ArticleAntimicrobial agents and chemotherapy2004
Functional cloning of Bacillus anthracis dihydrofolate reductase and confirmation of natural resistance to trimethoprim.
Article in Antimicrobial agents and chemotherapy, 2004. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 16 papers.
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Who cites it
16 citing papers in PubMed, 33 citations in OpenAlex.
- Exploration into the origins and mobilization of di-hydrofolate reductase genes and the emergence of clinical resistance to trimethoprim.Microbial genomics · 2020Article
- Evaluation ofJournal of veterinary science · 2019Article
- Modified 2,4-diaminopyrimidine-based dihydrofolate reductase inhibitors as potential drug scaffolds against Bacillus anthracis.Bioorganic & medicinal chemistry · 2015Article
- Identification of novel potential antibiotics against Staphylococcus using structure-based drug screening targeting dihydrofolate reductase.Journal of chemical information and modeling · 2014Article
- Synthesis and biological evaluation of 2,4-diaminopyrimidine-based antifolate drugs against Bacillus anthracis.Molecules (Basel, Switzerland) · 2014Article
- Utility of the Biosynthetic Folate Pathway for Targets in Antimicrobial Discovery.Antibiotics (Basel, Switzerland) · 2014Review
- Targeted mutations of Bacillus anthracis dihydrofolate reductase condense complex structure−activity relationships.Journal of medicinal chemistry · 2010Article
- Inhibition of antibiotic-resistant Staphylococcus aureus by the broad-spectrum dihydrofolate reductase inhibitor RAB1.Antimicrobial agents and chemotherapy · 2010Article
- Crystal structure of Bacillus anthracis dihydrofolate reductase with the dihydrophthalazine-based trimethoprim derivative RAB1 provides a structural explanation of potency and selectivity.Antimicrobial agents and chemotherapy · 2009Article
- X-ray structure of the ternary MTX.NADPH complex of the anthrax dihydrofolate reductase: a pharmacophore for dual-site inhibitor design.Journal of structural biology · 2009Article
- Synthetic and crystallographic studies of a new inhibitor series targeting Bacillus anthracis dihydrofolate reductase.Journal of medicinal chemistry · 2008Article
- Examination of intrinsic sulfonamide resistance in Bacillus anthracis: a novel assay for dihydropteroate synthase.Biochimica et biophysica acta · 2008Article
- In vitro efficacy of new antifolates against trimethoprim-resistant Bacillus anthracis.Antimicrobial agents and chemotherapy · 2007Article
- Evaluation of the metS and murB loci for antibiotic discovery using targeted antisense RNA expression analysis in Bacillus anthracis.Antimicrobial agents and chemotherapy · 2007Article
- Structure-activity relationships of Bacillus cereus and Bacillus anthracis dihydrofolate reductase: toward the identification of new potent drug leads.Antimicrobial agents and chemotherapy · 2006Article
- Cloning and characterization of a novel trimethoprim-resistant dihydrofolate reductase from a nosocomial isolate of Staphylococcus aureus CM.S2 (IMCJ1454).Antimicrobial agents and chemotherapy · 2005Article
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Authors and funding
3 authors at 1 institution in 1 country.
Funding
Abstract
Bacillus anthracis is reported to be naturally resistant to trimethoprim (TMP), a drug that inhibits dihydrofolate reductase (DHFR), a key enzyme in the folate pathway. A microdilution broth assay established that the MIC of TMP for B. anthracis Sterne is >2,048 but < or =4,096 microg/ml. A putative DHFR sequence was amplified from B. anthracis Sterne genomic DNA. The PCR product was cloned into the Invitrogen pCRT7/CT-TOPO vector, followed by transformation into Escherichia coli TOP10F' chemically competent cells. Plasmid DNA from a clone showing the correct construct with a thrombin cleavage site attached downstream from the terminus of the cloned PCR product was transformed into E. coli BL21 Star (DE3)pLysS competent cells for expression of the six-histidine-tagged fusion protein and purification on a His-Bind resin column. Functionality of the purified Sterne recombinant DHFR (Sterne rDHFR) was confirmed in an established enzyme assay. The 50% inhibitory concentrations of TMP and methotrexate for the Sterne rDHFR were found to be 77,233 and 12.2 nM, respectively. TMP resistance was observed with E. coli BL21 Star (DE3)pLysS competent cells transformed with the Sterne DHFR gene. Alignment of the amino acid sequence of the Sterne DHFR gene revealed 100% homology with various virulent strains of B. anthracis. These results confirm the natural resistance of B. anthracis to TMP and clarify that the resistance is correlated to a lack of selectivity for the chromosomally encoded gene product. These findings will assist in the development of narrow-spectrum antimicrobial agents for treatment of anthrax.
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