PUB - Publikationen an der Universität Bielefeld

Fast protein liquid chromatography on Superose 6 of partially purified FBPase II from Chlorella reveals a 1350 kDa-form at pH 6.0 and a 67 kDa-form at pH 8.5. Treatment of the large enzyme form with 5mM concentrations of Mg2+, F1,6P(2), DTT or ATP leads to dissociation into smaller ones of 215-470 kDa. Aggregation/dissoziation is a reversible process, as has been shown for the effect of F1,6P(2) and of pH, by rechromatography. The change in mole mass results in alterations of the activity and df the kinetic properties of the enzyme forms, obtained. Dissociation results in a 4-6 fold increase in activity, as can be shown for F1,6P(2)-treated samples. Halfsaturation constants, as well as the degree of cooperativity of the 67- and the 1350-kDa form, are different for substrate affinity, activation by Mg2+ and DTT, and for inhibition by ATP. Both enzyme forms hydrolyse fructose 1,6 bisphosphate and seduheptulose 1,7 bisphosphate better than other phosphorylated compounds. The ratio of F1,6P(2)- to SDP-cleavage is 100:58 for the small enzyme form and 100:84 for the large one. Activation of FBPase II in the light and inactivation in the dark is discussed on the basis of different oligomeric forms of the enzyme, generated by changes in the concentration of intermediates and effecters in the chloroplast stroma, leading to dissociation or aggregation. The conclusion is drawn that oligomerization of key enzymes, resulting in enzyme forms with different activities and different kinetic properties, might provide an effective mechanism for enzyme regulation in vivo.