Highly-Parallel Microfluidics-Based Force Spectroscopy on Single Cytoskeletal Motors.

Autor: Urbanska M; B CUBE - Center for Molecular Bioengineering, Technische Universität Dresden, 01069, Dresden, Germany., Lüdecke A; B CUBE - Center for Molecular Bioengineering, Technische Universität Dresden, 01069, Dresden, Germany., Walter WJ; B CUBE - Center for Molecular Bioengineering, Technische Universität Dresden, 01069, Dresden, Germany., van Oijen AM; Zernike Institute for Advanced Materials, University of Groningen, Groningen, AE, 9700, Netherlands.; Molecular Horizons, University of Wollongong, Illawarra Health and Medical Research Institute, Wollongong, NSW, 2522, Australia., Duderstadt KE; Zernike Institute for Advanced Materials, University of Groningen, Groningen, AE, 9700, Netherlands.; Structure and Dynamics of Molecular Machines, Max Planck Institute of Biochemistry, 82152, Martinsried, Germany.; Physics Department, Technische Universität München, 85748, Garching, Germany., Diez S; B CUBE - Center for Molecular Bioengineering, Technische Universität Dresden, 01069, Dresden, Germany.; Cluster of Excellence Physics of Life, Technische Universität Dresden, 01062, Dresden, Germany.; Max Planck Institute of Molecular Cell Biology and Genetics, 01307, Dresden, Germany.
Jazyk: angličtina
Zdroj: Small (Weinheim an der Bergstrasse, Germany) [Small] 2021 May; Vol. 17 (18), pp. e2007388. Date of Electronic Publication: 2021 Mar 23.
DOI: 10.1002/smll.202007388
Abstrakt: Cytoskeletal motors transform chemical energy into mechanical work to drive essential cellular functions. Optical trapping experiments have provided crucial insights into the operation of these molecular machines under load. However, the throughput of such force spectroscopy experiments is typically limited to one measurement at a time. Here, a highly-parallel, microfluidics-based method that allows for rapid collection of force-dependent motility parameters of cytoskeletal motors with two orders of magnitude improvement in throughput compared to currently available methods is introduced. Tunable hydrodynamic forces to stepping kinesin-1 motors via DNA-tethered beads and utilize a large field of view to simultaneously track the velocities, run lengths, and interaction times of hundreds of individual kinesin-1 molecules under varying resisting and assisting loads are applied. Importantly, the 16 µm long DNA tethers between the motors and the beads significantly reduces the vertical component of the applied force pulling the motors away from the microtubule. The approach is readily applicable to other molecular systems and constitutes a new methodology for parallelized single-molecule force studies on cytoskeletal motors.
(© 2021 The Authors. Small published by Wiley-VCH GmbH.)
Databáze: MEDLINE
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