title: Allosteric regulation of purine nucleoside phosphorylase.
abstract: Purine nucleoside phosphorylase (EC 2.4.2.1) from bovine spleen is allosterically regulated. With the substrate inosine the enzyme displayed complex kinetics: positive cooperativity vs inosine when this substrate was close to physiological concentrations, negative cooperativity at inosine concentrations greater than 60 microM, and substrate inhibition at inosine greater than 1 mM. No cooperativity was observed with the alternative substrate, guanosine. The activity of purine nucleoside phosphorylase toward the substrate inosine was sensitive to the presence of reducing thiols; oxidation caused a loss of cooperativity toward inosine, as well as a 10-fold decreased affinity for inosine. The enzyme also displayed negative cooperativity toward phosphate at physiological concentrations of Pi, but oxidation had no effect on either the affinity or cooperativity toward phosphate. The importance of reduced cysteines on the enzyme is thus specific for binding of the nucleoside substrate. The enzyme was modestly inhibited by the pyrimidine nucleotides CTP (Ki = 118 microM) and UTP (Ki = 164 microM), but showed greater sensitivity to 5-phosphoribosyl-1-pyrophosphate (Ki = 5.2 microM).
authors.affiliation: Department of Biochemistry and Biophysics, University of North Carolina School of Medicine, Chapel Hill 27599-7260.
authors.forename: P A
authors.initials: PA
authors.lastname: Ropp
authors.forename: T W
authors.initials: TW
authors.lastname: Traut
mesh_terms: D000494:Allosteric Regulation; D000495:Allosteric Site; D000818:Animals; D002417:Cattle; D007288:Inosine / Q000378:metabolism / Q000494:pharmacology*; D007700:Kinetics; D008433:Mathematics; D008962:Models, Theoretical; D010754:Phosphoribosyl Pyrophosphate / Q000494:pharmacology; D011683:Purine-Nucleoside Phosphorylase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013154:Spleen / Q000201:enzymology; D014544:Uridine Triphosphate / Q000494:pharmacology
chemical_list: D007288:Inosine; D010754:Phosphoribosyl Pyrophosphate; D011683:Purine-Nucleoside Phosphorylase; D014544:Uridine Triphosphate