title: Kinetic characterization of the weak binding states of myosin V.
abstract: Myosin V is a molecular motor shown to move processively along actin filaments. We investigated the properties of the weak binding states of monomeric myosin V containing a single IQ domain (MV 1IQ) to determine if the affinities of these states are increased as compared to conventional myosin. Further, using a combination of non-hydrolyzable nucleotide analogues and mutations that block ATP hydrolysis, we sought to probe the states that are populated during ATP-induced dissociation of actomyosin. MV 1IQ binds actin with a K(d) = 4 microM in the presence of ATP gamma S at 50 mM KCl, which is 10-20-fold tighter than that of nonprocessive class II myosins. Mutations within the switch II region trapped MV 1IQ in two distinct M.ATP states with very different actin binding affinities (K(d) = 0.2 and 2 microM). Actin binding may change the conformation of the switch II region, suggesting that elements of the nucleotide binding pocket will be in a different conformation when bound to actin than is seen in any of the myosin crystal structures to date.
authors.affiliation: University of Pennsylvania School of Medicine, Department of Physiology and Pennsylvania Muscle Institute, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Christopher M
authors.initials: CM
authors.lastname: Yengo
authors.forename: Enrique M
authors.initials: EM
authors.lastname: De la Cruz
authors.forename: Daniel
authors.initials: D
authors.lastname: Safer
authors.forename: E Michael
authors.initials: EM
authors.lastname: Ostap
authors.forename: H Lee
authors.initials: HL
authors.lastname: Sweeney
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000266:Adenylyl Imidodiphosphate / Q000493:pharmacokinetics; D000818:Animals; D001665:Binding Sites; D018076:DNA, Complementary; D007700:Kinetics; D051379:Mice; D024701:Myosin Type V / Q000737:chemistry / Q000378:metabolism*; D009218:Myosins / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013050:Spectrometry, Fluorescence
chemical_list: D018076:DNA, Complementary; D011994:Recombinant Proteins; D000266:Adenylyl Imidodiphosphate; D000255:Adenosine Triphosphate; D024701:Myosin Type V; D009218:Myosins