A variety of mutagenic/carcinogenic agents induce the formation of ring-opened, formamidopyrimidine dG and dA lesions, e.g., via ROS either spontaneously or with ionizing radiation. In spite of their potential importance, little is known about the biological significance of Fapy adducts. The principal investigator has developed some novel chemical strategies for preparing site-specifically modified oligonucleotides containing lesions that are sensitive to base deprotection, including Fapy(via Pd(0) deprotection and photolytic release from solid supports). Preliminary studies with another lesion (5OH-T) are described. Proposed are syntheses of oligos containing dA-Fapy and dG-Fapy, along with a variety of derivatives, which will be investigated for replication (in vitro polymerase bypass) (#1), as substrates for Fapy glycosylase (#2), as inhibitors of nucleases (#3), in thermodynamic studies (#4), and in structural analysis by NMR (#5). (1)5,6-Dihydro-5-hydroxythymine (5OH-T) was prepared by the principal investigator's newly developed, novel synthetic procedure. (2) Steady state polymerase bypass was investigated (Goodman approach), and kinetic parameters determined for incorporation of bases opposite the lesions and for bypass. 5OH-T appears to be a blocking but non-mutagenic lesion (e.g.) with Kf(exo-).
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