Improved Substrate Specificity of Fatty Acid Hydratases Enables Efficient and Selective Linoleic Acid Isolation

Area of Science:

• Biotechnology
• Biocatalysis
• Chemical Engineering

Background:

• Isolating linoleic acid (LA) from oleic acid (OA) is difficult due to similar properties.
• Traditional methods like distillation are energy-intensive and complex.
• Existing fatty acid hydratases (FAHs) lack sufficient specificity for OA.

Purpose of the Study:

• To engineer a fatty acid hydratase (FAH) from *Bifidobacterium breve* (BbMCRA2) for enhanced oleic acid (OA) selectivity.
• To develop an efficient and green method for isolating linoleic acid (LA).

Main Methods:

• Semirational design was used to optimize the substrate tunnel geometry of BbMCRA2.
• Mutants I206V and I202V/I209V were created and tested for OA selectivity.
• Reaction kinetics were optimized for the enzymatic hydration process.

Main Results:

• Engineered mutants showed 4.7- and 5.5-fold increased OA selectivity.
• Linoleic acid (LA) isolation achieved 94.3% purity and 81.7% recovery in a 5 L system.
• The enzymatic process significantly reduced energy consumption and processing time compared to conventional methods.

Conclusions:

• Engineered BbMCRA2 provides a highly selective biocatalyst for OA conversion.
• This enzymatic hydration strategy offers a green, cost-effective, and efficient method for industrial LA purification.
• The improved specificity of hydroxy fatty acid (HFA) production enables superior fatty acid isolation.