cAPK-mediated tris-phosphorylation abolishes the binding of cardiac myosin binding protein C to myosin-S2, decreasing affinity from a Kd of ~5 microM to undetectable levels.
Phosphorylation of cardiac myosin binding protein C by cAPK acts as a 'brake-off' mechanism by abolishing its interaction with myosin-S2, freeing myosin heads to increase cardiac contractility.
Myosin binding protein C is a protein of the myosin filaments of striated muscle which is expressed in isoforms specific for cardiac and skeletal muscle. The cardiac isoform is phosphorylated rapidly upon adrenergic stimulation of myocardium by cAMP-dependent protein kinase, and together with the phosphorylation of troponin-I and phospholamban contributes to the positive inotropy that results from adrenergic stimulation of the heart. Cardiac myosin binding protein C is phosphorylated by cAMP-dependent protein kinase on three sites in a myosin binding protein C specific N-terminal domain which binds to myosin-S2. This interaction with myosin close to the motor domain is likely to mediate the regulatory function of the protein. Cardiac myosin binding protein C is a common target gene of familial hypertrophic cardiomyopathy and most mutations encode N-terminal subfragments of myosin binding protein C. The understanding of the signalling interactions of the N-terminal region is therefore important for understanding the pathophysiology of myosin binding protein C associated cardiomyopathy. We demonstrate here by cosedimentation assays and isothermal titration calorimetry that the myosin-S2 binding properties of the myosin binding protein C motif are abolished by cAMP-dependent protein kinase-mediated tris-phosphorylation, decreasing the S2 affinity from a Kd of approximately 5 microM to undetectable levels. We show that the slow and fast skeletal muscle isoforms are no cAMP-dependent protein kinase substrates and that the S2 interaction of these myosin binding protein C isoforms is therefore constitutively on. The regulation of cardiac contractility by myosin binding protein C therefore appears to be a 'brake-off' mechanism that will free a specific subset of myosin heads from sterical constraints imposed by the binding to the myosin binding protein C motif.
Gruen et al. (1999) studied Familial hypertrophic cardiomyopathy. cAPK-mediated tris-phosphorylation vs. Unphosphorylated state was evaluated on Myosin-S2 binding affinity. cAPK-mediated tris-phosphorylation abolishes the binding of cardiac myosin binding protein C to myosin-S2, decreasing affinity from a Kd of ~5 microM to undetectable levels.