Purine biosynthesis is the bottleneck in trimethoprim‐treated <i>Bacillus subtilis</i>

Stepanek, Jennifer Janina; Schäkermann, Sina; Wenzel, Michaela; Prochnow, Pascal; Bandow, Julia E.

doi:10.1002/prca.201600039

Cited by 21 publications

(21 citation statements)

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“…To identify commonalities of the physiological impact of allicin to that of other antibacterial agents, we compared the proteomic response to allicin to a response library containing 44 proteomic responses of B. subtilis to known antimicrobial agents . Regarding the number of induced proteins, with 56 marker proteins, allicin evoked the most complex response in B. subtilis among the tested compounds.…”

Section: Resultsmentioning

confidence: 99%

Interspecies Comparison of the Bacterial Response to Allicin Reveals Species‐Specific Defense Strategies

et al. 2019

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Allicin, a broad‐spectrum antimicrobial agent from garlic, disrupts thiol and redox homeostasis, proteostasis, and cell membrane integrity. Since medicine demands antimicrobials with so far unexploited mechanisms, allicin is a promising lead structure. While progress is being made in unraveling its mode of action, little is known on bacterial adaptation strategies. Some isolates of Pseudomonas aeruginosa and Escherichia coli withstand exposure to high allicin concentrations due to as yet unknown mechanisms. To elucidate resistance and sensitivity‐conferring cellular processes, the acute proteomic responses of a resistant P. aeruginosa strain and the sensitive species Bacillus subtilis are compared to the published proteomic response of E. coli to allicin treatment. The cellular defense strategies share functional features: proteins involved in translation and maintenance of protein quality, redox homeostasis, and cell envelope modification are upregulated. In both Gram‐negative species, protein synthesis of the majority of proteins is downregulated while the Gram‐positive B. subtilis responded by upregulation of multiple regulons. A comparison of the B. subtilis proteomic response to a library of responses to antibiotic treatment reveals 30 proteins specifically upregulated by allicin. Upregulated oxidative stress proteins are shared with nitrofurantoin and diamide. Microscopy‐based assays further indicate that in B. subtilis cell wall integrity is impaired.

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Section: Resultsmentioning

confidence: 99%

Interspecies Comparison of the Bacterial Response to Allicin Reveals Species‐Specific Defense Strategies

et al. 2019

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“…Inhibition of THF biosynthesis leads to cell death as a result of purine auxotrophy and consequent thymine deficiency (“thymineless death”) (24). In B. subtilis , purine biosynthesis is the primary bottleneck caused by 10f-THF depletion after treatment with antifolates, such as trimethoprim (TMP) (25). 10f-THF is used as a formyl group donor at two steps in purine biosynthesis (Fig 1C).…”

Section: Resultsmentioning

confidence: 99%

Activation of a zinc metallochaperone by the alarmone ZTP

Chandrangsu

Huang

Helmann

2019

Preprint

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20Bacteria tightly regulate intracellular zinc levels to ensure sufficient zinc to support 21 essential functions, while preventing toxicity. The bacterial response to zinc limitation 22Here, we show that the Bacillus subtilis YciC (renamed ZagA) protein responds to ZTP 42 as an activating ligand rather than GTP, and interacts specifically with a Zn-dependent 43 enzyme critical for folate synthesis (FolE). Thus, under conditions of Zn limitation ZagA 44 is synthesized, and as folate synthesis fails, it selectively delivers Zn to FolE to sustain 45 folate synthesis. 46 47

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“…Even in the presence of CAA, all colonies were small, and only about 20% of those growing on trimethoprim (6 g/ml) carried a stable thyB insertion. Work in E. coli as well as in B. subtilis has shown that disruption of the folate pathway can lead to depletion of key amino acids as well as of purines and pyrimidines (33,34). Trimethoprim-mediated inactivation of dihydrofolic acid reductase (DHFR) would deplete intracellular levels of tetrahydrofolic acid (THF), methylenetetrahydrofolic acid (MTHF), and dihydrofolate (DHF).…”

Section: Resultsmentioning

confidence: 99%

Biological Containment of Genetically Modified Bacillus subtilis

Hosseini

Curilovs

Cutting

2018

Appl Environ Microbiol

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Genetic manipulation of bacterial spores of the genus has shown potential for vaccination and for delivery of drugs or enzymes. Remarkably, proteins displayed on the spore surface retain activity and generally are not degraded. The heat stability of spores coupled with their desiccation resistance makes them suitable for delivery to humans or to animals by the oral route. Despite these attributes one regulatory obstacle has remained regarding the fate of recombinant spores shed into the environment as viable spores. We have addressed the biological containment of spore GMOs by utilizing the concept of a 'thymine-less death', a phenomenon first reported six decades ago. Using we have inserted chimeric genes in the two thymidylate synthase genes, and, using a two-step process. Insertion is made first at followed by where resistance to trimethoprim enables selection of recombinants. Importantly, this method requires introduction of no new antibiotic resistance genes. Recombinant spores have a strict dependence on thymine (or thymidine) and in their absence cells lyse and die. Insertions are stable with no evidence for suppression or reversion. Using this system we have successfully created a number of spore vaccines as well as spores displaying active enzymes. Genetic manipulation of bacterial spores offers a number of exciting possibilities for public and animal health including their use as heat stable vehicles for delivering vaccines or enzymes. Despite this, one remaining problem is the fate of recombinant spores if released to the environment where they could survive in a dormant form indefinitely. We describe a solution whereby following genetic manipulation the bacterium is rendered dependent on thymine. As a consequence spores if released would produce bacteria unable to survive and they would exhibit a thymine-less death due to rapid cessation of metabolism. The method we describe has been validated using a number of exemplars and solves a problem for containing spore GMOs in the environment.

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Purine biosynthesis is the bottleneck in trimethoprim‐treated Bacillus subtilis

Abstract: The frequently prescribed antibiotic trimethoprim causes purine depletion in B. subtilis, which can be complemented by supplementing purines to the medium.

Cited by 21 publications

References 62 publications

Interspecies Comparison of the Bacterial Response to Allicin Reveals Species‐Specific Defense Strategies

Interspecies Comparison of the Bacterial Response to Allicin Reveals Species‐Specific Defense Strategies

Activation of a zinc metallochaperone by the alarmone ZTP

Biological Containment of Genetically Modified Bacillus subtilis

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