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Magnesium modulates the stress response of oral streptococci to environmental and antibiotic challenges by altering cell envelope and nutrient transport pathways

ABSTRACT: Magnesium supplementation increases the tolerance of various Streptococcus species to environmental stressors, specifically osmotic and Zn-excess. In order to understand the mechanism of Mg-dependent stress tolerance, a putative Mg2+-efflux pump gene, hlyX, has been deleted in Streptococcus mutans. The effects of Mg2+ supplementation on wild-type and Delta-hlyX strains regarding growth, stress tolerance, and intracellular metal concentrations were compared to elucidate the role of HlyX in Mg2+ efflux. Proteomic and lipidomic analyses of the wild-type and Delta-hlyX strains in the presence of 25 mM MgCl2 were compared with those grown in unsupplemented media. Proteomic and lipidomic analyses revealed altered levels of amino acid transporters, cell envelope proteins, and an increase in long-chain unsaturated fatty acids. Furthermore, modulating intracellular Mg2+ concentration, either by MgCl2 supplementation or by eliminating HlyX, impacted the efficacy of multiple cell wall-targeting antibiotics.

INSTRUMENT(S): ULTIMATE 3000 nano UHPLC

ORGANISM(S): Homo Sapiens (ncbitaxon:9606)

SUBMITTER: Surabhi Mishra

PROVIDER: MSV000099768 | MassIVE | Wed Nov 05 13:13:00 GMT 2025

SECONDARY ACCESSION(S): PXD070376

REPOSITORIES: MassIVE

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Project description:Extracellular membrane vesicles (EMVs) are produced by many Gram-positive organisms, but information regarding vesiculogenesis is incomplete. We used single gene deletions to evaluate the impacts on Streptococcus mutans EMV biogenesis of Sortase A (SrtA), which affects S. mutans EMV composition, and Sfp, a 4'-phosphopantetheinyl transferase that affects Bacillus subtilis EMV stability. ΔsrtA EMVs were notably larger than Δsfp and wild-type (WT) EMVs. EMV proteins identified from all three strains are known to be involved in cell wall biogenesis and cell architecture, bacterial adhesion, biofilm cell density and matrix development, and microbial competition. Notably, the AtlA autolysin was not processed to its mature active form in the ΔsrtA mutant. Proteomic and lipidomic analyses of all three strains revealed multiple dissimilarities between vesicular and corresponding cytoplasmic membranes (CMs). A higher proportion of EMV proteins are predicted substrates of the general secretion pathway (GSP). Accordingly, the GSP component SecA was identified as a prominent EMV-associated protein. In contrast, CMs contained more multi-pass transmembrane (TM) protein substrates of co-translational transport machineries than EMVs. EMVs from the WT, but not the mutant strains, were enriched in cardiolipin compared to CMs, and all EMVs were over-represented in polyketide flavonoids. EMVs and CMs were rich in long-chain saturated, monounsaturated, and polyunsaturated fatty acids, except for Δsfp EMVs that contained exclusively polyunsaturated fatty acids. Lipoproteins were less prevalent in EMVs of all three strains compared to their CMs. This study provides insight into biophysical characteristics of S. mutans EMVs and indicates discrete partitioning of protein and lipid components between EMVs and corresponding CMs of WT, ΔsrtA, and Δsfp strains. Data availability:The proteomics data have been deposited into the ProteomeXchange Consortium via the PRIDE partner repository with the data set identifier <a href='https://www.ebi.ac.uk/pride/archive/projects/PXD019825' rel='noopener noreferrer' target='_blank'>PXD019825</a>.

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Magnesium modulates the stress response of oral streptococci to environmental and antibiotic challenges by altering cell envelope and nutrient transport pathways

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