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991.
Mansour S. Alturki Ngolui Rene Fuanta Madison A. Jarrard Judith V. Hobrath Douglas C. Goodwin Thankhoe A. Rantso Angela I. Calderón 《Bioorganic & medicinal chemistry letters》2018,28(4):802-808
Single dose high-throughput screening (HTS) followed by dose-response evaluations is a common strategy for the identification of initial hits for further development. Early identification and exclusion of false positives is a cost-saving and essential step in early drug discovery. One of the mechanisms of false positive compounds is the formation of aggregates in assays. This study evaluates the mechanism(s) of inhibition of a set of 14 compounds identified previously as actives in Mycobacterium tuberculosis (Mt) cell culture screening and in vitro actives in Mt shikimate kinase (MtSK) assay. Aggregation of hit compounds was characterized using multiple experimental methods, LC-MS, 1HNMR, dynamic light scattering (DLS), transmission electron microscopy (TEM), and visual inspection after centrifugation for orthogonal confirmation. Our results suggest that the investigated compounds containing oxadiazole-amide and aminobenzothiazole moieties are false positive hits and non-specific inhibitors of MtSK through aggregate formation. 相似文献
992.
993.
Physiological response to anaerobicity of glycerol-3-phosphate dehydrogenase mutants of Saccharomyces cerevisiae. 总被引:3,自引:0,他引:3
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Mutants of Saccharomyces cerevisiae, in which one or both of the genes encoding the two isoforms of NAD-dependent glycerol-3-phosphate dehydrogenase had been deleted, were studied in aerobic batch cultures and in aerobic-anaerobic step change experiments. The respirofermentative growth rates under aerobic conditions with semisynthetic medium (20 g of glucose per liter) of two single mutants, gpd1 delta and gpd2 delta, and the parental strain (mu = 0.5 h-1) were almost identical, whereas the growth rate of a double mutant, gpd1 delta gpd2 delta, was approximately half that of the parental strain. Upon a step change from aerobic to anaerobic conditions in the exponential growth phase, the specific carbon dioxide evolution rates (CER) of the wild-type strain and the gpd1 delta strain were almost unchanged. The gpd2 delta mutant showed an immediate, large (> 50%) decrease in CER upon a change to anaerobic conditions. However, after about 45 min the CER increased again, although not to the same level as under aerobic conditions. The gpd1 delta gpd2 delta mutant showed a drastic fermentation rate decrease upon a transition to anaerobic conditions. However, the CER values increased to and even exceeded the aerobic levels after the addition of acetoin. High-pressure liquid chromatographic analyses demonstrated that the added acetoin served as an acceptor of reducing equivalents by being reduced to butanediol. The results clearly show the necessity of glycerol formation as a redox sink for S. cerevisiae under anaerobic conditions. 相似文献
994.
Emili Saló Ana Maria Muñoz-Mármol José Ramon Bayascas-Ramirez Jordi Garcia-Fernàndez Agusti Miralles Andreu Casali Montserrat Corominas Jaume Baguñá 《Hydrobiologia》1995,305(1-3):269-275
To identify potential pattern control and cell determination and/or differentiation genes in the freshwater planarian Dugesial (G.) tigrina, we searched for homeobox genes of different types in the genome of this primitive metazoan. We applied two basic approaches: 1) Screening the cDNA library with degenerate oligonucleotides corresponding to the most conserved amino acid sequence from helix-3 of the homeodomain of each family; and 2) PCR amplification of genomic DNA or cDNA, using two sets of degenerated oligonucleotides corresponding to helices 1 and 3 of the homeodomain or two specific domains of the POU family. Using the first strategy we have identified and characterized two tissue-specific cell determination and/or differentiation NK-type homeobox genes. Using the second strategy we have identified several homeobox genes that belong to the HOM/Hox, paired (prd) or POU families. 相似文献
995.
We have combined high-resolution two-dimensional (2-D) gel electrophoresis and mass spectrometry with the aim of identifying proteins represented in the 2-D gel database of the wing imaginal discs of Drosophila melanogaster. First, we obtained a high-resolution 2-D gel pattern of [35S]methionine + [35S]cysteine-labeled polypeptides of Schneider cells, a permanent cell line of Drosophila embryonic origin, and compared it with the standard pattern of polypeptides of the wing imaginal disc. These studies reveal qualitative and quantitative differences between the two samples, but have more than 600 polypeptides in common. Second, we carried out preparative 2-D polyacrylamide gel electrophoresis using Schneider cells mixed with radioactively labeled wing imaginal discs in order to isolate some of the shared polypeptides and characterize them by matrix-assisted laser desorption/ionization-time of flight MALDI-TOF analysis. Using this strategy we identified 100 shared proteins represented in the database, and in each case confirmed their identity by MALDI-TOF/TOF analysis. 相似文献
996.
Aqueous, methanol and dichloromethane extracts from Artemisia copa, Baccharis grisebachii, Baccharis incarum, Baccharis latifolia, Mutisia kurtzii and Pluchea sagittalis, plants used in the Traditional Medicine of South America, are studied for activity on the respiratory burst and the inducible heat shock protein of 72 kD (hsp72) synthesis. Activity on the production of reactive oxygen species (ROS) and reactive nitrogen species (RNS), as well as on hsp72 synthesis was measured by flow cytometry in human neutrophils. Cells were stimulated using hydrogen peroxide, phorbol-12-myristate-13-acetate (PMA) or formyl-methionyl-leucyl-phenylalanine (FMLP) for ROS generation, and sodium nitroprusside (SNP) or PMA in the presence of calmodulin inhibitor W-13 for RNS. The production of hsp72 was induced by heat, PMA, H2O2 and SNP. The best inhibitory activity was shown by the dichloromethane extracts of Baccharis grisebachii and Pluchea sagittalis that were active in all the assays. The aqueous extract of Pluchea sagittalis was also active in most assays. The aqueous extract from Mutisia kurtzii caused a clear increase of the hsp72 production and showed prooxidant activity. 相似文献
997.
Background
Observers respond more accurately to targets in visual search tasks that share properties with previously presented items, and transient attention can learn featural consistencies on a precue, irrespective of its absolute location.Methodology/Principal Findings
We investigated whether such attentional benefits also apply to temporal consistencies. Would performance on a precued Vernier acuity discrimination task, followed by a mask, improve if the cue-lead times (CLTs; 50, 100, 150 or 200 ms) remained constant between trials compared to when they changed? The results showed that if CLTs remained constant for a few trials in a row, Vernier acuity performance gradually improved while changes in CLT from one trial to the next led to worse than average discrimination performance. The results show that transient attention can quickly adjust to temporal regularities, similarly to spatial and featural regularities. Further experiments show that this form of learning is not under voluntary control.Conclusions/Significance
The results add to a growing literature showing how consistency in visual presentation improves visual performance, in this case temporal consistency. 相似文献998.
von Bülow R Schmidt B Dierks T von Figura K Usón I 《Journal of molecular biology》2001,305(2):269-277
Arylsulfatase A (ASA) belongs to the sulfatase family whose members carry a C(alpha)-formylglycine that is post-translationally generated by oxidation of a conserved cysteine or serine residue. The crystal structures of two arylsulfatases, ASA and ASB, and kinetic studies on ASA mutants led to different proposals for the catalytic mechanism in the hydrolysis of sulfate esters.The structures of two ASA mutants that lack the functional C(alpha)-formylglycine residue 69, in complex with a synthetic substrate, have been determined in order to unravel the reaction mechanism. The crystal structure of the inactive mutant C69A-ASA in complex with p-nitrocatechol sulfate (pNCS) mimics a reaction intermediate during sulfate ester hydrolysis by the active enzyme, without the covalent bond to the key side-chain FGly69. The structure shows that the side-chains of lysine 123, lysine 302, serine 150, histidine 229, the main-chain of the key residue 69 and the divalent cation in the active center are involved in sulfate binding. It is proposed that histidine 229 protonates the leaving alcoholate after hydrolysis.C69S-ASA is able to bind covalently to the substrate and hydrolyze it, but is unable to release the resulting sulfate. Nevertheless, the resulting sulfation is low. The structure of C69S-ASA shows the serine side-chain in a single conformation, turned away from the position a substrate occupies in the complex. This suggests that the double conformation observed in the structure of wild-type ASA is more likely to correspond to a formylglycine hydrate than to a twofold disordered aldehyde oxo group, and accounts for the relative inertness of the C69S-ASA mutant. In the C69S-ASA-pNCS complex, the substrate occupies the same position as in the C69A-ASA-pNCS complex, which corresponds to the non-covalently bonded substrate. Based on the structural data, a detailed mechanism for sulfate ester cleavage is proposed, involving an aldehyde hydrate as the functional group. 相似文献
999.
Sayós J Martínez-Barriocanal A Kitzig F Bellón T López-Botet M 《Biochemical and biophysical research communications》2004,324(2):640-647
The CD85j inhibitory receptor (also termed ILT2 or LIR-1) is a type-I transmembrane protein that belongs to the Ig superfamily and is expressed by different leukocyte lineages. The extracellular region of CD85j binds HLA class I molecules and its cytoplasmic domain displays four immunoreceptor tyrosine-based inhibition motifs (ITIM). Upon tyrosine phosphorylation CD85j recruits the SHP-1 tyrosine phosphatase, involved in negative signaling. In order to identify other molecules to which CD85j might interact with in a phosphotyrosine-dependent manner, a cDNA B-cell library was screened in a three-hybrid system in yeast using the CD85j cytoplasmic tail as bait in the presence of the Src-kinase c-fyn420, 531Y-F, 176R-Q mutant. In this system, the C-terminal Src kinase (Csk) was shown to interact with CD85j. Phosphorylation-dependent recruitment of Csk to the CD85j cytoplasmic tail was confirmed in CD85j-transfected mammalian cells by immunoprecipitation and Western blot analysis. Mutational analyses and phospho-peptide mapping suggested that the SH2 domain of Csk may preferentially bind to ITIM Y562 of CD85j; yet, mutation to phenylalanine of Y533, Y614, and Y644 also significantly reduced Csk recruitment by CD85j. Even though CD85j was detected in both anti-SHP1 and CSK immunoprecipitates, these two molecules did not co-precipitate together with CD85j. Our data support the possibility that Csk regulates the function of CD85j. 相似文献
1000.
Duplicated sequence motif in the long terminal repeat of maedi-visna virus extends cell tropism and is associated with neurovirulence 总被引:2,自引:0,他引:2
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Oskarsson T Hreggvidsdóttir HS Agnarsdóttir G Matthíasdóttir S Ogmundsdóttir MH Jónsson SR Georgsson G Ingvarsson S Andrésson OS Andrésdóttir V 《Journal of virology》2007,81(8):4052-4057
Maedi-visna virus (MVV) is a lentivirus of sheep causing chronic inflammatory disease of the lungs (maedi) and the nervous system (visna). We have previously shown that a duplicated sequence in the long terminal repeat (LTR) of MVV is a determinant of cell tropism. Here, we demonstrate that deletion of a CAAAT sequence from either one of the repeats resulted in poor virus growth in sheep choroid plexus cells. A duplication in the LTR encompassing the CAAAT sequence was found in four neurological field cases that were sequenced, but no duplication was present in the LTRs from seven maedi cases; one maedi isolate was mixed. These results indicate that the duplication in the LTR is associated with neurovirulence. 相似文献