Genome-scale analysis of the metabolic networks of oleaginous Zygomycete fungi Wanwipa Vongsangnak a, ⁎, Rawisara Ruenwai b , Xin Tang c , Xinjie Hu c , Hao Zhang c , Bairong Shen a , Yuanda Song c , Kobkul Laoteng d, ⁎⁎ a Center for Systems Biology, Soochow University, Suzhou 215006, China b School of Agriculture and Natural Resources, University of Phayao, Maeka, Muang, Phayao 56000, Thailand c State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi, Jiangsu, China d Bioresources Technology Unit, National Center for Genetic Engineering and Biotechnology, National Sciences and Technology Development Agency, Khong Luang, Pathum Thani 12120, Thailand abstract article info Article history: Accepted 7 March 2013 Available online 27 March 2013 Keywords: Gene annotation Oleaginous fungi Lipid Metabolic function Genome-scale metabolic network Microbial lipids are becoming an attractive option for the industrial production of foods and oleochemicals. To investigate the lipid physiology of the oleaginous microorganisms, at the system level, genome-scale metabolic networks of Mortierella alpina and Mucor circinelloides were constructed using bioinformatics and systems biol- ogy. As scaffolds for integrated data analysis focusing on lipid production, consensus metabolic routes governing fatty acid synthesis, and lipid storage and mobilisation were identified by comparative analysis of developed metabolic networks. Unique metabolic features were identified in individual fungi, particularly in NADPH metab- olism and sterol biosynthesis, which might be related to differences in fungal lipid phenotypes. The frameworks detailing the metabolic relationship between M. alpina and M. circinelloides generated in this study is useful for further elucidation of the microbial oleaginicity, which might lead to the production improvement of microbial oils as alternative feedstocks for oleochemical industry. © 2013 Elsevier B.V. All rights reserved. 1. Introduction In living cells, lipid metabolism is associated with various metabolic pathways at different levels of regulation. For a large number of catalysed metabolic reactions involving enzymes, it is generally difficult to identify key metabolic and regulatory modules contributing to lipid physiology (Nielsen, 2009). Many efforts have been devoted to identify biochemical events involved in lipogenesis in the so-called oleaginous microorganisms, which accumulate lipids equal to at least 20% of their biomass (Ageitos et al., 2011; Ratledge, 2004). This biochemical knowl- edge is certainly important for the channelling of carbon flux to lipid biosynthesis by carbon regulation under limited nitrogen conditions as well as for enhancing lipid production by process optimisation of in- dustrial fermentation (Ratledge and Wynn, 2002). Although microbial lipids or single cell oils have been considered as a potential feedstock for oleochemicals and the biodiesel industry, applications are realisti- cally constrained by the criteria of production yield and cost. Further- more, relatively little information is known regarding the overall metabolic process inherent to lipid metabolism and the genetic back- grounds of oleaginous strains. Very recently, the orthologous sequences of oleaginous yeast and fungi were identified by comparative genome analysis, showing that orthologous sequences are distributed across several metabolic cellular processes. Research has also exhibited a rela- tionship among carbohydrate, lipid and amino acid metabolisms in the biosynthesis of acetyl-CoA, which is the initial precursor necessary for fatty acid biosynthesis (Vorapreeda et al., 2012). Furthermore, other metabolic processes crucial to oleaginicity exist, such as lipid storage and mobilisation as well as acetyl-CoA synthesis and amino acid metab- olism (Vorapreeda et al., 2012). Discrimination in the content of intra- cellular lipids produced by oleaginous microorganisms, which depends on strain and culture conditions, remains somewhat unclear. Indeed, addressing how oleaginous microorganisms maintain lipid ho- meostasis under particular conditions poses a research challenge. Of the oleaginous strains, the Zygomycetes fungi, such as Mortierella spp. and Mucor spp., have been considered as important oleaginous model organisms of industrial interest due to their ability Gene 521 (2013) 180–190 Abbreviations: ACP, acyl carrier protein; ACOAA, acetyl-CoA acyltransferase; ACC, acetyl-CoA carboxylase; AOX, acyl-CoA oxidase; ARA, arachidonic acid; ACLY, ATP-citrate lyase; CoA, Coenzyme A; DGAT, diacylglycerol O-acyltransferase; DGLA, dihomo γ-linolenic acid; DHAPAT, dihydroxyacetone phosphate acyltransferase; FADS3, ω3-fatty acid desaturase; FADS5, Δ 5 -fatty acid desaturase; FMN, flavin mononucleotide; EPA, eicosapentaenoic acid; FAS, fatty acid synthase; G6PD, glucose-6-phospate 1-dehydrogenase; GLA, γ-linolenic acid; GLUD, glutamate dehydrogenase; GPAT, glycerol-3-phosphate acyltransferase; GPR, gene-protein-reaction; ICDH, isocitrate dehydro- genase; LPA, lysophosphatidic acid; LPC, lysophosphatidylcholine; LPAAT, lysophosphatidic acid acyltransferase; MBOP, long-chain-enoyl-CoA hydratase; ME, malic enzyme; MCD, malonyl-CoA decarboxylase; NADPH, nicotinamide-adenine dinucleotide phosphate; PA, phosphatidic acid; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PG, phosphatidylglycerol; PS, phosphatidylserine; PI, phosphatidylinositol; PDAT, phospholipid: diacylglycerol acyltransferase; PDH, pyruvate dehydrogenase; PPAP, phosphatidate phos- phatase; SE, sterol esterase; SI, sterol isomerase; SR, sterol reductase; TAG, triacylglycerol; TCA, tricarboxylic acid; TGL, triglyceride lipase; zHAD, 3-hydroxyacyl-CoA dehydrogenae. ⁎ Corresponding author. Tel.: +86 512 65222093; fax: +86 512 65110951. ⁎⁎ Corresponding author. Tel.: +66 2 5646700; fax: +66 2 5646707. E-mail addresses: wanwipa@suda.edu.cn (W. Vongsangnak), kobkul@biotec.or.th (K. Laoteng). 0378-1119/$ – see front matter © 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.gene.2013.03.012 Contents lists available at SciVerse ScienceDirect Gene journal homepage: www.elsevier.com/locate/gene