Resolution of an ATP-metal chelate from metal-free ATP by reverse-phase high-performance liquid chromatography

Jahngen, Jessica H.; Rossomando, Edward F.

doi:10.1016/0003-2697(83)90608-5

Cited by 13 publications

(5 citation statements)

References 25 publications

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“…It also appears that this channel exhibits different permeability states (Yan et al, 2008), which further complicates the understanding of coupling of conformational changes in the orthosteric binding domains with the corresponding changes at the TM channel gate. The binding of Ca 2+ and Mg 2+ to β- and γ-phosphate groups of ATP is a well-established phenomenon (Jahngen and Rossomando, 1983), but at the present time it is unknown which form of ATP (ATP 4− and/or Mg/Ca-ATP) acts as an orthosteric agonist for this receptor (Jiang, 2009). It has also not been clarified whether these two macroelements affect the receptor function by acting as allosteric modulators, as earlier data suggest (Virginio et al, 1997), and/or by altering the permeability dynamics, as shown for P2X4R (Khakh et al, 1999).…”

Section: Introductionmentioning

confidence: 99%

Calcium-dependent block of P2X7 receptor channel function is allosteric

Yan

Khadra

Sherman

et al. 2011

Journal of General Physiology

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Among purinergic P2X receptor (P2XR) channels, the P2X7R exhibits the most complex gating kinetics; the binding of orthosteric agonists at the ectodomain induces a conformational change in the receptor complex that favors a gating transition from closed to open and dilated states. Bath Ca2+ affects P2X7R gating through a still uncharacterized mechanism: it could act by reducing the adenosine triphosphate4− (ATP4−) concentration (a form proposed to be the P2X7R orthosteric agonist), as an allosteric modulator, and/or by directly altering the selectivity of pore to cations. In this study, we combined biophysical and mathematical approaches to clarify the role of calcium in P2X7R gating. In naive receptors, bath calcium affected the activation permeability dynamics indirectly by decreasing the potency of orthosteric agonists in a concentration-dependent manner and independently of the concentrations of the free acid form of agonists and status of pannexin-1 (Panx1) channels. Bath calcium also facilitated the rates of receptor deactivation in a concentration-dependent manner but did not affect a progressive delay in receptor deactivation caused by repetitive agonist application. The effects of calcium on the kinetics of receptor deactivation were rapid and reversible. A438079, a potent orthosteric competitive antagonist, protected the rebinding effect of 2’(3′)-O-4-benzoylbenzoyl)ATP on the kinetics of current decay during the washout period, but in the presence of A438079, calcium also increased the rate of receptor deactivation. The corresponding kinetic (Markov state) model indicated that the decrease in binding affinity leads to a decrease in current amplitudes and facilitation of receptor deactivation, both in an extracellular calcium concentration–dependent manner expressed as a Hill function. The results indicate that calcium in physiological concentrations acts as a negative allosteric modulator of P2X7R by decreasing the affinity of receptors for orthosteric ligand agonists, but not antagonists, and not by affecting the permeability dynamics directly or indirectly through Panx1 channels. We expect these results to generalize to other P2XRs.

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Section: Introductionmentioning

confidence: 99%

Calcium-dependent block of P2X7 receptor channel function is allosteric

Yan

Khadra

Sherman

et al. 2011

Journal of General Physiology

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“…Hon et al 31 previously showed that magnesium stimulates the activity of T. saccharolyticum aldehyde dehydrogenase. Since ATP is known to chelate magnesium 32 , it is possible that the observed inhibition is due to chelation of magnesium present in the cell extracts. Whether such chelation could happen following a spike in ATP levels in vivo is unknown, though a recent structural study of ATP inhibition of a human aldehyde dehydrogenase showed a complex interplay of ATP and magnesium binding 33 .…”

Section: Resultsmentioning

confidence: 99%

Growth-uncoupled propanediol production in a Thermoanaerobacterium thermosaccharolyticum strain engineered for high ethanol yield

Herring

Ajie

Lynd

2023

Sci Rep

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Cocultures of engineered thermophilic bacteria can ferment lignocellulose without costly pretreatment or added enzymes, an ability that can be exploited for low cost biofuel production from renewable feedstocks. The hemicellulose-fermenting species Thermoanaerobacterium thermosaccharolyticum was engineered for high ethanol yield, but we found that the strains switched from growth-coupled production of ethanol to growth uncoupled production of acetate and 1,2-propanediol upon growth cessation, producing up to 6.7 g/L 1,2-propanediol from 60 g/L cellobiose. The unique capability of this species to make 1,2-propanediol from sugars was described decades ago, but the genes responsible were not identified. Here we deleted genes encoding methylglyoxal reductase, methylglyoxal synthase and glycerol dehydrogenase. Deletion of the latter two genes eliminated propanediol production. To understand how carbon flux is redirected in this species, we hypothesized that high ATP levels during growth cessation downregulate the activity of alcohol and aldehyde dehydrogenase activities. Measurements with cell free extracts show approximately twofold and tenfold inhibition of these activities by 10 mM ATP, supporting the hypothesized mechanism of metabolic redirection. This result may have implications for efforts to direct and maximize flux through alcohol dehydrogenase in other species.

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“…Therefore, the involvement of ATP in cell death is difficult to comprehend. However, ATP is known to bind and chelate various divalent metal ions (48–50) and intriguingly, a study reported growth inhibitory activity of exogenously supplied ATP on multiple bacterial species, which was rescued upon co-treatment with magnesium and iron salts (51). Based on these studies, we hypothesized that the ATP surge observed upon antibiotic treatment may have sequestered and deprived the macromolecules and cellular processes of their essential divalent metal ions, resulting in cell death.…”

Section: Resultsmentioning

confidence: 99%

ATP burst is the dominant driver of antibiotic lethality in Mycobacteria

Lodhiya,

Palande,

Veeram

et al. 2023

Preprint

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Antibiotic-tolerant bacteria, due to their unique physiology, are refractory to antimicrobial killing and pose challenges for successful therapy.M. tuberculosis, the causative agent of tuberculosis (TB), is exceptionally difficult to control due to antibiotic tolerance. Incomplete knowledge of how bactericidal antibiotics work has impeded our understanding of the development of antibiotic tolerance in mycobacteria. In this study, using quantitative proteomics,13C-isotopomer analysis, and biochemical assays, we investigated the physiological response ofM. smegmatischallenged with sub-lethal doses of aminoglycoside and fluoroquinolone antibiotics. Two distinct classes of antibiotics elicited remarkably similar bacterial responses in central dogma, cell division, and central carbon metabolic processes. Both antibiotics increased TCA cycle flux, resulting in enhanced respiration, reactive oxygen species (ROS) production, and ATP burst. While both ROS and ATP burst were found to contribute to antibiotic lethality, our findings indicate that ATP burst, and not ROS, is the dominant cause of antibiotic-induced cell death in mycobacteria.13C isotope tracing experiments indicated metabolic bifurcation of the TCA cycle flux from the oxidative arm into glutamate-glutamine-GABA as a potential bacterial adaptive mechanism to reduce the production of reactive oxygen species (ROS) and ATP burst. In addition, antibiotic-adapted mycobacteria have an activated intrinsic drug resistance mechanism, a higher mutation rate, and are prone to developing antibiotic resistance. Our study provides a novel understanding of the intricate mechanisms of antibiotic-induced cell death, and has significance for the design of combinatorial interventions and adjuvant therapy against one of the most successful infectious diseases.

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Resolution of an ATP-metal chelate from metal-free ATP by reverse-phase high-performance liquid chromatography

Cited by 13 publications

References 25 publications

Calcium-dependent block of P2X7 receptor channel function is allosteric

Calcium-dependent block of P2X7 receptor channel function is allosteric

Growth-uncoupled propanediol production in a Thermoanaerobacterium thermosaccharolyticum strain engineered for high ethanol yield

ATP burst is the dominant driver of antibiotic lethality in Mycobacteria

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