Enhancing LC-PUFA production in Thalassiosira pseudonana by overexpressing the endogenous fatty acid elongase genes
Enhancing LC-PUFA production in Thalassiosira pseudonana by overexpressing the endogenous fatty acid elongase genes
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DOI：
10.1007/s10811-015-0617-2
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发表时间：
2016-04-01
影响因子：
3.3
通讯作者：
Hildebrand, Mark
中科院分区：
文献类型：
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作者：
Cook, Orna;Hildebrand, Mark
有益健康的ω-3长链多不饱和脂肪酸（LC-PUFA）、二十碳五烯酸（EPA）和二十二碳六烯酸（DHA）由硅藻通过脂肪酸延伸酶和去饱和酶的连续步骤天然合成。在海链藻中，这些脂肪酸占总脂肪酸的约10- 20%，EPA的积累比DHA高5 - 10倍。为了确定T.为了控制EPA和DHA的产生，我们构建了过表达每个T。长链脂肪酸延长酶基因。将全长蛋白质与GFP融合，并产生转基因株系。此外，测试了没有GFP融合的过表达的天然蛋白。确定每种延伸酶蛋白的亚细胞定位。然后，我们检查了每个转基因系与野生型相比的脂质总量并分析了脂肪酸谱。具有过表达的延伸酶的品系显示EPA增加高达1.4倍，DHA增加高达4.5倍，并且增加的脂肪酸的类型（EPA与DHA）取决于过表达的延伸酶的类型。这些数据为未来的代谢工程方法提供了信息，以进一步提高硅藻中的EPA和DHA含量。
The health beneficial omega-3 long-chain polyunsaturated fatty acids (LC-PUFAs), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA) are naturally synthesized by diatoms through consecutive steps of fatty acid elongase and desaturase enzymes. In Thalassiosira pseudonana, these fatty acids constitute about 10-20 % of the total fatty acids, with EPA accumulation being five to ten times higher than DHA. In order to identify the subcellular localization of enzymes in the pathway of LC-PUFA biosynthesis in T. pseudonana and to manipulate the production of EPA and DHA, we generated constructs for overexpressing each of the T. pseudonana long-chain fatty acid elongase genes. Full-length proteins were fused to GFP, and transgenic lines were generated. In addition, overexpressed native proteins with no GFP fusion were tested. The subcellular localization of each elongase protein was determined. We then examined the total amount of lipids and analyzed the fatty acid profile in each of the transgenic lines compared to wild type. Lines with overexpressed elongases showed an increase of up to 1.4-fold in EPA and up to 4.5-fold in DHA, and the type of fatty acid that was increased (EPA vs. DHA) depended on the type of elongase that was overexpressed. This data informs future metabolic engineering approaches to further improve EPA and DHA content in diatoms.