Characterization of the 7,8-dihydro-8-oxo-deoxyguanosine Repair System of Pseudomonas aeruginosa

MORERO NR, ARGARAÑA CE

Mar del Plata (Buenos Aires, Argentina)

Congreso; XLIII Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2007

The 7,8-dihydro-8-oxo-deoxyguanosine (8-oxodG) Repair System prevents or repairs the mutations produced by the oxidized base. This system has been mainly studied in E. coli where it involves the action of three proteins: the pyrophosphohydrolase MutT, and the DNA glycosylases MutM and MutY. MutT converts 8-oxodGTP to 8-oxodGMP, preventing incorporation of the oxidized base into the DNA. MutM removes 8-oxodG mispaired with A, and MutY removes A mispaired with G or 8-oxodG. We aim to describe this system in P. aeruginosa, by studying the influence of these three genes in the mutagenesis rates, and the biochemical characterization of the enzymes. In this work, we used DmutT, DmutM and DmutY P. aeruginosa strains, and the three WT genes cloned in expression vectors for complementation assays. The mutation rates for mutants and complemented strains were determined by the Rifampicin and Ciprofloxacin resistance tests. Rifampicin resistance mutation frequencies for DmutT, DmutM and DmutY strains showed, respect to the WT strain, an increase of 44-, 3.8- and 26-fold, respectively.  Interestingly, the emergence of Ciprofloxacin resistant mutants was, compared with the WT strain, 17- and 136-fold in DmutM and DmutY strains, and more than 800-times in DmutT. This strain was complemented by the mutT gene confirming that the mutation frequency was dependent of the presence of oxidized bases.