Trypanosoma cruzi contains a single detectable uracil-DNA glycosylase and repairs uracil exclusively via short patch base excision repair.
Enzymes involved in genomic maintenance of human parasites are attractive targets for parasite-specific drugs. The parasitic protozoan Trypanosoma cruzi contains at least two enzymes involved in the protection against potentially mutagenic uracil, a deoxyuridine triphosphate nucleotidohydrolase (dUTPase) and a uracil-DNA glycosylase belonging to the highly conserved UNG-family. Uracil-DNA glycosylase activities excise uracil from DNA and initiate a multistep base-excision repair (BER) pathway to restore the correct nucleotide sequence. Here we report the biochemical characterisation of T.cruzi UNG (TcUNG) and its contribution to the total uracil repair activity in T.cruzi. TcUNG is shown to be the major uracil-DNA glycosylase in T.cruzi. The purified recombinant TcUNG exhibits substrate preference for removal of uracil in the order ssU>U:G>U:A, and has no associated thymine-DNA glycosylase activity. T.cruzi apparently repairs U:G DNA substrate exclusively via short-patch BER, but the DNA polymerase involved surprisingly displays a vertebrate POLdelta-like pattern of inhibition. Back-up UDG activities such as SMUG, TDG and MBD4 were not found, underlying the importance of the TcUNG enzyme in protection against uracil in DNA and as a potential target for drug therapy.
Peña-Diaz
Javier
J
Instituto de Parasitologia y Biomedicina "Lopez Neyra", Consejo Superior de Investigaciones Cientificas, C/Ventanilla 11, 18001 Granada, Spain.
Akbari
Mansour
M
Sundheim
Ottar
O
Farez-Vidal
M Esther
ME
Andersen
Sonja
S
Sneve
Ragnhild
R
Gonzalez-Pacanowska
Dolores
D
Krokan
Hans E
HE
Slupphaug
Geir
G
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D014355:Chagas Disease / Q000188:drug therapy / Q000469:parasitology; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D045647:DNA Glycosylases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D004260:DNA Repair; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005816:Genetic Complementation Test; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D014349:Trypanosoma cruzi / Q000201:enzymology* / Q000235:genetics; D014498:Uracil / Q000378:metabolism*; D051981:Uracil-DNA Glycosidase
D011994:Recombinant Proteins; D014498:Uracil; D004247:DNA; D045647:DNA Glycosylases; D051981:Uracil-DNA Glycosidase