Functional identification and regulatory analysis of Δ6-fatty acid desaturase from the oleaginous fungus Mucor sp. EIM-10
2017
Jiang, Xianzhang | Liu, Hongjiao | Niu, Yongchao | Qi, Feng | Zhang, Mingliang | Huang, Jianzhong
OBJECTIVES: To enlarge the diversity of the desaturases associated with PUFA biosynthesis and to better understand the transcriptional regulation of desaturases, a Δ⁶-desaturase gene (Md6) from Mucor sp. and its 5′-upstream sequence was functionally identified in Saccharomyces cerevisiae. RESULTS: Expression of the Δ⁶-fatty acid desaturase (Md6) in S. cerevisiae showed that Md6 could convert linolenic acid to γ-linolenic acid. Computational analysis of the promoter of Md6 suggested it contains several eukaryotic fundamental transcription regulatory elements. In vivo functional analysis of the promoter showed the 5′-upstream sequence of Md6 could initiate expression of GFP and Md6 itself in S. cerevisiae. A series deletion analysis of the promoter suggested that sequence between −919 to −784 bp (relative to start site) named as eMd6 is the key factor for high activity of Δ⁶-desaturase. The activity of Δ⁶-desaturase was increased by 2.8-fold and 2.5-fold when the eMd6 sequence was placed upstream of −434 with forward or reverse orientations respectively. CONCLUSION: To our best knowledge, the native promoter of Md6 from Mucor is the strongest promoter for Δ⁶-desaturase reported so far and the sequence between −919 to −784 bp is an enhancer for Δ⁶-desaturase activity.
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