Purification and characterization of the RecA protein from Neisseria gonorrhoeae

Elizabeth A. Stohl, Marielle C. Gruenig, Michael M. Cox, H. Steven Seifert

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12 Scopus citations

Abstract

The strict human pathogen Neisseria gonorrhoeae is the only causative agent of the sexually transmitted infection gonorrhea. The recA gene from N. gonorrhoeae is essential for DNA repair, natural DNA transformation, and pilin antigenic variation, all processes that are important for the pathogenesis and persistence of N. gonorrhoeae in the human population. To understand the biochemical features of N. gonorrhoeae RecA (RecANg), we overexpressed and purified the RecANg and SSBNg proteins and compared their activities to those of the well-characterized E. coli RecA and SSB proteins in vitro. We observed that RecANg promoted more strand exchange at early time points than RecAEc through DNA homologous substrates, and exhibited the highest ATPase activity of any RecA protein characterized to date. Further analysis of this robust ATPase activity revealed that RecANg is more efficient at displacing SSB from ssDNA and that RecANg shows higher ATPase activity during strand exchange than RecAEc. Using substrates created to mimic the cellular processes of DNA transformation and pilin antigenic variation we observed that RecAEc catalyzed more strand exchange through a 100 bp heterologous insert, but that RecANg catalyzed more strand exchange through regions of microheterology. Together, these data suggest that the processes of ATP hydrolysis and DNA strand exchange may be coupled differently in RecANg than in RecAEc. This difference may explain the unusually high ATPase activity observed for RecANg with the strand exchange activity between RecANg and RecAEc being more similar.

Original languageEnglish (US)
Article numbere17101
JournalPloS one
Volume6
Issue number2
DOIs
StatePublished - 2011

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)
  • General

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