Role of non-PTS dependent glucose permease (GlcU) in maintaining the fitness cost during acquisition of nisin resistance by <i>Enterococcus faecalis</i>

Kumar, Sandeep; Narayan, Kapil Singh; Shandilya, Shruti; Sood, S. K.; Kapila, Suman

doi:10.1093/femsle/fnz230

Cited by 4 publications

(6 citation statements)

References 36 publications

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“…1658003) from Biorad was used for PAGE and blotting was performed using semi-dry blotting unit (Scie-Plas Ltd., UK). Sensitive and resistant E. faecalis (Nis R -147) bacteria were previously isolated from raw buffalo milk in our lab [ 5 , 23 ]. The nisin-resistant strain used here was found to be resistant to chloramphenicol, ampicillin, ciprofloxacin, rifampicin, vancomycin, carbenicillin, linezolid, oxacillin, and fosfomycin with minimum inhibitory concentrations (MICs) varying between 4 µg/mL (ciprofloxacin) and 512 µg/mL (fosfomycin) [ 5 ].…”

Section: Methodsmentioning

confidence: 99%

“…The crude sample was loaded on 12% SDS-PAGE without heating. Anti-phosphoserine antibody was used to detect the serine phosphorylation of the HPr protein as described in [ 23 ]. The experiment was performed in triplicate and relative density was analysed using ImageJ (1.51v 9).…”

Section: Methodsmentioning

confidence: 99%

“…Therefore, bacterial kinases are also gaining pronounced attention as potential targets for antimicrobial drug discovery [ 11 , 21 , 22 ]. In our previous study, we observed an enhanced expression of hprK along with HPr-Ser46 phosphorylation in antimicrobial resistant E. faecalis strains [ 23 ]. This suggests that HPrK/P plays an important role in the fitness of antibiotic resistant E. faecalis .…”

Section: Introductionmentioning

confidence: 99%

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Identification of HPr kinase/phosphorylase inhibitors: novel antimicrobials against resistant Enterococcus faecalis

Kumar

Bhadane

Shandilya

et al. 2022

J Comput Aided Mol Des

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Enterococcus faecalis, a gram-positive bacterium, is among the most common nosocomial pathogens due to its limited susceptibility to antibiotics and its reservoir of the genes coding for virulence factors. Bacterial enzymes such as kinases and phosphorylases play important roles in diverse functions of a bacterial cell and, thus, are potential antibacterial drug targets. In Gram-positive bacteria, HPr Kinase/Phosphorylase (HPrK/P), a bifunctional enzyme is involved in the regulation of carbon catabolite repression by phosphorylating/dephosphorylating the histidine-containing phosphocarrier protein (HPr) at Ser46 residue. Deficiencies in HPrK/P function leads to severe defects in bacterial growth. This study aimed at identifying novel inhibitors of E. faecalis HPrK/P from a commercial compound library using structure-based virtual screening. The hit molecules were purchased and their effect on enzyme activity and growth of resistant E. faecalis was evaluated in vitro. Furthermore, docking and molecular dynamics simulations were performed to study the interactions of the hit compounds with HPrK/P. Among the identified hit molecules, two compounds inhibited the phosphorylation of HPr as well as significantly reduced the growth of resistant E. faecalis in vitro. These identified potential HPrK/P inhibitors open new research avenues towards the development of novel antimicrobials against resistant Gram-positive bacteria.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Identification of HPr kinase/phosphorylase inhibitors: novel antimicrobials against resistant Enterococcus faecalis

Kumar

Bhadane

Shandilya

et al. 2022

J Comput Aided Mol Des

Self Cite

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“…In both Staphylococcus aureus (58) and Streptococcus pyogenes (59), GlcU has been shown to contribute to glucose uptake under low affinity conditions, and a GlcU homolog is also present in the V583 genome (ef0928). Recent work by Kumar et al (60) showed that GlcU expression compensates for PTS-dependent glucose transport when E. faecalis is exposed to the lantibiotic nisin. Their findings indicated that glucose was shuttled through the pentose phosphate shunt pathway under GlcU-dependent conditions, as opposed to the conventional glycolytic pathway.…”

Section: Figmentioning

confidence: 99%

Influence of the Alternative Sigma Factor RpoN on Global Gene Expression and Carbon Catabolism in Enterococcus faecalis V583

Keffeler

Iyer

Parthasarathy

et al. 2021

mBio

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The alternative sigma factor σ54 has been shown to regulate the expression of a wide array of virulence-associated genes, as well as central metabolism, in bacterial pathogens. In Gram-positive organisms, the σ54 is commonly associated with carbon metabolism. In this study, we show that the Enterococcus faecalis alternative sigma factor σ54 (RpoN) and its cognate enhancer binding protein MptR are essential for mannose utilization and are primary contributors to glucose uptake through the Mpt phosphotransferase system. To gain further insight into how RpoN contributes to global transcriptional changes, we performed microarray transcriptional analysis of strain V583 and an isogenic rpoN mutant grown in a chemically defined medium with glucose as the sole carbon source. Transcripts of 340 genes were differentially affected in the rpoN mutant; the predicted functions of these genes mainly related to nutrient acquisition. These differentially expressed genes included those with predicted catabolite-responsive element (cre) sites, consistent with loss of repression by the major carbon catabolite repressor CcpA. To determine if the inability to efficiently metabolize glucose/mannose affected infection outcome, we utilized two distinct infection models. We found that the rpoN mutant is significantly attenuated in both rabbit endocarditis and murine catheter-associated urinary tract infection (CAUTI). Here, we examined a ccpA mutant in the CAUTI model and showed that the absence of carbon catabolite control also significantly attenuates bacterial tissue burden in this model. Our data highlight the contribution of central carbon metabolism to growth of E. faecalis at various sites of infection. IMPORTANCE Hospital-acquired infections account for 2 billion dollars annually in increased health care expenses and cause more than 100,000 deaths in the United States alone. Enterococci are the second leading cause of hospital-acquired infections. They form biofilms at surgical sites and are often associated with infections of the urinary tract following catheterization. Nutrient uptake and growth are key factors that influence their ability to cause disease. Our research identified a large set of genes that illuminate nutrient uptake pathways in enterococci. Perturbation of the metabolic circuit reduces virulence in a rabbit endocarditis model, as well as in catheter-associated urinary tract infection in mice. Targeting metabolic pathways that are important in infection may lead to new treatments against multidrug-resistant enterococcal infections.

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“…CHCC13235 has an SNP in a gene annotated as EAM type sugar transporter (glcU) (302C to 302T, resulting in amino acid change V101A). GlcU encodes a novel non-PTS glucose permease (Castro et al, 2009), which previously has been shown to play a role in fitness cost in nisin-resistant variants of Streptococcus faecalis (Kumar et al, 2019). In addition, CHCC13235 carries an SNP in the promoter region upstream of a LacI type regulator potentially regulating the neighboring gene maltodextrin phosphorylase, whose role in milk fermentations is unknown but was previously found to be upregulated in S. thermophilus under acidic growth conditions (Wu et al, 2019).…”

Section: Analysis Of Genetic Changes In Streptococcus Thermophilus Strains Resistant To D-cycloserinementioning

confidence: 99%

Use of Cell Envelope Targeting Antibiotics and Antimicrobial Agents as a Powerful Tool to Select for Lactic Acid Bacteria Strains With Improved Texturizing Ability in Milk Fermentations

Sørensen

Kjærbølling

Neves

et al. 2021

Front. Bioeng. Biotechnol.

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Many antibiotics and antimicrobial agents have the bacterial cell envelope as their primary target, interfering with functions such as synthesis of peptidoglycan, membrane stability and permeability, and attachment of surface components. The cell envelope is the outermost barrier of the bacterial cell, conferring protection against environmental stresses, and maintaining structural integrity and stability of the growing cell, while still allowing for required metabolism. In this work, inhibitory concentrations of several different cell envelope targeting antibiotics and antimicrobial agents were used to select for derivatives of lactic acid bacteria (LAB) with improved properties for dairy applications. Interestingly, we observed that for several LAB species a fraction of the isolates had improved milk texturizing capabilities. To further improve our understanding of the mechanisms underlying the improved rheology and to validate the efficacy of this method for strain improvement, genetic and physiological characterization of several improved derivatives was performed. The results showed that the identified genetic changes are diverse and affect also other cellular functions than the targeted cell surface. In short, this study describes a new versatile and powerful toolbox based on targeting of the cell envelope to select for LAB derivatives with improved phenotypic traits for dairy applications.

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Role of non-PTS dependent glucose permease (GlcU) in maintaining the fitness cost during acquisition of nisin resistance by Enterococcus faecalis

Cited by 4 publications

References 36 publications

Identification of HPr kinase/phosphorylase inhibitors: novel antimicrobials against resistant Enterococcus faecalis

Identification of HPr kinase/phosphorylase inhibitors: novel antimicrobials against resistant Enterococcus faecalis

Influence of the Alternative Sigma Factor RpoN on Global Gene Expression and Carbon Catabolism in Enterococcus faecalis V583

Use of Cell Envelope Targeting Antibiotics and Antimicrobial Agents as a Powerful Tool to Select for Lactic Acid Bacteria Strains With Improved Texturizing Ability in Milk Fermentations

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