title: AP sites are not significantly involved in mutagenesis by the (+)-anti diol epoxide of benzo[a]pyrene: the complexity of its mutagenic specificity is likely to arise from adduct conformational polymorphism.
abstract: In previous work, mutations induced by the (+)-anti diol epoxide of benzo[a]pyrene [(+)-anti-B[a]PDE] were scored in the supF gene of the Escherichia coli plasmid pUB3 [Rodriguez & Loechler (1993) Biochemistry 32, 1759]. pUB3 was reacted with (+)-anti-B[a]PDE and then either (1) transformed immediately into E. coli or (2) heated at 80 degrees C for 10 min prior to transformation. Heating only released a small fraction of adducts (approximately 5%) and did not significantly affect the mutagenic pattern at most sites in supF. However, at the major base substitution hotspot, G115, principally G-->T mutations (87%) were obtained prior to heating, while after heating, G-->T mutations decreased (45%) and G-->A (21%) and G-->C (33%) mutations became more prevalent. One model for this result is that prior to heating a heat-labile adduct at G115 causes one pattern of mutagenesis, but after heating the labile adduct is hydrolyzed to an apurinic site (AP site), which causes a second mutational pattern. To test this, a role for AP sites generated from labile adducts by heating at 80 degrees C for 10 min is investigated. It is shown that when plasmid pUB3 contains 22 (+)-anti-B[a]PDE adducts, 0.6% (or fewer) are converted to AP sites as determined in an assay based upon the action of an AP-endonuclease. In a separate line of investigation not involving (+)-anti-B[a]PDE adducts, mutation frequency (MF) per AP site is estimated. (In these experiments, AP sites were introduced into pUB3 by the classic procedure of heating at 70 degrees C/pH 5.0 to hydrolyze purines.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Biology, Boston University, Massachusetts 02215.
authors.forename: E E
authors.initials: EE
authors.lastname: Drouin
authors.forename: E L
authors.initials: EL
authors.lastname: Loechler
mesh_terms: D015123:7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide / Q000633:toxicity*; D001080:Apurinic Acid / Q000378:metabolism*; D004269:DNA, Bacterial / Q000187:drug effects*; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D043223:Deoxyribonuclease IV (Phage T4-Induced); D004706:Endodeoxyribonucleases / Q000378:metabolism*; D004926:Escherichia coli / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics; D029968:Escherichia coli Proteins; D005798:Genes, Bacterial; D007700:Kinetics; D015394:Molecular Structure; D016296:Mutagenesis; D009153:Mutagens / Q000633:toxicity*; D009690:Nucleic Acid Conformation; D010957:Plasmids / Q000187:drug effects; D013237:Stereoisomerism
chemical_list: D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D009153:Mutagens; D001080:Apurinic Acid; D015123:7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide; D004706:Endodeoxyribonucleases; D043223:Deoxyribonuclease IV (Phage T4-Induced); C468972:endonuclease IV, E coli; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase