Phosphoenolpyruvate carboxylase during grape berry development: protein level, enzyme activity and regulation

The protein level and regulation of phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31, involved in malic acid synthesis) was studied during the fruit development of two grape (Vitis vinifera L.) varieties, 'Cabernet Sauvigno n' and 'Gora Chirine', with berries of normal and low organic acid content, respectively. The protein level and in vitro activity were higher in the l ow-acid variety than in the normal-acid variety for most stages. In vivo PE PC activity, measured using (CO2)-C-14 labelling, was significantly higher in the low-acid variety than in the normal-acid variety about 1 week before and 1 week after veraison (the day which corresponds to the onset of ripen ing). However, partitioning into malate was the same for both varieties. An tibodies raised against the N-terminal part of Sorghum PEPC recognised the grape berry PEPC, indicating the presence of the consensus phosphorylation site involved in PEPC regulation. PEPC phosphorylation status was estimated by studying sensitivity to pH and malate. Grape berry PEPC appeared more s ensitive to low pH and malate during ripening (IC50 malate, 0.2-0.7 mM) com pared to during the earlier stages of development (IC50 malate, 1.2-2 mM) f or both varieties. Therefore, in the normal-acid variety, PEPC seems to par ticipate in controlling malic acid accumulation but does not seem to contro l the differences in malic acid concentration observed between the two vari eties.