Molecular Pharmacology, Vol 8, 731-739, Copyright © 1972 by the American Society for Pharmacology and Experimental Therapeutics

Some Properties of a Pyrimidine Phosphoribosyltransferase from Murine Leukemia Cells

¹ Department of Pharmacology and Division of Oncology, University of Rochester School of Medicine and Dentistry, Rochester, New York 14642

A pyrimidine phosphoribosyltransferase (approximate mol wt 100,000) with a sharply defined specificity was partially purified from ascitic cells of the P388/38280 murine leukemia. This enzyme is involved in the conversion of the antineoplastic drug 5-fluorouracil (K_m = 100 µM) to pharmacologically active nucleotides. The lowest K_m value was obtained with orotic acid as substrate (K_m 50 µM). The enzyme could also utilize 5-fluoroorotate (K_m 85 µM) and uracil (K_m = 5 mM). Inhibition studies, using fluorouracil as substrate, indicate that this enzyme has a strong affinity for pyrimidines with a carboxyl or amino group at position 6, or a fluorine (but not a larger halogen) at position 5. A methyl group at position 5 markedly decreases affinity of the enzyme for all pyrimidines. The affinity of the enzyme for 6-carboxypyrimidines was greatly increased in the presence of dimethylsulfoxide, but the rate of the enzyme-catalyzed reaction was markedly decreased. The enzyme requires Mg²⁺ and phosphoribosyl pyrophosphate; the latter promotes stability at all temperatures. Enzyme extracted from a cell line made resistant to fluorouracil showed a decreased capacity to utilize fluorouracil as a substrate.