A novel method of extraction of TnC from skeletal muscle myofibrils

Incubation of mechanically skinned barnacle myofibrillar bundles in 10 mM o rthovanadate (pH 6.6) results in the loss of Ca2+-dependent force generatio n, which reduces to 0.98+/-0.006% (mean +/- SEM, n=25) of control levels. A nalysis of myofibrillar bundles by gel electrophoresis showed that tension loss is primarily due to the extraction of troponin C (TnC) (65.4+/-5.04% m ean +/-SEM, n=5). This is a novel finding, since treating cardiac fibres wi th orthovanadate results in the removal of both TnC and troponin I (TnI) (2 8). Ca2+ dependence was restored to the myofibrillar bundles following reco nstitution with either native isoform of barnacle TnC (BTnC(1): 78.72+/-12. 8%, n=9, BTnC(2): 82.73+/-20.3%, n=3). The reversible loss of Ca2+-dependen t tension generation following the removal and replacement of TnC indicates that the regulation of contraction in the barnacle is controlled by thin-f ilament regulatory proteins.