Binding specificity of escherichia coli single-stranded DNA binding protein for the χ subunit of DNA pol III holoenzyme and PriA helicase

Alexander G. Kozlov, Maria J. Jezewska, Wlodzimierz Bujalowski, Timothy M. Lohman

Research output: Contribution to journalArticle

36 Citations (Scopus)

Abstract

The Escherichia coli single-stranded DNA binding protein (SSB) plays a central role in DNA metabolism through its high affinity interactions with ssDNA, as well as its interactions with numerous other proteins via its unstructured C-termini. Although SSB interacts with at least 14 other proteins, it is not understood how SSB might recruit one protein over another for a particular metabolic role. To probe the specificity of these interactions, we have used isothermal titration calorimetry to examine the thermodynamics of binding of SSB to two E. coli proteins important for DNA replication, the χ subunit of DNA polymerase III holoenzyme and the PriA helicase. We find that an SSB tetramer can bind up to four molecules of either protein primarily via interactions with the last ∼9 amino acids in the conserved SSB C-terminal tails (SSB-Ct). We observe intrinsic specificity for the binding of an isolated SSB-Ct peptide to PriA over χ due primarily to a more favorable enthalpic component. PriA and χ also bind with weaker affinity to SSB (in the absence of ssDNA) than to isolated SSB-Ct peptides, indicating an inhibitory effect of the SSB protein core. Although the binding affinity of SSB for both χ and PriA is enhanced if SSB is prebound to ssDNA, this effect is larger with PriA indicating a further enhancement of SSB specificity for PriA. These results also suggest that DNA binding proteins such as PriA, which also interact with SSB, could use this interaction to gain access to ssDNA by first interacting with the SSB C-termini.

Original languageEnglish (US)
Pages (from-to)3555-3566
Number of pages12
JournalBiochemistry
Volume49
Issue number17
DOIs
StatePublished - May 4 2010

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DNA Polymerase III
Holoenzymes
Protein Subunits
DNA-Binding Proteins
Escherichia coli
Proteins
Peptides
Calorimetry
Escherichia coli Proteins
DNA
DNA Replication
Titration
Thermodynamics
Metabolism
Amino Acids
Molecules

ASJC Scopus subject areas

  • Biochemistry

Cite this

Binding specificity of escherichia coli single-stranded DNA binding protein for the χ subunit of DNA pol III holoenzyme and PriA helicase. / Kozlov, Alexander G.; Jezewska, Maria J.; Bujalowski, Wlodzimierz; Lohman, Timothy M.

In: Biochemistry, Vol. 49, No. 17, 04.05.2010, p. 3555-3566.

Research output: Contribution to journalArticle

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