TopR2, the second reverse gyrase of Sulfolobus solfataricus, exhibits unusual properties.

Whereas reverse gyrase is considered as a strong marker of thermophily, the function of this peculiar type IA topoisomerase still remains to be elucidated. The archaeon Sulfolobus solfataricus encodes two reverse gyrases, TopR1 and TopR2. This duplication seems to be important because most of Crenarcheota exhibit two copies of reverse gyrase. However, to date, while TopR1 has been well characterized, no characterization of TopR2 has been reported. In this study, we describe for the first time the activity of S. solfataricus TopR2 that appears as a new reverse gyrase. Indeed, in spite of the sequence similarities between TopR1 and TopR2, we evidence unexpected great differences between the two enzymes. While TopR1 exhibits ATP-independent relaxation activity, TopR2 does not, and its activity is strictly dependent on the presence of ATP. Whereas TopR1 is a distributive topoisomerase, TopR2 exhibits an amazing high intrinsic processivity compared to all the topoisomerases studied so far. TopR2 is able to introduce a very high number of positive superturns in DNA, while TopR1 generates weakly positively supercoiled DNA. Finally, TopR2 behaves differently from TopR1 when incubated at different assay temperatures. All the results presented in this study indicate that TopR1 and TopR2 have, in vitro, different activities suggesting different functions in vivo.