysed using Leica microsystem. All data were expressed as means ± SEM. Statistical comparisons were performed with Graph Pad instat Software using the Student's two-sided t-test. Pearson correlation coefficient was calculated to assess the association. Value of p < 0.05 was considered statistically significant.ResultsThe prediabetes group showed a markedly high oxidative stress as indicated by significantly increased NADH oxidase 1 and malondialdehyde while superoxide dismutase and glutathione peroxide were decreased compared to non-prediabetes group. There was no statistical difference between cardiac troponin I and T in the non-prediabetes and prediabetes groups. Cardiac troponins had a weak positive association with glycated haemoglobin.ConclusionThe findings of this study demonstrate that prediabetes is associated with myocardial injury through oxidative stress. Future studies are to investigate cardiac contractile function and include more cardiac biomarkers.

Altered mitochondria oscillations and circadian changes in NADH levels in freshwaters mussels exposed to cadmium

The effects of contaminants on biological rhythms are largely unknown. The purpose of this study was to examine the effects of cadmium (Cd) and time of day on levels of defence mechanism (metallothionein-MT), mitochondria NADH oxidase activity and oscillations in NADH levels in freshwater mussels Dreissena bugensis. Following exposure of mussels to 50 and 100 μg/L Cd for 7 days, mussels were sampled at various times of the day (9, 12, 15, 18, 21 and 24 h) and immediately stored at −85 °C. Tissues were analyzed for MT levels, NADH oxidase and oscillatory changes in NADH in mitochondria assessments. The data revealed that exposure to Cd readily increased MT and NADH oxidase activity at each time of day. In control mussels, NADH oxidase activity changed during the day reaching a minimum and maximum at 15 h and 24 h respectively as with MT levels but the maximum occurred at 12 h. Exposure to Cd displaced the maximum values in NADH oxidase towards the night (≥18 h) while no change in the pattern in MT changes during the day albeit remained at high levels. NADH oscillations in mitochondria were complex but main changes occurred at periods of 60–90 s in control mussels. The strength of association (β) between amplitudes and frequencies of these oscillations was negatively associated with MT levels suggesting that activation of defence mechanism limited the propagation of amplitude changes at higher times. The study revealed that Cd exposure could disrupt not only the intensity of responses but at the levels of rhythms commonly found in biological systems.

Antifungal Activity of β-Carboline Alkaloids Compound and Its Resistance Mechanism on Peronophythora Litchii

ABSTRACT Peronophythora litchii (P. litchii) is the pathogenic factor of litchi downy blight. Derivatives extracted from harmine officinalis and modified, N-(2-pyridyl)-1-phenyl-9 H-pyrido [3, 4-b] indole-3-formamide (PPPIF), could inhibited P. litchii growth and development. Scanning Electron Microscopy (SEM) and Transmission Electron Microscopy (TEM) analyses indicated that PPPIF severely damaged the mitochondria, cell wall and endomembrane system of P. litchii cells, thus abnormal morphology, as well as deformed mycelia. PPPIF could suppress mycelial growth, sporulation, sporangia germination and germ tube elongation as well. PPPIF also caused a series of physiological and biochemical changes of P. litchii, including the serious deviation of extracellular pH, the obvious increase of the content of extracellular reducing sugar and malondialdehyde, the notable decrease of soluble protein and the activity of NADH oxidase. Furthermore, PPPIF seriously raised openness of membrane permeability transporter (MPTP) and markedly decreased the transmembrane potential (Δψm) and the activity of enzyme complex I–complex V in respiratory chain of mitochondria. The current research suggests that PPPIF may act upon the mitochondrion of P. litchii, then respiratory chain complex activity was blocked and energy metabolism disrupted or inhibited, resulting in the growth inhibition of P. litchii. Above results have strengthened our understanding of P. litchii resistance mechanisms and may help in the development of more potent inhibitors against plant diseases in the fields.

Inhibition of respiratory complex I by 6-ketocholestanol: Relevance to recoupling action in mitochondria

6-Ketocholestanol (kCh) is known as a mitochondrial recoupler, i.e. it abolishes uncoupling of mitochondria by such potent agents as carbonyl cyanide m-chlorophenyl hydrazone (CCCP) and 3,5-di(tert-butyl)-4-hydroxybenzylidenemalononitril (SF6847) [Starkov et al., 1997]. Here, we report data on the kCh-induced inhibition of both NADH-oxidase and NADH-ubiquinone oxidoreductase activities of the respiratory complex I in bovine heart submitochondrial particles (SMP). Based on the absence of such inhibition with hexaammineruthenium (III) (HAR) as the complex I electron acceptor, the kCh effect could be associated with the ubiquinone-binding centre of this respiratory enzyme. In isolated rat liver mitochondria (RLM), kCh inhibited oxygen consumption with the glutamate/malate, substrates of NAD-linked dehydrogenases, while no inhibition of RLM respiration was observed with succinate, in agreement with the absence of the kCh effect on the succinate oxidase activity in SMP. Three kCh analogs (cholesterol, 6α-hydroxycholesterol, and 5α,6α-epoxycholesterol) exhibited no effect on the NADH oxidase activities in both SMP and RLM. Importantly, the kCh analogs were ineffective in the recoupling of RLM treated with CCCP or SF6847. Therefore, interaction of kCh with the complex I may be involved in the kCh-mediated mitochondrial recoupling.

Characterization of four diol dehydrogenases for enantioselective synthesis of chiral vicinal diols

Enantiopure vicinal diols are important building blocks used in the synthesis of fine chemicals and pharmaceutical compounds. Diol dehydrogenase (DDH) mediated stereoselective oxidation of racemic vicinal is an efficient way to prepare enantiopure vicinal diols. In this study, four new bacterial DDHs (AnDDH from Anoxybacillus sp. P3H1B, HcDDH from Hazenella coriacea, GzDDH from Geobacillus zalihae and LwDDH from Leptotrichia wadei ) were mined from the GenBank database and expressed in E. coli T7. The four DDHs were purified and biochemically characterized for oxidation activity toward ( R )-1-phenyl-1,2-ethanediol, with the optimal reaction condition of pH9.0 (AnDDH), 10.0 (HcDDH) and 11.0 (GzDDH and LwDDH) and the temperatures at 40 °C (AnDDH), 50 °C (HcDDH) and 60 °C (GzDDH and LwDDH), respectively. The four enzymes were stable at the pH from 7.0 to 9.0 and below 40 °C. Kinetic parameters of four DDHs showed that the HcDDH from Hazenella coriacea had high activity toward a broad range of vicinal diols. A series of racemic vicinal diols were successfully resolved by recombinant E. coli (HcDDH-NOX) resting cells co-expression of an NADH oxidase (NOX), affording ( S )-diols and (1 S , 2 S )- trans -diols in ≥99% ee. The synthetic potential of HcDDH was proved by E. coli (HcDDH-NOX) via kinetic resolution of racemic trans -1,2-indandiol on a 100 ml scale reaction, ( S , S )- trans -1,2-indandiol was prepared in 46.7% yield and >99% ee. In addition, asymmetric reduction of four α-hydroxy ketones (10–300 mmol·L −1 ) by E. coli (HcDDH-GDH) resting cells resulted in >99% ee and 69–98% yields of ( R )-vicinal diols. The current research expands the toolbox of DDHs to synthesize chiral vicinal diols and demonstrated that the mined HcDDH is a potential enzyme in the synthesis of a broad range of chiral vicinal diols.

A natural fusion of flavodiiron, rubredoxin, and rubredoxin oxidoreductase domains is a self-sufficient water-forming oxidase of Trichomonas vaginalis

Microaerophilic pathogens such as Giardia lamblia, Entamoeba histolytica, and Trichomonas vaginalis have robust oxygen consumption systems to detoxify oxygen and maintain intracellular redox balance. This oxygen consumption results from H2O-forming NADH oxidase (NOX) activity of two distinct flavin-containing systems: H2O-forming NOXes and multicomponent flavodiiron proteins (FDPs). Neither system is membrane bound, and both recycle NADH into oxidized NAD+ while simultaneously removing O2 from the local environment. However, little is known about the specific contributions of these systems in T. vaginalis. In this study, we use bioinformatics and biochemical analyses to show that T. vaginalis lacks a NOX–like enzyme and instead harbors three paralogous genes (FDPF1–3), each encoding a natural fusion product between the N-terminal FDP, central rubredoxin (Rb), and C-terminal NADH:Rb oxidoreductase domains. Unlike a “stand-alone” FDP that lacks Rb and oxidoreductase domains, this natural fusion protein with fully populated flavin redox centers directly accepts reducing equivalents of NADH to catalyze the four-electron reduction of oxygen to water within a single polypeptide with an extremely high turnover. Furthermore, using single-particle cryo-EM, we present structural insights into the spatial organization of the FDP core within this multidomain fusion protein. Together, these results contribute to our understanding of systems that allow protozoan parasites to maintain optimal redox balance and survive transient exposure to oxic conditions.

Biocatalytic Production of a Nylon 6 Precursor from Caprolactone in Continuous Flow.

6‐Aminocaproic acid (6ACA) is a key building block and an attractive precursor of caprolactam, which is used to synthesize nylon 6, one of the most common polymers manufactured nowadays. (Bio)‐production of platform chemicals from renewable feedstocks is instrumental to tackle climate change and decrease fossil fuel dependence. Here, the cell‐free biosynthesis of 6ACA from 6‐hydroxycaproic acid was achieved using a co‐immobilized multienzyme system based on horse liver alcohol dehydrogenase, Halomonas elongata transaminase, and Lactobacillus pentosus NADH oxidase for in‐situ cofactor recycling, with >90 % molar conversion (m.c.) The integration of a step to synthesize hydroxy‐acid from lactone by immobilized Candida antarctica lipase B resulted in >80 % m.c. of ϵ‐caprolactone to 6ACA, >20 % of δ‐valerolactone to 5‐aminovaleric acid, and 30 % of γ‐butyrolactone to γ‐aminobutyric acid in one‐pot batch reactions. Two serial packed‐bed reactors were set up using these biocatalysts and applied to the continuous‐flow synthesis of 6ACA from ϵ‐caprolactone, achieving a space‐time yield of up to 3.31 g6ACA h−1 L−1 with a segmented liquid/air flow for constant oxygen supply.

Ginkgo biloba Extract Attenuates Light-Induced Photoreceptor Degeneration by Modulating CAV-1-Redoxosome Signaling.

The clinical potential of Ginkgo biloba extract (GBE) in the prevention and/or treatment of retinal degenerative diseases has been widely explored; however, the underlying molecular mechanism is poorly understood. Photoreceptor degeneration is the hallmark of retinal degenerative diseases and leads to vision impairment or loss. In this study, the effect of GBE against white light (WL) illumination-induced photoreceptor degeneration was investigated, as well as its underlying mechanism. To evaluate the in vitro activity of GBE, analysis of cell viability, cell apoptosis, oxidative stress, NOX (NADH oxidase) activity and mitochondrial membrane potential (MMP), as well as Western blotting and transcriptome sequencing and analysis, were conducted. To evaluate the in vivo activity of GBE, HE staining, electroretinography (ERG), Terminal-deoxynucleoitidyl transferase (TdT)-mediated nick end labeling (TUNEL) assay and immunofluorescence analysis were conducted. Our study showed that GBE treatment significantly attenuated WL illumination-induced oxidative damage in photoreceptor 661W cells—a finding that was also verified in C57BL/6J mice. Further molecular study revealed that WL illumination downregulated caveolin-1 (CAV-1) expression, interrupted CAV-1-NOX2 interaction, re-located NOX2 from the cell membrane to the cytoplasm and induced the formation of redoxosomes, which led to cell death. However, these cytotoxic events were significantly alleviated by GBE treatment. Interestingly, CAV-1 overexpression showed a consistent protective effect with GBE, while CAV-1 silencing impacted the protective effect of GBE against WL illumination-induced oxidative damage in in vitro and in vivo models. Thus, GBE was identified to prevent photoreceptor cell death due to CAV-1-dependent redoxosome activation, oxidative stress and mitochondrial dysfunction resulting from WL illumination. Overall, our study reveals the protective effect of GBE on photoreceptors against WL illumination-induced oxidative damage in in vitro and in vivo models, which effect is mediated through the modulation of CAV-1-redoxosome signaling. Our findings contribute to better understanding the therapeutic effect of GBE in preventing photoreceptor degeneration in retinal degenerative diseases, and GBE may become a novel therapeutic agent that is effective in reducing the morbidity of these diseases.

Immobilization of Alcohol Dehydrogenase, Acetaldehyde Lyase, and NADH Oxidase for Cascade Enzymatic Conversion of Ethanol to Acetoin

Acetoin, a four-carbon hydroxyl-keto compound, is used in the food, pharmaceutical, and chemical industries. The cascade enzymatic production is considered a promising and efficient method to produce acetoin. However, the stability and compatibility of the enzymes under the same catalytic conditions are challenges that need to be resolved. In this work, alcohol dehydrogenase, acetaldehyde lyase, and NADH oxidase were selected to work at the same conditions to efficiently convert ethanol into acetoin. These three enzymes were immobilized on epoxy-modified magnetic nanomaterials to obtain highly stable biocatalysts. The stability and the immobilization conditions, including temperature, pH, enzyme–carrier ratio, and immobilization time, were optimized to obtain the immobilized enzymes with a high catalytic activity. The cascade reactions catalyzed by the immobilized enzymes yielded a high conversion of 90%, suggesting that the use of immobilized enzymes is a promising way to produce acetoin.

Association of cerebral microvascular dysfunction and white matter injury in Alzheimer's disease.

Patients with Alzheimer's disease (AD) often have cerebral white matter (WM) hyperintensities on MRI and microinfarcts of presumed microvascular origin pathologically. Here, we determined if vasodilator dysfunction of WM-penetrating arterioles is associated with pathologically defined WM injury and disturbances in quantitative MRI-defined WM integrity in patients with mixed microvascular and AD pathology. We analyzed tissues from 28 serially collected human brains from research donors diagnosed with varying degrees of AD neuropathologic change (ADNC) with or without cerebral microinfarcts (mVBI). WM-penetrating and pial surface arteriolar responses to the endothelium-dependent agonist bradykinin were quantified ex vivo with videomicroscopy. Vascular endothelial nitric oxide synthase (eNOS) and NAD(P)H-oxidase (Nox1,2 and 4 isoforms) expression were measured with quantitative PCR. Glial fibrillary acidic protein (GFAP)-labeled astrocytes were quantified by unbiased stereological approaches in regions adjacent to the sites of WM-penetrating vessel collection. Post-mortem diffusion tensor imaging (DTI) was used to measure mean apparent diffusion coefficient (ADC) and fractional anisotropy (FA), quantitative indices of WM integrity. In contrast to pial surface arterioles, white matter-penetrating arterioles from donors diagnosed with high ADNC and mVBI exhibited a significantly reduced dilation in response to bradykinin when compared to the other groups. Expression of eNOS was reduced, whereas Nox1 expression was increased in WM arterioles in AD and mVBI cases. WM astrocyte density was increased in AD and mVBI, which correlated with a reduced vasodilation in WM arterioles. Moreover, in cases with low ADNC, bradykinin-induced WM arteriole dilation correlated with lower ADC and higher FA values. Comorbid ADNC and mVBI appear to synergistically interact to selectively impair bradykinin-induced vasodilation in WM-penetrating arterioles, which may be related to reduced nitric oxide- and excess reactive oxygen species-mediated vascular endothelial dysfunction. WM arteriole vasodilator dysfunction is associated with WM injury, as supported by reactive astrogliosis and MRI-defined disrupted WM microstructural integrity.

REDD1 interacts with AIF and regulates mitochondrial reactive oxygen species generation in the keratinocyte response to UVB

Non-melanoma skin cancer (NMSC) incidence is rising, especially in high-risk, immunocompromised groups such as organ transplant patients, who often develop numerous, aggressive cutaneous squamous cell carcinomas. Identifying the pathways that support NMSC development will result in new approaches for prevention and therapy. Our goal is to define the function of REDD1 (Regulated in DNA Damage and Development 1) in the UVB stress response. REDD1 is rapidly induced by a variety of stressors to repress mechanistic target of rapamycin complex I (mTORC1), and it has been reported that REDD1 loss causes dysfunctional mitochondria with increased reactive oxygen species (ROS) and impaired oxidative phosphorylation (OXPHOS). We now show that knockout of REDD1 in human keratinocytes sensitizes them to UVB-induced apoptosis in an mTORC1-independent manner and intensifies mitochondrial ROS generation. Upon REDD1 knockout, we observe reduced levels of apoptosis inducing factor (AIF), a mitochondrial intermembrane space NADH oxidase that is required for electron transport chain Complex I biogenesis. Further, we show that keratinocyte REDD1 interacts with both AIF and the mitochondrial import protein CHCHD4, a direct binding partner of AIF that ensures functional OXPHOS. Our results support the hypothesis that REDD1 is part of a mitochondrial complex that protects cells from UVB-induced ROS toxicity and suggest novel therapeutic targets for prevention and therapy of NMSC.

Cu/nucleotide coordination self-assembling to in situ regenerate NAD(P)+ and co-immobilize dehydrogenase with robust activity and stability

Enzyme is an efficient green catalyst, but its vulnerability and difficulties of separation from products limited the application in industry. Furthermore, a large number of oxidase enzymes rely on dihydronicotinamide adenine dinucleotide (NADH) or dihydronicotinamide adenine dinucleotide phosphate (NADPH) cofactors. These cofactors are too expensive to be used in industrial processes. Thus, cofactor regeneration is an important method to avoid the consumption of large quantities of oxidized cofactor NAD(P)+ in enzyme-catalyzed reactions. In this paper, Cu based metal nucleotides coordination polymers (MNCPs) with NAD(P)H oxidase activity were constructed by self-assembly of copper ions and nucleotides. Cu2+/AMP and Cu2+/GMP showed high catalytic activity and high reusability. Then, Cu2+/AMP with NADH oxidase activity was combined with NAD+ dependent alcohol dehydrogenase (ADH) to construct a bienzyme cascade system with circulating cofactor, which applied to the conversion of benzyl alcohol to benzaldehyde. In this system, ADH exhibited high activity due to the increase of substrate concentration and the inhibition of the diffusion of intermediate products. Besides, the bienzyme cascade system also showed high stability and reusability. After 8 cycles, the immobilized ADH remained more than 57% of the initial activity. The prepared bienzyme cascade system can extensively extended its application in industrial production.

Antimicrobial activity, molecular typing and in vitro safety assessment of Lactococcus garvieae isolates from healthy cultured rainbow trout (Oncorhynchus mykiss, Walbaum) and rearing environment

Lactococcus garvieae is the aetiological agent of lactococcosis, an ichthyopathology affecting rainbow trout (Oncorhynchus mykiss, Walbaum) farming and having recently been considered as a potential food-borne zoonotic agent. This work aims to characterise a population of 53 L. garvieae isolates from healthy cultured rainbow trout and rearing environment by using microbiological, biochemical and genetic assays to unveil the potential impact that their presence may have, not only for fish, but also for consumers. The lactococci exhibited a broad antimicrobial spectrum against Gram-negative ichthyopathogens, likely due to production of organic acids. The 53 L. garvieae isolates were grouped into two and five genetic clusters by PFGE and ERIC-PCR, respectively. All the lactococcal isolates produce α-haemolysis and harbour, at least, 10 putative virulence factors (hly-1, hly-2, hly-3, NADH oxidase, sod, pgm, adhPsaA, eno, LPxTG-3 and adhCI). While all the lactococci were resistant to streptomycin and clindamycin, no resistance genes were found. The manuscript concludes that the non-existence of active lactococcosis does not guarantee the absence of potentially virulent L. garvieae, which also exhibit a great capacity for adaptation and persistence in the environment. Furthermore, the antibiotic resistance profile shown may lead to re-evaluate the treatments of choice in human patients.

Melatonin Maintains Fruit Quality and Reduces Anthracnose in Postharvest Papaya via Enhancement of Antioxidants and Inhibition of Pathogen Development.

Papaya fruit is widely grown in tropical regions because of its sweet taste, vibrant color, and the huge number of health benefits it provides. Melatonin is an essential hormone that governs many plants′ biological processes. In the current study, the impact of melatonin on fruit ripening and deterioration in postharvest papaya fruit was explored. An optimum melatonin dose (400 μmol L−1, 2 h) was found to be effective in delaying fruit softening and reducing anthracnose incidence. Melatonin enhanced antioxidant activity and decreased fruit oxidative injury by lowering superoxide anion, hydrogen peroxide, and malondialdehyde content by enhancing the enzymatic and non-enzymatic antioxidants, and by improving the antioxidant capacity of papaya fruit. Melatonin increased catalase, ascorbate peroxidase, NADH oxidase, glutathione reductase, polyphenol oxidase, superoxide dismutase, and peroxidase activity, as well as induced total phenol, total flavonoid, and ascorbic acid accumulation. Melatonin also enhanced the activity of defense-related enzymes, such as chitinase, 4-coumaric acid-CoA-ligase, and phenylalanine ammonia lyase, while it repressed lipid metabolism. Additionally, melatonin inhibited the development of anthracnose in vitro and in vivo. These findings suggest that exogenous melatonin application improves papaya fruit quality by boosting antioxidant and defense-related mechanisms.

The Effect of Benzyl Isothiocyanate on the Expression of Genes Encoding NADH Oxidase and Fibronectin-Binding Protein in Oral Streptococcal Biofilms.

Recent studies have shown that antimicrobial treatment results in up- or down regulation of several virulence-associated genes in bacterial biofilms. The genes encoding NADH oxidase (nox) and fibronectin-binding protein (fbp) are known to play important roles in biofilm growth of some oral bacterial species. The objective was to study the effect of benzyl isothiocyanate (BITC), an antimicrobial agent from Miswak plant, on the expression of nox and fbp genes in some oral streptococci. The biofilms were treated with BITC and mRNA expression of nox and fbp genes was measured by comparative ΔΔCt method. The highest amount of biofilm mass was produced by A. defectiva, followed by S. gordonii, S. mutans, G. elegans and G. adiacens. Upon treatment with BITC, S. gordonii biofilms showed highest folds change in mRNA expression for both fbp and nox genes followed by S. mutans, A. defectiva, and G. adiacens. G. elegans mRNA levels for nox were extremely low. In conclusion, BITC treatment of the biofilms caused an upregulation of biofilm-associated genes fbp and nox genes in most of the tested species suggesting the significance of these genes in biofilm lifestyle of these oral bacteria and needs further investigation to understand if it contributes to antimicrobial resistance.

Listeria monocytogenes requires cellular respiration for NAD+ regeneration and pathogenesis.

Cellular respiration is essential for multiple bacterial pathogens and a validated antibiotic target. In addition to driving oxidative phosphorylation, bacterial respiration has a variety of ancillary functions that obscure its contribution to pathogenesis. We find here that the intracellular pathogen Listeria monocytogenes encodes two respiratory pathways which are partially functionally redundant and indispensable for pathogenesis. Loss of respiration decreased NAD+ regeneration, but this could be specifically reversed by heterologous expression of a water-forming NADH oxidase (NOX). NOX expression fully rescued intracellular growth defects and increased L. monocytogenes loads >1000-fold in a mouse infection model. Consistent with NAD+ regeneration maintaining L. monocytogenes viability and enabling immune evasion, a respiration-deficient strain exhibited elevated bacteriolysis within the host cytosol and NOX expression rescued this phenotype. These studies show that NAD+ regeneration represents a major role of L. monocytogenes respiration and highlight the nuanced relationship between bacterial metabolism, physiology, and pathogenesis.

NADH oxidase of Mycoplasma hyopneumoniae functions as a potential mediator of virulence

BackgroundMycoplasma hyopneumoniae (M. hyopneumoniae) is the etiological agent of enzootic pneumonia, a highly infectious swine respiratory disease that distributed worldwide. The pathogenesis and virulence factors of M. hyopneumoniae are not fully clarified. As an important virulence factor of bacteria, nicotinamide adenine dinucleotide (NADH) oxidase (NOX) participates in host-pathogen interaction, however, the function of NOX involved in the pathogenesis of M. hyopneumoniae is not clear.ResultsIn this study, significant differences in NOX transcription expression levels among different strains of M. hyopneumoniae differed in virulence were identified, suggesting that NOX may be correlated with M. hyopneumoniae virulence. The nox gene of M. hyopneumoniae was cloned and expressed in Escherichia coli, and polyclonal antibodies against recombinant NOX (rNOX) were prepared. We confirmed the enzymatic activity of rNOX based on its capacity to oxidize NADH to NAD+. Flow cytometry analysis demonstrated the surface localization of NOX, and subcellular localization analysis further demonstrated that NOX exists in both the cytoplasm and cell membrane. rNOX was depicted to mediate adhesion to immortalized porcine bronchial epithelial cells (hTERT-PBECs). Pre-neutralizing M. hyopneumoniae with anti-rNOX antibody resulted in a more than 55% reduction in the adhesion rate of high- and low-virulence M. hyopneumoniae strains to hTERT-PBECs. Moreover, a significant difference appeared in the decline in CCU50 titer between virulent (168) and virulence-attenuated (168L) strains. NOX not only recognized and interacted with host fibronectin but also induced cellular oxidative stress and apoptosis in hTERT-PBECs. The release of lactate dehydrogenase by NOX in hTERT-PBECs was positively correlated with the virulence of M. hyopneumoniae strains.ConclusionsNOX is considered to be a potential virulence factor of M. hyopneumoniae and may play a significant role in mediating its pathogenesis.

Cymbopogon flexuosus (nees ex steud.) w. watson essential oil effect on mitochondrial bioenergetics

ABSTRACT Cymbopogon flexuosus is a medicinal plant with various biological activities. Although its essential oil (EO) is widely used, the effect of C. flexuosus EO on mitochondrial bioenergetics has not been completely elucidated. Here, we report the effect of C. flexuosus EO on electron transport through the mitochondrial respiratory chain using isolated mitochondria from liver rats. The EO decreases the oxygen consumption rate up to −20% at 10 ppm, and −17% at 25 ppm using glutamate and succinate, respectively. The C. flexuosus EO inhibits the NADH oxidase (34%), succinate oxidase (53%), succinate dehydrogenase (44%), and cytochrome c oxidase (69%) activities at 10 ppm, which indicates a robust inhibitory effect on the electron transport chain. Our results show the inhibitory effect of C. flexuosus EO on electron transport through complex II and IV and suggest that this EO could be used as a potential drug to treat mitochondrial diseases, including cancer.

Cadmium sulfide as bifunctional mimics of NADH oxidase and cytochrome c reductase takes effect at physiological pH

Cadmium sulfide as bifunctional mimics of NADH oxidase and cytochrome c reductase takes effect at physiological pH

Stereo-Divergent Enzyme Cascades to Convert Racemic 4-Phenyl-2-Butanol into either (S)- or (R)-Corresponding Chiral Amine.

The synthesis of enantiopure chiral amines from racemic alcohols is a key transformation in the chemical industry, e. g., in the production of active pharmaceutical ingredients (APIs). However, this reaction remains challenging. In this work, we propose a one‐pot enzymatic cascade for the direct conversion of a racemic alcohol into either (S)‐ or (R)‐enantiomers of the corresponding amine, with in‐situ cofactor recycling. This enzymatic cascade consists of two enantio‐complementary alcohol dehydrogenases, both NADH and NADPH oxidase for in‐situ recycling of NAD(P)+ cofactors, and either (S)‐ or (R)‐enantioselective transaminase. This cell‐free biocatalytic system has been successfully applied to the conversion of racemic 4‐phenyl‐2‐butanol into the high value (S)‐ or (R)‐enantiomers of the amine reaching good (73 % (S)) and excellent (>99 % (R)) enantioselectivities.