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Subunit exchange and DNA‐binding dynamics of Escherichia coli single‐stranded DNA binding protein (SSB)
Author(s) -
Mason Claire Elise,
Jergic Slobodan,
Dixon Nicholas,
Beck Jennifer
Publication year - 2010
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.24.1_supplement.492.11
Subject(s) - dna , chemistry , protein subunit , dna replication , escherichia coli , biophysics , replication protein a , oligonucleotide , dna binding protein , microbiology and biotechnology , biochemistry , biology , gene , transcription factor
Single‐stranded DNA binding proteins (SSBs) are ubiquitous in all organisms, and are responsible for binding to and maintaining the structural integrity of single‐stranded DNA (ssDNA) produced during DNA replication, recombination and repair processes. Escherichia coli SSB is a homotetrameric protein, and its interaction with ssDNA has been well‐characterised in vitro , but the mechanism by which it is able to move rapidly upon DNA during dynamic processes such as replication is largely unclear. In this work, nanoelectrospray mass spectrometry was used to investigate aspects of the E. coli SSB‐ssDNA interaction. By examining subunit exchange between unlabeled SSB and uniformly labeled 15 N‐SSB, it was found that ssDNA binding has a strongly inhibitory effect on subunit exchange, which occurs slowly but extensively in the absence of DNA. Furthermore, using mass spectrometry and surface plasmon resonance, intact SSB tetramers, under certain conditions, were found to be able to transfer directly between discrete ssDNA strands. Whether or not transfer occurs is dependent on both oligonucleotide length and salt concentration. This work shows that direct transfer is a possible mechanism of SSB mobility on DNA. This work was funded by an Australian Research Council Discovery Grant