@article{3000806,
    title = "Insight on specificity of uracil permeases of the NAT/NCS2 family from analysis of the transporter encoded in the pyrimidine utilization operon of Escherichia coli",
    author = "Botou, M. and Lazou, P. and Papakostas, K. and Lambrinidis, G. and Evangelidis, T. and Mikros, E. and Frillingos, S.",
    journal = "Molecular Microbiology",
    year = "2018",
    volume = "108",
    number = "2",
    pages = "204-219",
    publisher = "Wiley-Blackwell Publishing Ltd",
    issn = "0950-382X, 1365-2958",
    doi = "10.1111/mmi.13931",
    keywords = "alanine;  bacterial protein;  fluorouracil;  nucleobase ascorbate transporter;  nucleobase cation symporter 2;  oxipurinol;  permease UraA;  phenylalanine;  RutG protein;  thymine;  unclassified drug;  uracil;  xanthine;  carrier protein;  Escherichia coli protein;  pyrimidine;  pyrimidine derivative;  UraA protein, E coli, Acinetobacter calcoaceticus;  Aeromonas veronii;  amino acid substitution;  Article;  bacterial cell;  binding affinity;  binding site;  controlled study;  enzyme specificity;  Escherichia coli;  molecular docking;  molecular model;  molecular phylogeny;  nonhuman;  priority journal;  protein determination;  protein function;  protein protein interaction;  site directed mutagenesis;  structural homology;  bacterium;  chemistry;  classification;  enzyme specificity;  enzymology;  Escherichia coli;  genetics;  metabolism;  multigene family;  operon;  phylogeny, Bacteria;  Escherichia coli;  Escherichia coli Proteins;  Membrane Transport Proteins;  Molecular Docking Simulation;  Multigene Family;  Operon;  Phylogeny;  Pyrimidines;  Substrate Specificity;  Thymine;  Uracil",
    abstract = "The uracil permease UraA of Escherichia coli is a structurally known prototype for the ubiquitous Nucleobase-Ascorbate Transporter (NAT) or Nucleobase-Cation Symporter-2 (NCS2) family and represents a well-defined subgroup of bacterial homologs that remain functionally unstudied. Here, we analyze four of these homologs, including RutG of E. coli which shares 35% identity with UraA and is encoded in the catabolic rut (pyrimidine utilization) operon. Using amplified expression in E. coli K-12, we show that RutG is a high-affinity permease for uracil, thymine and, at low efficiency, xanthine and recognizes also 5-fluorouracil and oxypurinol. In contrast, UraA and the homologs from Acinetobacter calcoaceticus and Aeromonas veronii are permeases specific for uracil and 5-fluorouracil. Molecular docking indicates that thymine is hindered from binding to UraA by a highly conserved Phe residue which is absent in RutG. Site-directed replacement of this Phe with Ala in the three uracil-specific homologs allows high-affinity recognition and/or transport of thymine, emulating the RutG profile. Furthermore, all RutG orthologs from enterobacteria retain an Ala at this position, implying that they can use both uracil and thymine and, possibly, xanthine as substrates and provide the bacterial cell with a range of catabolizable nucleobases. © 2018 John Wiley & Sons Ltd"
}