Involvement of the 50-kDa peptide of myosin heads in the ATPase activity revealed by fluorescent modification with 4-fluoro-7-nitrobenz-2-oxa-1,3-diazole.
The fluorescent reagent 4-fluoro-7-nitrobenz-2-oxa-1,3-diazole (NBD-F) reacted specifically with 1.9 lysyl residues/mol of the myosin subfragment-1 (S-1) ATPase. When 1.9 lysyl residues were modified, the K+- and Ca2+-ATPase activities were almost completely inhibited, whereas the Mg2+-ATPase activity was increased to 180% of original activity. The actin-activated Mg2+-ATPase activity was decreased to 30% of original activity by this modification. However, affinity of S-1 for actin in the presence of ATP was unchanged. The NBD fluorescence of the modified S-1 was quenched on addition of ATP, suggesting that ATP induced conformational changes around the NBD groups attached to S-1. Tryptic digestion of the modified S-1 revealed that the NBD groups are attached mainly to the 50-kDa peptide of S-1, more precisely the 45-kDa peptide. These results confirm the recent reports that the 50-kDa peptide of S-1 is involved in the myosin ATPase reaction (Körner, M., Thiem, N. V., Cardinaud, R., and Lacombe, G. (1983) Biochemistry 22, 5843-5847; Hiratsuka, T. (1986) Biochemistry 25, in press).
D009327:4-Chloro-7-nitrobenzofurazan / Q000031:analogs & derivatives; D000251:Adenosine Triphosphatases / Q000032:analysis*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D005453:Fluorescence; D007700:Kinetics; D008239:Lysine / Q000032:analysis; D015879:Myosin Subfragments; D009218:Myosins / Q000032:analysis*; D010069:Oxadiazoles; D010446:Peptide Fragments / Q000032:analysis*; D011487:Protein Conformation; D011817:Rabbits
D015879:Myosin Subfragments; D010069:Oxadiazoles; D010446:Peptide Fragments; C032930:7-fluoro-4-nitrobenzo-2-oxa-1,3-diazole; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D009218:Myosins; D009327:4-Chloro-7-nitrobenzofurazan; D008239:Lysine