Structure of C protein purified from cardiac muscle.

Abstract

C protein is a component of the thick filament of striated muscles. Although the function of C protein remains unknown, a variety of evidence suggests that C protein may regulate actin-myosin interaction or be involved in structural support or elasticity of the sarcomere. We have previously proposed (Hartzell, H. C., 1984, J. Gen. Physiol., 83:563-588) that C protein is involved in regulating twitch relaxation in cardiac muscle. To gain further insight into the function of C protein, we have studied the structure of C protein purified from chicken heart. C protein was purified from extracts of detergent-washed myofibrils by sequential hydroxylapatite and DEAE-Sephacel chromatography. C protein was judged >95% pure by SDS PAGE. The polypeptide subunit had a molecular weight of 155,000 and the native molecule sedimented on linear sucrose or glycerol gradients at 4-5S. For electron microscopy, purified C protein was dialyzed and diluted into a volatile buffer in 50% glycerol, aspirated onto mica, dried under vacuum, and rotary platinum-shadowed. Replicas revealed particles of relatively homogeneous overall dimensions. Over half of the particles were Vshaped. The "arm" lengths of the V-shaped particles were 22 + 4.5 nm (SD). Gel filtration on Sephacryl S-300 demonstrated that purified C protein had a Stokes' radius of 5.07 nm. Measurements of viscosity gave an intrinsic viscosity of 16.5 cm3/g. These data are consistent with the electron microscopic data and suggest that C protein in heart muscle is asymmetric. The C protein molecule is large enough to extend from the surface of a thick filament to adjacent thin or thick filaments.

Support for a trimeric collar of myosin binding protein C in cardiac and fast skeletal muscle, but not in slow skeletal muscle

“…At physiological ionic strength the sedimentation co-efficient of fsMyBPC increases with rising concentrations [9], consistent with dimerisation and multimerisation of the protein, a phenomenon also reported for cardiac MyBPC [10]. Purified fsMyBPC also takes on a similar V-shaped structure as cMyBPC [10,11].…”

Myosin binding protein C: Structural abnormalities in familial hypertrophic cardiomyopathy

“…It is not clear whether this unusual packing occurs in the full length MyBPC, or if these 10 residues actually form a short linker between domains 4 and 5, as the sequence alignments would predict. Electron micrographs of isolated cardiac MyBPC revealed over half of the protein molecules were V-shaped, with arms of 22±4.5 nm [60], implying a point of flexibility occurs between Motifs 4 and 5 [9]. Electron micrographs of skeletal MyBPC also showed V-shaped molecules [61].…”

“…The first, P873H [60], changes a proline that is highly conserved across species and isoforms of MyBPC and is at the position that defines the N-terminal boundary in FnIII domains. The mutation P873H is unique, being found in only one patient with a mild phenotype.…”

Isolation and characterization of a cDNA clone encoding avian skeletal muscle C-protein: an intracellular member of the immunoglobulin superfamily.

“…A flexible hinge region, which may stabilize homophilic interactions, is located between these sets of domains in N-CAM (53). Given that C-protein contains a similar flexible region located in a pericentric position along the molecule (52,54), it is conceivable that its hinge also occurs at this site and serves a comparable function.…”