1,721,029 research outputs found
Genotoxic effects of boric acid and borax in zebrafish, Danio rerio using alkaline comet assay
No full textThe present study is conducted to determine the potential mechanisms of Boron compounds, boric acid (BA) and borax (BX), on genotoxicity of zebrafish Danio rerio for 24, 48, 72 and 96-hours acute exposure (level:1, 4, 16, 64 mg/l BA and BX) in semi-static bioassay experiment. For that purpose, peripheral erythrocytes were drawn from caudal vein and Comet assay was applied to assess genotoxicity. Acute (96 hours) exposure and high concentrations of boric acid and borax increases % tail DNA and Olive tail moment. Genotoxicity was found for BA as concentration-dependent and BX as concentration and time dependent manner. In general, significant effects (P < 0,05) on both concentrations and exposure times were observed in experimental groups. DNA damage was highest at 96 h and 24 h for all BX and BA concentrations, respectively in peripheral blood of D. rerio. For the first time, our study demonstrates the effect of waterborne BA and BX exposure on genotoxicity at the molecular level, which may contribute to understanding the mechanism of boric acid and borax-induced genotoxicity in fish
Going Beyond Counting First Authors in Author Co-citation Analysis
Get PDFThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Functional analyses of dipeptide and pentapeptide insertions on Theileria annulata enolase by site-directed mutagenesis and in silico approaches
No full textTheileria annulata enolase (TaENO) could be assessed as a druggable target for tropical theileriosis treatment. The parasite enzyme plays an important role in many cellular functions and carries some structural differences like dipeptide (262EK263) and pentapeptide ((103)EWGYC(107)) insertions from the host enzyme, Bos taurus enolase. In this study, the functional effects of these insertions on TaENO activity were analyzed by in vitro site-directed mutagenesis and in silico molecular docking analyses for the first time in the literature. In vitro results showed that, although the deletion of the pentapeptide insertion (TaENO Delta EWGYC) reduced the enzyme activity slightly, the removal of the dipeptide insertion (TaENO Delta EK) halted it. Also, molecular docking results revealed that the deletion of these insertions affected the substrate binding affinity of the mutant enzymes. The active site of TaENO Delta EK exhibited a small decrease of substrate binding affinity compared to the active site of TaENO Delta EWGYC relative to the wild type TaENO. Although we conclude that both regions could be evaluated as possible drug-binding sites to inhibit TaENO in further studies, these results indicate that the dipeptide insertion could be a more promising drug binding site than the pentapeptide insertion
Cloning, expression and characterization of the gene encoding the enolase from Fusobacterium nucleatum
No full textThe gene encoding enolase from Fusobacterium nucleatum (FnENO) was cloned and analyzed for the first time. The gene comprises of 1302 nucleotide base pairs and encodes 433 amino acids. The gene sequence alignment demonstrated the presence of several distinct insertions and deletions, compared with the human enzyme. The gene for recombinant FnENO was inserted into the pLATE 31 vector system and expressed in E. coli BL21(DE3) cells as a soluble protein. The protein was purified by affinity chromatography using a Ni-NTA agarose matrix and shown on SDS-PAGE to be a 46 kDa protein. The molecular weight of the octameric form of the purified recombinant protein was determined as being 375 kDa by size exclusion chromatography. Optimal enzyme activity was observed at pH 8.5 and the enzyme remained stable at a range of different temperatures from 30 to 60°C. Using 2-phosphoglyceric acid as substrate for the purified enzyme, KM, kcat and kcat/KM were determined as 0.48 mM, 20.4 s–1 and 4.22 × 104 M–1s–1, respectively. Potential drug binding sites of FnENO were detected using homology modeling. These data could facilitate the design of new inhibitors of F. nucleatum which has already been shown to be resistant to several known antibiotics. © 2016, Pleiades Publishing, Inc
Kinetic Analysis of the Amino Terminal End of Active Site Loop of Lactate Deyhdrogenase from Plasmodium Vivax
No full textObjective: In this study, kinetic analysis was performed to understand the functional importance of the amino terminal of the active site of previously mutated Plasmodium vivax Lactate Dehydrogenase enzyme by mimicking Toxoplasma gondii I, II, Eimeria acervulina and Eimeria tenella LDH's. Material and Methods: Mutant LDH genes were amplified by PCR and 6xHistag was added to the C-terminal of the enzymes. Then LDH enzymes are overproduced as recombinant in E. coli cells, purified by Ni-NTA agarose matrix and kinetic properties were analysed. Results: Observing increase of K-m values of mutant enzymes showed that mutations in this place caused decreasing affinity of enzyme for its substrate. However k(cat) values were about the same throughout all mutant proteins. Conclusion: Sensitivity of the studied region emphasizes the significance of this site for drug design studies for both Plasmodium and some other Apicomplexans
Assessment of mitogen-activated protein kinases as therapeutic targets for the treatment of babesiosis and theileriosis
No full textThe Piroplasmida order comprises parasitic protozoa including the Theileria and Babesia species that are transmitted by vector ticks and can cause severe diseases in domestic and wild animals. Because of limited therapies and available drug resistance, the discovery of new, effective, and safer drugs for veterinary use is important. Mitogen-activated protein kinases (MAPK) are a group of serine-threonine protein kinases found in diverse species, including animals and protozoa that conduct vital cellular functions. Therefore, they have been at the centre of drug design studies for many years. Computer-aided structure-based drug design is a fast and effective way in drug discovery efforts to identify candidate compounds. In this study, we conducted comparative sequence analysis of MAPK proteins from the Theileria (T. annulata, T. parva., T. orientalis, and T. equi) and Babesia species (B. bigemina, B. microti, and B. bovis). Three-dimensional protein structures from relevant species (T. annulata and B. bovis) were modelled and compounds were screened for interaction. Results showed that the inhibitors designed for human use could also be potent against Prioplasmida MAPKs. Furthermore, the structural differences between Prioplasmida and mammalian MAPKs could be a way for researchers to better instigate selective drug design
Functional and structural characterization of the pentapeptide insertion of Theileria annulata lactate dehydrogenase by site-directed mutagenesis, comparative modeling and molecular dynamics simulations
No full textLactate dehydrogenase (LDH) is an important metabolic enzyme in glycolysis and it has been considered as the main energy source in many organisms including apicomplexan parasites. Differences at the active site loop of the host and parasite LDH's makes this enzyme an attractive target for drug inhibitors. In this study, five amino acid insertions in the active site pocket of Theileria annulata LDH (TaLDH) were deleted by PCR-based site-directed mutagenesis, expression and activity analysis of mutant and wild type TaLDH enzymes were performed. Removal of the insertion at the active site loop caused production of an inactive enzyme. Furthermore, structures of wild and mutant enzymes were predicted by comparative modeling and the importance of the insertions at the active site loop were also assigned by molecular docking and dynamics simulations in order to evaluate essential role of this loop for the enzymatic activity. Pentapeptide insertion removal resulted in loss of LDH activity due to deletion of Trp96 and conformational change of Arg98 because of loop instability. Analysis of wild type and mutant enzymes with comparative molecular dynamics simulations showed that the fluctuations of the loop residues increase in mutant enzyme. Together with in silico studies, in vitro results revealed that active site loop has a vital role in the enzyme activity and our findings promise hope for the further drug design studies against theileriosis and other apicomplexan parasite diseases. (C) 2017 Elsevier Inc. All rights reserved
Identification of potential inhibitors of Trichomonas vaginalis iron-containing superoxide dismutase by computer-aided drug design approach
No full textTrichomonas vaginalis is a human protozoan parasite that causes trichomoniasis, a common sexually transmitted disease. Metronidazole is a commonly used drug for the treatment of T. vaginalis infections. However, it was reported that the parasite has developed resistance to this drug. Therefore, a necessity of discovering new drugs that have different modes of action against T. vaginalis has emerged. In this computational study, T. vaginalis iron-containing superoxide dismutase (TvSOD) was selected as target protein because of its vital role in converting superoxide to oxygen and hydrogen peroxide and protecting the parasite against toxic reactive oxygen species (ROS). TvSOD was modeled using two different protein modeling programs, MODELLER and SWISS-MODEL, and then, small drug-like chemicals were screened for interaction with three different druggable pockets of the enzyme. The best interacting chemicals were then evaluated through molecular dynamics simulations (MDSs) for ligand stability. As a result, ligand-129817054 (7-(6-amino-1,2,3,4,5-pentahydroxyhexyl)-4-methylchromen-2-one) was determined to be a viable drug candidate based on docking scores and MDS results. Additional in vitro inhibition studies are necessary for the evaluation and assessment of the compound of interest as an effective TvSOD inhibitor
Identification of novel compounds against Acinetobacter baumannii 3-oxoacyl-[acyl-carrier-protein] synthase I (FabB) via comprehensive structure-based computational approaches.
No full textAcinetobacter baumannii is one of the most serious opportunistic pathogens according to WHO. The difference
between bacterial and mammalian fatty acid biosynthesis pathways makes FASII enzymes attractive targets in
drug discovery. 3-oxoacyl-[acyl-carrier-protein] synthase I (FabB) from the FAS II pathway catalyze the
condensation of malonyl ACP with acyl-ACP, and elongates the fatty acid chain by two carbons. To investigate
potential inhibitors of the A. baumannii FabB, we used computational approaches including homology modeling,
high-throughput virtual screening, molecular docking, molecular dynamics simulations, and MM-GBSA free
energy calculations. After the high-throughput virtual screening, the resulting ligands were further screened
using the QM-polarized ligand docking (QPLD) and induced fit docking (IFD) approaches. Molecular dynamics
simulations were performed for 100 ns. And according to binding free energy calculations, we have identified
nine compounds with the best binding affinities. Three of these compounds were selected for an additional 1 μs
MD simulation to assess ligand stability. Two of them named L6 and L7 showed promised stability and affinity to
the target. Here, we present novel compounds against A. baumannii FabB via structure-based computational
approaches. These compounds might pave the way for the design of new lead structures and inhibitors for
multidrug-resistant A. baumannii
- …
