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Increased fatty acid unsaturation and production of arachidonic acid by homologous over-expression of the mitochondrial malic enzyme in Mortierella alpina.

ABSTRACT: Malic enzyme (ME) catalyses the oxidative decarboxylation of L-malate to pyruvate and provides NADPH for intracellular metabolism, such as fatty acid synthesis. Here, the mitochondrial ME (mME) gene from Mortierella alpina was homologously over-expressed. Compared with controls, fungal arachidonic acid (ARA; 20:4 n-6) content increased by 60 % without affecting the total fatty acid content. Our results suggest that enhancing mME activity may be an effective mean to increase industrial production of ARA in M. alpina.

SUBMITTER: Hao G

PROVIDER: S-EPMC4386067 | biostudies-literature | 2014 Sep

REPOSITORIES: biostudies-literature

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Increased fatty acid unsaturation and production of arachidonic acid by homologous over-expression of the mitochondrial malic enzyme in Mortierella alpina.

Hao Guangfei G Chen Haiqin H Du Kai K Huang Xiaoyun X Song Yuanda Y Gu Zhennan Z Wang Lei L Zhang Hao H Chen Wei W Chen Yong Q YQ

Biotechnology letters 20140527 9

Malic enzyme (ME) catalyses the oxidative decarboxylation of L-malate to pyruvate and provides NADPH for intracellular metabolism, such as fatty acid synthesis. Here, the mitochondrial ME (mME) gene from Mortierella alpina was homologously over-expressed. Compared with controls, fungal arachidonic acid (ARA; 20:4 n-6) content increased by 60 % without affecting the total fatty acid content. Our results suggest that enhancing mME activity may be an effective mean to increase industrial production ...[more]

PMID: 24863290

Similar Datasets

Role of malic enzyme during fatty acid synthesis in the oleaginous fungus Mortierella alpina.

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Expression analysis for genes involved in arachidonic acid biosynthesis in Mortierella alpina CBS 754.68.

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An Online Respiratory Quotient-Feedback Strategy of Feeding Yeast Extract for Efficient Arachidonic Acid Production by Mortierella alpina.

Project description:Mortierella alpina (M. alpina) is well known for arachidonic acid (ARA) production. However, low efficiency and unstableness are long existed problems for industrial production of ARA by M. alpina due to the lack of online regulations. The aim of the present work is to develop an online-regulation strategy for efficient and stable ARA production in industry. The strategy was developed in 50 L fermenters and then applied in a 200 m3 fermenter. Results indicated that yeast extract (YE) highly increased cell growth in shake flask, it was then used in bioreactor fermentation by various feeding strategies. Feeding YE to control respiratory quotient (RQ) at 1.1 during 0-48 h and at 1.5 during 48-160 h, dry cell weight, and ARA titer reached 53.1 and 11.49 g/L in 50 L fermenter, which were increased by 79.4 and 36.9% as compared to that without YE feeding, respectively. Then, the online RQ-feedback strategy was applied in 200 m3 bioreactor fermentation and an average ARA titer of 16.82 g/L was obtained from 12 batches, which was 41.0% higher than the control batches. This is the first report on successful application of online RQ-feedback control of YE in ARA production, especially in an industrial scale of 200 m3 fermentation. It could be applied to other industrial production of microbial oil by oleaginous microorganisms.

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Production and enhancement of omega-3 fatty acid from Mortierella alpina CFR-GV15: its food and therapeutic application.

Project description:Mortierella sp. has been known to produce polyunsaturated fatty acids (PUFAs) such as GLA and AA under normal growth medium conditions. Similarly, under the stress condition, this fungus produces EPA and DHA in their mycelial biomass. Among the 67 soil samples screened from the Western Ghats of India, 11 Mortierella isolates showed the presence of omega-6 and omega-3 fatty acid, mainly GLA, AA, EPA, and DHA in starch, yeast-extract medium. Nile red and TTC strains were used for screening their qualitative oleaginesity. Among the representative isolates, when Mortierella sp. is grown in a fat-producing basal medium, a maximum lipid content of 42.0 ± 1.32% in its mycelia, 6.72 ± 0.5% EPA, and 4.09 ± 0.1% DHA was obtained. To understand the Mortierella sp. CFR-GV15, to the species level, its morphology was seen under the light microscope and scanning electron microscope, respectively. These microscopic observations showed that isolate Mortierella sp. CFR-GV15 produced coenocytic hyphae. Later on, its 18S rRNA and the internal transcribed spacer (ITS) sequences were cloned, sequenced, and analyzed phylogenetically to 18S rRNA and ITS1 and ITS4 sequences of related fungi. This newly isolated Mortierella alpina CFR-GV15 was found to be promising culture for the development of an economical method for commercial production of omega-3 fatty acid for food and therapeutical application.

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NADPH-Cytochrome P450 Reductase Mediates the Fatty Acid Desaturation of ω3 and ω6 Desaturases from <i>Mortierella alpina</i>.

Project description:Fatty acid desaturases play an important role in maintaining the appropriate structure and function of biological membranes. The biochemical characterization of integral membrane desaturases, particularly ω3 and ω6 desaturases, has been limited by technical difficulties relating to the acquisition of large quantities of purified proteins, and by the fact that functional activities of these proteins were only tested in an NADH-initiated reaction system. The main aim of this study was to reconstitute an NADPH-dependent reaction system in vitro and investigate the kinetic properties of Mortierella alpina ω3 and ω6 desaturases in this system. After expression and purification of the soluble catalytic domain of NADPH-cytochrome P450 reductase, the NADPH-dependent fatty acid desaturation was reconstituted for the first time in a system containing NADPH, NADPH-cytochrome P450 reductase, cytochrome b5, M. alpina ω3 and ω6 desaturase and detergent. In this system, the maximum activity of ω3 and ω6 desaturase was 213.4 ± 9.0 nmol min-1 mg-1 and 10.0 ± 0.5 nmol min-1 mg-1, respectively. The highest kcat/Km value of ω3 and ω6 desaturase was 0.41 µM-1 min-1 and 0.09 µM-1 min-1 when using linoleoyl CoA (18:2 ω6) and oleoyl CoA (18:1 ω9) as substrates, respectively. M. alpina ω3 and ω6 desaturases were capable of using NADPH as reductant when mediated by NADPH-cytochrome P450 reductase; although, their efficiency is distinguishable from NADH-dependent desaturation. These results provide insights into the mechanisms underlying ω3 and ω6 fatty acid desaturation and may facilitate the production of important fatty acids in M. alpina.

| S-EPMC9164069 | biostudies-literature

Influence of Supplementation of Vegetable Oil Blends on Omega-3 Fatty Acid Production in <i>Mortierella alpina</i> CFR-GV15.

Project description:Objectives of this study were designed for improved production of mycelial omega-3 fatty acids with particular reference to EPA and DHA from the oleaginous fungus Mortierella alpina CFR-GV15 under submerged low temperatures fermentation supplemented with linseed oil and garden cress oil as an additional energy source. The fungus was grown at 20°C temperature for four days initially followed by 12°C temperature for next five days. The basal medium contained starch, yeast extract, and a blend of linseed oil (LSO) and garden cress oil (GCO) in the ratio 1 : 1. Results of the study revealed that, after nine days of total incubation period, the enhancement of biomass was up to 16.7 g/L dry weight with a total lipid content of 55.4% (v/w). Enrichment of omega-3 fatty acids indicated a significant increase in fatty acid bioconversion (ALA 32.2 ± 0.42%, EPA 7.9 ± 0.1%, and DHA 4.09 ± 0.2%) by 2.5-fold. The two-stage temperature cultivation alters the fatty acid profile due to activation of the desaturase enzyme in the cellular levels due to which arachidonic acid (AA) content reduced significantly. It can be concluded that Mortierella alpina CFR-GV15 is a fungal culture suitable for commercial production of PUFAs with enriched EPA and DHA.

| S-EPMC5390627 | biostudies-literature

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