title: Phosphorylation of tropomyosin extends cooperative binding of myosin beyond a single regulatory unit.
abstract: Tropomyosin (Tm) is one of the major phosphoproteins comprising the thin filament of muscle. However, the specific role of Tm phosphorylation in modulating the mechanics of actomyosin interaction has not been determined. Here we show that Tm phosphorylation is necessary for long-range cooperative activation of myosin binding. We used a novel optical trapping assay to measure the isometric stall force of an ensemble of myosin molecules moving actin filaments reconstituted with either natively phosphorylated or dephosphorylated Tm. The data show that the thin filament is cooperatively activated by myosin across regulatory units when Tm is phosphorylated. When Tm is dephosphorylated, this "long-range" cooperative activation is lost and the filament behaves identically to bare actin filaments. However, these effects are not due to dissociation of dephosphorylated Tm from the reconstituted thin filament. The data suggest that end-to-end interactions of adjacent Tm molecules are strengthened when Tm is phosphorylated, and that phosphorylation is thus essential for long range cooperative activation along the thin filament.
authors.affiliation: Department of Biomedical Engineering, University of Virginia, Charlottesville, VA 22908, USA.
authors.forename: Vijay S
authors.initials: VS
authors.lastname: Rao
authors.forename: Ellisha N
authors.initials: EN
authors.lastname: Marongelli
authors.forename: William H
authors.initials: WH
authors.lastname: Guilford
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism; D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D001696:Biomechanical Phenomena; D009218:Myosins / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D051381:Rats; D014335:Tropomyosin / Q000378:metabolism*
chemical_list: D010750:Phosphoproteins; D014335:Tropomyosin; D000469:Alkaline Phosphatase; D009218:Myosins