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This study investigated 96 h median lethal concentration of endosulfan (99%, pure α: β ratio of 7:3) by conducting static non-renewable acute toxicity bio-assay in Chanos chanos juvenile with average weight (110±5.65 g). Further, the effect of different definitive doses (18.5, 19.5, 20.5, 21.5 and 22.5 µg/L) of endosulfan on metabolic, heamato-immunoligcal and histopathological response were probed. Anti-oxidative enzymes CAT, SOD and GST showed significant (p<0.01) increase of activity in the liver, gill and brain during exposure to endosulfan in a dose and time dependent manner. The brain AChE activity showed significant (p<0.01) inhibition from 18.5 to 22.5 µg/L exposure of endosulfan than the control group. LDH and MDH activity gradually increased with consequent increasing dose of endosulfan exposure in the liver, gill and brain. Similarly, ALT, AST and G6PDH activities in both liver and gill increased with consequent increases in the dose of endosulfan exposure. Immunological profile such as blood glucose and serum cortisol level significantly enhanced while respiratory burst activity declined with consequent increasing doses of endosulfan exposure. Histopathological alteration in the gill demonstrated curling of secondary lamellae, thickening of primary epithelium, shorting of secondary lamellae, epithelial hyperplasia, fusion of secondary lamellae, aneurism, and collapsed secondary lamellae due to dose dependent exposure of endosulfan. Liver histology illustrated cloudy swelling and necrosis with pyknotic nuclei to the moderate dose of endosulfan, whereas higher dose of endosulfan (21.5 µg/L) displayed severe necrosis of hepatic cells. Overall results clearly indicate that acute exposure of endosulfan led to pronounced deleterious alterations on biochemical, heamato-immunological, and histopathological responses of C. chanos juvenile. [ABSTRACT FROM AUTHOR]

The present study attempts to integrate phyco-remediation and enhanced lipid productivity using microalgae-bacterial consortium enriched from wastewater fed aquaculture pond. Metagenomic analyses and microscopic images of the consortium revealed the presence of Chlorella variabilis, Parachlorella kessleri, Thermosynechococcus elongatus, Chlamydomonas, Phaeodactylum tricornutum, Oscillatoriales, Synechocystis sp. , Microcystis aeruginosa, Nostocales, Naviculales, Stramenopiles, other members of Chlorophyceae, Trebouxiophyceae, and Chroococcales along with potential bacterial bioremediants. During a 30 days trial run (15 days stabilization and 14 days remediation studies) for phyco-remediation drastic reduction in the nutrient and COD content from the tested wastewater samples was seen. There was up to 93% and 87.2% reduction in chemical oxygen demand (COD) and ammonium concentration, respectively. Further, almost 100% removal of nitrates and phosphates from the dairy wastewater upon 48 h of treatment with polyculture under ambient temperature (25 ± 2 °C) with 6309 lux illumination and mild aeration, was observed for all the seven cycles. Interestingly, the nutrient and COD concentrations in the treated water were below the discharge standards as per Central Pollution Control Board (CPCB) norms. In additions, biomass (reported as dry cell weight) was enhanced by 67% upon treatment with ammonia-rich dairy wastewater exhibiting 42% lipid, 55% carbohydrate, and 18.6% protein content enhancement. The polyculture mainly grown as attached biofilm to the surface, offered an easy harvesting and separation of grown biomass from the treated wastewater. Overall, dairy wastewater was found to be a potential nutrient source for microalgae-bacteria cultivation thereby making the treatment process sustainable and eco-friendly. [Display omitted] • 42% lipid and 1.7-fold algal biomass enhancement during dairy wastewater treatment. • Wastewater treatment within 48 h incubation for discharge and reuse. • An integrated approach for algal biomass production and wastewater treatment. [ABSTRACT FROM AUTHOR]

• First purification protocol of an intact mycobacterial F-ATP synthase. • First low resolution structure of an entire mycobacterial F-ATP synthase. • Purification of the fused mycobacterial b -δ 24-446 construct. • The novel 111-residues domain of mycobacterial subunit δ. • The potential of mycobacterial subunit δ as a drug target. In contrast to other prokaryotes, the Mycobacterial F 1 F O ATP synthase (α 3 :β 3 :γ:δ:ε: a : b : b' : c 9) is essential for growth. The mycobacterial enzyme is also unique as a result of its 111 amino acids extended δ subunit, whose gene is fused to the peripheral stalk subunit b. Recently, the crystallographic structures of the mycobacterial α 3 :β 3 :γ:ε-domain and c subunit ring were resolved. Here, we report the first purification protocol of the intact M. smegmatis F 1 F O ATP synthase including the F 1 -domain, the entire membrane-embedded F O sector, and the stator subunits b' and the fused b -δ. This enzyme purification enabled the determination of the first projected 2D- and 3D structure of the intact M. smegmatis F 1 F O ATP synthase by electron microscopy (EM) and single particle analysis. Expression and purification of the fused mycobacterial b -δ 24-446 construct, excluding the membrane-embedded N-terminal amino acids, provided insight into its secondary structure. By combining these data with homology and ab-initio modeling techniques, a model of the mycobacterial peripheral stalk subunits b -δ and b ' was generated. Superposition of the 3D M. smegmatis F-ATP synthase EM-structure, the α 3 :β 3 :γ:ε and c -ring, and the derived structural models of the peripheral stalk enabled a clear assignment of all F-ATP synthase subunits, in particular with respect to the unique mycobacterial peripheral stalk subunit b' and the elongated δ fused with subunit b. The arrangement of δ relative to the N-termini of the catalytic α 3 β 3 -headpiece and its potential as a drug target are discussed. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • 3D reconstruction from electron micrographs of the α chi 3 :β 3 :γ complex. • Low resolution solution structure of the mycobacterial F-ATP synthase subunit α. • Critical residues of subunit ε for energy coupling of mycobacterial F-ATP synthase. • Residue 121 of mycobacterial subunit ε affects MgADP inhibition. • Differences of the N-terminus of mycobacterial ε to its prokaryotic counterparts. Abstract The Mycobacterium tuberculosis (Mtb) F 1 F O -ATP synthase (α 3 :β 3 :γ:δ:ε: a : b : b' : c 9) is an essential enzyme that supplies energy for both the aerobic growing and the hypoxic dormant stage of the mycobacterial life cycle. Employing the heterologous F-ATP synthase model system α chi 3 :β 3 :γ we showed previously, that transfer of the C-terminal domain (CTD) of Mtb subunit α (Mt α 514-549) to a standard F-ATP synthase α subunit suppresses ATPase activity. Here we determined the 3D reconstruction from electron micrographs of the α chi 3 :β 3 :γ complex reconstituted with the Mtb subunit ε (Mt ε), which has been shown to crosstalk with the CTD of Mt α. Together with the first solution shape of Mtb subunit α (Mt α), derived from solution X-ray scattering, the structural data visualize the extended C-terminal stretch of the mycobacterial subunit α. In addition, Mt ε mutants Mt εR62L, Mt εE87A, Mt ε 6-121 , and Mt ε 1-120 , reconstituted with α chi 3 :β 3 :γ provided insight into their role in coupling and in trapping inhibiting MgADP. NMR solution studies of Mt εE87A gave insights into how this residue contributes to stability and crosstalk between the N-terminal domain (NTD) and the CTD of Mt ε. Analyses of the N-terminal mutant Mt ε 6-121 highlight the differences of the NTD of mycobacterial subunit ε to the well described Geobacillus stearothermophilus or Escherichia coli counterparts. These data are discussed in context of a crosstalk between the very N-terminal amino acids of Mt ε and the loop region of one c subunit of the c -ring turbine for coupling of proton-translocation and ATP synthesis activity. [ABSTRACT FROM AUTHOR]

We herein report the protective role of pyridoxine in enhancing thermal tolerance of Milkfish Chanos chanos reared under endosulfan-induced stress. Four isocaloric and isonitrogenous diets were prepared with graded levels of pyridoxine (0, 50, 75 and 100 mg/kg). Two hundred and twenty five fishes were randomly distributed into four treatment groups in triplicate, reared under endosulfan-treated water, which were fed with pyridoxine supplemented diet, while the negative control group was reared without endosulfan-treatment and control fed. The concentration of endosulfan in treated water was maintained at a level of 1/40th of LC 50 i.e. 0.52 μg/L. Dietary pyridoxine supplementation had significant (p < 0.01) effect on temperature tolerance viz. CTmax (Critical temperature maxima), LTmax (Lethal temperature maxima), CTmin (Critical temperature minima) and LTmin (Lethal temperature minima) of milkfish. The positive correlation was observed between CT max and LTmax (Y = −1.54 + 15.6x, R 2 , 0.943) as well as CTmin and LTmin (Y = −1.44 + 1.021x, R 2 , 0.941). At the end of the thermal tolerance study, antioxidative status and HSP 70 were significantly reduced in pyridoxine supplemented groups, whereas brain AChE was significantly (p < 0.01) elevated compared to positive and negative control. It is concluded that CTmax, LTmax, CTmin and LTmin, antioxidative status, neurotransmitter enzyme and HSP 70 strengthened the enhancement of thermal tolerance of Milkfish. [ABSTRACT FROM AUTHOR]

Introduction: Consensus has formed that hydrogen sulfide (H2S) has robust cytoprotective actions in multiple organ systems. The pro-survival actions and biological profiles of H2S are remarkably similar to those of nitric oxide (NO). Although there is recent evidence of cross-talk between H2S and NO, these molecules are thought to modulate independent signaling pathways. Methods/Results: Mice devoid of the key H2S producing enzyme, cystathionine gamma-lyase (CSE KO), exhibited impaired endothelial nitric oxide synthase (eNOS) function and diminished circulating nitrite (0.41 uM vs 0.73μM, p <0.05) nitrosylated protein (3.37nM vs 15.03nM, p <0.05), and cyclic GMP (9.3 vs 28.05pmol/ml, p <0.01) levels compared to wild-type (WT) mice. Further studies revealed that H2S therapy in CSE KO mice restored eNOS function and NO levels. eNOS KO mice and mice with phospho-dead eNOS (S1179A) were subjected to 45min of ischemia, followed by 24h of reperfusion. The H2S donor, Na2S (100μg/kg), or vehicle (0.9% NaCl) was administered 5min before reperfusion. There was no significant change in myocardial infarction in the H2S-treated group compared to the vehicle-treated group. Conclusion: These findings reveal that H2S modulates eNOS activity and nitric oxide signaling. Moreover, the inability of H2S therapy to protect eNOS dysfunctional mice against myocardial ischemia/reperfusion injury indicates that the cardioprotective actions of H2S are dependent on eNOS function and NO signaling. [Copyright &y& Elsevier]

Two novel metalloproteases from Arabidopsis thaliana, termed AtPrePI and AtPrePII, were recently identified and shown to degrade targeting peptides in mitochondria and chloroplasts using an ambiguous targeting peptide. AtPrePI and AtPrePII are classified as dually targeted proteins as they are targeted to both mitochondria and chloroplasts. Both proteases harbour an inverted metal binding motif and belong to the pitrilysin subfamily A. Here we have investigated the subsite specificity of AtPrePI and AtPrePII by studying their proteolytic activity against the mitochondrial F1β pre-sequence, peptides derived from the F1β pre-sequence as well as non-mitochondrial peptides and proteins. The degradation products were analysed, identified by MALDI-TOF spectrometry and superimposed on the 3D structure of the F1β pre-sequence. AtPrePI and AtPrePII cleaved peptides that are in the range of 10 to 65 amino acid residues, whereas folded or longer unfolded peptides and small proteins were not degraded. Both proteases showed preference for basic amino acids in the P1 position and small, uncharged amino acids or serine residues in the position. Interestingly, both AtPrePI and AtPrePII cleaved almost exclusively towards the ends of the α-helical elements of the F1β pre-sequence. However, AtPrePI showed a preference for the N-terminal amphiphilic α-helix and positively charged amino acid residues and degraded the F1β pre-sequence into 10–16 amino acid fragments, whereas AtPrePII did not show any positional preference and degraded the F1β pre-sequence into 10–23 amino acid fragments. In conclusion, despite the high sequence identity between AtPrePI and AtPrePII and similarities in cleavage specificities, cleavage site recognition differs for both proteases and is context and structure dependent. [Copyright &y& Elsevier]

Introduction: Hydrogen sulfide (H2S) is the endogenous signaling gasotransmitter, offering protective effects on oxidative stress including ischemia–reperfusion and hypertrophy/heart failure (HF). CSE is believed to be the major H2S-generating enzyme in the heart. Our previous study investigated that CSE KO mice are more susceptible to oxidative stress. NF-kappaB is a marker of cellular oxidative stress and its activation has been found to increase in ischemia–reperfusion and hypertrophy/HF. The purpose of the present study was to investigate the role of NF-kappaB in response to CSE. Methods/results: We found that both mRNA and protein levels of NF-kappaB-p65 were significantly increased in the heart tissues of CSE KO mice compared with control as they were judged by RT-qPCR and immunoblot, respectively. Interestingly, phosphorylation of p65 was increased (1.4 fold, p <0.05) in CSE KO mice. By utilizing electrophoretic mobility shift assay, we found increased NF-kappaB DNA binding activity in CSE KO mice compared to control. In addition, biomarkers of oxidative stress such as malondialdehyde (1.3 fold, p <0.05) and protein carbonyl (2 fold, p <0.01) formation were increased in heart tissues of CSE KO mice compared to wild type. Conclusion: Since we observed that the activation of NF-kappaB occurred in the heart tissues of CSE KO mice, our findings suggest that CSE may protect the heart from oxidative injury, such as that which occurs during ischemia–reperfusion, hypertrophy/HF. [Copyright &y& Elsevier]

Introduction: CBS, CSE and 3-MST are the enzymatic source of endogenous hydrogen sulfide (H2S), a gaseous signaling molecule implicated in a wide range of physiological processes. We investigated the status of CBS, CSE, and 3-MST as well as the extent of oxidative stress in skeletal muscle tissue obtained from patients suffering from critical limb ischemia (CLI) ± diabetes mellitus (Db). Methods/Results: We found that both mRNA and protein levels of CSE, CBS, and 3-MST were significantly decreased in skeletal muscle of CLI (>2 fold, p <0.05) and CLI +Db (2 fold, p <0.05) patients as judged by RT-qPCR and immunoblot, respectively. We also observed reduced expression of nuclear factor-erythriod 2-related factor (Nrf2), a factor responsible for the expression of antioxidant response element-regulated genes (2 fold, p <0.05) and Cu, Zn-super oxide dismutase, an antioxidant enzyme (2 fold, P <0.05) in skeletal muscle of both patients. Biomarkers of oxidative stress, such as malondialdehyde and protein carbonyl formation were significantly increased (2 fold) in skeletal muscle of CLI and CLI +Db patients as compared with age-matched controls. Conclusion: The present study suggests that the induction of oxidative stress in skeletal muscle of both CLI and CLI +Db patients may directly down-regulate the expression of the endogenous H2S-generating enzymes: CSE, CBS, and 3-MST. Increased oxidative stress and reductions in H2S bioavailability may contribute to the pathogenesis of CLI. [Copyright &y& Elsevier]