The reverse gyrase helicase-like domain is a nucleotide-dependent switch that is attenuated by the topoisomerase domain.
Reverse gyrase is a topoisomerase that introduces positive supercoils into DNA in an ATP-dependent manner. It is unique to hyperthermophilic archaea and eubacteria, and has been proposed to protect their DNA from damage at high temperatures. Cooperation between its N-terminal helicase-like and the C-terminal topoisomerase domain is required for positive supercoiling, but the precise role of the helicase-like domain is currently unknown. Here, the characterization of the isolated helicase-like domain from Thermotoga maritima reverse gyrase is presented. We show that the helicase-like domain contains all determinants for nucleotide binding and ATP hydrolysis. Its intrinsic ATP hydrolysis is significantly stimulated by ssDNA, dsDNA and plasmid DNA. During the nucleotide cycle, the helicase-like domain switches between high- and low-affinity states for dsDNA, while its affinity for ssDNA in the ATP and ADP states is similar. In the context of reverse gyrase, the differences in DNA affinities of the nucleotide states are smaller, and the DNA-stimulated ATPase activity is strongly reduced. This inhibitory effect of the topoisomerase domain decelerates the progression of reverse gyrase through the nucleotide cycle, possibly providing optimal coordination of ATP hydrolysis with the complex reaction of DNA supercoiling.
del Toro Duany
Yoandris
Y
University of Basel, Biozentrum, Biophysical Chemistry, Klingelbergstrasse 70, 4056 Basel, Switzerland.
Jungblut
Stefan P
SP
Schmidt
Andreas S
AS
Klostermeier
Dagmar
D
D000227:Adenine Nucleotides / Q000378:metabolism*; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004264:DNA Topoisomerases, Type I / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000378:metabolism; D009154:Mutation; D011072:Poly U / Q000378:metabolism; D017434:Protein Structure, Tertiary; D020124:Thermotoga maritima / Q000201:enzymology
D000227:Adenine Nucleotides; D004277:DNA, Single-Stranded; D011072:Poly U; D000255:Adenosine Triphosphate; D004247:DNA; D000251:Adenosine Triphosphatases; C041813:DNA reverse gyrase; D004264:DNA Topoisomerases, Type I
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