Substructure and Communication in Myosin and Actin

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The contraction of skeletal, cardiac, and smooth muscles requires cyclic interactions between the two main muscle proteins, myosin and actin. The essential steps in these interactions include the activation of the myosin ATPase activity by actin and, according to prevalent views, conformational changes on one or both of these proteins. A major goal of the proposed research is to clarify the molecular basis of these events. To achieve aim, actin sites necessary for the activation of the myosin ATPase activity and the motility of actin filaments will be mapped. The experimental materials to be used in the mapping of functional sites on actin will include peptide antibodies, actin mutants, and modified actins. These materials will be used in the in vitro motility assays, and in biochemical and biophysical assays of actomyosin interactions. In another part of this program, the structural determinants of the essential steps in actomyosin interactions will be clarified by comparing the properties of monomeric and polymeric complexes of actin and myosin. Peptides, protein fragments, cross- linked protein complexes, peptide antibodies, and actin mutants will be used in this project. The role of specific sites on actin in actomyosin interactions will be assessed via chemical modifications, proteolytic digestions, fluorescence and light scattering measurements, and other measurements carried out on these materials. Additional insight into actomyosin interactions will be obtained by clarifying the mechanism of G-actin polymerization by myosin. %%% The proposed studies should lead to the elucidation of the basic mechanism of actomyosin interactions and thus, a better understanding of how the muscle functions and how chemical energy is transduced into work in biological systems.