Enzyme regulation in C4 photosynthesis. Purification and properties of thioredoxin-linked fructose bisphosphatase and sedoheptulose bisphosphatase from corn leaves.

Homogeneous preparations of thioredoxin-linked fructose-l,6-bisphosphatase and sedoheptulose-l,7-bisphosphatase were prepared from leaves of corn (Zea mays), a classical C4 plant. Fructose-1,6-bisphosphatase had a molecular weight of 184,000 and consisted of four apparently identical subunits of M, = 46,000. Sedoheptulose-1,7-bisphosphatase, by contrast, was composed of two identical subunits of M, = 38,000 (total M, = 76,000). Antibody experiments revealed that the two enzymes were immunologically distinct. The fructose-1,6-bisphosphatase and sedoheptulosel,7-bisphosphatase were substrate-specific and required M&’ for activity. Both enzymes were also activated specifically by thioredoxin f , which could be reduced either photochemically by the ferredoxin/thioredoxin system or chemically by dithiothreitol. The substrates, fructose l,(r-bisphosphate and sedoheptulose l,7-bisphosphate, acted synergistically with reduced thioredoxin f in the activation of fructose-1,6bisphosphatase and sedoheptulose-l,7-bisphosphatase, respectively. Activation by reduced thioredoxin f effected a shift of the pH optimum for fructose-1,6-bisphosphatase (pH 8.2 + pH 7.7), but not for sedoheptulose-l,7-bisphosphatase. Sedoheptulose-1,7-bisphosphatase, but not fructose-1,6-bisphosphatase, showed an absolute requirement for a dithiol, Le., reduced thioredoxin fo r dithiothreitol. Selected oxidants, including dehydroascorbate and oxidized glutathione, deactivated both enzymes following activation by reduced thioredoxin f. Both enzymes showed hysteretic behavior in activation as well as deactivation. The results demonstrate that fructose-1,6-bisphosphatase and sedoheptulose-l,7-bisphosphatase from corn leaves are distinct enzymes, which, like their spinach counterparts, share in common the capacity for photoactivation by the ferredoxin/thioredoxin system. Activation of each of the enzymes by reduced thioredoxin fis enhanced by the presence of the appropriate sugar bisphosphate substrate. The results support the earlier expressed view that the ferredoxin/thioredoxin system functions jointly with effector metabolites in light-actuated enzyme regulation during photosynthesis.