Hoechst 33258 binds to G-quadruplex in the promoter region of human c-myc
Hoechst 33258 binds to G-quadruplex in the promoter region of human c-myc
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DOI：
10.1016/j.bbrc.2003.09.052
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发表时间：
2003-10-17
影响因子：
3.1
通讯作者：
Chowdhury, S
中科院分区：
文献类型：
--
作者：
Maiti, S;Chaudhury, NK;Chowdhury, S
用平衡吸收、荧光和动力学表面等离子体共振法研究了Hoechst 33258在体内外与人c-myc，d(GG、GGAGGG、TGG、GGA、GGG、TGG、GGA、AGG、TGG、GGA、AGG、TGG、GG)启动子区域的结合。四链体存在时，Hoechst的紫外吸收光谱的减色效应和荧光发射峰的蓝移表明Hoechst与四链体结合，UV和荧光滴定数据分析表明Hoechst与四链体的结合亲和力为10(6)数量级。各向异性测量和由时间分辨衰变实验获得的较高寿命表明，四链束缚的Hoechst在比缓冲介质更弱的极化性环境中受到旋转限制。从表面等离子体共振研究，我们得到了1.23+/-0.04x10(5)M~(-1)S(-1)和0.686+/-0.009 S(~(-1))的动力学缔合(k(A))和解离(k(D))。由于Hoechst与双链DNA的A-T富集区相结合，因此我们在这里提出了Hoechst与四链的AAGGT环相互作用的可能性。(C)2003 Elsevier Inc.保留所有权利。
In vitro binding of Hoechst 33258 to the promoter region of human c-myc, d(GG GGAGGG TGG GGA GGG TGG GGA AGG TGG GG) which forms G-quadruplex, both in vitro and in vivo in the presence of metal ions, was investigated by equilibrium absorption, fluorescence, and kinetic surface plasmon resonance methods. Hypochromic effect in UV absorption spectra and blue shift in fluorescence emission maxima of Hoechst in the presence of quadruplex revealed that Hoechst binds to the quadruplex, Analysis of UV and fluorescence titration data revealed that Hoechst binds to quadruplex with binding affinity of the order of 10(6). Anisotropy measurements and higher lifetime obtained from time-resolved decay experiments revealed that quadruplex-bound Hoechst is rotationally restricted in a less polar environment than the bulk buffer medium. From surface plasmon resonance studies, we obtained kinetic association (k(a)) and dissociation (k(d)) of 1.23 +/- 0.04 x 10(5) M-1 s(-1) and 0.686 +/- 0.009 s(-1), respectively. As Hoechst is known to bind A-T-rich region of duplex DNA, here we propose the likelihood of Hoechst interacting with the AAGGT loop of the quadruplex. (C) 2003 Elsevier Inc. All rights reserved.