Novel DNA glycosylases from Mycobacterium tuberculosis.
Oxidized bases are removed from DNA of Escherichia coli by enzymes formamidopyrimidine DNA glycosylase (Eco-Fpg) and endonuclease VIII (Eco-Nei) of the same structural family Fpg/Nei. New homologs of these enzymes not characterized earlier have been found in genomes of Actinobacteria. We have cloned and expressed two paralogs (Mtu-Nei2 and Mtu-Fpg2) from 36KAZ and KHA94 isolates of Mycobacterium tuberculosis and studied their ability to participate in DNA repair. Under heterologous expression in E. coli, Mtu-Nei2 decreased the rate of spontaneous mutagenesis in the rpoB gene, whereas Mtu-Fpg2 moderately increased it, possibly due to absence of residues crucially important for catalysis in this protein. Mtu-Nei2 was highly active toward double-stranded DNA substrates containing dihydrouracil residues and apurine-apyrimidine sites and was less efficient in cleavage of substrates containing 8-oxoguanine and uracil residues. These lesions, as well as 8-oxoadenine residues, were also recognized and removed by the enzyme from single-stranded DNA. Fpg and Nei homologs from M. tuberculosis can play an important role in protection of bacteria against genotoxic stress caused by oxidative burst in macrophages.
Sidorenko
V S
VS
Institute of Chemical Biology and Fundamental Medicine, Siberian Division of the Russian Academy of Sciences, Novosibirsk 630090, Russia.
Rot
M A
MA
Filipenko
M L
ML
Nevinsky
G A
GA
Zharkov
D O
DO
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D003001:Cloning, Molecular; D004260:DNA Repair; D043382:DNA-Formamidopyrimidine Glycosylase / Q000737:chemistry / Q000235:genetics*; D043224:Deoxyribonuclease (Pyrimidine Dimer) / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D005816:Genetic Complementation Test; D008969:Molecular Sequence Data; D009169:Mycobacterium tuberculosis / Q000201:enzymology* / Q000235:genetics; D011994:Recombinant Proteins / Q000302:isolation & purification; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D001426:Bacterial Proteins; D011994:Recombinant Proteins; D043224:Deoxyribonuclease (Pyrimidine Dimer); D043382:DNA-Formamidopyrimidine Glycosylase