Two Co-Expression Strategies for Eicosapentaenoic Acid Production in Mortierella alpina

Xin Tang,Yong Q Chen,Zhennan Gu,Haiqin Chen,Xiaoke Zhang,Wei Chen,Hao Zhang

doi:10.3844/ajbbsp.2019.208.217

Xin Tang, Yong Q Chen + Show 5 more

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https://doi.org/10.3844/ajbbsp.2019.208.217

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Abstract

The omega-3 long-chain Polyunsaturated Fatty Acid (PUFA) Eicosapentaenoic Acid (EPA) has beneficial effects on human health, leading to its use in health products and foods. The oleaginous microorganism Mortierella alpina has been used in the industrial production of Arachidonic Acid (AA). The omega-3 desaturase oPpFADS17 is a key enzyme in the bioconversion of AA to EPA from Phytophthora parasitica and Glucose-6-Phosphate Dehydrogenase (G6PD2) is a critical enzyme that provide reducing power NADPH for lipid biosynthesis. In this study, we used a double expression cassette and 2A peptide strategies to co-express the these two critical enzymes. Subsequently, we investigated the effects of these strategies on total lipid and EPA accumulation. The recombination strains generated using the above-described strategies exhibited no differences in cell growth, compared with the control strain. Recombination strains generated using the double expression cassette exhibited an increase in total lipids to 43% of the cell dry weight, but did not accumulate EPA. Recombination strains generated using the 2A peptide strategy exhibited increased EPA accumulation, such that this PUFA accounted for 30% of the total lipid content. These co-expression strategies provide the improvements of multi-gene modification in PUFA accumulation in M. alpina.

Highlights

Eicosapentaenoic Acid (EPA, C20:5), an omega-3 long-chain Polyunsaturated Fatty Acid (LC-PUFA), has attracted considerable attention due to its beneficial effects on human health, including the prevention and treatment of diabetes, immune disorders, cardiovascular disease and cancer (Hirahashi et al, 2014; Maehre et al, 2015; Xue et al, 2013)
EPA is mainly accumulated in marine fish
When expressed in a recombinant strain of M. alpina, oPpFADS17 increased the proportion of EPA among the Total Fatty Acids (TFA) content to 31.5% while the control

Summary

Introduction

Eicosapentaenoic Acid (EPA, C20:5), an omega-3 long-chain Polyunsaturated Fatty Acid (LC-PUFA), has attracted considerable attention due to its beneficial effects on human health, including the prevention and treatment of diabetes, immune disorders, cardiovascular disease and cancer (Hirahashi et al, 2014; Maehre et al, 2015; Xue et al, 2013). Various microorganisms, including Mortierella alpina and Yarrowia lipolytica, have been investigated as potential hosts for the sustainable commercial production of EPA (Ando et al, 2009; Xue et al, 2013). M. alpina, an oleaginous filamentous fungus currently used to produce commercial Arachidonic Acid (AA, C20:4), can generate EPA from AA via an omega-3 desaturase (EC 1.14.19.-) (Ge et al, 2017; Ji et al, 2014; Okuda et al, 2015). Our latest research identified a new omega-3 desaturase, oPpFADS17, from Phytophthora parasitica. This enzyme converts AA to EPA at a conversion rate of 70%. When expressed in a recombinant strain of M. alpina, oPpFADS17 increased the proportion of EPA among the Total Fatty Acids (TFA) content to 31.5% while the control

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