title: Structural basis of mapping the spontaneous mutations with 5-flurouracil in uracil phosphoribosyltransferase from Mycobacterium tuberculosis.
abstract: Tuberculosis (TB) remains the second leading cause of death from an infectious disease globally, despite the incessant efforts to control it. Research and development into new TB medicines is imperative for effective TB control; however, new strategies for the rational use of existing drugs, such as through the identification of new drug targets, could also significantly enhance this process. Key enzymes involved in the essential metabolic and regulatory pathways are usually sought in the pursuit of potential drug targets. Uracil phosphoribosyltransferase (UPRT) is a key salvage pathway enzyme in the synthesis of uridine 5'-monophosphate (UMP) and a probable target of 5-fluorouracil (5-FU) in Mycobacterium tuberculosis (Mtb). To date, there is no structure available for UPRT from Mtb (MtUPRT) that would assist in the identification of appropriate inhibitors for the enzyme. Here we report the structure of MtUPRT along with its spontaneous mutational studies in the presence of 5-FU. We further mapped these four single nucleotide polymorphisms (SNPs) onto the MtUPRT structure, with two residues found to be conserved among the MtUPRT homologs. Notably, none of these SNPs are located in the 5-FU binding pocket. However, the mutants harboring these mutations showed increased MICs (minimum inhibitory concentration) as compared to wild type strains. The present study will aid in the screening of inhibitors of MtUPRT and thus assist in TB drug design and development.
authors.affiliation: Department of Biological Sciences, National University of Singapore, 14 Science Drive 4, Singapore, 117543, Singapore; Novartis Institute for Tropical Diseases, 10 Biopolis Road, Singapore, 138670, Singapore.
authors.forename: Pramila
authors.initials: P
authors.lastname: Ghode
authors.affiliation: Department of Biological Sciences, National University of Singapore, 14 Science Drive 4, Singapore, 117543, Singapore.
authors.forename: Chacko
authors.initials: C
authors.lastname: Jobichen
authors.affiliation: Department of Biological Sciences, National University of Singapore, 14 Science Drive 4, Singapore, 117543, Singapore.
authors.forename: Sarath
authors.initials: S
authors.lastname: Ramachandran
authors.affiliation: Novartis Institute for Tropical Diseases, 10 Biopolis Road, Singapore, 138670, Singapore.
authors.forename: Pablo
authors.initials: P
authors.lastname: Bifani
authors.affiliation: Department of Biological Sciences, National University of Singapore, 14 Science Drive 4, Singapore, 117543, Singapore. Electronic address: dbsjayar@nus.edu.sg.
authors.forename: J
authors.initials: J
authors.lastname: Sivaraman
mesh_terms: D000595:Amino Acid Sequence; D005472:Fluorouracil / Q000494:pharmacology*; D008969:Molecular Sequence Data; D009154:Mutation; D009169:Mycobacterium tuberculosis / Q000201:enzymology* / Q000235:genetics; D010430:Pentosyltransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid
chemical_list: D010430:Pentosyltransferases; C023269:uracil phosphoribosyltransferase; D005472:Fluorouracil
keywords: 5-fluorouracil (5-FU); Mycobacterium tuberculosis (Mtb); Spontaneous mutants; Uracil phosphoribosyltransferase (UPRT); upp (Rv3309c)