Crystal structure of a covalent intermediate in DNA cleavage and rejoining by Escherichia coli DNA topoisomerase

Zhongtao Zhang, Bokun Cheng, Yuk Ching Tse-Dinh

Research output: Contribution to journalArticle

38 Scopus citations

Abstract

DNA topoisomerases control DNA topology by breaking and rejoining DNA strands via covalent complexes with cleaved DNA substrate as catalytic intermediates. Here we report the structure of Escherichia coli topoisomerase I catalytic domain (residues 2-695) in covalent complex with a cleaved single-stranded oligonucleotide substrate, refined to 2.3-Å resolution. The enzyme-substrate intermediate formed after strand cleavage was captured due to the presence of the D111N mutation. This structure of the covalent topoisomerase-DNA intermediate, previously elusive for type IA topoisomerases, shows distinct conformational changes from the structure of the enzyme without bound DNA and provides detailed understanding of the covalent catalysis required for strand cleavage to take place. The portion of cleaved DNA 5′ to the site of cleavage is anchored tightly with extensive noncovalent protein-DNA interactions as predicted by the "enzyme-bridged" model. Distortion of the scissile strand at the -4 position 5′ to the cleavage site allows specific selectivity of a cytosine base in the binding pocket. Many antibacterial and anticancer drugs initiate cell killing by trapping the covalent complexes formed by topoisomerases. We have demonstrated in previous mutagenesis studies that accumulation of the covalent complex of bacterial topoisomerase I is bactericidal. This structure of the covalent intermediate provides the basis for the design of novel antibiotics that can trap the enzyme after formation of the covalent complex.

Original languageEnglish (US)
Pages (from-to)6939-6944
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume108
Issue number17
DOIs
StatePublished - Apr 26 2011
Externally publishedYes

    Fingerprint

Keywords

  • Cytosine recognition
  • DNA relaxation
  • Protein-DNA complex
  • TopA

ASJC Scopus subject areas

  • General

Cite this