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Facilitated cross-bridge interactions with thin filaments by familial hypertrophic cardiomyopathy mutations in ?-tropomyosin.

ABSTRACT: Familial hypertrophic cardiomyopathy (FHC) is a disease of cardiac sarcomeres. To identify molecular mechanisms underlying FHC pathology, functional and structural differences in three FHC-related mutations in recombinant ?-Tm (V95A, D175N, and E180G) were characterized using both conventional and modified in vitro motility assays and circular dichroism spectroscopy. Mutant Tm's exhibited reduced ?-helical structure and increased unordered structure. When thin filaments were fully occupied by regulatory proteins, little or no motion was detected at pCa 9, and maximum speed (pCa 5) was similar for all tropomyosins. Ca(2+)-responsiveness of filament sliding speed was increased either by increased pCa(50) (V95A), reduced cooperativity n (D175N), or both (E180G). When temperature was increased, thin filaments with E180G exhibited dysregulation at temperatures ~10°C lower, and much closer to body temperature, than WT. When HMM density was reduced, thin filaments with D175N required fewer motors to initiate sliding or achieve maximum sliding speed.

SUBMITTER: Wang F

PROVIDER: S-EPMC3237018 | biostudies-literature | 2011

REPOSITORIES: biostudies-literature

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Facilitated cross-bridge interactions with thin filaments by familial hypertrophic cardiomyopathy mutations in α-tropomyosin.

Wang Fang F Brunet Nicolas M NM Grubich Justin R JR Bienkiewicz Ewa A EA Asbury Thomas M TM Compton Lisa A LA Mihajlović Goran G Miller Victor F VF Chase P Bryant PB

Journal of biomedicine & biotechnology 20111201

Familial hypertrophic cardiomyopathy (FHC) is a disease of cardiac sarcomeres. To identify molecular mechanisms underlying FHC pathology, functional and structural differences in three FHC-related mutations in recombinant α-Tm (V95A, D175N, and E180G) were characterized using both conventional and modified in vitro motility assays and circular dichroism spectroscopy. Mutant Tm's exhibited reduced α-helical structure and increased unordered structure. When thin filaments were fully occupied by re ...[more]

PMID: 22187526

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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

Facilitated cross-bridge interactions with thin filaments by familial hypertrophic cardiomyopathy mutations in ?-tropomyosin.

Publications

Facilitated cross-bridge interactions with thin filaments by familial hypertrophic cardiomyopathy mutations in α-tropomyosin.

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