Sorption of heavy metals from electroplating effluent using immobilized biomass Trichoderma viride in a continuous packed-bed column

The sorption of heavy metals ions by immobilized Trichoderma viride biomass in a packed-bed column was studied. Fungal biomass T. viride was immobilized to Ca-alginate used for removal of Cr(VI), Ni(II) and Zn(II) ions from synthetic solutions and electroplating effluent. The experiments were conducted to study the effect of important design parameters such as bed height, flow rate and initial concentration of metal ions. The maximum sorption capacity was observed at flow rate 5 ml/min, bed height 20 cm and metal ions concentration 50 mg/L with immobilized biomass. Whereas, breakthrough time and saturation time decreased with increase flow rate and metal ions concentration and an inverse condition was found in bed height. The bed depth service time (BDST) Adams-Bohart model was used to analyze the experimental data. The regeneration efficiency was observed 40.1%, 75% and 53% for Cr(VI), Ni(II) and Zn(II) without any significant alteration in sorption capacity after 5th sorption-desorption cycles.     

N-terminal Gly(224)-Gly(411) domain in Listeria adhesion protein interacts with host receptor Hsp60

Background

Listeria adhesion protein (LAP) is a housekeeping bifunctional enzyme consisting of N-terminal acetaldehyde dehydrogenase (ALDH) and C-terminal alcohol dehydrogenase (ADH). It aids Listeria monocytogenes in crossing the epithelial barrier through a paracellular route by interacting with its host receptor, heat shock protein 60 (Hsp60). To gain insight into the binding interaction between LAP and Hsp60, LAP subdomain(s) participating in the Hsp60 interaction were investigated.

Methods

Using a ModBase structural model, LAP was divided into 4 putative subdomains: the ALDH region contains N1 (Met₁–Pro₂₂₃) and N2 (Gly₂₂₄–Gly₄₁₁), and the ADH region contains C1 (Gly₄₁₂–Val₆₄₈) and C2 (Pro₆₄₉–Val₈₆₆). Each subdomain was cloned and overexpressed in Escherichia coli and purified. Purified subdomains were used in ligand overlay, immunofluorescence, and bead-based epithelial cell adhesion assays to analyze each domain''s affinity toward Hsp60 protein or human ileocecal epithelial HCT-8 cells.

Results

The N2 subdomain exhibited the greatest affinity for Hsp60 with a K _D of 9.50±2.6 nM. The K _D of full-length LAP (7.2±0.5 nM) to Hsp60 was comparable to the N2 value. Microspheres (1 µm diameter) coated with N2 subdomain showed significantly (P<0.05) higher binding to HCT-8 cells than beads coated with other subdomains and this binding was inhibited when HCT-8 cells were pretreated with anti-Hsp60 antibody to specifically block epithelial Hsp60. Furthermore, HCT-8 cells pretreated with purified N2 subdomain also reduced L. monocytogenes adhesion by about 4 log confirming its involvement in interaction with epithelial cells.

Conclusion

These data indicate that the N2 subdomain in the LAP ALDH domain is critical in initiating interaction with mammalian cell receptor Hsp60 providing insight into the molecular mechanism of pathogenesis for the development of potential anti-listerial control strategies.     

Antiplasmodial and cytotoxicity evaluation of 3-functionalized 2-azetidinone derivatives

3-Azido-, 3-amino- and 3-(1,2,3-triazol-1-yl)-β-lactams were synthesized and evaluated for their antiplasmodial activity against four strains of Plasmodium falciparum and KB cells for their cytotoxicity profiles. The presence of a cyclohexyl substituent at N-1 and a phenyl group on the triazole ring markedly improved the activity profiles of triazole-tethered β-lactam exhibiting IC₅₀ values of 1.13, 1.21 and 1.00 μM against 3D7, K1 and W2 strains respectively.     

Synthesis, molecular modeling and bio-evaluation of cycloalkyl fused 2-aminopyrimidines as antitubercular and antidiabetic agents

An economical and efficient one step synthesis of a series of 8-(arylidene)-4-(aryl)-5,6,7,8-tetrahydro-quinazolin-2-ylamines and 9-(arylidene)-4-(aryl)-6,7,8,9-tetrahydro-5H-cycloheptapyrimidin-2-ylamines by the reaction of bis-benzylidene cycloalkanones and guanidine hydrochloride in presence of NaH has been developed. All the synthesized compounds were evaluated against Mycobacterium tuberculosis H₃₇Rv strain and the α-glucosidase and glycogen phosphorylase enzymes. Few of the compounds have shown interesting in vitro activity with MIC up to 3.12 μg/mL against M. tuberculosis and very good inhibition of α-glucosidase and glycogen phosphorylase enzymes. The most potent non toxic compound 40 exhibited about 58% ex vivo activity at MIC of 3.12 μg/mL. The present study opens a new gate to synthesize antitubercular agents for diabetic TB patients. In silico docking studies indicate that mycobacterial dihydrofolate reductase is the possible target of these compounds.     

Biodegradation of endosulfan and endosulfan sulfate by <Emphasis Type="Italic">Achromobacter xylosoxidans</Emphasis> strain C8B in broth medium

Endosulfan is one of the most widely used wide spectrum cyclodiene organochlorine insecticide. In environment, endosulfan can undergo either oxidation or hydrolysis reaction to form endosulfan sulfate and endosulfan diol respectively. Endosulfan sulfate is as toxic and as persistent as its parent isomers. In the present study, endosulfan degrading bacteria were isolated from soil through selective enrichment technique using sulfur free medium with endosulfan as sole sulfur source. Out of the 8 isolated bacterial strains, strain C8B was found to be the most efficient endosulfan degrader, degrading 94.12% α-endosulfan and 84.52% β-endosulfan. The bacterial strain was identified as Achromobacter xylosoxidans strain C8B on the basis of 16S rDNA sequence similarity. Achromobacter xylosoxidans strain C8B was also found to degrade 80.10% endosulfan sulfate using it as sulfur source. No known metabolites were found to be formed in the culture media during the entire course of degradation. Besides, the bacterial strain was found to degrade all the known endosulfan metabolites. There was marked increase in the quantity of released CO₂ from the culture media with endosulfan as sulfur source as compared to MgSO₄ suggesting that the bacterial strain, Achromobacter xylosoxidans strain C8B probably degraded endosulfan completely through the formation of endosulfan ether.     

Sodium Phenylbutyrate and Edaravone Abrogate Chronic Restraint Stress-Induced Behavioral Deficits: Implication of Oxido-Nitrosative,Endoplasmic Reticulum Stress Cascade,and Neuroinflammation

Chronic stress exposure can produce deleterious effects on the hippocampus (HC) which eventually leads to cognitive impairment and depression. Endoplasmic reticulum (ER) stress has been reported as one of the major culprits in the development of stress-induced cognitive impairment and depression. We investigated the neuroprotective efficacy of sodium phenylbutyrate (SPB), an ER stress inhibitor, and edaravone, a free radical scavenger, against chronic restraint stress (CRS)-induced cognitive deficits and anxiety- and depressive-like behavior in mice. Adult male Swiss albino mice were restrained for 6 h/day for 28 days and injected (i.p.) with SPB (40 and 120 mg/kg) or edaravone (3 and 10 mg/kg) for the last seven days. After stress cessation, the anxiety- and depressive-like behavior along with spatial learning and memory were examined. Furthermore, oxido-nitrosative stress, proinflammatory cytokines, and gene expression level of ER stress-related genes were assessed in HC and prefrontal cortex (PFC). CRS-exposed mice showed anxiety- and depressive-like behavior, which was significantly improved by SPB and edaravone treatment. In addition, SPB and edaravone treatment significantly alleviated CRS-induced spatial learning and memory impairment. Furthermore, CRS-evoked oxido-nitrosative stress, neuroinflammation, and depletion of Brain-derived neurotrophic factor were significantly ameliorated by SPB and edaravone treatment. We found significant up-regulation of ER stress-related genes in both HC and PFC regions, which were suppressed by SPB and edaravone treatment in CRS mice. Our study provides evidence that SPB and edaravone exerted neuroprotective effects on CRS-induced cognitive deficits and anxiety- and depressive-like behavior, which is possibly coupled with inhibition of oxido-nitrosative stress, neuroinflammation, and ER stress cascade.     

Co-Factor Binding Confers Substrate Specificity to Xylose Reductase from Debaryomyces hansenii

Binding of substrates into the active site, often through complementarity of shapes and charges, is central to the specificity of an enzyme. In many cases, substrate binding induces conformational changes in the active site, promoting specific interactions between them. In contrast, non-substrates either fail to bind or do not induce the requisite conformational changes upon binding and thus no catalysis occurs. In principle, both lock and key and induced-fit binding can provide specific interactions between the substrate and the enzyme. In this study, we present an interesting case where cofactor binding pre-tunes the active site geometry to recognize only the cognate substrates. We illustrate this principle by studying the substrate binding and kinetic properties of Xylose Reductase from Debaryomyces hansenii (DhXR), an AKR family enzyme which catalyzes the reduction of carbonyl substrates using NADPH as co-factor. DhXR reduces D-xylose with increased specificity and shows no activity towards “non-substrate” sugars like L-rhamnose. Interestingly, apo-DhXR binds to D-xylose and L-rhamnose with similar affinity (K_d∼5.0–10.0 mM). Crystal structure of apo-DhXR-rhamnose complex shows that L-rhamnose is bound to the active site cavity. L-rhamnose does not bind to holo-DhXR complex and thus, it cannot competitively inhibit D-xylose binding and catalysis even at 4–5 fold molar excess. Comparison of K_d values with K_m values reveals that increased specificity for D-xylose is achieved at the cost of moderately reduced affinity. The present work reveals a latent regulatory role for cofactor binding which was previously unknown and suggests that cofactor induced conformational changes may increase the complimentarity between D-xylose and active site similar to specificity achieved through induced-fit mechanism.     

Upregulation of the Tim-3/Galectin-9 Pathway of T Cell Exhaustion in Chronic Hepatitis B Virus Infection

The S-type lectin galectin-9 binds to the negative regulatory molecule Tim-3 on T cells and induces their apoptotic deletion or functional inactivation. We investigated whether galectin-9/Tim-3 interactions contribute to the deletion and exhaustion of the antiviral T cell response in chronic hepatitis B virus infection (CHB). We found Tim-3 to be expressed on a higher percentage of CD4 and CD8 T cells from patients with CHB than healthy controls (p<0.0001) and to be enriched on activated T cells and those infiltrating the HBV-infected liver. Direct ex vivo examination of virus-specific CD8 T cells binding HLA-A2/peptide multimers revealed that Tim-3 was more highly upregulated on HBV-specific CD8 T cells than CMV-specific CD8 T cells or the global CD8 T cell population in patients with CHB (p<0.001) or than on HBV-specific CD8 after resolution of infection. T cells expressing Tim-3 had an impaired ability to produce IFN-γ and TNF-α upon recognition of HBV-peptides and were susceptible to galectin-9-triggered cell death in vitro. Galectin-9 was detectable at increased concentrations in the sera of patients with active CHB-related liver inflammation (p = 0.02) and was strongly expressed by Kupffer cells within the liver sinusoidal network. Tim-3 blockade resulted in enhanced expansion of HBV-specific CD8 T cells able to produce cytokines and mediate cytotoxicity in vitro. Blocking PD-1 in combination with Tim-3 enhanced the number of patients from whom functional antiviral responses could be recovered and/or the strength of responses, indicating that these co-inhibitory molecules play a non-redundant role in driving T cell exhaustion in CHB. Patients taking antivirals able to potently suppress HBV viraemia continued to express Tim-3 on their T cells and respond to Tim-3 blockade. In summary, both Tim-3 and galectin-9 are increased in CHB and may contribute to the inhibition and deletion of T cells as they infiltrate the HBV-infected liver.     

In vivo active antimalarial isonitriles

Building on the lead from antimalarial isonitriles 1-4 of marine origin, several easily accessible synthetic isonitriles were assessed for their antimalarial activity against Plasmodium falciparum (in vitro) and multidrug resistant Plasmodium yoelii in Swiss mice model (in vivo). Isonitrile 11 has shown promising activity in both these assays.     

Ca(2+) and Mg(2+)/ATP independently trigger homotypic membrane fusion in gastric secretory membranes

Exocytic activation of gastric parietal cells represents a massive transformation. We studied a step in this process, homotypic fusion of H,K-ATPase-containing tubulovesicles, using R18 dequenching. Ca²⁺ and Mg²⁺/ATP each caused dramatic dequenching, reflecting a change in R18 distribution from 5% to 65–90% of the assay's membranes in 2.5 min. These stimuli also triggered fusion between tubulovesicles and liposomes. Independent confirmation that dequenching represented membrane fusion was established by separating tubulovesicle–liposome fusion products on density gradients. Only agents that trigger fusion allowed the transmembrane H,K-ATPase to move to low-density fractions along with R18. EC₅₀ for Ca²⁺-triggered fusion was 150 n m and for Mg²⁺/ATP-triggered fusion 1 m m , the latter having a Hill coefficient of 2.5. ATP-triggered fusion was specific for Mg²⁺/ATP, required ATP hydrolysis, and was insensitive to inhibition of NSF and/or H,K-ATPase. Fusion initiated by either trigger caused tubulovesicles to become resistant to subsequent challenge by either trigger. Ca²⁺-and Mg²⁺/ATP-triggered fusion required protein component(s) in tubulovesicles, though this was required in only one of the fusing membranes since tubulovesicles fused well with liposomes containing no proteins. Our data suggest that exocytosis in parietal cells is triggered by separate but interacting pathways and is regulated by self-inhibition.