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Structure of the SSB–DNA polymerase III interface and its role in DNA replication
Author(s) -
Marceau Aimee H,
Bahng Soon,
Massoni Shawn C,
George Nicholas P,
Sandler Steven J,
Marians Kenneth J,
Keck James L
Publication year - 2011
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1038/emboj.2011.305
Subject(s) - replisome , biology , dna clamp , dna replication , dna polymerase , dna polymerase delta , dna polymerase ii , primase , polymerase , primer (cosmetics) , microbiology and biotechnology , prokaryotic dna replication , dna polymerase i , dna , dnag , genetics , eukaryotic dna replication , gene , polymerase chain reaction , reverse transcriptase , chemistry , organic chemistry
Interactions between single‐stranded DNA‐binding proteins (SSBs) and the DNA replication machinery are found in all organisms, but the roles of these contacts remain poorly defined. In Escherichia coli , SSB's association with the χ subunit of the DNA polymerase III holoenzyme has been proposed to confer stability to the replisome and to aid delivery of primers to the lagging‐strand DNA polymerase. Here, the SSB‐binding site on χ is identified crystallographically and biochemical and cellular studies delineate the consequences of destabilizing the χ/SSB interface. An essential role for the χ/SSB interaction in lagging‐strand primer utilization is not supported. However, sequence changes in χ that block complex formation with SSB lead to salt‐dependent uncoupling of leading‐ and lagging‐strand DNA synthesis and to a surprising obstruction of the leading‐strand DNA polymerase in vitro , pointing to roles for the χ/SSB complex in replisome establishment and maintenance. Destabilization of the χ/SSB complex in vivo produces cells with temperature‐dependent cell cycle defects that appear to arise from replisome instability.