Both DNA gyrase and reverse gyrase are present in the hyperthermophilic bacterium Thermotoga maritima

Abstract

Like all hyperthermophiles yet tested, the bacterium Thermotoga maritima contains a reverse gyrase. Here we show that it contains also a DNA gyrase. The genes top2A and top2B encoding the two subunits of a DNA gyraselike enzyme have been cloned and sequenced. The Top2A (type II DNA topoisomerase A protein) is more similar to GyrA (DNA gyrase A protein) than to ParC [topoisomerase IV (Topo IV) C protein]. The difference is especially striking at the C-terminal domain, which differentiates DNA gyrases from Topo IV. DNA gyrase activity was detected in T. maritima and purified to homogeneity using a novobiocin-Sepharose column. This hyperhermophilic DNA gyrase has an optimal activity around 82-86°C. In contrast to plasmids from hyperthermophilic archaea, which are from relaxed to positively supercoiled, we found that the plasmid pRQ7 from Thermotoga sp. RQ7 is negatively supercoiled. pRQ7 became positively supercoiled after addition of novobiocin to cell cultures, indicating that its negative supercoiling is due to the DNA gyrase of the host strain. The findings concerning DNA gyrase and negative supercoiling in Thermotogales put into question the role of reverse gyrase in hyperthermophiles.Prokaryotes possess atypical DNA topoisomerases, DNA gyrase, and reverse gyrase, which can produce negative or positive DNA supercoiling, respectively (1). Up to now, reverse gyrase has been found in all organisms with an optimal growth temperature of 80°C or above (hyperthermophiles) (2, 3), whereas DNA gyrase has been found only in mesophilic organisms. Reverse gyrase is present in both Archaea and Bacteria (the two prokaryotic domains), whereas DNA gyrase has only been isolated from bacteria, although putative DNA gyrase genes are present in halophilic archaea (4). The existence of reverse gyrase in hyperthermophilic archaea has been correlated with a unique topological state of their genetic elements (plasmid and viral DNA) which are from relaxed to positively supercoiled, whereas plasmids in mesophilic bacteria and archaea are negatively supercoiled (5-8).Reverse gyrase and DNA gyrase appear to be specialized DNA topoisomerases. Reverse gyrase is a unique combination of a classical type I DNA topoisomerase (Topo I) and a protein module with helicase signatures (9-12). The putative ATPdependent unwinding activity of this module probably induces the formation of a compensatory positive superturn, which is then stabilized by formation of a new topological link introduced by the topoisomerase moiety (9, 12). By contrast, DNA gyrase belongs to the classical family of Topo II (Topo IIA, see below), but it has the unique ability to wrap a DNA segment into a positive toroidal superturn (13). This enzyme is composed of two subunits, GyrA and GyrB (DNA gyrase A and B proteins), which are homologous to the C and E proteins of Topo IV (ParC and ParE), and to the C-and N-terminal halves of the eukaryotic Topo II, respectively (14). The wrapping of DNA around DNA gyrase is dependent on the C-terminal domain of the GyrA subunit (15...

Reverse gyrase from hyperthermophiles: probable transfer of a thermoadaptation trait from Archaea to Bacteria

“…Recently, a thermophilic type II topoisomerase has been isolated from T. maritima and determined to be a gyrase (19). These results raise the question of the function of these enzymes.…”

Plasmid pGS5 from the Hyperthermophilic Archaeon Archaeoglobus profundus Is Negatively Supercoiled

“…Accordingly, the idea that DNA was stabilized in organisms living at very high temperatures by an overall linking excess seemed to be supported (6). However, this hypothesis was called into question when plasmid pRQ7 from the hyperthermophilic bacterium Thermotoga maritima was found to be negatively supercoiled (7). In addition to reverse gyrase, T. maritima possesses DNA gyrase, a typical bacterial topoisomerase that introduces negative supercoils into DNA and is responsible for the negative supercoiling of pRQ7 (7).…”

“…However, this hypothesis was called into question when plasmid pRQ7 from the hyperthermophilic bacterium Thermotoga maritima was found to be negatively supercoiled (7). In addition to reverse gyrase, T. maritima possesses DNA gyrase, a typical bacterial topoisomerase that introduces negative supercoils into DNA and is responsible for the negative supercoiling of pRQ7 (7). Two alternative possibilities could be then considered to explain the different plasmid topologies in hyperthermophiles.…”

DNA-DNA Recognition: From Tight Contact to Fatal Attraction

“…Indeed, in contrast to life at high temperature that can tolerate various topological states of DNA -from negative supercoiled DNA to slightly positively supercoiled DNA-(BrochierArmanet and Forterre, 2006;Charbonnier and Forterre, 1994;Guipaud et al, 1997;LopezGarcia et al, 2000;Marguet and Forterre, 1994), adaptation to mesophilic life is much more contraining on the topology of DNA: the genome of mesophilic organisms, including bacteria, archaea and eukarya, is systematically (-) supercoiled. All mesophilic bacteria have a DNA gyrase that introduce (-) supercoiling in a plectonemic form (Forterre and Gadelle, 2009).…”