Sugar-Phosphate Toxicities

Boulanger, Erin F; Sabag-Daigle, Anice; Thirugnanasambantham, Pankajavalli; Gopalan, Venkat; Ahmer, Brian M. M.

doi:10.1128/mmbr.00123-21

Cited by 30 publications

(24 citation statements)

References 226 publications

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“…Deletion mutants in ptsI completely rescued the Δ pgi SF sensitivity phenotype and morphological defects ( Fig.4B-C ), while Δ treB promoted partial restoration of morphology and cell viability, with complete restoration of intrinsic resistance as measured by ZOI assay ( Fig.4B-C ). Consistent with previous work in E. coli and S. enterica (29), our suppressor analysis thus overwhelmingly suggests that the pgi mutant suffers from sugar phosphate toxicity, which can be mitigated by curbing sugar uptake.…”

Section: Resultssupporting

confidence: 90%

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Disrupting central carbon metabolism increases antibiotic susceptibility inVibrio cholerae

Keller

Han

Dörr

2022

Preprint

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Antibiotic tolerance, the ability of bacteria to sustain viability in the presence of typically bactericidal antibiotics for extended time periods, is an understudied contributor to treatment failure. The Gram-negative pathogen Vibrio cholerae, the causative agent of cholera disease, becomes highly tolerant to β-lactam antibiotics (penicillin and related compounds) in a process requiring the two-component system VxrAB. VxrAB is induced by exposure to cell wall damage conditions, which results in the differential regulation of >100 genes. While the effectors of VxrAB are relatively well-known, VxrAB environment-sensing and activation mechanisms remain a mystery. Here, we used transposon mutagenesis to screen for mutants that spontaneously upregulate VxrAB signaling. This screen was answered by genes known to be required for proper cell envelope homeostasis, validating the approach. Unexpectedly, we also uncovered a new connection between central carbon metabolism and antibiotic tolerance. Inactivation of pgi (vc0374, coding for Glucose-6-phosphate isomerase) resulted in an intracellular accumulation of glucose-6-phosphate and fructose-6-phosphate, concomitant with a marked cell envelope defect, resulting in VxrAB induction. Deletion of pgi also increased sensitivity to β-lactams and conferred a growth defect on salt-free LB; phenotypes that could be suppressed by deleting sugar uptake systems and by supplementing cell wall precursors in the growth medium. Our data suggest an important connection between central metabolism and cell envelope integrity and highlight a potential new target for developing novel antimicrobial agents.

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Section: Resultssupporting

confidence: 90%

“…4B-C). Consistent with previous work in E. coli and S. enterica (29), our suppressor analysis thus overwhelmingly suggests that the pgi mutant suffers from sugar phosphate toxicity, which can be mitigated by curbing sugar uptake.…”

Section: Suppressor Mutations In Sugar Transport System Components Re...supporting

confidence: 90%

Disrupting central carbon metabolism increases antibiotic susceptibility inVibrio cholerae

Keller

Han

Dörr

2022

Preprint

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“…In this context, the GlcN-1P switch-on mechanism exerted by the corresponding enzyme activators (specifically GlcN-6P) might be a strategy to cope with temporary GlcN phosphates accumulation. It has been demonstrated that increased intracellular hexose-P concentrations could be toxic and detrimental to cell viability ( Boulanger et al, 2021 ).…”

Section: Discussionmentioning

confidence: 99%

Carbohydrate Metabolism in Bacteria: Alternative Specificities in ADP-Glucose Pyrophosphorylases Open Novel Metabolic Scenarios and Biotechnological Tools

et al. 2022

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We explored the ability of ADP-glucose pyrophosphorylase (ADP-Glc PPase) from different bacteria to use glucosamine (GlcN) metabolites as a substrate or allosteric effectors. The enzyme from the actinobacteria Kocuria rhizophila exhibited marked and distinctive sensitivity to allosteric activation by GlcN-6P when producing ADP-Glc from glucose-1-phosphate (Glc-1P) and ATP. This behavior is also seen in the enzyme from Rhodococcus spp., the only one known so far to portray this activation. GlcN-6P had a more modest effect on the enzyme from other Actinobacteria (Streptomyces coelicolor), Firmicutes (Ruminococcus albus), and Proteobacteria (Agrobacterium tumefaciens) groups. In addition, we studied the catalytic capacity of ADP-Glc PPases from the different sources using GlcN-1P as a substrate when assayed in the presence of their respective allosteric activators. In all cases, the catalytic efficiency of Glc-1P was 1–2 orders of magnitude higher than GlcN-1P, except for the unregulated heterotetrameric protein (GlgC/GgD) from Geobacillus stearothermophilus. The Glc-1P substrate preference is explained using a model of ADP-Glc PPase from A. tumefaciens based on the crystallographic structure of the enzyme from potato tuber. The substrate-binding domain localizes near the N-terminal of an α-helix, which has a partial positive charge, thus favoring the interaction with a hydroxyl rather than a charged primary amine group. Results support the scenario where the ability of ADP-Glc PPases to use GlcN-1P as an alternative occurred during evolution despite the enzyme being selected to use Glc-1P and ATP for α-glucans synthesis. As an associated consequence in such a process, certain bacteria could have improved their ability to metabolize GlcN. The work also provides insights in designing molecular tools for producing oligo and polysaccharides with amino moieties.

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“…We also hypothesize that modulation of purine biosynthesis can be one direction to counteract redox imbalance. Another interesting implication of our work is that reductive stress can also result in the accumulation of non-canonical sugar phosphates, which are known to be toxic (59).…”

Section: Discussionmentioning

confidence: 94%

A genetically encoded tool to increase cellular NADH/NAD⁺ ratio in living cells

Heacock

Abdulaziz

Pan

et al. 2022

Preprint

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Impaired reduction/oxidation (redox) metabolism is a key contributor to the etiology of many diseases, including primary mitochondrial disorders, cancer, neurodegeneration, and aging. However, mechanistic studies of redox imbalance remain challenging due to limited strategies which can perturb cellular redox metabolism and model pathology in various cellular, tissue, or organismal backgrounds without creating additional and potentially confounding metabolic perturbations. To date, most studies involving impaired redox metabolism have focused on oxidative stress and reactive oxygen species (ROS) production; consequently, less is known about the settings where there is an overabundance of reducing equivalents, termed reductive stress. NADH reductive stress has been modeled using pharmacologic inhibition of the electron transport chain (ETC) and ethanol supplementation. Still, both these methods have significant drawbacks. Here, we introduce a soluble transhydrogenase from E. coli (EcSTH) as a novel genetically encoded tool to promote NADH overproduction in living cells. When expressed in mammalian cells, EcSTH, and a mitochondrially-targeted version (mitoEcSTH), can elevate the NADH/NAD+ ratio in a compartment-specific manner. Using this tool, we determine the metabolic and transcriptomic signatures of NADH reductive stress in mammalian cells. We also find that cellular responses to NADH reductive stress, including blunted proliferation, are dependent on cellular background and identify the metabolic reactions that sense changes in the cellular NADH/NAD+ balance. Collectively, our novel genetically encoded tool represents an orthogonal strategy to perturb redox metabolism and characterize the impact on normal physiology and disease states.

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Sugar-Phosphate Toxicities

Cited by 30 publications

References 226 publications

Disrupting central carbon metabolism increases antibiotic susceptibility inVibrio cholerae

Disrupting central carbon metabolism increases antibiotic susceptibility inVibrio cholerae

Carbohydrate Metabolism in Bacteria: Alternative Specificities in ADP-Glucose Pyrophosphorylases Open Novel Metabolic Scenarios and Biotechnological Tools

A genetically encoded tool to increase cellular NADH/NAD⁺ ratio in living cells

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Sugar-Phosphate Toxicities

Cited by 30 publications

References 226 publications

Disrupting central carbon metabolism increases antibiotic susceptibility inVibrio cholerae

Disrupting central carbon metabolism increases antibiotic susceptibility inVibrio cholerae

Carbohydrate Metabolism in Bacteria: Alternative Specificities in ADP-Glucose Pyrophosphorylases Open Novel Metabolic Scenarios and Biotechnological Tools

A genetically encoded tool to increase cellular NADH/NAD+ ratio in living cells

Contact Info

Product

Resources

About

A genetically encoded tool to increase cellular NADH/NAD⁺ ratio in living cells