title: Effective utilization of N2-ethyl-2'-deoxyguanosine triphosphate during DNA synthesis catalyzed by mammalian replicative DNA polymerases.
abstract: Acetaldehyde is produced by metabolic oxidation of ethanol after drinking alcoholic beverages. This agent reacts with nucleosides and nucleotides, resulting in the formation of N2-ethyl-guanine residues. N2-ethyl-2'-deoxyguanosine (N2-ethyl-dG) adduct has been detected in the lymphocyte DNA of alcoholic patients [Fang, J. L., and Vaca, C. E. (1997) Carcinogenesis 18, 627-632]. Thus, the nucleotide pool is also expected to be modified by acetaldehyde. N2-Ethyl-2'-deoxyguanosine triphosphate (N2-ethyl-dGTP) was chemically synthesized. The utilization of N2-ethyl-dGTP during DNA synthesis was determined by steady-state kinetic studies. N2-Ethyl-dGTP was efficiently incorporated opposite template dC in reactions catalyzed by mammalian DNA polymerase alpha and delta. When pol alpha was used, the insertion frequency of N2-ethyl-dGTP was 400 times less than that of dGTP, but 320 times higher than that of 7,8-dihydro-8-oxo-2'-deoxyguanosine triphosphate (8-oxo-dGTP), an oxidative damaged nucleotide. Using pol delta, the insertion frequency of N2-ethyl-dGTP was only 37 times less than that of dGTP. The chain extension from dC:N2-ethyl-dG pair occurred much more rapidly: the extension frequencies for pol alpha and pol delta were only 3.8 times and 6.3 times, respectively, lower than that of dC:dG pair. We also found that N2-ethyl-dG can be detected in urine samples obtained from healthy volunteers who had abstained from drinking alcohol for 1 week before urine collection. This indicates that humans are exposed constantly to acetaldehyde even without drinking alcoholic beverages. Incorporation of N2-ethyl-dG adducts into DNA may cause mutations and may be related to the development of alcohol- and acetaldehyde-induced human cancers.
authors.affiliation: Research Center for Environmental Quality Control, Kyoto University, Shiga, Japan. matsuda@biwa.eqc.kyoto-u.ac.jp
authors.forename: T
authors.initials: T
authors.lastname: Matsuda
authors.forename: I
authors.initials: I
authors.lastname: Terashima
authors.forename: Y
authors.initials: Y
authors.lastname: Matsumoto
authors.forename: H
authors.initials: H
authors.lastname: Yabushita
authors.forename: S
authors.initials: S
authors.lastname: Matsui
authors.forename: S
authors.initials: S
authors.lastname: Shibutani
mesh_terms: D000328:Adult; D002384:Catalysis; D004247:DNA / Q000096:biosynthesis* / Q000737:chemistry; D018736:DNA Adducts / Q000138:chemical synthesis* / Q000652:urine; D004256:DNA Polymerase I / Q000737:chemistry*; D004258:DNA Polymerase III / Q000737:chemistry*; D003848:Deoxyguanine Nucleotides / Q000138:chemical synthesis / Q000737:chemistry* / Q000652:urine; D003849:Deoxyguanosine / Q000031:analogs & derivatives / Q000652:urine; D005260:Female; D006801:Humans; D007700:Kinetics; D008297:Male
chemical_list: D018736:DNA Adducts; D003848:Deoxyguanine Nucleotides; C117229:N(2)-ethyl-2'-deoxyguanosine triphosphate; C105669:N-7-ethyl-2'-deoxyguanosine 3'-monophosphate; D004247:DNA; D004256:DNA Polymerase I; D004258:DNA Polymerase III; D003849:Deoxyguanosine