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  • 1
    Electronic Resource
    Electronic Resource
    Structure and function of proteins of the phosphotransferase system and of 6-phospho-β-glycosidases in Gram-positive bacteria (1993)
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology reviews 12 (1993), S. 0 
    Details
    ISSN: 1574-6976
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Abstract: New information about the proteins of the phosphotransferase system (PTS) and of phosphoglycosidases of homofermentative lactic acid bacteria and related species is presented. Tertiary structures were elucidated from soluble PTS components. They help to understand regulatory processes and PTS function in lactic acid bacteria. A tertiary structure of a membrane-bound enzyme II is still not available, but expression of Gram-positive genes encoding enzymes II can be achieved in Escherichia coli and enables the development of effective isolation procedures which are necessary for crystallization experiments. Considerable progress was made in analysing the functions of structural genes which are in close vicinity of the genes encoding the sugar-specific PTS components, such as the genes encoding the tagatose-6-P pathway and the 6-phospho-β-glycosidases. These phosphoglycosidases belong to a subfamily of the β-glycosidase family I among about 300 different glycosidases. The active site nucleophile was recently identified to be Glu 358 in Agrobacteriumβ-glucosidase. This corresponds to Glu 375 in staphylococcal and lactococcal 6-phospho-β-galactosidase. This enzyme is inactivated by mutating Glu 375 to Gln. Diffracting crystals of the lactococcal 6-P-β-galactosidase allow the elucidation of its tertiary structure which helps to derive the structures for the entire glycosidase family 1. In addition, a fusion protein with 6-phospho-β-galactosidase and staphylococcal protein A was constructed.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1574-6976.1993.tb00016.x
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  • 2
    Electronic Resource
    Electronic Resource
    Fibroblast growth factor 2, heparin and suramin reduce epithelial ulcer development in experimental HSV-1 keratitis (1997)
    Springer
    Graefe's archive for clinical and experimental ophthalmology 235 (1997), S. 733-740 
    Details
    ISSN: 1435-702X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract • Background: We have previously shown that basic fibroblast growth factor (FGF-2) enhances corneal epithelial healing in different experimental models in vivo. In order to study the healing effect of this growth factor in pathological conditions of the cornea, we investigated whether topical application of FGF-2 could affect herpes keratitis in rabbits. Since HSV-1 infection is prevented in vitro by incubation with heparin, we also topically applied heparin and suramin, considering the similar interaction of herpes simplex virus and FGF-2 with cell membrane-anchored heparan sulfate. • Methods: After virus inoculation with a human BEY.2 strain, rabbits were treated with either FGF-2 (200 ng to 2 μg/application), heparin (250 μg/application) or suramin (250 μg/application) 4 times daily until day 14. Acyclovir and placebo administrations served as controls (n=48 rabbits). Computerized ulcer surface analysis, clinical observations and virus recovery assays were performed. • Results: Topical FGF-2, heparin and suramin treatment revealed a significant reduction in peak ulcer sizes, and complete epithelial healing was achieved earlier than in placebo-treated corneas. However, no significant antiviral effect of FGF-2, heparin and suramin was detectable in plaque assays from conjunctival swabs. • Conclusions: These experiments demonstrate that FGF-2 is effective in promoting herpetic epithelial ulcer healing, either due to its proliferative effects on epithelial cells or indirectly by occupying the sites on cell surface heparan sulfate necessary for the attachment of the virion. The latter mechanism of action is presumably the reason for the similar effect of heparin and suramin.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF01880673
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  • 3
    Electronic Resource
    Electronic Resource
    A new, rapid colorimetric assay for quantitative determination of cellular proliferation, growth inhibition, and viability (1993)
    Springer
    Methods in cell science 15 (1993), S. 37-41 
    Details
    ISSN: 1573-0603
    Keywords: Janus green ; spectrophotometry ; tissue culture ; cellular viability
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A sensitive spectrophotometric test has been developed to rapidly quantitate cell growth and viability on monolayer cultures. The method consists of staining fixed cells with the supravital dye Janus green, extraction of the dye from these cells with absolute alcohol, spectrophotometric measurement of the eluant, and referring optical density (OD) readings to a previously established standard curve such that measurements can be expressed directly as number of surviving cells. Staining of cells with the same colorant before fixation allows microscopical or colorimetric quantitation or both of damaged cells still adherent to the substratum. Both steps can be conveniently carried out on the same cell culture plate. More rapid and easier to perform than the usual cell counting or tritiated thymidine assays, this technique may prove to be useful in evaluation of the effect of growth promoting substances, growth inhibitors, or cytotoxic agents on monolayer cultures.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF02387288
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  • 4
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    Fully automated analysis of chemically induced γH2AX foci in human peripheral blood mononuclear cells by indirect immunofluorescence (2013)
    Wiley-Blackwell
    Details
    Publication Date: 2013-09-06
    Description: Analysis of phosphorylated histone protein H2AX (γH2AX) foci is currently the most sensitive method to detect DNA double-strand breaks (DSB). This protein modification has the potential to become an individual biomarker of cellular stress, especially in the diagnosis and monitoring of neoplastic diseases. To make γH2AX foci analysis available as a routine screening method, different software approaches for automated immunofluorescence pattern evaluation have recently been developed. In this study, we used novel pattern recognition algorithms on the AKLIDES ® platform to automatically analyze immunofluorescence images of γH2AX foci and compared the results with visual assessments. Dose- and time-dependent γH2AX foci formation was investigated in human peripheral blood mononuclear cells (PBMCs) treated with the chemotherapeutic drug etoposide (ETP). Moreover, the AKLIDES system was used to analyze the impact of different immunomodulatory reagents on γH2AX foci formation in PBMCs. Apart from γH2AX foci counting the use of novel pattern recognition algorithms allowed the measurement of their fluorescence intensity and size, as well as the analysis of overlapping γH2AX foci. The comparison of automated and manual foci quantification showed overall a good correlation. After ETP exposure, a clear dose-dependent increase of γH2AX foci formation was evident using the AKLIDES as well as Western blot analysis. Kinetic experiments on PBMCs incubated with 5 μM ETP demonstrated a peak in γH2AX foci formation after 4 to 8 h, while a removal of ETP resulted in a strong reduction of γH2AX foci after 1 to 4 h. In summary, this study demonstrated that the AKLIDES system can be used as an efficient automatic screening tool for γH2AX foci analysis by providing new evaluation features and facilitating the identification of drugs which induce or modulate DNA damage. © 2013 International Society for Advancement of Cytometry
    Electronic ISSN: 1552-4930
    Topics: Biology , Medicine
    Published by Wiley-Blackwell
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  • 5
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    Unconventional Peptide Presentation by Major Histocompatibility Complex (MHC) Class I Allele HLA-A*02:01 [Immunology] (2017)
    The American Society for Biochemistry and Molecular Biology (ASBMB)
    Details
    Publication Date: 2017-04-01
    Description: Peptide antigen presentation by major histocompatibility complex (MHC) class I proteins initiates CD8+ T cell-mediated immunity against pathogens and cancers. MHC I molecules typically bind peptides with 9 amino acids in length with both ends tucked inside the major A and F binding pockets. It has been known for a while that longer peptides can also bind by either bulging out of the groove in the middle of the peptide or by binding in a zigzag fashion inside the groove. In a recent study, we identified an alternative binding conformation of naturally occurring peptides from Toxoplasma gondii bound by HLA-A*02:01. These peptides were extended at the C terminus (PΩ) and contained charged amino acids not more than 3 residues after the anchor amino acid at PΩ, which enabled them to open the F pocket and expose their C-terminal extension into the solvent. Here, we show that the mechanism of F pocket opening is dictated by the charge of the first charged amino acid found within the extension. Although positively charged amino acids result in the Tyr-84 swing, amino acids that are negatively charged induce a not previously described Lys-146 lift. Furthermore, we demonstrate that the peptides with alternative binding modes have properties that fit very poorly to the conventional MHC class I pathway and suggest they are presented via alternative means, potentially including cross-presentation via the MHC class II pathway.
    Print ISSN: 0021-9258
    Electronic ISSN: 1083-351X
    Topics: Biology , Chemistry and Pharmacology
    Published by The American Society for Biochemistry and Molecular Biology (ASBMB)
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