Structure of the anthramycin-d(ATGCAT)2 adduct from one- and two-dimensional proton NMR experiments in solution.
Structure of the anthramycin-d(ATGCAT)2 adduct from one- and two-dimensional proton NMR experiments in solution.
复制标题
来自溶液中一维和二维质子 NMR 实验的蒽霉素-d(ATGCAT)2 加合物的结构。
DOI：
10.1021/bi00347a011
复制
发表时间：
1985
期刊：
影响因子：
2.9
通讯作者：
Krugh,TR
中科院分区：
文献类型：
--
作者：
Graves,DE;Stone,MP;Krugh,TR
罗彻斯特大学化学系，罗彻斯特，纽约 14627 1984 年 12 月 28 日收稿；修订稿于 1985 年 6 月 19 日收到 摘要：一维和二维 400 MHz 质子 NMR 实验用于检查蒽霉素甲醚与自互补脱氧寡核苷酸 d (ATGCAT) 2 相互作用形成的共价加合物的溶液结构。利用化学位移的浓度依赖性和核奥豪瑟增强 (NOE) 实验来分配内的腺嘌呤 H2 质子游离 d (ATGCAT) 2 和加合物的小沟。这些研究表明，四个腺嘌呤 H2 质子之一与结合的蒽霉素非常接近，与游离双链体的腺嘌呤 H2 质子相比，这导致其上场位移为 0.3 ppm。二维自相关 (COSY) NMR 技术证明，蒽霉素与 d (ATGCAT) 2 双链体共价连接的作用导致位于加合物小沟内的选定脱氧核糖质子的屏蔽增加。利用二维 NOE (NOESY) 技术测定共价连接的蒽霉素和 d-(ATGCAT) 2 双链体的质子之间的相互作用。对这些数据的分析揭示了小沟内蒽霉素甲基、H6和H7质子与特定脱氧寡核苷酸质子之间的NOE交叉峰，从而允许确定小沟内药物的方向。非选择性反转恢复 (7j) 弛豫实验用于探测 anthramycin-d (ATGCAT) 2 加合物的结构和动态特性。这些数据表明，蒽霉素的结合改变了 d (ATGCAT) 2 双链体的相关时间，并在溶剂交换方面稳定了两个内部 A* T 碱基对。从 NMR 数据推导出的安-thramycin-d (ATGCAT) 2 加合物的溶液构象与模型构建研究一致 [Hurley, LH, & Petrusek, R. L.(1979) Nature (London) 282, 529-531; Petrusek, R. L., Anderson, G. L., Garner, T. F., Fannin, Q. L., Kaplan, D. J., Zimmer, S. G., & Hurley, LH (1981) Biochemistry 20, 1111-1119], e 蒽霉素的抗肿瘤活性（图 1A）归因于其与 DNA 相互作用的能力，从而抑制其生物合成核酸（Kohn 等人，1968；Stefanovic，1968；Horwitz 等人，1971；Glaubiger 等人，1974）。这种与 DNA 相互作用的确切性质在过去几年中一直是众多研究的主题。这些研究表明，蒽霉素与 DNA 形成不稳定的共价附着，跨越大约三个碱基对（Glaubiger 等人，1974 年；Kohn 和 Spears，1970 年；Kohn 等人，1974 年）。该键的稳定性取决于 DNA 二级结构的维持，并且在通过加热、酶消化或将 pH 降低至 < 7.0 使 DNA 变性时会丢失。蒽霉素与 DNA 的结合具有高度选择性，因此这项工作得到了国家癌症研究所拨款 CA-35251 和 CA-17865 的支持。
Department of Chemistry, University of Rochester, Rochester, New York 14627 Received December 28, 1984; Revised Manuscript Received June 19, 1985 abstract: One-and two-dimensional 400-MHz proton NMR experiments are used to examine the solution structure of the covalent adduct formed by the interaction of anthramycin methyl ether with the self-complementary deoxyoligonucleotide d (ATGCAT) 2. The concentration dependence of chemical shifts and nuclear Overhauser enhancement (NOE) experiments are utilized to assign the adenine H2 protons within the minor groove for both free d (ATGCAT) 2 and the adduct. These studies demonstrate that one of the four adenine H2 protons is in close proximity to the bound anthramycin and this results in itsupfield shift of 0.3 ppm compared to the adenine H2 protons of the free duplex. Effects of the covalent attachment of anthramycin to the d (ATGCAT) 2 duplex result in an increased shielding of selected deoxyribose protons located within the minor groove of the adduct, as demonstrated by two-dimensional autocorrelated (COSY) NMR techniques. Interactions between the protons of the covalently attached anthramycin and the d-(ATGCAT) 2 duplex are determined by utilizing two-dimensional NOE (NOESY) techniques. Analysis of these data reveals NOE cross-peaks between the anthramycin methyl, H6, and H7 protons with specific deoxyoligonucleotide protons within the minor groove, thus allowing the orientation of the drugwithin the minor groove to be determined. Nonselective inversion recovery (7j) relaxation experiments are used to probe the structural and dynamic properties of the anthramycin-d (ATGCAT) 2 adduct. These data suggest that the binding of anthramycin alters the correlation time of the d (ATGCAT) 2 duplex and stabilizes both of the internal A* T base pairs with respect to solvent exchange. The solutionconformation of the an-thramycin-d (ATGCAT) 2 adduct, as deduced from the NMR data, is in agreement with model-building studies [Hurley, LH, & Petrusek, R. L.(1979) Nature (London) 282, 529-531; Petrusek, R. L., Anderson, G. L., Garner, T. F., Fannin, Q. L., Kaplan, D. J., Zimmer, S. G., & Hurley, LH (1981) Biochemistry 20, 1111-1119], e antitumor activity of anthramycin (Figure 1A) has been attributed to its ability to interact with DNA resulting in the inhibition of the biosynthesis of nucleic acids (Kohn et al., 1968; Stefanovic, 1968; Horwitz et al., 1971; Glaubiger et al., 1974). The exact nature of this interaction with DNA has been the subject of numerous studies over the past several years. From these studies, anthramycin has been shown to form a labile covalent attachment to DNA spanning approximately three base pairs (Glaubiger et al., 1974; Kohn & Spears, 1970; Kohn et al., 1974). The stabilityof this bond is dependent upon the maintenance of the secondary structure of the DNA and is lost upon denaturation of the DNA by heating, by enzymatic digestion, or by lowering the pH to< 7.0. Anthramycin is highly selective in binding to DNA, requiring fThis work was supported by National Cancer Institute Grants CA-35251 and CA-17865.