Biochemical and genetic studies on two different types of erythromycin resistant mutants of Escherichia coli with altered ribosomal proteins

Hg, Wittmann; Stöffler, Georg; Apirion, David; Rosen, Larry; Tanaka, Kentaro; Tamaki, Mikio; Takata, Renkichi; Dekio, Satoshi; Otaka, Eiko

doi:10.1007/bf00333665

Cited by 167 publications

(124 citation statements)

References 34 publications

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“…This feature of drug efflux pump deficiency is intuitively obvious, but recent findings for the ribosome targeting macrolide antibiotic erythromycin suggest the existence of more subtle aspects of the interplay between drug efflux pump efficiency and drug susceptibility. An Escherichia coli mutant with an amino acid deletion in protein L22 of the large ribosomal subunit (10,11), causing reduced affinity of erythromycin to the ribosome (12), displayed reduced susceptibility to erythromycin in relation to a ribosome WT strain in a drug efflux pump proficient but the same susceptibility in a drug efflux pump deficient background (12,13). These experiments demonstrated how a mutation reducing drug affinity to a vital intracellular target may give a distinct growth advantage to drug exposed bacteria in a drug efflux pump proficient background but virtually no advantage in a drug efflux pump deficient background.…”

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confidence: 99%

Drug efflux pump deficiency and drug target resistance masking in growing bacteria

Fange

Nilsson²,

Tenson³

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

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Recent experiments have shown that drug efflux pump deficiency not only increases the susceptibility of pathogens to antibiotics, but also seems to ''mask'' the effects of mutations, that decrease the affinities of drugs to their intracellular targets, on the growth rates of drug-exposed bacteria. That is, in the presence of drugs, the growth rates of drug-exposed WT and target mutated strains are the same in a drug efflux pump deficient background, but the mutants grow faster than WT in a drug efflux pump proficient background. Here, we explain the mechanism of target resistance masking and show that it occurs in response to drug efflux pump inhibition among pathogens with high-affinity drug binding targets, low cell-membrane drug-permeability and insignificant intracellular drug degradation. We demonstrate that target resistance masking is fundamentally linked to growth-bistability, i.e., the existence of 2 different steady state growth rates for one and the same drug concentration in the growth medium. We speculate that target resistance masking provides a hitherto unknown mechanism for slowing down the evolution of target resistance among pathogens.antibiotic resistance ͉ efflux pump inhibition ͉ macrolides

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confidence: 99%

Drug efflux pump deficiency and drug target resistance masking in growing bacteria

Fange

Nilsson²,

Tenson³

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

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“…The inhibitory activity of macrolides, including erythromycin, depends on binding to a site near the polypeptide exit tunnel of the large ribosomal subunit (3, 4). Because macrolides do not bind to ribosomes with an occupied exit tunnel and cause the synthesis of 2-10 residue peptides in translation assays in vitro, it has been proposed that drug binding physically blocks elongation of nascent proteins beyond this size (5-7).Some macrolide-resistance mutations alter the ribosomal target site and prevent binding (4,8). Intriguingly, other mutations confer resistance despite the fact that macrolides still bind the mutant ribosome well (8-10).…”

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confidence: 99%

“…The same mutation makes Haemophilus influenzae resistant to numerous macrolides (14); different L22 mutations also confer macrolide resistance in other bacterial species (2). When binding has been measured, ribosomes with macrolideresistant alterations in L22 bind erythromycin with near wild-type affinity (8,10,12,15).…”

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confidence: 99%

“…Some macrolide-resistance mutations alter the ribosomal target site and prevent binding (4,8). Intriguingly, other mutations confer resistance despite the fact that macrolides still bind the mutant ribosome well (8)(9)(10).…”

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confidence: 99%

“…Intriguingly, other mutations confer resistance despite the fact that macrolides still bind the mutant ribosome well (8)(9)(10). For example, deletion of the M 82 K 83 R 84 sequence in Escherichia coli ribosomal protein L22 (⌬MKR) allows growth in the presence of high levels of erythromycin and other macrolides (11)(12)(13).…”

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confidence: 99%

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Revisiting the mechanism of macrolide-antibiotic resistance mediated by ribosomal protein L22

Moore

Sauer²

2008

Proc. Natl. Acad. Sci. U.S.A.

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Bacterial antibiotic resistance can occur by many mechanisms. An intriguing class of mutants is resistant to macrolide antibiotics even though these drugs still bind to their targets. For example, a 3-residue deletion (⌬MKR) in ribosomal protein L22 distorts a loop that forms a constriction in the ribosome exit tunnel, apparently allowing nascent-chain egress and translation in the presence of bound macrolides. Here, however, we demonstrate that ⌬MKR and wild-type ribosomes show comparable macrolide sensitivity in vitro. In Escherichia coli, we find that this mutation reduces antibiotic occupancy of the target site on ribosomes in a manner largely dependent on the AcrAB-TolC efflux system. We propose a model for antibiotic resistance in which ⌬MKR ribosomes alter the translation of specific proteins, possibly via changes in programmed stalling, and modify the cell envelope in a manner that lowers steady-state macrolide levels.ermC ͉ erythromycin ͉ ribosome ͉ TolC M any antibiotics inhibit bacterial protein synthesis. Understanding how microbes become antibiotic resistant is important both for developing effective treatment regimens and designing new therapeutics. Macrolides consist of a 14-to 16-member lactone ring with different appended sugars and comprise a key group of inhibitors of bacterial translation (1, 2). The inhibitory activity of macrolides, including erythromycin, depends on binding to a site near the polypeptide exit tunnel of the large ribosomal subunit (3, 4). Because macrolides do not bind to ribosomes with an occupied exit tunnel and cause the synthesis of 2-10 residue peptides in translation assays in vitro, it has been proposed that drug binding physically blocks elongation of nascent proteins beyond this size (5-7).Some macrolide-resistance mutations alter the ribosomal target site and prevent binding (4,8). Intriguingly, other mutations confer resistance despite the fact that macrolides still bind the mutant ribosome well (8-10). For example, deletion of the M 82 K 83 R 84 sequence in Escherichia coli ribosomal protein L22 (⌬MKR) allows growth in the presence of high levels of erythromycin and other macrolides (11-13). The same mutation makes Haemophilus influenzae resistant to numerous macrolides (14); different L22 mutations also confer macrolide resistance in other bacterial species (2). When binding has been measured, ribosomes with macrolideresistant alterations in L22 bind erythromycin with near wild-type affinity (8,10,12,15).In a crystal structure of the E. coli ribosome, the MKR sequence is part of an extended L22 loop, which together with a similar loop in protein L4 forms a narrow constriction in the exit tunnel (Fig. 1A) (16, 17). Cryo-EM studies initially revealed a widened exit tunnel in E. coli ⌬MKR ribosomes (18). This loop is also displaced to create an expanded tunnel in structures of ⌬MKR ribosomes from Thermus thermophilus and Haloarcula marismortui (4, 19). These results explain the altered chemical reactivity in E. coli ⌬MKR ribosomes of 23S-RNA bases (13). Together,...

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Molecular Mechanisms of Antibiotic Resistance: The Need for Novel Antimicrobial Therapies

Dale‐Skinner

Bonev

2008

New Strategies Combating Bacterial Infection

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Despite the enormous success of antibiotics as chemotherapeutic agents, infectious diseases remain a leading cause of mortality worldwide. Bacteria coevolving with infectious microorganisms have been driven to develop protection against environmental bioactive chemicals, and to resist their own antibiotics and defense compounds. Such resistance in pathogenic microorganisms provides protection against chemotherapeutic intervention and can lead to infections that are notoriously difficult to manage. Here, we introduce briefly the molecular mechanisms of action for common antibiotic classes and the structural determinants of bacterial resistance to antibiotics. Bacterial resistance to antibiotics interfering with cell wall biosynthesis is discussed with examples from b-lactams and glycopeptides. The molecular determinants of bacterial tolerance to protein biosynthesis inhibitors are analyzed with examples from aminoglycosides, marcolides and tetracyclines. Fluoroquinolone tolerance is described in connection with DNA regulation in the presence of inhibitors. The action of b-lactam and glycopeptides antibiotics, which target cell wall biosynthesis, is evaded through target modifications and antibiotic deactivation. bLactamases can be deployed to deactivate b-lactam antibiotics, while penicillinbinding proteins with altered binding sites provide an example of modification of the antibiotic target. Alteration of peptidoglycan peptide termini helps protect cell wall synthesis from glycopeptides, while reduced cross-linking and thickened cell wall obstruct glycopeptides access at the cell wall periphery. Specific molecular changes within ribosomal structures can foil aminoglycoside, tetracycline or macrolide attack. Further protection can be achieved by positioning proteins to protect protein synthesizing machinery from the action of tetracyclines. Important resistance mechanisms protecting bacterial protein synthesis include enzymatic deactivation of aminoglycosides and efflux systems expelling incoming macrolides, aminoglycosides and tetracyclines. Renewed efforts to explore alternative strategies for infection management have brought to the top of scientific agenda defense peptides, lanthionine antibiotics, phage therapies and other antimicrobial techniNew Strategies Combating Bacterial Infection. Edited by Iqbal Ahmad and Farrukh Aqil

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Biochemical and genetic studies on two different types of erythromycin resistant mutants of Escherichia coli with altered ribosomal proteins

Cited by 167 publications

References 34 publications

Drug efflux pump deficiency and drug target resistance masking in growing bacteria

Drug efflux pump deficiency and drug target resistance masking in growing bacteria

Revisiting the mechanism of macrolide-antibiotic resistance mediated by ribosomal protein L22

Molecular Mechanisms of Antibiotic Resistance: The Need for Novel Antimicrobial Therapies

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