Studies on the glycosomal orotate phosphoribosyl transferase of Trypanosoma cruzi.
The orotate phosphoribosyltransferase of the epimastigote form of Trypanosoma cruzi was studied in its particulate state in preparations derived from glycosomes. Maximum activity was observed at pH 9. There was little activity in the absence of Mg2+; optimum [Mg2+] was related to [5'-phosphoribosyl-alpha-1-pyrophosphate]; Mn2+ could partially substitute for it. Kinetic analyses ruled out a substituted mechanism and suggested instead that the mechanism may be sequential. The apparent Km orotate was 2 microM; that for 5'-phosphoribosyl-alpha-1-pyrophosphate was 8 microM. The enzyme could not use uracil as substrate and was apparently not regulated by naturally-occurring nucleotides. It was, however, sensitive to inhibition by a wide range of pyrimidine analogues, the most active of which was 5-fluoroorotate. These inhibitors were as effective against the enzyme activity of intact glycosomes as broken preparations. This observation, when considered with an apparent lack of latency, suggests that the enzyme is located on the outside of the glycosome. The product of its reaction, orotidine 5'-phosphate, did not exchange readily with exogenous orotidine 5'-phosphate, suggesting that it is channeled directly to orotidine 5'-phosphate decarboxylase, the next enzyme in the pathway.
D000818:Animals; D014355:Chagas Disease / Q000201:enzymology* / Q000469:parasitology; D006863:Hydrogen-Ion Concentration; D008274:Magnesium / Q000378:metabolism; D008830:Microbodies / Q000201:enzymology*; D009940:Organoids / Q000201:enzymology*; D009962:Orotate Phosphoribosyltransferase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009963:Orotic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D010430:Pentosyltransferases / Q000378:metabolism*; D010754:Phosphoribosyl Pyrophosphate / Q000378:metabolism; D013379:Substrate Specificity; D014349:Trypanosoma cruzi / Q000201:enzymology / Q000254:growth & development
D009963:Orotic Acid; D010754:Phosphoribosyl Pyrophosphate; C001242:5-fluoroorotic acid; D010430:Pentosyltransferases; D009962:Orotate Phosphoribosyltransferase; D008274:Magnesium