Patel D J, Shapiro L
Biochimie. 1985 Jul-Aug;67(7-8):887-915. doi: 10.1016/s0300-9084(85)80181-4.
We have investigated intermolecular interactions and conformational features of the netropsin complexes with d(G1-G2-A3-A4-T5-T6-C7-C8) duplex (AATT 8-mer) and the d(G1-G2-T3-A4-T5-A6-C7-C8) duplex (TATA 8-mer) by one and two-dimensional NMR studies in solution. We have assigned the amide, pyrrole and methylene protons of netropsin and the base and sugar H1' protons of the nucleic acid from an analysis of the nuclear Overhauser effect (NOESY) and correlated (COSY) spectra of the complex at 25 degrees C. The directionality of the observed distance-dependent NOEs demonstrates that the 8-mer helices remain right-handed and that the arrangement of concave and convex face protons of netropsin are retained in the complexes. The observed changes in NOE patterns and chemical shift changes on complex formation suggest small conformational changes in the nucleic acid at the AATT and TATA antibiotic binding sites and possibly the flanking G.C base pairs. We observe intermolecular NOEs between all three amide and both pyrrole protons on the concave face of the antibiotic and the minor groove adenosine H2 proton of the two central A4.T5 base pairs of the AATT 8-mer and TATA 8-mer duplexes. The concave face pyrrole protons of the antibiotic also exhibit NOEs to the sugar H1' protons of residues 5 and 6 in the AATT and TATA 8-mer complexes. We also detect intermolecular NOEs between the guanidino and propioamidino methylene protons at either end of netropsin and the adenosine H2 proton of the two flanking A3.T6 base pairs in the AATT 8-mer and T3.A6 base pairs in the TATA 8-mer duplexes. These studies establish a set of nine contacts between the concave face of the antibiotic and the minor groove AATT segment and TATA segment of the 8-mer duplexes in solution. The observed magnitude of the NOEs require that there be no intervening water molecules sandwiched between the concave face of the antibiotic and the minor groove of the DNA so that release of the minor groove spine of hydration is a prerequisite for netropsin complex formation. The observed differences in the netropsin amide proton chemical shifts in the AATT 8-mer and TATA 8-mer complexes suggest differences in the strength and/or type of intermolecular hydrogen bonds at the AATT and TATA binding sites.(ABSTRACT TRUNCATED AT 400 WORDS)
我们通过溶液中的一维和二维核磁共振研究，调查了纺锤菌素与d(G1-G2-A3-A4-T5-T6-C7-C8)双链体（AATT 8聚体）和d(G1-G2-T3-A4-T5-A6-C7-C8)双链体（TATA 8聚体）复合物的分子间相互作用和构象特征。通过对25摄氏度下复合物的核Overhauser效应（NOESY）和相关（COSY）光谱的分析，我们归属了纺锤菌素的酰胺、吡咯和亚甲基质子以及核酸的碱基和糖H1'质子。观察到的距离依赖性核Overhauser效应的方向性表明，8聚体螺旋保持右手螺旋，并且纺锤菌素凹面和凸面质子的排列在复合物中得以保留。复合物形成时观察到的核Overhauser效应模式变化和化学位移变化表明，在AATT和TATA抗生素结合位点以及可能的侧翼G.C碱基对处，核酸存在小的构象变化。我们观察到抗生素凹面上的所有三个酰胺和两个吡咯质子与AATT 8聚体和TATA 8聚体双链体两个中央A_{4}·T_{5}碱基对的小沟腺苷H2质子之间存在分子间核Overhauser效应。抗生素凹面的吡咯质子在AATT和TATA 8聚体复合物中也与残基5和6的糖H1'质子表现出核Overhauser效应。我们还检测到纺锤菌素两端的胍基和丙脒基亚甲基质子与AATT 8聚体中两个侧翼A_{3}·T_{6}碱基对以及TATA 8聚体中T_{3}·A_{6}碱基对的腺苷H2质子之间存在分子间核Overhauser效应。这些研究确定了溶液中抗生素凹面与8聚体双链体的小沟AATT片段和TATA片段之间的一组九个接触点。观察到的核Overhauser效应大小要求在抗生素凹面和DNA小沟之间不存在夹在中间的水分子，因此释放小沟水合脊柱是纺锤菌素复合物形成的先决条件。在AATT 8聚体和TATA 8聚体复合物中观察到的纺锤菌素酰胺质子化学位移差异表明，AATT和TATA结合位点处分子间氢键的强度和/或类型存在差异。（摘要截短于400字）
