Perturbations of functional interactions with myosin induce long-range allosteric and cooperative structural changes in actin.

The role of the rotational dynamics of actin filaments in their interaction with myosin was studied by comparing the effect of myosin subfragment 1 (S1) with two other structural perturbations, which have substantial inhibitory effects on activation of myosin ATPase and in vitro motility of F-actin: (1) binding of the antibody fragment Fab(1-7) against the first seven N-terminal residues and (2) copolymerization with monomers treated with the zero-length cross-linker 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide (EDC), referred to as EDC-actin. The rotational motion of actin was measured by time-resolved phosphorescence anisotropy (TPA) of erythrosin iodoacetamide (ErIA) attached to Cys 374 on actin. The binding of S1 in a rigor complex (no nucleotide) induced intramonomer (allosteric) and intermonomer (cooperative) structural changes that increased the residual anisotropy of labeled F-actin, indicating a conformational change in the region of the C terminus. Similar allosteric and cooperative changes were induced by binding of Fab(1-7) and by copolymerization of the ErIA-labeled actin monomers with EDC-actin. This suggests that the functional perturbations transform actin to a form resembling the rigor actomyosin complex. The correlation of the perturbation-induced changes in TPA of actin with the functional effects suggests that the actomyosin interaction can be inhibited by stabilization of actin in one of its structural intermediates.