A central role for SSB in Escherichia coli RecQ DNA helicase function.
RecQ DNA helicases are critical components of DNA replication, recombination, and repair machinery in all eukaryotes and bacteria. Eukaryotic RecQ helicases are known to associate with numerous genome maintenance proteins that modulate their cellular functions, but there is little information regarding protein complexes involving the prototypical bacterial RecQ proteins. Here we use an affinity purification scheme to identify three heterologous proteins that associate with Escherichia coli RecQ: SSB (single-stranded DNA-binding protein), exonuclease I, and RecJ exonuclease. The RecQ-SSB interaction is direct and is mediated by the RecQ winged helix subdomain and the C terminus of SSB. Interaction with SSB has important functional consequences for RecQ. SSB stimulates RecQ-mediated DNA unwinding, whereas deletion of the C-terminal RecQ-binding site from SSB produces a variant that blocks RecQ DNA binding and unwinding activities, suggesting that RecQ recognizes both the SSB C terminus and DNA in SSB.DNA nucleoprotein complexes. These findings, together with the noted interactions between human RecQ proteins and Replication Protein A, identify SSB as a broadly conserved RecQ-binding protein. These results also provide a simple model that explains RecQ integration into genome maintenance processes in E. coli through its association with SSB.
Shereda
Robert D
RD
Department of Biomolecular Chemistry, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin 53706-1532, USA.
Bernstein
Douglas A
DA
Keck
James L
JL
D001665:Binding Sites; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005090:Exodeoxyribonucleases / Q000378:metabolism; D017434:Protein Structure, Tertiary; D053484:RecQ Helicases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity
D004269:DNA, Bacterial; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C476430:SSB protein, E coli; D005090:Exodeoxyribonucleases; C489467:RecJ protein, E coli; C506724:RecQ protein, E coli; D053484:RecQ Helicases