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Glycal formation in crystals of uridine phosphorylase.

Paul D, O'Leary SE, Rajashankar K, Bu W, Toms A, Settembre EC, Sanders JM, Begley TP, Ealick SE
Biochemistry 49 3499-509 (2010)
Related entries: 3ku4, 3kuk, 3kvv, 3kvy

Cited: 12 times
EuropePMC logo PMID: 20364833

Abstract

Uridine phosphorylase is a key enzyme in the pyrimidine salvage pathway. This enzyme catalyzes the reversible phosphorolysis of uridine to uracil and ribose 1-phosphate (or 2'-deoxyuridine to 2'-deoxyribose 1-phosphate). Here we report the structure of hexameric Escherichia coli uridine phosphorylase treated with 5-fluorouridine and sulfate and dimeric bovine uridine phosphorylase treated with 5-fluoro-2'-deoxyuridine or uridine, plus sulfate. In each case the electron density shows three separate species corresponding to the pyrimidine base, sulfate, and a ribosyl species, which can be modeled as a glycal. In the structures of the glycal complexes, the fluorouracil O2 atom is appropriately positioned to act as the base required for glycal formation via deprotonation at C2'. Crystals of bovine uridine phosphorylase treated with 2'-deoxyuridine and sulfate show intact nucleoside. NMR time course studies demonstrate that uridine phosphorylase can catalyze the hydrolysis of the fluorinated nucleosides in the absence of phosphate or sulfate, without the release of intermediates or enzyme inactivation. These results add a previously unencountered mechanistic motif to the body of information on glycal formation by enzymes catalyzing the cleavage of glycosyl bonds.

Articles - 3kvr mentioned but not cited (1)

  1. Glycal formation in crystals of uridine phosphorylase. Paul D, O'Leary SE, Rajashankar K, Bu W, Toms A, Settembre EC, Sanders JM, Begley TP, Ealick SE. Biochemistry 49 3499-3509 (2010)


Articles citing this publication (11)

  1. The products of 5-fluorouridine by the action of the pseudouridine synthase TruB disfavor one mechanism and suggest another. Miracco EJ, Mueller EG. J. Am. Chem. Soc. 133 11826-11829 (2011)
  2. A novel structural mechanism for redox regulation of uridine phosphorylase 2 activity. Roosild TP, Castronovo S, Villoso A, Ziemba A, Pizzorno G. J. Struct. Biol. 176 229-237 (2011)
  3. The Pseudouridine Synthases Proceed through a Glycal Intermediate. Veerareddygari GR, Singh SK, Mueller EG. J. Am. Chem. Soc. 138 7852-7855 (2016)
  4. Uridine phosphorylase from Trypanosoma cruzi: kinetic and chemical mechanisms. Silva RG, Schramm VL. Biochemistry 50 9158-9166 (2011)
  5. Active site conformational dynamics in human uridine phosphorylase 1. Roosild TP, Castronovo S. PLoS ONE 5 e12741 (2010)
  6. The crystal structure of Streptococcus pyogenes uridine phosphorylase reveals a distinct subfamily of nucleoside phosphorylases. Tran TH, Christoffersen S, Allan PW, Parker WB, Piskur J, Serra I, Terreni M, Ealick SE. Biochemistry 50 6549-6558 (2011)
  7. Crystal Structure of Schistosoma mansoni Adenosine Phosphorylase/5'-Methylthioadenosine Phosphorylase and Its Importance on Adenosine Salvage Pathway. Torini JR, Brandão-Neto J, DeMarco R, Pereira HD. PLoS Negl Trop Dis 10 e0005178 (2016)
  8. High-syn conformation of uridine and asymmetry of the hexameric molecule revealed in the high-resolution structures of Shewanella oneidensis MR-1 uridine phosphorylase in the free form and in complex with uridine. Safonova TN, Mikhailov SN, Veiko VP, Mordkovich NN, Manuvera VA, Alekseev CS, Kovalchuk MV, Popov VO, Polyakov KM. Acta Crystallogr. D Biol. Crystallogr. 70 3310-3319 (2014)
  9. Probing the catalytic mechanism of bovine CD38/NAD+ glycohydrolase by site directed mutagenesis of key active site residues. Kuhn I, Kellenberger E, Cakir-Kiefer C, Muller-Steffner H, Schuber F. Biochim. Biophys. Acta 1844 1317-1331 (2014)
  10. Human 2'-Deoxynucleoside 5'-Phosphate N-Hydrolase 1: Mechanism of 2'-Deoxyuridine 5'-Monophosphate Hydrolysis. Devi S, Carberry AE, Zickuhr GM, Dickson AL, Harrison DJ, da Silva RG. Biochemistry 62 2658-2668 (2023)
  11. Structural and catalytic analysis of two diverse uridine phosphorylases in Phytophthora capsici. Yang C, Li J, Huang Z, Zhang X, Gao X, Zhu C, Morris PF, Zhang X. Sci Rep 10 9051 (2020)


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