Catalytic DNA Polymerization Can Be Expedited by Active Product Release

The sequence-specific hybridization of DNA facilitates its use as a building block for designer nanoscale structures and reaction networks that perform computations. However, the strong binding energy of Watson-Crick base pairing that underlies this specificity also causes the DNA dehybridization rate to depend sensitively on sequence length and temperature. This strong dependency imposes stringent constraints on the design of multi-step DNA reactions, because a small deviation from the optimal conditions slows down the process dramatically. Here we show how an ATP-dependent helicase, Rep-X, can drive certain dehybridization reactions in designed DNA reaction networks at rates independent of sequence length, thereby decoupling the rates of hybridization and dehybridization. To illustrate this principle, we show that Rep-X extends the range of conditions where the primer exchange reaction, which catalytically adds a domain provided by a hairpin template to a DNA substrate, proceeds rapidly: in the strong substrate-hairpin binding regime, Rep-X expedites the reaction almost one hundred-fold. Our results provide an example of how ATP consumption can drive specific dehybridization reactions in designed DNA reaction networks and how this consumption can be harnessed to expedite reactions beyond their equilibrium rates.