Mutations in the RecQ DNA helicase gene BLM give rise to Bloom's syndrome, which is a rare autosomal recessive disorder characterized by genetic instability and cancer predisposition. BLM helicase is highly active in binding and unwinding G-quadruplexes (G4s), which are physiological targets for BLM, as revealed by genome-wide characterizations of gene expression of cells from BS patients. With smFRET assays, we studied the molecular mechanism of BLM-catalyzed G4 unfolding and showed that ATP is required for G4 unfolding. Surprisingly, depending on the molecular environments of G4, BLM unfolds G4 through different mechanisms: unfolding G4 harboring a 3′-ssDNA tail in three discrete steps with unidirectional translocation, and unfolding G4 connected to dsDNA by ssDNA in a repetitive manner in which BLM remains anchored at the ss/dsDNA junction, and G4 was unfolded by reeling in ssDNA. This indicates that one BLM molecule may unfold G4s in different molecular environments through different mechanisms.

Escherichia coli UvrD is a non-ring-shaped model helicase, displaying a 3 0 -5 0 polarity in DNA unwinding. Using a transverse magnetic tweezer and DNA hairpins, we measured the unwinding kinetics of UvrD at various DNAdestabilizing forces. The multiform patterns of unwinding bursts and the distributions of the off-times favour the mechanism that UvrD unwinds DNA as a dimer. The two subunits of the dimer coordinate to unwind DNA processively. They can jointly switch strands and translocate backwards on the other strand to allow slow (B40 bp/s) rewinding, or unbind simultaneously to allow quick rehybridization. Partial dissociation of the dimer results in pauses in the middle of the unwinding or increases the translocation rate from B40 to B150 nt/s in the middle of the rewinding. Moreover, the unwinding rate was surprisingly found to decrease from B45 to B10 bp/s when the force is increased from 2 to 12 pN. The results lead to a strained-inchworm mechanism in which a conformational change that bends and tenses the ssDNA is required to activate the dimer.