CHARACTERIZATION OF THE STRUCTURE OF THE ANTHRAMYCIN-D(ATGCAT)2 ADDUCT BY NMR AND MOLECULAR MODELING STUDIES - DETERMINATION OF THE STEREOCHEMISTRY OF THE COVALENT LINKAGE SITE, ORIENTATION IN THE MINOR GROOVE OF DNA, AND EFFECT ON LOCAL DNA-STRUCTURE
CHARACTERIZATION OF THE STRUCTURE OF THE ANTHRAMYCIN-D(ATGCAT)2 ADDUCT BY NMR AND MOLECULAR MODELING STUDIES - DETERMINATION OF THE STEREOCHEMISTRY OF THE COVALENT LINKAGE SITE, ORIENTATION IN THE MINOR GROOVE OF DNA, AND EFFECT ON LOCAL DNA-STRUCTURE
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DOI：
10.1021/ja00165a004
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发表时间：
1990-04-25
影响因子：
15
通讯作者：
HURLEY, LH
中科院分区：
文献类型：
--
作者：
BOYD, FL;CHEATHAM, SF;HURLEY, LH
红霉素是抗肿瘤抗生素中吡咯[1,4]苯二氮卓类的一员。先前的研究表明，在DNA的小凹槽内，红霉素通过鸟嘌呤的N-2共价结合，产生相对不扭曲的DNA加合物。从炭素-d(ATGCAT)2加合物的质子核磁共振波谱（NOESY）中，我们得到的结果明确地将药物分子定向在DNA的小凹槽中。在共价或非共价修饰链上，红霉素质子和核苷酸质子之间的四组NOE交叉峰表明，该药物特异性地将芳香环定向到共价修饰的鸟嘌呤的3”侧。用j相关光谱（COSY）不能明确地确定蒽环类药物d(ATGCAT)2附着侧的几何形状。然而，当结合分子力学程序AMBER的建模结果时，可以自信地预测该位点的11S立体化学。核磁共振31P研究表明，炭素-d(ATGCAT)2加合物中的两个共振信号明显下移。两个下场31P核磁共振信号通过17O同位素标记和1H-31P二维j相关实验进行了分配，结果表明它们对应于共价修饰的脱氧鸟嘌呤5”侧的磷酸基团和相反链上的脱氧胞嘧啶。利用二维傅里叶变换核磁共振方法（COSY和NOESY）对红霉素-d(ATGCAT)2的非交换性碱和糖质子的共振信号进行了赋值。通过水溶液中二维核磁共振波谱的交叉峰的相对强度，得到了糖囊的构象细节、糖苷二面角以及蒽醌键对双链二级结构的影响。所有符合这种分析的糖都具有与b型DNA一致的构象。用AMBER进行分子力学计算，预测了与d(ATGCAT)2结合的红霉素的取向和立体化学性质。在共价键位点具有11S立体化学的物种，并且与红霉素的芳香环定向到共价修饰的红霉素-d(ATGCAT)2的鸟嘌呤的3‘’侧，似乎比其他三个可能的物种更受青睐。这是因为该物种的分子间结合更大，而不是螺旋畸变能更低。分子模型也符合实验确定的anthramycin-d(ATGCAT)2加合物的不扭曲性质。
Anthramycin is a member of the pyrrolo[1,4]benzodiazepine group of antitumor antibiotics. Previous studies have demonstrated that anthramycin binds covalently through N-2 of guanine within the minor groove of DNA, resulting in a relatively nondistortive DNA adduct. From the nuclear Overhauser effect spectroscopy (NOESY) proton NMR spectra of the anthramycin-d(ATGCAT)2 adduct, we have obtained results that unambiguously assign the orientation of the drug molecule in the minor groove of DNA. Four sets of NOE cross-peaks between anthramycin protons and nucleotide protons on either the covalently or the noncovalently modified strands reveal that the drug is specifically oriented with the aromatic ring to the 3''-side of the covalently modified guanine. Unequivocal assignment of the geometry at the side of attachment of anthramycin to d(ATGCAT)2 cannot be made by J-correlated spectroscopy (COSY). However, when combined with the results of modeling with the molecular mechanics program AMBER, and 11S stereochemistry at this site can be confidently predicted. 31P NMR studies show that two of the resonance signals in the anthramycin-d(ATGCAT)2 adduct have moved significantly downfield. Both downfield 31P NMR signals have been assigned by 17O isotopic labeling and 1H-31P two-dimensional J-correlation experiments and shown to correspond to the phosphates on the 5''-sides of the covalently modified deoxyguanine and the deoxycytosine on the opposite strand. Assignment of resonance signals of nonexchangeable base and sugar protons of the anthramycin-d(ATGCAT)2 has been made with two-dimensional Fourier transform NMR methods (COSY and NOESY). Conformational details about the sugar puckers, the glycosidic dihedral angles, and the effect of anthramycin bonding on secondary structure of the duplex have been obtained from the relative intensities of cross-peaks in the two-dimensional NMR spectra in aqueous solution. All of the sugars that are amenable to this analysis possess a conformation consistent with B-type DNA. Molecular mechanics calculations with AMBER are predictive of the orientation and stereochemistry of anthramycin bound to d(ATGCAT)2. The species having an 11S stereochemistry at the covalent bonding site and oriented with the aromatic ring of anthramycin to the 3''-side of the covalently modified guanine of anthramycin-d(ATGCAT)2 appears to be favored over the three other possible species. This is because of the greater intermolecular binding for this species rather than lower helix distortion energies. The molecular modeling is also in accord with the experimentally determined nondistortive nature of the anthramycin-d(ATGCAT)2 adduct.