brivudine and thymidine-5--triphosphate

The Drosophila melanogaster multisubstrate deoxyribonucleoside kinase (dNK; EC 2.7.1.145) has a high turnover rate and a wide substrate range that makes it a very good candidate for gene therapy. This concept is based on introducing a suicide gene into malignant cells in order to activate a prodrug that eventually may kill the cell. To be able to optimize the function of dNK, it is vital to have structural information of dNK complexes. In this study we present crystal structures of dNK complexed with four different nucleoside analogs (floxuridine, brivudine, zidovudine and zalcitabine) and relate them to the binding of substrate and feedback inhibitors. dCTP and dGTP bind with the base in the substrate site, similarly to the binding of the feedback inhibitor dTTP. All nucleoside analogs investigated bound in a manner similar to that of the pyrimidine substrates, with many interactions in common. In contrast, the base of dGTP adopted a syn-conformation to adapt to the available space of the active site.

Topics: Adenosine Diphosphate; Animals; Antimetabolites; Bromodeoxyuridine; Cytarabine; Cytidine Triphosphate; Drosophila melanogaster; Drosophila Proteins; Feedback; Floxuridine; Guanosine Triphosphate; Hydrogen Bonding; Inhibitory Concentration 50; Kinetics; Models, Chemical; Models, Molecular; Phosphotransferases (Alcohol Group Acceptor); Protein Binding; Protein Structure, Secondary; Structure-Activity Relationship; Thymine Nucleotides; X-Ray Diffraction; Zalcitabine; Zidovudine

The effect of E-5-(2-bromovinyl)-1-beta-D-arabinofuranosyluracil (BVaraU) on herpes simplex virus (HSV) replication was examined and compared with that of E-5-(2-bromovinyl)-2'-deoxyuridine (BVdUrd). The 50% inhibitory dose against HSV type 1 (HSV-1) was 0.1 microgram/ml compared with 0.008 microgram/ml for BVdUrd; the antimetabolic 50% inhibitory dose of BVaraU ranged from 20 to 95 micrograms/ml. The addition of 50 micrograms of BVaraU per ml to HSV-1-infected Vero cells decreased the synthesis of viral and cellular DNA by 37 and 28%, respectively. The 5'-triphosphate (BVaraUTP) competed with dTTP in DNA synthesis by the herpes-viral and cellular DNA polymerases; the apparent Ki values of HSV-1 DNA polymerase, DNA polymerase alpha, and DNA polymerase beta were 0.14, 0.32, and 5 microM, respectively. Thus, BVaraU was a less effective antiherpesvirus agent than BVdUrd; unlike BVdUrd, it did not appear to be internally incorporated into replicating DNA in virus-infected cells.

Topics: Animals; Bromodeoxyuridine; Cells, Cultured; DNA; DNA, Viral; Kinetics; Nucleic Acid Synthesis Inhibitors; Rabbits; Simplexvirus; Thymine Nucleotides; Virus Replication