四个脂肪酸合成酶基因在哺乳动物细胞中的超表达提高长链多不饱和脂肪酸生物合成效率

Overexpression of four fatty acid synthase genes elevated the efficiency of long-chain polyunsaturated fatty acids biosynthesis in mammalian cells

摘要:

DHA (22:6n-3)、EPA (20:5n-3) 和ARA (20:4n-6) 三种长链多不饱和脂肪酸在生物体内活性最强，它们在促进大脑发育和功能维持以及在预防和治疗心血管疾病、炎症、癌症等多种疾病方面有着重要作用。然而，尽管哺乳动物体内有完整的长链多不饱和脂肪酸合成酶系，但哺乳动物合成这些长链多不饱和脂肪酸的效率很低而主要依赖于食物获取。本研究应用转基因方法，将哺乳动物来源的Δ6和Δ5脂肪酸去饱和酶以及Δ6和Δ5脂肪酸延长酶这4种酶的编码基因构建成为一个多基因表达载体，然后转染哺乳动物细胞HEK293T，实现了4个目的基因的超表达，再通过气质联用 (GC-MS) 分析证实了DHA、EPA和ARA等长链多不饱和脂肪酸的合成效率及水平显著增加，DHA的水平更是提高了2.5倍。由此可见，哺乳动物具有某种抑制长链多不饱和脂肪酸高水平合成的机制，但通过Δ6和Δ5脂肪酸去饱和酶以及Δ6和Δ5脂肪酸延长酶的超表达，能够打破哺乳动物这种抑制机制，从而显著提高DHA、EPA、ARA等的合成水平。同时，本研究的思路也为在转基因动物中生产长链多不饱和脂肪酸提供了重要的启示。

Abstract:

Three long-chain polyunsaturated fatty acids, docosahexaenoic acid (DHA, 22:6n-3), eicosapentaenoic acid (EPA, 20:5n-3) and arachidonic acid (ARA, 20:4n-6), are the most biologically active polyunsaturated fatty acids in the body. They are important in developing and maintaining the brain function, and in preventing and treating many diseases such as cardiovascular disease, inflammation and cancer. Although mammals can biosynthesize these long-chain polyunsaturated fatty acids, the efficiency is very low and dietary intake is needed to meet the requirement. In this study, a multiple-genes expression vector carrying mammalian △6/△5 fatty acid desaturases and △6/△5 fatty acid elongases coding genes was used to transfect HEK293T cells, then the overexpression of the target genes was detected. GC-MS analysis shows that the biosynthesis efficiency and level of DHA, EPA and ARA were significantly increased in cells transfected with the multiple-genes expression vector. Particularly, DHA level in these cells was 2.5 times higher than in the control cells. This study indicates mammal possess a certain mechanism for suppression of high level of biosynthesis of long chain polyunsaturated fatty acids, and the overexpression of △6/△5 fatty acid desaturases and △6/△5 fatty acid elongases broke this suppression mechanism so that the level of DHA, EPA and ARA was significantly increased. This study also provides a basis for potential applications of this gene construct in transgenic animal to produce high level of these long-chain polyunsaturated fatty acid.