Determination of FAT-1 desaturase activity and substrate preference

Area of Science:

• Biochemistry
• Molecular Biology
• Genetics

Background:

• Omega-3 polyunsaturated fatty acids (PUFAs) offer significant health advantages.
• The FAT-1 desaturase enzyme, found in C. elegans, converts omega-6 PUFAs to omega-3 PUFAs.
• Previous studies utilized transgenic FAT-1 to enhance omega-3 levels in pigs, fish, and mice, but its precise activity and substrate preference were not fully understood.

Purpose of the Study:

• To elucidate the mechanism behind FAT-1 desaturase activity and substrate preference.
• To provide a structural understanding of FAT-1 using computational methods.
• To explore the potential of FAT-1 in improving omega-3 fatty acid content.

Main Methods:

• Utilized AlphaFold for predicting the structure of FAT-1.
• Analyzed the functional domains and key amino acid residues essential for FAT-1 activity.
• Performed experiments in both C. elegans and mammalian cells to assess FAT-1's conversion capabilities.

Main Results:

• AlphaFold predicted FAT-1 as an endoplasmic reticulum integral membrane protein with distinct transmembrane, CYTB5, and desaturase domains.
• A specific region (amino acids 210-217), particularly G212, G216, and S217, was identified as crucial for FAT-1 activity.
• FAT-1 demonstrated the ability to convert all tested n-6 PUFAs to n-3 PUFAs in both C. elegans and mammalian cells, with a potential preference for converting arachidonic acid (C20:4n6) to EPA (C20:5n3).

Conclusions:

• The study reveals the structural basis and key active sites of FAT-1 desaturase.
• FAT-1 exhibits broad substrate specificity for n-6 PUFA conversion to n-3 PUFAs.
• These findings enhance our understanding of FAT-1's mechanism, supporting its future transgenic applications for nutritional benefits.