Bioinformatical analysis and preliminary study of the role of lipase in lipid metabolism in Mucor circinelloides

Abstract

The filamentous fungus Mucor circinelloides has been widely used as a model organism to investigate the mechanisms of lipid accumulation. Although a lot of work has been done to analyze and explore many of the essential enzymes/genes related to lipid accumulation in M. circinelloides, the function of lipase in this fungus has not been studied at all. In this study, we report some important characteristics of all 30 lipases, and especially 4 lipases Lip6, Lip10, Lip19 and Lip24 with functional analyses in Mucor circinelloides CBS 277.49 based on the genome sequences. Transcriptional analyses revealed that the Lip6 or Lip10 gene expression increased significantly during 24 h to 72 h of lipid accumulation while Lip19 and Lip24 genes were down-regulated along with the increase in lipid accumulation. Over-expression of either lipase Lip6 or Lip10 led to a slight increase in cell dry weight, but no significant effect on lipid accumulation.More interestingly over-expression of Lip6 resulted in a 17% increase in C18:3 content of the lipids.However, over-expression of lipase Lip19 and Lip24 decreased the cell dry weight by 9.08-18.8% and lipid content by 37.86-41.42%, respectively. The fatty acid profiles of strains with Lip19 and Lip24 overexpression were also significantly changed as compared to control. Analysis of lipase genes revealed that the sequences of Lip6 and Lip10 not only contained the typical lipase motif (G/A) XSXG, but also the consensus sequence motif HXXXXD (a typical of acyltransferase motif). These results suggest that Lip6 or Lip10 may control acyltransferase activity in Mucor circienlloides and play a role in fatty acid reconstruction of TAG, while Lip19 or Lip24 may work as a TAG lipase involved in TAG metabolism/ degradation. † Electronic supplementary information (ESI) available: Table S1 sequence of primers used in quantitative RT-PCR. Table S2 sequence of primers used in gene over-expression of Lip6 (161426), Lip10 (115761), Lip19 (143450) and Lip24 (107413). Table S3 the sub-cellular location, and the presence and location of signal peptide of lipases from M. circinelloides CBS 277.49. See

Extract of Syzygium aromaticum suppress eEF1A protein expression and fungal growth

“…Total RNA of S. cerevisiae was extracted using TRIzol reagent after grinding under liquid N 2 (Zan et al . ) and reverse‐transcribed using the Prime Script RT reagent kit (Takara) according to the manufacturer's instructions. About 2 μ g of total RNA was reverse‐transcribed to cDNA by PrimeScript1st strand cDNA Synthesis Kit (Takara).…”

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

“…Media and growth conditions for M. circinelloides have been previously described in our previous study. 29,30,32 Lipase genes were overexpressed in the M. circinelloides uridine auxotroph pleu-MU402. 33 This strain could grow at 30 °C in YPG media containing 3 g/L yeast extract, 10 g/L tryptone, 20 g/L glucose, and 200 μg/mL uridine at pH 4.5.…”

“…However, the model of Cdk1/Cdc28-dependent activation of the lipase activity of Tgl4p is a good reference to study the molecular mechanism balancing catabolic and anabolic activities of multifunctional lipases. Previous studies from our group showed that the lipidproducing fungus M. circinelloides lipases Lip6 and Lip10 harbored both a typical lipase catalytic site motif GXSXG and an acyltransferase signature motif H-(X) 4 -D. 29,30 Overexpression of either lipase Lip6 or Lip10 promoted cell growth and increased lipid production. 30 These results suggested that Lip6 or Lip10 may function both as an acyltransferase and a lipase in M. circinelloides.…”

“…Previous studies from our group showed that the lipidproducing fungus M. circinelloides lipases Lip6 and Lip10 harbored both a typical lipase catalytic site motif GXSXG and an acyltransferase signature motif H-(X) 4 -D. 29,30 Overexpression of either lipase Lip6 or Lip10 promoted cell growth and increased lipid production. 30 These results suggested that Lip6 or Lip10 may function both as an acyltransferase and a lipase in M. circinelloides. Further analysis proved that Lip6 has a low lipolysis activity but a higher phospholipid/DAG acyltransferase (PDAT) activity.…”

Mucor circinelloides: a model organism for oleaginous fungi and its potential applications in bioactive lipid production

“…M. circinelloides grows well in culture-containing triacylglycerol, indicating a potential source of extracellular lipases [ 89 ]. Many scientific studies from a hereditary perspective have ascertained that M. circinelloides produces many lipase genes [ 35 , 46 , 91 ] .…”