Biotechnologically relevant enzymes and proteins

Applied Microbiology and Biotechnology

, Volume 97, Issue 5, pp 1973-1985

Open Access This content is freely available online to anyone, anywhere at any time.

Identification and characterization of new Δ-17 fatty acid desaturases

  • Zhixiong XueAffiliated withBiochemical Science and Engineering, Central Research and Development, E. I. DuPont de Nemours Email author 
  • , Hongxian HeAffiliated withInformation and Computer Technology, E. I. DuPont de Nemours
  • , Dieter HollerbachAffiliated withBiochemical Science and Engineering, Central Research and Development, E. I. DuPont de Nemours
  • , Daniel J. MacoolAffiliated withBiochemical Science and Engineering, Central Research and Development, E. I. DuPont de Nemours
  • , Narendra S. YadavAffiliated withBiochemical Science and Engineering, Central Research and Development, E. I. DuPont de Nemours
  • , Hongxiang ZhangAffiliated withBiochemical Science and Engineering, Central Research and Development, E. I. DuPont de Nemours
  • , Bogdan SzostekAffiliated withCorporate Center for Analytical Sciences, E. I. DuPont de Nemours
  • , Quinn ZhuAffiliated withBiochemical Science and Engineering, Central Research and Development, E. I. DuPont de Nemours

Abstract

ω-3 fatty acid desaturase is a key enzyme for the biosynthesis of ω-3 polyunsaturated fatty acids via the oxidative desaturase/elongase pathways. Here we report the identification of three ω-3 desaturases from oomycetes, Pythium aphanidermatum, Phytophthora sojae, and Phytophthora ramorum. These new ω-3 desaturases share 55 % identity at the amino acid level with the known Δ-17 desaturase of Saprolegnia diclina, and about 31 % identity with the bifunctional Δ-12/Δ-15 desaturase of Fusarium monoliforme. The three enzymes were expressed in either wild-type or codon optimized form in an engineered arachidonic acid producing strain of Yarrowia lipolytica to study their activity and substrate specificity. All three were able to convert the ω-6 arachidonic acid to the ω-3 eicosapentanoic acid, with a substrate conversion efficiency of 54–65 %. These enzymes have a broad ω-6 fatty acid substrate spectrum, including both C18 and C20 ω-6 fatty acids although they prefer the C20 substrates, and have strong Δ-17 desaturase activity but weaker Δ-15 desaturase activity. Thus, they belong to the Δ-17 desaturase class. Unlike the previously identified bifunctional Δ-12/Δ-15 desaturase from F. monoliforme, they lack Δ-12 desaturase activity. The newly identified Δ-17 desaturases could use fatty acids in both acyl-CoA and phospholipid fraction as substrates. The identification of these Δ-17 desaturases provides a set of powerful new tools for genetic engineering of microbes and plants to produce ω-3 fatty acids, such as eicosapentanoic acid and docosahexanoic acid, at high levels.

Keywords

ω-3 Desaturase Δ-17 Desaturase Long-chain polyunsaturated fatty acids ω-3 Fatty acids Yarrowia lipolytica Metabolic engineering