Structure of Escherichia coli DNA gyrase with chirally wrapped DNA supports ratchet-and-pawl mechanism for an ATP-powered supercoiling motor
crossref(2024)
Abstract
Gyrase is essential for replication and transcription in bacteria, and as such is an important target for antibiotics including fluoroquinolones. Gyrase is a molecular machine that channels the energy of ATP hydrolysis into negative supercoiling of DNA. The mechanism proposed more than 40 years ago involves stabilising and inverting a chiral DNA loop; however, the molecular basis for this is poorly understood, as the loop was never directly observed. We present high-resolution cryoEM structures of the Escherichia coli gyrase - 217 bp DNA holocomplex, and of the moxifloxacin-bound gyrase complex with cleaved 217 bp DNA. Each structure constrains an intact figure-of-eight positively supercoiled DNA loop, poised for strand passage. The loop is stabilised by a GyrA β-pinwheel domain which how we here show has a structure of a flat disc, and functions akin to a mini-nucleosome. Our data implies that during catalysis the ATPase domains of the enzyme undergo a large movement to push the transported DNA segment through the break in DNA. By comparing the catalytic site between native drug-free and moxifloxacin-bound gyrase structures that both contain a single metal ion we demonstrate that the enzyme is observed in a native pre-catalytic state. Finally, we propose a ‘ratchet and pawl’ mechanism for energy coupling in gyrase. These unexpected findings call for re-evaluation of existing data and offer a framework for further experiments designed to dissect the details of how gyrase molecular motor converts chemical energy into mechanical tension.
### Competing Interest Statement
The authors have declared no competing interest.
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