Mechanism-Based Post-Translational Modification and Inactivation in Terpene Synthases

Area of Science:

• Biochemistry
• Enzymology
• Natural Product Biosynthesis

Background:

• Terpenes are vital natural products with diverse biological roles.
• Terpene synthases (TPSs) catalyze complex reactions involving reactive carbocationic intermediates.
• The active sites of TPSs are shaped to control terpene biosynthesis.

Purpose of the Study:

• To investigate the mechanism of enzyme inactivation in terpene biosynthesis.
• To identify the role of carbocationic intermediates in TPS activity.
• To explore evolutionary pressures on terpene synthase evolution.

Main Methods:

• Proteomic analysis to identify modified residues.
• X-ray crystallography to visualize enzyme-substrate interactions.
• Mutagenesis studies to assess the impact of active site modifications.

Main Results:

• Conserved active site residues capture cationic intermediates, leading to self-alkylation and inactivation.
• Inactivation levels correlate with active site mutations, substrate variations, and increased temperature.
• Multi-product TPSs exhibit lower self-alkylation rates compared to specific TPSs.

Conclusions:

• Mechanism-based alkylation is a significant factor in terpene synthase evolution.
• Understanding self-alkylation provides insights into the regulation of terpene biosynthesis.
• This study highlights an overlooked evolutionary pressure on enzymes utilizing carbocationic intermediates.