title: Metal cation controls myosin and actomyosin kinetics.
abstract: We have perturbed myosin nucleotide binding site with magnesium-, manganese-, or calcium-nucleotide complexes, using metal cation as a probe to examine the pathways of myosin ATPase in the presence of actin. We have used transient time-resolved FRET, myosin intrinsic fluorescence, fluorescence of pyrene labeled actin, combined with the steady state myosin ATPase activity measurements of previously characterized D.discoideum myosin construct A639C:K498C. We found that actin activation of myosin ATPase does not depend on metal cation, regardless of the cation-specific kinetics of nucleotide binding and dissociation. The rate limiting step of myosin ATPase depends on the metal cation. The rate of the recovery stroke and the reverse recovery stroke is directly proportional to the ionic radius of the cation. The rate of nucleotide release from myosin and actomyosin, and ATP binding to actomyosin depends on the cation coordination number.
authors.affiliation: Department of Physics and Optical Science, University of North Carolina, Charlotte, North Carolina, 28223; Engelhardt Institute of Molecular Biology RAS, Moscow, 119991, Russia.
authors.forename: Yaroslav V
authors.initials: YV
authors.lastname: Tkachev
authors.forename: Jinghua
authors.initials: J
authors.lastname: Ge
authors.forename: Igor V
authors.initials: IV
authors.lastname: Negrashov
authors.forename: Yuri E
authors.initials: YE
authors.lastname: Nesmelov
mesh_terms: D000199:Actins / Q000378:metabolism*; D000205:Actomyosin / Q000737:chemistry* / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D002118:Calcium; D020134:Catalytic Domain; D002412:Cations / Q000378:metabolism*; D007700:Kinetics; D008274:Magnesium; D008345:Manganese; D008670:Metals / Q000378:metabolism*; D009218:Myosins / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D015398:Signal Transduction
chemical_list: D000199:Actins; D002412:Cations; D008670:Metals; D008345:Manganese; D000205:Actomyosin; D009218:Myosins; D008274:Magnesium; D002118:Calcium
keywords: FRET; calcium; fluorescence; magnesium; manganese; myosin; nucleotide