Double-Stranded DNA Binding Characteristics and Subcellular Distribution of a Minor Groove Binding Diphenyl Ether Bisbenzimidazole

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• 作者：Alexander L. Satz ; Christine M. White ; Terry A. Beerman ; and Thomas C. Bruice
• 刊名：Biochemistry
• 出版年：2001
• 出版时间：May 29, 2001
• 年：2001
• 卷：40
• 期：21
• 页码：6465 - 6474
• 全文大小：198K
• 年卷期：v.40,no.21(May 29, 2001)
• ISSN：1520-4995

文摘

The interactions of Hoechst 33377 (H1) with 20 different oligomeric duplexes have beeninvestigated via spectrofluorometric titrations and/or thermal denaturation experiments. H1 is shown toform 2:1 complexes with dsDNA binding sites of at least four contiguous A/T base pairs. H1 is alsoshown to possess the rare ability to meaningfully distinguish between different A·T rich sequences. Forexample, 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 believedthat the 5'-TpA-3' dinucleotide step in 16 disrupts the rigid "A-tract" conformation of 15 and discouragesminor groove binding by agents capable of recognizing longer dsDNA sequences. Molecular models arepresented which elucidate the structure of the (H1)₂-dsDNA minor groove complex. The two H1 moleculesbind 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 theminor groove of G/C base pairs. Fluorescence microscopy studies using NIH3T3 cells indicate that H1is capable of traversing the cytoplasmic membrane and selectively localizing to nuclear DNA. H1 alsodemonstrated 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 LD₅₀ of3.5 M.