Zegar I S, Kim S J, Johansen T N, Horton P J, Harris C M, Harris T M, Stone M P
Center in Molecular Toxicology, Vanderbilt University, Nashville, Tennessee 37235, USA.
Biochemistry. 1996 May 21;35(20):6212-24. doi: 10.1021/bi9524732.
The structure of the (-)-(7S,8R,9S,10R)-N6-[10-(7,8,910-tetrahydrobenzo [a]pyrenyl)]-2'-deoxyadenosyl adduct at X6 of 5'-d(CGGACXAGAAG)-3'-5'-d(CTTCTTGTCCG)-3', derived from trans addition of the exocyclic N6-amino group of dA to (-)-(7S,8R,9R,10S)-7, 8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene [(-)-DE2], was determined using molecular dynamics simulations restrained by 369 NOEs from 1H NMR. This was named the SRSR(61,2) adduct, derived from the N-ras protooncogene at and adjacent to the nucleotides encoding amino acid 61 (underlined) of the p21 gene product. NOEs between C5, S.R.S.R A6, and A7 were disrupted, as were those between T17 and G18. NOEs between benzo[a]pyrene and DNA protons were localized on the two faces of the pyrenyl ring. The benzo[a]pyrene H3-H6 protons showed NOEs to T17 CH3, while H1, H2, and H3 showed NOEs to T17 deoxyribose; the latter protons and H4 showed NOEs to T17 H2', H2" and to T17 H6. Noes were observed between H11 and H12 and C5 H]',H2', H2". G18 N1H showed NOEs to both faces of benzo[a]pyrene. Upfield shifts of 2.6 ppm for T17 N3H and 1.8 ppm for G18 N1H. 1 ppm for T17 H6 and CH3, and 0.75 ppm for C5 H5, with a smaller shift for C5 H6, and a 1.5 ppm dispersion of the pyrenyl protons suggested that benzo[a]pyrene intercalated above the 5'-face of S.R.S.R A6. The precision of the refined structures was monitored by pairwise root mean square deviations. which were < 1.5 A; accuracy was measured by complete relaxation matrix calculations, which yielded a sixth root R factor of 8.1 x 10(-2). Interstrand stacking between the pyrenyl ring and the T17 pyrimidine and G18 purine rings was enhanced by the bay ring. Changes of +30 degrees and -25 degrees in buckle for C5.G18 and S.R.S.R A6.T17, respectively, were calculated, as was a -40 degrees change in propeller twist for C5.G18. The rise between C5.G18 and S.R.S.R A6.T17 was calculated to be 7 A. The work extended the pattern for adenine N6 benzo[a]pyrene adducts, in which the R stereochemistry at C10 predicted 5'-intercalation of the pyrenyl moiety.
通过分子动力学模拟确定了5'-d(CGGACXAGAAG)-3'-5'-d(CTTCTTGTCCG)-3'中X6处(-)-(7S,8R,9S,10R)-N6-[10-(7,8,9,10-四氢苯并[a]芘基)]-2'-脱氧腺苷加合物的结构，该加合物源自dA的环外N6-氨基与(-)-(7S,8R,9R,10S)-7,8-二羟基-9,10-环氧-7,8,9,10-四氢苯并[a]芘[(-)-DE2]的反式加成，使用了来自1H NMR的369个NOE进行约束。这被命名为SRSR(61,2)加合物，源自p21基因产物编码氨基酸61（下划线）的核苷酸处及相邻的N-ras原癌基因。C5、S.R.S.R A6和A7之间的NOE以及T17和G18之间的NOE被破坏。苯并[a]芘与DNA质子之间的NOE定位于芘基环的两个面上。苯并[a]芘的H3-H6质子与T17 CH3显示NOE，而H1、H2和H3与T17脱氧核糖显示NOE；后一组质子和H4与T17 H2'、H2"以及T17 H6显示NOE。在H11和H12与C� H1'、H2'、H2"之间未观察到NOE。G18 N1H与苯并[a]芘的两个面都显示NOE结果。T17 N3H有2.6 ppm的高场位移，G18 N1H有1.8 ppm的高场位移，T17 H6和CH3有1 ppm的高场位移，C5 H5有0.75 ppm的高场位移，C5 H6位移较小，芘基质子有1.5 ppm的分散，表明苯并[a]芘插入到S.R.S.R A6的5'-面上方。通过成对均方根偏差监测精制结构的精度小于1.5 Å；通过完全弛豫矩阵计算测量准确性，得到六次根R因子为8.1×10⁻²。芘基环与T17嘧啶和G18嘌呤环之间的链间堆积通过湾区环得到增强。计算得出C5.G18的扣环变化为+30°，S.R.S.R A6.T17的扣环变化为-25°，C5.G18的螺旋扭转变化为-40°。计算得出C5.G18和S.R.S.R A6.T17之间的上升为7 Å。这项工作扩展了腺嘌呤N6苯并[a]芘加合物的模式，其中C10处的R立体化学预测了芘基部分的5'-插入。
