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

Bhan, A.K.; Malhotra, A.

Archives of Biochemistry and Biophysics 174(1): 27-35

1976


ISSN/ISBN: 0003-9861
PMID: 132895
DOI: 10.1016/0003-9861(76)90320-9
Accession: 068521503

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Abstract
Trypsin digestion of dog cardiac myosin leads to the formation of 2 dissimilar types of enzymatically active species based on the elution pattern of Sephadex G-200 columns. When the digestion was performed in 0.6 M KCl the major protein peak was eluted at the exclusion limit of the column. Sodium dodecyl sulfate (SDS)-gel electrophoresis of this peak showed the heterogeneity of the heavy chain component, indicating multiple sites of cleavage by trypsin. When the trypsinization was carried out in 0.15 M KCl in the presence of EDTA and .beta.-mercaptoethanol, the major protein peak (retarded on the Sephadex G-200 column) had a high Ca2+-ATPase activity. On SDS-gel electrophoresis it showed only 2 major bands with corresponding MW of 58,000 and 28,000, respectively. The 28,000-MW band apparently corresponds to cardiac light chain 1 of native myosin. With trypsinization of myosin in 0.15 M KCl, only a limited number of sites were 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 (2 heads) of configuration.