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Nuclear localization and in situ DNA damage by Mycobacterium tuberculosis nucleoside-diphosphate kinase.

Authors
  • Saini, Adesh Kumar
  • Maithal, Kapil
  • Chand, Prem
  • Chowdhury, Shantanu
  • Vohra, Reena
  • Goyal, Anita
  • Dubey, Gyanendra P
  • Chopra, Puneet
  • Chandra, Ramesh
  • Tyagi, Anil K
  • Singh, Yogendra
  • Tandon, Vibha
Type
Published Article
Journal
Journal of Biological Chemistry
Publisher
American Society for Biochemistry and Molecular Biology
Publication Date
Nov 26, 2004
Volume
279
Issue
48
Pages
50142–50149
Identifiers
PMID: 15377659
Source
Medline
License
Unknown

Abstract

Nucleoside-diphosphate kinase of Mycobacterium tuberculosis (mNdK) is a secretory protein, but the rationale behind secreting an enzyme involved in the maintenance of cellular pool of nucleoside triphosphates is not clearly understood. To elucidate the biological significance of mNdK secretion, we expressed mNdK fused to green fluorescent protein in HeLa and COS-1 cells. Interestingly, mNdK was detected in the nuclei of HeLa and COS-1 cells. Incubation of mNdK with nuclei isolated from HeLa and COS-1 cells led to in situ damage of chromosomal DNA. Surface plasmon resonance studies demonstrated that mNdK binds supercoiled plasmid DNA lacking apurinic/apyrimidinic sites with a dissociation constant of 30 +/- 3.2 mum. Plasmid cleavage by mNdK was found to be dependent on the specific divalent metal ion and inhibited by a metal ion chelator. Moreover, the metal ion-dependent DNA cleavage by mNdK was mediated by superoxide radicals as detected by electron paramagnetic resonance. The cleavage reaction was inhibited under nitrogen atmosphere confirming the necessity of molecular oxygen for DNA cleavage. In view of the findings that mNdK is secreted by intracellular mycobacteria and damages the nuclear DNA, it can be postulated that mNdK may cause cell death that could help in the dissemination of the pathogen.

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