Metabolic phenotype of phosphoglucose isomerase mutants of Corynebacterium glutamicum
Metabolic phenotype of phosphoglucose isomerase mutants of Corynebacterium glutamicum
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DOI：
10.1016/s0168-1656(03)00153-6
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发表时间：
2003-09-04
影响因子：
4.1
通讯作者：
de Graaf, AA
中科院分区：
文献类型：
--
作者：
Marx, A;Hans, S;de Graaf, AA
利用通量组学作为一种有效的功能基因组学工具的一系列实验报道表明，谷氨酸棒状杆菌菌株MH20-22B的l -赖氨酸产量与细胞内NADPH供应的程度相关。考虑了一些可替代的代谢工程策略来增加C谷氨酸菌株DSM5715细胞内NADPH的供应，最后赞成通过戊糖磷酸途径重定向碳通量，通过两个NADPH产生酶促反应。此外，利用基因工程技术构建了C - glutamum菌株DSM5715的磷酸葡萄糖异构酶（Pgi）零突变体及其代谢表型的一些方面。最有趣的是，结果表明，零突变株不仅l -赖氨酸的生成量增加了1.7倍，而且副产物浓度也大大降低。在这篇文章中，我们详细讨论了这种代谢表型，并提供了关于副产品形成以及产量考虑的额外数据。基于同位素代谢通量分析的结果结合NADPH代谢的考虑，明确排除了C谷氨酸菌株DSM5715中Pgi同工酶的存在。分析了pgi基因所在的基因组区域。不能排除极性效应可能是由pgi基因的破坏引起的，并可能导致C谷氨酸pgi突变体的代谢表型。我们展示了一个工业赖氨酸生产菌株的Pgi突变体的生长特性。观察到生长速率降低和双相生长行为。讨论了NADPH氧化对Pgi突变体良好平衡生长的重要性。文献中描述了C谷氨酸的另一种磷酸葡萄糖异构酶突变体，其l -赖氨酸产量从42%增加到52%。这一发现强调了代谢通量重定向到戊糖磷酸途径的普遍潜力，可用于(1)芳香族氨基酸和(2)合成依赖于细胞内NADPH供应的化学物质的生物技术合成的代谢工程。(C) 2003 Elsevier B.V.版权所有
A series of experiments reported in the literature using fluxomics as an efficient functional genomics tool revealed that the L-lysine production of the Corynebacterium glutamicum strain MH20-22B correlates with the extent of intracellular NADPH supply. Some alternative metabolic engineering strategies to increase intracellular NADPH supply in the C glutamicum strain DSM5715 were considered and finally the redirection of carbon flux through the pentose phosphate pathway with two NADPH generating enzymatic reactions was favored. Elsewhere, the construction of a phosphoglucose isomerase (Pgi) null mutant of the C glutamicum strain DSM5715 has been described by utilizing genetic engineering as well as some aspects of its metabolic phenotype. Most interestingly, it was shown that not only could the L-lysine formation be increased by 1.7-fold but the by-product concentration for the null mutant strain was also able to be drastically reduced. In this publication we discuss this metabolic phenotype in detail and present additional data on by-product formation as well as yield considerations. Results from isotope based metabolic flux analysis in combination with considerations on NADPH metabolism clearly exclude the existence of Pgi isoenzymes in C glutamicum strain DSM5715. The genome region containing the pgi gene was analyzed. It cannot be excluded that polar effects might have been caused by the disruption of the pgi gene and might have contributed to the observed metabolic phenotype of C glutamicum Pgi mutants. We illustrate growth characteristics of a Pgi mutant of an industrial L-lysine production strain. A reduced growth rate and a biphasic growth behavior was observed. The importance of NADPH reoxidation for well balanced growth in Pgi mutants is discussed. Another phosphoglucose isomerase mutant of C glutamicum has been described in literature with which an increase in L-lysine yield from 42 to 52% was observed. This finding highlights the general potential of metabolic flux redirection towards the pentose phosphate pathway, which could be used for metabolic engineering of the biotechnological synthesis of (1) aromatic amino acids and (2) chemicals whose synthesis depends on intracellular NADPH supply. (C) 2003 Elsevier B.V. All rights reserved.