In vitro and in vivo single myosin step-sizes in striated muscle

Thomas P. Burghardt, Xiaojing Sun, Yihua Wang, Katalin Ajtai

Research output: Contribution to journalArticle

4 Scopus citations

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.

Original languageEnglish (US)
Pages (from-to)463-477
Number of pages15
JournalJournal of Muscle Research and Cell Motility
Volume36
Issue number6
DOIs
StatePublished - Dec 1 2015

Keywords

  • Cardiac myosin step-frequency
  • Cardiac myosin step-size
  • High throughput Qdot assay
  • In vivo single myosin imaging
  • Second characterization
  • Skeletal muscle myosin mechanics

ASJC Scopus subject areas

  • Biochemistry
  • Physiology
  • Cell Biology

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