Functional analysis of Pseudomonas aeruginosa MutS-betaclamp interaction

MONTI MR; MIGUEL V; BORGOÑO V; ARGARAÑA CE

Puerto Madryn

Congreso; XLVI Reunión Anual de SAIB; 2010

SAIB

We previously demonstrated that overexpression of the mismatch repair protein MutS in P. aeruginosa induces cell death and filamentation. These effects were not detected when overexpressed a C-terminal deletion MutS mutant. In addition, we found that the Cterminal domain copurified with the processivity factor beta-clamp. The aim of this study is to determine if MutS interacts with beta-clamp and analyze the functional role of this interaction. First, we mutated the putative beta-clamp binding motif QSDLF located at the C-terminal domain of MutS generating the MutS-beta mutant. At difference of MutS, this mutant was not able to bind to beta-clamp  in vitro. In concordance, overexpression of MutS-beta did not produce lethality and morphological changes. Since it has been proposed that beta-clamp could be involved in the repair of DNA replication errors, we analyzed the proficiency for mismatch repair activity in vivo of a  mutant strain harboring the chromosomal mutSBeta mutant gene. This mutant strain presented a  mutation frequency similar to the parental strain in the presence or absence of the mutagen  2-aminopurine. On the other hand, the analysis of growth kinetic indicated that the mutSbeta strain showed a more extended lag period and a lower exponential doubling time compared to the parental strain. Thus, ours findings suggest that the MutS-beta clamp interaction may be implicated in the regulation of cell cycle in P. Aeruginosa.