Magnesium regulates both the nucleotide binding and the enzyme activity of isolated chloroplast coupling factor 1.

The inactivation and the activation of the ATPase of isolated CF1 as assayed by the hydrolysis of ATP in 10 s depended on prior binding of ADP-Mg and ATP-Mg. The effects of Mg2+ on the nucleotide binding kinetics were studied by monitoring the time courses of UV spectral changes induced by the interaction between CF1 and ADP or ATP using a rapid-scan spectrophotometer equipped with a stopped-flow cell. The apparent rate constant of ADP binding to the two high-affinity sites on CF1 (designated sites B and C in the previous report [Hisabori, T. & Sakurai, H. (1984) Plant Cell Physiol. 25, 483-493]) was drastically increased by prior binding of Mg2+ to CF1, but not ATP. The inhibitory effect of Mg2+ was attributed to a marked increase in kon for the inhibitory ADP binding at the high-affinity sites induced by the previous binding of Mg2+ to the enzyme. The location of site B is suggested to be on the beta subunit based on the difference spectral change induced by binding of the ADP analog 2',3'-O-(2,4,6-trinitrophenyl)ADP to CF1.