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Probing the DNA sequence specificity of Escherichia coli RECA protein.

Publication ,  Journal Article
Rajan, R; Wisler, JW; Bell, CE
Published in: Nucleic Acids Res
2006

Escherichia coli RecA protein catalyzes the central DNA strand-exchange step of homologous recombination, which is essential for the repair of double-stranded DNA breaks. In this reaction, RecA first polymerizes on single-stranded DNA (ssDNA) to form a right-handed helical filament with one monomer per 3 nt of ssDNA. RecA generally binds to any sequence of ssDNA but has a preference for GT-rich sequences, as found in the recombination hot spot Chi (5'-GCTGGTGG-3'). When this sequence is located within an oligonucleotide, binding of RecA is phased relative to it, with a periodicity of three nucleotides. This implies that there are three separate nucleotide-binding sites within a RecA monomer that may exhibit preferences for the four different nucleotides. Here we have used a RecA coprotease assay to further probe the ssDNA sequence specificity of E.coli RecA protein. The extent of self-cleavage of a lambda repressor fragment in the presence of RecA, ADP-AlF4 and 64 different trinucleotide-repeating 15mer oligonucleotides was determined. The coprotease activity of RecA is strongly dependent on the ssDNA sequence, with TGG-repeating sequences giving by far the highest coprotease activity, and GC and AT-rich sequences the lowest. For selected trinucleotide-repeating sequences, the DNA-dependent ATPase and DNA-binding activities of RecA were also determined. The DNA-binding and coprotease activities of RecA have the same sequence dependence, which is essentially opposite to that of the ATPase activity of RecA. The implications with regard to the biological mechanism of RecA are discussed.

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Published In

Nucleic Acids Res

DOI

EISSN

1362-4962

Publication Date

2006

Volume

34

Issue

8

Start / End Page

2463 / 2471

Location

England

Related Subject Headings

  • Viral Regulatory and Accessory Proteins
  • Viral Proteins
  • Repressor Proteins
  • Repetitive Sequences, Nucleic Acid
  • Rec A Recombinases
  • Protein Binding
  • Oligonucleotides
  • Escherichia coli Proteins
  • Escherichia coli
  • Developmental Biology
 

Citation

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Rajan, R., Wisler, J. W., & Bell, C. E. (2006). Probing the DNA sequence specificity of Escherichia coli RECA protein. Nucleic Acids Res, 34(8), 2463–2471. https://doi.org/10.1093/nar/gkl302
Rajan, Rakhi, James W. Wisler, and Charles E. Bell. “Probing the DNA sequence specificity of Escherichia coli RECA protein.Nucleic Acids Res 34, no. 8 (2006): 2463–71. https://doi.org/10.1093/nar/gkl302.
Rajan R, Wisler JW, Bell CE. Probing the DNA sequence specificity of Escherichia coli RECA protein. Nucleic Acids Res. 2006;34(8):2463–71.
Rajan, Rakhi, et al. “Probing the DNA sequence specificity of Escherichia coli RECA protein.Nucleic Acids Res, vol. 34, no. 8, 2006, pp. 2463–71. Pubmed, doi:10.1093/nar/gkl302.
Rajan R, Wisler JW, Bell CE. Probing the DNA sequence specificity of Escherichia coli RECA protein. Nucleic Acids Res. 2006;34(8):2463–2471.
Journal cover image

Published In

Nucleic Acids Res

DOI

EISSN

1362-4962

Publication Date

2006

Volume

34

Issue

8

Start / End Page

2463 / 2471

Location

England

Related Subject Headings

  • Viral Regulatory and Accessory Proteins
  • Viral Proteins
  • Repressor Proteins
  • Repetitive Sequences, Nucleic Acid
  • Rec A Recombinases
  • Protein Binding
  • Oligonucleotides
  • Escherichia coli Proteins
  • Escherichia coli
  • Developmental Biology