Two opposite effects of ATP on the chymotryptic cleavages in smooth muscle myosin head. Determination of cleavable points and their characterization.
The locations of ATP-induced structural change in gizzard myosin were examined by analyzing the changes in the chymotryptic fragmentation pattern. From the time course of fragmentation, and by fractionation of the produced fragments and detection of the masked amino terminal fragment, the original positions of the six different fragments in the parent myosin molecule were assigned. A reconstituted model of myosin based on the above assignment showed the presence of three cleavable sites in the heavy chain of myosin. ATP accelerated the cleavage at site 1, 5 K daltons from the masked amino terminus, while it inhibited cleavage at site 2, 71 K daltons from the N terminus. These opposite effects of ATP on sites 1 and 2 were remarkable, but ATP had no significant effect on cleavage at site 3, 100 K daltons from the carboxyl terminus. These results indicate that two distant regions in the myosin head, 66 K daltons apart in the primary sequence, were exposed or buried with the progress of the ATPase reaction. In addition, prolonged chymotryptic digestion failed to produce any subfragment-1, irrespective of the presence or absence of divalent cations in the digestion medium, but produced a protease-resistant and relatively long (100 K daltons) light meromyosin. This suggests a distinctive feature of the neck and hinge regions in gizzard myosin.
D000255:Adenosine Triphosphate / Q000494:pharmacology*; D000818:Animals; D002645:Chickens; D002918:Chymotrypsin / Q000378:metabolism*; D005895:Gizzard, Avian / Q000378:metabolism; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D009130:Muscle, Smooth / Q000378:metabolism*; D015879:Myosin Subfragments / Q000378:metabolism; D009218:Myosins / Q000378:metabolism*; D010446:Peptide Fragments / Q000032:analysis; D011487:Protein Conformation
D046911:Macromolecular Substances; D015879:Myosin Subfragments; D010446:Peptide Fragments; D000255:Adenosine Triphosphate; D002918:Chymotrypsin; D009218:Myosins