Production Of Long Chain Monounsaturated Fatty Acids By Saccharomyces Cerevisiae Regu-Lated By Transcription Factor Mga2

Long-chain monounsaturated fatty acids,which are important components in fat,participate in a variety of metabolic processes.Palmitoleic acid(C16:1^Δ9)has value in nutrition and medicine,and has a certain effect on diabetes and cardiovascular disease.At present,only several wild plants and marine fishes are rich in palmitoleic acid.The short of the source limits the production of palmitoleic acid.Therefore,it is necessary to develop the method for biosynthesis of palmitoleic acid by microbes.In this study,we used Saccharomyces cerevisiae as the microbial platform.We investigated the function of transcription factor Mga2 on the C16:1^Δ9 fatty acid synthesis and metabolism.And we optimized the fermentation conditions to produce C16:1^Δ9 in Saccharomyces cerevisiae.1.The MGA2 overexpression Saccharomyces cerevisiae was constructed.The regulatory effect of Mga2 on the synthesis of unsaturated fatty acids in Saccharomyces cerevisiae was initially determined,through transcriptome analysis.The results showed that overexpression of MGA2 resulted the upregulation of key fatty acid synthesis genes such as △9 fatty acid desaturase gene OLE1,acetyl-CoA carboxylase gene ACC1,and fatty acid synthase gene FAS1.2.We explored the effect of overexpressed or truncated MGA2 on biomass growth and monounsaturated fatty acid synthesis in Saccharomyces cerevisiae.The stress tolerance results showed that the overexpressed MGA2 strain has better growth on ethanol and NaCl stress condition,and has more resistance to starvation and antibiotic,is more sensitive to hydrogen peroxide stress.Moreover,the truncated MGA2 strains showed more tolerance to acetic acid stress.The shake flask fermentation results showed that the overexpressed MGA2 strain contained less C16:1^Δ9 than that in control strain.By contrast,the truncated MGA2 strains contained more monounsaturated fatty acids.The content of monounsaturated fatty acids in truncated MGA2 strains YS58MGA2_（529aa）can up reach to 83.61%（v/v）.Comparing with control strain,the C16:1 content of MGA2_（706aa）and MGA2_（529aa）strains increased by about 9.70%.3.We explored the effect of co-expression of MGA2 and exogenous fatty acid elongase gene KCS on the accumulation of C16:1^Δ9 in Saccharomyces cerevisiae.The co-expression of KCS and MGA2 increased the content of C16:1^Δ9 in Saccharomyces cerevisiae significantly.Compare with the control strain,the co-expression strain increased 21.41%in C16:1^Δ9 content,with 47.34%（v/v）.The co-expression of KCS、FAE and MGA2 has less influence to accumulation of C16:1^Δ9.In the truncated MGA2 and KCS co-expression strain,the content of C16:1^Δ9 reached 48.17%（v/v）,which was 23.36%higher than that of the control strain.4.The fermentation conditions of the engineered strains were optimized.The single-factor experiments results showed the initial pH value,the addition of biotin and Asn had a positive impact on the output of palmitoleic acid.And the optimal fermentation conditions were built by response surface analysis.Under the optimal condition,the content of palmitoleic acid in engineered strain can reach up to 53.12%（v/v）.