Tan Li, Li Shue, Zhang Xiaoyu, Ma Fuying
Key Laboratory of Molecular Biophysics of MOE, College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, 430074, P.R. China.
J Sci Food Agric. 2015 Dec;95(15):3077-83. doi: 10.1002/jsfa.7043. Epub 2015 Jan 8.
Δ6-desaturase belonging to membrane-bound enzyme is a key enzyme involved in the synthesis of polyunsaturated fatty acids (PUFAs). This study aimed to clone and characterise Δ6-desaturase gene and its upstream regulatory region of Mortierella sp. AGED.
Glucose and soybean meal are best for lipid and arachidonic acid accumulation of Mortierella sp. AGED. A 1375-bp Δ6-desaturase gene AGfad6 which contains a 1275-bp open reading frame encoding 424 amino acids without signal peptide was cloned. The putative protein contained three conserved histidine-rich motifs and a conserved cytochrome b5 HPGG (H: Histine, P: Proline, G: Glycine, G: Glycine) motif, with a mass of 48.3 kDa and an isoelectric point of 5.96. AGfad6 was successfully expressed in Pichia pastoris GS115, which exerted the effect on converting linoleic acid to γ-linolenic acid. The 1712-bp upstream region contained basic transcriptional elements including TATA, GC and GATA box, putative target-binding sites for transcription factors such as TATA binding protein, transcription activator, CCAAT-enhancer-binding protein, activator protein 1, alcohol dehydrogenase gene regulator 1 and metabolic regulators p40x in fungi, stress-related elements including GT-1 (light-responsive, salicylic acid-inducible), stress response element, heat stress-responsive element, which might participate in regulation of PUFAs synthesis.
The present finding could enable us to understand the evolution and regulatory mechanism of Δ6-desaturase gene.
Δ6-去饱和酶属于膜结合酶，是参与多不饱和脂肪酸（PUFAs）合成的关键酶。本研究旨在克隆并鉴定深黄被孢霉Δ6-去饱和酶基因及其上游调控区域。
葡萄糖和豆粕最有利于深黄被孢霉脂质和花生四烯酸的积累。克隆得到一个1375 bp的Δ6-去饱和酶基因AGfad6，其包含一个1275 bp的开放阅读框，编码424个无信号肽的氨基酸。推测的蛋白质包含三个保守的富含组氨酸基序和一个保守的细胞色素b5 HPGG（H：组氨酸，P：脯氨酸，G：甘氨酸，G：甘氨酸）基序，分子量为48.3 kDa，等电点为5.96。AGfad6在毕赤酵母GS115中成功表达，具有将亚油酸转化为γ-亚麻酸的作用。1712 bp的上游区域包含基本转录元件，如TATA、GC和GATA框，以及真菌中TATA结合蛋白、转录激活因子、CCAAT增强子结合蛋白、激活蛋白1、乙醇脱氢酶基因调节因子1和代谢调节因子p40x等转录因子的假定靶结合位点，还有应激相关元件，如GT-1（光响应、水杨酸诱导）、应激反应元件、热应激反应元件，这些元件可能参与多不饱和脂肪酸合成调控。
本研究结果有助于我们了解Δ6-去饱和酶基因的进化及调控机制。
