English
 
Help Privacy Policy Disclaimer
  Advanced SearchBrowse

Item

ITEM ACTIONSEXPORT

Released

Journal Article

Multiplex flow magnetic tweezers reveal rare enzymatic events with single molecule precision

MPS-Authors
/persons/resource/persons205435

Agarwal,  Rohit
Duderstadt, Karl / Structure and Dynamics of Molecular Machines, Max Planck Institute of Biochemistry, Max Planck Society;

/persons/resource/persons192507

Duderstadt,  Karl E.
Duderstadt, Karl / Structure and Dynamics of Molecular Machines, Max Planck Institute of Biochemistry, Max Planck Society;

External Resource
Fulltext (public)

s41467-020-18456-y.pdf
(Publisher version), 5MB

Supplementary Material (public)

41467_2020_18456_MOESM1_ESM.pdf
(Supplementary material), 2MB

Citation

Agarwal, R., & Duderstadt, K. E. (2020). Multiplex flow magnetic tweezers reveal rare enzymatic events with single molecule precision. Nature Communications, 11(1): 4714. doi:10.1038/s41467-020-18456-y.


Cite as: http://hdl.handle.net/21.11116/0000-0007-4CC4-7
Abstract
The application of forces and torques on the single molecule level has transformed our understanding of the dynamic properties of biomolecules, but rare intermediates have remained difficult to characterize due to limited throughput. Here, we describe a method that provides a 100-fold improvement in the throughput of force spectroscopy measurements with topological control, which enables routine imaging of 50,000 single molecules and a 100 million reaction cycles in parallel. This improvement enables detection of rare events in the life cycle of the cell. As a demonstration, we characterize the supercoiling dynamics and drug-induced DNA break intermediates of topoisomerases. To rapidly quantify distinct classes of dynamic behaviors and rare events, we developed a software platform with an automated feature classification pipeline. The method and software can be readily adapted for studies of a broad range of complex, multistep enzymatic pathways in which rare intermediates have escaped classification due to limited throughput. Single molecule force measurements have shed light on dynamic biological events, but rare events escape notice owing to low throughput of the methods. Here, the authors combine an array of magnetic tweezers with lateral flow to increase throughput 100-fold, and detect rare DNA breaks induced by gyrase.