31P NMR study of erythrocytes from a patient with hereditary pyrimidine-5'-nucleotidase deficiency.

Abstract

The composition of phosphate metabolites and the intracellular pH in erythrocytes from a patient with hereditary pyrimidine-5''-nucleotidase deficiency were examined using 31P NMR spectroscopy. Several resonances were identified in spectra from intact cells and from extracts. The 2,3-bisphosphoglycerate line intensities were normal, but the NTP [nucleotide triphosphate] resonances were about twice normal due to the presence of millimolar quantities of pyrimidine phosphates. Several intense resonances were also observed in the diphosphodiester region of the spectrum. One compound contributing to these lines was identified as cytidine diphosphocholine. The resonances of NTP were in a position indicating that the additional triphosphates were also bound by Mg2+. Direct measurement shows that there is a nearly proportional increase in total cell Mg2+ in the patient''s cells, in agreement with the interpretation of the spectra. The intracellular pH was .apprx. 0.2 unit lower in the patient''s erythrocytes. This lower pH is due to the elevation in intracellular fixed negative charges and the shift in permeable anions consequent to the Donnan equilibrium. The lower intracellular pH may explain the lower oxygen affinity of these cells in the presence of otherwise normal 2,3-bisphosphoglycerate levels and the increased Mg2+ triphosphates level, because the Mg2+ form of NTP is known to not alter the O2 affinity of Hb under physiologic conditions. The lower intracellular pH can also explain the abnormalities in glycolytic intermediates observed for these cells.