title: Pyrimidine nucleotides impair phosphoribosylpyrophosphate (PRPP) synthetase subunit aggregation by sequestering magnesium. A mechanism for the decreased PRPP synthetase activity in hereditary erythrocyte pyrimidine 5'-nucleotidase deficiency.
abstract: The activity of phosphoribosylpyrophosphate (PRPP) synthetase (ATP: D-ribose-5-phosphate pyrophosphotransferase, EC 2.7.6.1) is decreased in the erythrocyte in hereditary pyrimidine 5'-nucleotidase (P5N) deficiency. Given the increased pyrimidine nucleotide content of the P5N-deficient erythrocyte, we evaluated the effects of prototypic pyrimidine nucleotides on the activity of PRPP synthetase. In normal hemolysate a 1.0 mM combination of cytidine tri-, di- and monophosphate (CTP/CDP/CMP) inhibited PRPP synthetase activity and changed the ribose 5-phosphate (R5P) saturation curve from a hyperbola to a biphasic shape. Untreated crude hemolysate from P5N-deficient erythrocytes showed a biphasic R5P kinetic curve. Since the activity of PRPP synthetase is dependent on its state of subunit aggregation, we examined PRPP synthetase subunit aggregation using gel permeation chromatography. P5N-deficient erythrocytes had a decreased absolute amount of aggregated PRPP synthetase and almost a total loss of disaggregated PRPP synthetase. Using normal hemolysate, 1 mM CTP/CDP/CMP interfered with the ability of 1.0 mM ATP and 2.0 mM MgCl2 to promote PRPP synthetase subunit aggregation. Increasing the MgCl2 to 6.0 mM overcame the inhibitory effect of CTP/CDP/CMP. Thus, the decreased PRPP synthetase activity of the P5N-deficient erythrocyte is due, at least in part, to the ability of the accumulated pyrimidine nucleotides to sequester magnesium and to interfere with the subunit aggregation of PRPP synthetase.
authors.affiliation: Department of Medicine, Harbor-UCLA Medical Center, UCLA School of Medicine, Torrance 90502.
authors.forename: N A
authors.initials: NA
authors.lastname: Lachant
authors.forename: C R
authors.initials: CR
authors.lastname: Zerez
authors.forename: K R
authors.initials: KR
authors.lastname: Tanaka
mesh_terms: D015720:5'-Nucleotidase; D000249:Adenosine Monophosphate / Q000097:blood; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000744:Anemia, Hemolytic, Autoimmune / Q000201:enzymology; D003565:Cytidine Diphosphate / Q000494:pharmacology; D003568:Cytidine Monophosphate / Q000494:pharmacology; D003570:Cytidine Triphosphate / Q000494:pharmacology; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans; D007700:Kinetics; D046911:Macromolecular Substances; D008274:Magnesium / Q000097:blood* / Q000494:pharmacology; D015636:Magnesium Chloride; D009708:Nucleotidases / Q000172:deficiency*; D010770:Phosphotransferases / Q000097:blood*; D011742:Pyrimidine Nucleotides / Q000494:pharmacology*; D012268:Ribose-Phosphate Pyrophosphokinase / Q000037:antagonists & inhibitors / Q000097:blood*; D012267:Ribosemonophosphates / Q000097:blood
chemical_list: D046911:Macromolecular Substances; D011742:Pyrimidine Nucleotides; D012267:Ribosemonophosphates; D015636:Magnesium Chloride; D000249:Adenosine Monophosphate; C031626:ribose-5-phosphate; D003565:Cytidine Diphosphate; D003570:Cytidine Triphosphate; D000255:Adenosine Triphosphate; D010770:Phosphotransferases; D012268:Ribose-Phosphate Pyrophosphokinase; D009708:Nucleotidases; D015720:5'-Nucleotidase; D003568:Cytidine Monophosphate; D008274:Magnesium