JOURNAL OF BIOTECHNOLOGY
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2020
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IF 3.503
Hengqian Lu,Haiqin Chen,Xin Tang,Qin Yang,Hao Zhang,Yong Q. Chen,Wei Chen
Lipid hyperaccumulation in oleaginous microorganisms is generally induced by nitrogen limitation, while oxygen supply can influence biomass growth and cell metabolism. Although strategies based on nitrogen limitation or oxygen control have been extensively explored and applied in various oleaginous microorganisms, the role of oxygen supply in nitrogen limitation induced lipid hyperaccumulation still remains unclear. Here, we systematically surveyed the effects of oxygen supply on the oleaginous fungus M. alpina cultured in nitrogen limited conditions through integration of physiochemical parameters and metabolomics analysis. Our results indicated that a high oxygen supply promoted carbon/nitrogen consumption and was used for rapid biomass synthesis, while either high or low oxygen supply conditions were adverse to lipid and ARA accumulation. Different oxygen supply level significantly affected the balance between fermentation for lipid synthesis and respiration for energy generation. Under nitrogen limitation, a suitable oxygen supply promoted the recycling of preformed nitrogen and increased the redirection of carbon towards fatty acid synthesis through the hub centred around glutamic acid coupled to the intermediate metabolism of carbon in the TCA cycle, while a high oxygen supply favored the respiration process and enhanced the degradation of LC-PUFAs, rather than fermentation for fatty acid synthesis. This system-level insight reveals the underlying metabolic mechanism of oxygen control in nitrogen limitation induced lipid accumulation, and provides theoretical support for the integration of oxygen control with nutrient supply for efficient microbial oil production.
研究领域:
微生物学
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代谢生物学
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DOI
PubMed
高影响因子
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
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2019
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IF 3.571
Hengqian Lu,Haiqin Chen,Xin Tang,Qin Yang,Hao Zhang,Yong Q. Chen,Wei Chen
The objective of the present study was to reveal the effects of four types of nitrogen sources (soymeal, yeast extract, KNO3, and ammonium tartrate) on the lipid metabolism of the oleaginous fungus Mortierella alpina using untargeted lipidomics, targeted fatty acid, and reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis. Our results showed clear differences in the contents and compositions of lipids between four types of nitrogen sources. Soymeal and ammonium tartrate supplementation favored the accumulation of triglycerides with arachidonic acid (ARA) and C16–18 fatty acids, respectively. These results were further validated by our targeted fatty acid analysis. RT-qPCR analysis of related genes in M. alpina between the four nitrogen source conditions found that soymeal supplementation dramatically increased the expression of GPAT, ELOVL, and Δ12/Δ6 desaturase. Our findings provided new insights into the regulation of lipid biosynthesis in M. alpina and potential avenues for genetic manipulation and highlighted the importance of an optimal nitrogen source for ARA-rich oil production.
研究领域:
生物化学
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微生物学
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分子生物学
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产品关联
DOI
PubMed
高影响因子
APPLIED MICROBIOLOGY AND BIOTECHNOLOGY
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2023
·Vol. 107
·期 18
·5761-5774
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IF 5
Li Jun,Chen Haiqin,Chang Lulu,Wu Chen,Zhang Hao,Chen Yong Q.,Chen Wei
Abstract Mortierella alpina produces various polyunsaturated fatty acids in the form of triacylglycerols (TAG). Diacylglycerol acyltransferase (DGAT) catalyzes the binding of acyl-CoA to diacylglycerol to form TAG and is the key enzyme involved in TAG synthesis. A variety of DGATs are present in M. alpina ; however, comparative analysis of the functional properties and substrate selectivity of these DGATs is insufficient. In this study, DGAT1 (MaDGAT1A/1B/1C) and DGAT2 (MaDGAT2A/2B) isoforms from M. alpina were analyzed and heterologously expressed in S. cerevisiae H1246. The results showed that MaDGAT1A/1B/2A/2B were able to restore TAG synthesis, and the corresponding TAG content in recombinant yeasts was 2.92 ± 0.42%, 3.62 ± 0.22%, 0.86 ± 0.34%, and 0.18 ± 0.09%, respectively. In S. cerevisiae H1246, MaDGAT1A preferred C16:1 among monounsaturated fatty acids, MaDGAT1B preferred C16:0 among saturated fatty acids (SFAs), and MaDGAT2A/2B preferred C18:0 among SFAs. Under exogenous addition of polyunsaturated fatty acids (PUFAs), MaDGAT1A and 2A preferentially assembled linoleic acid into TAG, and MaDGAT2B had substrate selectivity for eicosapentaenoic and linoleic acids in ω-6 PUFAs. In vitro, MaDGAT1A showed no obvious acyl-CoA selectivity and MaDGAT1B preferred C20:5-CoA. MaDGAT1A/1B preferred C18:1/C18:1-DAG compared with C20:4/C20:4-DAG. This study indicates that MaDGATs have the potential to be used in the production of LA/EPA-rich TAG and provide a reference for improving the production of TAGs in oleaginous fungi. Key points
• MaDGAT1A preferred C16:1 among MUFAs, MaDGAT1B and MaDGAT2A/2B preferred C16:0 and C18:0 among SFAs, respectively
• MaDGAT1A/2A preferentially assembled linoleic acid into TAG, and MaDGAT2B has substrate selectivity for eicosapentaenoic acid and linoleic acid in ω-6 PUFAs
• MaDGAT1A showed no obvious acyl-CoA selectivity, and MaDGAT1B preferred C20:5-CoA. MaDGAT1A/1B preferred to select C18:1/C18:1-DAG compared with C20:4/C20:4-DAG
研究领域:
生物化学
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微生物学
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脂质代谢
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产品关联
DOI
PubMed
高影响因子
Metabolomics
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2019
·Vol. 15
·期 4
·1-10
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IF 3.167
Lu Hengqian,Chen Haiqin,Tang Xin,Yang Qin,Zhang Hao,Chen Yong Q.,Chen Wei
Introduction 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. Objective 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 . Methods 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. Results 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 . Conclusion 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.
研究领域:
生物化学
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微生物学
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DOI
PubMed
高影响因子
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
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2020
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IF 4.192
Lulu Chang,Xin Tang,Hao Zhang,Yong Q. Chen,Haiqin Chen,Wei Chen
Sensing nutrient levels and coordinating metabolism are requisites for all living organisms. In eukaryotes, heterotrimeric adenosine monophosphate-activated protein kinase/sucrose nonfermenting 1 (SNF1) is an energy monitor that primarily functions by regulating cell metabolism with its γ-subunit being responsible for energy sensing. Because of its strong lipogenesis capacity and dependence on nutrient availability, Mortierella alpina is an ideal model to investigate the SNF1 role. Knockdown of the M. alpina SNF1-γ-subunit (MaSnf4) abolished the energy preservation mode. In a low glucose medium (15 g/L), the fatty acid content in the MaSnf4-knockdown strain was similar to that in a high glucose medium (50 g/L), comprising 16 ± 1.17% of the dry cell weight after 96 h of culture (1.59 g/L), together with 1.41 ± 0.13 and 4.15 ± 0.19 fold increased acetyl-CoA carboxylase 1 and ATP-citrate lyase enzymatic activities, respectively. Metabolite analysis confirmed that knocking down MaSnf4 enhanced amino acid recycling and repressed the tricarboxylic acid cycle. In this case, more carbon skeleton acetyl-CoA and reductive nicotinamide adenine dinucleotide phosphate were rerouted into the fatty acid synthesis pathway. These findings provide new insight into the correlation between energy preservation and MaSnf4-regulated lipogenesis, which may enhance further development of cost-effective strategies to enhance lipid productivity in M. alpina.
研究领域:
微生物学
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代谢工程
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生物技术
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DOI
PubMed
高影响因子
ACS Synthetic Biology
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2023
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IF 4.7
Lulu Chang,Hanqin Chen,Bo Yang,Haiqin Chen,Wei Chen
Increasing carbon flux toward target metabolites is important in improving microbial productivity and economic value. To improve the efficiency of lipid production in Mortierella alpina, we knocked down genes for trehalose-6-phosphate synthetase (Matps) and phosphoenolpyruvate carboxykinase (Mapepck) in the major pathways for saccharide synthesis. The knockdown of Matps reduced trehalose content by an average of 31.87%, while the knockdown of Mapepck reduced the total saccharide content by 28.6%, and both recombinant strains showed more than 20% increased lipid yield. Trehalose plays a vital role in stress resistance, but a higher polyunsaturated fatty acid-rich lipid content was found to partly compensate for the loss of trehalose after Matps knockdown. As compared with Matps knockdown, the knockdown of Mapepck gave better lipid production by bringing forward the time to maximum lipid yield by three days in a scale-up test. The arachidonic acid yield after the Mapepck knockdown reached 1.23 g/L, which was 39.9% higher than that of the original strain. The present research provided an efficient strategy for redistributing carbon flux among different metabolites and therefore promoted microbial lipid yield in a shorter fermentation period.
研究领域:
微生物学
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生物技术
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代谢工程
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DOI
PubMed
高影响因子
Systems Microbiology and Biomanufacturing
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2024
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IF 0
Feng Yuanbang,Zhang Xin,Yang Hua,Feng Xue,Chen Ming
To achieve high production of arachidonic acid (ARA) through industrial fermentation of Mortierella alpina , the correlation between metal ions, mycelium pellet morphology and ARA production was investigated. By adding different metal ions to the culture medium, observing the morphological differences of mycelium pellet, total oil production and ARA content and established a correlation between them. The results showed that adding sodium ions increase the density of mycelium pellet, while reducing the diameter of mycelium pellet, resulting in a 41.59% increase in ARA production. Potassium ions, magnesium ions, calcium ions and iron ions slightly reduce the diameter of fungal hyphae and increase ARA production by 7.47%, 17.81%, 36.05% and 9.37%, respectively. At the same time, calcium ions promoted the branching growth of mycelium, which solve the problem of excessive free mycelium in fermentation broth within a certain concentration range. Zinc ions had a negative impact on bacterial growth, leading to the formation of mycelium clumps in the fermentation broth. Manganese ions and ferrous ions could transform the morphology of fungal mycelium into a smooth spherical surface, which was not conducive to the growth of M . alpina and the accumulation of oil. The comprehensive results showed that the optimal fermentation mycelium form for M . alpina was a dense central and loosely packed mycelium pellet. The yield of ARA was directly proportional to the density of mycelium pellet and inversely proportional to the diameter of mycelium pellet.
研究领域:
微生物学
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生物技术
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发酵工程
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脂类代谢
✓ 已验证
DOI
危险品化学品经营许可证（带存储）