Production of nitrite ions from trinitrophenyl myosin and from trinitrophenyl subfragment-1

When trinitrophenyl (TNP) myosin of either chicken breast muscle or porcine cardiac muscle was left to stand in an alkaline medium at 20 degrees C for several hours, nitrite ions were found to be gradually produced. The nitrite production from myosin trinitrophenylated in the presence of PPi occurred at the same rate and to the same extent as that from myosin trinitrophenylated in the absence of PPi. The nitrite production was significantly reduced when thiols of myosin were modified with 2-nitro-5-thiocyanobenzoate. Four different preparations of TNP subfragment-1, S1(Aa), S1(Ab), S1(Ba), and S1(Bb), were obtained from chymotryptic digest of chicken breast myosin trinitrophenylated in the absence of PPi. When these preparations of TNP-S1 were left to stand at alkaline pH, a significant amount of nitrite was produced from S1(Ab) and S1(Bb), but very little from S1(Aa) and S1(Ba). In our previous report (J. Biochem. 97, 965-968, 1985), S1(Aa) and S1(Ba) were suggested to correspond to "non-burst" heads of myosin, and S1(Ab) and S1(Bb) to "burst" heads of the myosin molecule (Inoue et al. (1980) Adv. Biophys. 13, 1-194). Therefore, the present findings described above strongly suggest that the nitrite production involves some interaction of TNP groups with thiols, and that it occurs at the "burst" heads.