OXIDATIVE MODIFICATION AND BREAKDOWN OF RIBULOSE-1,5-BISPHOSPHATE CARBOXYLASE OXYGENASE INDUCED IN EUGLENA-GRACILIS BY NITROGEN STARVATION

When photoheterotrophic Euglena gracilis Z Pringsheim was subjected to nitrogen (N)-deprivation, the abundant photosynthetic enzyme ribulose -1,5-bisphosphate carboxylase/oxygenase (Rubisco, EC 4.1.1.39) was rap idly and selectively degraded. The breakdown began after a 4-h lag per iod and continued for a further 8 h at a steady rate. After 12 h of st arvation, when the amount of Rubisco was reduced to 40%, the proteolys is of this enzyme slowed down while degradation of other proteins star ted at a similar pace. This resulted in a decline of culture growth, c hloroplast disassembly - as witnessed by chlorophyll (Chl) loss and ce ll bleaching. Experiments with spectinomycin, an inhibitor of chloropl astic translation, indicated that there was an absolute increase in th e rate of Rubisco degradation in the N-deprived culture as compared wi th control conditions, where no significant carboxylase breakdown was detected. Oxidative aggregation of Rubisco (as detected by non-reducti ve electrophoresis) and association of the enzyme to membranes increas ed with time of N-starvation. Fluorescent labeling of oxidized cystein e (Cys) residues with monobromobimane indicated a progressive oxidatio n of Cys throughout the first hours of N-deprivation. It is concluded that Rubisco acts as an N store in Euglena, being first oxidized, and then degraded, during N-starvation. The mobilization of Rubisco allows sustained cell growth and division, at almost the same rate as the co ntrol (non-starved) culture, during 12 h of N-deprivation. Afterwards, breakdown is extended to other photosynthetic structures and the whol e chloroplast is dismantled while cell growth is greatly reduced.