Measurement of nucleoside diphosphate kinase-Nm23 activity by anion-exchange high-performance liquid chromatography.
A first-order assay to detect the activity of nucleoside diphosphate kinase (NDP-kinase; EC 2.7.4.6) was developed. In this assay, the activity of NDP-kinase is measured using various deoxy- and ribonucleotide triphosphates as phosphate donors and dADP as phosphate acceptor. The enzyme activity is determined by quantifying, after anion-exchange HPLC, the amount of newly synthesized dATP. Contrary to the most common coupled enzymic assays or isotopic assays the use of different donor-acceptor pairs is not restricted. The resolution of the procedure described is limited only by the chromatographic separation of substrate and product pairs participating in the reaction.
Pulido-Cejudo
G
G
National Laboratory for Viral Oncology, LCDC, Health Canada, Ottawa, Ontario.
Gagnon
J
J
Leclerc
J M
JM
Jamison
K
K
Gordon
J
J
Campione-Piccardo
J
J
D000818:Animals; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D003600:Cytosol / Q000201:enzymology; D006160:Guanosine Triphosphate / Q000737:chemistry; D006801:Humans; D007700:Kinetics; D008099:Liver / Q000201:enzymology; D009363:Neoplasm Proteins / Q000096:biosynthesis; D009701:Nucleoside-Diphosphate Kinase / Q000032:analysis*; D010710:Phosphates / Q000737:chemistry; D012265:Ribonucleotides / Q000138:chemical synthesis / Q000302:isolation & purification; D014407:Tumor Cells, Cultured
D009363:Neoplasm Proteins; D010710:Phosphates; D012265:Ribonucleotides; D006160:Guanosine Triphosphate; D009701:Nucleoside-Diphosphate Kinase