Herpes simplex virus type 1 uracil-DNA glycosylase: isolation and selective inhibition by novel uracil derivatives.
We have purified Herpes simplex type 1 (HSV1) uracil-DNA glycosylase from the nuclei of HSV1-infected HeLa cells harvested 8 h post-infection, at which time the induction of the enzyme is a maximum. The enzyme has been shown to be distinct from the host enzyme, isolated from HeLa cells, by its lack of sensitivity to a monoclonal antibody to human uracil-DNA glycosylase. Furthermore, several uracil analogues were synthesized and screened for their capacity to discriminate between the viral and human uracil-DNA glycosylases. Both enzymes were inhibited by 6-(p-alkylanilino)uracils, but the viral enzyme was significantly more sensitive than the HeLa enzyme to most analogues. Substituents providing the best inhibitors of HSV1 uracil-DNA glycosylase were found to be in the order: p-n-butyl < p-n-pentl = p-n-hexyl < p-n-heptyl < p-n-octyl. The most potent HSV1 enzyme inhibitor, 6-(p-n-octylanilino)uracil (OctAU), with an IC50 of 8 microM, was highly selective for the viral enzyme. Short-term [3H]thymidine incorporation into the DNA of HeLa cells in culture was partially inhibited by OctAU, whereas it was unchanged when 6-(p-n-hexylanilino)uracil was present at concentrations that completely inhibited HSV1 uracil-DNA glycosylase activity. These compounds represent the first class of inhibitors that inhibit HSV1 uracil-DNA glycosylase at concentrations in the micromolar range. The results suggest their possible use to evaluate the functional role of HSV1 uracil-DNA glycosylase in viral infections and re-activation in nerve cells.
Focher
F
F
Istituto di Genetica Biochimica ed Evoluzionistica, CNR, Pavia, Italy.
Verri
A
A
Spadari
S
S
Manservigi
R
R
Gambino
J
J
Wright
G E
GE
D002467:Cell Nucleus / Q000201:enzymology; D045647:DNA Glycosylases; D004260:DNA Repair; D004591:Electrophoresis, Polyacrylamide Gel; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D009699:N-Glycosyl Hydrolases / Q000037:antagonists & inhibitors* / Q000302:isolation & purification*; D018139:Simplexvirus / Q000201:enzymology*; D013329:Structure-Activity Relationship; D013936:Thymidine / Q000378:metabolism; D014498:Uracil / Q000031:analogs & derivatives* / Q000494:pharmacology*; D051981:Uracil-DNA Glycosidase
C095137:6-(N-octylanilino)uracil; D014498:Uracil; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases; D051981:Uracil-DNA Glycosidase; D013936:Thymidine