Smooth muscle and nonmuscle myosins are turned on by phosphorylation of the regulatory light chain, which is required to activate contraction in vivo. This phosphorylation event also stabilizes myosin filaments, the conformation from which force is generated. However, if unphosphorylated, these myosin isoforms can adopt and intra-molecularly folded conformation in which the tail domain is bent in thirds and interacts with the head domains. This folded conformation called 10S cannot bind to actin and has a very low ATP turnover rate. It may serve as a reservoir of fully synthesized myosin in the cell from which filaments can form depending upon the need for active filamentous myosin. We are interested in the structure of this inhibited 10S myosin. We have used photocross-linking and chemical cross-linking in concert with mass spectrometry to determine the interaction of the tail domain with the regulatory domain in an effort to understand the molecular basis of self-inhibition. Recently, it has been shown that other myosin isoforms and kinesin can also form similar intra-molecularly folded conformations. Therefore, the self-inhibitory structural mechanism may be common to motor proteins.