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首页> 外文期刊>Applied Microbiology and Biotechnology >Variation of the by-product spectrum during a-ketoglutaric acid production from raw glycerol by overexpression of fumarase and pyruvate carboxylase genes in Yarrowia lipolytica
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Variation of the by-product spectrum during a-ketoglutaric acid production from raw glycerol by overexpression of fumarase and pyruvate carboxylase genes in Yarrowia lipolytica

机译：在解脂耶氏酵母中富马酸酶和丙酮酸羧化酶基因的过表达导致粗甘油生产α-酮戊二酸过程中副产物谱的变化

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The yeast Yarrowia lipolytica secretes high amounts of various organic acids, like citric, isocitric, pyruvic (PA), and a-ketoglutaric (KGA) acids, triggered by growth limitation and excess of carbon source. This is leading to an increased interest in this non-conventional yeast for biotechnological applications. To improve the KGA production by Y. lipolytica for an industrial application, it is necessary to reduce the amounts of by-products, e.g., fumarate (FU) and PA, because production of by-products is a main disadvantage of the KGA production by this yeast.We have examined whether the concentration of secreted organic acids (main product KGA and PA as major by-product and FU, malate (MA), and succinate (SU) as minor by-products) can be influenced by a gene-dose-dependent overexpression of fumarase (FUM) or pyruvate carboxylase (PYC) genes under KGA production conditions. Recombinant Y. lipolytica strains were constructed, which harbor multiple copies of the respective FUM1, PYC1 or FUM1, and PYC1 genes. Overexpression of the genes FUM1 and PYC1 resulted in strongly increased specific enzyme activities during cultivation of these strains on raw glycerol as carbon source in bioreactors. The recombinant Y. lipolytica strains showed different product selectivity of the secreted organic acids KGA, PA, FU, MA, and SU. Concentrations of the by-products FU, MA, SU, and PA decreased significantly at overproduction of FUM and increased at overproduction of PYC and also of FUM and PYC simultaneously. In contrast, the production of KGAwith the multicopy strains H355A(FUM1) and H355A(FUM1- PYC1) was comparable with the wild-Type strain H355 or slightly lower in case of H355(PYC1). KGA productivity was not changed significantly compared with strain H355 whereas product selectivity of the main product KGA was increased in H355A(FUM1).
机译：酵母解脂耶氏酵母（Yarrowia lipolytica）会分泌大量各种有机酸，例如柠檬酸，柠檬酸，丙酮酸（PA）和α-酮戊二酸（KGA），这是由于生长受限和碳源过多引起的。这导致人们对该生物技术应用的非常规酵母越来越感兴趣。为了改善脂解耶氏酵母在工业上的KGA产量，有必要减少副产物的量，例如富马酸盐（FU）和PA，因为副产物的产生是KGA生产KGA的主要缺点。我们检查了分泌的有机酸（主要副产物KGA和PA以及次要副产物FU，苹果酸（MA）和琥珀酸酯（SU））的浓度是否受以下基因影响：在KGA生产条件下，富马酸酶（FUM）或丙酮酸羧化酶（PYC）基因的剂量依赖性过表达。构建了重组解脂耶氏酵母菌株，其中包含各自FUM1，PYC1或FUM1和PYC1基因的多个副本。基因FUM1和PYC1的过表达导致这些菌株在粗甘油上作为生物反应器中的碳源进行培养期间比酶活性大大增加。重组解脂耶氏酵母菌株显示了分泌的有机酸KGA，PA，FU，MA和SU的不同产物选择性。副产物FU，MA，SU和PA的浓度在FUM过量生产时显着降低，而在PYC以及FUM和PYC同时过量时则升高。相反，多拷贝菌株H355A（FUM1）和H355A（FUM1-PYC1）的KGA产量与野生型菌株H355相当，在H355（PYC1）情况下稍低。与菌株H355相比，KGA生产率没有显着变化，而在H355A（FUM1）中，主要产物KGA的产物选择性增加了。

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