A study of troponin, a myofibrillar protein from rabbit skeletal muscle.

Precipitation at pH 5.6 is found to improve the rabbit muscle troponin preparation. Further purification of troponin is achieved by treating troponin preparations with 0.1 M dithiothreitol, followed by the removal of contaminating tropomyosin by gel filtration. This is possible because through these purification procedures the molecular weight of tropomyosin as estimated by gel filtration remains fairly constant (approximately 130,000), while that of troponin preparations is gradually reduced from approximately 150,000 via 90,000 to 44,000. The sulfhydryl and tryptophan contents of troponin thus purified are 5.4 and 4.0 moles/IO6 g protein, respectively. Evidence is presented which indicates that tropomyosin and troponin form a complex which behaves exactly like the relaxing protein, and that the relaxing protein preparation is actually dissociated by 5 M urea into troponin and tropomyosin. The weight ratio of troponin to tropomyosin in the relaxing protein as estimated from the density of Amido black staining on gel electrophoresis is approximately 1:2.5. Precipitation of troponin by calcium and disaggregation of troponin by dithiothreitol support the view that, subsequent to calcium binding, troponin undergoes conformational change which is in turn transmitted through tropomyosin to the actin-myosin interaction.