The Experts below are selected from a list of 414 Experts worldwide ranked by ideXlab platform
Alexander S Mankin - One of the best experts on this subject based on the ideXlab platform.
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Oxazolidinone resistance mutations in 23s rrna of escherichia coli reveal the central region of domain v as the primary site of drug action
Journal of Bacteriology, 2000Co-Authors: Liqun Xiong, Dean L Shinabarger, Patricia Kloss, Stephen Douthwaite, Niels Moller Andersen, Steven Swaney, Alexander S MankinAbstract:Oxazolidinone Antibiotics inhibit bacterial protein synthesis by interacting with the large ribosomal subunit. The structure and exact location of the Oxazolidinone binding site remain obscure, as does the manner in which these drugs inhibit translation. To investigate the drug-ribosome interaction, we selected Escherichia coli Oxazolidinone-resistant mutants, which contained a randomly mutagenized plasmid-borne rRNA operon. The same mutation, G2032 to A, was identified in the 23S rRNA genes of several independent resistant isolates. Engineering of this mutation by site-directed mutagenesis in the wild-type rRNA operon produced an Oxazolidinone resistance phenotype, establishing that the G2032A substitution was the determinant of resistance. Engineered U and C substitutions at G2032, as well as a G2447-to-U mutation, also conferred resistance to Oxazolidinone. All the characterized resistance mutations were clustered in the vicinity of the central loop of domain V of 23S rRNA, suggesting that this rRNA region plays a major role in the interaction of the drug with the ribosome. Although the central loop of domain V is an essential integral component of the ribosomal peptidyl transferase, Oxazolidinones do not inhibit peptide bond formation, and thus these drugs presumably interfere with another activity associated with the peptidyl transferase center.
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Oxazolidinone Resistance Mutations in 23S rRNA of Escherichia coli Reveal the Central Region of Domain V as the Primary Site of Drug Action
2000Co-Authors: Steven Swaney, Dean L Shinabarger, Alexander S MankinAbstract:Oxazolidinone Antibiotics inhibit bacterial protein synthesis by interacting with the large ribosomal subunit. The structure and exact location of the Oxazolidinone binding site remain obscure, as does the manner in which these drugs inhibit translation. To investigate the drug-ribosome interaction, we selected Escherichia coli Oxazolidinone-resistant mutants, which contained a randomly mutagenized plasmid-borne rRNA operon. The same mutation, G2032 to A, was identified in the 23S rRNA genes of several independent resistant isolates. Engineering of this mutation by site-directed mutagenesis in the wild-type rRNA operon produced an oxazo-lidinone resistance phenotype, establishing that the G2032A substitution was the determinant of resistance. Engineered U and C substitutions at G2032, as well as a G2447-to-U mutation, also conferred resistance to Oxazolidinone. All the characterized resistance mutations were clustered in the vicinity of the central loop of domain V of 23S rRNA, suggesting that this rRNA region plays a major role in the interaction of the drug with the ribosome. Although the central loop of domain V is an essential integral component of the ribosomal peptidyl transferase, Oxazolidinones do not inhibit peptide bond formation, and thus these drugs presumably interfere with another activity associated with the peptidyl transferase center. During the course of evolution, a disproportionately large number of natural Antibiotics have been selected to act upo
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resistance mutations in 23 s rrna identify the site of action of the protein synthesis inhibitor linezolid in the ribosomal peptidyl transferase center
Journal of Molecular Biology, 1999Co-Authors: Patricia Kloss, Dean L Shinabarger, Liqun Xiong, Alexander S MankinAbstract:Oxazolidinones represent a novel class of Antibiotics that inhibit protein synthesis in sensitive bacteria. The mechanism of action and location of the binding site of these drugs is not clear. A new representative of Oxazolidinone Antibiotics, linezolid, was found to be active against bacteria and against the halophilic archaeon Halobacterium halobium. The use of H. halobium, which possess only one chromosomal copy of rRNA operon, allowed isolation of a number of linezolid-resistance mutations in rRNA. Four types of linezolid-resistant mutants were isolated by direct plating of H. halobium cells on agar medium containing antibiotic. In addition, three more linezolid-resistant mutants were identified among the previously isolated mutants of H. halobium containing mutations in either 16 S or 23 S rRNA genes. All the isolated mutants were found to contain single-point mutations in 23 S rRNA. Seven mutations affecting six different positions in the central loop of domain V of 23 S rRNA were found to confer resistance to linezolid. Domain V of 23 S rRNA is known to be a component of the ribosomal peptidyl transferase center. Clustering of linezolid-resistance mutations within this region strongly suggests that the binding site of the drug is located in the immediate vicinity of the peptidyl transferase center. However, the antibiotic failed to inhibit peptidyl transferase activity of the H. halobium ribosome, supporting the previous conclusion that linezolid inhibits translation at a step different from the catalysis of the peptide bond formation.
Dean L Shinabarger - One of the best experts on this subject based on the ideXlab platform.
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in vitro activity of tr 700 the antibacterial moiety of the prodrug tr 701 against linezolid resistant strains
Antimicrobial Agents and Chemotherapy, 2008Co-Authors: Karen Joy Shaw, Dean L Shinabarger, Ronda D. Schaadt, S Poppe, Vickie Browndriver, John T Finn, Chris M Pillar, Gary E ZurenkoAbstract:TR-701 is the orally active prodrug of TR-700, a novel Oxazolidinone that demonstrates four- to eightfoldgreater activity than linezolid (LZD) against Staphylococcus and Enterococcus spp. In this study evaluating the in vitro sensitivity of LZD-resistant isolates, TR-700 demonstrated 8- to 16-fold-greater potency than LZD against all strains tested, including methicillin-resistant Staphylococcus aureus (MRSA), strains of MRSA carrying the mobile cfr methyltransferase gene, and vancomycin-resistant enterococci. The MIC90 for TR-700 against LZD-resistant S. aureus was 2 g/ml, demonstrating the utility of TR-700 against LZD-resistant strains. A model of TR-700 binding to 23S rRNA suggests that the increased potency of TR-700 is due to additional target site interactions and that TR-700 binding is less reliant on target residues associated with resistance to LZD. Oxazolidinone Antibiotics are one of the newest classes of Antibiotics developed within the past 30 years, with linezolid (LZD) representing the only marketed member of this class. In 2000, LZD (Zyvox) was granted approval for the treatment of infections associated with vancomycin-resistant Enterococcus faecium, nosocomial pneumonia, community-acquired pneumonia due to Streptococcus pneumoniae and methicillin-sensitive Staphylococcus aureus (MSSA), and complicated skin and skin structure infections, including cases due to methicillinresistant Staphylococcus aureus (MRSA) (1). Later approvals included pediatric use, pneumonia due to multidrugresistant S. pneumoniae, and treatment of diabetic foot infections, without osteomyelitis, caused by gram-positive bacteria. These approvals represent important milestones for the novel Oxazolidinone class in the treatment of serious infections. Oxazolidinones have been shown to bind to the 50S ribosomal subunit and inhibit protein translation (31). A model of the binding of LZD to the 23S rRNA peptidyl transferase region has been previously published, based upon in vivo crosslinking experiments (18). This model predicts that LZD would specifically interfere with the binding of the amino acid portion of the aminoacyl tRNA to the ribosomal A site. The recent crystal structure of LZD bound to the 50S ribosomal subunit confirms these findings and suggests that the mechanism of inhibition involves competition with the incoming A site substrates (13). Mutations in the 23S rRNA central loop of domain V, the peptidyl transferase center (PTC), are associated with the development of LZD resistance. LZD-resistant S. aureus, Staphylococcus epidermidis, Enterococcus faecium, and Enterococcus faecalis mutants have been isolated infrequently in the clinic and can be selected for in the
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Oxazolidinone Antibiotics Target the P Site on Escherichia coli Ribosomes
Antimicrobial agents and chemotherapy, 2002Co-Authors: Hiroyuki Aoki, Dean L Shinabarger, Susan M. Poppe, Toni J. Poel, Elizabeth A. Weaver, Robert C. Gadwood, Richard C. Thomas, M. Clelia GanozaAbstract:The Oxazolidinones are a novel class of antimicrobial agents that target protein synthesis in a wide spectrum of gram-positive and anaerobic bacteria. The Oxazolidinone PNU-100766 (linezolid) inhibits the binding of fMet-tRNA to 70S ribosomes. Mutations to Oxazolidinone resistance in Halobacterium halobium, Staphylococcus aureus, and Escherichia coli map at or near domain V of the 23S rRNA, suggesting that the Oxazolidinones may target the peptidyl transferase region responsible for binding fMet-tRNA. This study demonstrates that the potency of Oxazolidinones corresponds to increased inhibition of fMet-tRNA binding. The inhibition of fMet-tRNA binding is competitive with respect to the fMet-tRNA concentration, suggesting that the P site is affected. The fMet-tRNA reacts with puromycin to form peptide bonds in the presence of elongation factor P (EF-P), which is needed for optimum specificity and efficiency of peptide bond synthesis. Oxazolidinone inhibition of the P site was evaluated by first binding fMet-tRNA to the A site, followed by translocation to the P site with EF-G. All three of the Oxazolidinones used in this study inhibited translocation of fMet-tRNA. We propose that the Oxazolidinones target the ribosomal P site and pleiotropically affect fMet-tRNA binding, EF-P stimulated synthesis of peptide bonds, and, most markedly, EF-G-mediated translocation of fMet-tRNA into the P site.
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Oxazolidinone resistance mutations in 23s rrna of escherichia coli reveal the central region of domain v as the primary site of drug action
Journal of Bacteriology, 2000Co-Authors: Liqun Xiong, Dean L Shinabarger, Patricia Kloss, Stephen Douthwaite, Niels Moller Andersen, Steven Swaney, Alexander S MankinAbstract:Oxazolidinone Antibiotics inhibit bacterial protein synthesis by interacting with the large ribosomal subunit. The structure and exact location of the Oxazolidinone binding site remain obscure, as does the manner in which these drugs inhibit translation. To investigate the drug-ribosome interaction, we selected Escherichia coli Oxazolidinone-resistant mutants, which contained a randomly mutagenized plasmid-borne rRNA operon. The same mutation, G2032 to A, was identified in the 23S rRNA genes of several independent resistant isolates. Engineering of this mutation by site-directed mutagenesis in the wild-type rRNA operon produced an Oxazolidinone resistance phenotype, establishing that the G2032A substitution was the determinant of resistance. Engineered U and C substitutions at G2032, as well as a G2447-to-U mutation, also conferred resistance to Oxazolidinone. All the characterized resistance mutations were clustered in the vicinity of the central loop of domain V of 23S rRNA, suggesting that this rRNA region plays a major role in the interaction of the drug with the ribosome. Although the central loop of domain V is an essential integral component of the ribosomal peptidyl transferase, Oxazolidinones do not inhibit peptide bond formation, and thus these drugs presumably interfere with another activity associated with the peptidyl transferase center.
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Oxazolidinone Resistance Mutations in 23S rRNA of Escherichia coli Reveal the Central Region of Domain V as the Primary Site of Drug Action
2000Co-Authors: Steven Swaney, Dean L Shinabarger, Alexander S MankinAbstract:Oxazolidinone Antibiotics inhibit bacterial protein synthesis by interacting with the large ribosomal subunit. The structure and exact location of the Oxazolidinone binding site remain obscure, as does the manner in which these drugs inhibit translation. To investigate the drug-ribosome interaction, we selected Escherichia coli Oxazolidinone-resistant mutants, which contained a randomly mutagenized plasmid-borne rRNA operon. The same mutation, G2032 to A, was identified in the 23S rRNA genes of several independent resistant isolates. Engineering of this mutation by site-directed mutagenesis in the wild-type rRNA operon produced an oxazo-lidinone resistance phenotype, establishing that the G2032A substitution was the determinant of resistance. Engineered U and C substitutions at G2032, as well as a G2447-to-U mutation, also conferred resistance to Oxazolidinone. All the characterized resistance mutations were clustered in the vicinity of the central loop of domain V of 23S rRNA, suggesting that this rRNA region plays a major role in the interaction of the drug with the ribosome. Although the central loop of domain V is an essential integral component of the ribosomal peptidyl transferase, Oxazolidinones do not inhibit peptide bond formation, and thus these drugs presumably interfere with another activity associated with the peptidyl transferase center. During the course of evolution, a disproportionately large number of natural Antibiotics have been selected to act upo
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resistance mutations in 23 s rrna identify the site of action of the protein synthesis inhibitor linezolid in the ribosomal peptidyl transferase center
Journal of Molecular Biology, 1999Co-Authors: Patricia Kloss, Dean L Shinabarger, Liqun Xiong, Alexander S MankinAbstract:Oxazolidinones represent a novel class of Antibiotics that inhibit protein synthesis in sensitive bacteria. The mechanism of action and location of the binding site of these drugs is not clear. A new representative of Oxazolidinone Antibiotics, linezolid, was found to be active against bacteria and against the halophilic archaeon Halobacterium halobium. The use of H. halobium, which possess only one chromosomal copy of rRNA operon, allowed isolation of a number of linezolid-resistance mutations in rRNA. Four types of linezolid-resistant mutants were isolated by direct plating of H. halobium cells on agar medium containing antibiotic. In addition, three more linezolid-resistant mutants were identified among the previously isolated mutants of H. halobium containing mutations in either 16 S or 23 S rRNA genes. All the isolated mutants were found to contain single-point mutations in 23 S rRNA. Seven mutations affecting six different positions in the central loop of domain V of 23 S rRNA were found to confer resistance to linezolid. Domain V of 23 S rRNA is known to be a component of the ribosomal peptidyl transferase center. Clustering of linezolid-resistance mutations within this region strongly suggests that the binding site of the drug is located in the immediate vicinity of the peptidyl transferase center. However, the antibiotic failed to inhibit peptidyl transferase activity of the H. halobium ribosome, supporting the previous conclusion that linezolid inhibits translation at a step different from the catalysis of the peptide bond formation.
Liqun Xiong - One of the best experts on this subject based on the ideXlab platform.
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Oxazolidinone resistance mutations in 23s rrna of escherichia coli reveal the central region of domain v as the primary site of drug action
Journal of Bacteriology, 2000Co-Authors: Liqun Xiong, Dean L Shinabarger, Patricia Kloss, Stephen Douthwaite, Niels Moller Andersen, Steven Swaney, Alexander S MankinAbstract:Oxazolidinone Antibiotics inhibit bacterial protein synthesis by interacting with the large ribosomal subunit. The structure and exact location of the Oxazolidinone binding site remain obscure, as does the manner in which these drugs inhibit translation. To investigate the drug-ribosome interaction, we selected Escherichia coli Oxazolidinone-resistant mutants, which contained a randomly mutagenized plasmid-borne rRNA operon. The same mutation, G2032 to A, was identified in the 23S rRNA genes of several independent resistant isolates. Engineering of this mutation by site-directed mutagenesis in the wild-type rRNA operon produced an Oxazolidinone resistance phenotype, establishing that the G2032A substitution was the determinant of resistance. Engineered U and C substitutions at G2032, as well as a G2447-to-U mutation, also conferred resistance to Oxazolidinone. All the characterized resistance mutations were clustered in the vicinity of the central loop of domain V of 23S rRNA, suggesting that this rRNA region plays a major role in the interaction of the drug with the ribosome. Although the central loop of domain V is an essential integral component of the ribosomal peptidyl transferase, Oxazolidinones do not inhibit peptide bond formation, and thus these drugs presumably interfere with another activity associated with the peptidyl transferase center.
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resistance mutations in 23 s rrna identify the site of action of the protein synthesis inhibitor linezolid in the ribosomal peptidyl transferase center
Journal of Molecular Biology, 1999Co-Authors: Patricia Kloss, Dean L Shinabarger, Liqun Xiong, Alexander S MankinAbstract:Oxazolidinones represent a novel class of Antibiotics that inhibit protein synthesis in sensitive bacteria. The mechanism of action and location of the binding site of these drugs is not clear. A new representative of Oxazolidinone Antibiotics, linezolid, was found to be active against bacteria and against the halophilic archaeon Halobacterium halobium. The use of H. halobium, which possess only one chromosomal copy of rRNA operon, allowed isolation of a number of linezolid-resistance mutations in rRNA. Four types of linezolid-resistant mutants were isolated by direct plating of H. halobium cells on agar medium containing antibiotic. In addition, three more linezolid-resistant mutants were identified among the previously isolated mutants of H. halobium containing mutations in either 16 S or 23 S rRNA genes. All the isolated mutants were found to contain single-point mutations in 23 S rRNA. Seven mutations affecting six different positions in the central loop of domain V of 23 S rRNA were found to confer resistance to linezolid. Domain V of 23 S rRNA is known to be a component of the ribosomal peptidyl transferase center. Clustering of linezolid-resistance mutations within this region strongly suggests that the binding site of the drug is located in the immediate vicinity of the peptidyl transferase center. However, the antibiotic failed to inhibit peptidyl transferase activity of the H. halobium ribosome, supporting the previous conclusion that linezolid inhibits translation at a step different from the catalysis of the peptide bond formation.
Patricia Kloss - One of the best experts on this subject based on the ideXlab platform.
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Oxazolidinone resistance mutations in 23s rrna of escherichia coli reveal the central region of domain v as the primary site of drug action
Journal of Bacteriology, 2000Co-Authors: Liqun Xiong, Dean L Shinabarger, Patricia Kloss, Stephen Douthwaite, Niels Moller Andersen, Steven Swaney, Alexander S MankinAbstract:Oxazolidinone Antibiotics inhibit bacterial protein synthesis by interacting with the large ribosomal subunit. The structure and exact location of the Oxazolidinone binding site remain obscure, as does the manner in which these drugs inhibit translation. To investigate the drug-ribosome interaction, we selected Escherichia coli Oxazolidinone-resistant mutants, which contained a randomly mutagenized plasmid-borne rRNA operon. The same mutation, G2032 to A, was identified in the 23S rRNA genes of several independent resistant isolates. Engineering of this mutation by site-directed mutagenesis in the wild-type rRNA operon produced an Oxazolidinone resistance phenotype, establishing that the G2032A substitution was the determinant of resistance. Engineered U and C substitutions at G2032, as well as a G2447-to-U mutation, also conferred resistance to Oxazolidinone. All the characterized resistance mutations were clustered in the vicinity of the central loop of domain V of 23S rRNA, suggesting that this rRNA region plays a major role in the interaction of the drug with the ribosome. Although the central loop of domain V is an essential integral component of the ribosomal peptidyl transferase, Oxazolidinones do not inhibit peptide bond formation, and thus these drugs presumably interfere with another activity associated with the peptidyl transferase center.
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resistance mutations in 23 s rrna identify the site of action of the protein synthesis inhibitor linezolid in the ribosomal peptidyl transferase center
Journal of Molecular Biology, 1999Co-Authors: Patricia Kloss, Dean L Shinabarger, Liqun Xiong, Alexander S MankinAbstract:Oxazolidinones represent a novel class of Antibiotics that inhibit protein synthesis in sensitive bacteria. The mechanism of action and location of the binding site of these drugs is not clear. A new representative of Oxazolidinone Antibiotics, linezolid, was found to be active against bacteria and against the halophilic archaeon Halobacterium halobium. The use of H. halobium, which possess only one chromosomal copy of rRNA operon, allowed isolation of a number of linezolid-resistance mutations in rRNA. Four types of linezolid-resistant mutants were isolated by direct plating of H. halobium cells on agar medium containing antibiotic. In addition, three more linezolid-resistant mutants were identified among the previously isolated mutants of H. halobium containing mutations in either 16 S or 23 S rRNA genes. All the isolated mutants were found to contain single-point mutations in 23 S rRNA. Seven mutations affecting six different positions in the central loop of domain V of 23 S rRNA were found to confer resistance to linezolid. Domain V of 23 S rRNA is known to be a component of the ribosomal peptidyl transferase center. Clustering of linezolid-resistance mutations within this region strongly suggests that the binding site of the drug is located in the immediate vicinity of the peptidyl transferase center. However, the antibiotic failed to inhibit peptidyl transferase activity of the H. halobium ribosome, supporting the previous conclusion that linezolid inhibits translation at a step different from the catalysis of the peptide bond formation.
Paola Fucini - One of the best experts on this subject based on the ideXlab platform.
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the Oxazolidinone Antibiotics perturb the ribosomal peptidyl transferase center and effect trna positioning
Proceedings of the National Academy of Sciences of the United States of America, 2008Co-Authors: Daniel N. Wilson, Frank Schluenzen, Joerg Harms, Agata L Starosta, Sean R Connell, Paola FuciniAbstract:The Oxazolidinones represent the first new class of Antibiotics to enter into clinical usage within the past 30 years, but their binding site and mechanism of action has not been fully characterized. We have determined the crystal structure of the Oxazolidinone linezolid bound to the Deinococcus radiodurans 50S ribosomal subunit. Linezolid binds in the A site pocket at the peptidyltransferase center of the ribosome overlapping the aminoacyl moiety of an A-site bound tRNA as well as many clinically important Antibiotics. Binding of linezolid stabilizes a distinct conformation of the universally conserved 23S rRNA nucleotide U2585 that would be nonproductive for peptide bond formation. In conjunction with available biochemical data, we present a model whereby Oxazolidinones impart their inhibitory effect by perturbing the correct positioning of tRNAs on the ribosome.
