Dresden 2014 – wissenschaftliches Programm

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BP: Fachverband Biologische Physik

BP 10: Posters: Molecular Motors

BP 10.6: Poster

Dienstag, 1. April 2014, 09:30–12:30, P1

Measurements of single fluorescent motor proteins: The Right Way — •Felix Ruhnow, Linda Kloß, and Stefan Diez — B CUBE, Technische Universität Dresden

Cytoskeletal motor proteins are required in many cellular processes, such as intracellular transport and mitosis. Therefore, the biophysical characterization of motor protein movement along their filamentous tracks is essential. Commonly, stepping motility assays are used to determine the stepping and detachment rates of various molecular motor proteins by measuring their speed, run length and interaction time. However, comparison of these results proved to be difficult because the experimental setup (e.g. bead assay vs. single-molecule fluorescence assay), the experimental conditions (e.g. temperature, buffer or filament preparation) and data analysis (e.g. normal vs. exponential distribution) can influence the results. Here, we describe a method to evaluate traces of fluorescent motor proteins and propose an algorithm to correct the measurements for photobleaching and the limited length of the filaments. Additionally, bootstrapping is used to estimate statistical errors of the evaluation method. The method was tested with numerical simulations as well as with experimental data from kinesin-1 stepping experiments to show that the run length of kinesin-1 is independent of the microtubule length distribution. Our work will not only improve the evaluation of experimental data, but will also allow for better statistical comparison of two or more populations of motor proteins (e.g. motors with distinct mutations or motors linked to different cargos).