Metabolic impact of redox cofactor perturbations in Saccharomyces cerevisiae
Metabolic impact of redox cofactor perturbations in Saccharomyces cerevisiae
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DOI：
10.1016/j.ymben.2009.05.001
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发表时间：
2009-07-01
影响因子：
8.4
通讯作者：
Vemuri, Goutham N.
中科院分区：
文献类型：
--
作者：
Hou, Jin;Lages, Nuno F.;Vemuri, Goutham N.
氧化还原辅因子在代谢过程中偶联催化剂与拮抗剂和能量产生中起关键作用。这些辅因子的细胞内水平存在微妙的平衡，以确定最佳的代谢输出。因此，辅因子正在成为诱导代谢广泛变化的有吸引力的靶标。我们提出了一个详细的分析，在细胞质或线粒体中的氧化还原辅因子的扰动对葡萄糖和能量代谢的影响，在酿酒酵母，以帮助代谢工程决策，涉及辅因子工程。我们通过引入NADH氧化酶或替代氧化酶来增强NADH氧化，通过NADH激酶增强其ATP介导的向NADPH的转化，以及通过可溶性、非质子转运的细菌转氢酶增强不依赖于ATP的NADH和NADPH的相互转化。降低胞质NADH水平降低甘油产量，而降低线粒体NADH降低乙醇产量。然而，当这些反应与NADPH的产生相结合时，代谢变化更加缓和。这些扰动的直接后果可以在辅因子的细胞内浓度的变化中看到。产物谱和细胞内代谢物水平的变化与生物质合成的ATP需求和氧化磷酸化的效率密切相关，如从简单的化学计量模型估计的。本文的结果将为定量理解和预测代谢工程应用中细胞对氧化还原扰动的反应提供有价值的见解。(C)2009 Elsevier Inc. All rights reserved.
Redox cofactors play a pivotal role in coupling catabolism with anabolism and energy generation during metabolism. There exists a delicate balance in the intracellular level of these cofactors to ascertain an optimal metabolic output. Therefore, cofactors are emerging to be attractive targets to induce widespread changes in metabolism. We present a detailed analysis of the impact of perturbations in redox cofactors in the cytosol or mitochondria on glucose and energy metabolism in Saccharomyces cerevisiae to aid metabolic engineering decisions that involve cofactor engineering. We enhanced NADH oxidation by introducing NADH oxidase or alternative oxidase, its ATP-mediated conversion to NADPH using NADH kinase as well as the interconversion of NADH and NADPH independent of ATP by the soluble, non-proton-translocating bacterial transhydrogenase. Decreasing cytosolic NADH level lowered glycerol production, while decreasing mitochondrial NADH lowered ethanol production. However, when these reactions were coupled with NADPH production, the metabolic changes were more moderated. The direct consequence of these perturbations could be seen in the shift of the intracellular concentrations of the cofactors. The changes in product profile and intracellular metabolite levels were closely linked to the ATP requirement for biomass synthesis and the efficiency of oxidative phosphorylation, as estimated from a simple stoichiometric model. The results presented here will provide valuable insights for a quantitative understanding and prediction of cellular response to redox-based perturbations for metabolic engineering applications. (C) 2009 Elsevier Inc. All rights reserved.