The process of displacing the single-stranded DNA-binding protein from single-stranded DNA by RecO and RecR proteins.
The regions of single-stranded (ss) DNA that result from DNA damage are immediately coated by the ssDNA-binding protein (SSB). RecF pathway proteins facilitate the displacement of SSB from ssDNA, allowing the RecA protein to form protein filaments on the ssDNA region, which facilitates the process of recombinational DNA repair. In this study, we examined the mechanism of SSB displacement from ssDNA using purified Thermus thermophilus RecF pathway proteins. To date, RecO and RecR are thought to act as the RecOR complex. However, our results indicate that RecO and RecR have distinct functions. We found that RecR binds both RecF and RecO, and that RecO binds RecR, SSB and ssDNA. The electron microscopic studies indicated that SSB is displaced from ssDNA by RecO. In addition, pull-down assays indicated that the displaced SSB still remains indirectly attached to ssDNA through its interaction with RecO in the RecO-ssDNA complex. In the presence of both SSB and RecO, the ssDNA-dependent ATPase activity of RecA was inhibited, but was restored by the addition of RecR. Interestingly, the interaction of RecR with RecO affected the ssDNA-binding properties of RecO. These results suggest a model of SSB displacement from the ssDNA by RecF pathway proteins.
Inoue
Jin
J
RIKEN Discovery Research Institute, 2-1, Hirosawa, Wako, Saitama 351-0198, Japan.
Honda
Masayoshi
M
Ikawa
Shukuko
S
Shibata
Takehiko
T
Mikawa
Tsutomu
T
D000251:Adenosine Triphosphatases / Q000378:metabolism; D001426:Bacterial Proteins / Q000302:isolation & purification / Q000378:metabolism*; D001667:Binding, Competitive; D004277:DNA, Single-Stranded / Q000378:metabolism* / Q000648:ultrastructure; D004268:DNA-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism* / Q000648:ultrastructure; D011938:Rec A Recombinases / Q000378:metabolism; D016963:Thermus thermophilus / Q000378:metabolism*
D001426:Bacterial Proteins; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; C065831:RecR protein, Bacteria; C066218:recF protein, Bacteria; D011938:Rec A Recombinases; D000251:Adenosine Triphosphatases