scholarly journals A Comprehensive Assessment of the Genetic Determinants inSalmonellaTyphimurium for Resistance to Hydrogen Peroxide Using Proteogenomics

2017 ◽  
Author(s):  
Sardar Karash ◽  
Rohana Liyanage ◽  
Abdullah Qassab ◽  
Jackson O. Lay ◽  
Young Min Kwon

AbstractSalmonellais an intracellular pathogen that infects a wide range of hosts and can survive in macrophages. An essential mechanism uses by the macrophages to eradicateSalmonellais production of reactive oxygen species. Here, we used proteogenomics to determine the candidate genes and proteins that have a role in resistance ofS.Typhimurium to H2O2. For Tn-seq, a highly saturated Tn5 insertion library was grownin vitrounder either 2.5 (H2O2L) or 3.5 mM H2O2(H2O2H). We identified two sets of overlapping genes that are required for resistance ofS. Typhimurium to H2O2L and H2O2H, and the results were validated via phenotypic evaluation of 50 selected mutants. The enriched pathways for resistance to H2O2included DNA repair, aromatic amino acid biosynthesis (aroBK), Fe-S cluster biosynthesis, iron homeostasis and a putative iron transporter system (ybbKLM), flagellar genes (fliBC), H2O2scavenging enzymes, and DNA adenine methylase. Proteomics revealed that the majority of essential proteins, including ribosomal proteins, were downregulated upon exposure to H2O2. A subset of proteins identified by Tn-seq were analyzed by targeted proteomics, and 70% of them were upregulated upon exposure to H2O2. The identified candidate genes will deepen our understanding about mechanisms ofS. Typhimurium survival in macrophages, and can be exploited to develop new antimicrobial drugs.

2014 ◽  
Vol 27 (11) ◽  
pp. 1226-1240 ◽  
Author(s):  
Emmanuel Koen ◽  
Pauline Trapet ◽  
Daphnée Brulé ◽  
Anna Kulik ◽  
Agnès Klinguer ◽  
...  

β-Aminobutyric acid (BABA) is a nonprotein amino acid inducing resistance in many different plant species against a wide range of abiotic and biotic stresses. Nevertheless, how BABA primes plant natural defense reactions remains poorly understood. Based on its structure, we hypothesized and confirmed that BABA is able to chelate iron (Fe) in vitro. In vivo, we showed that it led to a transient Fe deficiency response in Arabidopsis thaliana plants exemplified by a reduction of ferritin accumulation and disturbances in the expression of genes related to Fe homeostasis. This response was not correlated to changes in Fe concentrations, suggesting that BABA affects the availability or the distribution of Fe rather than its assimilation. The phenotype of BABA-treated plants was similar to those of plants cultivated in Fe-deficient conditions. A metabolomic analysis indicated that both BABA and Fe deficiency induced the accumulation of common metabolites, including p-coumaroylagmatine, a metabolite previously shown to be synthesized in several plant species facing pathogen attack. Finally, we showed that the protective effect induced by BABA against Botrytis cinerea was mimicked by Fe deficiency. In conclusion, the Fe deficiency response caused by BABA could bring the plant to a defense-ready state, participating in the plant resistance against the pathogens.


Author(s):  
R.A. Milligan ◽  
P.N.T. Unwin

A detailed understanding of the mechanism of protein synthesis will ultimately depend on knowledge of the native structure of the ribosome. Towards this end we have investigated the low resolution structure of the eukaryotic ribosome embedded in frozen buffer, making use of a system in which the ribosomes crystallize naturally.The ribosomes in the cells of early chicken embryos form crystalline arrays when the embryos are cooled at 4°C. We have developed methods to isolate the stable unit of these arrays, the ribosome tetramer, and have determined conditions for the growth of two-dimensional crystals in vitro, Analysis of the proteins in the crystals by 2-D gel electrophoresis demonstrates the presence of all ribosomal proteins normally found in polysomes. There are in addition, four proteins which may facilitate crystallization. The crystals are built from two oppositely facing P4 layers and the predominant crystal form, accounting for >80% of the crystals, has the tetragonal space group P4212, X-ray diffraction of crystal pellets demonstrates that crystalline order extends to ~ 60Å.


1991 ◽  
Vol 30 (01) ◽  
pp. 35-39 ◽  
Author(s):  
H. S. Durak ◽  
M. Kitapgi ◽  
B. E. Caner ◽  
R. Senekowitsch ◽  
M. T. Ercan

Vitamin K4 was labelled with 99mTc with an efficiency higher than 97%. The compound was stable up to 24 h at room temperature, and its biodistribution in NMRI mice indicated its in vivo stability. Blood radioactivity levels were high over a wide range. 10% of the injected activity remained in blood after 24 h. Excretion was mostly via kidneys. Only the liver and kidneys concentrated appreciable amounts of radioactivity. Testis/soft tissue ratios were 1.4 and 1.57 at 6 and 24 h, respectively. Testis/blood ratios were lower than 1. In vitro studies with mouse blood indicated that 33.9 ±9.6% of the radioactivity was associated with RBCs; it was washed out almost completely with saline. Protein binding was 28.7 ±6.3% as determined by TCA precipitation. Blood clearance of 99mTc-l<4 in normal subjects showed a slow decrease of radioactivity, reaching a plateau after 16 h at 20% of the injected activity. In scintigraphic images in men the testes could be well visualized. The right/left testis ratio was 1.08 ±0.13. Testis/soft tissue and testis/blood activity ratios were highest at 3 h. These ratios were higher than those obtained with pertechnetate at 20 min post injection.99mTc-l<4 appears to be a promising radiopharmaceutical for the scintigraphic visualization of testes.


1997 ◽  
Vol 77 (04) ◽  
pp. 725-729 ◽  
Author(s):  
Mario Colucci ◽  
Silvia Scopece ◽  
Antonio V Gelato ◽  
Donato Dimonte ◽  
Nicola Semeraro

SummaryUsing an in vitro model of clot lysis, the individual response to a pharmacological concentration of recombinant tissue plasminogen activator (rt-PA) and the influence on this response of the physiological variations of blood parameters known to interfere with the fibrinolytic/thrombolytic process were investigated in 103 healthy donors. 125I-fibrin labelled blood clots were submersed in autologous plasma, supplemented with 500 ng/ml of rt-PA or solvent, and the degree of lysis was determined after 3 h of incubation at 37° C. Baseline plasma levels of t-PA, plasminogen activator inhibitor 1 (PAI-1), plasminogen, α2-anti-plasmin, fibrinogen, lipoprotein (a), thrombomodulin and von Willebrand factor as well as platelet and leukocyte count and clot retraction were also determined in each donor. rt-PA-induced clot lysis varied over a wide range (28-75%) and was significantly related to endogenous t-PA, PAI-1, plasminogen (p <0.001) and age (p <0.01). Multivariate analysis indicated that both PAI-1 antigen and plasminogen independently predicted low response to rt-PA. Surprisingly, however, not only PAI-1 but also plasminogen was negatively correlated with rt-PA-ginduced clot lysis. The observation that neutralization of PAI-1 by specific antibodies, both in plasma and within the clot, did not potentiate clot lysis indicates that the inhibitor, including the platelet-derived form, is insufficient to attenuate the thrombolytic activity of a pharmacological concentration of rt-PA and that its elevation, similarly to the elevation of plasminogen, is not the cause of clot resistance but rather a coincident finding. It is concluded that the in vitro response of blood clots to rt-PA is poorly influenced by the physiological variations of the examined parameters and that factors other than those evaluated in this study interfere with clot dissolution by rt-PA. In vitro clot lysis test might help to identify patients who may be resistant to thrombolytic therapy.


Growth regulators, phytohormones, both natural and artificial, are the main means to control plant ontogenesis. They are involved in regulating the processes of cell differentiation and cell divisions, the formation of tissues and organs, the changes in the rate of growth and development, the duration of the certain stages of ontogenesis. The main classes of phytohormones used in plant biotechnology, in particular, in the induction of haploid structures, are auxins and cytokinins. The mechanism of action of phytohormones on a cell is rather complicated and may have a different character. Understanding the characteristics of the action of phytohormones is complicated by the fact that the system of hormonal regulation of plant life is multicomponent. This is manifested in the fact that the same physiological process is most often influenced not by one, but by several phytohormones, covering a wide range of aspects of cell metabolism. In connection with the foregoing, the purpose of our work was to test a set of nutrient media with different basic composition and different proportions of phytohormones to determine the patterns of their influence on the processes of haploid structure induction in rape anther culture using accessions, developed at the Institute of Oilseed Crops NAAS. The material used was two accessions of winter rapeseed (No. 1 and No. 2) and one sample of spring rapeseed, provided by the Rapeseed Breeding laboratory of the Institute of Oilseed Crops. Incised inflorescences were kept against the background of low temperature of 6–8 ° C for several days, and then, under aseptic conditions, anthers with unripe pollen grains were isolated and planted on nutrient media differing in both basic mineral composition and content of phytohormones. MS (Murashige & Skoog 1962) and B5 (Gamborg et al 1968) media were used as basic media. Phytohormones were added to the basic media in various combinations – BA, 2,4-D, NAA at the concentrations of 0.1-0.6 mg/l. In each treatment up to 300 anthers were cultivated. Differences between treatments were evaluated using standard t-test. Studies have shown that in the anther culture of rapeseed on the tested nutrient media, morphogenic structures of different types (embryoids and callus) were originated. Synthetic auxin 2,4-D, regardless of the composition of the basic medium, caused the formation of structures of both types, though with a low frequency. Phytohormone BA of the cytokinin type had a similar effect. In this case, the frequency of structures was slightly higher, and the developed structures were represented mainly by embryoids. The joint action of cytokinin and auxin was the most favorable for the initiation of morphogenic structures. Such combination of phytohormones caused the formation of these structures with a frequency of 24.5-14.7% in the studied genotypes of winter rape. A similar effect of phytohormones on the induction and development of morphogenic structures was also observed in spring rape. In this case, a single basic MS medium was used. The experiment included treatments where phytohormones were absent (control), as well as various combinations of auxin and cytokinin. In the control treatment, the formation of new structures was not noted. In treatments with phytohormones, in addition to the medium with the combination of auxin and cytokinin, the medium in which only cytokinin was present was also rather effective. The treatment in which the action of auxin 2,4-D was combined with the action of another auxin, NAA, turned out to be practically ineffective. Thus, it was found that for the induction of morphogenic structures from microspores in rape anther culture of the tested genotypes, the combination of cytokinin with auxin, or the use of only single cytokinin BA without other phytohormones, had the most positive effect.


Author(s):  
Roohi Mohi-ud-din ◽  
Reyaz Hassan Mir ◽  
Prince Ahad Mir ◽  
Saeema Farooq ◽  
Syed Naiem Raza ◽  
...  

Background: Genus Berberis (family Berberidaceae), which contains about 650 species and 17 genera worldwide, has been used in folklore and various traditional medicine systems. Berberis Linn. is the most established group among genera with around 450-500 species across the world. This comprehensive review will not only help researchers for further evaluation but also provide substantial information for future exploitation of species to develop novel herbal formulations. Objective: The present review is focussed to summarize and collect the updated review of information of Genus Berberis species reported to date regarding their ethnomedicinal information, chemical constituents, traditional/folklore use, and reported pharmacological activities on more than 40 species of Berberis. Conclusion: A comprehensive survey of the literature reveals that various species of the genus possess various phytoconstituents mainly alkaloids, flavonoid based compounds isolated from different parts of a plant with a wide range of pharmacological activities. So far, many pharmacological activities like anti-cancer, anti-hyperlipidemic, hepatoprotective, immunomodulatory, anti-inflammatory both in vitro & in vivo and clinical study of different extracts/isolated compounds of different species of Berberis have been reported, proving their importance as a medicinal plant and claiming their traditional use.


2019 ◽  
Vol 20 (14) ◽  
pp. 1203-1212
Author(s):  
Abdelmonaem Messaoudi ◽  
Manel Zoghlami ◽  
Zarrin Basharat ◽  
Najla Sadfi-Zouaoui

Background & Objective: Pseudomonas aeruginosa shows resistance to a large number of antibiotics, including carbapenems and third generation cephalosporin. According to the World Health Organization global report published in February 2017, Pseudomonas aeruginosa is on the priority list among resistant bacteria, for which new antibiotics are urgently needed. Peptidoglycan serves as a good target for the discovery of novel antimicrobial drugs. Methods: Biosynthesis of peptidoglycan is a multi-step process involving four mur enzymes. Among these enzymes, UDP-N-acetylmuramate-L-alanine ligase (MurC) is considered to be an excellent target for the design of new classes of antimicrobial inhibitors in gram-negative bacteria. Results: In this study, a homology model of Pseudomonas aeruginosa MurC ligase was generated and used for virtual screening of chemical compounds from the ZINC Database. The best screened inhibitor i.e. N, N-dimethyl-2-oxo-2,3-dihydro-1H-1,3-benzodiazole-5-sulfonamide was then validated experimentally through inhibition assay. Conclusion: The presented results based on combined computational and in vitro analysis open up new horizons for the development of novel antimicrobials against this pathogen.


2019 ◽  
Vol 20 (12) ◽  
pp. 1227-1243
Author(s):  
Hina Qamar ◽  
Sumbul Rehman ◽  
D.K. Chauhan

Cancer is the second leading cause of morbidity and mortality worldwide. Although chemotherapy and radiotherapy enhance the survival rate of cancerous patients but they have several acute toxic effects. Therefore, there is a need to search for new anticancer agents having better efficacy and lesser side effects. In this regard, herbal treatment is found to be a safe method for treating and preventing cancer. Here, an attempt has been made to screen some less explored medicinal plants like Ammania baccifera, Asclepias curassavica, Azadarichta indica, Butea monosperma, Croton tiglium, Hedera nepalensis, Jatropha curcas, Momordica charantia, Moringa oleifera, Psidium guajava, etc. having potent anticancer activity with minimum cytotoxic value (IC50 >3μM) and lesser or negligible toxicity. They are rich in active phytochemicals with a wide range of drug targets. In this study, these medicinal plants were evaluated for dose-dependent cytotoxicological studies via in vitro MTT assay and in vivo tumor models along with some more plants which are reported to have IC50 value in the range of 0.019-0.528 mg/ml. The findings indicate that these plants inhibit tumor growth by their antiproliferative, pro-apoptotic, anti-metastatic and anti-angiogenic molecular targets. They are widely used because of their easy availability, affordable price and having no or sometimes minimal side effects. This review provides a baseline for the discovery of anticancer drugs from medicinal plants having minimum cytotoxic value with minimal side effects and establishment of their analogues for the welfare of mankind.


Author(s):  
Shangfei Wei ◽  
Tianming Zhao ◽  
Jie Wang ◽  
Xin Zhai

: Allostery is an efficient and particular regulatory mechanism to regulate protein functions. Different from conserved orthosteric sites, allosteric sites have distinctive functional mechanism to form the complex regulatory network. In drug discovery, kinase inhibitors targeting the allosteric pockets have received extensive attention for the advantages of high selectivity and low toxicity. The approval of trametinib as the first allosteric inhibitor validated that allosteric inhibitors could be used as effective therapeutic drugs for treatment of diseases. To date, a wide range of allosteric inhibitors have been identified. In this perspective, we outline different binding modes and potential advantages of allosteric inhibitors. In the meantime, the research processes of typical and novel allosteric inhibitors are described briefly in terms of structureactivity relationships, ligand-protein interactions and in vitro and in vivo activity. Additionally, challenges as well as opportunities are presented.


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