Mao H, Deng Z, Wang F, Harris T M, Stone M P
Department of Chemistry and Center in Molecular Toxicology, Vanderbilt University, Nashville, Tennessee 37235, USA.
Biochemistry. 1998 Mar 31;37(13):4374-87. doi: 10.1021/bi9718292.
The structure of a formamidopyrimidine (FAPY) adduct arising from imidazole ring opening of the initially formed trans-8, 9-dihydro-8-(N7-guanyl)-9-hydroxyaflatoxin B1 adduct under basic conditions and positioned in the 5'-d(CTATFAPYGATTCA)-3'5'-d(TGAATCATAG)-3' oligodeoxynucleotide was determined. The FAPY adduct may be a major progenitor of aflatoxin B1-induced mutations in DNA. The freshly prepared sample showed biphasic melting, with transitions at 28 and 56 degreesC. NMR initially showed multiple subspectra. Over a period of several days at 4 degreesC, the sample converted to a single species with a Tm of 56 degreesC, 15 degrees C greater than the unmodified duplex. The deoxyribose was in the beta configuration about the anomeric carbon, evidenced by NOEs between FAPYG5 H3', H2', H2", and H1'. FAPY formation resulted in the loss of the guanine H8 proton, and the introduction of the formyl proton, which showed NOEs to FAPYG5 H1' and A6 N6Ha. A total of 31 NOEs from AFB1 to DNA protons were observed, mostly to the 5'-neighboring base, T4 in the modified strand. Sequential NOEs were interrupted between T4 and FAPYG5 in the modified strand, between C16 and A17 in the complementary strand, and between T4 N3H and FAPYG5 N1H. An NOE between FAPYG5 N1H and C16 N4H showed intact hydrogen bonding at FAPYG5C16. Upfield chemical shifts were observed for T4 H6 and A17 H8. Molecular dynamics calculations converged with pairwise rmsd differences of <0.9 A. The sixth root residual was 8.7 x 10(-2). The AFB1 moiety intercalated from the major groove between FAPYG5 and T4A17, and stacked with T4 and FAPYG5 and partially stacked with A17. The base step between T4A17 and FAPYG5*C16 was increased from 3.4 to 7 A. The duplex unwound by about 15 degrees. The FAPY formyl group was positioned to form a hydrogen bond with A6 N6Ha. Strong stacking involving the AFB1 moiety, and this hydrogen bond explains the thermal stabilization of four base pairs by this adduct, and may be a significant factor in its processing.
确定了在碱性条件下，最初形成的反式-8,9-二氢-8-(N7-鸟嘌呤基)-9-羟基黄曲霉毒素B1加合物经咪唑环开环产生的甲酰胺嘧啶（FAPY）加合物在5'-d(CTATFAPYGATTCA)-3'5'-d(TGAATCATAG)-3'寡聚脱氧核苷酸中的结构。FAPY加合物可能是黄曲霉毒素B1诱导DNA突变的主要前体。新制备的样品显示出双相熔解，转变温度分别为28℃和56℃。核磁共振最初显示有多个子谱。在4℃下放置几天，样品转变为单一物种，其熔解温度为56℃，比未修饰的双链体高15℃。脱氧核糖在异头碳处呈β构型，可以通过FAPYG5的H3'、H2'、H2"和H1'之间的核Overhauser效应（NOE）来证明。FAPY的形成导致鸟嘌呤H8质子的丢失，并引入了甲酰基质子，该质子与FAPYG5的H1'和A6的N6Ha显示出NOE。总共观察到31个从黄曲霉毒素B1到DNA质子的NOE，大部分是到修饰链中5'-相邻碱基T4的。修饰链中T4和FAPYG5之间、互补链中C16和A17之间以及T4的N3H和FAPYG5的N1H之间的顺序NOE被打断。FAPYG5的N1H和C16的N4H之间的NOE表明FAPYG5C16处的氢键完整。观察到T4的H6和A17的H8有高场化学位移。分子动力学计算收敛，成对均方根偏差小于0.9 Å。六次方根残差为8.7×10⁻²。黄曲霉毒素B1部分从FAPYG5和T4A17之间的大沟插入，并与T4和FAPYG5堆积，部分与A17堆积。T4A17和FAPYG5*C16之间的碱基步长从3.4 Å增加到7 Å。双链体解开约15°。FAPY甲酰基的位置可与A6的N6Ha形成氢键。涉及黄曲霉毒素B1部分的强堆积以及这种氢键解释了该加合物对四个碱基对的热稳定作用，并且可能是其加工过程中的一个重要因素。
