title: Biochemical and structural characterization of the hypoxanthine-guanine-xanthine phosphoribosyltransferase from Pyrococcus horikoshii.
abstract: The 6-oxopurine phosphoribosyltransferase (HPRT, EC 2.4.2.8) from the hyperthermophile Pyrococcus horikoshii was expressed in Escherichia coli and purified. Steady-state kinetic studies indicated that the enzyme is able to use hypoxanthine, guanine and xanthine. The first two substrates showed similar catalytic efficiencies, and xanthine presented a much lower value (around 20 times lower), but the catalytic constant was comparable to that of hypoxanthine. The enzyme was not able to bind to GMP-agarose, but was able to bind the other reverse reaction substrate, inorganic pyrophosphate, with low affinity (K(d) of 4.7+/-0.1 mM). Dynamic light scattering and analytical gel filtration suggested that the enzyme exists as a homohexamer in solution.
authors.affiliation: Laboratorio de Genômica e Expressão, Departamento de Genética e Evolução, IB-UNICAMP, Caixa Postal 6109, CEP 13083-970, Campinas, Brazil.
authors.forename: Deyse
authors.initials: D
authors.lastname: de Souza Dantas
authors.forename: Camila
authors.initials: C
authors.lastname: Ramos Dos Santos
authors.forename: Gonçalo Amarante
authors.initials: GA
authors.lastname: Guimarães Pereira
authors.forename: Francisco Javier
authors.initials: FJ
authors.lastname: Medrano
mesh_terms: D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002850:Chromatography, Gel; D002942:Circular Dichroism; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D006147:Guanine / Q000378:metabolism; D006157:Guanosine Monophosphate / Q000378:metabolism; D019271:Hypoxanthine / Q000378:metabolism; D008969:Molecular Sequence Data; D010430:Pentosyltransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D043902:Pyrococcus horikoshii / Q000201:enzymology* / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D019820:Xanthine / Q000378:metabolism
chemical_list: D019843:Archaeal Proteins; D019820:Xanthine; D019271:Hypoxanthine; D006147:Guanine; D006157:Guanosine Monophosphate; D010430:Pentosyltransferases; C083867:hypoxanthine-guanine-xanthine phosphoribosyltransferase