title: Kinetics of association of myosin subfragment-1 to unlabeled and pyrenyl-labeled actin.
abstract: The kinetics of reaction of myosin subfragment-1 (S1) with F-actin have been monitored by the changes in light scattering and in pyrenyl-actin fluorescence at 20 degrees C, pH 7.5, and physiological ionic strength. The association rate constant of S1 to F-actin decreases about 10-fold as the molar ratio of bound S1 increases from 0 to 1. This decrease in k+ is most likely due to the steric hindrance of available binding sites by initially bound S1. The apparent rate constant for association of S1 to bare filaments is 9 microM-1 s-1, a value 1 order of magnitude higher than the one previously estimated from experiments in which S1 was in excess over F-actin. The anticooperative binding kinetics of S1 to F-actin are consistent with the negative cooperativity displayed in the equilibrium binding curves of S1 to pyrenyl-F-actin. Fluorescence titration curves of partially labeled pyrenyl-F-actin by S1 are sigmoidal, consistent with a 4-fold higher affinity of S1 for unlabeled than for labeled action. This conclusion is strengthened by kinetic data of S1 binding to partially labeled F-actin, which exhibit a biphasic behavior due to the slower dissociation of S1 from unlabeled than from labeled actin.
authors.affiliation: Laboratoire d'Enzymolgie, CNRS, 91198 Gif-sur-Yvette, France.
authors.forename: L
authors.initials: L
authors.lastname: Blanchoin
authors.forename: D
authors.initials: D
authors.lastname: Didry
authors.forename: M F
authors.initials: MF
authors.lastname: Carlier
authors.forename: D
authors.initials: D
authors.lastname: Pantaloni
mesh_terms: D000199:Actins / Q000737:chemistry* / Q000378:metabolism; D000818:Animals; D007700:Kinetics; D008027:Light; D015879:Myosin Subfragments / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D011721:Pyrenes / Q000737:chemistry*; D011817:Rabbits; D012542:Scattering, Radiation; D013696:Temperature
chemical_list: D000199:Actins; D015879:Myosin Subfragments; D011721:Pyrenes