PURIFICATION AND CHARACTERIZATION OF NADP(-LINKED ISOCITRATE DEHYDROGENASE FROM SCOTS PINE - EVIDENCE FOR DIFFERENT PHYSIOLOGICAL ROLES OF THE ENZYME IN PRIMARY DEVELOPMENT())

NADP(+)-isocitrate dehydrogenase (NADP(+)-IDH; EC 1.1.1.42) is involve d in the supply of 2-oxoglutarate for ammonia assimilation and glutama te synthesis in higher plants through the glutamine synthetase/glutama te synthase (GS/GOGAT) cycle. Only one NADP(+)-IDH form of cytosolic l ocalization was detected in green cotyledons of pine (Pinus spp.) seed lings. The pine enzyme was purified and exhibited molecular and kineti c properties similar to those described for NADP(+)-IDH from angiosper m, with a higher catalytic efficiency (10(5) M-1 s(-1)) than the deduc ed efficiencies for GS and GOGAT in higher plants. A polyclonal antise rum was raised against pine NADP(+)-IDH and used to assess protein exp ression in the seedlings. Steady-state levels of NADP(+)-IDH were coor dinated with CS during seed germination and were associated with GS/GO GAT enzymes during chloroplast biogenesis, suggesting that NADP(+)-IDH is involved in the provision of carbon skeletons for the synthesis of nitrogen-containing molecules. However, a noncoordinated pattern of N ADP(+)-IDH and GS/GOGAT was observed in advanced stages of cotyledon d evelopment and in the hypocotyl. A detailed analysis in hypocotyl sect ions revealed that NADP(+)-IDH abundance was inversely correlated with the presence of CS, GOGAT, and ribulose-1,5-bisphosphate carboxylase/ oxygenase but was associated with the differentiation of the organ. Th ese results cannot be explained by the accepted role of the enzyme in nitrogen assimilation and strongly suggest that NADP(+)-IDH may have o ther, as-yet-unknown, biological functions.