Department of Biotechnology and Food Microbiology, Wroclaw University of Environmental and Life Sciences, Chełmońskiego 37, 51-630, Wrocław, Poland.
Microb Cell Fact. 2020 Jul 11;19(1):138. doi: 10.1186/s12934-020-01398-x.
During the pentose phosphate pathway (PPP), two important components, NADPH and pentoses, are provided to the cell. Previously it was shown that this metabolic pathway is a source of reducing agent for lipid synthesis from glucose in the yeast Yarrowia lipolytica. Y. lipolytica is an attractive microbial host since it is able to convert untypical feedstocks, such as glycerol, into oils, which subsequently can be transesterified to biodiesel. However, the lipogenesis process is a complex phenomenon, and it still remains unknown which genes from the PPP are involved in lipid synthesis.
To address this problem we overexpressed five genes from this metabolic pathway: transaldolase (TAL1, YALI0F15587g), transketolase (TKL1, YALI0E06479g), ribulose-phosphate 3-epimerase (RPE1, YALI0C11880g) and two dehydrogenases, NADP-dependent glucose-6-phosphate dehydrogenase (ZWF1, YALI0E22649g) and NADP-dependent 6-phosphogluconate dehydrogenase (GND1, YALI0B15598g), simultaneously with diacylglycerol acyltransferase (DGA1, YALI0E32769g) and verified each resulting strain's ability to synthesize fatty acid growing on both glycerol and glucose as a carbon source. Our results showed that co-expression of DGA1 and TKL1 results in higher SCO synthesis, increasing lipid content by 40% over the control strain (DGA1 overexpression).
Simultaneous overexpression of DGA1 and TKL1 genes results in a higher lipid titer independently from the fermentation conditions, such as carbon source, pH and YE supplementation.
在戊糖磷酸途径（PPP）中，细胞提供了两个重要的组成部分，NADPH 和戊糖。此前的研究表明，该代谢途径是酵母解脂耶氏酵母从葡萄糖合成脂质的还原剂来源。解脂耶氏酵母是一种有吸引力的微生物宿主，因为它能够将非典型的原料，如甘油，转化为油，随后可以将其酯化为生物柴油。然而，脂肪生成过程是一个复杂的现象，目前仍不清楚 PPP 中的哪些基因参与了脂质的合成。
为了解决这个问题，我们过量表达了这个代谢途径中的五个基因：转醛醇酶（TAL1，YALI0F15587g）、转酮醇酶（TKL1，YALI0E06479g）、核酮糖-5-磷酸 3-差向异构酶（RPE1，YALI0C11880g）和两个脱氢酶，NADP 依赖性葡萄糖-6-磷酸脱氢酶（ZWF1，YALI0E22649g）和 NADP 依赖性 6-磷酸葡萄糖酸脱氢酶（GND1，YALI0B15598g），同时过量表达二酰基甘油酰基转移酶（DGA1，YALI0E32769g），并验证了每个产生的菌株在甘油和葡萄糖作为碳源时合成脂肪酸的能力。我们的结果表明，DGA1 和 TKL1 的共表达导致更高的 SCO 合成，与对照菌株（DGA1 过表达）相比，脂质含量增加了 40%。
无论发酵条件（如碳源、pH 值和 YE 补充）如何，DGA1 和 TKL1 基因的同时过表达都会导致更高的脂质产量。
