Engineering Circular Gliding of Actin Filaments Along Myosin-Patterned DNA Nanotube Rings To Study Long-Term Actin–Myosin Behaviors
| Autor: | Rizal F. Hariadi, Abhinav J. Appukutty, Sivaraj Sivaramakrishnan |
|---|---|
| Rok vydání: | 2016 |
| Předmět: |
0301 basic medicine
Nanotube General Physics and Astronomy Nanotechnology macromolecular substances 02 engineering and technology Myosins Biology Article Protein filament 03 medical and health sciences Myosin DNA nanotechnology Molecular motor General Materials Science Actin Nanotubes General Engineering DNA Processivity 021001 nanoscience & nanotechnology Actins Molecular machine Actin Cytoskeleton 030104 developmental biology Biophysics Biological Assay 0210 nano-technology |
| Zdroj: | ACS Nano. 10:8281-8288 |
| ISSN: | 1936-086X 1936-0851 |
| DOI: | 10.1021/acsnano.6b01294 |
| Popis: | Nature has evolved molecular motors that are critical in cellular processes occurring over broad timescales, ranging from seconds to years. Despite the importance of the long-term behavior of molecular machines, topics such as enzymatic lifetime are underexplored due to the lack of a suitable approach for monitoring motor activity over long time periods. Here, we developed an “O”-shaped Myosin-Empowered Gliding Assay (OMEGA) that utilizes engineered micron-scale DNA nanotube rings with precise arrangements of myosin VI to trap gliding actin filaments. This circular gliding assay platform allows the same individual actin filament to glide over the same myosin ensemble (50–1000 motors per ring) multiple times. First, we systematically characterized the formation of DNA nanotubes rings with 4, 6, 8, and 10 helix circumferences. Individual actin filaments glide along the nanotube rings with high processivity for up to 12.8 revolutions or 11 minutes in run time. We then show actin gliding speed is robust to variation in motor number and independent of ring curvature within our sample space (ring diameter of 0.5–4 μm). As a model application of OMEGA, we then analyze motor-based mechanical influence on “stop-and-go” gliding behavior of actin filaments, revealing that the stop-to-go transition probability is dependent on motor flexibility. Our circular gliding assay may provide a closed-loop platform for monitoring long-term behavior of broad classes of molecular motors and enable characterization of motor robustness and long timescale nanomechanical processes. |
| Databáze: | OpenAIRE |
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