title: N2-Substituted 2'-Deoxyguanosine Triphosphate Derivatives as Selective Substrates for Human DNA Polymerase κ.
abstract: N2-Alkyl-2'-deoxyguanosine triphosphate (N2-alkyl-dGTP) derivatives with methyl, butyl, benzyl, or 4-ethynylbenzyl substituents were prepared and tested as substrates for human DNA polymerases. N2-Benzyl-dGTP was equal to dGTP as a substrate for DNA polymerase κ (pol κ), but was a poor substrate for pols β, δ, η, ι, or ν. In vivo reactivity was evaluated through incubation of N2-4-ethynylbenzyl-dG with wild-type and pol κ deficient mouse embryonic fibroblasts. CuAAC reaction with 5(6)-FAM-azide demonstrated that only cells containing pol κ were able to incorporate N2-4-ethynylbenzyl-dG into the nucleus. This is the first instance of a Y-family-polymerase-specific dNTP, and this method could be used to probe the activity of pol κ in vivo.
authors.affiliation: Department of Biochemistry and Molecular Biology, Pennsylvania State University, 500 University Dr., Hershey, PA, 17033, USA.
authors.forename: A S Prakasha
authors.identifier: 0000-0002-0055-7103
authors.initials: AS
authors.lastname: Gowda
authors.affiliation: Department of Biochemistry and Molecular Biology, New York Medical College, Valhalla, NY, 10595, USA.
authors.forename: Marietta
authors.identifier: 0000-0001-8696-6962
authors.initials: M
authors.lastname: Lee
authors.affiliation: Department of Biochemistry and Molecular Biology, Pennsylvania State University, 500 University Dr., Hershey, PA, 17033, USA.
authors.forename: Thomas E
authors.identifier: 0000-0002-6805-3729
authors.initials: TE
authors.lastname: Spratt
mesh_terms: D000818:Animals; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D003848:Deoxyguanine Nucleotides / Q000138:chemical synthesis / Q000737:chemistry*; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D013379:Substrate Specificity; D000099270:Y-Family DNA Polymerases
chemical_list: D004259:DNA-Directed DNA Polymerase; D003848:Deoxyguanine Nucleotides; C029603:deoxyguanosine triphosphate; C404302:POLK protein, human; D000099270:Y-Family DNA Polymerases
keywords: DNA; DNA polymerases; fluorescent labelling; nucleotide analogues; nucleotides