Oleaginous Fungus Research Articles

Rare fungus, Mortierella capitata, promotes crop growth by stimulating primary metabolisms related genes and reshaping rhizosphere bacterial community

Rhizosphere and endosphere microorganisms are recognized as an extended plant genome. Changes in rhizosphere and endosphere microbial community structure are closely linked to plant nutrition uptake, development and immunity. Mortierella is a saprophytic oleaginous fungus, with various agricultural benefits, which have triggered interest in recent years. Here, we conducted Mortierella capitata inoculation experiments in a climate chamber. M. capitata inoculation increased biomass, chlorophyll and gibberellic acid content in maize. The concentration of available soil phosphorus was also increased, while O2 concentration and soil density decreased. Further, the prokaryotic and eukaryotic microbial structure was evaluated by MiSeq sequencing. M. capitata inoculants increased rhizosphere bacterial diversity and significantly altered rhizosphere bacterial community composition. Yet, the influence of M. capitata on fungal community structure was not significant according to ANOSIM analysis. RNA-seq was used to ascertain differentially expressed genes (DEGs) in maize roots following M. capitata inoculation. Genes encoding chitinase, peroxidase, lipid transfer protein, sugars exported transporter, and various transcription factor proteins were significantly up-regulated. All the DEGs, soil and plant basic properties were used to construct a co-occurrence network for modularity analysis. The whole network was divided into five modules based on Louvain algorithm, including organic substances metabolism and hormone regulation, nucleobases metabolism, stimulus response, primary metabolism and an unknown module. The stimulus response module showed the closest correlation with biomass, soil oxygen, and M. capitata, while genes in the primary metabolism module were closely associated with plant gibberellic acid and available soil phosphorus. Our results highlight that the rare fungus M. capitata can promote crop growth directly by altering the root gene expression levels, and indirectly via interaction with indigenous rhizosphere bacteria.

FAMEs Profile of Oil Produced by Oleaginous Fungi Isolated from Fermented Beverage Wastewaters and Soil

Fungal strains isolated from fermented maize (ogi) (PW) and sorghum-based brewery wastewaters (BW) and two soil isolates were evaluated for oleaginicity. The fungal isolates from the wastewater that had lipid content of at least 20% of their biomass were identified by both culture methods and internal transcribed spacer (ITS) 1-5.8S-ITS2 ribosomal DNA sequencing. The isolates were identified to be Aspergillus fumigatus (PW8), Aspergillus flavus (PW10), Candida tropicalis (PW16) and Aspergillus tubingensis (PW3), Trichosporon luoberi (BW7), Aspergillus sp. (BW4) and Candida tropicalis (BW1; BW3). FAMEs composition was determined for the four strains with the highest lipid content by acid-catalyzed transesterification and analyzed by Gas Chromatography-Flame Ionization Detector (GC-FID). Palmitoleic acid was the dominant fatty acid in M. circinelloides and T. reesei, and the best producers of capric and lauric acids were Aspergillus fumigatus and Aspergillus sp. (BW4), respectively. These fatty acids are beneficial in making cosmetics and pharmaceuticals (antimicrobials and dietary supplements). The analysis of the FAMEs profile in the species indicated low amounts or absence of some key long chain fatty acid (LCFA) constituents of biodiesels. Based on the FAMEs profile of M. circinelloides investigated, this strain could hold promise for use as feedstock for biodiesel with genetic engineering and a tailored lipid production favouring enrichment of LCFA.
 Keywords: Fungal lipids, wastewater, fatty acid methyl ester, GC-FID

Enhancement of lipid productivity from a promising oleaginous fungus <em>Aspergillus</em> sp. strain EM2018 for biodiesel production: Optimization of culture conditions and identification

Oleaginous fungi have recently gained increasing attention among different microorganisms due to their ability for lipid production for the preparation of biofuel. In the present study, a locally isolated fungus E45, identified genetically as Aspergillus sp. strain EM2018, was found to produce 25.2% of the total lipids content of its dry cell weight (DCW). Optimization of culture conditions was performed and lipid accumula­tion increased by about 2.4 fold (from 25.2% to 60.1% of DCW) when the fungus was grown for seven days in the potato dextrose (50 g/L) liquid medium at pH 5.0, incubation temperature at 30 ºC and inoculum size of 2 × 106 spore/mL. Supplementation of the medium with yeast extract and NaNO3 at a concentration of 0.05% as organic and inorganic nitrogen sources, respectively, increased lipid production (53.3% lipid/dry biomass). Gas chromatography analysis of fungal lipids revealed the presence of saturated (mainly palmitic acid C16:0 (33%) and lignoceric acid C24:0 (15%)) and unsaturated fatty acids in different proportions (mainly linoleic acid C18:2 (24.4%), oleica cid C18:1 (14%) and arachidonic C20:4 (7.4%). These findings suggest this new oleaginous fungus as a promising feedstock for various industrial applications and for the preparation of biodiesel.

Consolidated bioprocesses for efficient bioconversion of palm biomass wastes into biodiesel feedstocks by oleaginous fungi and yeasts

Consolidated bioprocesses for bioconversion of lignocellulosic biomass into biodiesel feedstocks were developed. Palm empty fruit bunch (EFB) was biologically pretreated coupling with fungal lipid production (121.4 ± 2.7 mg/g-EFB) by lignocellulolytic oleaginous fungi prior to lipid production by oleaginous yeasts. In subsequent separate hydrolysis and fermentation (SHF) of fungal pretreated EFB (FPEFB), the oleaginous yeast with the maximum lipid yield of 37.0 ± 0.1 mg/g-FPEFB was screened. While a higher lipid yield of 47.9 ± 1.5 mg/g-FPEFB was achieved in simultaneous saccharification and fermentation (SSF) with less enzyme requirement. Fed-batch SSF of non-sterile FPEFB was proven as a practical and efficient strategy to increase lipid yield up to 53.4 ± 0.5 mg/g-FPEFB. Total lipid yield by both fungi and yeast was 165.0 ± 4.4 mg/g-EFB. Interestingly, the consolidated bioprocesses of enzyme and lipid production also achieved comparable total lipid yield of 149.3 ± 6.6 mg/g-EFB. These strategies may contribute greatly to cost-effective and sustainable bioconversion of lignocellulosic biomass into biodiesel feedstocks.

Time-resolved multi-omics analysis reveals the role of nutrient stress-induced resource reallocation for TAG accumulation in oleaginous fungus Mortierella alpina

BackgroundGlobal resource reallocation is an established critical strategy through which organisms deal with environmental stress. The regulation of intracellular lipid storage or utilization is one of the most important strategies for maintaining energy homeostasis and optimizing growth. Oleaginous microorganisms respond to nitrogen deprivation by inducing lipid hyper accumulation; however, the associations between resource allocation and lipid accumulation are poorly understood.ResultsHere, the time-resolved metabolomics, lipidomics, and proteomics data were generated in response to nutrient availability to examine how metabolic alternations induced by nitrogen deprivation drive the triacylglycerols (TAG) accumulation in M. alpina. The subsequent accumulation of TAG under nitrogen deprivation was a consequence of the reallocation of carbon, nitrogen sources, and lipids, rather than an up-regulation of TAG biosynthesis genes. On one hand, nitrogen deprivation induced the down-regulation of isocitrate dehydrogenase level in TCA cycle and redirected glycolytic flux of carbon from amino acid biosynthesis into fatty acids’ synthesis; on the other hand, nitrogen deprivation induced the up-regulation of cell autophagy and ubiquitin-mediated protein proteolysis which resulted in a recycling of preformed protein nitrogen and carbon. Combining with the up-regulation of glutamate decarboxylase and succinic semialdehyde dehydrogenase in GABA shunt, and the phosphoenolpyruvate carboxykinase in the central hub involving pyruvate/phosphoenolpyruvate/oxaloacetate, the products from nitrogen-containing compounds degradation were recycled to be intermediates of TCA cycle and be shunted toward de novo biosynthesis of fatty acids. We found that nitrogen deprivation increased the protein level of phospholipase C/D that contributes to degradation of phosphatidylcholine and phosphatidylethanolamine, and supplied acyl chains for TAG biosynthesis pathway. In addition, ATP from substrate phosphorylation was presumed to be a critical factor regulation of the global resource allocation and fatty acids’ synthesis rate.ConclusionsThe present findings offer a panoramic view of resource allocation by M. alpina in response to nutrient stress and revealed a set of intriguing associations between resource reallocation and TAG accumulation. This system-level insight provides a rich resource with which to explore in-depth functional characterization and gain information about the strategic combination of strain development and process integration to achieve optimal lipid productivity under nutrient stress.

Sugarcane by-products within the biodiesel production chain: Vinasse and molasses as feedstock for oleaginous fungi and conversion to ethyl esters

Oleaginous microorganisms pose a number of technical advantages when compared to first generation oil crops, but a common drawback to their utilization is often linked to the high operational costs involved in media preparation, using mostly industrial-grade sugars as carbon sources. Considering the abundance of by-products generated by the sugarcane industry, the combined use of molasses and vinasse is suggested in this work as culture medium for the fungal strain Mucor circinelloides URM 4182. Results demonstrate that a 3:1 vol% medium (vinasse to diluted molasses ratio) with minor supplementation of organic nitrogen can provide values within the range of 5 g L−1 of oil-rich biomass with 25 wt% of lipids. The microbial oil was then analyzed in terms of fatty acid profile, demonstrating a favorable composition for biodiesel production, and was subjected to simultaneous esterification and transesterification assays using ethanol and heterogeneous catalyst (H3PMo/Al2O3) in a pressurized reactor, demonstrating satisfactory conversions (>95%). The purified ethyl esters possessed viscosity and density values within the regulating standards and lack of triacylglycerols, suggesting a potential biodiesel production system that could be coupled within sugarcane ethanol facilities.

Lowering the inhibition of sugarcane vinasse as a culture medium for oleaginous fungi through oxidative pre‐treatment aiming at the degradation of toxic compounds

AbstractBACKGROUNDThe utilization of vinasse as a feasible culture medium for fungi is hindered by the presence of inhibitory compounds, such as phenols and other recalcitrant structures. This work analyzes the effect of different homogeneous advanced oxidation processes (AOP) to lower the concentration of inhibitory phenolic compounds in vinasse, and to test the effects of the pre‐treatment on the growth of an oleaginous fungal strain. Vinasse was treated using a batch system with the addition of ferrous ion (Fe2+) (Fenton), in the presence of UV, and with the addition of hydrogen peroxide (H2O2).RESULTSAll of the reaction systems were able to partially degrade the chemical oxygen demand, ranging from 18.4% up to 54.1% in the factorial results. The pre‐treated vinasse samples were assayed for the growth performance of Mucor circinelloides, demonstrating that the control value of 3.54 g L−1 was increased to values as high as 4.84 g L−1, also followed by an increase of ≈73% in terms of lipid productivity.CONCLUSIONDifferent AOP were tested to degrade phenolic compounds present in vinasse, demonstrating that the optimum condition (Fe2+/UV at alkaline pH) provided a culture medium for M. circinelloides that increased the biomass accumulation by ≈35% and the lipid productivity by >70% when compared to the control. © 2020 Society of Chemical Industry

The role of MTHFDL in mediating intracellular lipogenesis in oleaginous Mortierella alpina.

The oleaginous fungus Mortierella alpina can synthesize a variety of polyunsaturated fatty acids, which are used extensively in industry for the production of arachidonic acid (AA). NADPH is the limiting factor and critical reducing agent in lipid biosynthesis. In the folate cycle, methylenetetrahydrofolate dehydrogenase (MTHFDL) catalyzes the conversion of methylene tetrahydrofolate into 10-formyl-tetrahydrofolate with the reduction of NADP+ to NADPH. MTHFDL RNAi was used to investigate the role of the folate cycle in lipogenesis. Gene knockdown decreased the transcript levels of MTHFDL by about 50 % and attenuated cell fatty acid synthesis. The observation of decreased NADPH levels and downregulated NADPH-producing genes in response to MTHFDL RNAi indicates a novel aspect of the NADPH regulatory mechanism. Thus, our study demonstrates that MTHFDL plays key role in the mediation of NADPH in lipogenesis in M. alpina.

Production of Value-Added Microbial Metabolites: Oleaginous Fungus as a Tool for Valorization of Dairy By-products

Mucor circinelloides is a filamentous fungus capable of producing mycelium-bound lipase and lipids and able to assimilate a wide array of substrates, like agricultural wastes and industrial effluents. A biotechnological valorization of cheese whey is described herein, in which simultaneous reduction of the organic loading and conversion of the various compounds of the by-product into added-value microbial metabolites is obtained. M. circinelloides URM 4182 was tested for the ability of producing biomass containing lipids and mycelium-bound lipase using raw cheese whey as culture medium. The fungal cultivation assays were performed in non-supplemented cheese whey at pH 4.5, 26 °C, 250 rpm for 120 h providing biomass concentration of 4.7 g L−1 containing lipids (22.5% of dry biomass weight) and lipolytic activities up to 160 U g−1. The mycelium-bound lipase was characterized in terms of its biochemical and kinetic parameters and tested on biogas production assays of cheese whey as a way to enhance the lipid hydrolysis, the limiting step in anaerobic digestion. Results demonstrate that the addition of M. circinelloides lipase on cheese whey reduced the lag phase of anaerobic digestion up to 43.7%, and the biogas production was increased by up to ninefold when compared with the control experiments.

Tetrahydrobiopterin Plays a Functionally Significant Role in Lipogenesis in the Oleaginous Fungus Mortierella alpina.

Tetrahydrobiopterin (BH4) is well-known as a cofactor of phenylalanine hydroxylase (PAH) and nitric oxide synthase (NOS), but its exact role in lipogenesis is unclear. In this study, the GTP cyclohydrolase I (GTPCH) gene was overexpressed to investigate the role of BH4 in lipogenesis in oleaginous fungus Mortierella alpina. Transcriptome data analysis reveal that GTPCH expression was upregulated when nitrogen was exhausted, resulting in lipid accumulation. Significant changes were also found in the fatty acid profile of M. alpina grown on medium that contained a GTPCH inhibitor relative to that of M. alpina grown on medium that lacked the inhibitor. GTPCH overexpression in M. alpina (the MA-GTPCH strain) led to a sevenfold increase in BH4 levels and enhanced cell fatty acid synthesis and poly-unsaturation. Increased levels of nicotinamide adenine dinucleotide phosphate (NADPH) and upregulated expression of NADPH-producing genes in response to enhanced BH4 levels were also observed, which indicate a novel aspect of the NADPH regulatory mechanism. Increased BH4 levels also enhanced phenylalanine hydroxylation and nitric oxide synthesis, and the addition of an NOS or a PAH inhibitor in the MA-GTPCH and control strain cultures decreased fatty acid accumulation, NADPH production, and the transcript levels of NADPH-producing genes. Our research suggests an important role of BH4 in lipogenesis and that the phenylalanine catabolism and arginine–nitric oxide pathways play an integrating role in translating the effects of BH4 on lipogenesis by regulating the cellular NADPH pool. Thus, our findings provide novel insights into the mechanisms of efficient lipid biosynthesis regulation in oleaginous microorganisms and lay a foundation for the genetic engineering of these organisms to optimize their dietary fat yield.

Evaluation of the Oleaginous Rhizopus stolonifer capability for Lipid Production Using Agricultural Wastes as cheap nutritional sources for Biodiesel production.

Depletion of fossil fuels and increasing the environmental concerns against pollution accelerate the research in renewable energy with much efforts directed for the production of biodiesel by the promising oil-rich biomass, like oleaginous fungi, growing on inexpensive agricultural and industrial wastes. Rhizopus stolonifer was found to be the most potent lipid producer among the tested fungi in the present study; its growth was optimized at pH=6, glucose as C-source, NaNO3 as N-source, with agitation rate of 120 rpm, incubation temperature of 28 C after 6 days growth; which gave an observable lipid content of 57.9% dry wt. on non-detoxified liquid hydrolysate (NDLH) medium with saturated aliphatic long chains as revealed by GC-MS analysis. R. stolonifer gave noticeable activity of released sugar consumption (92.2%) on flax straw as a row material higher than rice, wheat straws and orange peel, forming high lipid content (54.33%) on flax; that was transesterified into biodiesel with cetane number of 61.48, that was higher than cetane number of the standard non-processed liquid diesel (< 47). So R. stolonifer can be considered as a good lipid producer on the depolymerized flax straw as a cheap source for biodiesel production.

Valorization of palm biomass wastes for biodiesel feedstock and clean solid biofuel through non-sterile repeated solid-state fermentation

Palm biomass wastes are currently considered as promising solid biofuels. However, their high potassium content leads to formation of slag in combustion chambers and causes frequent power-plant shutdowns for maintenance. Therefore, this study aimed to develop a low-cost practical biological pretreatment for these wastes. Oleaginous fungi Aspergillus tubingensis TSIP9, which originates from palm wastes, was used to pretreat biomass wastes and simultaneously produce oils through non-sterile solid state fermentation (SoSF). The operating conditions were optimized through response surface methodology. The fungi could grow and produce oils with good biodiesel fuel properties. After SoSF, potassium content in biomass wastes was reduced by 90% and cellulose content increased to >57%, making it suitable as clean solid biofuel. Repeated-SoSF with 90% substrate replacement was highly effective in continuously pretreating biomass wastes and producing fungal oils. This study demonstrates the cost-effective and environmentally friendly process for production of clean renewable energy through zero-waste strategy.

Novel Dual-Functional Enzyme Lip10 Catalyzes Lipase and Acyltransferase Activities in the Oleaginous Fungus Mucor circinelloides.

Lipases or triacylglycerol (TAG) lipases belong to the α/β-hydrolases superfamily, which are enzymes capable of catalyzing the hydrolysis of the ester bond between fatty acids and glycerol. Interestingly, some lipases have been found to not only possess hydrolysis activity but also acyltransferase activity in yeasts and microalgae. Our present study reported a novel dual-functional Mucor circinelloides lipase Lip10 with a slight lipolysis activity but a noteworthy phospholipid/diacylglycerol acyltransferase (PDAT) activity. The purified Lip10 mutants prefer to utilize phosphatidyl serine to form TAG over phosphatidyl ethanolamine and phosphatidylcholine. Site-directed mutagenesis indicated that the histidine residue in the acyltransferase motif H-(X)4-D is indispensable for the PDAT activity of Lip10. Overexpression of the acyltransferase motif of Lip10 promoted cell growth by 12% and increased lipid production by 14% compared to the control, whilst overexpression of the lipase motif induced lipid degradation in M. circinelloides.

Eco-Green Conversion of Watermelon Peels to Single Cell Oils Using a Unique Oleaginous Fungus: Lichtheimia corymbifera AH13

Watermelon peel waste (WPW) is being used for first time in single cell oils (SCOs) production via a promising oleaginous fungus Lichtheimia corymbifera which was isolated from Egyptian ecosystem. Pretreatments of WPW were carried out by mechanical, physical and chemical methods; the most potent pretreatment was selected according to total reducing sugar and total lipid production. Accordingly, the mechanical ptetreatment of WPW was distinctly the best pretreatment method for SCOs production from L. corymbifera. Taguchi design clarified that the most optimal culture conditions were 35 °C and pH 7.0 for 4 days by which the highest potential of SCOs and lipid content was yielded (2.93 gl−1 and 39.56% respectively). Mechanical treatment revealed that the dominant fatty acid was palmitic and oleic acids with 41.98 and 34.65% respectively with appearance of γ linolenic acid (GLA) at low concentration 1.43%. Finally, this study showed that WPW was used as a natural, effective, economic, ecofriendly and integrated substrate without adding any outsource nutrients to produce sustainable SCOs with low cost production.

Production of polyunsaturated fatty acids from animal fat emulsion by oleaginous filamentous fungi

Production of polyunsaturated fatty acids from animal fat emulsion by oleaginous filamentous fungi

Effect of phosphorus source on the lipid accumulation in oleaginous fungi grown under nitrogen limitation

Effect of phosphorus source on the lipid accumulation in oleaginous fungi grown under nitrogen limitation

Ultra Performance Liquid Chromatography–Q Exactive Orbitrap/Mass Spectrometry-Based Lipidomics Reveals the Influence of Nitrogen Sources on Lipid Biosynthesis of Mortierella alpina

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.

13C metabolic flux analysis on roles of malate transporter in lipid accumulation of Mucor circinelloides

BackgroundMitochondrial and cytoplasmic malate transporter proteins are responsible for transmembrane transport of malate, thereby linking malate metabolism in various subcellular regions of the cell. These transporters play an important role in fatty acid biosynthesis of oleaginous microorganisms. Our previous studies have found that lipid content of the recombinant Mucor circinelloides (M. circinelloides) strain with mitochondrial malate transporter (mt) gene overexpression was increased by 70%, while that of strain with mt gene knockout was decreased by 27%. However, the mechanism of malate transporter promoting the transport of mitochondrial malate and citrate related to lipid accumulation is not clear. Therefore, 13C-labeled glucose metabolic flux analysis was carried out to identify the metabolic network topology and estimate intracellular fluxes of genetically engineered M. circinelloides strains for the purpose of better understanding the roles of malate transporters in citrate transport systems and lipid accumulation.ResultsThe metabolic flux distribution analysis suggested that tricarboxylic acid (TCA) cycle flux ratio of mt-overexpression strains was decreased compared to that of the control strain, but in contrast, glyoxylic acid (GOX) cycle flux ratio was increased. Accordingly, the mt-knockout strain showed an opposite phenomenon with a higher TCA cycle flux ratio and a lower GOX cycle flux ratio than the control strain. GOX cycle might be more effective than TCA cycle in producing malate and oxaloacetate replenishment. Moreover, a relatively higher flux ratio of the pentose phosphate (PP) pathway was obtained in mt-overexpression strains, but no significant difference in the malic enzyme flux between recombinant strains and the control strain. Our results confirmed that PP pathway might play an important role for supplying NADPH and malic enzyme is not a limiting factor for fatty acid synthesis in oleaginous fungus M. circinelloides strains.ConclusionIntracellular metabolic flux information suggested that mt-overexpression strains had higher flux in PP pathway and GOX cycle, lower flux in TCA cycle, and no difference in malic enzyme cycle. Together, the role of malate transporter was assumed to further participate in transporting cycle of acetyl-CoA and drive PP pathway to supply NADPH required for lipid accumulation in recombinant M. circinelloides strains.

Morphologically engineered strain of Aspergillus oryzae as a cell chassis for production development of functional lipids

Cell morphology of the oleaginous fungus, Aspergillus oryzae BCC7051, was genetically engineered by disruption of non-essential genes involved in cell wall biosynthesis. Comparative phenotypic analysis of two disruptant strains defective either in α-1,3-glucan synthase 1 (ΔAoAgs1) or chitin synthase B (ΔAoChsB), and the wild type showed that the ΔAoAgs1 strain had no alterations in colonial growth and sporulation when grown on agar medium whereas the ΔAoChsB disruptant showed growth retardation and lower sporulation. However, tiny and loose pellets were found in the ΔAoAgs1 culture grown in liquid medium, where fungal pellet size was decreased by 35–50% of the wild type size. Further investigation of the ΔAoAgs1 mutant grown under stress-induced conditions, including high salt concentration, ionic strength and osmolarity, showed that its growth and development remained similar to that of the wild type. When cultivating the ΔAoAgs1 strain in a stirred-tank bioreactor, lipid production in terms of titer and productivity was significantly improved. As compared to the wild type, an increase of triacylglycerol and ergosterol contents with a proportional decrease in steryl ester content was observed in the ΔAoAgs1 strain. These results suggest that the morphologically engineered strain of A. oryzae is a robust cell chassis useful for exploitation in further production development of functional lipids with industrial significance.

Role of Adenosine Monophosphate Deaminase during Fatty Acid Accumulation in Oleaginous Fungus Mortierella alpina.

In oleaginous micro-organisms, nitrogen limitation activates adenosine monophosphate deaminase (AMPD) and promotes lipogenesis via the inhibition of isocitrate dehydrogenase. We found that the overexpression of homologous AMPD in Mortierella alpina favored lipid synthesis over cell growth. Total fatty acid content in the recombinant strain was 15.0-34.3% higher than that in the control, even though their biomass was similar. During the early fermentation stage, the intracellular AMP level reduced by 40-60%, together with a 1.9-2.7-fold increase in citrate content compared with the control, therefore provided more precursors for fatty acid synthesis. Moreover, the decreased AMP level resulted in metabolic reprogramming, reflected by the blocked TCA cycle and reduction of amino acids, distributing more carbon to lipid synthesis pathways. By coupling the energy balance with lipogenesis, this study provides new insights into cell metabolism under nitrogen-limited conditions and targets the regulation of fatty acid accumulation in oleaginous micro-organisms.