title: Harmonic force spectroscopy measures load-dependent kinetics of individual human β-cardiac myosin molecules.
abstract: Molecular motors are responsible for numerous cellular processes from cargo transport to heart contraction. Their interactions with other cellular components are often transient and exhibit kinetics that depend on load. Here, we measure such interactions using 'harmonic force spectroscopy'. In this method, harmonic oscillation of the sample stage of a laser trap immediately, automatically and randomly applies sinusoidally varying loads to a single motor molecule interacting with a single track along which it moves. The experimental protocol and the data analysis are simple, fast and efficient. The protocol accumulates statistics fast enough to deliver single-molecule results from single-molecule experiments. We demonstrate the method's performance by measuring the force-dependent kinetics of individual human β-cardiac myosin molecules interacting with an actin filament at physiological ATP concentration. We show that a molecule's ADP release rate depends exponentially on the applied load, in qualitative agreement with cardiac muscle, which contracts with a velocity inversely proportional to external load.
authors.affiliation: 1] Department of Biochemistry, Stanford University School of Medicine, Stanford, California 94305, USA [2] Department of Applied Physics, Stanford University, Stanford, California 94305, USA.
authors.forename: Jongmin
authors.initials: J
authors.lastname: Sung
authors.affiliation: Department of Biochemistry, Stanford University School of Medicine, Stanford, California 94305, USA.
authors.forename: Suman
authors.initials: S
authors.lastname: Nag
authors.affiliation: Department of Micro- and Nanotechnology, Technical University of Denmark, DK-2800 Kongens Lyngby, Denmark.
authors.forename: Kim I
authors.initials: KI
authors.lastname: Mortensen
authors.affiliation: Department of Micro- and Nanotechnology, Technical University of Denmark, DK-2800 Kongens Lyngby, Denmark.
authors.forename: Christian L
authors.identifier: 000000015329475X
authors.initials: CL
authors.lastname: Vestergaard
authors.affiliation: Department of Biochemistry, Stanford University School of Medicine, Stanford, California 94305, USA.
authors.forename: Shirley
authors.initials: S
authors.lastname: Sutton
authors.affiliation: 1] Department of Biochemistry, Stanford University School of Medicine, Stanford, California 94305, USA [2] Department of Pediatrics (Cardiology), Stanford University School of Medicine, Stanford, California 94305, USA.
authors.forename: Kathleen
authors.initials: K
authors.lastname: Ruppel
authors.affiliation: Department of Micro- and Nanotechnology, Technical University of Denmark, DK-2800 Kongens Lyngby, Denmark.
authors.forename: Henrik
authors.identifier: 0000000216919367
authors.initials: H
authors.lastname: Flyvbjerg
authors.affiliation: Department of Biochemistry, Stanford University School of Medicine, Stanford, California 94305, USA.
authors.forename: James A
authors.initials: JA
authors.lastname: Spudich
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism*; D000244:Adenosine Diphosphate / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D007834:Lasers; D013057:Spectrum Analysis; D024723:Ventricular Myosins / Q000378:metabolism*
chemical_list: D000244:Adenosine Diphosphate; D024723:Ventricular Myosins