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Characterization of the glmS Ribozymes from Listeria Monocytogenes and Clostridium Difficile.

ABSTRACT: The glmS ribozyme regulates the expression of the essential GlmS enzyme being involved in cell wall biosynthesis. While >450 variants of the glmS ribozyme were identified by in silico approaches and homology searches, only a few have yet been experimentally investigated. Herein, we validate and characterize the glmS ribozymes of the human pathogens Clostridium difficile and Listeria monocytogenes. Both ribozymes, as their previous characterized homologs rely on glucosamine-6-phosphate as co-factor and the presence of divalent cations for exerting the cleavage reaction. The observed EC₅₀ values in turn were found to be in the submicromolar range, at least an order of magnitude lower than observed for glmS ribozymes from other bacteria. The glmS ribozyme of L. monocytogenes was further shown to bear unique properties. It discriminates between co-factors very stringently and other than the glmS ribozyme of C. difficile retains activity at low temperatures. This finding illustrates that albeit being highly conserved, glmS ribozymes have unique characteristics.

SUBMITTER: Esser A

PROVIDER: S-EPMC10099748 | biostudies-literature | 2023 Jan

REPOSITORIES: biostudies-literature

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Characterization of the glmS Ribozymes from Listeria Monocytogenes and Clostridium Difficile.

Esser Anna A Mayer Günter G

Chemistry (Weinheim an der Bergstrasse, Germany) 20221122 3

The glmS ribozyme regulates the expression of the essential GlmS enzyme being involved in cell wall biosynthesis. While >450 variants of the glmS ribozyme were identified by in silico approaches and homology searches, only a few have yet been experimentally investigated. Herein, we validate and characterize the glmS ribozymes of the human pathogens Clostridium difficile and Listeria monocytogenes. Both ribozymes, as their previous characterized homologs rely on glucosamine-6-phosphate as co-fact ...[more]

PMID: 36194523

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Project description:BackgroundListeria monocytogenes is a common cause of bacterial meningitis. We developed an animal model of listerial meningitis.MethodsIn survival studies, C57BL/6 mice received intracisternal injections with different L. monocytogenes sequence type 1 (ST1) colony forming units per milliliter (CFU; n = 48, 105, 106, 107, 108, and 109 CFU/ml). Second, mice were inoculated with 108 CFU/ml ST1 and sacrificed at 6 h and 24 h (n = 12/group). Outcome parameters were clinical score, CFUs, cyto- and chemokine levels, and brain histopathology. Third, 84 mice were inoculated (109 CFU/ml ST1) to determine optimal antibiotic treatment with different doses of amoxicillin and gentamicin. Fourth, mice were inoculated with 109 CFU/ml ST1, treated with amoxicillin, and sacrificed at 16 h and 24 h (n = 12/group) for outcome assessment. Finally, time point experiments were repeated with ST6 (n = 24/group).ResultsMedian survival time for inoculation with 108 and 109 CFU/ml ST1 was 46 h and 40 h; lower doses of bacteria led to minimal clinical signs of disease. Brain levels of IL-6, IL-17A, and IFN-γ were elevated at 24 h, and IL-1β, IL-6, IL-10, IFN-γ, and TNF-α were elevated in blood at 6 h and 24 h. Histopathology showed increased meningeal infiltration, vascular inflammation of meningeal vessels, hemorrhages, and ventriculitis. In the treatment model, brain levels of IL-6 and IL-17A and blood levels of IL-6 and IFN-γ were elevated. Compared to ST6, infection with ST1 led initially to higher levels of IL-1β and TNF-α in blood and more profound neuropathological damage. At 16 h post inoculation, IL-1β, IL-10, and TNF-α in blood and IL-6, IL17A, TNF-α, and IFN-γ levels in brain were higher in ST1 compared to ST6 without differences in CFUs between STs. At 24 h, neuropathology score was higher in ST1 compared to ST6 (p = 0.002) infected mice.ConclusionsWe developed and validated a murine model of listerial meningitis. ST1-infected mice had a more severe inflammatory response and brain damage as compared to ST6-infected mice.

Dataset Information

Characterization of the glmS Ribozymes from Listeria Monocytogenes and Clostridium Difficile.

Publications

Characterization of the glmS Ribozymes from Listeria Monocytogenes and Clostridium Difficile.

Similar Datasets

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