title: 5-Fluoro-2'deoxyuridine 5'-(p-azidophenyl phosphate), a potential photoaffinity label of thymidylate synthetase.
abstract: 5-Fluoro-2'-deoxyuridine 5'-(p-azidophenyl phosphate) (1), a potential photoaffinity labeling reagent for thymidylate synthetase from a methotrexate-resistant strain of Lactobacillus casei, has been synthesized and characterized. UV254 irradiation of mixtures of thymidylate synthetase with 1, containing 14C-labeled phenyl and 3H-labeled pyrimidine rings, in the presence of excess 5,10-methylenetetrahydrofolate, the cofactor for the reaction, produced two complexes, separable from the native enzyme by polyacrylamide gel electrophoresis, in which only the 3H-containing moiety was bound to the protein. When mixtures of enzyme and 1 were irradiated in the absence of cofactor, complexes separable from the native enzyme were not observed. However, the 14C-containing component of 1 was now bound to the protein in the absence of the 3H-containing portion. The results are discussed in terms of the topography of the enzyme active site.
mesh_terms: D000345:Affinity Labels / Q000138:chemical synthesis*; D001386:Azides; D055598:Chemical Phenomena; D002621:Chemistry; D003856:Deoxyuracil Nucleotides / Q000138:chemical synthesis*; D005468:Fluorodeoxyuridylate / Q000031:analogs & derivatives / Q000138:chemical synthesis*; D007780:Lacticaseibacillus casei / Q000201:enzymology; D008780:Methyltransferases / Q000037:antagonists & inhibitors*; D010782:Photolysis; D013940:Thymidylate Synthase / Q000037:antagonists & inhibitors*
chemical_list: C021192:5-fluoro-2'-deoxyuridine 5'-(4-azidophenyl phosphate); D000345:Affinity Labels; D001386:Azides; D003856:Deoxyuracil Nucleotides; D005468:Fluorodeoxyuridylate; D008780:Methyltransferases; D013940:Thymidylate Synthase