Marsch G A, Ward R L, Colvin M, Turteltaub K W
Biology and Biotechnology Research Program, Lawrence Livermore National Laboratory, Livermore, CA 94551.
Nucleic Acids Res. 1994 Dec 11;22(24):5408-15. doi: 10.1093/nar/22.24.5408.
The cooked meat mutagen 2-amino-1-methyl-6-phenyl-imidazo[4,5-b]pyridine (PhIP) is metabolized in vivo to electrophilic intermediates that covalently bind to DNA guanines. Here we address the mechanism of PhIP's non-covalent interaction with DNA by using spectroscopic and computational methodologies. NMR methodologies indicated that upon addition of DNA, PhIP aromatic protons underwent a small, 0.11-0.12 p.p.m. upfield shift. DNA phosphorus resonances of non-covalent PhIP-DNA complexes broadened and slightly shifted upfield, while DNA base imino proton resonances shifted slightly downfield relative to DNA alone. UV and fluorescence spectra of PhIP titrated with DNA showed no detectable shifting and hypochromism of absorbance or fluorescence bands. In the presence of DNA, PhIP fluorescence was efficiently quenched by acrylamide, but not by silver ion. Further, the NMR spectra suggest that PhIP is in fast exchange with the DNA, and is slightly specific for adenine-thymine (A-T) sequences. Finally, structural arguments based on quantum chemistry calculations suggested that PhIP and its metabolites are unlikely to intercalate into DNA. These data collectively indicate that PhIP non-covalently binds in a groove of DNA.
熟肉诱变剂2-氨基-1-甲基-6-苯基咪唑并[4,5-b]吡啶（PhIP）在体内代谢为亲电子中间体，这些中间体与DNA鸟嘌呤共价结合。在此，我们通过光谱和计算方法研究PhIP与DNA非共价相互作用的机制。核磁共振（NMR）方法表明，加入DNA后，PhIP的芳香族质子经历了一个小的、0.11 - 0.12 ppm的场向位移。非共价PhIP - DNA复合物的DNA磷共振变宽并略微向场向位移，而DNA碱基亚氨基质子共振相对于单独的DNA略微向低场位移。用DNA滴定的PhIP的紫外和荧光光谱显示吸光度或荧光带没有可检测到的位移和减色现象。在DNA存在下，PhIP荧光被丙烯酰胺有效猝灭，但不被银离子猝灭。此外，NMR光谱表明PhIP与DNA处于快速交换状态，并且对腺嘌呤 - 胸腺嘧啶（A - T）序列略有特异性。最后，基于量子化学计算的结构论证表明，PhIP及其代谢产物不太可能嵌入DNA。这些数据共同表明PhIP以非共价方式结合在DNA的一个凹槽中。
