A plasmid-encoded dihydrofolate reductase from trimethoprim-resistant bacteria has a novel D2-symmetric active site

@article{Narayana1995APD,
  title={A plasmid-encoded dihydrofolate reductase from trimethoprim-resistant bacteria has a novel D2-symmetric active site},
  author={N. Narayana and D. Matthews and E. Howell and N. Xuong},
  journal={Nature Structural Biology},
  year={1995},
  volume={2},
  pages={1018-1025}
}
Bacteria expressing R67-plasmid encoded dihydrofolate reductase (R67 DHFR) exhibit high-level resistance to the antibiotic trimethoprim. Native R67 DHFR is a 34,000 Mr homotetramer which exists in equilibrium with an inactive dimeric form. The structure of native R67 DHFR has now been solved at 1.7 Å resolution and is unrelated to that of chromosomal DHFR. Homotetrameric R67 DHFR has an unusual pore, 25 Å in length, passing through the middle of the molecule. Two folate molecules bind… Expand
Mechanistic Studies of R67 Dihydrofolate Reductase
TLDR
Results indicate protonated dihydrofolate (pKa = 2.59) is the productive substrate and that R67 DHFR does not possess a proton donor. Expand
Redesigning the Quaternary Structure of R67 Dihydrofolate Reductase
TLDR
The production of a fully active, monomeric R67 DHFR variant will enable the design of more meaningful site-directed mutants where single substitutions per active site pore can be generated. Expand
Novel crystallization conditions for tandem variant R67 DHFR yield a wild-type crystal structure.
TLDR
A tandem dimer construct was created that linked two monomeric R67 DHFR subunits together and mutated the sequence of residues 66-69 of the first subunit from VQIY to INSF, and it was demonstrated that the variant protomer was selectively degraded by chymotrypsin, although no canonical chymosynthesis site had been introduced by these mutations. Expand
Integron-sequestered dihydrofolate reductase: a recently redeployed enzyme.
TLDR
This information supports the ideas that DfrB DHFR is a poorly adapted catalyst and has recently been recruited to perform a novel enzymatic activity in response to selective pressure. Expand
Asymmetric mutations in the tetrameric R67 dihydrofolate reductase reveal high tolerance to active‐site substitutions
TLDR
The results suggest that the presence of two native protomers in the R67 DHFR tetramer is sufficient to provide native‐like catalytic rate and thus ensure cellular proliferation, and suggests a high tolerance for active‐site substitutions. Expand
Directed Evolution and Osmolyte Studies of R67 Dihydrofolate Reductase
TLDR
An in vivo study was conducted monitoring the effect of the osmolyte sorbitol plus TMP on the growth of wild-type R67 DHFR and several mutants and the effects of heavily mutating a variant of this enzyme were investigated, supporting in vitro findings about the importance of available water to this enzyme. Expand
Role of Lys-32 Residues in R67 Dihydrofolate Reductase Probed by Asymmetric Mutations*
TLDR
The K32M:1+3 mutant data suggest this interaction is an ionic interaction between Lys-32 and the charged tail of dihydrofolate, which arises from the 222 symmetry imposed on the single active site pore. Expand
Multiple ligand-binding modes in bacterial R67 dihydrofolate reductase
TLDR
Results suggest that multipe binding modes of the ligands are possible within R67 DHFR, a bacterial plasmid-encoded enzyme associated with resistance to the drug trimethoprim. Expand
Searching Sequence Space: Two Different Approaches to Dihydrofolate Reductase Catalysis
  • E. Howell
  • Biology, Medicine
  • Chembiochem : a European journal of chemical biology
  • 2005
TLDR
This review focuses on two dihydrofolate reductases (DHFRs) that have disparate structures and discusses how the catalytic strategies of these two DHFRs are driven by their respective scaffolds. Expand
Tales of Dihydrofolate Binding to R67 Dihydrofolate Reductase
TLDR
Results indicate a balance between folate tail disorder, which helps the enzyme bind substrate while dynamics facilitates catalysis, and decreased dynamics and/or suboptimal positioning of the cross-linked folate for catalysis. Expand
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TLDR
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TLDR
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TLDR
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TLDR
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TLDR
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  • Biology, Medicine
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TLDR
Data suggest that the enzyme from R plasmid R67 possesses a pyridine nucleotide binding site different from that of other dihydrofolate reductases and dehydrogenases. Expand
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TLDR
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