Structural Refinement of the 8,9-Dihydro-8-(N7-guanyl)-9-hydroxy-aflatoxin B₁ Adduct in a 5‘-Cp^AFBG-3‘ Sequence

Abstract

The structure of the cationic 8,9-dihydro-8-(N7-guanyl)-9-hydroxy-aflatoxin B(1) adduct embedded in a 5'-CpG-3' sequence context and paired with deoxycytosine in the oligodeoxynucleotide d(ACATC(AFB)GATCT) x d(AGATCGATGT) was refined using molecular dynamics calculations restrained by NOE data and dihedral angle restraints obtained from NMR data. The aflatoxin moiety intercalated above the 5' face of the modified guanine. It stacked between C(5) x G(16) and (AFB)G(6) x C(15). The AFB(1) H5, OCH(3), and methylene protons faced into the minor groove, with the methylene protons oriented between the C(15) and G(16) nucleobases. The aflatoxin B(1) H6a, H8, H9, and H9a protons faced the major groove, with H6a and H9a pointing toward the 5' direction from the lesion site. The refined structure was compared to the structure of the aflatoxin B(1) adduct embedded in a 5'-ATGCAT-3' sequence in the oligodeoxynucleotide d(TAT(AFB)GCATA)(2) [Jones, W. R., Johnston, D. S., and Stone, M. P. (1998) Chem. Res. Toxicol.11, 873-881]. The structure of the intercalated aflatoxin B(1) lesion in the ATC(AFB)GAT sequence is similar to its structure in the d(AT(AFB)GCAT) sequence. This is consistent with a mechanism in which the precovalent intercalation of aflatoxin-8,9-exo-epoxide on the 5' face of guanine places the epoxide in close proximity and in the proper orientation to the N7 position of guanine, thus facilitating an S(N)2 reaction. The data provides additional insight into the nature of the disruption of the B-DNA duplex induced by aflatoxin B(1) intercalation. Overall, the results suggest that sequence contributes a minor role in modulating the structure of the cationic guanine N7 AFB(1) lesion in duplex DNA. On the other hand, structural differences are observed when the correctly paired structure is compared to the structure of the cationic AFB(1) adduct mispaired with dA [Giri, I., Johnston, D. S., and Stone, M. P. (2002) Biochemistry 41, 5462-5472].

Thermal stabilization of the DNA duplex by adducts of aflatoxin B1

“…Figure 3 shows an example using the sitespecifically modified duplex d(ACATC AFB GATCT)⅐ d(AGATCGATGT) duplex. 61,62 In this instance, guanine H8 of of the 5Ј-neighbor base pair. It did, however, exhibit an NOE to the H6a proton of the intercalated AFB 1 moiety, which provided the missing connectivity to C5 H6.…”

“…For example, in the d(ACATC AFB GATCT)⅐ d(AGATCGATGT) duplex a total of 39 NOEs were observed between the AFB 1 adduct and the DNA. 61,62 Most were intrastrand connectivities, although, significantly, five were cross-strand NOEs. Intranucleotide NOEs were observed between the AFB 1 4-OCH 3 …”

“…To solve this problem, the d(AT AFB GCAT) 2 sequence motif was embedded into the corresponding 8-mer 2 , which allowed for structural refinement of the cationic adduct placed opposite deoxycytosine in the doubly adducted duplex. 43 Similarly, the solution structure of the cationic guanine N7 adduct placed opposite dC embedded in d(ACATC AFB GATCT)⅐d(AGATCGATGT) 61,62 was solved.…”

“…Comparison of the lesion site in the CpG 61,62 or the TpG 43 sequence, both from major groove and viewed from above ( Figure 5), revealed similar stacking interactions among the AFB 1 moiety and the DNA. In both instances, the cytosines complementary to AFB G were stacked into the helices, and the helices were spanned by the cationic AFB 1 adduct.…”

A Single Site-specific trans-Opened 7,8,9,10-Tetrahydrobenzo[a]pyrene 7,8-Diol 9,10-EpoxideN2-Deoxyguanosine Adduct Induces Mutations at Multiple Sites in DNA

“…In contrast, the 10R BPDE-dG adduct, which orients in the opposite direction toward the 3Ј-end of the modified strand in duplex DNA (16), was not found to cause any mutations at non-target bases on either side of the adduct. Interestingly, about 13% of the total mutations in SOS-induced E. coli caused by a dG adduct of aflatoxin B 1 8,9-epoxide (43), which intercalates into the helix on the 5Ј-side of the modified G base (47,48), were primarily C 3 T mutations at the dC immediately 5Ј to the lesion (G) in a 5ЈϳCGAϳ sequence. In contrast, an N-(deoxyguanosin-8-yl)-2-acetylaminofluorene lesion, whose hydrocarbon moiety displaces the modified G and intercalates in its place opposite the complementary C (49), gave rise to base misincorporation on the 3Ј-side of the lesion site (50).…”