A Bi-Enzymatic Cascade Pathway towards Optically Pure FAHFAs*

Area of Science:

• Biochemistry
• Enzymology
• Lipid Science

Background:

• Endogenous mammalian lipids, fatty acid esters of hydroxy fatty acids (FAHFAs), exhibit significant anti-inflammatory and anti-diabetic properties.
• The extremely low in vivo abundance of FAHFAs necessitates efficient synthesis methods for biological and clinical research.
• Developing scalable FAHFA synthesis is crucial for unlocking their therapeutic potential.

Purpose of the Study:

• To establish a feasible and effective enzymatic approach for synthesizing fatty acid esters of hydroxy fatty acids (FAHFAs).
• To create a novel in vitro bi-enzymatic cascade system for FAHFA production.
• To demonstrate the utility of this system for generating optically pure, structure-diverse FAHFAs.

Main Methods:

• Employed a bi-enzymatic cascade system utilizing hydratases from Lactobacillus acidophilus and Candida antarctica lipase A (CALA).
• Unsaturated fatty acids were converted to enantiomeric hydroxy fatty acids by hydratases.
• Hydroxy fatty acids were subsequently esterified with other fatty acids by CALA in a one-pot, two-step operation.

Main Results:

• Successfully synthesized various FAHFAs on a semi-preparative scale using the developed bi-enzymatic approach.
• The hydratase-CALA system demonstrated effective conversion of abundant fatty acids into desired FAHFAs.
• The method yielded optically pure and structure-diverse FAHFAs.

Conclusions:

• The novel hydratase-CALA bi-enzymatic system provides a promising and efficient route for FAHFA synthesis.
• This enzymatic approach overcomes the limitations of low natural abundance for FAHFA research and potential therapeutic applications.
• The developed method facilitates the production of diverse FAHFAs, enabling further investigation into their biological roles and clinical utility.