SOS INDUCTION IN MYCOBACTERIA - ANALYSIS OF THE DNA-BINDING ACTIVITY OF A LEXA-LIKE REPRESSOR AND ITS ROLE IN DNA-DAMAGE INDUCTION OF THE RECA GENE FROM MYCOBACTERIUM-SMEGMATIS

The protein encoded by the lexA gene from Mycobacterium leprae was ove rproduced in Escherichia coli. The recombinant protein bound to the pr omoter regions of the M. leprae lexA, M. leprae recA and M. smegmatis recA genes at sites with the sequences 5'-GAACACATGTTT and 5'-GAACAGGT GTTC, which belong to the 'Cheo box' family of binding sites recognize d by the SOS repressor from Bacillus subtilis. Gel mobility shift assa ys were used to confirm that proteins with the same site specificity o f DNA binding are also present in Mycobacterium tuberculosis and M. sm egmatis. Complex formation was impaired by mutagenic disruption of the dyad symmetry of the M. smegmatis recA Cheo box. LexA binding was als o inhibited by preincubation of the M. smegmatis and M. tuberculosis e xtracts with anti-M. leprae LexA antibodies, suggesting that the mycob acterial LexA proteins are functionally conserved at the level of DNA binding. Finally, exposure of M. smegmatis to DNA-damaging agents resu lted in induction of the M. smegmatis recA promoter with concomitant l oss of DNA binding of LexA to its Cheo box, confirming that this organ ism possesses the key regulatory elements of a functional SOS inductio n system.