Khanam, Taran and Afsar, Mohammad and Shukla, Ankita and Alam, Faiyaz and Kumar, Sanjay and Soyar, Horam and Dolma, Kunzes and Ashish, . and Pasupuleti, Mukesh and Srivastava, Kishore Kumar and Ampapathi, Ravi Sankar and Ramachandran, Ravishankar (2020) M. tuberculosis class II apurinic/ apyrimidinic-endonuclease/3'-5' exonuclease (XthA) engages with NAD+-dependent DNA ligase A (LigA) to counter futile cleavage and ligation cycles in base excision repair. Nucleic acids research, 48 (8). pp. 4325-4343. ISSN 1362-4962
Full text not available from this repository. (Request a copy)Abstract
Class-II AP-endonuclease (XthA) and NAD+-dependent DNA ligase (LigA) are involved in initial and terminal stages of bacterial DNA base excision repair (BER), respectively. XthA acts on abasic sites of damaged DNA to create nicks with 3'OH and 5'-deoxyribose phosphate (5'-dRP) moieties. Co-immunoprecipitation using mycobacterial cell-lysate, identified MtbLigA-MtbXthA complex formation. Pull-down experiments using purified wild-type, and domain-deleted MtbLigA mutants show that LigA-XthA interactions are mediated by the BRCT-domain of LigA. Small-Angle-X-ray scattering, 15N/1H-HSQC chemical shift perturbation experiments and mutational analysis identified the BRCT-domain region that interacts with a novel 104DGQPSWSGKP113 motif on XthA for complex-formation. Isothermal-titration calorimetry experiments show that a synthetic peptide with this sequence interacts with MtbLigA and disrupts XthA-LigA interactions. In vitro assays involving DNA substrate and product analogs show that LigA can efficiently reseal 3'OH and 5'dRP DNA termini created by XthA at abasic sites. Assays and SAXS experiments performed in the presence and absence of DNA, show that XthA inhibits LigA by specifically engaging with the latter's BRCT-domain to prevent it from encircling substrate DNA. Overall, the study suggests a coordinating function for XthA whereby it engages initially with LigA to prevent the undesirable consequences of futile cleavage and ligation cycles that might derail bacterial BER.
| Item Type: | Article |
|---|---|
| Additional Information: | Copyright of this article belongs to Oxford University Press. |
| Uncontrolled Keywords: | M. tuberculosis;genome integrity |
| Subjects: | Q Science > QR Microbiology |
| Depositing User: | Dr. K.P.S.Sengar |
| Date Deposited: | 04 Apr 2020 17:10 |
| Last Modified: | 20 Nov 2020 11:38 |
| URI: | http://crdd.osdd.net/open/id/eprint/2549 |
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