Tropomyosin Movement on F-actin Analyzed by Energy Landscape Determination

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Muscle contraction is regulated by movement of tropomyosin over the surface of actin filaments. At low-Ca2+, tropomyosin blocks myosin-binding on actin, whereas, in a two-step process, Ca2+-saturated troponin and myosin open the myosin-binding site leading to contraction. No obvious steric obstructions or geometrical barriers on actin limit such tropomyosin movement. However, lacking atomic models for these transitions, the pathways taken by tropomyosin during regulatory movements are uncertain. Here, end-points for regulatory transitions were determined by fitting tropomyosin to EM reconstructions. Reconstructions of negatively-stained low-Ca2+ thin filaments yield an atomic model very close to that described for troponin-free actin-tropomyosin by Li et al. (2011), while comparable high-Ca2+ filament maps suggest an azimuthal sliding of tropomyosin parallel to its superhelical path. Additionally, Raunser et al. (2012-Biophys. Soc. abst.) describe an atomic model of tropomyosin based on high-resolution cryo-EM reconstructions of myosin-decorated filaments, showing expected azimuthal movement and pronounced ∼15A axial displacement of tropomyosin toward the pointed-end of the thin filament. To evaluate transitions between these various regulatory positions, we explored the energy landscape between “end-states” over a comprehensive grid of 832 tropomyosin locations relative to F-actin coordinates. The position of tropomyosin was varied azimuthally and axially relative to F-actin, and then the structure energy-minimized. The resulting electrostatic energy landscape shows a wide energy basin with a minimum centered near the blocked-state. The width of this basin indicates that large azimuthal and axial oscillations of tropomyosin are possible. By contrast, in the myosin-induced open-position, tropomyosin is located at an energy peak, representing a region with no obvious complementary electrostatic interactions between tropomyosin and F-actin. Our results therefore suggest that the open-position is reached only because of tropomyosin interaction with myosin, while binding of tropomyosin to the F-actin surface becomes less important.