High-level expression and characterization of solvent-tolerant lipase

Area of Science:

• Biotechnology
• Enzyme Engineering
• Industrial Microbiology

Background:

• Lipases are crucial enzymes for various industrial applications, including biofuel production.
• Optimizing lipase expression and activity is key to improving biocatalytic efficiency.
• Proteus sp. SW1 lipase offers potential for industrial use but requires enhancement.

Purpose of the Study:

• To optimize the lipase gene sequence from Proteus sp. SW1 for enhanced expression.
• To characterize the biochemical properties and stability of the recombinant lipase.
• To evaluate the recombinant lipase's efficacy in biodiesel synthesis.

Main Methods:

• Codon optimization of the lipase gene sequence.
• Heterologous expression of the lipase in Pichia pastoris GS115.
• Purification and characterization of the recombinant lipase, including activity assays and stability tests in solvents and under varying conditions.
• Assessment of the lipase's performance in a solvent-free biodiesel synthesis system.

Main Results:

• Achieved a maximum enzyme yield of 387 mg/L in shake-flask cultures.
• The purified recombinant lipase showed a specific activity of 130 U/mg towards p-nitrophenyl Laurate.
• Optimal activity at pH 8.0 and 40°C, with high stability and activation in water-miscible organic solvents.
• Demonstrated 87% conversion rate in solvent-free biodiesel synthesis from palm oil with 80% methanol/ethanol.

Conclusions:

• Codon optimization and Pichia pastoris expression significantly enhanced lipase yield and activity.
• The recombinant lipase exhibits remarkable stability and activity in organic solvents and under high substrate concentrations.
• This engineered lipase is a promising biocatalyst for efficient and scalable biodiesel production.