Moleculer dynamics simulaiton revealed reciever domain of Acinetobacter baumannii BfmR enzyme as the hot spot for future antibiotics designing

Acinetobacter baumannii is an alarming nosocomial pathogen that is resistant to multiple drugs. The pathogen is forefront of scientific attention because of high mortality and morbidity found for its complications in the past decade. As a consequence, identification of novel drug candidates and subsequent designing of novel chemical scaffolds is an imperative need of time. In the present study, we used a recently reported structure of BfmR enzyme and performed structure based virtual screening, MD simulation and binding free energies calculations. MD simulation revealed a profound movement of the best-characterized inhibitor towards the α4-β5-α5 face of the enzyme receiver domain, thus indicating its high affinity for this site compared to phosphorylation. Furthermore, it was observed that the enzyme and enzyme-inhibitor complex have high structure stability with mean RMSD of 1.2 and 1.1 Å, respectively. Binding free energy calculations for the complex unraveled high stability with MMGBSA score of ?26.21?kcal/mol and MMPBSA score of ?1.47?kcal/mol. Van der Waal energy was found highly favorable with value of ?30.25?kcal/mol and dominated significantly the overall binding energy. Furthermore, a novel WaterSwap assay was used to circumvent the limitations of MMGB/PBSA that complements the inhibitor affinity for enzyme active pocket as depicted by the low convergence of Bennett, TI and FEP algorithms. Results yielded from this study will not only give insight into the phenomena of inhibitor movement towards the enzyme receiver domain, but will also provide a useful baseline for designing derivatives with improved biological and pharmacokinetics profiles.

Communicated by Ramaswamy H. Sarma     

Spectral lights trigger biomass accumulation and production of antioxidant secondary metabolites in adventitious root cultures of Stevia rebaudiana (Bert.)

Stevia rebaudiana (S. rebaudiana) is the most important therapeutic plant species and has been accepted as such worldwide. It has a tendency to accumulate steviol glycosides, which are 300 times sweeter than marketable sugar. Recently, diabetic patients commonly use this plant as a sugar substitute for sweet taste. In the present study, the effects of different spectral lights were investigated on biomass accumulation and production of secondary metabolites in adventitious root cultures of S. rebaudiana. For callus development, leaf explants were excised from seed-derived plantlets and inoculated on a Murashige and Skoog (MS) medium containing the combination of 2,4-dichlorophenoxy acetic acid (2, 4-D, 2.0 mg/l) and 6-benzyladenine (BA, 2.0 mg/l), while 0.5 mg/l naphthalene acetic acid (NAA) was used for adventitious root culture. Adventitious root cultures were exposed to different spectral lights (blue, green, violet, red and yellow) for a 30-day period. White light was used as control. The growth kinetics was studied for 30 days with 3-day intervals. In this study, the violet light showed the maximum accumulation of fresh biomass (2.495 g/flask) as compared to control (1.63 g/flask), while red light showed growth inhibition (1.025 g/flask) as compared to control. The blue light enhanced the highest accumulation of phenolic content (TPC; 6.56 mg GAE/g DW), total phenolic production (TPP; 101 mg/flask) as compared to control (5.44 mg GAE/g DW; 82.2 mg GAE/g DW), and exhibited a strong correlation with dry biomass. Blue light also improved the accumulation of total flavonoid content (TFC; 4.33 mg RE/g DW) and total flavonoid production (TFP; 65 mg/flask) as compared to control. The violet light showed the highest DPPH inhibition (79.72%), while the lowest antioxidant activity was observed for control roots (73.81%). Hence, we concluded that the application of spectral lights is an auspicious strategy for the enhancement of the required antioxidant secondary metabolites in adventitious root cultures of S. rebaudiana and of other medicinal plants.     

Antioxidant and anticholinesterase investigations of Rumex hastatus D. Don: potential effectiveness in oxidative stress and neurological disorders

Background

Rumex species are traditionally used for the treatment of neurological disorders including headache, migraine, depression, paralysis etc. Several species have been scientifically validated for antioxidant and anticholinestrase potentials. This study aims to investigate Rumex hastatus D. Don crude methanolic extract, subsequent fractions, saponins and flavonoids for acetylcholinestrase, butyrylcholinestrase inhibition and diverse antioxidant activities to validate its folkloric uses in neurological disorders. Rumex hastatus crude methanolic extract (Rh. Cr), subsequent fractions; n-hexane (Rh. Hex), chloroform (Rh. Chf), ethyl acetate (Rh. EtAc), aqueous fraction (Rh. Aq), crude saponins (Rh. Sp) and flavonoids (Rh. Fl) were investigated against acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) at various concentrations (125, 250, 500, 1000 μg/mL) using Ellman’s spectrophotometric analysis. Antioxidant potentials of Rh. Sp and Rh. Fl were evaluated using DPPH, H₂O₂ and ABTS free radical scavenging assays at 62.5, 125, 250, 500, 1000 μg/mL.

Results

All the test samples showed concentration dependent cholinesterase inhibition and radicals scavenging activity. The AChE inhibition potential of Rh. Sp and Rh. Fl were most prominent i.e., 81.67 ± 0.88 and 91.62 ± 1.67 at highest concentration with IC₅₀ 135 and 20 μg/mL respectively. All the subsequent fractions exhibited moderate to high AChE inhibition i.e., Rh. Cr, Rh. Hex, Rh. Chf, Rh. EtAc and Rh. Aq showed IC₅₀ 218, 1420, 75, 115 and 1210 μg/mL respectively. Similarly, against BChE various plant extracts i.e., Rh. Sp, Rh. Fl, Rh. Cr, Rh. Hex, Rh. Chf, Rh. EtAc and Rh. Aq resulted IC₅₀ 165, 175, 265, 890, 92, 115 and 220 μg/mL respectively. In DPPH free radical scavenging assay, Rh. Sp and Rh. Fl showed comparable results with the positive control i.e., 63.34 ± 0.98 and 76.93 ± 1.13% scavenging at 1 mg/mL concentration (IC₅₀ 312 and 104 μg/mL) respectively. The percent ABTS radical scavenging potential exhibited by Rh. Sp and Rh. Fl (1000 μg/mL) were 82.58 ± 0.52 and 88.25 ± 0.67 with IC₅₀ 18 and 9 μg/mL respectively. Similarly in H₂O₂ scavenging assay, the Rh. Sp and Rh. Fl exhibited IC₅₀ 175 and 275 μg/mL respectively.

Conclusion

The strong anticholinesterase and antioxidant activities of Rh. Sp, Rh. Fl and various fractions of R. hastatus support the purported ethnomedicinal uses and recommend R. hastatus as a possible remedy for the treatment of AD and neurodegenerative disorders.     

Synthesis and evaluation of antifungal properties of a series of the novel 2-amino-5-oxo-4-phenyl-5,6,7,8-tetrahydroquinoline-3-carbonitrile and its analogues

A series of 2-amino-5-oxo-4-phenyl-5,6,7,8-tetrahydroquinoline-3-carbonitrile and various analogues have been synthesized in excellent isolated yields starting from various arylidenemalononitrile and 3-amino-2-cyclohexen-1-one in 1-propanol as solvent at reflux temperature in the absence of any added catalyst. All the synthesized compounds were evaluated for their antifungal activity. The relationship between functional group variation and biological activity of the evaluated compounds is discussed in the article.     

Gibberellins and indole-3-acetic acid producing rhizospheric bacterium Leifsonia xyli SE134 mitigates the adverse effects of copper-mediated stress on tomato

Beneficial bacteria Supplemental data for this article can be accessed at https://doi.org/10.1080/17429145.2017.1370142.View all notesliving in the rhizosphere pose several implications on plant growth promotion and are highly desirable for sustainable agriculture. In the current study, we explored the ameliorative capacity of Leifsonia xyli SE134, a plant growth-promoting rhizobacteria (PGPR), against copper (Cu) stress on tomato grown under elevated Cu levels of 50 and 100?mM. Initially, L. xyli SE134 modulated innate gibberellins (GAs) and indole-3-acetic acid (IAA) metabolism in response to elevated Cu toxicity. The IAA contents increased, whereas that of bioactive GAs decreased in relation to Cu concentration gradient in the broth media. Furthermore, exposure to elevated Cu caused detrimental effects on the physiological attributes as revealed by attenuated shoot length, root length, stem diameter, shoot dry weight, root dry weight, and chlorophyll content in non-inoculated tomatoes as compared to L. xyli SE134 inoculated plants. The growth rescuing effect of L. xyli SE134 may be attributed to the modulation of endogenous amino acids contents in plants, such as glutamic acid, threonine, phenylalanine, glycine, proline, and arginine. Moreover, L. xyli SE134 inoculation stimulated total polyphenol and flavonoid content, reduced super oxide dismutase activity, strongly inhibited Cu, and increased phosphorus and iron content in plants grown under elevated Cu stress. In the absence of Cu toxicity, L. xyli SE134 significantly enhanced amino acid content, improved total flavonoids, and increased phosphorus content, thus resulting in higher plant growth.     

OipA plays a role in Helicobacter pylori-induced focal adhesion kinase activation and cytoskeletal re-organization

The initial signalling events leading to Helicobacter pylori infection associated changes in motility, cytoskeletal reorganization and elongation of gastric epithelial cells remain poorly understood. Because focal adhesion kinase (FAK) is known to play important roles in regulating actin cytoskeletal organization and cell motility we examined the effect of H. pylori in gastric epithelial cells co-cultured with H. pylori or its isogenic cag pathogenicity island (PAI) or oipA mutants. H. pylori induced FAK phosphorylation at distinct tyrosine residues in a dose- and time-dependent manner. Autophosphorylation of FAK Y397 was followed by phosphorylation of Src Y418 and resulted in phosphorylation of the five remaining FAK tyrosine sites. Phosphorylated FAK and Src activated Erk and induced actin stress fibre formation. FAK knock-down by FAK-siRNA inhibited H. pylori- mediated Erk phosphorylation and abolished stress fibre formation. Infection with oipA mutants reduced phosphorylation of Y397, Y576, Y577, Y861 and Y925, inhibited stress fibre formation and altered cell morphology. cag PAI mutants reduced phosphorylation of only FAK Y407 and had less effect on stress fibre formation than oipA mutants. We propose that activation of FAK and Src are responsible for H. pylori -induced induction of signalling pathways resulting in the changes in cell phenotype important for pathogenesis.     

Production enhancement and refolding of caprine growth hormone expressed in Escherichia coli

This study describes comparison between IPTG and lactose induction on expression of caprine growth hormone (cGH), enhancing cell densities of Escherichia coli cultures and refolding the recombinant cGH, produced as inclusion bodies, to biologically active state. 2–3 times higher cell densities were obtained in shake flask cultures when induction was done with lactose showing almost same level of expression as in case of IPTG induction. With lactose induction highest cell densities were achieved in TB (OD₆₀₀ 16.3) and M9NG (OD₆₀₀ 16.1) media, producing 885 and 892 mg cGH per liter of the culture, respectively. Lactose induction done at mid-exponential stage resulted in a higher cell density and thus higher product yield. cGH over-expressed as inclusion bodies was solubilized in 50 mM Tris–Cl buffer (pH 12.5) containing 2 M urea, followed by dilution and lowering the pH in a step-wise manner to obtain the final solution in 50 mM Tris–Cl (pH 9.5). The cGH was purified by Q-Sepharose chromatography followed by gel filtration with a recovery yield of 39% on the basis of total cell proteins. The product thus obtained showed a single band by SDS–PAGE analysis. MALDI-TOF analysis showed a single peak with a mass of 21,851 dalton, which is very close to its calculated molecular weight. A bioassay based on proliferation of Nb2 rat lymphoma cells showed that the purified cGH was biologically active.