title: Experimental and structural evidence that herpes 1 kinase and cellular DNA polymerase(s) discriminate on the basis of sugar pucker.
abstract: Two isomers of methanocarba (MC) thymidine (T), one an effective antiherpes agent with the pseudosugar moiety locked in the North (N) hemisphere of the pseudorotational cycle (1a, N-MCT) and the other an inactive isomer locked in the antipodean South (S) conformation (1b, S-MCT) were used to determine whether kinases and polymerases discriminate between their substrates on the basis of sugar conformation. A combined solid-state and solution conformational analysis of both compounds, coupled with the direct measurement of mono-, di-, and triphosphate levels in control cells, cells infected with the Herpes simplex virus, or cells transfected with the corresponding viral kinase gene (HSV-tk), suggests that kinases prefer substrates that adopt the S sugar conformation. On the other hand, the cellular DNA polymerase(s) of a murine tumor cell line transfected with HSV-tk incorporated almost exclusively the triphosphate of the locked N conformer (N-MCTTP), notwithstanding the presence of higher triphosphate levels of the S-conformer (S-MCTTP).
authors.affiliation: Laboratory of Medicinal Chemistry, Center for Cancer Research, National Cancer Institute at Frederick, National Institutes of Health, 376 Boyles St., Frederick, MD, 21702, USA. marquezv@dc37a.nci.nih.gov
authors.forename: Victor E
authors.initials: VE
authors.lastname: Marquez
authors.forename: Tsipi
authors.initials: T
authors.lastname: Ben-Kasus
authors.forename: Joseph J
authors.initials: JJ
authors.lastname: Barchi
authors.forename: Karen M
authors.initials: KM
authors.lastname: Green
authors.forename: Marc C
authors.initials: MC
authors.lastname: Nicklaus
authors.forename: Riad
authors.initials: R
authors.lastname: Agbaria
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D000998:Antiviral Agents / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D045744:Cell Line, Tumor; D002522:Chlorocebus aethiops; D004247:DNA / Q000378:metabolism; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry / Q000378:metabolism*; D018259:Herpesvirus 1, Human / Q000201:enzymology* / Q000235:genetics; D051379:Mice; D008958:Models, Molecular; D008968:Molecular Conformation; D019906:Nuclear Magnetic Resonance, Biomolecular; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D013936:Thymidine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology*; D013937:Thymidine Kinase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D014709:Vero Cells
chemical_list: D000970:Antineoplastic Agents; D000998:Antiviral Agents; D004247:DNA; D013937:Thymidine Kinase; D004259:DNA-Directed DNA Polymerase; C469995:(north)-methanocarbathymidine; D013936:Thymidine