Engineering isoprenoids production in metabolically versatile microbial host Pseudomonas putida.

Area of Science:

• Synthetic biology and metabolic engineering for sustainable bioproduction.
• Microbial biotechnology for renewable energy and chemical manufacturing.

Background:

• Increasing demand for microbial bioproduction to replace petrochemicals.
• Pseudomonas putida KT2440 is a promising industrial host due to its metabolic versatility and robustness.
• Need for efficient conversion of lignocellulosic carbon into biofuels and bioproducts.

Purpose of the Study:

• Engineer P. putida KT2440 for the production of isoprenoids, including biofuels isoprenol and epi-isozizaene.
• Optimize isoprenol production and identify strategies to prevent its native consumption by P. putida.
• Demonstrate the potential of P. putida as a microbial chassis for sustainable isoprenoid production from lignocellulosic biomass.

Main Methods:

• Engineering a heterologous mevalonate (MVA) pathway for isoprenoid synthesis in P. putida.
• Comparing three different isoprenoid pathways to maximize isoprenol production.
• Investigating methods to prevent isoprenol self-consumption, including L-glutamate supplementation.
• Utilizing metabolomics to analyze the metabolic state during isoprenol degradation.

Main Results:

• Achieved 104 mg/L of isoprenol production in batch flask experiments.
• Identified L-glutamate supplementation as an effective strategy to prevent isoprenol consumption.
• Metabolomics revealed energy limitation and redox imbalance during isoprenol degradation.
• Demonstrated P. putida's ability to produce isoprenol using aromatic substrates like p-coumarate.

Conclusions:

• Engineered P. putida is a viable platform for producing isoprenoids and biofuels from lignocellulosic materials.
• Preventing isoprenol self-consumption is crucial for efficient bioproduction.
• P. putida's metabolic flexibility supports sustainable production of valuable compounds from renewable resources.