Trypsin digestion of canine cardiac myosin: Low molecular weight fragment of myosin with high adenosine triphosphatase activity

Abstract

(1) Trypsin digestion of dog cardiac myosin leads to the formation of two dissimilar types of enzymatically active species based on the elution pattern of Sephadex G-200 columns. (2) When the digestion is performed in 0.6 m KCl the major protein peak is eluted at the exclusion limit of the column. Sodium dodecyl sulfate (SDS)-gel electrophoresis of this peak shows the heterogeneity of the heavy chain component, indicating multiple sites of cleavage by trypsin. (3) When the trypsinization is carried out in 0.15 m KCl in the presence of EDTA and β-mercaptoethanol, the major protein peak (retarded on the Sephadex G-200 column) has a high Ca 2+-ATPase activity. On SDS-gel electrophoresis it shows only two major bands with corresponding molecular weights of 58,000 and 28,000, respectively. The 28,000-molecular-weight band apparently corresponds to cardiac light chain 1 of native myosin. (4) The results suggest that, with trypsinization of myosin in 0.15 m KCl, only a limited number of sites is exposed to trypsin. The fragment isolated under these conditions differs from a papain digestion fragment with respect to its molecular weight and the composition of the heavy chain fraction. On the basis of the molecular weight of the undissociated fragment it seems likely that the fragment retains a heavy meromyosin type (two heads) of configuration.