Sarzynska Joanna, Nilsson Lennart, Kulinski Tadeusz
Institute of Bioorganic Chemistry, Polish Academy of Sciences, Poznań, Poland.
Biophys J. 2003 Dec;85(6):3445-59. doi: 10.1016/S0006-3495(03)74766-3.
Contributions of individual interactions in the GGCGCAAGCC hairpin containing a GCAA tetraloop were studied by computer simulations using base substitutions. The G in the first tetraloop position was replaced by inosine (I) or adenosine (A), and the G in the C-G basepair closing the tetraloop was replaced by I. These substitutions eliminate particular hydrogen bonds proposed in the nuclear magnetic resonance model of the GCAA tetraloop. Molecular dynamics simulations of the GCAA tetraloop in aqueous solvent displayed a well-defined hydrogen pattern between the first and last loop nucleotides (G and A) stabilized by a bridging water molecule. Substitution of G-->I in the basepair closing the tetraloop did not significantly influence the loop structure and dynamics. The ICAA loop maintained the overall structure, but displayed variation in the hydrogen-bond network within the tetraloop itself. Molecular dynamics simulations of the ACAA loop led to conformational heterogeneity of the resulting structures. Changes of hairpin formation free energy associated with substitutions of individual bases were calculated by the free energy perturbation method. The calculated decrease of the hairpin stability upon G-->I substitution in the C-G basepair closing the tetraloop was in good agreement with experimental thermodynamic data. Our theoretical estimates for G-->I and G-->A mutations located in the tetraloop suggest larger loop destabilization than corresponding experimental results. The extent of conformational sampling of the structures resulting from base substitutions and its impact on the calculated free energy was discussed.
通过使用碱基替换的计算机模拟，研究了含有GCAA四环的GGCGCAAGCC发夹中个体相互作用的贡献。四环第一个位置的G被次黄嘌呤（I）或腺嘌呤（A）取代，闭合四环的C-G碱基对中的G被I取代。这些替换消除了GCAA四环核磁共振模型中提出的特定氢键。在水性溶剂中对GCAA四环进行分子动力学模拟，显示第一个和最后一个环核苷酸（G和A）之间通过一个桥连水分子稳定存在明确的氢键模式。在闭合四环的碱基对中用I取代G，对环结构和动力学没有显著影响。ICAA环保持了整体结构，但在四环自身的氢键网络中表现出变化。对ACAA环的分子动力学模拟导致所得结构的构象异质性。通过自由能微扰法计算了与单个碱基替换相关的发夹形成自由能的变化。计算得出，在闭合四环的C-G碱基对中用I取代G时，发夹稳定性降低，这与实验热力学数据高度吻合。我们对位于四环中的G→I和G→A突变的理论估计表明，环的不稳定程度比相应的实验结果更大。讨论了碱基替换产生的结构的构象采样程度及其对计算自由能的影响。
