Determination of exponential rate constants for calpain-mediated degradation of myofibrillar complexed proteins

The purpose of this study was to test the hypothesis that an orderly sequence or time course of calpain action on purified myofibrillar/cytoskeletal complexes from cardiac and fast muscle tissue exists. To test the response of individual substrate proteins to degradation (i.e. susceptibility), exogenous calpain (1.5U/ml) was incubated with 40 ug of highly purified rat myofibrillar complexes from cardiac and gastrocnemius muscles (0 to 60 min), in vitro. Apparent molecular weights (SDS-PAGE) were used to compare and identify individual myofibril proteins. Myofibrillar yields for cardiac (n=9) and fast skeletal (n=9) muscles were 62.93 ± 6.58 and 98.42 ± 9.36 mg/g (p<0.05). Following 30 minutes of calpain treatment, the amount of cardiac desmin, C-protein, a-actinin and troponin-T remaining in complex were 0%, 19%, 23% and 68%, compared to controls (p<0.05). For fast skeletal muscle, the remaining desmin, C-protein, a-actinin and tropomyosin after 30 minutes of calpain digestion was 0%, 38%, 51% and 17%, respectively, compared to control (p<0.05). The estimated exponential rate constants (k) for degradation/loss for each protein (in both tissues) had the following order: desmin>C-protein>a -actinin (p<0.05). The results of this study support the hypothesis that there are selective degradation rates and an ordered sequence of degradation for myofibrillar complexed proteins. Factors contributing to the heterogeneity of k values for myofibril proteins, and therefore their susceptibility to calpain degradation, may be their spatial arrangement (peripheral -> central) within the myofibril and/or their primary amino acid residue composition.Education, Faculty ofKinesiology, School ofGraduat