Physicochemical studies on the creatine kinase M-line protein and its interaction with myosin and myosin fragments.
The M-line protein which is identical to the muscle form of creatine kinase was purified from rabbit skeletal muscle using ion exchange chromatography. Gel electrophoresis in the presence and absence of sodium dodecyl sulfate revealed the protein to be homogeneous. Sodium dodecyl sulfate gel electrophoresis gave 44 000 +/- 2000 as the minimum molecular weight while low speed sedimentation equilibrium experiments yielded a molecular weight of 84 000 +/- 4000, suggesting that the parent molecule is a dimer. Circular dichroism spectra revealed the presence of two negative dichroic bands located at 218 and 208 nm suggesting the presence of some beta-structure. Ellipticity values at these two wavelengths were -8000 +/- 400 and -9000 +/- 400 deg-cm2-dmol-1. Circular dichroism measurements indicated the protein to interact with myosin, heavy meromyosin and heavy meromyosin subfragment 1 (S1). The Ca2+-activated ATPase activities of myosin, heavy meromyosin and subfragment 1 were inhibited by the addition of M-line protein. When the protein was mixed with subfragment 1 in a 1:1 mole ratio in 0.15 M KC1, 50 mM Tris pH 8, low speed sedimentation equilibrium studies gave a molecular weight of 205 000 +/- 10 000 for the complex, indicative of an interaction of the two components. Both circular dichroism and sedimentation equilibrium studies indicated no interaction of M-line protein with light meromyosin.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001665:Binding Sites; D002942:Circular Dichroism; D003402:Creatine Kinase; D007700:Kinetics; D008970:Molecular Weight; D009132:Muscles / Q000201:enzymology; D015879:Myosin Subfragments; D009218:Myosins; D010446:Peptide Fragments; D011485:Protein Binding; D011487:Protein Conformation; D011817:Rabbits; D013056:Spectrophotometry, Ultraviolet
D000596:Amino Acids; D015879:Myosin Subfragments; D010446:Peptide Fragments; D003402:Creatine Kinase; D009218:Myosins