Substrate docking and molecular dynamic simulation for prediction of fungal enzymes from Trichoderma species-assisted extraction of nanocellulose from oil palm leaves

Abstract

Fungi of the Trichoderma species are valued industrial enzymes in support of the ‘zero-waste’ technology to convert agro-industrial biomass into valuable products, i.e. nanocellulose (NC). In this study, an in silico approach using substrate docking and molecular dynamic (MD) simulation was used to predict the order of which the multilayers of cellulosic polymers, i.e. lignin, hemicellulose and cellulose in oil palm leaves (OPL) are degraded by fungal enzymes, endocellulase and exocellulase. The study aimed to establish the catalytic tendencies of the enzymes to optimally degrade the cellulosic components of OPL for high yield production of NC. Energy minimized endocellulase and exocellulase models revealed satisfactory scores of PROCHECK (90.0% and 91.2%), Verify3D (97.23% and 98.85%) and ERRAT (95.24% and 91.00%) assessments. Active site prediction by blind docking, COACH meta-server and multiple sequence alignment indicated the catalytic triads for endocellulase and exocellulase were Ser116–His205–Glu249 and Ser382–Arg124–Asp385, respectively. Binding energy of endocellulase docked with hemicellulose (?6.0 ? kcal mol^?1) was the most favourable followed by lignin (?5.6 ? kcal mol^?1) and cellulose (?4.4 ? kcal mol^?1). Exocellulase, contrarily, bonded favorably with lignin (?8.7 ? kcal mol^?1), closely followed by cellulose (?8.5 ? kcal mol^?1) and hemicellulose (?8.4 ? kcal mol^?1). MDs simulations showed that interactions of complexes, endocellulase–hemicellulose and the exocellulase–cellulose being the most stable. Thus, the findings of the study successfully identified the specific actions of sugar-acting enzymes for NC production.

Communicated by Ramaswamy H. Sarma     

In silico characterization of a novel dehalogenase (DehHX) from the halophile <Emphasis Type="Italic">Pseudomonas halophila</Emphasis> HX isolated from Tuz Gölü Lake,Turkey: insights into a hypersaline-adapted dehalogenase

Halogenated compounds represent potential long-term threats to human well-being and health and, therefore, the quest for microorganisms capable of degrading these hazardous substances merits urgent consideration. We have isolated a novel dehalogenase-producing bacterium from the hypersaline environment of Tuz Gölü Lake, Turkey and subsequently identified this isolate as Pseudomonas halophila HX. Under optimal culture conditions (pH 8.0, 15% NaCl, 30 °C, 200 rpm, 96 h culture time), the strain almost completely degraded (99.3%) 2,2-dichloropropionic acid (20 mM). The dehalogenase gene (dehHX) of the bacterium was amplified by PCR, and the deduced amino acid sequence of the DehHX was found to belong to a Group I dehalogenase and to share an 82% sequence identity to the dehalogenase DehI of Pseudomonas putida strain PP3. Interestingly, the pI of DehHX was more acidic (pI 3.89) than those of the non-halophilic dehalogenases (average measured pI 5.95). Homology-based structural modeling revealed that the surface of DehHX was unusually negatively charged due to the higher presence of acidic residues, which accounts for the uncommonly low pI seen in DehHX and explains the mechanism of adaptation that contributes to the exceptional halotolerance of the enzyme. The excess surface acidic residues were beneficial in enhancing the water-binding capacity, a crucial feature for preserving the stability and solubility of DehHX in highly saline conditions. In summary, we suggest that bio-prospecting for halogenated compound-degrading microorganisms in highly saline environments is a practical and safe strategy for the bioremediation of contaminated coastal areas.     

Synthesis,structure–activity relationship and molecular docking of 3-oxoaurones and 3-thioaurones as acetylcholinesterase and butyrylcholinesterase inhibitors

The present study describes efficient and facile syntheses of varyingly substituted 3-thioaurones from the corresponding 3-oxoaurones using Lawesson’s reagent and phosphorous pentasulfide. In comparison, the latter methodology was proved more convenient, giving higher yields and required short and simple methodology. The structures of synthetic compounds were unambiguously elucidated by IR, MS and NMR spectroscopy. All synthetic compounds were screened for their inhibitory potential against in vitro acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) enzymes. Molecular docking studies were also performed in order to examine their binding interactions with AChE and BChE human proteins. Both studies revealed that some of these compounds were found to be good inhibitors against AChE and BChE.     

Characterization of Dahl salt-sensitive rats with genetic disruption of the A2B adenosine receptor gene: implications for A2B adenosine receptor signaling during hypertension

The A_2B adenosine receptor (AR) has emerged as a unique member of the AR family with contrasting roles during acute and chronic disease states. We utilized zinc-finger nuclease technology to create A_2BAR gene (Adora2b)-disrupted rats on the Dahl salt-sensitive (SS) genetic background. This strategy yielded a rat strain (SS-Adora2b mutant rats) with a 162-base pair in-frame deletion of Adora2b that included the start codon. Disruption of A_2BAR function in SS-Adora2b mutant rats was confirmed by loss of agonist (BAY 60-6583 or NECA)-induced cAMP accumulation and loss of interleukin-6 release from isolated fibroblasts. In addition, BAY 60-6583 produced a dose-dependent increase in glucose mobilization that was absent in SS-Adora2b mutants. Upon initial characterization, SS-Adora2b mutant rats were found to exhibit increased body weight, a transient delay in glucose clearance, and reduced proinflammatory cytokine production following challenge with lipopolysaccharide (LPS). In addition, blood pressure was elevated to a greater extent (∼15–20 mmHg) in SS-Adora2b mutants as they aged from 7 to 21 weeks. In contrast, hypertension augmented by Ang II infusion was attenuated in SS-Adora2b mutant rats. Despite differences in blood pressure, indices of renal and cardiac injury were similar in SS-Adora2b mutants during Ang II-augmented hypertension. We have successfully created and validated a new animal model that will be valuable for investigating the biology of the A_2BAR. Our data indicate varying roles for A_2BAR signaling in regulating blood pressure in SS rats, playing both anti- and prohypertensive roles depending on the pathogenic mechanisms that contribute to blood pressure elevation.     

Possible role of metal ionophore against zinc induced cognitive dysfunction in <Emphasis Type="SmallCaps">d-</Emphasis>galactose senescent mice

Metal ionophores are considered as potential anti-dementia agents, and some are currently undergoing clinical trials. Many metals are known to accumulate and distribute abnormally in the aging brain. Alterations in zinc metal homeostasis in the glutaminergic synapse could contribute to ageing and the pathophysiology of Alzheimer’s disease (AD). The present study was designed to investigate the effect of metal ionophores on long term administration of zinc in D-galactose induced senescent mice. The ageing model was established by combined administration of zinc and D-galactose to mice for 6 weeks. A novel metal ionophore, PBT-2 was given daily to zinc-induced d-galactose senescent mice. The cognitive behaviour of mice was monitored using the Morris Water Maze. The anti-oxidant status and amyloidogenic activity in the ageing mouse was measured by determining mito-oxidative parameters and deposition of amyloid β (Aβ) in the brain. Systemic administration of both zinc and D-galactose significantly produced memory deficits, mito-oxidative damage, heightened acetylcholinesterase enzymatic activity and deposition of amyloid-β. Treatment with PBT-2 significantly improved behavioural deficits, biochemical profiles, cellular damage, and curbed the deposition of APP in zinc-induced senescent mice. These findings suggest that PBT-2, acting as a metal protein attenuating compound, may be helpful in the prevention of AD or alleviation of ageing.     

Use of textile waste water along with liquid NPK fertilizer for production of wheat on saline sodic soils

A field experiment in collaboration with a private textile industry (Noor Fatima Fabrics Private (Ltd.), Faisalabad) was conducted to evaluate the effect of disposed water from bleaching unit, printing unit and end drain for improving growth and yield of wheat under saline sodic soil. Textile waste water along with canal water (control) was applied with and without liquid NPK fertilizer. The application of liquid NPK fertilizer with end drain waste water increased plant height, spike length, flag leaf length, root length, number of tillers (m^?2), number of fertile tillers (m^?2), 1000 grain weight, grain yield, straw yield and biological yield up to 21, 20, 20, 44, 17, 20, 14, 44, 40 and 41%, respectively compared to canal water (control). Similarly, the NPK uptake in grain was increased up to 15, 30 and 28%, respectively by liquid fertilizer treated end drain water as compare to canal water with liquid fertilizer. Moreover, concentration of different heavy metals particularly Cu, Cr, Pb and Cd was decreased in grains by application of waste water along with liquid NPK. The result may imply that waste water application along with liquid-NPK could be a novel approach for improving growth and yield of wheat in saline sodic soils.     

DNA photolyase of enterococci: possible explanation for its low sunlight inactivation rate

DNA photolyase is perhaps the most ancient and direct arsenal in curing the UV-induced dimers formed in the microbial genome. Out of two cofactors of the enzyme, catalytic and light harvesting, differences in the latter have provided basis for categorizing photolyases of prokaryotes as folate and deazaflavin types. In the present study, the homology modeling of DNA photolyase of Enterococcus faecalis was undertaken. The predicted models were structurally compared with the crystal structure coordinates of photolyases from Escherichia coli (folate type) and Anacystis nidulans (deazaflavin type). Discrepancies present in the multiple sequence alignment and tertiary structures, particularly at the light harvesting cofactor (methenyltetrahydrofolic acid, MTHF; 8-hydroxy-5-deazaflavin, 8-HDF) binding sites indicated the mechanistic nature of enterococcal photolyase. Concisely, despite the greater holistic homology with folate-type photolyase, enterococcal photolyase was characterized as deazaflavin-type. The presence of 8-HDF binding sites and groove architecture of substrate binding sites were also found supportive in this regard. The inter cofactor distance and/or orientation also implied to the efficient energy transfer in photolyase of Enterococcus in comparison with E. coli. In addition, we observed relatively high protein deformability in the enterococcal genome, which may favors the repair action of photolyase. The findings are expected to provide molecular insights into the difference in sunlight inactivation rate of two important fecal contamination indicators, namely Enterococcus and E. coli.     

Characterization and partial purification of <Emphasis Type="Italic">Candida albicans</Emphasis> Secretory IL-12 Inhibitory Factor

Background  

We have previously shown that supernatant from Candida albicans (CA) culture contains a Secretory Interleukin (IL)-12 Inhibitory Factor (CA-SIIF), which inhibits IL-12 production by human monocytes. However, the effect of CA-SIIF on secretion of other cytokines by monocytes is unknown, and detailed characterization of this factor has not been performed.     

Phosphorylation of the receptor for PTH and PTHrP is required for internalization and regulates receptor signaling.

We have previously shown that agonist-dependent phosphorylation of the PTH/PTHrP receptor occurs on its carboxyl-terminal tail. Using site-directed mutagenesis, phosphopeptide mapping, and direct sequencing of cyanogen bromide-cleaved fragments of phosphoreceptors, we report here that PTH-dependent phosphorylation occurs on the serine residues at positions 491, 492, 493, 495, 501, and 504, and that the serine residue at position 489 is required for phosphorylation. When these seven sites were mutated to alanine residues, the mutant receptor was no longer phosphorylated after PTH stimulation. The phosphorylation-deficient receptor, stably expressed in LLCPK-1 cells, was impaired in PTH-dependent internalization and showed an increased sensitivity to PTH stimulation; the EC(50) for PTH-stimulated cAMP accumulation was decreased by 7-fold. Furthermore, PTH stimulation of the phosphorylation-deficient PTH/PTHrP receptor caused a sustained elevation in intracellular cAMP levels. These data indicate that agonist-dependent phosphorylation of the PTH/PTHrP receptor plays an important role in receptor function.