title: Velocity-dependent actomyosin ATPase cycle revealed by in vitro motility assay with kinetic analysis.
abstract: The actomyosin interaction plays a key role in a number of cellular functions. Single-molecule measurement techniques have been developed to study the mechanism of the actomyosin contractile system. However, the behavior of isolated single molecules does not always reflect that of molecules in a complex system such as a muscle fiber. Here, we developed a simple method for studying the kinetic parameters of the actomyosin interaction using small numbers of molecules. This approach does not require the specialized equipment needed for single-molecule measurements, and permits us to observe behavior that is more similar to that of a complex system. Using an in vitro motility assay, we examined the duration of continuous sliding of actin filaments on a sparsely distributed heavy meromyosin-coated surface. To estimate the association rate constant of the actomyosin motile system, we compared the distribution of experimentally obtained duration times with a computationally simulated distribution. We found that the association rate constant depends on the sliding velocity of the actin filaments. This technique may be used to reveal new aspects of the kinetics of various motor proteins in complex systems.
authors.affiliation: Institute of Multidisciplinary Research for Advanced Materials, Tohoku University, Sendai, Japan.
authors.forename: Masaaki K
authors.initials: MK
authors.lastname: Sato
authors.forename: Takashi
authors.initials: T
authors.lastname: Ishihara
authors.forename: Hiroto
authors.initials: H
authors.lastname: Tanaka
authors.forename: Akihiko
authors.initials: A
authors.lastname: Ishijima
authors.forename: Yuichi
authors.initials: Y
authors.lastname: Inoue
mesh_terms: D008841:Actin Cytoskeleton / Q000187:drug effects / Q000378:metabolism; D000205:Actomyosin / Q000378:metabolism*; D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D054443:Cell Migration Assays / Q000379:methods*; D007700:Kinetics; D015879:Myosin Subfragments / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D011817:Rabbits
chemical_list: D015879:Myosin Subfragments; D000255:Adenosine Triphosphate; D000205:Actomyosin; D000251:Adenosine Triphosphatases