CHARACTERIZATION OF PHOSPHOENOLPYRUVATE CARBOXYKINASE FROM PANICUM-MAXIMUM

Phosphoenolpyruvate carboxykinase, EC 4.1.1.32 (PEPCK), was purified 43-fold from the grass P. maximum. Km were determined for the exchange reaction, the carboxylation reaction, and the decarboxylation reaction. The Km values for oxaloacetate and ATP in the decarboxylation reaction were lower than the Km values for the substrates used in the exchange reaction and in the carboxylation reaction. Phosphoenolpyruvate carboxylase was not detectable in the purified PEPCK preparation. Studies on the nucleotide specificity of the oxaloacetate decarboxylation reaction indicate that ATP serves as the best nucleotide for this reaction and that ADP is about 60% as effective as ATP. The pH optimum for decarboxylase activity is near 6.8. The decarboxylation reaction has a divalent cation requirement with both Mn2+ and Mg2+ needed for full activity. Temperature curves of the 3 PEPCK reactions indicate optimum activities between 38-45.degree. C. There is a pronounced drop in the decarboxylation and carboxylation activities as the temperature is decreased from these optima. Below 30.degree. C the energy of activation was 8.2 kcal/mol for the decarboxylation reaction. Under physiological conditions PEPCK apparently catalyzes the decarboxylation of oxaloacetate in the bundle sheath cells of P. maximum leaves during C4 dicarboxylic acid photosynthesis.