The substructure of the cardiac myosin molecule was examined by the limited proteolytic digestion of the parent molecule with (dialdehyde starch)-methylenedianiline–mercuripapain, S-MDA–mercuripapain, at low temperatures and neutral pH, using moderate enzyme to myosin ratios. Pertinent properties of the insoluble enzyme complex were also examined. Kinetic, ultracentrifugal, and chromatographic observations of the fragmentation process revealed that a single type of lytic reaction occurs during the early stages, predominantly releasing heavy meromyosin subfragment 1 (HMM-S1) and myosin rods. With further time of digestion, the rods are additionally cleaved yielding light meromyosin and HMM-S2, and HMM-S1 is found to be partially degraded. The major proteolytic subfragments were isolated, purified, and characterized with respect to their enzymatic, optical, amino acid, and physicochemical properties. Only HMM-S1 exhibited Ca2+-activated ATPase activity, and at a level three- to fourfold higher than that of native myosin. Moreover, its hydrodynamic properties suggest that it is globular in structure. On the other hand, light meromyosin-A (LMM-A) (which consists mainly of rods), and HMM-S2 appear to be highly asymmetric, rigid, α-helical molecules devoid of the amino acid proline. Strong similarities were evident in all aspects upon comparison of these results with documented information concerning the skeletal system. On the basis of the physical and chemical properties of the proteolytic subfragments relative to that of native myosin, it was further concluded that the cardiac myosin molecule is a double-stranded, α-helical rod ending in two subfragment 1 globules, of which only one may be enzymatically active at a time.
A Small Chemical Mimicking Actin Binding to Myosin and Putative Structural Changes of Myosin Molecule upon Ligand Binding