Thermodynamics shapes the in vivo enzyme burden of glycolytic pathways.

IF 4.7 1区 生物学 Q1 MICROBIOLOGY
mBio Pub Date : 2025-10-08 Epub Date: 2025-09-16 DOI:10.1128/mbio.01837-25
Daven B Khana, Annie Jen, Evgenia Shishkova, Kirsten Fisher, Eashant Thusoo, Jonathan Williams, Alex Henkel, David M Stevenson, Joshua J Coon, Daniel Amador-Noguez
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Abstract

Thermodynamically constrained reactions and pathways are hypothesized to impose greater protein demands on cells, requiring higher enzyme amounts to sustain a given flux compared to those with stronger thermodynamics. To test this, we quantified the absolute concentrations of glycolytic enzymes in three bacterial species-Zymomonas mobilis, Escherichia coli, and Clostridium thermocellum-which employ distinct glycolytic pathways with varying thermodynamic driving forces. By integrating enzyme concentration data with corresponding in vivo metabolic fluxes and ΔG measurements, we found that the highly favorable Entner-Doudoroff pathway in Z. mobilis requires only one-fourth the amount of enzymatic protein to sustain the same flux as the thermodynamically constrained pyrophosphate-dependent glycolytic pathway in C. thermocellum, with the Embden-Meyerhof-Parnas pathway in E. coli exhibiting intermediate thermodynamic favorability and enzyme demand. Across all three pathways, early reactions with stronger thermodynamic driving forces generally required lower enzyme investment than later, less favorable steps. Additionally, reflecting differences in glycolytic strategies, the highly reversible ethanol fermentation pathway in C. thermocellum requires 10-fold more protein to maintain the same flux as the irreversible, forward-driven ethanol fermentation pathway in Z. mobilis. Thus, protein investment across glycolytic pathways reflects differences in their thermodynamic favorability.IMPORTANCECells regulate metabolic fluxes to balance energy production, biosynthesis, and the efficient use of limited resources, including the finite capacity for synthesizing and maintaining metabolic enzymes. Here, we present in vivo evidence that strongly thermodynamically favorable metabolic pathways require significantly fewer enzyme resources to sustain a given flux compared to less thermodynamically favorable pathways. These findings underscore the connection between pathway thermodynamics, resource allocation, and enzyme burden, providing valuable insights for metabolic engineering strategies aimed at optimizing pathways for high flux with minimal protein cost.

热力学决定了糖酵解途径的体内酶负荷。
热力学约束的反应和途径被假设对细胞施加了更大的蛋白质需求,与热力学更强的反应和途径相比,需要更高的酶量来维持给定的通量。为了验证这一点,我们量化了三种细菌(活动单胞菌、大肠杆菌和热细胞梭菌)中糖酵解酶的绝对浓度,这三种细菌采用不同的糖酵解途径和不同的热力学驱动力。通过将酶浓度数据与相应的体内代谢通量和ΔG测量相结合,我们发现Z. mobilis中非常有利的enterner - doudoroff途径只需要四分之一的酶蛋白量就可以维持与C. thermocellum中热力学受限的焦磷酸盐依赖的糖酵解途径相同的通量,大肠杆菌中的embden - meyerhoff - parnas途径具有中等的热力学有利性和酶需求。在这三种途径中,具有更强热力学驱动力的早期反应通常比较晚的不利步骤需要更少的酶投入。此外,C. thermocellum中高度可逆的乙醇发酵途径需要比Z. mobilis中不可逆的、正向驱动的乙醇发酵途径多10倍的蛋白质来维持相同的通量,这反映了糖酵解策略的差异。因此,蛋白质在糖酵解途径中的投资反映了它们在热力学有利性上的差异。细胞调节代谢通量以平衡能量产生、生物合成和有限资源的有效利用,包括有限的合成和维持代谢酶的能力。在这里,我们提出的体内证据表明,与热力学不太有利的代谢途径相比,热动力学有利的代谢途径需要更少的酶资源来维持给定的通量。这些发现强调了途径热力学、资源分配和酶负荷之间的联系,为旨在以最小蛋白质成本优化高通量途径的代谢工程策略提供了有价值的见解。
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
mBio
mBio MICROBIOLOGY-
CiteScore
10.50
自引率
3.10%
发文量
762
审稿时长
1 months
期刊介绍: mBio® is ASM''s first broad-scope, online-only, open access journal. mBio offers streamlined review and publication of the best research in microbiology and allied fields.
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