title: The superfast human extraocular myosin is kinetically distinct from the fast skeletal IIa, IIb, and IId isoforms.
abstract: Humans express five distinct myosin isoforms in the sarcomeres of adult striated muscle (fast IIa, IId, the slow/cardiac isoform I/β, the cardiac specific isoform α, and the specialized extraocular muscle isoform). An additional isoform, IIb, is present in the genome but is not normally expressed in healthy human muscles. Muscle fibers expressing each isoform have distinct characteristics including shortening velocity. Defining the properties of the isoforms in detail has been limited by the availability of pure samples of the individual proteins. Here we study purified recombinant human myosin motor domains expressed in mouse C2C12 muscle cells. The results of kinetic analysis show that among the closely related adult skeletal isoforms, the affinity of ADP for actin·myosin (K(AD)) is the characteristic that most readily distinguishes the isoforms. The three fast muscle myosins have K(AD) values of 118, 80, and 55 μM for IId, IIa, and IIb, respectively, which follows the speed in motility assays from fastest to slowest. Extraocular muscle is unusually fast with a far weaker K(AD) = 352 μM. Sequence comparisons and homology modeling of the structures identify a few key areas of sequence that may define the differences between the isoforms, including a region of the upper 50-kDa domain important in signaling between the nucleotide pocket and the actin-binding site.
authors.affiliation: School of Biosciences, University of Kent, Canterbury, CT2 7NJ, United Kingdom.
authors.forename: Marieke J
authors.initials: MJ
authors.lastname: Bloemink
authors.affiliation: Department of Molecular, Cellular & Developmental Biology, University of Colorado, Boulder, Colorado 80309.
authors.forename: John C
authors.initials: JC
authors.lastname: Deacon
authors.affiliation: Department of Molecular, Cellular & Developmental Biology, University of Colorado, Boulder, Colorado 80309.
authors.forename: Daniel I
authors.initials: DI
authors.lastname: Resnicow
authors.affiliation: Department of Molecular, Cellular & Developmental Biology, University of Colorado, Boulder, Colorado 80309. Electronic address: leslie.leinwand@colorado.edu.
authors.forename: Leslie A
authors.initials: LA
authors.lastname: Leinwand
authors.affiliation: School of Biosciences, University of Kent, Canterbury, CT2 7NJ, United Kingdom. Electronic address: m.a.geeves@kent.ac.uk.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Geeves
mesh_terms: D000818:Animals; D001665:Binding Sites; D024722:Cardiac Myosins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002461:Cell Line, Transformed; D006801:Humans; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D051379:Mice; D018995:Myosin Heavy Chains / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D009801:Oculomotor Muscles / Q000737:chemistry* / Q000378:metabolism; D009928:Organ Specificity / Q000502:physiology; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D024744:Skeletal Muscle Myosins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D007527:Isoenzymes; D011994:Recombinant Proteins; D024722:Cardiac Myosins; D024744:Skeletal Muscle Myosins; D018995:Myosin Heavy Chains
keywords: Actin; Fluorescence; Homology Modeling; Kinetics; Muscle; Myosin; Protein Structure-Function; Sequence Alignment
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