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BP: Fachverband Biologische Physik

BP 49: Molecular motors

BP 49.2: Vortrag

Donnerstag, 19. März 2015, 17:15–17:30, H 1028

Microfluidic setup for highly-parallel force-velocity measurements on single motor proteins — •Marta Urbanska¹, Karl Duderstadt², Annemarie Lüdecke¹, Wim Walter³, Antoine van Oijen², and Stefan Diez¹ — ¹B CUBE, TU Dresden, Germany — ²Zernike Institute for Advanced Materials, University of Groningen, Netherlands — ³Biozentrum Klein Flottbek, Uni Hamburg, Germany

Cytoskeletal motor proteins are essential for long-range, directed transport within cells. In vitro, it has been shown that external loads alter the velocity with which these ATP-driven molecules step along their filamentous tracks. So far, such force-velocity studies have been performed almost exclusively using optical traps. While optical trapping provides the highest force and spatio-temporal accuracy, it allows for one measurement at a time only. Here, we describe an alternative method to simultaneously measure multiple force-velocity relations based on the application of calibrated hydrodynamic forces to stepping motors via DNA-tethered beads. In particular, 1-um beads were attached to the SNAP-tag-labelled tails of kinesin-1 motors via λ-DNA linker. Such motor-DNA-bead complexes were applied to surface-immobilized microtubules and controlled flow was used to exert forces onto the motors. By observing the bead positions over time we were able to simultaneously track stepping of hundreds of individual motors with nanometer precision. The highly parallel nature of the measurements enables efficient collection of statistically significant quantities of data. Moreover, our approach is readily applicable to other motors and constitutes a new methodology for single-molecule force studies.