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- Baquero, María-Rosario, et al.
(författare)
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Polymorphic mutation frequencies in Escherichia coli : emergence of weak mutators in clinical isolates
- 2004
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Ingår i: Journal of Bacteriology. - 0021-9193 .- 1098-5530. ; 186:16, s. 5538-5542
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Tidskriftsartikel (refereegranskat)abstract
- Polymorphisms in the rifampin resistance mutation frequency (f) were studied in 696 Escherichia coli strains from Spain, Sweden, and Denmark. Of the 696 strains, 23% were weakly hypermutable (4 x 10(-8) < or = f < 4 x 10(-7)), and 0.7% were strongly hypermutable (f > or = 4 x 10(-7)). Weak mutators were apparently more frequent in southern Europe and in blood isolates (38%) than in urinary tract isolates (25%) and feces of healthy volunteers (11%).
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| 4. |
- Chen, Yang, et al.
(författare)
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Structure of AadA from Salmonella enterica : a monomeric aminoglycoside (3'')(9) adenyltransferase
- 2015
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Ingår i: Acta Crystallographica Section D. - 0907-4449 .- 1399-0047. ; 71, s. 2267-2277
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Tidskriftsartikel (refereegranskat)abstract
- Aminoglycoside resistance is commonly conferred by enzymatic modification of drugs by aminoglycoside-modifying enzymes such as aminoglycoside nucleo\-tidyltransferases (ANTs). Here, the first crystal structure of an ANT(3$^\prime$$^\prime$)(9) adenyltransferase, AadA from Salmonella enterica, is presented. AadA catalyses the magnesium-dependent transfer of adenosine monophosphate from ATP to the two chemically dissimilar drugs streptomycin and spectinomycin. The structure was solved using selenium SAD phasing and refined to 2.5Å resolution. AadA consists of a nucleotidyltransferase domain and an α-helical bundle domain. AadA crystallizes as a monomer and is a monomer in solution as confirmed by small-angle X-ray scattering, in contrast to structurally similar homodimeric adenylating enzymes such as kanamycin nucleotidyltransferase. Isothermal titration calorimetry experiments show that ATP binding has to occur before binding of the aminoglycoside substrate, and structure analysis suggests that ATP binding repositions the two domains for aminoglycoside binding in the interdomain cleft. Candidate residues for ligand binding and catalysis were subjected to site-directed mutagenesis. In vivo resistance and in vitro binding assays support the role of Glu87 as the catalytic base in adenylation, while Arg192 and Lys205 are shown to be critical for ATP binding.
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| 6. |
- Guo, Xiaohu, et al.
(författare)
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Structure and mechanism of a phage-encoded SAM lyase revises catalytic function of enzyme family
- 2021
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Ingår i: eLIFE. - : eLife Sciences Publications Ltd. - 2050-084X. ; 10
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Tidskriftsartikel (refereegranskat)abstract
- The first S-adenosyl methionine (SAM) degrading enzyme (SAMase) was discovered in bacteriophage T3, as a counter-defense against the bacterial restriction-modification system, and annotated as a SAM hydrolase forming 5’-methyl-thioadenosine (MTA) and L-homoserine. From environmental phages, we recently discovered three SAMases with barely detectable sequence similarity to T3 SAMase and without homology to proteins of known structure. Here, we present the very first phage SAMase structures, in complex with a substrate analogue and the product MTA. The structure shows a trimer of alpha–beta sandwiches similar to the GlnB-like superfamily, with active sites formed at the trimer interfaces. Quantum-mechanical calculations, thin-layer chromatography, and nuclear magnetic resonance spectroscopy demonstrate that this family of enzymes are not hydrolases but lyases forming MTA and L-homoserine lactone in a unimolecular reaction mechanism. Sequence analysis and in vitro and in vivo mutagenesis support that T3 SAMase belongs to the same structural family and utilizes the same reaction mechanism.
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| 8. |
- Gustafsson, Ingegerd, et al.
(författare)
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Bacteria with increased mutation frequency and antibiotic resistance are enriched in the commensal flora of patients with high antibiotic usage
- 2003
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Ingår i: Journal of Antimicrobial Chemotherapy. - : Oxford University Press (OUP). - 0305-7453 .- 1460-2091. ; 52:4, s. 645-650
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: We examined how prolonged antibiotic treatment affected the resistance and mutation frequency of human microflora isolated from intestine (Escherichia coli, enterococci spp.), pharynx (alpha-streptococci) and nostril (coagulase-negative staphylococci, CoNS).METHODS: Samples were collected from patients at the Center of Cystic Fibrosis (n=18) and the haematology ward (n=18) of the University Hospital, Uppsala, Sweden. The individually used amount of antibiotics for 1 year was recorded as the defined daily dose (DDD). Primary health care patients (n=30), with no antibiotic treatment for 1 year before sampling, were used as controls. Three isolates of each bacterium from each patient were examined. Antibiotic susceptibilities were determined by disc diffusion. Mutation frequencies to rifampicin resistance were measured on 30 independent cultures of each bacterial species from each individual by plating on rifampicin agar plates. For alpha-streptococci the mutation frequency to streptomycin resistance was also determined.RESULTS: Isolates from patients with high antibiotic use showed a pronounced shift towards increased resistance and a small but significant increase in the mutation frequency compared with isolates from the controls. For E. coli, enterococci and CoNS the increase in geometric mean mutation frequency in the patient group was 3-, 1.8- and 1.5-fold, respectively (P values 0.0001, 0.016 and 0.012). For alpha-streptococci there was a significant difference in geometric mean mutation frequency between patient and control groups for streptomycin resistance (P=0.024) but not for rifampicin resistance (P=0.74).CONCLUSIONS: High antibiotic use selected for commensals with highly increased resistance and a slight increase in mutation frequency.
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| 9. |
- Jerlström-Hultqvist, Jon, 1982-, et al.
(författare)
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A bacteriophage enzyme induces bacterial metabolic perturbation that confers a novel promiscuous function
- 2018
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Ingår i: Nature Ecology & Evolution. - : Springer Science and Business Media LLC. - 2397-334X. ; 2:8, s. 1321-1330
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Tidskriftsartikel (refereegranskat)abstract
- One key concept in the evolution of new functions is the ability of enzymes to perform promiscuous side-reactions that serve as a source of novelty that may become beneficial under certain conditions. Here, we identify a mechanism where a bacteriophage-encoded enzyme introduces novelty by inducing expression of a promiscuous bacterial enzyme. By screening for bacteriophage DNA that rescued an auxotrophic Escherichia coli mutant carrying a deletion of the ilvA gene, we show that bacteriophage-encoded S-adenosylmethionine (SAM) hydrolases reduce SAM levels. Through this perturbation of bacterial metabolism, expression of the promiscuous bacterial enzyme MetB is increased, which in turn complements the absence of IlvA. These results demonstrate how foreign DNA can increase the metabolic capacity of bacteria, not only by transfer of bona fide new genes, but also by bringing cryptic bacterial functions to light via perturbations of cellular physiology.
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| 10. |
- Koskiniemi, Sanna, et al.
(författare)
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Activation of cryptic aminoglycoside resistance in Salmonella enterica
- 2011
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Ingår i: Molecular Microbiology. - : Wiley. - 0950-382X .- 1365-2958. ; 80:6, s. 1464-1478
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Tidskriftsartikel (refereegranskat)abstract
- Aminoglycoside resistance in bacteria can be acquired by several mechanisms, including drug modification, target alteration, reduced uptake and increased efflux. Here we demonstrate that increased resistance to the aminoglycosides streptomycin and spectinomycin in Salmonella enterica can be conferred by increased expression of an aminoglycoside adenyl transferase encoded by the cryptic, chromo-somally located aadA gene. During growth in rich medium the wild-type strain was susceptible but mutations that impaired electron transport and conferred a small colony variant (SCV) phenotype or growth in glucose/glycerol minimal media resulted in activation of the aadA gene and aminoglycoside resistance. Expression of the aadA gene was positively regulated by the stringent response regulator guanosine penta/tetraphosphate ((p) ppGpp). SCV mutants carrying stop codon mutations in the hemA and ubiA genes showed a streptomycin pseudo-dependent phenotype, where growth was stimulated by streptomycin. Our data suggest that this phenotype is due to streptomycin-induced readthrough of the stop codons, a resulting increase in HemA/UbiA levels and improved electron transport and growth. Our results demonstrate that environmental and mutational activation of a cryptic resistance gene can confer clinically significant resistance and that a streptomycin-pseudo-dependent phenotype can be generated via a novel mechanism that does not involve the classical rpsL mutations.
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