Structural Dynamics and Mechanochemical Coupling in DNA Gyrase

Basu, Aakash; Parente, Angelica C.; Bryant, Zev

doi:10.1016/j.jmb.2016.03.016

Cited by 23 publications

(21 citation statements)

References 62 publications

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“…In contrast to the canonical mechanism used by all other type II topoisomerases, gyrase uses a mechanism in which the C-terminal domain of the GyrA subunit wraps DNA, inducing a positive crossover between the G- and T-segments that mimics a positive supercoil (Figure 1 ) ( 6 , 13 – 18 ). This ‘wrapping’ mechanism has three important implications for gyrase activity.…”

Section: Introductionmentioning

confidence: 99%

Activities of gyrase and topoisomerase IV on positively supercoiled DNA

Ashley

Dittmore

McPherson

et al. 2017

Nucleic Acids Research

113

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Although bacterial gyrase and topoisomerase IV have critical interactions with positively supercoiled DNA, little is known about the actions of these enzymes on overwound substrates. Therefore, the abilities of Bacillus anthracis and Escherichia coli gyrase and topoisomerase IV to relax and cleave positively supercoiled DNA were analyzed. Gyrase removed positive supercoils ∼10-fold more rapidly and more processively than it introduced negative supercoils into relaxed DNA. In time-resolved single-molecule measurements, gyrase relaxed overwound DNA with burst rates of ∼100 supercoils per second (average burst size was 6.2 supercoils). Efficient positive supercoil removal required the GyrA-box, which is necessary for DNA wrapping. Topoisomerase IV also was able to distinguish DNA geometry during strand passage and relaxed positively supercoiled substrates ∼3-fold faster than negatively supercoiled molecules. Gyrase maintained lower levels of cleavage complexes with positively supercoiled (compared with negatively supercoiled) DNA, whereas topoisomerase IV generated similar levels with both substrates. Results indicate that gyrase is better suited than topoisomerase IV to safely remove positive supercoils that accumulate ahead of replication forks. They also suggest that the wrapping mechanism of gyrase may have evolved to promote rapid removal of positive supercoils, rather than induction of negative supercoils.

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Section: Introductionmentioning

confidence: 99%

Activities of gyrase and topoisomerase IV on positively supercoiled DNA

Ashley

Dittmore

McPherson

et al. 2017

Nucleic Acids Research

113

View full text Add to dashboard Cite

show abstract

“…In contrast, the C-terminal domain of GyrA wraps DNA, inducing a positive supercoil between the G- and T-segments. 1 , 4 , 9 − 14 Because these captured DNA segments are proximal to one another, 1 , 4 , 15 gyrase greatly favors the catalysis of intramolecular strand passage reactions. As a result, the enzyme can efficiently alter superhelical density but is very poor at removing knots and tangles.…”

mentioning

confidence: 99%

Recognition of DNA Supercoil Geometry byMycobacterium tuberculosisGyrase

Ashley¹,

Blower

Osheroff

2017

Biochemistry

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Mycobacterium tuberculosis encodes only a single type II topoisomerase, gyrase. As a result, this enzyme likely carries out the cellular functions normally performed by canonical gyrase and topoisomerase IV, both in front of and behind the replication fork. In addition, it is the sole target for quinolone antibacterials in this species. Because quinolone-induced DNA strand breaks generated on positively supercoiled DNA ahead of replication forks and transcription complexes are most likely to result in permanent genomic damage, the actions of M. tuberculosis gyrase on positively supercoiled DNA were investigated. Results indicate that the enzyme acts rapidly on overwound DNA and removes positive supercoils much faster than it introduces negative supercoils into relaxed DNA. Canonical gyrase and topoisomerase IV distinguish supercoil handedness differently during the DNA cleavage reaction: while gyrase maintains lower levels of cleavage complexes on overwound DNA, topoisomerase IV maintains similar levels of cleavage complexes on both over- and underwound substrates. M. tuberculosis gyrase maintained lower levels of cleavage complexes on positively supercoiled DNA in the absence and presence of quinolone-based drugs. By retaining this important feature of canonical gyrase, the dual function M. tuberculosis type II enzyme remains a safe enzyme to act in front of replication forks and transcription complexes. Finally, the N-terminal gate region of the enzyme appears to be necessary to distinguish supercoil handedness during DNA cleavage, suggesting that the capture of the transport segment may influence how gyrase maintains cleavage complexes on substrates with different topological states.

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“…The architecture of a full-length DNA Gyrase from T. thermophilus in complex with a 155 bp dsDNA and ciprofloxacin was first solved using cryoelectron microscopy (cryo-EM) at a resolution of 18 Å 12 . This first model provided a rationale for probing mechanistic questions based on biochemical, single molecule and FRET techniques [13][14][15][16] , but the low resolution model did not allow deconvolution of discreet conformations of the full-length enzyme, leaving many mechanistic questions unresolved. Furthermore, the atomic details of the structure and of the drug binding site in the context of the overall conformation of DNA gyrase were not available at this resolution.…”

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confidence: 99%

Cryo-EM structure of the completeE. coliDNA Gyrase nucleoprotein complex

Broeck

Ortiz

Lamour

2019

Preprint

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DNA Gyrase is an essential enzyme involved in the homeostatic control of DNA supercoiling and the target of successful antibacterial compounds. Despite extensive studies, the detailed architecture of DNA Gyrase from the model genetic organism E. coli, is still missing, impeding structure-function analysis of E. coli-specific catalytic regulation and limiting the study of conformational intermediates of this highly flexible macromolecule. Herein, we determined the complete molecular structure of the E. coli DNA Gyrase bound to a 180 bp DNA and the antibiotic Gepotidacin, using phase-plate single-particle cryo-electron microscopy. Our data unveil with unprecedented details the structural and spatial organization of the functional domains, their connections and the position of the conserved GyrA-box motif. The deconvolution of closed and pre-opening states of the DNA-binding domain provides a better understanding of the allosteric movements of the enzyme complex. In this region, the local atomic resolution reaching up to 3.0 Å enables the identification of the antibiotic density in the DNA complex. Altogether, this study paves the way for the cryo-EM determination of gyrase complexes with antibiotics and opens perspectives for targeting conformational intermediates.

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Structural Dynamics and Mechanochemical Coupling in DNA Gyrase

Cited by 23 publications

References 62 publications

Activities of gyrase and topoisomerase IV on positively supercoiled DNA

Activities of gyrase and topoisomerase IV on positively supercoiled DNA

Recognition of DNA Supercoil Geometry byMycobacterium tuberculosisGyrase

Cryo-EM structure of the completeE. coliDNA Gyrase nucleoprotein complex

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