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首页> 外文期刊>Applied Microbiology >Combined Fluxomics and Transcriptomics Analysis of Glucose Catabolism via a Partially Cyclic Pentose Phosphate Pathway in Gluconobacter oxydans 621H
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Combined Fluxomics and Transcriptomics Analysis of Glucose Catabolism via a Partially Cyclic Pentose Phosphate Pathway in Gluconobacter oxydans 621H

机译：在氧化葡糖杆菌621H中通过部分循环的磷酸戊糖途径进行葡萄糖代谢的通量组学和转录组学分析

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In this study, the distribution and regulation of periplasmic and cytoplasmic carbon fluxes in Gluconobacter oxydans 621H with glucose were studied by ~(13)C-based metabolic flux analysis (~(13)C-MFA) in combination with transcriptomics and enzyme assays. For ~(13)C-MFA, cells were cultivated with specifically ~(13)C-labeled glucose, and intracellular metabolites were analyzed for their labeling pattern by liquid chromatography-mass spectrometry (LC-MS). In growth phase I, 90% of the glucose was oxidized periplasmically to gluconate and partially further oxidized to 2-ketogluconate. Of the glucose taken up by the cells, 9% was phosphorylated to glucose 6-phosphate, whereas 91% was oxidized by cytoplasmic glucose dehydrogenase to gluconate. Additional gluconate was taken up into the cells by transport. Of the cytoplasmic gluconate, 70% was oxidized to 5-ketogluconate and 30% was phosphorylated to 6-phosphogluconate. In growth phase II, 87% of gluconate was oxidized to 2-ketogluconate in the periplasm and 13% was taken up by the cells and almost completely converted to 6-phosphogluconate. Since G. oxydans lacks phosphofructokinase, glucose 6-phosphate can be metabolized only via the oxidative pentose phosphate pathway (PPP) or the Entner-Doudoroff pathway (EDP). ~(13)C-MFA showed that 6-phosphogluconate is catabolized primarily via the oxidative PPP in both phases I and II (62% and 93%) and demonstrated a cyclic carbon flux through the oxidative PPP. The transcriptome comparison revealed an increased expression of PPP genes in growth phase II, which was supported by enzyme activity measurements and correlated with the increased PPP flux in phase II. Moreover, genes possibly related to a general stress response displayed increased expression in growth phase II.
机译：在这项研究中，通过基于〜（13）C的代谢通量分析（〜（13）C-MFA）结合转录组学和酶法研究了葡萄糖氧化葡糖杆菌621H中周质和细胞质碳通量的分布和调控。对于〜（13）C-MFA，将细胞用〜（13）C特异性标记的葡萄糖培养，并通过液相色谱-质谱（LC-MS）分析细胞内代谢产物的标记方式。在生长期I中，90％的葡萄糖被周质氧化为葡萄糖酸盐，部分被进一步氧化为2-酮葡萄糖酸盐。细胞吸收的葡萄糖中，有9％被磷酸化为6-磷酸葡萄糖，而91％被细胞质葡萄糖脱氢酶氧化为葡萄糖酸盐。通过运输将另外的葡萄糖酸酯吸收到细胞中。在细胞质葡萄糖酸中，有70％被氧化为5-酮葡萄糖酸，有30％被磷酸化为6-磷酸葡萄糖酸。在生长阶段II中，周质中87％的葡萄糖酸被氧化为2-酮葡萄糖酸，而13％的葡萄糖被细胞吸收并几乎完全转化为6-磷酸葡萄糖酸。由于氧化单孢菌缺乏磷酸果糖激酶，因此6-磷酸葡萄糖只能通过氧化戊糖磷酸途径（PPP）或Entner-Doudoroff途径（EDP）代谢。 〜（13）C-MFA表明6-磷酸葡萄糖酸酯主要通过氧化PPP在I和II阶段（62％和93％）分解代谢，并显示出通过氧化PPP的环状碳通量。转录组比较显示，在生长阶段II中PPP基因表达增加，这被酶活性测量所支持，并且与在生长阶段II中PPP通量增加相关。而且，可能与一般应激反应有关的基因在生长期II中显示出增加的表达。

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