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Master Thesis

Étude et comparaison de l'utilisation des sucres aminés chez E.coli et chez B.subtilis

Abstract : Amino sugars are essential in all organisms and they are part of many subcellular structures in eukaryotes and bacteria. Metabolic pathways involving amino sugars have not been studied in detail in other organisms besides E.coli. Analysis of the genomic sequence of B.subtilis has identified genes presumed to be involved in the use of N-acetylglucosamine (GlcNAc) and glucosamine (GlcN), namely two PTS (phosphotransferase system) transporters homologous to NagE of E.coli (GamP and NagP), two copies of NagB encoding the enzyme GlcN-6-phosphate (GlcN6P) deaminase (NagB and GamA) and a gene for GlcNAc6P deacetylase (NagA). These genes are located in two operons (hprK nagAB yvoA and ybgA-gamAP) and a isolated gene for nagP. Our ANR funded project "Study and comparison of the use of amino sugars in E coli and in B.subtilis" aims to produce a detailed description of the use of amino sugars in the two diverged bacterial species. In B.subtilis, unlike E. coli, growth on GlcN is faster than on GlcNAc. To study the functions of the nag and gam genes, we developed a system to make gene deletions using an exisable antibiotique resistance cassette which allows the accumulation of mutations in multiple loci. Our study showed that GamA is essential for the use of GlcN and NagA essential for the use of GlcNAc. GamP and NagP are required for the transport of GlcNAc but GamP or PtsG (PTS transporter for glucose) is sufficient for transport of GlcN. NagB seems to have little effect. These results were confirmed by the study of expression of nag and gam genes by primer extension, Northerns blots, fusions with β-galactosidase and Westerns blots. This study also showed that the transcriptional regulators of the GntR family, YbgA and YvoA, are transcriptional repressors of their respective operons and their binding sites were identified by DNaseI footprinting. The binding of YbgA seems sensitive to GlcN6P but the regulation by YvoA seems more complex. The gamAP operon, which exists only in B.subtilis, is now the best expressed of the nag related genes and GamA is responsible not only for growth on GlcN but also for most of the growth on GlcNAc
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Master Thesis
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2012.TH18200. Gaugué isabelle...
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Isabelle Gaugué. Étude et comparaison de l'utilisation des sucres aminés chez E.coli et chez B.subtilis. Alimentation et Nutrition. 2012. ⟨dumas-01076672⟩

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