E. Prochniewicz and D.D. Thomas
Department of Biochemistry, University of Minnesota, 4-225 Millard Hall, 435 Delaware St., SE Minneapolis, NM55455
Cooperativity within an actin filament in its interaction with myosin was examined by measuring the effects of structural and functional perturbations on the time-resolved anisotropy (TPA) of erythrosin iodoacetamide (ErIA) labeled actin (at Cys 374) and its complexes with S1. The first perturbation -- binding of Fab (1-7) to the N-terminus of actin - allowed us to examine the effects of changes in the sites of weak myosin interaction on actin rotational dynamics. The second perturbation -- copolymerization of ErIA-labeled monomers with unlabeled, EDC-treated ones -- allowed us to examine the effect of local changes in the filament on the interaction of the unmodified segments with myosin. In a parallel set of experiments, we studied the effects of perturbations of actin on the configuration of bound S1 labeled with ErIA. The functional effects of these perturbations were inhibition of the in vitro motility and activation of the myosin ATPase. The results are discussed in terms of the mechanism of regulation of functional interactions with myosin by long-range structural changes within the actin filament.