Effects of γ-turn and β-tail amino acids on sequence-specific recognition of DNA by hairpin polyamides
Effects of γ-turn and β-tail amino acids on sequence-specific recognition of DNA by hairpin polyamides
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DOI：
10.1021/ja9830905
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发表时间：
1999-02-17
影响因子：
15
通讯作者：
Dervan, PB
中科院分区：
文献类型：
--
作者：
Swalley, SE;Baird, EE;Dervan, PB
含有吡咯（Py）和咪唑（Im）氨基酸的三环聚酰胺通过转角特异性γ-氨基丁酸接头（γ-转角）共价偶联，形成六环发夹，识别DNA小沟中预定的5个碱基对（bp）序列。为了确定γ-转角和C-末端β-丙氨酸（β-尾）氨基酸的序列特异性，通过在含有8个可能的5 ′-ATGGCNA-3 ′和5 ′-ANGGCTA-3 ′位点（N = A、T、G或C; 5-bp发夹位点为斜体）的DNA限制性片段上进行足迹法和亲和切割，分析发夹聚酰胺ImPy-gamma-ImPyPy-beta-Dp的DNA结合性质。定量足迹滴定表明，γ-转角和β-尾氨基酸在这些位置上相对于G. C碱基对对对A. T/T. A具有> 200-400倍的偏好。通过在含有8个可能的5 ′-ATGGCTN-3 ′和5 ′-NTGGCTA-3 ′位点的DNA限制性片段上的足迹实验来分析邻近5-bp发夹结合位点的碱基对的影响，定量足迹滴定证明，相对于在这些位置处的G. C碱基对，转角和尾部氨基酸对A. T/T. A具有降低的特异性（3-20倍偏好）。这些结果表明，转角和尾部氨基酸不简单地作为中性接头残基，但事实上，是序列特异性识别元件与可预测的DNA结合特异性。
Three-ring polyamides containing pyrrole (Py) and imidazole (Im) amino acids covalently coupled by a turn-specific gamma-aminobutyric acid linker (gamma-turn) form six-ring hairpins that recognize predetermined 5-base pair (bp) sequences in the minor groove of DNA. To determine the sequence specificity of the gamma-turn and C-terminal beta-alanine (beta-tail) amino acids, the DNA-binding properties of the hairpin polyamide ImImPy-gamma-ImPyPy-beta-Dp were analyzed by footprinting and affinity cleavage on DNA-restriction fragments containing the eight possible 5'-ATGGCNA-3' and 5'-ANGGCTA-3' sites (N = A, T,G or C; 5-bp hairpin site is in italics). Quantitative footprint titrations demonstrate that both the gamma-turn and beta-tail amino acids have a > 200-400-fold preference for A.T/T.A relative to G.C base pairs at these positions, Effects of the base pairs adjacent to the 5-bp hairpin-binding site were analyzed by footprinting experiments on a DNA-restriction fragment containing the eight possible 5'-ATGGCTN-3' and 5'-NTGGCTA-3' sites, Quantitative footprint titrations demonstrate that the turn and tail amino acids have reduced specificity (3-20-fold preference) for A.T/T.A relative to G.C base pairs at these positions. These results indicate that the turn and tail amino acids do not simply act as neutral linker residues but, in fact, are sequence-specific recognition elements with predictable DNA-binding specificity.