title: In vitro and in vivo single myosin step-sizes in striated muscle.
abstract: Myosin in muscle transduces ATP free energy into the mechanical work of moving actin. It has a motor domain transducer containing ATP and actin binding sites, and, mechanical elements coupling motor impulse to the myosin filament backbone providing transduction/mechanical-coupling. The mechanical coupler is a lever-arm stabilized by bound essential and regulatory light chains. The lever-arm rotates cyclically to impel bound filamentous actin. Linear actin displacement due to lever-arm rotation is the myosin step-size. A high-throughput quantum dot labeled actin in vitro motility assay (Qdot assay) measures motor step-size in the context of an ensemble of actomyosin interactions. The ensemble context imposes a constant velocity constraint for myosins interacting with one actin filament. In a cardiac myosin producing multiple step-sizes, a "second characterization" is step-frequency that adjusts longer step-size to lower frequency maintaining a linear actin velocity identical to that from a shorter step-size and higher frequency actomyosin cycle. The step-frequency characteristic involves and integrates myosin enzyme kinetics, mechanical strain, and other ensemble affected characteristics. The high-throughput Qdot assay suits a new paradigm calling for wide surveillance of the vast number of disease or aging relevant myosin isoforms that contrasts with the alternative model calling for exhaustive research on a tiny subset myosin forms. The zebrafish embryo assay (Z assay) performs single myosin step-size and step-frequency assaying in vivo combining single myosin mechanical and whole muscle physiological characterizations in one model organism. The Qdot and Z assays cover "bottom-up" and "top-down" assaying of myosin characteristics.
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic Rochester, Rochester, MN, 55905, USA. burghardt@mayo.edu.|Department of Physiology and Biomedical Engineering, Mayo Clinic Rochester, Rochester, MN, 55905, USA. burghardt@mayo.edu.
authors.forename: Thomas P
authors.initials: TP
authors.lastname: Burghardt
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic Rochester, Rochester, MN, 55905, USA.
authors.forename: Xiaojing
authors.initials: X
authors.lastname: Sun
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic Rochester, Rochester, MN, 55905, USA.
authors.forename: Yihua
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic Rochester, Rochester, MN, 55905, USA.
authors.forename: Katalin
authors.initials: K
authors.lastname: Ajtai
mesh_terms: D000199:Actins / Q000378:metabolism; D000818:Animals; D018482:Muscle, Skeletal / Q000378:metabolism*; D054792:Muscle, Striated / Q000378:metabolism*; D018994:Myosin Light Chains / Q000378:metabolism*; D013552:Swine
chemical_list: D000199:Actins; D018994:Myosin Light Chains
keywords: Cardiac myosin step-frequency; Cardiac myosin step-size; High throughput Qdot assay; In vivo single myosin imaging; Second characterization; Skeletal muscle myosin mechanics