Single-molecule dynamics of DNA gyrase in evolutionarily distant bacteria Mycobacterium tuberculosis and E. coli.

Feb 13, 2023

Experts: Cooper J Galvin,Matthew Hobson,Jonathan Xianglong Meng,Athena Ierokomos,Ivan E Ivanov,James M Berger,Zev Bryant

DNA gyrase is an essential nucleoprotein motor present in all bacteria, and is a major target for antibiotic treatment of Mycobacterium tuberculosis (MTB) infection. Gyrase hydrolyzes ATP to add negative supercoils to DNA using a strand passage mechanism that has been investigated using biophysical and biochemical approaches. To analyze the dynamics of substeps leading to strand passage, single-molecule rotor bead tracking (RBT) has been used previously to follow real-time supercoiling and conformational transitions in Escherichia coli (EC) gyrase. However, RBT has not yet been applied to gyrase from other pathogenically relevant bacteria, and it is not known whether substeps are conserved across evolutionarily distant species. Here, we compare gyrase supercoiling dynamics between two evolutionarily distant bacterial species, MTB and E. coli (EC). We used RBT to measure supercoiling rates, processivities, and the geometries and transition kinetics of conformational states of purified gyrase proteins in complex with DNA. Our results show that E. coli and MTB gyrases are both processive, with the MTB enzyme displaying velocities ∼5.5X slower than the EC enzyme. Compared with EC gyrase, MTB gyrase also more readily populates an intermediate state with DNA chirally wrapped around the enzyme, in both the presence and absence of ATP. Our substep measurements reveal common features in conformational states of EC and MTB gyrases interacting with DNA, but also suggest differences in populations and transition rates that may reflect distinct cellular needs between these two species.
Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.

Author

admin

View all posts

ABOUT THE EXPERTS

Cooper J Galvin,Matthew Hobson,Jonathan Xianglong Meng,Athena Ierokomos,Ivan E Ivanov,James M Berger,Zev Bryant

Cooper J Galvin

Program in Biophysics, Stanford University, Stanford, CA, 94305, USA; Department of Bioengineering, Stanford University, Stanford, CA, 94305, USA.

Matthew Hobson

Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.

Jonathan Xianglong Meng

Department of Bioengineering, Stanford University, Stanford, CA, 94305, USA.

Athena Ierokomos

Program in Biophysics, Stanford University, Stanford, CA, 94305, USA; Department of Bioengineering, Stanford University, Stanford, CA, 94305, USA.

Ivan E Ivanov

Department of Bioengineering, Stanford University, Stanford, CA, 94305, USA; Department of Chemical Engineering, Stanford University, Stanford, CA, 94305, USA.

James M Berger

Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.

Zev Bryant

Program in Biophysics, Stanford University, Stanford, CA, 94305, USA; Department of Bioengineering, Stanford University, Stanford, CA, 94305, USA; Department of Structural Biology, Stanford University Medical Center, Stanford, CA 94305, USA. Electronic address: zevry@stanford.edu.