A Molecular Throttle The Recombination Hotspot χ Controls DNA Translocation by the RecBCD Helicase

AbstractRecBCD enzyme is a heterotrimeric helicase/nuclease that initiates homologous recombination at double-stranded DNA breaks. Several of its activities are regulated by the DNA sequence χ (5′-GCTGGTGG-3′), which is recognized in cis by the translocating enzyme. When RecBCD enzyme encounters χ, the intensity and polarity of its nuclease activity are changed, and the enzyme gains the ability to load RecA protein onto the χ-containing, unwound single-stranded DNA. Here, we show that interaction with χ also affects translocation by RecBCD enzyme. By observing translocation of individual enzymes along single molecules of DNA, we could see RecBCD enzyme pause precisely at χ. Furthermore, and more unexpectedly, after pausing at χ, the enzyme continues translocating but at approximately one-half the initial rate. We propose that interaction with χ results in an enzyme in which one of the two motor subunits, likely the RecD motor, is uncoupled from the holoenzyme to produce the slower translocase