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Nucleic Acids Res. 2016 May 19;44(9):4317-29. doi: 10.1093/nar/gkw262. Epub 2016 Apr 15.

Is a fully wrapped SSB-DNA complex essential for Escherichia coli survival?

Author information

1
Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis, MO, 660 S. Euclid Avenue, Box 8231, 63110-1093, USA.
2
Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis, MO, 660 S. Euclid Avenue, Box 8231, 63110-1093, USA lohman@biochem.wustl.edu.

Abstract

Escherichia coli single-stranded DNA binding protein (SSB) is an essential homotetramer that binds ssDNA and recruits multiple proteins to their sites of action during genomic maintenance. Each SSB subunit contains an N-terminal globular oligonucleotide/oligosaccharide binding fold (OB-fold) and an intrinsically disordered C-terminal domain. SSB binds ssDNA in multiple modes in vitro, including the fully wrapped (SSB)65 and (SSB)56 modes, in which ssDNA contacts all four OB-folds, and the highly cooperative (SSB)35 mode, in which ssDNA contacts an average of only two OB-folds. These modes can both be populated under physiological conditions. While these different modes might be used for different functions, this has been difficult to assess. Here we used a dimeric SSB construct with two covalently linked OB-folds to disable ssDNA binding in two of the four OB-folds thus preventing formation of fully wrapped DNA complexes in vitro, although they retain a wild-type-like, salt-dependent shift in cooperative binding to ssDNA. These variants complement wild-type SSB in vivo indicating that a fully wrapped mode is not essential for function. These results do not preclude a normal function for a fully wrapped mode, but do indicate that E. coli tolerates some flexibility with regards to its SSB binding modes.

PMID:
27084941
PMCID:
PMC4872115
DOI:
10.1093/nar/gkw262
[Indexed for MEDLINE]
Free PMC Article

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