Translation error clusters induced by aminoglycoside antibiotics.
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ABSTRACT:
SUBMITTER: Wohlgemuth I
PROVIDER: S-EPMC7987974 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Aminoglycoside antibiotics target the ribosome and induce mistranslation, yet which translation errors induce bacterial cell death is unclear. The analysis of cellular proteins by quantitative mass spectrometry shows that bactericidal aminoglycosides induce not only single errors, but also clusters of errors in full-length proteins in vivo with as much as four amino acid substitutions in a row. The downstream errorsin a cluster are up to 10,000-fold more frequent than the first error and independent of the intracellular aminoglycoside concentration. The prevalence, length, and composition of error clusters dependsnot only on the misreading propensity of a given aminoglycoside, but also on its ability to inhibit ribosome translocation along the mRNA. Error clusters constitute a new class of misreading events in vivo that may provide the predominant source of proteotoxic stress at low aminoglycoside concentration, which is particularly important for the autocatalytic uptake of the drugs.
2021-04-06 | PXD022098 | Pride
Project description:Aminoglycoside antibiotics (AGAs) target the ribosome and induce mistranslation, yet which translation errors induce bacterial cell death is unclear. The analysis of cellular proteins by quantitative mass spectrometry shows that bactericidal AGAs induce not only single translation errors, but also clusters of errors in full-length proteins in vivo with as much as four amino acid substitutions in a row. The downstream errors in a cluster are much more frequent than the first error (0.4 vs. 10-3, respectively) and independent of the intracellular AGA concentration. The prevalence, length, and composition of error clusters depends not only on the misreading propensity of a given AGA, but also on its ability to inhibit ribosome translocation along the mRNA. Error clusters constitute a new class of misreading events in vivo that may constitute the predominant source of proteotoxic stress at low AGA concentration, which is particularly important for the autocatalytic uptake of the drugs.
2021-04-06 | PXD019188 | Pride
Project description:Aminoglycoside antibiotics (AGAs) target the ribosome and induce mistranslation, yet which translation errors induce bacterial cell death is unclear. The analysis of cellular proteins by quantitative mass spectrometry shows that bactericidal AGAs induce not only single translation errors, but also clusters of errors in full-length proteins in vivo with as much as four amino acid substitutions in a row. The downstream errors in a cluster are much more frequent than the first error (0.4 vs. 10-3, respectively) and independent of the intracellular AGA concentration. The prevalence, length, and composition of error clusters depends not only on the misreading propensity of a given AGA, but also on its ability to inhibit ribosome translocation along the mRNA. Error clusters constitute a new class of misreading events in vivo that may constitute the predominant source of proteotoxic stress at low AGA concentration, which is particularly important for the autocatalytic uptake of the drugs.
2021-04-06 | PXD019328 | panorama
Project description:Aminoglycoside antibiotics interfere with the selection of cognate tRNAs during translation, resulting in the synthesis of aberrant proteins that are the ultimate cause of cell death. However, the toxic potential of aberrant proteins and how they avoid degradation by the cell's protein quality control (QC) machinery are not understood. Here we report that levels of the heat shock (HS) transcription factor σ32 increased sharply following exposure of Escherichia coli to the aminoglycoside kanamycin (Kan), suggesting that at least some of the aberrant proteins synthesized in these cells were recognized as substrates by DnaK, a molecular chaperone that regulates the HS response, the major protein QC pathway in bacteria. To further investigate aberrant protein toxic potential and interaction with cell QC factors, we studied an acutely toxic 48-residue polypeptide (ARF48) that is encoded by an alternate reading frame in a plant cDNA. As occurred in cells exposed to Kan, σ32 levels were strongly elevated following ARF48 expression, suggesting that ARF48 was recognized as a substrate by DnaK. Paradoxically, an internal 10-residue region that was tightly bound by DnaK in vitro also was required for the ARF48 toxic effect. Despite the increased levels of σ32, levels of several HS proteins were unchanged following ARF48 expression, suggesting that the HS response had been aborted. Nucleoids were condensed and cell permeability increased rapidly following ARF48 expression, together suggesting that ARF48 disrupts DNA-membrane interactions that could be required for efficient gene expression. Our results are consistent with earlier studies showing that aberrant proteins induced by aminoglycoside antibiotics disrupt cell membrane integrity. Insights into the mechanism for this effect could be gained by further study of the ARF48 model system.
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Project description:Antibiotic resistance is becoming the biggest threat to global health. At the same time, phage therapy is witnessing a return of interest. The therapeutic use of bacteriophages that infect and kill bacteria is a suitable strategy to combat antibiotic resistance. Furthermore, bacteriophages are increasingly used in combination with standard antibiotics against drug-resistant pathogens. Interestingly, we found that the engineered mycobacteriophage phAE159 and natural phage D29 cannot infect the Mycobacterium tuberculosis in the presence of kanamycin, hygromycin or streptomycin, but the phage infection was not affected in the presence of spectinomycin. Based on a series of studies and structural analysis of the above four aminoglycoside antibiotics, it could be speculated that the amino sugar group of aminoglycoside might selectively inhibit mycobacteriophage DNA replication. Our discovery that broad-spectrum antibiotics inhibit phage infection is of great value. This study will provide guidance for people to combine phage and antibiotics to treat M. tuberculosis.
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Project description:The condensing effect of aminoglycoside antibiotics on the structure of double-stranded DNA was examined. The selective condensation of DNA by small molecules is an interesting approach in biotechnology. Here, we present the interaction between calf thymus DNA and three types of antibiotic molecules: tobramycin, kanamycin, and neomycin. Several techniques were applied to study this effect. Atomic force microscopy, transmission electron microscopy images, and nuclear magnetic resonance spectra showed that the interaction of tobramycin with double-stranded DNA caused the rod, toroid, and sphere formation and very strong condensation of DNA strands, which was not observed in the case of other aminoglycosides used in the experiment. Studies on the mechanisms by which small molecules interact with DNA are important in understanding their functioning in cells, in designing new and efficient drugs, or in minimizing their adverse side effects. Specific interactions between tobramycin and DNA double helix was modeled using molecular dynamics simulations. Simulation study shows the aminoglycoside specificity to bend DNA double helix, shedding light on the origins of toroid formation. This phenomenon may lighten the ototoxicity or nephrotoxicity issues, but also other adverse reactions of aminoglycoside antibiotics in the human body.
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Project description:Global multidrug-resistant (MDR) bacteria are spreading rapidly and causing a great threat to human health due to the abuse of antibiotics. Determining how to resensitize MDR bacteria to conventional inefficient antibiotics is of extreme urgency. Here, a low-temperature photothermal treatment (PTT, 45 °C) is utilized with red phosphorus nanoparticles to resensitize methicillin-resistant Staphylococcus aureus (MRSA) to conventional aminoglycoside antibiotics. The antibacterial mechanism is studied by the proteomic technique and molecular dynamics (MD) simulation, which proves that the aminoglycoside antibiotics against MRSA can be selectively potentiated by low-temperature PTT. The catalytic activity of 2-aminoglycoside phosphotransferase (APH (2?))-a modifying enzyme-is demonstrated to be obviously inhibited via detecting the consumption of adenosine triphosphate (ATP) in the catalytic reaction. It is also found that the active site of aspartic acid (ASP) residues in APH (2?) is thermally unstable from the results of molecular dynamics simulation. Its catalytic ability is inhibited by preventing the deprotonating procedure for the target -OH of gentamycin. The combined therapy also exhibits great biocompatibility and successfully treats MRSA infections in vivo. This low-temperature PTT strategy has the potential to be an exogenous-modifying enzyme inhibitor for the treatment of MDR bacterial infection.
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Project description:The cochlea and kidney are susceptible to aminoglycoside-induced toxicity. The non-selective cation channel TRPV4 is expressed in kidney distal tubule cells, and hair cells and the stria vascularis in the inner ear. To determine whether TRPV4 is involved in aminoglycoside trafficking, we generated a murine proximal-tubule cell line (KPT2) and a distal-tubule cell line (KDT3). TRPV4 expression was confirmed in KDT3 cells but not in KPT2 cells. Removal of extracellular Ca(2+) significantly enhanced gentamicin-Texas-Red (GTTR) uptake by KDT3, indicative of permeation through non-selective cation channels. To determine whether TRPV4 is permeable to GTTR, stable cell lines were generated that express TRPV4 in KPT2 (KPT2-TRPV4). KPT2-TRPV4 cells took up more GTTR than control cell lines (KPT2-pBabe) in the absence of extracellular Ca(2+). TRPV4-dependent GTTR uptake was abolished by a point mutation within the crucial pore region of the channel, suggesting that GTTR permeates the TRPV4 channel. In an endolymph-like extracellular environment, clearance of GTTR was attenuated from KPT2-TRPV4 cells in a TRPV4-dependent fashion. We propose that TRPV4 has a role in aminoglycoside uptake and retention in the cochlea.
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Project description:Clinical use of 2-deoxystreptamine aminoglycoside antibiotics, which target the bacterial ribosome, is compromised by adverse effects related to limited drug selectivity. Here we present a series of 4',6'-O-acetal and 4'-O-ether modifications on glucopyranosyl ring I of aminoglycosides. Chemical modifications were guided by measuring interactions between the compounds synthesized and ribosomes harbouring single point mutations in the drug-binding site, resulting in aminoglycosides that interact poorly with the drug-binding pocket of eukaryotic mitochondrial or cytosolic ribosomes. Yet, these compounds largely retain their inhibitory activity for bacterial ribosomes and show antibacterial activity. Our data indicate that 4'-O-substituted aminoglycosides possess increased selectivity towards bacterial ribosomes and little activity for any of the human drug-binding pockets.
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Project description:The effects of aminoglycoside antibiotics on phospholipase D (PLD) activity were investigated in permeabilized NG108-15 cells and in isolated rat brain membranes. Neomycin inhibited guanosine 5'-[gamma-thio]triphosphate-stimulated PLD activity in digitonin-permeabilized NG108-15 cells in a concentration-dependent manner (50% inhibition at 100 microM). Neomycin similarly inhibited PLD activity present in rat brain membranes and assayed in vitro with [3H]phosphatidylcholine as substrate (50% inhibition at 65 microM). Other aminoglycosides tested (kanamycin, geneticin and streptomycin) were nearly equipotent inhibitors of rat brain PLD. These results indicate that aminoglycoside antibiotics inhibit phosphatidylcholine-PLD activity with comparable and sometimes greater potency than their well known inhibition of phosphoinositide-phospholipase C. The possibility that PLD inhibition could mediate some of the toxic side effects of aminoglycosides is suggested.
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