Caldesmon-actin-tropomyosin contains two types of binding sites for myosin S1.

Caldesmon inhibits the activation of myosin ATPase activity by actin-tropomyosin. Caldesmon also inhibits the binding of myosin to actin. There is disagreement as to the degree to which competitive displacement of myosin subfragment binding to actin is responsible for the inhibition of ATPase activity. We have examined the possibility that one or more molecules of S1 may bind to actin-tropomyosin-caldesmon without having the normal actin activation of ATPase activity. The effect of caldesmon on the binding and ATPase activity of S1 was measured at several initial levels of saturation of S1 to determine if a fraction of the bound S1 was resistant to displacement by caldesmon. In the case of both unmodified S1 and rhoPDM-modified S1, most, but not all, of the S1 was displaced by caldesmon. The results are consistent with a single molecule of S1 binding with low affinity for each seven actin monomers that are fully saturated with caldesmon and tropomyosin. This single S1 is not necessarily bound directly to actin but may be attached to the NH2-terminal region of caldesmon.