Site-directed mutagenesis of residues involved in G strand DNA binding by Escherichia coli DNA topoisomerase I

Bokun Cheng, Jingyang Feng, Vishwaroop Mulay, Sharvari Gadgil, Yuk Ching Tse-Dinh

Research output: Contribution to journalArticle

12 Scopus citations

Abstract

Crystal structures of complexes between type IA DNA topoisomerases and single-stranded DNA suggest that the residues Ser-192, Arg-195, and Gin-197 in a conserved region of Escherichia coli topoisomerase I may be important for direct interactions with phosphates on the G strand of DNA, which is the substrate for DNA cleavage and religation (Changela A., DiGate, R. J., and Mondragón, A. (2001) Nature 411, 1077-1081; Perry, K., and Mondragón, A. (2003) Structure 11, 1349-1358). Site-directed mutagenesis experiments altering these residues to alanines and other amino acids were carried out to probe the relevance of these interactions in the catalytic activities of the enzyme. The results show that the side chains of Arg-195 and Gin-197 are required for DNA cleavage by the enzyme and are likely to be important for positioning of the G strand of DNA at the active site prior to DNA cleavage. Mutation of Ser-192 did not affect DNA binding and cleavage but nevertheless decreased the overall rate of relaxation of supercoiled DNA probably because of its participation in a later step of the reaction pathway.

Original languageEnglish (US)
Pages (from-to)39207-39213
Number of pages7
JournalJournal of Biological Chemistry
Volume279
Issue number38
DOIs
StatePublished - Sep 17 2004
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Fingerprint Dive into the research topics of 'Site-directed mutagenesis of residues involved in G strand DNA binding by Escherichia coli DNA topoisomerase I'. Together they form a unique fingerprint.

  • Cite this