title: Effects of the N-terminal domains of myosin binding protein-C in an in vitro motility assay: Evidence for long-lived cross-bridges.
abstract: Myosin binding protein-C (MyBP-C) is a thick-filament protein whose precise function within the sarcomere is not known. However, recent evidence from cMyBP-C knock-out mice that lack MyBP-C in the heart suggest that cMyBP-C normally slows cross-bridge cycling rates and reduces myocyte power output. To investigate possible mechanisms by which cMyBP-C limits cross-bridge cycling kinetics we assessed effects of recombinant N-terminal domains of MyBP-C on the ability of heavy meromyosin (HMM) to support movement of actin filaments using in vitro motility assays. Here we show that N-terminal domains of cMyBP-C containing the MyBP-C "motif," a sequence of approximately 110 amino acids, which is conserved across all MyBP-C isoforms, reduced actin filament velocity under conditions where filaments are maximally activated (i.e. either in the absence of thin filament regulatory proteins or in the presence of troponin and tropomyosin and high [Ca2+]). By contrast, under conditions where thin filament sliding speed is submaximal (i.e. in the presence of troponin and tropomyosin and low [Ca2+]), proteins containing the motif increased filament speed. Recombinant N-terminal proteins also bound to F-actin and inhibited acto-HMM ATPase rates in solution. The results suggest that N-terminal domains of MyBP-C slow cross-bridge cycling kinetics by reducing rates of cross-bridge detachment.
authors.affiliation: Department of Bioengineering, University of Washington, Seattle, Washington 98195, USA.
authors.forename: Maria V
authors.initials: MV
authors.lastname: Razumova
authors.forename: Justin F
authors.initials: JF
authors.lastname: Shaffer
authors.forename: An-Yue
authors.initials: AY
authors.lastname: Tu
authors.forename: Galina V
authors.initials: GV
authors.lastname: Flint
authors.forename: Michael
authors.initials: M
authors.lastname: Regnier
authors.forename: Samantha P
authors.initials: SP
authors.lastname: Harris
mesh_terms: D000199:Actins / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002352:Carrier Proteins / Q000737:chemistry*; D002465:Cell Movement; D007700:Kinetics; D008297:Male; D009068:Movement; D009206:Myocardium / Q000378:metabolism; D015879:Myosin Subfragments / Q000737:chemistry; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D011817:Rabbits; D011994:Recombinant Proteins / Q000737:chemistry; D000099163:Myosin Binding Protein C
chemical_list: D000199:Actins; D002118:Calcium; D002352:Carrier Proteins; D015879:Myosin Subfragments; D020033:Protein Isoforms; D011994:Recombinant Proteins; D000099163:Myosin Binding Protein C