Double-stranded DNA binding characteristics and subcellular distribution of a minor groove binding diphenyl ether bisbenzimidazole.
Double-stranded DNA binding characteristics and subcellular distribution of a minor groove binding diphenyl ether bisbenzimidazole.
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双链 DNA 结合特性和小沟结合二苯醚双苯并咪唑的亚细胞分布。
DOI：
10.1021/bi0103415
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发表时间：
2001
期刊：
影响因子：
2.9
通讯作者：
Bruice,TC
中科院分区：
文献类型：
--
作者：
Satz,AL;White,CM;Beerman,TA;Bruice,TC
Hoechst 33377（H1）与20种不同的寡聚双链体的相互作用已通过荧光分光光度滴定和/或热变性实验进行了研究。H1显示与至少四个连续A/T碱基对的dsDNA结合位点形成2：1复合物。H1也被证明具有罕见的能力，有意义地区分不同的A·T丰富的序列。例如，发现寡聚双链体5 '-GCAATTGC-3'（15）与H1复合的组合平衡常数比双链体5 '-GCTTAAGC-3'（16）大110倍。据信，16中的5 '-TpA-3'二核苷酸步骤破坏了15的刚性“A道”构象，并阻碍了小沟与能够识别较长dsDNA序列的试剂的结合。分子模型阐明了（H1）2-dsDNA小沟复合物的结构。这两个H1分子结合到一个A/T丰富的序列6 bp的稍微交错，并排，和反平行的安排。有证据表明，哌嗪环的H1并排复合物能够休息在G/C碱基对的小沟。使用NIH 3 T3细胞的荧光显微镜研究表明，H1能够穿过细胞质膜并选择性地定位于核DNA。H1还表现出在微摩尔浓度下抑制NIH 3 T3细胞中c-fos基因的内源性转录的能力。采用相同细胞类型的细胞毒性研究显示，H1的LD 50为3.5 μM。
The interactions of Hoechst 33377 (H1) with 20 different oligomeric duplexes have been investigated via spectrofluorometric titrations and/or thermal denaturation experiments. H1 is shown to form 2:1 complexes with dsDNA binding sites of at least four contiguous A/T base pairs. H1 is also shown to possess the rare ability to meaningfully distinguish between different A·T rich sequences. For example, the combined equilibrium constants for complexation of the oligomeric duplex 5‘-GCAATTGC-3‘ (15) by H1 are found to be 110-fold greater than for the duplex 5‘-GCTTAAGC-3‘ (16). It is believed that the 5‘-TpA-3‘ dinucleotide step in16disrupts the rigid “A-tract” conformation of15and discourages minor groove binding by agents capable of recognizing longer dsDNA sequences. Molecular models are presented which elucidate the structure of the (H1)2−dsDNA minor groove complex. The two H1 molecules bind to an A/T rich sequence of 6 bp in a slightly staggered, side-by-side, and antiparallel arrangement. Evidence suggests that the piperazine rings of the H1 side-by-side complex are capable of resting in the minor groove of G/C base pairs. Fluorescence microscopy studies using NIH3T3 cells indicate that H1 is capable of traversing the cytoplasmic membrane and selectively localizing to nuclear DNA. H1 also demonstrated the ability to inhibit endogenous transcription of the c-fos gene in NIH3T3 cells at micromolar concentrations. Cytotoxicity studies employing the same cell type show H1 to possess an LD50of 3.5 μM.