Fox K R, Cons B M
Department of Physiology and Pharmacology, University of Southampton, U.K.
Biochemistry. 1993 Jul 20;32(28):7162-71. doi: 10.1021/bi00079a013.
We have studied the sequence-specific interaction of mithramycin with nucleosome core particles which have been reconstituted with various DNA fragments. Mithramycin binds to these DNAs without disrupting the integrity of the nucleosome and produces clear DNase I footprints centered around GC-rich regions. In some instances, the footprints produced on free DNA are resolved into two or more smaller sites when the DNA is complexed with the nucleosome core. In a few cases, novel footprints are produced in sequences which did not bind the drug in free DNA samples. The results are explained by suggesting mithramycin binds to GC-rich regions in which the minor groove faces away from the protein core, and which possess a wider than normal narrow groove on account of their location. Hydroxyl radical footprinting and diethyl pyrocarbonate modification confirm that mithramycin does not affect the rotational positioning of the nucleosome-bound DNA. Although distamycin and echinomycin induce novel DNase I digestion products in nucleosomal DNA which are consistent with the proposed change in DNA positioning [Low, C. M. L., Drew, H. R., & Waring, M. J. (1986) Nucleic Acids Res. 14, 6785-6801], hydroxyl radical footprinting and diethyl pyrocarbonate modification suggest these ligands do not change the rotational positioning of the DNA on the nucleosome cores.
我们研究了光神霉素与用各种DNA片段重构的核小体核心颗粒的序列特异性相互作用。光神霉素与这些DNA结合而不破坏核小体的完整性，并在富含GC的区域周围产生清晰的DNase I足迹。在某些情况下，当DNA与核小体核心复合时，游离DNA上产生的足迹会分解为两个或更多较小的位点。在少数情况下，在游离DNA样品中不与药物结合的序列中会产生新的足迹。结果表明，光神霉素结合到富含GC的区域，其中小沟背离蛋白质核心，并且由于其位置而具有比正常窄沟更宽的沟。羟基自由基足迹法和焦碳酸二乙酯修饰证实，光神霉素不影响核小体结合DNA的旋转定位。尽管偏端霉素和棘霉素在核小体DNA中诱导出与DNA定位的提议变化一致的新的DNase I消化产物[Low, C. M. L., Drew, H. R., & Waring, M. J. (1986) Nucleic Acids Res. 14, 6785 - 6801]，但羟基自由基足迹法和焦碳酸二乙酯修饰表明这些配体不会改变DNA在核小体核心上的旋转定位。
