Enhancing Bulge Stabilization through Linear Extension of C8-Aryl-Guanine Adducts to Promote Polymerase Blockage or Strand Realignment to Produce a C:C Mismatch

Abstract

Aryl radicals can react at the C8-site of 2'-deoxyguanosine (dG) to produce DNA adducts with a C8-C linkage (denoted C-linked). Such adducts are structurally distinct from those possessing a flexible amine (N-linked) or ether (O-linked) linkage, which separates the C8-aryl moiety from the guanine nucleobase. In the current study, two model C-linked C8-dG adducts, namely, C8-benzo[b]thienyl-dG ([BTh]G) and C8-(pyren-1-yl)-dG ([Py]G), were incorporated into the NarI (12mer, NarI(12) and 22mer, NarI(22)) hotspot sequence for frameshift mutations in bacteria. For the first time, C-linked C8-dG adducts are shown to stabilize the -2 deletion duplex within the NarI sequence. Primer-elongation assays employing Sulfolobus solfataricus P2 DNA polymerase IV (Dpo4) demonstrates the influence of C8-aryl ring size and shape in promoting Dpo4 blockage or strand realignment to produce a C:C mismatch downstream of the adduct site. Molecular dynamics simulations of the -2 deletion duplex suggest that both anti and syn adduct structures are energetically accessible. These findings provide a rationale for describing the biochemical outcome induced by C-linked C8-dG adducts when processed by Dpo4.