Phosphoenolpyruvate carboxykinase activity in grape berries

Area of Science:

• Biochemistry
• Plant Physiology
• Enzymology

Background:

• Grape berry development involves complex metabolic pathways.
• Enzymes like PEP carboxykinase and PEP carboxylase play roles in carbon fixation and metabolism.
• Understanding these enzymes is crucial for grape quality and yield.

Purpose of the Study:

• To characterize the activity and kinetic properties of PEP carboxykinase in Pinot noir grape berries.
• To investigate the changes in PEP carboxykinase and PEP carboxylase activity during grape fruit development.
• To compare the relative activities of these two key carboxylating enzymes.

Main Methods:

• Extraction and partial purification of enzymes from grape berry crude extracts.
• Enzyme assays to determine optimal conditions (pH, temperature, cofactors) and kinetic parameters (Km).
• Weekly measurements of enzyme activity throughout the entire fruit development period.

Main Results:

• Phosphoenolpyruvate (PEP) carboxykinase activity was detected in Pinot noir grape berries, requiring ATP, Mn(2+)/Mg(2+), pH 6.6, and 40°C for maximal activity.
• The enzyme exhibited optimal oxaloacetic acid concentration between 5-10 mM, with a Km for HCO(3)(-) of 26.8 mM.
• PEP carboxykinase activity peaked 4 weeks post-flowering and remained stable, while PEP carboxylase activity was 2-4 times higher and decreased sharply at ripening onset.

Conclusions:

• PEP carboxykinase is present and active in developing grape berries, with specific optimal conditions.
• Enzyme activity profiles suggest distinct roles during fruit development, with PEP carboxylase dominating early stages.
• These findings contribute to understanding carbon metabolism during grape ripening.