Theruvathu Jacob A, Kim Cherine H, Rogstad Daniel K, Neidigh Jonathan W, Sowers Lawrence C
Department of Basic Sciences, School of Medicine, Loma Linda University, Loma Linda, California 92350, USA.
Biochemistry. 2009 Aug 11;48(31):7539-46. doi: 10.1021/bi9007947.
Inflammation-mediated reactive molecules can damage DNA by oxidation and chlorination. The biological consequences of this damage are as yet incompletely understood. In this paper, we have constructed oligonucleotides containing 5-chlorouracil (ClU), one of the known inflammation damage products. The thermodynamic stability, base pairing configuration, and duplex conformation of oligonucleotides containing ClU paired opposite adenine have been examined. NMR spectra reveal that the ClU-A base pair adopts a geometry similar to that of the T-A base pair, and the ClU-A base pair-containing duplex adopts a normal B-form conformation. The line width of the imino proton of the ClU residue is substantially greater than that of the corresponding T imino proton; however, this difference is not attributed to a reduced thermal or thermodynamic stability or to an increased level of proton exchange with solvent. While the NMR studies reveal an increased level of chemical exchange for the ClU imino proton of the ClU-A base pair, the ClU residue is not a target for removal by the Escherichia coli mispaired uracil glycosylase, which senses damage-related helix instability. The results of this study are consistent with previous reports indicating that the DNA of replicating cells can tolerate substantial substitution with ClU. The fraudulent, pseudo-Watson-Crick ClU-A base pair is sufficiently stable to avoid glycosylase removal and, therefore, might constitute a persistent form of cellular DNA damage.
炎症介导的反应性分子可通过氧化和氯化作用损伤DNA。这种损伤的生物学后果尚未完全明了。在本文中，我们构建了含有5-氯尿嘧啶（ClU）（一种已知的炎症损伤产物）的寡核苷酸。对与腺嘌呤配对的含ClU寡核苷酸的热力学稳定性、碱基配对构型和双链构象进行了研究。核磁共振光谱显示，ClU-A碱基对采用的几何结构类似于T-A碱基对，且含ClU-A碱基对的双链体采用正常的B型构象。ClU残基亚氨基质子的线宽明显大于相应T亚氨基质子的线宽；然而，这种差异并非归因于热稳定性或热力学稳定性的降低，也不是由于与溶剂的质子交换水平增加。虽然核磁共振研究表明ClU-A碱基对中ClU亚氨基质子的化学交换水平增加，但ClU残基并不是大肠杆菌错配尿嘧啶糖基化酶识别损伤相关螺旋不稳定性后进行切除的靶点。本研究结果与先前的报道一致，即复制细胞的DNA能够耐受大量ClU的取代。这种虚假的、类似沃森-克里克的ClU-A碱基对足够稳定，可避免糖基化酶的切除，因此可能构成细胞DNA损伤的一种持续形式。
