Protofilament Bundles Created by Mechanical Splitting of Microtubules by Surface-Tethered Kinesin-1

Abstract

Microtubules are hollow cytoskeletal filaments composed of about 13 protofilaments, which are made of α,β-tubulin dimers connected end to end. Motor proteins, such as kinesin-1, walk along microtubules in accordance with the polarity prescribed by the anisotropy of the microtubule's tubulin dimer building blocks. Surface-tethered kinesin can transport microtubules across surfaces, and have been proposed as molecular shuttles for capture and transport of analytes in microdevices. In device applications, microtubules, which are highly dynamic in vivo, are stabilized with the anti-cancer drug paclitaxel. Even so, molecular wear and breakage into shorter microtubules result from the mechanical forces exerted on the microtubules during transport. We observe that microtubule can also be split laterally into two protofilament bundles. These protofilament bundles continue to be transported across the surface by the tethered kinesin, although they follow trajectories with a much high curvature than MTs and can curl up into submicron diameter rings. We characterized these bundles using fluorescence microscopy, AFM, and SEM. We also investigated the effect of the kinesin surface density and the kinesin-surface linker on the rate of protofilament bundle formation.Sandia National Laboratories is a multi-mission laboratory managed and operated by Sandia Corporation, a wholly owned subsidiary of Lockheed Martin Corporation, for the U.S. Department of Energy's National Nuclear Security Administration under contract DE-AC04-94AL85000.