Criticality of a conserved tyrosine residue in the <scp>SpeG</scp> protein from <scp><i>Escherichia coli</i></scp>

Le, Van Thi Bich; Dang, Joseph; Lim, Ee Qi; Kuhn, Misty L.

doi:10.1002/pro.4078

Cited by 6 publications

(4 citation statements)

References 32 publications

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“…Therefore, if the ∆ rv1877 mutant still displays a sensitivity towards Spm despite the upregulation of rv3065 (that is supposed to detoxify Spm), then Rv1877 is indeed involved in the detoxification of Spm, though it is not the only protein involved (explaining the low sensitivity). Compensation can also occur by other mechanism of Spm inactivation/detoxification such as acetylation by SpeG as reported in other bacteria ( Limsuwun and Jones, 2000 ; Filippova et al, 2019 ; Le et al, 2021 ; Kumar et al, 2022 ). In our previous studies, we found that M.tb has a SpeG ortholog that is Rv3034c ( Sao Emani and Reiling, 2023 ).…”

Section: Discussionmentioning

confidence: 76%

The efflux pumps Rv1877 and Rv0191 play differential roles in the protection of Mycobacterium tuberculosis against chemical stress

Sao Emani,

Reiling

2024

Front. Microbiol.

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BackgroundIt was previously shown that GlnA3sc enabled Streptomyces coelicolor to survive in excess polyamines. However, subsequent studies revealed that Rv1878, the corresponding Mycobacterium tuberculosis (M.tb) ortholog, was not essential for the detoxification of spermine (Spm), in M.tb. On the other hand, the multi-drug efflux pump Rv1877 was previously shown to enable export of a wide range of compounds, while Rv0191 was shown to be more specific to chloramphenicol.RationaleTherefore, we first wanted to determine if detoxification of Spm by efflux can be achieved by any efflux pump, or if that was dependent upon the function of the pump. Next, since Rv1878 was found not to be essential for the detoxification of Spm, we sought to follow-up on the investigation of the physiological role of Rv1878 along with Rv1877 and Rv0191.ApproachTo evaluate the specificity of efflux pumps in the mycobacterial tolerance to Spm, we generated unmarked ∆rv1877 and ∆rv0191 M.tb mutants and evaluated their susceptibility to Spm. To follow up on the investigation of any other physiological roles they may have, we characterized them along with the ∆rv1878 M.tb mutant.ResultsThe ∆rv1877 mutant was sensitive to Spm stress, while the ∆rv0191 mutant was not. On the other hand, the ∆rv1878 mutant grew better than the wild-type during iron starvation yet was sensitive to cell wall stress. The proteins Rv1877 and Rv1878 seemed to play physiological roles during hypoxia and acidic stress. Lastly, the ∆rv0191 mutant was the only mutant that was sensitive to oxidative stress.ConclusionThe multidrug MFS-type efflux pump Rv1877 is required for Spm detoxification, as opposed to Rv0191 which seems to play a more specific role. Moreover, Rv1878 seems to play a role in the regulation of iron homeostasis and the reconstitution of the cell wall of M.tb. On the other hand, the sensitivity of the ∆rv0191 mutant to oxidative stress, suggests that Rv0191 may be responsible for the transport of low molecular weight thiols.

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Section: Discussionmentioning

confidence: 76%

The efflux pumps Rv1877 and Rv0191 play differential roles in the protection of Mycobacterium tuberculosis against chemical stress

Sao Emani,

Reiling

2024

Front. Microbiol.

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“…However, we searched and found Rv3170 (putative flavin-containing monoamine oxidase AofH) and Rv1751 (probable oxidoreductase) as the closest orthologs to the eukaryotic SPO when blasted against the M.tb proteome. Acetylation of Spm by SpeG and excretion of the acetylated product are another route used by Escherichia coli ( 47 , 51 – 53 ) to detoxify Spm and Spd. We found that the closest ortholog of E. coli SpeG in M.tb is Rv3034c ( Fig.…”

Section: Resultsmentioning

confidence: 99%

Spermine enhances the activity of anti-tuberculosis drugs

Sao Emani,

Reiling

2024

Microbiol Spectr

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The discovery of molecules that could shorten the treatment period of tuberculosis (TB) holds the promise of improving the current regimen. Polyamines are organic polycationic alkylamines found in millimolar concentrations in all living cells. Spermine (Spm) is a polyamine that was shown (more than 70 years ago) to be toxic to Mycobacterium tuberculosis (M.tb). In this study, we used various biochemical assays to assess the mechanism of action of Spm. Furthermore, using various drug susceptibility testing methods, we investigated the ability of Spm to enhance the activity of first-line TB drugs, isoniazid (INH) and rifampicin (RIF), as well as second-line TB drugs, para-aminosalicylic acid (PAS) and bedaquiline (BDQ). Our findings revealed that at toxic concentrations, Spm induced the formation of reactive oxygen species (ROS), a reaction that was influenced by the pH and the iron concentration of the media. Furthermore, the generation of ROS by Spm was associated with increased ferric iron and NADP levels. Moreover, the ROS produced by Spm synergized with the ROS generated by the lipophilic peroxide cumene hydroperoxide (CuOOH), as well as with the ROS generated by RIF and PAS, while enhancing their antimycobacterial activity. Additionally, we found that Spm enhanced the combined activity of INH and RIF against M.tb. Lastly, Spm was also able to enhance the activity of BDQ due to another mechanism of action. This study provides data and findings that would fuel future studies on the suitability of Spm as a potential adjunct TB therapy. The enhanced antimycobacterial activity of TB drugs in combination with Spm could offer promising new avenues for improving TB treatment outcomes. IMPORTANCE This is the first study that attempted to demonstrate the mechanisms of reactive oxygen species (ROS) generation by spermine (Spm) in Mycobacterium tuberculosis (M.tb). Furthermore, this is the first study to demonstrate that it is able to enhance the activity of currently available and World Health Organization (WHO)-approved tuberculosis (TB) drugs. Spermine can easily be obtained since it is already found in our diet. Moreover, as opposed to conventional antibiotics, it is less toxic to humans since it is found in millimolar concentrations in the body. Finally, with the difficulty of curing TB with conventional antibiotics, this study suggests that less toxic molecules, such as Spm, could in a long-term perspective be incorporated in a TB regimen to boost the treatment.

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“…40 In addition to GLO1, we also found that speG displayed upregulation in ESBLs-EC compared to susceptible Escherichia coli , which is at least partially consistent with the results from recent two studies. 41 , 42 In the future, we will continue with the study with other three upregulated genes.…”

Section: Discussionmentioning

confidence: 99%

GLO1 Contributes to the Drug Resistance of Escherichia coli Through Inducing PER Type of Extended-Spectrum β-Lactamases

Ma¹,

Lai²,

Zan³

et al. 2022

IDR

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Background Escherichia coli -associated antimicrobial resistance (AMR) issue so far needs urgent considerations. This study aims to screen the potent genes associated with extended-spectrum β-lactamases (ESBLs) in drug-resistant Escherichia coli and elucidate the specific drug-resistant mechanism. Methods Clinical ESBLs-EC samples were obtained based on the microbial identification, and the whole genome was sequenced. In combination with the significantly enriched pathways, several differently expressed genes were screened and verified by RT-PCR. Furthermore, through knocking out glyoxalase 1 (GLO1) gene and transfecting overexpressed plasmids, the potential relationship between GLO1 and ESBLs was then investigated. Lastly, the concentrations of β-lactamases in bacteria and supernatant from different groups were examined by enzyme-linked immunosorbent assay (ELISA). Results After successful isolation and identification of ESBLs-EC, the whole genome and eighteen differential metabolic pathways were analyzed to select differently expressed genes, including add, deoD, guaD, speG, GLO1, VNN1, etc. RT-PCR results showed that there were no differences in these genes between the standard bacteria and susceptible Escherichia coli . Remarkably, the relative levels of four genes including speG, Hdac10, GLO1 and Ppcdc were significantly increased in ESBLs-EC in comparison with susceptible strains, whereas other gene expression was decreased. Further experiments utilizing gene knockout and overexpression strains confirmed the role of GLO1. At last, a total of 10 subtypes of β-lactamases were studied using ELISA, including BES-, CTX-M1-, CTX-M2-, OXA1-, OXA2-, OXA10-, PER-, SHV-, TEM-, and VEB-ESBLs, and results demonstrated that GLO1 gene expression only affected PER-β-lactamases but had no effects on other β-lactamases. Conclusion SpeG, Hdac10, GLO1 and Ppcdc might be associated with the drug-resistant mechanism of Escherichia coli . Of note, this study firstly addressed the role of GLO1 in the drug resistance of ESBLs-EC, and this effect may be mediated by increasing PER-β-lactamases.

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Criticality of a conserved tyrosine residue in the SpeG protein from Escherichia coli

Cited by 6 publications

References 32 publications

The efflux pumps Rv1877 and Rv0191 play differential roles in the protection of Mycobacterium tuberculosis against chemical stress

The efflux pumps Rv1877 and Rv0191 play differential roles in the protection of Mycobacterium tuberculosis against chemical stress

Spermine enhances the activity of anti-tuberculosis drugs

GLO1 Contributes to the Drug Resistance of Escherichia coli Through Inducing PER Type of Extended-Spectrum β-Lactamases

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