The role of calcium in the adensine triphosphatase activity of myofibrils and in the mechanism of the relaxing factor system of muscle.

Several factors tend to create the impression that the removal of Ca from the myofibrils plays an important role in the mechanism of relaxation. Among these factors are the inhibition of the activity of the relaxing factor system of muscle by Ca (2) ; the similarity between the effect of ethylenediaminetetraacetate and the relaxing factor system in producing inhibition of myofibrillar adenosine triphosphatase (3) and reversal of the adenosine triphosphate-induced contraction of glycerinated muscle fiber bundles (4, 5); the binding of Ca to the so called granules (a constituent of the relaxing factor system), which, as recently shown, requires the presence of ATP’ (cf. (1)) ; and the dependence of the myofibrillar ATPase on the presence of small amounts of Ca (6). Although Bozler has shown that about half of the Ca bound to muscle fibers is removed by EDTA2 when relaxation occurs, after washing, even in the absence of Gaff, relaxed fibers contract again upon the addition of ATP (4,7). He therefore interpreted relaxation in terms of the combination of relaxing agents with a site associated with firmly bound Ca. Our studies, involving the comparison of the effect of Ca on both the complete relaxing factor system and the recently described soluble relaxing substance (8)) suggest that it is the latter which is inhibited by Ca, or Ca-ATP, and that granules, in addition to producing the relaxing substance, combine with Ca in maintaining conditions favorable for the activity of the relaxing substance. In the present report, direct evidence is presented to show that treatment with granules or EDTA does not lead to the removal from myofibrils of a Ca fraction essential to their ATPase activity, and that the mechanism of relaxing activity is most likely to depend on the binding of the relaxing substance or EDTA to the contractile proteins. The experiments show the presence of a readily removable and exchangeable, but apparently