Role of cyanobacterial phosphoketolase in energy regulation and glucose secretion under dark anaerobic and osmotic stress conditions

Chuang, Derrick S.; Liao, James C.

doi:10.1016/j.ymben.2020.12.004

Cited by 21 publications

(25 citation statements)

References 25 publications

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“…Similar evidence concerning this fine-tuned network for Pi was manifested in S. elongatus PCC 7942. 67 Additionally, product-specific reaction rates that contributed to the flow of Pi were identified. For example, inorganic diphosphatase (PPA) was shown to have a high contribution for isoprene biosynthesis, by pulling out diphosphate – the byproduct of isoprene synthase.…”

Section: Resultsmentioning

confidence: 99%

Optimal energy and redox metabolism in the cyanobacterium Synechocystis sp. PCC 6803

Kugler¹,

Stensjö²

2022

Preprint

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Cyanobacteria represent an attractive platform for the sustainable production of chemicals and fuels. However, the obtained rates, yields, and titers are below those required for commercial application. Carbon metabolism alone cannot achieve maximal accumulation of end-products, since an efficient production of target molecules entails energy and redox balance, in addition to carbon flow. The interplay between cofactor regeneration and heterologous metabolite overproduction in cyanobacteria is not fully explored. Here, we applied stoichiometric metabolic modelling of the cyanobacterium Synechocystis sp. PCC 6803, in order to investigate the optimality of energy and redox metabolism, while overproducing bio-alkenes - isobutene, isoprene, ethylene and 1-undecene. Our network-wide analysis indicates that the rate of NADP+ reduction, rather than ATP synthesis, controls ATP/NADPH ratio, and thereby chemical production. The simulation implies that energy and redox balance necessitates gluconeogenesis, and that acetate metabolism via phosphoketolase serves as an efficient carbon- and energy-recycling pathway. Furthermore, we show that an auxiliary pathway, composed of serine, one-carbon and glycine metabolism, supports cellular redox homeostasis and ATP cycling, and that the Synechocystis metabolism is controlled by few key reactions carrying a high flux. The study also revealed non-intuitive metabolic pathways to enhance isoprene, ethylene and 1-undecene production. We conclude that metabolism of ATP and NAD(P)H is entwined with carbon and nitrogen metabolism, and cannot be assessed in isolation. We envision that the presented here in-depth metabolic analysis will guide the a priori design of Synechocystis as a host strain for an efficient manufacturing of target products.

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Section: Resultsmentioning

confidence: 99%

Optimal energy and redox metabolism in the cyanobacterium Synechocystis sp. PCC 6803

Kugler¹,

Stensjö²

2022

Preprint

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“…The low cellular energy charge of the PCCK strain during the early phase of fermentation, probably accompanied by the repression of high acetate production concomitant with ATP production, may reflect low pool sizes of sugar phosphates as ATP is considered a major allosteric inhibitor that represses the activities of glycolytic enzymes such as pyruvate kinase 36 and phosphoketolase. 37 The PCCK strain produces less lactate and more malate (Figure 3), suggesting that the Synechocystis PCCK strain utilizes its reducing power more for the reductive TCA branch. Accumulation of acetyl-CoA and the low pool size of citrate in the PCCK strain clearly explain the interruption of citrate biosynthesis from oxaloacetate and acetyl-CoA, leading to a low flux of the oxidative TCA branch.…”

Section: Discussionmentioning

confidence: 99%

Metabolic and Microbial Community Engineering for Four-Carbon Dicarboxylic Acid Production from CO₂-Derived Glycogen in the Cyanobacterium Synechocystis sp. PCC6803

et al. 2022

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The four-carbon (C4) dicarboxylic acids, fumarate, malate, and succinate, are the most valuable targets that must be exploited for CO2-based chemical production in the move to a sustainable low-carbon future. Cyanobacteria excrete high amounts of C4 dicarboxylic acids through glycogen fermentation in a dark anoxic environment. The enhancement of metabolic flux in the reductive TCA branch in the Cyanobacterium Synechocystis sp. PCC6803 is a key issue in the C4 dicarboxylic acid production. To improve metabolic flux through the anaplerotic pathway, we have created the recombinant strain PCCK, which expresses foreign ATP-forming phosphoenolpyruvate carboxykinase (PEPck) concurrent with intrinsic phosphoenolpyruvate carboxylase (Ppc) overexpression. Expression of PEPck concurrent with Ppc led to an increase in C4 dicarboxylic acids by autofermentation. Metabolome analysis revealed that PEPck contributed to an increase in carbon flux from hexose and pentose phosphates into the TCA reductive branch. To enhance the metabolic flux in the reductive TCA branch, we examined the effect of corn-steep liquor (CSL) as a nutritional supplement on C4 dicarboxylic acid production. Surprisingly, the addition of sterilized CSL enhanced the malate production in the PCCK strain. Thereafter, the malate and fumarate excreted by the PCCK strain are converted into succinate by the CSL-settling microorganisms. Finally, high-density cultivation of cells lacking the acetate kinase gene showed the highest production of malate and fumarate (3.2 and 2.4 g/L with sterilized CSL) and succinate (5.7 g/L with non-sterile CSL) after 72 h cultivation. The present microbial community engineering is useful for succinate production by one-pot fermentation under dark anoxic conditions.

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“…However, in nature the PK mainly cooperates with acetate kinase (AK), and even forms gene clusters that convert AcP into acetate and ATP for energy supply (Fig. S22 and S23) 35,36 . Since the intricate sugar molecules produced through formose reactions provided an important source of carbohydrate for early life [37][38][39] , the ability to catabolize various carbohydrates by PK might have been of great significance for life survival during the ancient reductive world.…”

Section: Main Textmentioning

confidence: 99%

Designing a Minimal Artificial Glycolytic Pathway

Yang

Liu

Zhang

et al. 2022

Preprint

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The canonical glycolysis generates two molecules of acetyl-coenzyme A (acetyl-CoA) from one glucose through eleven cascade biochemical reactions. Here, we designed and constructed a Minimal Artificial Glycolytic (MAG) pathway consisting of only three types of biochemical reactions, with phosphoketolase as the core, combined with phosphatase and isomerase as auxiliary enzymes. It could theoretically achieve a 100% carbon yield to acetyl-CoA from any monosaccharide by integrating one-carbon condensation reaction. We tested the MAG pathway in vitro and in vivo, demonstrating the catabolism of typical C1-C6 carbohydrates to acetyl-CoA with yields from 82% to 95%. This novel glycolytic pathway provides a promising route for biomanufacturing with stoichiometric productivity from multiple carbon sources in the future.

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Role of cyanobacterial phosphoketolase in energy regulation and glucose secretion under dark anaerobic and osmotic stress conditions

Cited by 21 publications

References 25 publications

Optimal energy and redox metabolism in the cyanobacterium Synechocystis sp. PCC 6803

Optimal energy and redox metabolism in the cyanobacterium Synechocystis sp. PCC 6803

Metabolic and Microbial Community Engineering for Four-Carbon Dicarboxylic Acid Production from CO₂-Derived Glycogen in the Cyanobacterium Synechocystis sp. PCC6803

Designing a Minimal Artificial Glycolytic Pathway

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Role of cyanobacterial phosphoketolase in energy regulation and glucose secretion under dark anaerobic and osmotic stress conditions

Cited by 21 publications

References 25 publications

Optimal energy and redox metabolism in the cyanobacterium Synechocystis sp. PCC 6803

Optimal energy and redox metabolism in the cyanobacterium Synechocystis sp. PCC 6803

Metabolic and Microbial Community Engineering for Four-Carbon Dicarboxylic Acid Production from CO2-Derived Glycogen in the Cyanobacterium Synechocystis sp. PCC6803

Designing a Minimal Artificial Glycolytic Pathway

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Metabolic and Microbial Community Engineering for Four-Carbon Dicarboxylic Acid Production from CO₂-Derived Glycogen in the Cyanobacterium Synechocystis sp. PCC6803