Structural and energetic characterization of the major DNA adduct formed from the food mutagen ochratoxin A in the NarI hotspot sequence: influence of adduct ionization on the conformational preferences and implications for the NER propensity

Abstract

The nephrotoxic food mutagen ochratoxin A (OTA) produces DNA adducts in rat kidneys, the major lesion being the C8-linked-2′-deoxyguanosine adduct (OTB-dG). Although research on other adducts stresses the importance of understanding the structure of the associated adducted DNA, site-specific incorporation of OTB-dG into DNA has yet to be attempted. The present work uses a robust computational approach to determine the conformational preferences of OTB-dG in three ionization states at three guanine positions in the NarI recognition sequence opposite cytosine. Representative adducted DNA helices were derived from over 2160 ns of simulation and ranked via free energies. For the first time, a close energetic separation between three distinct conformations is highlighted, which indicates OTA-adducted DNA likely adopts a mixture of conformations regardless of the sequence context. Nevertheless, the preferred conformation depends on the flanking bases and ionization state due to deviations in discrete local interactions at the lesion site. The structural characteristics of the lesion thus discerned have profound implications regarding its repair propensity and mutagenic outcomes, and support recent experiments suggesting the induction of double-strand breaks and deletion mutations upon OTA exposure. This combined structural and energetic characterization of the OTB-dG lesion in DNA will encourage future biochemical experiments on this potentially genotoxic lesion.

Adenine versus guanine DNA adducts of aristolochic acids: role of the carcinogen–purine linkage in the differential global genomic repair propensity

“…Most importantly, our work points toward the effects of the intrinsic adduct conformation on GGR recognition and repair propensity. If verified by future experimental studies, our results will likely expand the list of previously identified factors that affect the structure and repair propensity of adducted DNA, such as substitution at the carcinogen–base linkage ( 47 ), stereochemistry ( 29 , 48 – 49 ), adduct ionization state ( 50 ), sequence context ( 35 , 47 ) and the identity of the partner base ( 51 ).…”

“…To the best of our knowledge, no example exists in the literature of N2-dG adducted DNA in such a conformation. Nevertheless, this structure is analogous to the syn base−displaced intercalated conformation observed for damaged DNA corresponding to several C8-dG adducts ( 47 , 50 , 71 – 72 ) in terms of the (major groove) extrahelical opposing cytosine. However, damaged guanine is displaced towards the major groove in C8-dG adducted DNA versus the minor groove in ALII-N 2 -dG adducted DNA.…”

“…Although potential inaccuracies in free energy calculations may arise due to the theoretical simplicity of the MM-PBSA approach (e.g., implicit treatment of solvent), as well as limitations in our conformational sampling, the MM-PBSA free energies are expected to provide at least qualitative estimates of the energetic differences between possible conformations of damaged DNA ( 64 ). Furthermore, the choice of this method for the present application stems from our desire to compare the conformational preferences of ALII-N 2 -dG adducted DNA with those of ALII-N 6 -dA adducted DNA determined using the same protocol ( 46 ), as well as the successful applications of this method in previous studies of damaged DNA ( 50 , 65 – 67 ). Full details of the computational protocol are provided in Supplementary Data (Section S1).…”

Structural and biochemical impact of C8-aryl-guanine adducts within the NarI recognition DNA sequence: influence of aryl ring size on targeted and semi-targeted mutagenicity

“…The resulting N-linked C8-dG adducts contain a flexible amine tether separating the dG component from the aryl ring and there is a strong established relationship between the resulting conformational heterogeneity and biological effects ( 5 , 6 ). The structurally related aromatic organic carcinogens, such as polycyclic aromatic hydrocarbons (PAHs) ( 7 – 9 ), arylhydrazines ( 10 – 12 ) and phenols ( 13 – 16 ), may also undergo bioactivation, producing aryl radical species that covalently attach to the C8-site of dG to produce carbon-linked C8-aryl-dG adducts. In these cases, the adducts are similar in structure to their N-linked counterparts in terms of aryl ring size and shape, but do not have an amine linkage, thereby altering the orientation of the modification in the DNA duplex as well as reducing conformational flexibility.…”

Chlorine Functionalization of a Model Phenolic C8-Guanine Adduct Increases Conformational Rigidity and Blocks Extension by a Y-Family DNA Polymerase

“…25 In contrast to N-linked C8-dG adducts, C-linked lesions tend to induce B-type conformation, although conformational heterogeneity is observed depending on the size of the C8-aryl ring, 26 the nature and placement of substituents on the C8-aryl ring, 27 and the adduct ionization state. 28 Thus, C-and Nlinked adducts have different mutagenic outcomes. For example, in contrast to the nonmutagenic N-linked C8-aniline-dG adduct, 25 the single-ringed C8-phenyl-dG ([Ph]G) adduct induces transversion and frameshift mutations 26,29 and strongly blocks the progress of DNA replication by high-fidelity DNA polymerases: Klenow fragment exo − (Kf − ) 26,29 and DNA pol α.…”