The Role of Sodium in the Conversion of Pyruvate to Phosphoenolpyruvate in Mesophyll Chloroplasts of C₄ Plants

Abstract

PEP formation from pyruvate was determined in mesophyll chloroplasts mechanically isolated from sodium-deficient and sodium-replete plants of the NADP malic enzyme-type C₄ species, Kochia trichophylla. An extremely sensitive method for estimating PEP was developed which permitted determination of picomole quantities of PEP in small samples taken sequentially from the mesophyll chloroplast suspension concurrently with observations on oxygen evolution. It was shown that PEP formation requires light and depends upon the intactness of the chloroplasts. The rate of formation of PEP from pyruvate increased in the presence of the Hill reagent, oxaloacetate, thus indicating its dependence on non-cyclic photophosphorylation for the supply of ATP required in the conversion of pyruvate to PEP. The optimum inorganic phosphate concentration for PEP formation was approximately 16 mM. The rates of oxygen evolution and PEP formation were equivalent at concentrations of pyruvate up to 20 mM, suggesting tight coupling between electron transport and phosphorylation. In both Kochia trichophylla and the NAD malic enzyme-type, Panicum miliaceum, the rates of PEP formation were greater in the mesophyll chloroplasts from sodium-replete than from sodium-deficient plants. Chloroplasts resuspended in 'sodium-free'media containing less than 30 μM (0.7 ppm) sodium showed reduced rates of PEP formation compared with chloroplasts resuspended in media to which 1.0 mM (23 ppm) sodium had been added. Both media contained 10 mM 'sodium-free' KCI. These results indicate that sodium ions may be required to maintain the functional integrity of the mesophyll chloroplasts and that irreversible damage is sustained when sodium is absent during their isolation.