Characterization of DNA substrate specificities of apurinic/apyrimidinic endonucleases from Mycobacterium tuberculosis

Sailau Abeldenov, Ibtissam Talhaoui, Dmitry O. Zharkov, Alexander A. Ishchenko, Erlan Ramanculov, Murat Saparbaev, Bekbolat Khassenov

Research output: Contribution to journalArticlepeer-review

9 Citations (Scopus)


Apurinic/apyrimidinic (AP) endonucleases are key enzymes involved in the repair of abasic sites and DNA strand breaks. Pathogenic bacteria Mycobacterium tuberculosis contains two AP endonucleases: MtbXthA and MtbNfo members of the exonuclease III and endonuclease IV families, which are exemplified by Escherichia coli Xth and Nfo, respectively. It has been shown that both MtbXthA and MtbNfo contain AP endonuclease and 3'→5' exonuclease activities. However, it remains unclear whether these enzymes hold 3'-repair phosphodiesterase and nucleotide incision repair (NIR) activities. Here, we report that both mycobacterial enzymes have 3'-repair phosphodiesterase and 3'-phosphatase, and MtbNfo contains in addition a very weak NIR activity. Interestingly, depending on pH, both enzymes require different concentrations of divalent cations: 0.5mM MnCl2 at pH 7.6 and 10mM at pH 6.5. MtbXthA requires a low ionic strength and 37°C, while MtbNfo requires high ionic strength (200mM KCl) and has a temperature optimum at 60°C. Point mutation analysis showed that D180 and N182 in MtbXthA and H206 and E129 in MtbNfo are critical for enzymes activities. The steady-state kinetic parameters indicate that MtbXthA removes 3'-blocking sugar-phosphate and 3'-phosphate moieties at DNA strand breaks with an extremely high efficiency (kcat/KM=440 and 1280μM-1·min-1, respectively), while MtbNfo exhibits much lower 3'-repair activities (kcat/KM=0.26 and 0.65μM-1·min-1, respectively). Surprisingly, both MtbXthA and MtbNfo exhibited very weak AP site cleavage activities, with kinetic parameters 100- and 300-fold lower, respectively, as compared with the results reported previously. Expression of MtbXthA and MtbNfo reduced the sensitivity of AP endonuclease-deficient E. coli xth nfo strain to methylmethanesulfonate and H2O2 to various degrees. Taken together, these data establish the DNA substrate specificity of M. tuberculosis AP endonucleases and suggest their possible role in the repair of oxidative DNA damage generated by endogenous and host- imposed factors.

Original languageEnglish
Pages (from-to)1-16
Number of pages16
JournalDNA Repair
Publication statusPublished - Sep 1 2015
Externally publishedYes


  • 3'-repair phosphodiesterases
  • AP endonucleases
  • Abasic sites
  • DNA repair
  • Oxidative DNA damage

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology


Dive into the research topics of 'Characterization of DNA substrate specificities of apurinic/apyrimidinic endonucleases from Mycobacterium tuberculosis'. Together they form a unique fingerprint.

Cite this