Uridine phosphorylase from Schistosoma mansoni.
Uridine phosphorylase is the only pyrimidine nucleoside cleaving activity that can be detected in extracts of Schistosoma mansoni. The enzyme is distinct from the two purine nucleoside phosphorylases contained in this parasite. Although Urd is the preferred substrate, uridine phosphorylase can also catalyze the reversible phosphorolysis of dUrd and dThd, but not Cyd, dCyd, or orotidine. The enzyme was purified 170-fold to a specific activity of 2.76 nmol/min/mg of protein with a 16% yield. It has a Mr of 56,000 as determined by molecular sieving on Sephadex G-100. The mechanism of uridine phosphorylase is sequential. When Urd was the substrate, the KUrd = 13 microM and the KPi = 533 +/- 78 microM. When dThd was used as a substrate, the KdThd = 54 microM and the KPi = 762 +/- 297 microM. The Vmax with dThd was 53 +/- 9.8% that of Urd. dThd was a competitive inhibitor when Urd was used as a substrate. The enzyme showed substrate inhibition by Urd, dThd (greater than 0.125 mM) and phosphate (greater than 10 mM). 5-(Benzyloxybenzyloxybenzyl)acyclouridine was identified as a potent and specific inhibitor of parasite (Ki = 0.98 microM) but not host uridine phosphorylase. Structure-activity relationship studies suggest that uridine phosphorylase from S. mansoni has a hydrophobic pocket adjacent to the 5-position of the pyrimidine ring and indicate differences between the binding sites of the mammalian and parasite enzymes. These differences may be useful in designing specific inhibitors for schistosomal uridine phosphorylase which will interfere selectively with nucleic acids synthesis in this parasite.
el Kouni
M H
MH
Division of Biology and Medicine, Brown University, Providence, Rhode Island 02912.
Naguib
F N
FN
Niedzwicki
J G
JG
Iltzsch
M H
MH
Cha
S
S
D000818:Animals; D007700:Kinetics; D008970:Molecular Weight; D010430:Pentosyltransferases / Q000378:metabolism*; D012550:Schistosoma mansoni / Q000201:enzymology*; D013329:Structure-Activity Relationship; D014498:Uracil / Q000031:analogs & derivatives / Q000494:pharmacology; D014543:Uridine Phosphorylase / Q000378:metabolism*
C034753:5-benzylacyclouridine; C055762:5-(benzyloxybenzyloxybenzyl)acyclouridine; D014498:Uracil; D010430:Pentosyltransferases; D014543:Uridine Phosphorylase