Methods of NMR structure refinement: molecular dynamics simulations improve the agreement with measured NMR data of a C-terminal peptide of GCN4-p1
Methods of NMR structure refinement: molecular dynamics simulations improve the agreement with measured NMR data of a C-terminal peptide of GCN4-p1
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DOI：
10.1007/s10858-010-9425-9
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发表时间：
2010-07-01
影响因子：
2.7
通讯作者：
van Gunsteren, Wilfred F.
中科院分区：
文献类型：
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作者：
Dolenc, Jozica;Missimer, John H.;van Gunsteren, Wilfred F.
c端触发序列在GCN4-p1的线圈-线圈形成中是必不可少的；因此，它的构象性质对于在原子水平上理解这一过程是很重要的。几年前提出了包含GCN4-p1触发序列的肽GCN4p16-31的溶液核磁共振模型结构。采用基于172个核Overhauser效应（NOE）距离约束、14个氢键约束和11个中心点扭转角约束的标准单结构改进方案推导得到的20个核磁共振模型结构具有规则的α -螺旋结构。然而，该集合稍微违反了一些测量到的NOE界限，并且不能重现所有15个测量到的(3)J（H-N-H-C α）耦合常数，这表明溶液中可能存在GCN4p16-31的不同构象。为了解决与所有NOE上距离边界和j耦合常数兼容的结构，我们在两种力场下使用无约束和约束分子动力学（MD）模拟执行了几种结构优化方案。我们发现，只有同时施加时间平均NOE距离和(3)j -耦合常数约束的构型系综才能再现所有实验数据。此外，对模拟体系的分析表明，可用的187个核磁共振数据集所允许的溶液中GCN4p16-31的构象变异性大于核磁共振模型结构集所表示的构象变异性。GCN4p16-31在溶液中的构象不仅侧链的取向不同，而且主链的取向也不同。核磁共振模型结构与实测核磁共振数据之间的不一致是由于忽略了平均效应以及包含了氢键和扭转角约束，而这些约束在主要的核磁共振实测数据中几乎没有基础。
The C-terminal trigger sequence is essential in the coiled-coil formation of GCN4-p1; its conformational properties are thus of importance for understanding this process at the atomic level. A solution NMR model structure of a peptide, GCN4p16-31, encompassing the GCN4-p1 trigger sequence was proposed a few years ago. Derived using a standard single-structure refinement protocol based on 172 nuclear Overhauser effect (NOE) distance restraints, 14 hydrogen-bond and 11 I center dot torsional-angle restraints, the resulting set of 20 NMR model structures exhibits regular alpha-helical structure. However, the set slightly violates some measured NOE bounds and does not reproduce all 15 measured (3)J(H-N-H-C alpha)-coupling constants, indicating that different conformers of GCN4p16-31 might be present in solution. With the aim to resolve structures compatible with all NOE upper distance bounds and (3)J-coupling constants, we executed several structure refinement protocols employing unrestrained and restrained molecular dynamics (MD) simulations with two force fields. We find that only configurational ensembles obtained by applying simultaneously time-averaged NOE distance and (3)J-coupling constant restraining with either force field reproduce all the experimental data. Additionally, analyses of the simulated ensembles show that the conformational variability of GCN4p16-31 in solution admitted by the available set of 187 measured NMR data is larger than represented by the set of the NMR model structures. The conformations of GCN4p16-31 in solution differ in the orientation not only of the side-chains but also of the backbone. The inconsistencies between the NMR model structures and the measured NMR data are due to the neglect of averaging effects and the inclusion of hydrogen-bond and torsional-angle restraints that have little basis in the primary, i.e. measured NMR data.