Specificity of pyrimidine nucleoside phosphorylases and the phosphorolysis of 5-fluoro-2'-deoxyuridine.
Isoelectric focusing and studies with 1-(2'-deoxy-beta-D-glucopyranosyl)thymine (GPT), a specific inhibitor of uridine phosphorylase activity, were used to determine the substrate specificities of mammalian pyrimidine nucleoside phosphorylases and their cleavage of 5-fluoro-2'-deoxyuridine (FdUrd). Isoelectric focusing profiles for the cytosol fractions from Ehrlich ascites cells and from Novikoff hepatoma cells each consisted essentially of one peak of nucleoside phosphorylase activity [isoelectric points (pl) 5.4 and 5.8, respectively] that cleaved both uridine and thymidine (dThd), as well as FdUrd. By contrast, cytosol fractions from HeLa (S3) cells, mouse liver, and normal human leukocytes each exhibited a major peak of activity (pl 4.6, 6.5, and 4.9, respectively) that cleaved only dThd and FdUrd, while mouse liver exhibited a second peak (pl 5.2) that cleaved primarily uridine. To distinguish clearly between (a) uridine phosphorylases that cleave primarily uridine and that are inhibited by GPT and (b)dThd phosphorylases that cleave only deoxynucleosides and that are not inhibited by GPT, we propose the term "uridine-deoxyuridine phosphorylases" to define those pyrimidine nucleoside phosphorylases that cleave both uridine and dThd and that are inhibited by GPT. On the basis of this definition and studies with GPT in nonfocused cytosol preparations, we conclude that FdUrd is cleaved to 5-fluorouracil by uridine-deoxyuridine phosphorylase activity in Ehrlich ascites cells and in Novikoff hepatoma cells, and by dThd phosphorylases in mouse liver, in normal human leukocytes, and in HeLa (S3) cells.
D000818:Animals; D002286:Carcinoma, Ehrlich Tumor / Q000201:enzymology; D002478:Cells, Cultured; D005467:Floxuridine / Q000378:metabolism*; D006367:HeLa Cells / Q000201:enzymology; D006801:Humans; D007526:Isoelectric Point; D007962:Leukocytes / Q000201:enzymology; D008099:Liver / Q000201:enzymology; D008114:Liver Neoplasms, Experimental / Q000201:enzymology; D051379:Mice; D010430:Pentosyltransferases / Q000378:metabolism*; D011741:Pyrimidine Nucleosides / Q000378:metabolism; D057845:Pyrimidine Phosphorylases; D051381:Rats; D013379:Substrate Specificity; D013936:Thymidine / Q000031:analogs & derivatives / Q000494:pharmacology; D013939:Thymidine Phosphorylase / Q000378:metabolism; D014543:Uridine Phosphorylase / Q000037:antagonists & inhibitors / Q000378:metabolism
D011741:Pyrimidine Nucleosides; D005467:Floxuridine; C029221:1-(2'-deoxy-beta-D-glucopyranosyl)thymine; D010430:Pentosyltransferases; D057845:Pyrimidine Phosphorylases; D014543:Uridine Phosphorylase; D013939:Thymidine Phosphorylase; D013936:Thymidine