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Escherichia coli single-stranded DNA-binding protein: nanoESI-MS studies of salt-modulated subunit exchange and DNA binding transactions

Journal Article


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Abstract


  • Single-stranded DNA-binding proteins (SSBs) are ubiquitous oligomeric proteins that bind with very high affinity to single-stranded DNA and have a variety of essential roles in DNA metabolism. Nanoelectrospray ionization mass spectrometry (nanoESI-MS) was used to monitor subunit exchange in full-length and truncated forms of the homotetrameric SSB from Escherichia coli. Subunit exchange in the native protein was found to occur slowly over a period of hours, but was significantly more rapid in a truncated variant of SSB from which the eight C-terminal residues were deleted. This effect is proposed to result from C-terminus mediated stabilization of the SSB tetramer, in which the C-termini interact with the DNA-binding cores of adjacent subunits. NanoESI-MS was also used to examine DNA binding to the SSB tetramer. Binding of single-stranded oligonucleotides [one molecule of (dT)70, one molecule of (dT)35, or two molecules of (dT)35] was found to prevent SSB subunit exchange. Transfer of SSB tetramers between discrete oligonucleotides was also observed and is consistent with predictions from solution-phase studies, suggesting that SSB-DNA complexes can be reliably analyzed by ESI mass spectrometry.

Publication Date


  • 2013

Citation


  • Mason, C. E., Jergic, S., Lo, T., Wang, Y., Dixon, N. E. & Beck, J. L. (2013). Escherichia coli single-stranded DNA-binding protein: nanoESI-MS studies of salt-modulated subunit exchange and DNA binding transactions. Journal of the American Society for Mass Spectrometry, 24 (2), 274-285.

Scopus Eid


  • 2-s2.0-84877013291

Ro Full-text Url


  • http://ro.uow.edu.au/cgi/viewcontent.cgi?article=1767&context=smhpapers

Ro Metadata Url


  • http://ro.uow.edu.au/smhpapers/754

Has Global Citation Frequency


Number Of Pages


  • 11

Start Page


  • 274

End Page


  • 285

Volume


  • 24

Issue


  • 2

Place Of Publication


  • United States

Abstract


  • Single-stranded DNA-binding proteins (SSBs) are ubiquitous oligomeric proteins that bind with very high affinity to single-stranded DNA and have a variety of essential roles in DNA metabolism. Nanoelectrospray ionization mass spectrometry (nanoESI-MS) was used to monitor subunit exchange in full-length and truncated forms of the homotetrameric SSB from Escherichia coli. Subunit exchange in the native protein was found to occur slowly over a period of hours, but was significantly more rapid in a truncated variant of SSB from which the eight C-terminal residues were deleted. This effect is proposed to result from C-terminus mediated stabilization of the SSB tetramer, in which the C-termini interact with the DNA-binding cores of adjacent subunits. NanoESI-MS was also used to examine DNA binding to the SSB tetramer. Binding of single-stranded oligonucleotides [one molecule of (dT)70, one molecule of (dT)35, or two molecules of (dT)35] was found to prevent SSB subunit exchange. Transfer of SSB tetramers between discrete oligonucleotides was also observed and is consistent with predictions from solution-phase studies, suggesting that SSB-DNA complexes can be reliably analyzed by ESI mass spectrometry.

Publication Date


  • 2013

Citation


  • Mason, C. E., Jergic, S., Lo, T., Wang, Y., Dixon, N. E. & Beck, J. L. (2013). Escherichia coli single-stranded DNA-binding protein: nanoESI-MS studies of salt-modulated subunit exchange and DNA binding transactions. Journal of the American Society for Mass Spectrometry, 24 (2), 274-285.

Scopus Eid


  • 2-s2.0-84877013291

Ro Full-text Url


  • http://ro.uow.edu.au/cgi/viewcontent.cgi?article=1767&context=smhpapers

Ro Metadata Url


  • http://ro.uow.edu.au/smhpapers/754

Has Global Citation Frequency


Number Of Pages


  • 11

Start Page


  • 274

End Page


  • 285

Volume


  • 24

Issue


  • 2

Place Of Publication


  • United States