Effect of Single and Combined Expression of Lysophosphatidic Acid Acyltransferase, Glycerol-3-Phosphate Acyltransferase, and Diacylglycerol Acyltransferase on Lipid Accumulation and Composition in Neochloris oleoabundans
Effect of Single and Combined Expression of Lysophosphatidic Acid Acyltransferase, Glycerol-3-Phosphate Acyltransferase, and Diacylglycerol Acyltransferase on Lipid Accumulation and Composition in Neochloris oleoabundans
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DOI：
10.3389/fpls.2019.01573
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发表时间：
2019-11-29
影响因子：
5.6
通讯作者：
Wijffels, Rene H.
中科院分区：
文献类型：
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作者：
Munoz, Camilo F.;Weusthuis, Ruud A.;Wijffels, Rene H.
微藻脂质是有前途的食品和生物燃料原料。由于微藻生产脂质在经济上还不可行，因此基因工程正在成为实现更高脂质积累和生产率的有前途的策略。肯尼迪途径中涉及的酶，如甘油-3-磷酸酰基转移酶 (GPAT)、溶血磷脂酸酰基转移酶 (LPAT) 和二酰基甘油酰基转移酶 (DGAT) 催化三酰基甘油形成的关键步骤，三酰基甘油是 N. oleoabundans 中脂质的主要成分。这些酶在目标菌株中的过度表达具有进一步增加其三酰甘油含量的巨大潜力。我们在 N. oleoabundans 中过表达 Acutodesmus obliquus 的 LPAT、GPAT 和 DGAT 的单个和多个编码基因。过表达单基因的菌株产生高达52%和45%的g中心点gDW(-1)，这对应于总脂肪酸和三酰甘油分别增加1.3倍和1.4倍。三个基因的协调表达导致g中心点gDW(-1)分别增加49%和39%，总脂肪酸和三酰甘油增加了1.2倍。 LPAT、GPAT 和 DGAT 基因的单一表达导致饥饿期间更高的脂质生产率，而在充足条件下对生长和光合作用活性没有显着影响。另一方面，同时表达LPAT、GPAT和DGAT基因会导致生长率降低52%，光合活性降低14%，细胞直径增加4倍。此外，多基因表达系在氮饥饿条件下表现出碳水化合物和蛋白质含量的减少以及色素的增加。异源基因 LPAT、GPAT 和 DGAT 的单一和多重表达显示显着增强了 N. oleoabundans 中的脂质积累。单基因表达导致更高的脂质产量和生产力，而不对菌株的生理状态产生显着影响。这种方法显示了产生具有更高的脂质生产经济潜力的微藻菌株的潜力。
Microalgal lipids are promising feedstocks for food and biofuels. Since lipid production by microalgae is not yet economically feasible, genetic engineering is becoming a promising strategy to achieve higher lipid accumulation and productivities. Enzymes involved in the Kennedy pathway such as glycerol-3-phosphate acyltransferase (GPAT), lysophosphatidic acid acyltransferase (LPAT), and diacylglycerol acyltransferase (DGAT) catalyze key steps in the formation of triacylglycerol, which is the main constituent of lipids in N. oleoabundans. The overexpression of these enzymes in the targeted strain has a great potential to further increase their triacylglycerol content. We overexpressed single and multiple encoding genes for LPAT, GPAT, and DGAT from Acutodesmus obliquus in N. oleoabundans. Strains overexpressing single genes produced up to 52% and 45% g center dot gDW(-1), which corresponds to 1.3- and 1.4-fold increase in total fatty acids and triacylglycerols, respectively. The orchestrated expression of the three genes resulted in 49% and 39% g center dot gDW(-1), which is 1.2-folds increase in total fatty acids and triacylglycerols. Single expression of LPAT, GPAT, and DGAT genes resulted in higher lipid productivities during starvation without a significant effect on growth and photosynthetic activity during replete conditions. On the other hand, the simultaneous expression of LPAT, GPAT, and DGAT genes resulted in 52% lower growth rate, 14% lower photosynthetic activity and 4-folds increase in cell diameter. Moreover, the multigene expressing line showed a decrease in carbohydrates and protein content and an increase in pigments during nitrogen starved condition. The single and multiple expression of heterologous genes LPAT, GPAT, and DGAT showed to significantly enhanced the lipid accumulation in N. oleoabundans. Single gene expression resulted in higher lipid production and productivities without having a significant impact in the physiological status of the strains. This approach shows the potential for the generation of microalgal strains with higher economical potential for the production of lipids.