Current theories of the force generating step in muscle contraction involve a reorientation of the myosin head relative to the actin filament during the contractile cycle. Implicit in this model is the existence of more than one conformational state of the head region in myosin. A major aim of our research continues to be the elucidation of such putative structural changes in the heavy and light chain subunits of myosin. We will pursue two main directions to advance this problem: (1) Specific markers, such as fluorescent probes, monoclonal antibodies, and antibodies against small peptides will be introduced into well-defined sites on the light and heavy chains. The position of the antibody on the myosin head will be visualized by electron microscopy of shadow-cast preparations. Distances between fluorophores will be measured by Forster energy transfer. Any change in energy transfer with ATP and/or actin binding may be indicative of structural changes within the head. (2) Comparative studies on the structure and function of widely different myosin isozymes can provide important insights not obtainable from any one myosin. Among the myosins we will continue to study are fast and slow adult myosins, embryonic and neonatal myosins, and smooth muscle myosin from avian and mammalian sources. The effect of light chain removal or phosphorylation on the conformation of myosin will be explored. Techniques will include kinetic analysis, peptide mapping, immunological, electrophoretic, and sedimentation analysis. By investigating these myosin polymorphs both in the monomeric and filamentous state, we may eventually understand how this complex molecule works, and why muscle is composed of such a seemingly endless mosaic of fiber types.