Quinolone Resistance Mutations in
            <i>Streptococcus pneumoniae</i>
            GyrA and ParC Proteins: Mechanistic Insights into Quinolone Action from Enzymatic Analysis, Intracellular Levels, and Phenotypes of Wild-Type and Mutant Proteins

Pan, Xiao-Su; Yagüe, Genoveva; Fisher, L. Mark

doi:10.1128/aac.45.11.3140-3147.2001

Cited by 67 publications

(68 citation statements)

References 51 publications

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“…Four S. gallolyticus strains were resistant to enrofloxacin. This resistance, which is also encountered in the human pathogen Streptococcus pneumoniae (Pan et al, 2001), has not been reported earlier in the streptococci of animal origin. The E. coli strains in particular had acquired resistance against the most commonly used antimicrobials in pigeons.…”

Section: Discussionmentioning

confidence: 89%

Prevalence of antimicrobial resistance among pigeon isolates of Streptococcus gallolyticus , Escherichia coli and Salmonella enterica serotype Typhimurium

et al. 2002

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Thirty-three Streptococcus gallolyticus, 60 Escherichia coli and 18 Salmonella enterica serotype Typhimurium var. Copenhagen strains isolated from homing pigeons (Columba livia) were tested for susceptibility to the antimicrobials most commonly used to treat pigeons. Minimal inhibitory concentrations were determined using the agar dilution technique. Aminoglycosides (gentamicin and kanamycin), trimethoprim and flumequine were relatively inactive against the streptococci tested. Acquired tetracycline resistance amounted to 85%, and lincomycin and macrolide (erythromycin) resistance to 48 and 45%, respectively. Fluoroquinolone (enrofloxacin) resistance was found in four S. gallolyticus strains. All strains were susceptible to ampicillin. With the E. coli strains, resistance was found to all antibiotics tested. Over one-half of them were resistant to tetracycline and to broadspectrum penicillins (ampicillin); however, none showed extended spectrum b -lactamase activity, implying that the cephalosporins (ceftiofur) remained active. Resistance to trimethoprim, aminoglycosides and fluoroquinolone ranked next. In contrast to the S. gallolyticus and E. coli strains, the S. enterica strains were susceptible to all the antimicrobials tested.

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Section: Discussionmentioning

confidence: 89%

Prevalence of antimicrobial resistance among pigeon isolates of Streptococcus gallolyticus , Escherichia coli and Salmonella enterica serotype Typhimurium

et al. 2002

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“…Each of these positions once mutated, either alone or in association with other positions located in the vicinity (Table 1), resulted in the particular phenotype of resistance to FQs and hypersusceptibility to novobiocin observed with the His103Tyr or His103Phe substitution. None of these positions is located in the QRDR, mutations of which are thought to decrease the affinity of the topoisomerases for quinolones, a partial explanation of the mechanism of resistance to FQs (5,40). Interestingly, in S. aureus, the Asn470Asp substitution near the conserved motif of the QRDR of ParE led to a similar phenotype, with decreased susceptibility to FQs and increased susceptibility to novobiocin (14).…”

Section: Vol 185 2003 Atp-bound Conformation Of Topoisomerase IV 6143mentioning

confidence: 96%

ATP-Bound Conformation of Topoisomerase IV: a Possible Target for Quinolones in Streptococcus pneumoniae

et al. 2003

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Topoisomerase IV, a C 2 E 2 tetramer, is involved in the topological changes of DNA during replication. This enzyme is the target of antibacterial compounds, such as the coumarins, which target the ATP binding site in the ParE subunit, and the quinolones, which bind, outside the active site, to the quinolone resistance-determining region (QRDR). After site-directed and random mutagenesis, we found some mutations in the ATP binding site of ParE near the dimeric interface and outside the QRDR that conferred quinolone resistance to Streptococcus pneumoniae, a bacterial pathogen. Modeling of the N-terminal, 43-kDa ParE domain of S. pneumoniae revealed that the most frequent mutations affected conserved residues, among them His43 and His103, which are involved in the hydrogen bond network supporting ATP hydrolysis, and Met31, at the dimeric interface. All mutants showed a particular phenotype of resistance to fluoroquinolones and an increase in susceptibility to novobiocin. All mutations in ParE resulted in resistance only when associated with a mutation in the QRDR of the GyrA subunit. Our models of the closed and open conformations of the active site indicate that quinolones preferentially target topoisomerase IV of S. pneumoniae in its ATP-bound closed conformation.Topoisomerase IV and DNA gyrase are bacterial type II topoisomerases, which modify DNA topology during the replication process by unlinking DNA and facilitating chromosome segregation (59). Topoisomerase IV forms a C 2 E 2 tetramer involved in segregation of the chromosome at cell division (1, 30, 59). The ParC subunit contains the site of topoisomerization catalyzing the double-stranded DNA break (30, 53), while the ParE subunit catalyzes the hydrolysis of ATP, providing the free energy necessary for these reactions (2, 6, 11). The ParE and ParC subunits share extensive sequence homology with, respectively, the GyrB and GyrA subunits of the DNA gyrase A 2 B 2 tetramer, which catalyzes negative DNA supercoiling during the initiation and elongation processes of DNA replication (16,53).Both topoisomerases are the targets of antibacterial molecules, such as the quinolones and the coumarins. The coumarins inhibit supercoiling and enzyme turnover by preventing the binding and hydrolysis of ATP (45). The quinolones form a ternary complex with the topoisomerases in the presence of DNA, resulting in lethal double-stranded DNA breaks (11). Enzymatic studies and binding assays have also shown that the quinolones can form a complex with the topoisomerases before binding DNA (15, 29); however, some binding data indicate the occurrence of a specific and higher level of binding to the enzyme-DNA complex rather than to the enzyme alone (43,56). These families of molecules, in particular, the fluoroquinolones (FQs), are under continuous development as resistance to antibiotics in pathogenic bacteria has dramatically increased during the last decade (20,22,49).

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“…Mutants resistant to quinolones most frequently harbor mutations in the quinolone resistance-determining regions (QRDR) of GyrA and ParC and less frequently in that of ParE (7). The level of quinolone resistance increases when a mutation is present in the secondary target in addition to one in the primary target (21,29). In S. pneumoniae, in vitro-selected resistance to novobiocin is associated with the S127L substitution in the GyrB subunit (17).…”

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confidence: 99%

“…In Streptococcus pneumoniae, it has been shown that the primary and the secondary targets of quinolones are either the gyrase or the topoisomerase IV, depending upon the molecule (21,29). Mutants resistant to quinolones most frequently harbor mutations in the quinolone resistance-determining regions (QRDR) of GyrA and ParC and less frequently in that of ParE (7).…”

mentioning

confidence: 99%

“…Catalytic activities of topoisomerase IV were tested as described previously (20,21). For the decatenation assay, 0.4 g of kinetoplast DNA (kDNA; TOPOGEN, Inc., Columbus, Ohio) and 2 U of topoisomerase IV were incubated at 37°C for 1 h. For the relaxation assay, 0.4 g of supercoiled pBR322 (Roche-Boehringer, Mannheim, Germany) and 2 U of topoisomerase IV were incubated at 37°C for 1 h. The specific activities of WT ParE and mutant H103Y ParE were first determined using an excess of ParC and an excess of ATP (Ն1 mM).…”

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confidence: 99%

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Contribution of the ATP Binding Site of ParE to Susceptibility to Novobiocin and Quinolones in Streptococcus pneumoniae

et al. 2005

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In Streptococcus pneumoniae, an H103Y substitution in the ATP binding site of the ParE subunit of topoisomerase IV was shown to confer quinolone resistance and hypersensitivity to novobiocin when associated with an S84F change in the A subunit of DNA gyrase. We reconstituted in vitro the wild-type topoisomerase IV and its ParE mutant. The ParE mutant enzyme showed a decreased activity for decatenation at subsaturating ATP levels and was more sensitive to inhibition by novobiocin but was as sensitive to quinolones. These results show that the ParE alteration H103Y alone is not responsible for quinolone resistance and agree with the assumption that it facilitates the open conformation of the ATP binding site that would lead to novobiocin hypersensitivity and to a higher requirement of ATP.Topoisomerase IV and DNA gyrase are bacterial type II topoisomerases that modify DNA topology during the replication process by unlinking DNA and facilitating chromosome segregation (33). DNA gyrase, encoded by gyrA and gyrB, forms an A 2 B 2 tetramer that removes the positive supercoils introduced during DNA replication and helps to maintain the steady-state balance of negative supercoiling of the bacterial DNA (30). Topoisomerase IV, encoded by parC and parE, forms a C 2 E 2 tetramer. Its primary role is the decatenation of daughter chromosomes following DNA replication, and other functions are DNA relaxation and double-strand DNA breakage (1,6,14,33). Both enzymes are the target of two antibiotic families: the quinolones, such as ciprofloxacin, sparfloxacin, and grepafloxacin, and the coumarins, such as novobiocin. The quinolones form a ternary complex with topoisomerases in the presence of DNA, resulting in double-stranded breaks (4). The coumarins inhibit ATP-dependent functions of these enzymes by preventing binding and hydrolysis of ATP (27).In Streptococcus pneumoniae, it has been shown that the primary and the secondary targets of quinolones are either the gyrase or the topoisomerase IV, depending upon the molecule (21, 29). Mutants resistant to quinolones most frequently harbor mutations in the quinolone resistance-determining regions (QRDR) of GyrA and ParC and less frequently in that of ParE (7). The level of quinolone resistance increases when a mutation is present in the secondary target in addition to one in the primary target (21, 29). In S. pneumoniae, in vitro-selected resistance to novobiocin is associated with the S127L substitution in the GyrB subunit (17). This position is located upstream from and close to positions 136 and 164 described in the ATP binding domain of GyrB in Escherichia coli and also involved in coumarin resistance (3). The substitution in ParE of E. coli at a position equivalent to that of position 136 in GyrB resulted in a decreased inhibition of topoisomerase IV by novobiocin (6).In previous studies (11, 26), we showed in S. pneumoniae that the H103Y substitution in the N terminus of the ParE subunit of the topoisomerase IV resulted in an original phenotype of resistance only if it was as...

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Quinolone Resistance Mutations in Streptococcus pneumoniae GyrA and ParC Proteins: Mechanistic Insights into Quinolone Action from Enzymatic Analysis, Intracellular Levels, and Phenotypes of Wild-Type and Mutant Proteins

Cited by 67 publications

References 51 publications

Prevalence of antimicrobial resistance among pigeon isolates of Streptococcus gallolyticus , Escherichia coli and Salmonella enterica serotype Typhimurium

Prevalence of antimicrobial resistance among pigeon isolates of Streptococcus gallolyticus , Escherichia coli and Salmonella enterica serotype Typhimurium

ATP-Bound Conformation of Topoisomerase IV: a Possible Target for Quinolones in Streptococcus pneumoniae

Contribution of the ATP Binding Site of ParE to Susceptibility to Novobiocin and Quinolones in Streptococcus pneumoniae

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