Transcriptional and Metabolic Response of a Strain of Escherichia coli PTS− to a Perturbation of the Energetic Level by Modification of [ATP]/[ADP] Ratio

IF 2.7 Q3 BIOTECHNOLOGY & APPLIED MICROBIOLOGY
BioTech Pub Date : 2024-04-10 DOI:10.3390/biotech13020010
Sandra Soria, Ofelia E. Carreón-Rodríguez, R. de Anda, N. Flores, A. Escalante, Francisco Bolívar
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Abstract

The intracellular [ATP]/[ADP] ratio is crucial for Escherichia coli’s cellular functions, impacting transport, phosphorylation, signaling, and stress responses. Overexpression of F1-ATPase genes in E. coli increases glucose consumption, lowers energy levels, and triggers transcriptional responses in central carbon metabolism genes, particularly glycolytic ones, enhancing carbon flux. In this contribution, we report the impact of the perturbation of the energetic level in a PTS− mutant of E. coli by modifying the [ATP]/[ADP] ratio by uncoupling the cytoplasmic activity of the F1 subunit of the ATP synthase. The disruption of [ATP]/[ADP] ratio in the evolved strain of E. coli PB12 (PTS−) was achieved by the expression of the atpAGD operon encoding the soluble portion of ATP synthase F1-ATPase (strain PB12AGD+). The analysis of the physiological and metabolic response of the PTS− strain to the ATP disruption was determined using RT–qPCR of 96 genes involved in glucose and acetate transport, glycolysis and gluconeogenesis, pentose phosphate pathway (PPP), TCA cycle and glyoxylate shunt, several anaplerotic, respiratory chain, and fermentative pathways genes, sigma factors, and global regulators. The apt mutant exhibited reduced growth despite increased glucose transport due to decreased energy levels. It heightened stress response capabilities under glucose-induced energetic starvation, suggesting that the carbon flux from glycolysis is distributed toward the pentose phosphate and the Entner–Duodoroff pathway with the concomitant. Increase acetate transport, production, and utilization in response to the reduction in the [ATP]/[ADP] ratio. Upregulation of several genes encoding the TCA cycle and the glyoxylate shunt as several respiratory genes indicates increased respiratory capabilities, coupled possibly with increased availability of electron donor compounds from the TCA cycle, as this mutant increased respiratory capability by 240% more than in the PB12. The reduction in the intracellular concentration of cAMP in the atp mutant resulted in a reduced number of upregulated genes compared to PB12, suggesting that the mutant remains a robust genetic background despite the severe disruption in its energetic level.
大肠杆菌 PTS- 菌株对改变[ATP]/[ADP]比例以扰乱能量水平的转录和代谢反应
细胞内[ATP]/[ADP]比率对大肠杆菌的细胞功能至关重要,影响着运输、磷酸化、信号传递和应激反应。在大肠杆菌中过表达 F1-ATP 酶基因会增加葡萄糖消耗,降低能量水平,并引发碳代谢中心基因(尤其是糖酵解基因)的转录反应,从而提高碳通量。在这篇论文中,我们报告了通过解除 ATP 合酶 F1 亚基细胞质活性的耦合,改变 [ATP] /[ADP] 比率,从而扰乱大肠杆菌 PTS- 突变体能量水平的影响。通过表达编码 ATP 合成酶 F1-ATP 酶可溶性部分的 atpAGD 操作子(菌株 PB12AGD+),大肠杆菌进化菌株 PB12(PTS-)的[ATP]/[ADP]比率被破坏。利用 RT-qPCR 对参与葡萄糖和乙酸盐转运、糖酵解和葡萄糖生成、磷酸戊糖途径 (PPP)、TCA 循环和乙醛酸分流的 96 个基因、几个无性繁殖、呼吸链和发酵途径基因、sigma 因子和全局调控因子进行了分析,确定了 PTS- 菌株对 ATP 干扰的生理和代谢反应。由于能量水平下降,尽管葡萄糖转运增加,但apt突变体却表现出生长减弱。在葡萄糖诱导的能量饥饿条件下,它的应激反应能力增强,这表明糖酵解产生的碳通量被分配到磷酸戊糖和 Entner-Duodoroff 途径,并伴随着。随着[ATP]/[ADP]比率的降低,乙酸盐的运输、产生和利用也随之增加。编码 TCA 循环和乙醛酸分流的几个基因以及几个呼吸基因的上调表明呼吸能力增强,同时可能还增加了 TCA 循环中电子供体化合物的可用性,因为该突变体的呼吸能力比 PB12 提高了 240%。与 PB12 相比,atp 突变体细胞内 cAMP 浓度的降低导致上调基因数量的减少,这表明尽管突变体的能量水平受到了严重破坏,但它仍然是一个稳健的遗传背景。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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来源期刊
BioTech
BioTech Immunology and Microbiology-Applied Microbiology and Biotechnology
CiteScore
3.70
自引率
0.00%
发文量
51
审稿时长
11 weeks
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