Light modulation of maize leaf phosphoenolpyruvate carboxylase

Area of Science:

• Biochemistry
• Plant Physiology
• Enzymology

Background:

• Phosphoenolpyruvate carboxylase (PEPC) plays a crucial role in carbon fixation in plants like maize (Zea mays L.).
• Previous studies indicated differences in malate sensitivity between PEPC extracted from light and dark-harvested maize leaves.

Purpose of the Study:

• To investigate the structural and functional basis for differential malate sensitivity in maize leaf PEPC.
• To determine if light/dark conditions induce stable structural changes in PEPC.

Main Methods:

• Partial purification of PEPC from maize leaves harvested in light and dark using ammonium sulfate fractionation and gel filtration.
• Monitoring of PEPC activity, malate sensitivity, and protein levels (immunochemically) throughout purification.
• Analysis of enzyme behavior during gel filtration (Sephacryl S-300) and characterization of purified polypeptides via immunoblotting.

Main Results:

• PEPC from dark-harvested leaves exhibited higher malate sensitivity than that from light-harvested leaves.
• Purification in the presence of malate stabilized these differential characteristics.
• Gel filtration revealed a single high molecular weight peak for PEPC, indicating a tetrameric structure of a single 100,000 Da polypeptide, with slight differences in elution volume between light and dark enzymes.
• Specific activity and malate sensitivity varied within the elution peak for the dark enzyme, correlating with elution volume.
• Extraction and purification without malate abolished the observed differences between light and dark enzymes.

Conclusions:

• Maize leaf PEPC exists in situ as a stable tetramer composed of a single polypeptide type.
• Subtle conformational changes, influenced by extraction conditions (presence/absence of malate) and potentially light/dark harvesting, affect PEPC's specific activity and malate sensitivity.