Hubei Key Laboratory of Agricultural Bioinformatics, College of Informatics, Huazhong Agricultural University, Wuhan, 430070, People's Republic of China.
J Mol Model. 2021 Jul 15;27(8):227. doi: 10.1007/s00894-021-04840-y.
DNA methylation is important in regulation of gene expression and normal development because it alters the interplay between protein and DNA. Experiments have shown that a single 5-methylcytosine at different CpG sites (mCpG) might have different effects on specific recognition, but the atomistic origin and dynamic details are largely unclear. In this work, we investigated the mechanism of monomethylation at different CpG sites in the cognate motif and the cooperativity of full methylation. By constructing four models of c-Jun/Jun protein binding to the 5[Formula: see text]-XGAGTCA-3[Formula: see text] (X represents C or methylated C) motif, we characterized the dynamics of the contact interface using the all-atom molecular dynamics method. Free energy analysis of MM/GBSA suggests that regardless of whether the C12pG13 site of the bottom strand is methylated, the effects from mC25 of the top strand are dominant and can moderately enhance the binding by [Formula: see text] 31 kcal/mol, whereas mC12 showed a relatively small contribution, in agreement with the experimental data. Remarkably, we found that this spatial-specific influence was induced by different regulatory rules. The influence of the mC25 site is mainly mediated by steric hindrance. The additional methyl group leads to the conformational changes in nearby residues and triggers an obvious structural bending in the protein, which results in the formation of a new T-Asn-C triad that enhances the specific recognition of TCA half-sites. The substitution of the methyl group at the mC12 site of the bottom strand breaks the original H-bonds directly. Such changes in electrostatic interactions also lead to the remote allosteric effects of protein by multifaceted interactions but have negligible contributions to binding. Although these two influence modes are different, they can both fine-tune the local environment, which might produce remote allosteric effects through protein-protein interactions. Further analysis reveals that the discrepancies in these two modes are primarily due to their location. Moreover, when both sites are methylated, the major determinant of binding specificity depends on the context and the location of the methylation site, which is the result of crosstalk and cooperativity.
DNA 甲基化在基因表达调控和正常发育中很重要，因为它改变了蛋白质与 DNA 之间的相互作用。实验表明，在不同的 CpG 位点（mCpG）上的单个 5-甲基胞嘧啶可能对特定识别有不同的影响，但原子起源和动态细节在很大程度上尚不清楚。在这项工作中，我们研究了在同源基序中不同 CpG 位点单甲基化的机制以及完全甲基化的协同作用。通过构建 c-Jun/Jun 蛋白与 5'-XGAGTCA-3'（X 代表 C 或甲基化的 C）基序结合的四个模型，我们使用全原子分子动力学方法对接触界面的动力学进行了表征。MM/GBSA 的自由能分析表明，无论底部链的 C12pG13 位点是否被甲基化，顶部链的 mC25 的影响都是占主导地位的，可以适度增强结合作用[Formula: see text]31 kcal/mol，而 mC12 的贡献相对较小，这与实验数据一致。值得注意的是，我们发现这种空间特异性影响是由不同的调控规则引起的。mC25 位点的影响主要是由空间位阻介导的。额外的甲基基团导致附近残基的构象变化，并在蛋白质中引发明显的结构弯曲，从而形成新的 T-Asn-C 三联体，增强了 TCA 半位点的特异性识别。底部链 mC12 位点的甲基取代直接破坏了原来的氢键。这种静电相互作用的变化也通过多方面的相互作用导致蛋白质的远程变构效应，但对结合的贡献可以忽略不计。尽管这两种影响模式不同，但它们都可以微调局部环境，通过蛋白质-蛋白质相互作用产生远程变构效应。进一步的分析表明，这两种模式的差异主要是由于它们的位置不同。此外，当两个位点都被甲基化时，结合特异性的主要决定因素取决于甲基化位点的上下文和位置，这是串扰和协同作用的结果。
