ATPASE AND PHOSPHATASE-ACTIVITIES ARE DIFFERENTIALLY INHIBITED BY PHOTOOXIDATION OF THE SARCOPLASMIC-RETICULUM CA2-ATPASE( C)

We have already described that photo-oxidation of the sarcoplasmic ret iculum Ca2+-ATPase with the halogenated dye erythrosin B produces inhi bition of the ATPase activity (J.A. Mignaco et al., Biochemistry 35 (1 996) 3886-3891), We now show that the Ca2+-dependent and Ca2+-independ ent p-nitrophenylphosphatase activities are also inhibited by this tre atment. Modification of rapidly (< 10 min) oxidized residue(s) is resp onsible: for the major loss of ATPase activity, whereas photo-inhibiti on of the phosphatase activities occurs more slowly (t(1/2) 20-30 min) . Here we have focused on photo-inhibition of the Ca2+-independent pNP Pase activity. and the counteracting effects of ATP and FITC. Followin g photo-oxidation, the Ca2+-independent pNPPase activity decreases mon otonically, ATP partially protects against the inactivation of the pNP Pase, whereas labeling the enzyme with FITC does not. However, the pro tective effect of ATP is completely abolished by the attached FITC. Th ese data are interpreted in terms of two different sites that art: sus ceptible to photo-oxidation and are involved in different events relat ed to substrate hydrolysis. (C) 1997 Elsevier Science B.V.