Department of Chemistry, James Franck Institute and Institute for Biophysical Dynamics, University of Chicago, Chicago, Illinois.
Department of Chemistry, James Franck Institute and Institute for Biophysical Dynamics, University of Chicago, Chicago, Illinois.
Biophys J. 2018 Jun 19;114(12):2820-2832. doi: 10.1016/j.bpj.2018.05.003.
Characterizing ensembles of intrinsically disordered proteins is experimentally challenging because of the ill-conditioned nature of ensemble determination with limited data and the intrinsic fast dynamics of the conformational ensemble. Amide I two-dimensional infrared (2D IR) spectroscopy has picosecond time resolution to freeze structural ensembles as needed for probing disordered-protein ensembles and conformational dynamics. Also, developments in amide I computational spectroscopy now allow a quantitative and direct prediction of amide I spectra based on conformational distributions drawn from molecular dynamics simulations, providing a route to ensemble refinement against experimental spectra. We performed a Bayesian ensemble refinement method on Ala-Ala-Ala against isotope-edited Fourier-transform infrared spectroscopy and 2D IR spectroscopy and tested potential factors affecting the quality of ensemble refinements. We found that isotope-edited 2D IR spectroscopy provides a stringent constraint on Ala-Ala-Ala conformations and returns consistent conformational ensembles with the dominant ppII conformer across varying prior distributions from many molecular dynamics force fields and water models. The dominant factor influencing ensemble refinements is the systematic frequency uncertainty from spectroscopic maps. However, the uncertainty of conformer populations can be significantly reduced by incorporating 2D IR spectra in addition to traditional Fourier-transform infrared spectra. Bayesian ensemble refinement against isotope-edited 2D IR spectroscopy thus provides a route to probe equilibrium-complex protein ensembles and potentially nonequilibrium conformational dynamics.
表征无规卷曲蛋白质的集合体在实验上具有挑战性，因为在有限的数据条件下，集合体的确定具有病态特征，而且构象集合体的固有快速动力学也会影响其确定。酰胺 I 二维红外（2D IR）光谱具有皮秒时间分辨率，可以根据需要冻结结构集合体，从而用于探测无规卷曲蛋白质集合体和构象动力学。此外，酰胺 I 计算光谱学的发展现在允许根据从分子动力学模拟中得出的构象分布，对酰胺 I 光谱进行定量和直接预测，为与实验光谱进行集合体细化提供了一种途径。我们对 Ala-Ala-Ala 进行了贝叶斯集合体细化方法，以对抗同位素编辑傅里叶变换红外（FTIR）光谱和 2D IR 光谱，并测试了影响集合体细化质量的潜在因素。我们发现，同位素编辑的 2D IR 光谱对 Ala-Ala-Ala 构象提供了严格的约束，并返回了一致的构象集合体，其中包括来自许多分子动力学力场和水模型的不同先验分布的主要 ppII 构象。影响集合体细化的主要因素是光谱图的系统频率不确定性。然而，通过结合传统的傅里叶变换红外光谱和 2D IR 光谱，可以显著降低构象群体的不确定性。因此，对同位素编辑的 2D IR 光谱进行贝叶斯集合体细化，为探测平衡复杂蛋白质集合体和潜在的非平衡构象动力学提供了一种途径。
