Bacterial cell killing mediated by topoisomerase I DNA cleavage activity

Bokun Cheng, Shikha Shukla, Sarinnapha Vasunilashorn, Somshuvra Mukhopadhyay, Yuk Ching Tse-Dinh

Research output: Contribution to journalArticle

38 Scopus citations

Abstract

DNA topoisomerases are important clinical targets for antibacterial and anticancer therapy. At least one type IA DNA topoisomerase can be found in every bacterium, making it a logical target for antibacterial agents that can convert the enzyme into poison by trapping its covalent complex with DNA. However, it has not been possible previously to observe the consequence of having such a stabilized covalent complex of bacterial topoisomerase I in vivo. We isolated a mutant of recombinant Yersinia pestis topoisomerase I that forms a stabilized covalent complex with DNA by screening for the ability to induce the SOS response in Escherichia coli. Overexpression of this mutant topoisomerase I resulted in bacterial cell death. From sequence analysis and site-directed mutagenesis, it was determined that a single amino acid substitution in the TOPRIM domain changing a strictly conserved glycine residue to serine in either the Y. pestis or E. coli topoisomerase I can result in a mutant enzyme that has the SOS-inducing and cell-killing properties. Analysis of the purified mutant enzymes showed that they have no relaxation activity but retain the ability to cleave DNA and form a covalent complex. These results demonstrate that perturbation of the active site region of bacterial topoisomerase I can result in stabilization of the covalent intermediate, with the in vivo consequence of bacterial cell death. Small molecules that induce similar perturbation in the enzyme-DNA complex should be candidates as leads for novel antibacterial agents.

Original languageEnglish (US)
Pages (from-to)38489-38495
Number of pages7
JournalJournal of Biological Chemistry
Volume280
Issue number46
DOIs
StatePublished - Dec 1 2005
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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  • Cite this

    Cheng, B., Shukla, S., Vasunilashorn, S., Mukhopadhyay, S., & Tse-Dinh, Y. C. (2005). Bacterial cell killing mediated by topoisomerase I DNA cleavage activity. Journal of Biological Chemistry, 280(46), 38489-38495. https://doi.org/10.1074/jbc.M509722200