Identification of two novel microalgal enzymes involved in the conversion of the ω3-fatty acid, eicosapentaenoic acid, into docosahexaenoic acid

Abstract

Marine microalgae such as Pavlova and Isochrysis produce abundant amounts of the omega3-PUFAs (polyunsaturated fatty acids), EPA (eicosapentaenoic acid, 20:5n-3) and DHA (docosahexaenoic acid, 22:6n-3). The pathway leading to the conversion of EPA into DHA in these lower eukaryotes is not well established although it is predicted to involve an elongation step, catalysed by an elongating enzyme complex, leading to the conversion of EPA into omega3-DPA (omega-3-docosapentaenoic acid, 22:5n-3); followed by a desaturation step, catalysed by a Delta4-desaturase, which results in the conversion of DPA into DHA. To date, the enzymes involved in the elongation of EPA have not been identified from any lower eukaryote. In the present study, we describe the identification of microalgal genes involved in the two-step conversion of EPA into DHA. By expressed sequence tag analysis, a gene (pavELO) encoding a novel elongase was identified from Pavlova, which catalysed the conversion of EPA into omega3-DPA in yeast. Unlike any previously identified elongase from higher or lower eukaryotes, this enzyme displayed unique substrate specificity for both n-6 and n-3 C20-PUFA substrates, with no activity towards any C18- or C22-PUFA substrates. In addition, a novel Delta4-desaturase gene (IgD4) was isolated from Isochrysis, which was capable of converting omega3-DPA into DHA, as well as adrenic acid (22:4n-6) into omega6-DPA. Yeast co-expression studies, with pavELO and IgD4, revealed that these genes were capable of functioning together to carry out the two-step conversion of EPA into DHA.

Isolation and functional characterization of fatty acid Δ5‐elongase gene from the liverwort Marchantia polymorpha L

“…To date, D4-desaturase genes have been identified from a marine fungus and several microalgae [8][9][10][11]. Genes encoding an enzyme with D5-elongating activity have also been isolated from marine microalgae [11,12] and several vertebrates [5]. Their amino acid sequences show similarity to Saccharomyces cerevisiae ELO1, ELO2, and ELO3 [13,14] and other ELO-like enzymes including D6-elongase [5].…”

“…In lower eukaryotes, DHA is synthesized more simply by D5-elongation from EPA to docosapentaenoic acid (DPA, 22:5D 7,10,13,16,19 ), followed by D4-desaturation. To date, D4-desaturase genes have been identified from a marine fungus and several microalgae [8][9][10][11]. Genes encoding an enzyme with D5-elongating activity have also been isolated from marine microalgae [11,12] and several vertebrates [5].…”

“…Very-long-chain polyunsaturated fatty acids (VLCPUFAs) represented by arachidonic acid (AA; 20:4D 5,8,11,14 ), eicosapentaenoic acid (EPA; 20:5D 5,8,11,14,17 ), and docosahexaenoic (DHA; 22:6D 4,7,10,13,16,19 ) acid are widely recognized, because of their beneficial roles in human health. In particular, DHA is an essential component of membrane lipids of brain neurons, testes, and the external segments of photoreceptors in the retina [1].…”

Characterization of membrane-bound fatty acid desaturases

“…It was isolated from Thraustochytrium sp. (Qui et al 2001;Zank et al 2005), Schizochytrium aggregatum (Kinney et al 2004), Isochrysis galbana (Pereira et al 2004), Thalassiosira pseudonana (Tonon et al 2005), Pavlova viridis (Xu et al 2011). It has typical sign for 'front-end' desaturase -Q in 3 rd His box instead of H, cytochrome b 5 domain on N-terminal, but in 2 nd His box contains one more amino acid residue (Fig.…”

The production of very-long-chain PUFA biosynthesis in transgenic plants: towards a sustainable source of fish oils

“…The cytochrome b 5 -fusion C 22 D 4 -desaturase has been identified and functionally characterised from several aquatic micro-organisms (Qiu et al 2001). More recently, the specific elongating activity required for the synthesis of 22: 5 (n-3) from EPA has been characterised (Meyer et al 2004;Pereira et al 2004). However, it might be expected that the previouslymentioned constraints on efficient elongation associated with poor acyl exchange between glycerolipids and the acyl-CoA pool might also apply to the synthesis of DHA, with a potential bottleneck occurring via the accumulation of EPA in phosphatidylcholine (see Fig.…”