Pilch D S, Kirolos M A, Breslauer K J
Department of Chemistry, Rutgers, State University of New Jersey, New Brunswick 08903, USA.
Biochemistry. 1995 Dec 12;34(49):16107-24. doi: 10.1021/bi00049a026.
Berenil is an antitrypanosomal agent that binds to nucleic acid duplexes. Recently, we reported that this drug can bind to both DNA and RNA duplexes, while exhibiting properties characteristic of both intercalation and groove binding [Pilch, D. S., Kirolos, M. A., Liu, X., Plum, G. E., & Breslauer, K. J. (1995) Biochemistry 34, 9962-9976]. In this work, we use spectroscopic, calorimetric, and hydrodynamic techniques to demonstrate that berenil also can bind to DNA and RNA triplexes. Our results reveal the following significant features: (i) Berenil binds to the poly(dA).2poly(dT) DNA triplex and to the poly(rA).2poly(rU) RNA triplex without displacing the major groove-bound third strands. (ii) Both berenil-bound triplexes melt via two distinct transitions: initial conversion of the triplex to the duplex state, with the berenil remaining bound, followed by denaturation of the duplex to its component single strands. (iii) The magnitude and even the direction of the impact of berenil binding on the thermal stability of the DNA triplex depends on both the Na+ concentration and the drug binding density (the [base triplet]/[total berenil] ratio). Specifically, at Na+ concentrations < or = 0.08 M, the DNA triplex to duplex transition is thermally stabilized, while at Na+ concentrations > or = 0.125 M it is thermally destabilized. Between these two salt concentrations, berenil binding either enhances or diminishes the thermal stability of the DNA triplex in a manner that depends on the [base triplet]/[total berenil] ratio. (iv) The effect of berenil binding on the thermal stability of the RNA triplex to duplex equilibrium also depends on the [base triplet]/[total berenil] ratio, having a weakly destabilizing effect on this equilibrium at [base triplet]/[total berenil] ratios > 5, while thermally stabilizing this equilibrium at [base triplet]/[total berenil] ratios < 5. (v) The apparent "site sizes" associated with berenil binding to the triplexes range from approximately 1 to 12 base triplets per bound berenil and depend, in part, on the host triplex. One of the site sizes common to both triplexes is consistent with berenil binding to the minor groove. (vi) Berenil exhibits a higher apparent binding affinity for the DNA triplex relative to the RNA triplex. (vii) Viscometric data reveal nonintercalative binding properties when berenil complexes with both triplexes, consistent with a minor groove binding mode. (viii) Berenil binding to either the DNA or the RNA triplex is enthalpically more favorable than berenil binding to the corresponding duplex. (ix) Berenil binding to both triplexes decreases the cooperativity of the triplex to duplex melting event.(ABSTRACT TRUNCATED AT 400 WORDS)
贝尼尔是一种与核酸双链体结合的抗锥虫剂。最近，我们报道了这种药物既能与DNA双链体结合，也能与RNA双链体结合，同时展现出嵌入和沟槽结合的特性[皮尔希，D. S.，基罗洛斯，M. A.，刘，X.，普拉姆，G. E.，& 布雷斯劳尔，K. J.（1995年）《生物化学》34卷，9962 - 9976页]。在这项研究中，我们运用光谱、量热和流体动力学技术来证明贝尼尔也能与DNA和RNA三链体结合。我们的研究结果揭示了以下显著特征：（i）贝尼尔能与聚（dA）·2聚（dT）DNA三链体以及聚（rA）·2聚（rU）RNA三链体结合，且不会取代与大沟结合的第三条链。（ii）两种贝尼尔结合的三链体都通过两个不同的转变过程解链：首先三链体转变为双链体状态，贝尼尔仍保持结合状态，随后双链体解链为其组成的单链。（iii）贝尼尔结合对DNA三链体热稳定性的影响程度甚至方向，取决于Na⁺浓度和药物结合密度（[碱基三联体]/[总贝尼尔]比值）。具体而言，当Na⁺浓度≤0.08 M时，DNA三链体到双链体的转变在热稳定性上得到增强，而当Na⁺浓度≥0.125 M时则热稳定性降低。在这两个盐浓度之间，贝尼尔结合对DNA三链体热稳定性的影响增强或减弱，取决于[碱基三联体]/[总贝尼尔]比值。（iv）贝尼尔结合对RNA三链体到双链体平衡热稳定性的影响也取决于[碱基三联体]/[总贝尼尔]比值，当[碱基三联体]/[总贝尼尔]比值＞5时，对该平衡有微弱的热稳定性降低作用，而当[碱基三联体]/[总贝尼尔]比值＜5时则热稳定性增强。（v）与贝尼尔结合到三链体相关的表观“结合位点大小”范围为每个结合的贝尼尔约1至12个碱基三联体，并且部分取决于宿主三链体。两种三链体共有的一种结合位点大小与贝尼尔结合到小沟一致。（vi）相对于RNA三链体，贝尼尔对DNA三链体表现出更高的表观结合亲和力。（vii）粘度测定数据显示，当贝尼尔与两种三链体形成复合物时具有非嵌入结合特性，这与小沟结合模式一致。（viii）贝尼尔与DNA或RNA三链体的结合在焓变上比与相应双链体的结合更有利。（ix）贝尼尔与两种三链体结合都会降低三链体到双链体解链过程的协同性。（摘要截取自400字）
