Erythrocyte pyrimidine 5'-nucleotidase and deoxynucleotidase isozymes: metallosensitivity and kinetics.
Erythrocytes from a subject with classical pyrimidine 5'-nucleotidase (P5N) deficiency (PND) possessed typically low nucleotidase activity with uridine 5'-monophosphate (UMP) and cytidine 5'-monophosphate (CMP) as the substrates with subnormal dephosphorylation of 2'-deoxy thymidine 5'-monophosphate (dTMP) and 2'-deoxyuridine 5'-monophosphate (dUMP) and intermediate activity with 2'-deoxycytidine 5'-monophosphate (dCMP). Contrary to previously reported data, there was enzyme activity with all five substrates. The metallosensitivities of the P5N isozymes using UMP, dUMP, dCMP and dTMP as the substrates were defined. Nucleotidase (P5N) is slightly more sensitive than 2'-deoxy-pyrimidine 5'-nucleotidase (dP5N) to inhibition by lead, copper and mercury. Chromium stimulated more enzyme activity with dCMP than other substrates. Apparent Michaelis constants (Km) were calculated for UMP, CMP, dUMP, dCMP, and dTMP for PND and control subjects. Optimal activity was at pH 7.0-7.8 when using UMP and CMP as the substrates but at pH 5.5-6.5 with dUMP, dCMP and dTMP as the substrates. The Km, pH optima and metallosensitivities were, however, consistent with electrophoretic evidence for two and probably three isozymes: P5N with maximal affinity for UMP and CMP; dP5N with maximal affinity for dUMP and dTMP; and a closely related dCMPase. The relative activity with representative substrates can be used to distinguish homozygotes and heterozygotes for P5N deficiency and subjects with heavy metal exposure.
Cook
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Department of Pediatrics, University of Nebraska Medical Center, Omaha 68105.
Kubitschek
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Stohs
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Angle
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D015720:5'-Nucleotidase; D000328:Adult; D002851:Chromatography, High Pressure Liquid; D002857:Chromium / Q000633:toxicity; D003300:Copper / Q000633:toxicity; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000097:blood / Q000172:deficiency; D007700:Kinetics; D007854:Lead / Q000633:toxicity; D008297:Male; D008670:Metals / Q000633:toxicity*; D009708:Nucleotidases / Q000037:antagonists & inhibitors / Q000097:blood* / Q000172:deficiency; D013379:Substrate Specificity
D007527:Isoenzymes; D008670:Metals; D002857:Chromium; D007854:Lead; D003300:Copper; D009708:Nucleotidases; D015720:5'-Nucleotidase