Dataset Information

Ca2+-induced movement of tropomyosin on native cardiac thin filaments revealed by cryoelectron microscopy.

ABSTRACT: Muscle contraction relies on the interaction of myosin motors with F-actin, which is regulated through a translocation of tropomyosin by the troponin complex in response to Ca2+ The current model of muscle regulation holds that at relaxing (low-Ca2+) conditions tropomyosin blocks myosin binding sites on F-actin, whereas at activating (high-Ca2+) conditions tropomyosin translocation only partially exposes myosin binding sites on F-actin so that binding of rigor myosin is required to fully activate the thin filament (TF). Here we used a single-particle approach to helical reconstruction of frozen hydrated native cardiac TFs under relaxing and activating conditions to reveal the azimuthal movement of the tropomyosin on the surface of the native cardiac TF upon Ca2+ activation. We demonstrate that at either relaxing or activating conditions tropomyosin is not constrained in one structural state, but rather is distributed between three structural positions on the surface of the TF. We show that two of these tropomyosin positions restrain actomyosin interactions, whereas in the third position, which is significantly enhanced at high Ca2+, tropomyosin does not block myosin binding sites on F-actin. Our data provide a structural framework for the enhanced activation of the cardiac TF over the skeletal TF by Ca2+ and lead to a mechanistic model for the regulation of the cardiac TF.

SUBMITTER: Risi C

PROVIDER: S-EPMC5495243 | biostudies-literature | 2017 Jun

REPOSITORIES: biostudies-literature

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Ca2+-induced movement of tropomyosin on native cardiac thin filaments revealed by cryoelectron microscopy.

Risi Cristina C Eisner Jamie J Belknap Betty B Heeley David H DH White Howard D HD Schröder Gunnar F GF Galkin Vitold E VE

Proceedings of the National Academy of Sciences of the United States of America 20170612 26

Muscle contraction relies on the interaction of myosin motors with F-actin, which is regulated through a translocation of tropomyosin by the troponin complex in response to Ca2+ The current model of muscle regulation holds that at relaxing (low-Ca2+) conditions tropomyosin blocks myosin binding sites on F-actin, whereas at activating (high-Ca2+) conditions tropomyosin translocation only partially exposes myosin binding sites on F-actin so that binding of rigor my ...[more]

PMID: 28607071

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Project description:Complete description of thin filament conformational transitions accompanying muscle regulation requires ready access to atomic structures of actin-bound tropomyosin-troponin. To date, several molecular-docking protocols have been employed to identify troponin interactions on actin-tropomyosin because high-resolution experimentally determined structures of filament-associated troponin are not available. However, previously published all-atom models of the thin filament show chain separation and corruption of components during our molecular dynamics simulations of the models, implying artifactual subunit organization, possibly due to incorporation of unorthodox tropomyosin-TnT crystal structures and complex FRET measurements during model construction. For example, the recent Williams et al. (2016) atomistic model of the thin filament displays a paucity of salt bridges and hydrophobic complementarity between the TnT tail (TnT1) and tropomyosin, which is difficult to reconcile with the high, 20 nM Kd binding of TnT onto tropomyosin. Indeed, our molecular dynamics simulations show the TnT1 component in their model partially dissociates from tropomyosin in under 100 ns, whereas actin-tropomyosin and TnT1 models themselves remain intact. We therefore revisited computational work aiming to improve TnT1-thin filament models by employing unbiased docking methodologies, which test billions of trial rotations and translations of TnT1 over three-dimensional grids covering end-to-end bonded tropomyosin alone or tropomyosin on F-actin. We limited conformational searches to the association of well-characterized TnT1 helical domains and either isolated tropomyosin or actin-tropomyosin yet avoided docking TnT domains that lack known or predicted structure. The docking programs PIPER and ClusPro were used, followed by interaction energy optimization and extensive molecular dynamics. TnT1 docked to either side of isolated tropomyosin but uniquely onto one location of actin-bound tropomyosin. The antiparallel interaction with tropomyosin contained abundant salt bridges and intimately integrated hydrophobic networks joining TnT1 and the tropomyosin N-/C-terminal overlapping domain. The TnT1-tropomyosin linkage yields well-defined molecular crevices. Interaction energy measurements strongly favor this TnT1-tropomyosin design over previously proposed models.

Dataset Information

Ca2+-induced movement of tropomyosin on native cardiac thin filaments revealed by cryoelectron microscopy.

Publications

Ca<sup>2+</sup>-induced movement of tropomyosin on native cardiac thin filaments revealed by cryoelectron microscopy.

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