Evaluation of Abelmoschus moschatus extracts for antioxidant, free radical scavenging, antimicrobial and antiproliferative activities using in vitro assays

<p>Abstract</p> <p>Background</p> <p><it>Abelmoschus moschatus </it>Medik. leaves and seeds are considered as valuable traditional medicine. The aromatic seeds of this plant are aphrodisiac, ophthalmic, cardio tonic, antispasmodic and used in the treatment of intestinal complaints and check queasiness. To give a scientific basis for traditional usage of this medicinal plant, the seed and leaf extracts were evaluated for their antioxidant, free radical scavenging, antimicrobial and antiproliferative activities.</p> <p>Methods</p> <p>In this study, antioxidant, antimicrobial and antiproliferative activities of <it>A. moschatus </it>extracts were evaluated in a series of <it>in vitro </it>assay involving free radicals, reactive oxygen species and their IC₅₀values were also determined. The antioxidant activities of the seed and leaf extracts of <it>A. moschatus </it>were determined by total antioxidant, DPPH, and ferrous reducing antioxidant property (FRAP) methods. In addition, the antiproliferative activity was also evaluated using colorectal adenocarcinoma and retinoblastoma human cancer cell lines. Moreover, six bacterial reference strains, two gram-positive (<it>Bacillus subtilis </it>and <it>Staphylococcus aureus</it>), four gram-negative (<it>Escherichia coli, Pseudomonas aeruginosa, Proteus vulgaris </it>and <it>Salmonella enterica paratyphi</it>) and one fungal strain (<it>Candida albicans</it>) were used to evaluate its antimicrobial activity.</p> <p>Results</p> <p>The results from this study showed that the antioxidant activities of <it>A. moschatus </it>as determined by the total phenol, flavonoids, total antioxidant and FRAP methods were higher in leaf than that of the seed extracts. On the other hand, the aqueous overnight seed extract (AMS-I) has shown significant radical scavenging activity as in 1, 1- Diphenyl-2-picrylhydrazyl (DPPH), hydrogen peroxide, hydroxyl radical, superoxide and lipid peroxidation as compared to other seed and leaf extracts. The AMS-I and AML-IV have shown activity against six and seven microorganisms respectively. Simulteneously, AMS-IV and AML-IV have demonstrated potential antiproliferative activity against two human cell lines - Colorectal adenocarcinoma (COLO-205) and retinoblastoma (Y79).</p> <p>Conclusion</p> <p>The seed and leaf extracts of <it>A. moschatus </it>possess significant antioxidant activity and could serve as free radical inhibitors or scavenger, or substitute, probably as primary antioxidants. The plant possesses moderate antibacterial activity against bacterial strains used in this study. Hydroalcoholic seed and leaf extracts also exhibited antiproliferative activity against two human cancer cell lines. <it>A. moschatus </it>may therefore, be a good candidate for functional foods as well as pharmaceutics.</p

Mycobacterium tuberculosis DosR Regulon Gene Rv0079 Encodes a Putative, ‘Dormancy Associated Translation Inhibitor (DATIN)’

Mycobacterium tuberculosis is a major human pathogen that has evolved survival mechanisms to persist in an immune-competent host under a dormant condition. The regulation of M. tuberculosis metabolism during latent infection is not clearly known. The dormancy survival regulon (DosR regulon) is chiefly responsible for encoding dormancy related functions of M. tuberculosis. We describe functional characterization of an important gene of DosR regulon, Rv0079, which appears to be involved in the regulation of translation through the interaction of its product with bacterial ribosomal subunits. The protein encoded by Rv0079, possibly, has an inhibitory role with respect to protein synthesis, as revealed by our experiments. We performed computational modelling and docking simulation studies involving the protein encoded by Rv0079 followed by in vitro translation and growth curve analysis experiments, involving recombinant E. coli and Bacille Calmette Guérin (BCG) strains that overexpressed Rv0079. Our observations concerning the interaction of the protein with the ribosomes are supportive of its role in regulation/inhibition of translation. We propose that the protein encoded by locus Rv0079 is a ‘dormancy associated translation inhibitor’ or DATIN

Concurrent Proinflammatory and Apoptotic Activity of a Helicobacter pylori Protein (HP986) Points to Its Role in Chronic Persistence

Helicobacter pylori induces cytokine mediated changes in gastroduodenal pathophysiology, wherein, the activated macrophages at the sub-mucosal space play a central role in mounting innate immune response against the antigens. The bacterium gains niche through persistent inflammation and local immune-suppression causing peptic ulcer disease or chronic gastritis; the latter being a significant risk factor for the development of gastric adenocarcinoma. What favors persistence of H. pylori in the gastric niches is not clearly understood. We report detailed characterization of a functionally unknown gene (HP986), which was detected in patient isolates associated with peptic ulcer and gastric carcinoma. Expression and purification of recombinant HP986 (rHP986) revealed a novel, ∼29 kDa protein in biologically active form which associates with significant levels of humoral immune responses in diseased individuals (p<0.001). Also, it induced significant levels of TNF-α and Interleukin-8 in cultured human macrophages concurrent to the translocation of nuclear transcription factor-κB (NF-κB). Further, the rHP986 induced apoptosis of cultured macrophages through a Fas mediated pathway. Dissection of the underlying signaling mechanism revealed that rHP986 induces both TNFR1 and Fas expression to lead to apoptosis. We further demonstrated interaction of HP986 with TNFR1 through computational and experimental approaches. Independent proinflammatory and apoptotic responses triggered by rHP986 as shown in this study point to its role, possibly as a survival strategy to gain niche through inflammation and to counter the activated macrophages to avoid clearance

AI is a viable alternative to high throughput screening: a 318-target study

: High throughput screening (HTS) is routinely used to identify bioactive small molecules. This requires physical compounds, which limits coverage of accessible chemical space. Computational approaches combined with vast on-demand chemical libraries can access far greater chemical space, provided that the predictive accuracy is sufficient to identify useful molecules. Through the largest and most diverse virtual HTS campaign reported to date, comprising 318 individual projects, we demonstrate that our AtomNet® convolutional neural network successfully finds novel hits across every major therapeutic area and protein class. We address historical limitations of computational screening by demonstrating success for target proteins without known binders, high-quality X-ray crystal structures, or manual cherry-picking of compounds. We show that the molecules selected by the AtomNet® model are novel drug-like scaffolds rather than minor modifications to known bioactive compounds. Our empirical results suggest that computational methods can substantially replace HTS as the first step of small-molecule drug discovery

Purification, identification and preliminary crystallographic studies of an allergenic protein from Lathyrus sativus

A 24 kDa protein was purified from the seeds of L. sativus by ammonium sulfate fractionation and ion-exchange chromatography. Crystals were obtained by the hanging-drop vapour-diffusion method

Crystallographic structure of ubiquitin in complex with cadmium ions

Abstract Background Ubiquitination plays a critical role in regulating many cellular processes, from DNA repair and gene transcription to cell cycle and apoptosis. It is catalyzed by a specific enzymatic cascade ultimately leading to the conjugation of ubiquitin to lysine residues of the target protein that can be the ubiquitin molecule itself and to the formation of poly-ubiquitin chains. Findings We present the crystal structure at 3.0 Å resolution of bovine ubiquitin crystallized in presence of cadmium ions. Two molecules of ubiquitin are present in the asymmetric unit. Interestingly this non-covalent dimeric arrangement brings Lys-6 and Lys-63 of each crystallographically-independent monomer in close contact with the C-terminal ends of the other monomer. Residues Leu-8, Ile-44 and Val-70 that form a hydrophobic patch at the surface of the Ub monomer are trapped at the dimer interface. Conclusions The structural basis for signalling by poly-Ub chains relies on a visualization of conformations of alternatively linked poly-Ub chains. This arrangement of ubiquitin could illustrate how linkages involving Lys-6 or Lys-63 of ubiquitin are produced in the cell. It also details how ubiquitin molecules can specifically chelate cadmium ions.</p

Antioxidant and antiproliferative activities of Abrus precatorius leaf extracts - an in vitro study

BACKGROUND: The use of traditional medicine at the primary health care level is widespread and plant-based treatments are being recommended for curing various diseases by traditional medical practitioners all over the world. The phytochemicals present in the fruits, vegetables and medicinal plants are getting attention day-by-day for their active role in the prevention of several human diseases. Abrus precatorius is a widely distributed tropical medicinal plant with several therapeutic properties. Therefore in the present study, A. precatorius leaf extracts were examined for their antioxidant and cytotoxic properties in vitro in order to discover resources for new lead structures or to improve the traditional medicine. METHODS: In this study, antioxidant and antiproliferative properties of the different leaf extracts (hexane, ethyl acetate, ethanol and water) from A. precatorius were investigated along with the quantification of the polyphenol and flavonoid contents. The ability of deactivating free radicals was extensively investigated with in vitro biochemical methods like DPPH(•), (•)OH, NO, SO(2-) scavenging assays and inhibition capability of Fe(II)-induced lipid peroxidation. Furthermore, antiproliferative activities using different human cancer cell lines and primary cell line was carried out by MTT method. RESULTS: Total phenolic content and total flavonoid content of the extracts were found in the range of 1.65 ± 0.22 to 25.48 ± 0.62 GAE mg/g dw and 6.20 ± 0.41 to 17.16 ± 1.04 QE mg/g dw respectively. The experimental results further revealed that A. precatorius extracts showed strong antiradical properties, capable to chelate Fe(2+) and possess good inhibition ability of lipid peroxidation. In addition, as a first step towards the identification of phytoconstituents endowed with potent chemopreventive activities, we evaluated the inhibitory effects of A. precatorius extracts on the proliferation of four different human tumour cell lines such as human colon adenocarcinoma cells (Colo-205), human retinoblastoma cancer cells (Y79), human hepatocellular carcinoma cells (HepG2) and Leukemia cells (SupT1). Ethanol extract (APA) and ethyl acetate extract (APE) of A. precatorius had apparent capabilities of inhibiting the survival of tested human cancer cell lines. Moreover, it was observed that the A. precatorius extracts did not inhibit the growth of mice peritoneal macrophages, thus confirming that plants extracts are selective against the cancer cell lines. CONCLUSION: This work provides a scientific support for the high antioxidant and antiproliferative activity of this plant and thus it may find potential applications in the treatment of the diseases caused by ROS. Further studies are needed to confirm in vivo anti-tumorgenicity and subsequent chemical characterization of the active molecule(s)