Etude structurale et fonctionnelle de la reconnaissance et de la métabolisation de lésions puriques et pyrimidiques dans l'ADN par la Formamidopyrimidine-ADN glycosylase

Abstract : Oxidations on nucleic bases constitute one of the major sources of DNA lesions appearance, which can be mutagenic or lethal for cells in the absence of DNA repair. The prokaryotic Formamidopyrimidine-DNA glycosylase (Fpg), a base excision DNA repair (BER) enzyme, initiate the repair of a wide range of such lesions via its DNA glycosylase (excision of the oxidized base) and AP lyase (cleavage of the AP site by ß,d-elimination) activities. We carried out functional studies by biochemical techniques and structural studies by X-ray crystallography so as to state Fpg’s substrate specificity and catalytic mechanism. Thus, we have been able to underline the structural determinants enabling this enzyme to accommodate lesions of very different sizes in its active site, in this case 2,6-diamino-4-hydroxy-5-formamidopyrimidine (FapyG) residues N7-substituted or not by bulky adducts. On the other hand, we structurally and functionally characterized the recognition and excision by Fpg of a pyrimidic lesion, the 5-hydroxy-5-methyl-hydantoin (Hyd). Thus, we have shown that this lesion paired with a cytosine was a good substrate for the enzyme, and stated structurally the recognition mode of Hyd by Fpg. On the other hand, we have underlined an unexpected behaviour of the enzyme on this substrate. In this case, we have biochemically and structurally shown that a covalent link was formed in sizeable quantities between Fpg and Hyd in physiological conditions.
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Yann-Vaï Le Bihan. Etude structurale et fonctionnelle de la reconnaissance et de la métabolisation de lésions puriques et pyrimidiques dans l'ADN par la Formamidopyrimidine-ADN glycosylase. Sciences agricoles. Université d'Orléans, 2009. Français. ⟨NNT : 2009ORLE2004⟩. ⟨tel-00488816⟩

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