Kinetic and regulatory properties of myosin adenosinetriphosphatase purified from arterial smooth muscle.

Abstract

Myosin was prepared from bovine arterial smooth muscle, and the properties of its ATPase [EC 3.6.1.3] reaction were compared with those of skeletal muscle myosin. A protein fraction, called Fraction-4 (Fr-4), was isolated from a crude arterial myosin preparation, and its effects on the ATPase activity of a complex of arterial myosin with skeletal muscle F-actin were examined. The following results were obtained. The Ca2+- and EDTA-ATPase activities of arterial myosin were almost equal to those of skeletal muscle myosin. Furthermore, no significant differences were detected between arterial and skeletal muscle myosins in the rates of formation of reaction intermediates, in the Mg2+-ATPase reaction. The amounts of initial P1-brst of these myosins were equal, i.e. about 1 mol/4.8 × 105 g. The Mg2+-ATPase activity of arterial myosin was activated up to tenfold by adding skeletal muscle F-actin. However, the ATPase activity of reconstituted arterial actomyosin was much lower than that of skeletal muscle actomyosin. The sensitivity of the ATPase activity of arterial myosin B to Ca2+was unaffected by the addition of skeletal muscle F-actin, but was eliminated by the addition of skeletal muscle myosin. These results suggested that the Ca2+ regulating system of arterial muscle is linked with myosin. Actomyosin reconstituted from purified arterial myosin and skeletal muscle F-actin showed a low ATPase activity with low Ca2+-sensitivity When the ATPase reaction was started by adding ATP in the presence of the protein fraction Fr-4, actomyosin showed a high ATPase activity with high Ca2+-sensitivity Arterial myosin was incubated with ATP in the presence of Ca2+ ions and Fr-4, and then myosin was isolated. Myosin thus “modified” showed a high actin-activated ATPase activity. The high ATPase activity of “modified” actomyosin was unaffected by removal of Ca2+ ions even in the presence of Fr-4, but was markedly inhibited by removal of Ca2+ ions when the skeletal muscle tropomyosin-troponin system was added.