Post-graduate theses
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| Identifier |
000369221 |
| Title |
Βιοχημικός χαρακτηρισμός και γενετική ανάλυση του ενζύμου ΒΑ0150 από τον παθογόνο μικροοργανισμό Bacillus anthracis |
| Alternative Title |
Biochemical characterization and genetic analysis of the enzyme BA0150 from the pathogen microorganism Bacillus anthracis |
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Author
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Γκέκας, Σωτήριος
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Thesis advisor
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Μπουριώτης, Βασίλης
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| Abstract |
Peptidoglycan is the basic polysaccharide of the bacterial cell wall and it is responsible for the stability and viability of the bacterial cell. It is a polymer of a repeating disaccharide-peptide unit, Ν-acetyl-glucosamine (GlcNAc) and N-acetyl- muramic acid (MurNAc) cross-linked with a small group of amino acids. Peptidoglycan is a dynamic molecule and plays a crucial role in many biological functions of bacterial cells (bacterial growth, division, autolysis and spore formation in some species) as well as in pathogenesis, through modifications of peptidoglycan from the procedures above. Such a modification is the deacetylation of peptidoglycan via the specialized enzymes peptidoglycan deacetylases (PDAs). Peptidoglycan deacetylases belong to carbohydrate esterase family 4 (CE4) and de-N-acetylate the N-acetyl-glucosamine and the N-acetyl-muramic acid residues of the di-sugar repeats in bacterial peptidoglycan.
The genome of Bacillus anthracis contains 11 open reading frames (ORFs) encoding for putative polyssacharide deacetylases. Six of these have been proposed to be peptidoglycan N-acetylglucosamine deacetylases and the other five polyssacharide deacetylases. The gene ba0150 from B. anthracis codes for a putative polyssacharide deacetylase and is homologous to PdaB (61% identity), a putative polyssacharide deacetylase from B. subtilis, that is involved in sporulation.
In this study, ba0150 was cloned and expressed in E. coli cells. The recombinant enzyme was purified to homogeneity employing Ni-NTA agarose and Sephacryl S200 HR chromatography and was further characterized. The protein was not active on glycol- chitin, GMDP and N-acetyl chitooligomers substrates.
In order to elucidate the biological role of BA0150, genetic analysis (knock out) and cellular localization experiments were performed. So far we have isolated B. anthracis strain with single recombination events. The mutant showed abnormally elongated-curved cells and a delay in autolysis and spore germination. Furthermore, the mutant was resistant against lysozyme, but more sensitive against mutanolysin.
Considering the interest on B. anthracis as a potential bioweapon, these studies may set a basis for potential drug design applications targeting enzymes involved in biosynthesis and modification of B. anthracis polyssacharides.
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| Language |
Greek |
| Subject |
Deacetylates |
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Peptidoglycan |
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Peptidoglycan polysacchacharide |
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Απακετυλάσες πεπτιδογλυκάνης |
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Απακετυλάσες πολυσακχαριτών |
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Πεπτιδοκλυκάνη |
| Issue date |
2011-11-18 |
| Collection
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School/Department--School of Sciences and Engineering--Department of Biology--Post-graduate theses
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Type of Work--Post-graduate theses
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| Permanent Link |
https://elocus.lib.uoc.gr//dlib/e/6/7/metadata-dlib-t59pl9s2hfmuuqisimdv36sal1_1319532845.tkl
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| Views |
273 |
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