The Acidic C-terminal Tail of GyrA moderates the DNA Supercoiling Activity of B. subtilis Gyrase [DNA and Chromosomes]

February 20th, 2014 by Lanz, M. A., Farhat, M., Klostermeier, D.

Gyrase is a type II DNA topoisomerase that introduces negative supercoils into DNA in an ATP-dependent reaction. It consists of a topoisomerase core, formed by the N-terminal domains of the two GyrA subunits and the two GyrB subunits, that catalyzes double-strand DNA cleavage and passage of a second double-stranded DNA through the gap in the first. The C-terminal domains (CTDs) of the GyrA subunit form a β-pinwheel, and bind DNA around their positively charged perimeter. As a result, DNA is bound as a positive supercoil that is converted into a negative supercoil by strand passage. The CTDs contain a conserved seven-amino-acid motif that connects blades one and six of the β-pinwheel, and is a hallmark feature of gyrases. Deletion of the GyrA-box abrogates DNA bending by the CTDs and DNA-induced narrowing of the N-gate, affects T-segment presentation, reduces the coupling of DNA binding to ATP hydrolysis, and leads to supercoiling deficiency. Recently, a severe loss of supercoiling activity of E. coli gyrase upon deletion of the non-conserved acidic C-tail of the CTDs has been reported. We show here that, in contrast to E. coli gyrase, the C-tail is a very moderate negative regulator of B. subtilis gyrase activity. The C-tail reduces the degree of bending by the CTDs, but has no effect on DNA-induced conformational changes of gyrase that precede strand passage, and reduces DNA-stimulated ATPase and DNA supercoiling activities only two-fold. Our results are in agreement with species-specific, differential regulatory effects of the C-tail in gyrases from different organisms.