REGULATION OF RIBULOSE 1,5-BISPHOSPHATE CARBOXYLASE OXYGENASE ACTIVATION BY INORGANIC-PHOSPHATE THROUGH STIMULATING THE BINDING OF THE ACTIVATOR CO2 TO THE ACTIVATION SITES

The mechanism of the regulation of the activation of ribulose 1,5-bisphosphate carboxylase/oxygenase (RuBisCO) by inorganic phosphate (P-i) in the presence of limiting concentrations of CO2 was explored. The activation state of RuBisCO increased sigmoidally following a biphasic kinetics against the concentration of P-i in the activation mixture with an intermediary plateau at 2 to 3 mM P-i when the enzyme was activated for 30 min. The intermediary plateau could not be seen when the preincubation time was 10 min and the activation was completed at 10 mM P-i. RuBisCO from Euglena also showed a quite similar activation kinetics. The activation was not due to the contaminating CO2 included in the stock P-i solution or in the activation buffer containing the enzyme. The experiments with 2-carboxyarabinitol 1,5-bisphosphate showed that the P-i-stimulated activation was due to the promotion of binding of the activator CO2 to the activation sites. It was also found that P-i increased the affinity of RuBisCO for the activator CO2 5.4-fold accompanied by a decrease of the half-saturating concentration of CO2 to 1.6 mu M at 20 mM MgCl2. Physiological significance of the effects of P-i on the activation of RuBisCO is discussed.