Modifying the N-terminus of polyamides: PyImPyIm has improved sequence specificity over f-ImPyIm
Modifying the N-terminus of polyamides: PyImPyIm has improved sequence specificity over f-ImPyIm
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DOI：
10.1016/j.bmc.2008.03.008
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发表时间：
2008-05-01
影响因子：
3.5
通讯作者：
Lee, Moses
中科院分区：
文献类型：
--
作者：
Brown, Toni;Mackay, Hilary;Lee, Moses
合成了7个N-末端修饰的小沟槽结合聚酰胺衍生物(f-ImPyIm，1)，并对其进行了生化和生物物理化学评价。合成这些化合物的目的是获得比f-ImPyIm(1)更高水平的序列选择性，f-ImPyIm(1)是以前发表的一种强小沟槽粘结剂。两个N-端含有呋喃或苯并呋喃部分的化合物在同源ACGCGT位点上的足迹为0.5µM(由DNase I足迹确定)；然而，这些化合物的特异性并未得到提高。相比之下，PyImPyIm(4)产生了0.5µM的足迹，但使用相同的技术显示了更好的特异性。当用ACGCGT和非同源AAATTT序列进行热熔融实验和圆二色谱评价时，所有化合物都被证明结合在DNA的小槽中，并且稳定同源序列比非同源序列好得多(非氨基化合物不像预期的那样与这两个序列结合)。PyImPyIm(4)很有趣，因为该化合物的Delta T(M)只有4摄氏度，但足迹非常有选择性。没有观察到该化合物与第三个DNA(非同源DNA，ACCGGT)结合。ITC对化合物4的研究表明，化合物4与ACGCGT发生放热结合，在将化合物滴定到另外两个DNA序列时没有观察到热变化。由疏水相互作用和SASA计算得到的PIPI/ACGCGT络合物的热容(Delta C(P))与ITC的实验值(-146cal mol(-1)K(-1))相当。SPR结果提供了可靠的证据。确认了PyImPyIm(4)的序列特异性，同源序列的K(Eq)值为7.1×10(6)M(-1)，未观察到与AAATTT和ACCGGT的结合。分子动力学模拟证实，PyImPyIm(4)以二聚体的形式以重叠构象结合，并紧密结合在ACGCGT寡核苷酸的小凹槽中。PyImPyIm(4)是一个特别有趣的分子，因为虽然结合亲和力略有降低，但相对于f-ImPyIm(1)的特异性显著提高。(C)2008爱思唯尔有限公司。保留所有权利。
Seven N-terminus modified derivatives of a previously published minor-groove binding polyamide (f-ImPyIm, 1) were synthesized and the biochemical and biophysical chemistry evaluated. These compounds were synthesized with the aim of attaining a higher level of sequence selectivity over f-ImPyIm (1), a previously published strong minor-groove binder. Two compounds possessing a furan or a benzofuran moiety at the N-terminus showed a footprint of 0.5 mu M at the cognate ACGCGT site (determined by DNase I footprinting); however, the specificity of these compounds was not improved. In contrast, PyImPyIm (4) produced a footprint of 0.5 mu M but showed a superior specificity using the same technique. When evaluated by thermal melting experiments and circular dichroism using ACGCGT and the non-cognate AAATTT sequence, all compounds were shown to bind in the minor-groove of DNA and stabilize the cognate sequence much better than the non-cognate (except for the non-amido-compound that did not bind either sequence, as expected). PyImPyIm (4) was interesting as the Delta T(m) for this compound was only 4 degrees C but the footprint was very selective. No binding was observed for this compound with a third DNA (non-cognate, ACCGGT). ITC studies on compound 4 showed exothermic binding with ACGCGT and no heat change was observed for titrating the compound to the other two DNA sequences. The heat capacity (Delta C(p)) of the PIPI/ACGCGT complex calculated from the hydrophobic interactions and SASA calculations was comparable to the experimental value obtained from ITC (-146 cal mol(-1) K(-1)). SPR results provided con. firmation of the sequence specificity of PyImPyIm (4), with a K(eq) value determined to be 7.1 x 10(6) M(-1) for the cognate sequence and no observable binding to AAATTT and ACCGGT. Molecular dynamic simulations affirmed that PyImPyIm (4) binds as a dimer in an overlapped conformation, and it fits snugly in the minor-groove of the ACGCGT oligonucleotide. PyImPyIm (4) is an especially interesting molecule, because although the binding affinity is slightly reduced, the specificity with respect to f-ImPyIm (1) is significantly improved. (C) 2008 Elsevier Ltd. All rights reserved.