Mechanism for oxygen exchange in the chloroplast photophosphorylation system.

The oxygen exchange that occurs between water and the gamma-PO3 of ATP in light-activated chloroplast lamellae was found to proceed with close to full equilibration of the oxygens before ATP returned to the medium. This is in contrast to the entry of approximately one water oxygen when ATP is synthesized from ADP and P1 in the same system. In the latter case, the limitation is kinetic, however, not steric, as shown by the presence of some molecules containing more than one water-derived oxygen in the gamma-PO3. The different extents of exchange can be explained by a relatively faster rate of dissociation of ATP from the chloroplast coupling factor during synthesis from ADP and P1 relative to its dissociation in the absence of net phosphorylation. To determine the mechanism of gamma-PO3:H2O exchange, its rate was compared with the rate of reversible cleavage of ATP as detected by betagamma bridge to beta nonbridge 18O scrambling in [Pbeta-18O-Pgamma]ATP (Midelfort, C. F., and Rose, I. A. (1976) J. Biol. Chem. 251, 5881-5887). The scrambling reaction, which depends on cleavage of the PbetaO--Pgamma bond, was found to occur in nearly the same fraction of ATP molecules that experienced gamma-PO3:H2O exchange in the same incubation, suggesting that the latter is due to multiple cycles of reversible ATP hydrolysis on the chloroplast coupling factor, i.e. [ATP-H2O in equilibrium ADP-Pi].