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Abstract
Previous studies using solubilized fragments of myosins have shown that an ATP analogue, 8-bromoadenosine triphosphate (8-Br-ATP) is a poor substrate for fast skeletal myosin isoform. We further characterized the analogue by using vertebrate skeletal muscle fibers. In the absence of calcium, the rate of 8-Br-ATP hydrolysis by fibers was higher than that for ATP, but it kept the fibers relaxed. The X-ray diffraction patterns of fibers relaxed by 8-Br-ATP were also indistinguishable from those of fibers relaxed by ATP, but higher concentrations were needed to keep the fibers relaxed. In the presence of calcium, the fibers exhibited force development and active shortening to varying extents. Although some of the energy for the observed contractility could be ascribed to the trace ATP in the reagents, the fibers activated in 8-Br-ATP performed much more mechanical work than expected from the energy of the trace ATP alone. The results suggest that most of the hydrolytic products of 8-Br-ATP dissociate from myosin prematurely, but a small fraction of myosin with these products does enter the calcium-dependent work-producing pathway and complete the normal process of chemo-mechanical conversion.