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Sakayu Shimizu - One of the best experts on this subject based on the ideXlab platform.
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Arachidonic acid production by the oleaginous fungus Mortierella alpina 1S-4: A review
Elsevier, 2018Co-Authors: Hiroshi Kikukawa, Akinori Ando, Sakayu ShimizuAbstract:The filamentous fungus Mortierella alpina 1S-4 is capable of accumulating a large amount of triacylglycerol containing C20 polyunsaturated fatty acids (PUFAs). Indeed, triacylglycerol production by M. alpina 1S-4 can reach 20 g/L of culture broth, and the critical cellular signaling and structural PUFA arachidonic acid (ARA) comprises 30%–70% of the total fatty acid. The demonstrated health benefits of functional PUFAs have in turn encouraged the search for rich sources of these compounds, including fungal strains showing enhanced production of specific PUFAs. Screening for mutants and targeted gene manipulation of M. alpina 1S-4 have elucidated the functions of various enzymes involved in PUFA biosynthesis and established lines with improved PUFA productivity. In some cases, these strains have been used for indistrial-scale production of PUFAs, including ARA. In this review, we described practical ARA production through mutant breeding, functional analyses of genes encoding enzymes involved in PUFA biosynthesis, and recent advances in the production of specific PUFAs through molecular breeding of M. alpina 1S-4. Keywords: Arachidonic acid, Mortierella alpina, Molecular breeding, Fatty acid desaturas
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eicosapentaenoic acid epa production by an oleaginous fungus mortierella alpina expressing heterologous the δ17 desaturase gene under ordinary temperature
European Journal of Lipid Science and Technology, 2015Co-Authors: Tomoyo Okuda, Eiji Sakuradani, Sakayu Shimizu, Hiroshi Kikukawa, Akinori Ando, Hiroaki Negoro, Tatsuya Muratsubaki, Takaiku SakamotoAbstract:The oleaginous fungus Mortierella alpina is known to accumulate eicosapentaenoic acid (EPA) only when cultivated at a low temperature (below 15°C). Here, we investigated EPA production at an ordinary temperature (28°C) by expressing the Saprolegnia diclina Δ17 desaturase gene (sdd17m) in M. alpina ST1358, an ω3-desaturation activity-defective mutant derived from M. alpina 1S-4. Expression of the exogenous gene was confirmed by EPA accumulation in transformants at both 28 and 12°C. The EPA content in total lipids produced by transformants was over 20% at 28°C. Bench-scale fermentation with a 5-L jar fermentor showed that EPA content reached 26.4% of total fatty acids, and EPA production reached 1.8 g/L. This is the first study to report the accumulation of EPA in M. alpina at an ordinary temperature, and provide a platform technology for the industrial production of EPA using M. alpina as a promising source for EPA. Practical applications: This achievement shows the potential of M. alpina for industrial production of EPA at lower cost compared to conventional transgenic organisms such as plants and yeasts. The composition of fatty acids produced by M. alpina transformants was simple and did not contain unusual fatty acids compared with that of fish oils, so it may be acceptable for general consumers. Low-cost, large-scale, and high-purity EPA supply by M. alpina will also lead to development of academic research on physiological functions of EPA and its derivatives. The oleaginous fungus Mortierella alpina accumulates eicosapentaenoic acid (EPA) only when cultivated at a low temperature (below 15°C). Here, we investigated EPA production at an ordinary temperature (28°C) by expressing the Saprolegnia diclina Δ17 desaturase gene (sdd17m) in M. alpina. The transformants with the sdd17m gene produced EPA (>20% in total lipids) at 28°C.
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omega 3 eicosatetraenoic acid production by molecular breeding of the mutant strain s14 derived from mortierella alpina 1s 4
Journal of Bioscience and Bioengineering, 2015Co-Authors: Tomoyo Okuda, Eiji Sakuradani, Sakayu Shimizu, Hiroshi Kikukawa, Akinori Ando, Hiroaki Negoro, Takaiku Sakamoto, Jun OgawaAbstract:We investigated the omega-3 eicosatetraenoic acid (ETA) production by molecular breeding of the oleaginous fungus Mortierella alpina, which can slightly accumulate ETA only when cultivated at a low temperature. The endogenous ω3-desaturase gene or the heterologous Saprolegnia diclina Δ17 (sdd17m) desaturase gene were overexpressed in M. alpina S14, a Δ5-desaturation activity-defective mutant derived from M. alpina 1S-4. M. alpina S14 transformants introduced with the endogenous ω3-desaturase gene showed ETA at 42.1% content in the total lipids that was 84.2-fold and 3.2-fold higher than that of the wild-type strain 1S-4 and host strain S14, respectively, when cultivated at 12°C. No accumulation of ETA was observed at 28°C. In contrast, transformants with the heterologous sdd17m gene showed 24.9% of the content of total lipids at 28°C. These results indicated that these M. alpina S14 transformants are promising strains for the production of ETA, which is hard to obtain from natural sources.
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establishment of agrobacterium tumefaciens mediated transformation of an oleaginous fungus mortierella alpina 1s 4 and its application for eicosapentaenoic acid producer breeding
Applied and Environmental Microbiology, 2009Co-Authors: Akinori Ando, Yosuke Sumida, Hiroaki Negoro, Dian Anggraini Suroto, Sakayu ShimizuAbstract:Gene manipulation tools for an arachidonic-producing filamentous fungus, Mortierella alpina 1S-4, have not been sufficiently developed. In this study, Agrobacterium tumefaciens-mediated transformation (ATMT) was investigated for M. alpina 1S-4 transformation, using the uracil-auxotrophic mutant (ura5− strain) of M. alpina 1S-4 as a host strain and the homologous ura5 gene as a selectable marker gene. Furthermore, the gene for ω3-desaturase, catalyzing the conversion of n-6 fatty acid to n-3 fatty acid, was overexpressed in M. alpina 1S-4 by employing the ATMT system. As a result, we revealed that the frequency of transformation surpassed 400 transformants/108 spores, most of the integrated T-DNA appeared as a single copy at a random position in chromosomal DNA, and most of the transformants (60 to 80%) showed mitotic stability. Moreover, the accumulation of n-3 fatty acid in transformants was observed under the conditions of optimal ω3-desaturase gene expression. In particular, eicosapentaenoic acid (20:5n-3), an end product of n-3 fatty acids synthesized in M. alpina 1S-4, reached a maximum of 40% of total fatty acids. In conclusion, the ATMT system was found to be effective and suitable for the industrial strain Mortierella alpina 1S-4 and will be a useful tool for basic mutagenesis research and for industrial breeding of this strain.
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functional analysis of a fatty acid elongase from arachidonic acid producing mortierella alpina 1s 4
Applied Microbiology and Biotechnology, 2008Co-Authors: Eiji Sakuradani, Shoichi Murata, Hiroyuki Kanamaru, Sakayu ShimizuAbstract:We describe the isolation and characterization of a gene (MAELO) that encodes a fatty acid elongase from arachidonic acid-producing fungus Mortierella alpina 1S-4. Although the homologous MAELO gene had already been isolated from M. alpina ATCC 32221, its function had not yet been identified. The MAELO gene from M. alpina 1S-4 was confirmed to encode a fatty acid elongase by its expression in yeast Saccharomyces cerevisiae. Analysis of the fatty acid composition of the yeast transformant revealed the accumulation of 22-, 24-, and 26-carbon saturated fatty acids. On the other hand, RNA interference of the MAELO gene in M. alpina 1S-4 was carried out. The gene-silenced strain obtained on RNA interference exhibited low contents of 20-, 22-, and 24-carbon saturated fatty acids and a high content of stearic acid (18 carbons), compared with those in the wild strain. The enzyme encoded by the MAELO gene was demonstrated to be involved in the biosynthesis of 20-, 22-, and 24-carbon saturated fatty acids in M. alpina 1S-4.
Eiji Sakuradani - One of the best experts on this subject based on the ideXlab platform.
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eicosapentaenoic acid epa production by an oleaginous fungus mortierella alpina expressing heterologous the δ17 desaturase gene under ordinary temperature
European Journal of Lipid Science and Technology, 2015Co-Authors: Tomoyo Okuda, Eiji Sakuradani, Sakayu Shimizu, Hiroshi Kikukawa, Akinori Ando, Hiroaki Negoro, Tatsuya Muratsubaki, Takaiku SakamotoAbstract:The oleaginous fungus Mortierella alpina is known to accumulate eicosapentaenoic acid (EPA) only when cultivated at a low temperature (below 15°C). Here, we investigated EPA production at an ordinary temperature (28°C) by expressing the Saprolegnia diclina Δ17 desaturase gene (sdd17m) in M. alpina ST1358, an ω3-desaturation activity-defective mutant derived from M. alpina 1S-4. Expression of the exogenous gene was confirmed by EPA accumulation in transformants at both 28 and 12°C. The EPA content in total lipids produced by transformants was over 20% at 28°C. Bench-scale fermentation with a 5-L jar fermentor showed that EPA content reached 26.4% of total fatty acids, and EPA production reached 1.8 g/L. This is the first study to report the accumulation of EPA in M. alpina at an ordinary temperature, and provide a platform technology for the industrial production of EPA using M. alpina as a promising source for EPA. Practical applications: This achievement shows the potential of M. alpina for industrial production of EPA at lower cost compared to conventional transgenic organisms such as plants and yeasts. The composition of fatty acids produced by M. alpina transformants was simple and did not contain unusual fatty acids compared with that of fish oils, so it may be acceptable for general consumers. Low-cost, large-scale, and high-purity EPA supply by M. alpina will also lead to development of academic research on physiological functions of EPA and its derivatives. The oleaginous fungus Mortierella alpina accumulates eicosapentaenoic acid (EPA) only when cultivated at a low temperature (below 15°C). Here, we investigated EPA production at an ordinary temperature (28°C) by expressing the Saprolegnia diclina Δ17 desaturase gene (sdd17m) in M. alpina. The transformants with the sdd17m gene produced EPA (>20% in total lipids) at 28°C.
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omega 3 eicosatetraenoic acid production by molecular breeding of the mutant strain s14 derived from mortierella alpina 1s 4
Journal of Bioscience and Bioengineering, 2015Co-Authors: Tomoyo Okuda, Eiji Sakuradani, Sakayu Shimizu, Hiroshi Kikukawa, Akinori Ando, Hiroaki Negoro, Takaiku Sakamoto, Jun OgawaAbstract:We investigated the omega-3 eicosatetraenoic acid (ETA) production by molecular breeding of the oleaginous fungus Mortierella alpina, which can slightly accumulate ETA only when cultivated at a low temperature. The endogenous ω3-desaturase gene or the heterologous Saprolegnia diclina Δ17 (sdd17m) desaturase gene were overexpressed in M. alpina S14, a Δ5-desaturation activity-defective mutant derived from M. alpina 1S-4. M. alpina S14 transformants introduced with the endogenous ω3-desaturase gene showed ETA at 42.1% content in the total lipids that was 84.2-fold and 3.2-fold higher than that of the wild-type strain 1S-4 and host strain S14, respectively, when cultivated at 12°C. No accumulation of ETA was observed at 28°C. In contrast, transformants with the heterologous sdd17m gene showed 24.9% of the content of total lipids at 28°C. These results indicated that these M. alpina S14 transformants are promising strains for the production of ETA, which is hard to obtain from natural sources.
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Improved production of various polyunsaturated fatty acids through filamentous fungus Mortierella alpina breeding
Applied Microbiology and Biotechnology, 2009Co-Authors: Eiji Sakuradani, Akinori Ando, Jun OgawaAbstract:Studies on the application of functional lipids such as polyunsaturated fatty acids (PUFAs) have proceeded in various fields regarding health and dietary requirements in a search for novel and rich sources. Filamentous fungus Mortierella alpina 1S-4 produces triacylglycerols rich in arachidonic acid, ones reaching 20 g/L and containing 30–70% arachidonic acid as to the total fatty acids. Mutants derived from M. alpina 1S-4, defective in Δ5 and Δ6 desaturases, accumulate triacylglycerols rich in unique PUFAs, i.e., dihomo-γ-linolenic acid and Mead acid, respectively. Furthermore, various mutants derived from M. alpina 1S-4 have led to the production of oils containing n −1, n −3, n −4, n −6, n −7, and n −9 PUFAs. A variety of genes encoding fatty acid desaturases and elongases involved in PUFA biosynthesis in M. alpina 1S-4 has been isolated and characterized. Molecular breeding of M. alpina strains by means of manipulation of these genes facilitates improvement of PUFA productivity and elucidation of the functions of enzymes involved in PUFA biosynthesis.
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functional analysis of a fatty acid elongase from arachidonic acid producing mortierella alpina 1s 4
Applied Microbiology and Biotechnology, 2008Co-Authors: Eiji Sakuradani, Shoichi Murata, Hiroyuki Kanamaru, Sakayu ShimizuAbstract:We describe the isolation and characterization of a gene (MAELO) that encodes a fatty acid elongase from arachidonic acid-producing fungus Mortierella alpina 1S-4. Although the homologous MAELO gene had already been isolated from M. alpina ATCC 32221, its function had not yet been identified. The MAELO gene from M. alpina 1S-4 was confirmed to encode a fatty acid elongase by its expression in yeast Saccharomyces cerevisiae. Analysis of the fatty acid composition of the yeast transformant revealed the accumulation of 22-, 24-, and 26-carbon saturated fatty acids. On the other hand, RNA interference of the MAELO gene in M. alpina 1S-4 was carried out. The gene-silenced strain obtained on RNA interference exhibited low contents of 20-, 22-, and 24-carbon saturated fatty acids and a high content of stearic acid (18 carbons), compared with those in the wild strain. The enzyme encoded by the MAELO gene was demonstrated to be involved in the biosynthesis of 20-, 22-, and 24-carbon saturated fatty acids in M. alpina 1S-4.
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a novel fungal ω3 desaturase with wide substrate specificity from arachidonic acid producing mortierella alpina 1s 4
Applied Microbiology and Biotechnology, 2005Co-Authors: Eiji Sakuradani, Takahiro Abe, Keita Iguchi, Sakayu ShimizuAbstract:A filamentous fungus, Mortierella alpina 1S-4, is capable of producing not only arachidonic acid (AA; 20:4n-6) but also eicosapentaenoic acid (EPA; 20:5n-3) below a cultural temperature of 20°C. Here, we describe the isolation and characterization of a gene (maw3) that encodes a novel ω3-desaturase from M. alpina 1S-4. Based on the conserved sequence information for M. alpina 1S-4 Δ12-desaturase and Saccharomyces kluyveri ω3-desaturase, the ω3-desaturase gene from M. alpina 1S-4 was cloned. Homology analysis of protein databases revealed that the amino acid sequence showed 51% identity, at the highest, with M. alpina 1S-4 Δ12-desaturase, whereas it exhibited 36% identity with Sac. kluyveri ω3-desaturase. The cloned cDNA was confirmed to encode the ω3-desaturase by its expression in the yeast Sac. cerevisiae. Analysis of the fatty acid composition of the yeast transformant demonstrated that 18-carbon and 20-carbon n-3 polyunsaturated fatty acids (PUFAs) were accumulated through conversion of exogenous 18-carbon and 20-carbon n-6 PUFAs. The substrate specificity of the M. alpina 1S-4 ω3-desaturase differs from those of the known fungal ω3-desaturases from Sac. kluyveri and Saprolegnia diclina. Plant, cyanobacterial and Sac. kluyveri ω3-desaturases desaturate 18-carbon n-6 PUFAs, Spr. diclina ω3-desaturase desaturates 20-carbon n-6 PUFAs and Caenorhabditis elegans ω3-desaturase prefers 18-carbon n-6 PUFAs as substrates rather than 20-carbon n-6 PUFAs. The substrate specificity of M. alpina 1S-4 ω3-desaturase is rather similar to that of C. elegans ω3-desaturase, but the M. alpina ω3-desaturase can more effectively convert AA into EPA when expressed in yeast. The M. alpina 1S-4 ω3-desaturase is the first known fungal desaturase that uses both 18-carbon and 20-carbon n-6 PUFAs as substrates.
Akinori Ando - One of the best experts on this subject based on the ideXlab platform.
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Arachidonic acid production by the oleaginous fungus Mortierella alpina 1S-4: A review
Elsevier, 2018Co-Authors: Hiroshi Kikukawa, Akinori Ando, Sakayu ShimizuAbstract:The filamentous fungus Mortierella alpina 1S-4 is capable of accumulating a large amount of triacylglycerol containing C20 polyunsaturated fatty acids (PUFAs). Indeed, triacylglycerol production by M. alpina 1S-4 can reach 20 g/L of culture broth, and the critical cellular signaling and structural PUFA arachidonic acid (ARA) comprises 30%–70% of the total fatty acid. The demonstrated health benefits of functional PUFAs have in turn encouraged the search for rich sources of these compounds, including fungal strains showing enhanced production of specific PUFAs. Screening for mutants and targeted gene manipulation of M. alpina 1S-4 have elucidated the functions of various enzymes involved in PUFA biosynthesis and established lines with improved PUFA productivity. In some cases, these strains have been used for indistrial-scale production of PUFAs, including ARA. In this review, we described practical ARA production through mutant breeding, functional analyses of genes encoding enzymes involved in PUFA biosynthesis, and recent advances in the production of specific PUFAs through molecular breeding of M. alpina 1S-4. Keywords: Arachidonic acid, Mortierella alpina, Molecular breeding, Fatty acid desaturas
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eicosapentaenoic acid epa production by an oleaginous fungus mortierella alpina expressing heterologous the δ17 desaturase gene under ordinary temperature
European Journal of Lipid Science and Technology, 2015Co-Authors: Tomoyo Okuda, Eiji Sakuradani, Sakayu Shimizu, Hiroshi Kikukawa, Akinori Ando, Hiroaki Negoro, Tatsuya Muratsubaki, Takaiku SakamotoAbstract:The oleaginous fungus Mortierella alpina is known to accumulate eicosapentaenoic acid (EPA) only when cultivated at a low temperature (below 15°C). Here, we investigated EPA production at an ordinary temperature (28°C) by expressing the Saprolegnia diclina Δ17 desaturase gene (sdd17m) in M. alpina ST1358, an ω3-desaturation activity-defective mutant derived from M. alpina 1S-4. Expression of the exogenous gene was confirmed by EPA accumulation in transformants at both 28 and 12°C. The EPA content in total lipids produced by transformants was over 20% at 28°C. Bench-scale fermentation with a 5-L jar fermentor showed that EPA content reached 26.4% of total fatty acids, and EPA production reached 1.8 g/L. This is the first study to report the accumulation of EPA in M. alpina at an ordinary temperature, and provide a platform technology for the industrial production of EPA using M. alpina as a promising source for EPA. Practical applications: This achievement shows the potential of M. alpina for industrial production of EPA at lower cost compared to conventional transgenic organisms such as plants and yeasts. The composition of fatty acids produced by M. alpina transformants was simple and did not contain unusual fatty acids compared with that of fish oils, so it may be acceptable for general consumers. Low-cost, large-scale, and high-purity EPA supply by M. alpina will also lead to development of academic research on physiological functions of EPA and its derivatives. The oleaginous fungus Mortierella alpina accumulates eicosapentaenoic acid (EPA) only when cultivated at a low temperature (below 15°C). Here, we investigated EPA production at an ordinary temperature (28°C) by expressing the Saprolegnia diclina Δ17 desaturase gene (sdd17m) in M. alpina. The transformants with the sdd17m gene produced EPA (>20% in total lipids) at 28°C.
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omega 3 eicosatetraenoic acid production by molecular breeding of the mutant strain s14 derived from mortierella alpina 1s 4
Journal of Bioscience and Bioengineering, 2015Co-Authors: Tomoyo Okuda, Eiji Sakuradani, Sakayu Shimizu, Hiroshi Kikukawa, Akinori Ando, Hiroaki Negoro, Takaiku Sakamoto, Jun OgawaAbstract:We investigated the omega-3 eicosatetraenoic acid (ETA) production by molecular breeding of the oleaginous fungus Mortierella alpina, which can slightly accumulate ETA only when cultivated at a low temperature. The endogenous ω3-desaturase gene or the heterologous Saprolegnia diclina Δ17 (sdd17m) desaturase gene were overexpressed in M. alpina S14, a Δ5-desaturation activity-defective mutant derived from M. alpina 1S-4. M. alpina S14 transformants introduced with the endogenous ω3-desaturase gene showed ETA at 42.1% content in the total lipids that was 84.2-fold and 3.2-fold higher than that of the wild-type strain 1S-4 and host strain S14, respectively, when cultivated at 12°C. No accumulation of ETA was observed at 28°C. In contrast, transformants with the heterologous sdd17m gene showed 24.9% of the content of total lipids at 28°C. These results indicated that these M. alpina S14 transformants are promising strains for the production of ETA, which is hard to obtain from natural sources.
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establishment of agrobacterium tumefaciens mediated transformation of an oleaginous fungus mortierella alpina 1s 4 and its application for eicosapentaenoic acid producer breeding
Applied and Environmental Microbiology, 2009Co-Authors: Akinori Ando, Yosuke Sumida, Hiroaki Negoro, Dian Anggraini Suroto, Sakayu ShimizuAbstract:Gene manipulation tools for an arachidonic-producing filamentous fungus, Mortierella alpina 1S-4, have not been sufficiently developed. In this study, Agrobacterium tumefaciens-mediated transformation (ATMT) was investigated for M. alpina 1S-4 transformation, using the uracil-auxotrophic mutant (ura5− strain) of M. alpina 1S-4 as a host strain and the homologous ura5 gene as a selectable marker gene. Furthermore, the gene for ω3-desaturase, catalyzing the conversion of n-6 fatty acid to n-3 fatty acid, was overexpressed in M. alpina 1S-4 by employing the ATMT system. As a result, we revealed that the frequency of transformation surpassed 400 transformants/108 spores, most of the integrated T-DNA appeared as a single copy at a random position in chromosomal DNA, and most of the transformants (60 to 80%) showed mitotic stability. Moreover, the accumulation of n-3 fatty acid in transformants was observed under the conditions of optimal ω3-desaturase gene expression. In particular, eicosapentaenoic acid (20:5n-3), an end product of n-3 fatty acids synthesized in M. alpina 1S-4, reached a maximum of 40% of total fatty acids. In conclusion, the ATMT system was found to be effective and suitable for the industrial strain Mortierella alpina 1S-4 and will be a useful tool for basic mutagenesis research and for industrial breeding of this strain.
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Improved production of various polyunsaturated fatty acids through filamentous fungus Mortierella alpina breeding
Applied Microbiology and Biotechnology, 2009Co-Authors: Eiji Sakuradani, Akinori Ando, Jun OgawaAbstract:Studies on the application of functional lipids such as polyunsaturated fatty acids (PUFAs) have proceeded in various fields regarding health and dietary requirements in a search for novel and rich sources. Filamentous fungus Mortierella alpina 1S-4 produces triacylglycerols rich in arachidonic acid, ones reaching 20 g/L and containing 30–70% arachidonic acid as to the total fatty acids. Mutants derived from M. alpina 1S-4, defective in Δ5 and Δ6 desaturases, accumulate triacylglycerols rich in unique PUFAs, i.e., dihomo-γ-linolenic acid and Mead acid, respectively. Furthermore, various mutants derived from M. alpina 1S-4 have led to the production of oils containing n −1, n −3, n −4, n −6, n −7, and n −9 PUFAs. A variety of genes encoding fatty acid desaturases and elongases involved in PUFA biosynthesis in M. alpina 1S-4 has been isolated and characterized. Molecular breeding of M. alpina strains by means of manipulation of these genes facilitates improvement of PUFA productivity and elucidation of the functions of enzymes involved in PUFA biosynthesis.
Jun Ogawa - One of the best experts on this subject based on the ideXlab platform.
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omega 3 eicosatetraenoic acid production by molecular breeding of the mutant strain s14 derived from mortierella alpina 1s 4
Journal of Bioscience and Bioengineering, 2015Co-Authors: Tomoyo Okuda, Eiji Sakuradani, Sakayu Shimizu, Hiroshi Kikukawa, Akinori Ando, Hiroaki Negoro, Takaiku Sakamoto, Jun OgawaAbstract:We investigated the omega-3 eicosatetraenoic acid (ETA) production by molecular breeding of the oleaginous fungus Mortierella alpina, which can slightly accumulate ETA only when cultivated at a low temperature. The endogenous ω3-desaturase gene or the heterologous Saprolegnia diclina Δ17 (sdd17m) desaturase gene were overexpressed in M. alpina S14, a Δ5-desaturation activity-defective mutant derived from M. alpina 1S-4. M. alpina S14 transformants introduced with the endogenous ω3-desaturase gene showed ETA at 42.1% content in the total lipids that was 84.2-fold and 3.2-fold higher than that of the wild-type strain 1S-4 and host strain S14, respectively, when cultivated at 12°C. No accumulation of ETA was observed at 28°C. In contrast, transformants with the heterologous sdd17m gene showed 24.9% of the content of total lipids at 28°C. These results indicated that these M. alpina S14 transformants are promising strains for the production of ETA, which is hard to obtain from natural sources.
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Improved production of various polyunsaturated fatty acids through filamentous fungus Mortierella alpina breeding
Applied Microbiology and Biotechnology, 2009Co-Authors: Eiji Sakuradani, Akinori Ando, Jun OgawaAbstract:Studies on the application of functional lipids such as polyunsaturated fatty acids (PUFAs) have proceeded in various fields regarding health and dietary requirements in a search for novel and rich sources. Filamentous fungus Mortierella alpina 1S-4 produces triacylglycerols rich in arachidonic acid, ones reaching 20 g/L and containing 30–70% arachidonic acid as to the total fatty acids. Mutants derived from M. alpina 1S-4, defective in Δ5 and Δ6 desaturases, accumulate triacylglycerols rich in unique PUFAs, i.e., dihomo-γ-linolenic acid and Mead acid, respectively. Furthermore, various mutants derived from M. alpina 1S-4 have led to the production of oils containing n −1, n −3, n −4, n −6, n −7, and n −9 PUFAs. A variety of genes encoding fatty acid desaturases and elongases involved in PUFA biosynthesis in M. alpina 1S-4 has been isolated and characterized. Molecular breeding of M. alpina strains by means of manipulation of these genes facilitates improvement of PUFA productivity and elucidation of the functions of enzymes involved in PUFA biosynthesis.
Wei Chen - One of the best experts on this subject based on the ideXlab platform.
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Effects of Agrobacterium tumefaciens strain types on the Agrobacterium-mediated transformation efficiency of filamentous fungus Mortierella alpina.
Letters in applied microbiology, 2020Co-Authors: Shunxian Wang, Haiqin Chen, Wei Chen, Hao Zhang, Xin Tang, Yuzhuo Wang, Caixing Pan, Yong Q. ChenAbstract:Four Agrobacterium tumefaciens strains AGL‐1, C58C1, EHA105 and LBA4404 were tested for the effects of strain types on the transformation efficiency in Mortierella alpina. Results showed that AGL‐1, C58C1 and EHA105 transformed M. alpina successfully. Among them, A. tumefaciens EHA105 was first proven successful transformation of M. alpina. AGL‐1 and EHA105 had the highest transformation efficiency among the four strains, while LBA4404 failed to transform M. alpina. The reason leading to the transformation efficiency difference among the four strains was explored by determining transcription levels of the virulence (vir) gene in the induction medium. Results showed that the expressions of virD1, virD2, virD4 and virE1 genes were obviously induced by acetosyringone in all the strains, and their transcriptional levels as well as virA’s of AGL‐1, C58C1 and EHA105 were higher than that of LBA4404, suggesting high transcriptional levels of vir genes were important for successful transformation. The study selected A. tumefaciens with high transformation efficiency of M. alpina, and would accelerate the genetic management of M. alpina. SIGNIFICANCE AND IMPACT OF THE STUDY: Oleaginous filamentous fungus Mortierella alpina is a commercial strain for the production of arachidonic acid. Genetic manipulation of M. alpina requires highly efficient transformation method. In this study, we explore the effect of Agrobacterium tumefaciens strain types on the transformation efficiency of M. alpina and select A. tumefaciens with the highest transformation efficiency, which accelerates the genetic manipulation of M. alpina. Besides, high transcriptional levels of virulence genes in A. tumefaciens were proven to play an important role for successful transformation.
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evaluation of metabolome sample preparation and extraction methodologies for oleaginous filamentous fungi mortierella alpina
Metabolomics, 2019Co-Authors: Haiqin Chen, Yong Q. Chen, Hao Zhang, Xin Tang, Qin Yang, Wei ChenAbstract:Metabolomics has been successfully applied to guide the rational engineering of industrial strains and improve the performance of bioprocesses. Mortierella alpina has traditionally been one of the most popular industrial strains for the production of polyunsaturated fatty acids. However, a systematic comparison and optimisation of the metabolomic analysis methods of M. alpina has not yet been reported. We sought to identify potential weaknesses that are important for accurate metabolomic analysis. We also aimed to determine an efficient sample preparation protocol for metabolomics studies in the oleaginous filamentous fungus M. alpina. In this study, using GC-MS, we evaluated three sample preparation protocols and five solvent mixtures by assessment of the metabolite profile differences, the sum of peak intensities and the reproducibility of metabolite quantification. The freeze-dried biomass had better reproducibility and recovery than fresh biomass for metabolite extraction and data normalisation that is part of a metabolomics analysis of filamentous fungi M. alpina. Methanol:water (1:1) was superior for the profiling of metabolites in oleaginous fungi M. alpina. The unbiased metabolite profiling difference between the growth phase and lipids synthesis phase revealed that the degradation of amino acids were critical nodes for the efficient synthesis of lipids in M. alpina. The use of freeze-dried biomass for metabolite extraction and data normalisation was more efficient at measuring the active state of the intracellular metabolites in M. alpina. We recommend extracting the intracellular metabolites with methanol:water (1:1). An important role of amino acid oxidation in the nitrogen limitation-mediated lipid accumulation was found.
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Genome Characterization of the Oleaginous Fungus Mortierella alpina
2016Co-Authors: Wei Chen, Haiqin Chen, Hongchao Wang, Yun Feng, Yan Ren, Michael J. Thomas, Baixi Zhang, Isabelle M. BerquinAbstract:Mortierella alpina is an oleaginous fungus which can produce lipids accounting for up to 50 % of its dry weight in the form of triacylglycerols. It is used commercially for the production of arachidonic acid. Using a combination of high throughput sequencing and lipid profiling, we have assembled the M. alpina genome, mapped its lipogenesis pathway and determined its major lipid species. The 38.38 Mb M. alpina genome shows a high degree of gene duplications. Approximately 50 % of its 12,796 gene models, and 60 % of genes in the predicted lipogenesis pathway, belong to multigene families. Notably, M. alpina has 18 lipase genes, of which 11 contain the class 2 lipase domain and may share a similar function. M. alpina’s fatt
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Genome Characterization of the Oleaginous Fungus Mortierella alpina
PloS one, 2011Co-Authors: Wei Chen, Haiqin Chen, Hongchao Wang, Yun Feng, Yan Ren, Michael J. Thomas, Baixi Zhang, Isabelle M. BerquinAbstract:Mortierella alpina is an oleaginous fungus which can produce lipids accounting for up to 50% of its dry weight in the form of triacylglycerols. It is used commercially for the production of arachidonic acid. Using a combination of high throughput sequencing and lipid profiling, we have assembled the M. alpina genome, mapped its lipogenesis pathway and determined its major lipid species. The 38.38 Mb M. alpina genome shows a high degree of gene duplications. Approximately 50% of its 12,796 gene models, and 60% of genes in the predicted lipogenesis pathway, belong to multigene families. Notably, M. alpina has 18 lipase genes, of which 11 contain the class 2 lipase domain and may share a similar function. M. alpina's fatty acid synthase is a single polypeptide containing all of the catalytic domains required for fatty acid synthesis from acetyl-CoA and malonyl-CoA, whereas in many fungi this enzyme is comprised of two polypeptides. Major lipids were profiled to confirm the products predicted in the lipogenesis pathway. M. alpina produces a complex mixture of glycerolipids, glycerophospholipids and sphingolipids. In contrast, only two major sterol lipids, desmosterol and 24(28)-methylene-cholesterol, were detected. Phylogenetic analysis based on genes involved in lipid metabolism suggests that oleaginous fungi may have acquired their lipogenic capacity during evolution after the divergence of Ascomycota, Basidiomycota, Chytridiomycota and Mucoromycota. Our study provides the first draft genome and comprehensive lipid profile for M. alpina, and lays the foundation for possible genetic engineering of M. alpina to produce higher levels and diverse contents of dietary lipids.