Please use this identifier to cite or link to this item: http://repositorio.unicamp.br/jspui/handle/REPOSIP/55526
Type: Artigo de periódico
Title: Biochemical Characterization of Uracil Phosphoribosyltransferase from Mycobacterium tuberculosis
Author: Villela, AD
Ducati, RG
Rosado, LA
Bloch, CJ
Prates, MV
Goncalves, DC
Ramos, CHI
Basso, LA
Santos, DS
Abstract: Uracil phosphoribosyltransferase (UPRT) catalyzes the conversion of uracil and 5-phosphoribosyl-alpha-1-pyrophosphate (PRPP) to uridine 5'-monophosphate (UMP) and pyrophosphate (PPi). UPRT plays an important role in the pyrimidine salvage pathway since UMP is a common precursor of all pyrimidine nucleotides. Here we describe cloning, expression and purification to homogeneity of upp-encoded UPRT from Mycobacterium tuberculosis (MtUPRT). Mass spectrometry and N-terminal amino acid sequencing unambiguously identified the homogeneous protein as MtUPRT. Analytical ultracentrifugation showed that native MtUPRT follows a monomer-tetramer association model. MtUPRT is specific for uracil. GTP is not a modulator of MtUPRT ativity. MtUPRT was not significantly activated or inhibited by ATP, UTP, and CTP. Initial velocity and isothermal titration calorimetry studies suggest that catalysis follows a sequential ordered mechanism, in which PRPP binding is followed by uracil, and PPi product is released first followed by UMP. The pH-rate profiles indicated that groups with pK values of 5.7 and 8.1 are important for catalysis, and a group with a pK value of 9.5 is involved in PRPP binding. The results here described provide a solid foundation on which to base upp gene knockout aiming at the development of strategies to prevent tuberculosis.
Country: EUA
Editor: Public Library Science
Rights: aberto
Identifier DOI: 10.1371/journal.pone.0056445
Date Issue: 2013
Appears in Collections:Artigos e Materiais de Revistas Científicas - Unicamp

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