The Three-Dimensional Model Of The Tropomodulin Assembly At The Pointed End Of Actin Filament

Actin filaments have the pointed (slow growing) end and the barbed (fast growing) end. Interactions with actin-binding proteins at each end enable control of the length of actin filaments. Tropomodulin (Tmod) modulates actin filament length by interacting with actin and tropomyosin (Tpm) at the pointed end of the filament. Tmod binds two Tpm molecules through two distinct Tpm binding sites – site 1 and site 2. Establishing the structure of each Tmod-Tpm binding interface is critical to understand the mechanism of regulation of the pointed end dynamics by Tmod. In this study, we built three-dimensional models of the complexes by using (1) structural information obtained from NMR data for complexes of Tpm and Tmod fragments representing the binding sites, and (2) secondary structure predictions. To refine the models, we employed simulated annealing molecular dynamics followed by conventional molecular dynamics simulations. The refined modeled structures reveal similarities between the complexes of Tmod2s1 and Tmod2s2 (representing sites 1 and 2, respectively) with the Tpm fragment. In both complexes, Tmod2s1 and Tmod2s2 form an α-helical hairpin arranged with the Tpm coiled coil in a four-α-helix bundle. Positioning of Tmod2s1 and Tmod2s2 over the N-terminus of Tpm blocks further Tpm polymerization and prevents elongation of the Tpm cable. Both complexes are stabilized by a hydrophobic core formed by residues that are conserved between the two sites. The two models and known structures of actin binding sites were used to dock Tmod to the pointed end of the actin filament associated with tropomyosin cables. The model of the pointed-end Tmod assembly offers new insights into how Tmod binding ensures tight control over the pointed end dynamics.