大肠杆菌同时利用葡萄糖、木糖和阿拉伯糖生产（R）-3-羟基丁酸的培养策略。
Cultivation strategies for production of (R)-3-hydroxybutyric acid from simultaneous consumption of glucose, xylose and arabinose by Escherichia coli.
作者信息
Jarmander Johan, Belotserkovsky Jaroslav, Sjöberg Gustav, Guevara-Martínez Mónica, Pérez-Zabaleta Mariel, Quillaguamán Jorge, Larsson Gen
机构信息
School of Biotechnology, Division of Industrial Biotechnology, KTH Royal Institute of Technology, SE 106 91, Stockholm, Sweden.
Center of Biotechnology, Faculty of Science and Technology, Universidad Mayor de San Simón, Cochabamba, Bolivia.
出版信息
Microb Cell Fact. 2015 Apr 11;14:51. doi: 10.1186/s12934-015-0236-2.
BACKGROUND
Lignocellulosic waste is a desirable biomass for use in second generation biorefineries. Up to 40% of its sugar content consist of pentoses, which organisms either take up sequentially after glucose depletion, or not at all. A previously described Escherichia coli strain, PPA652ara, capable of simultaneous consumption of glucose, xylose and arabinose was in the present work utilized for production of (R)-3-hydroxybutyric acid (3HB) from a mixture of glucose, xylose and arabinose.
RESULTS
The Halomonas boliviensis genes for 3HB production were for the first time cloned into E. coli PPA652ara, leading to product secretion directly into the medium. Process design was based on comparisons of batch, fed-batch and continuous cultivation, where both excess and limitation of the carbon mixture was studied. Carbon limitation resulted in low specific productivity of 3HB (<2 mg g(-1) h(-1)) compared to carbon excess (25 mg g(-1) h(-1)), but the yield of 3HB/cell dry weight (Y3HB/CDW) was very low (0.06 g g(-1)) during excess. Nitrogen-exhausted conditions could be used to sustain a high specific productivity (31 mg g(-1) h(-1)) and to increase the yield of 3HB/cell dry weight to 1.38 g g(-1). Nitrogen-limited fed-batch process design led to further increased specific productivity (38 mg g(-1) h(-1)) but also to additional cell growth (Y3HB/CDW=0.16 g g(-1)). Strain PPA652ara did under all processing conditions simultaneously consume glucose, xylose and arabinose, which was not the case for a reference wild type E. coli, which also gave a higher carbon flux to acetic acid.
CONCLUSIONS
It was demonstrated that by using E. coli PPA652ara, it was possible to design a production process for 3HB from a mixture of glucose, xylose and arabinose where all sugars were consumed. An industrial 3HB production process is proposed to be divided into a growth and a production phase, and nitrogen depletion/limitation is a potential strategy to maximize the yield of 3HB/CDW in the latter. The specific productivity of 3HB reported here from glucose, xylose and arabinose by E. coli is further comparable to the current state of the art for production from glucose sources.
背景
木质纤维素废料是用于第二代生物精炼厂的理想生物质。其糖含量中高达40%由戊糖组成，而生物体要么在葡萄糖耗尽后依次利用这些戊糖，要么根本不利用。在本研究中，利用先前描述的能够同时消耗葡萄糖、木糖和阿拉伯糖的大肠杆菌菌株PPA652ara，从葡萄糖、木糖和阿拉伯糖的混合物中生产（R）-3-羟基丁酸（3HB）。
结果
首次将玻利维亚嗜盐单胞菌的3HB生产基因克隆到大肠杆菌PPA652ara中，从而使产物直接分泌到培养基中。工艺设计基于分批培养、补料分批培养和连续培养的比较，研究了碳源混合物的过量和限量情况。与碳源过量（25 mg g⁻¹ h⁻¹）相比，碳源限量导致3HB的比生产率较低（<2 mg g⁻¹ h⁻¹），但在碳源过量期间，3HB/细胞干重的产率非常低（0.06 g g⁻¹）。氮耗尽条件可用于维持高比生产率（31 mg g⁻¹ h⁻¹），并将3HB/细胞干重的产率提高到1.38 g g⁻¹。氮限量补料分批工艺设计导致比生产率进一步提高（38 mg g⁻¹ h⁻¹），但也导致额外的细胞生长（3HB/细胞干重 = 0.16 g g⁻¹）。在所有处理条件下，PPA652ara菌株均能同时消耗葡萄糖、木糖和阿拉伯糖，而对照野生型大肠杆菌则不然，其向乙酸的碳通量也更高。
结论
结果表明，利用大肠杆菌PPA652ara，有可能设计出一种从葡萄糖、木糖和阿拉伯糖的混合物中生产3HB的工艺，其中所有糖类都被消耗。提出将工业3HB生产工艺分为生长阶段和生产阶段，氮耗尽/限量是在后一阶段使3HB/细胞干重产率最大化的潜在策略。此处报道的大肠杆菌利用葡萄糖、木糖和阿拉伯糖生产3HB的比生产率进一步与目前从葡萄糖源生产的技术水平相当。
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