Synthesis and turnover of ribulose biphosphate carboxylase and of its subunits during the cell cycle of Chlamydomonas reinhardtii.
The chloroplast enzyme ribulose-1,5-bisphosphate (Ru-1,5-P2) carboxylase (EC 4.1 1.39) is made up ot two nonidentical subunits, one synthesized in the chloroplast and the other outside. Both of these subunits of the assembled enzyme are synthesized in a stepwise manner during the synchronous cell cycle of the green alga Chlamydomonas reinhardtii. The activity of this enzyme increases in the light and this increase is due to de novo protein synthesis as shown by the measurement of the amount of protein and by the pulse incorporation of radioactive arginine in the 18S enzyme peak in linear sucrose density gradients. During the dark phase of the cell cycle, there is little change in the enzymatic activity as well as in the amount of this enzyme. Pulse-labeling studies using radioactive arginine indicated that there is a slow but detectable rate of synthesis of the carboxylase and of its subunits in the dark. Ru-1,5-P2 carboxylase, prelabeled with radioactive arginine throughout the entire light period, shows a similarly slow rate of degradation in the following dark period. This slow turnover of the enzyme in the dark accounts for the steady levels of carboxylase protein and of enzymatic activity during this period. A wide variety of inhibitors of protein synthesis by 70S and 80S ribosomes abolished the incorporation of [3H]arginine into total Ru-1,5-P2 carboxylase during short-term incubation. These results suggest a tight-coordinated control of the biosynthesis of the small and large subunits of the enzyme. This stringent control is further substantiated by the finding that both subunits are synthesized in sychrony with each other, that the ratio of radioactivity of the small to the large subunit remains constant throughout the entire light-dark cycle, and that the rates of synthesis and of degradation of both subunits are similar to that of the assembled enzyme.
D002262:Carboxy-Lyases / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D002455:Cell Division; D002499:Centrifugation, Density Gradient; D002696:Chlamydomonas / Q000201:enzymology*; D002736:Chloroplasts / Q000201:enzymology; D004591:Electrophoresis, Polyacrylamide Gel; D046911:Macromolecular Substances; D010428:Pentosephosphates; D012266:Ribose; D013997:Time Factors
D046911:Macromolecular Substances; D010428:Pentosephosphates; D012266:Ribose; D002262:Carboxy-Lyases