INACTIVATION OF ECTO-ATPASE ACTIVITY OF RAT-BRAIN SYNAPTOSOMES

The ecto-ATPase activity of synaptosomes plasma membrane decays expone ntially as a function of time from 0.35 +/- 0.05 to 0.08 +/- 0.02 mu m ol ATP hydrolyzed per min per mg synaptosome protein. The first-order rate constant of inactivation is dependent on the Mg-ATP concentration varying from 0.042 +/- 0.001 min(-1) with 30 mu M ATP up to 0.216 +/- 0.003 min(-1) with 2 mM ATP. The non-hydrolyzable ATP analogue, beta- y-methyleneadenosine 5'-triphosphate, did not produce inactivation of the ecto-ATPase activity. Thus, the inactivation of the ecto-ATPase ac tivity requires hydrolysis of ATP. Product inhibition can he excluded because ADP, AMP, adenosine and inorganic phosphate up to 1 mM had no effect on the inactivation of the ecto-ATPase. Concanavalin A partiall y protected against the ATP-dependent inactivation. The ecto-ATPase in activation produced by Mg-ATP is partially reverted by centrifugation, removal of the supernatant and resuspension of synaptosomes in a fres h medium. This partial reversion occurs in parallel to the release to the supernatant of phosphorylated protein(s) of 90-95 kDa. Alkaline ph osphatase treatment fully reverts the ecto-ATPase inactivation, We con clude that the ATP-induced inactivation is mediated, at least partiall y, by phosphorylation of membrane proteins.