MULTIFREQUENCY ESR SPECTRA OF SPIN-LABELED PROTEIN FROM MD SIMULATIONS

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 多频电子自旋共振（ESR）光谱提供了丰富的结构和动力学信息的自旋标签的本地环境，并间接有关的蛋白质，它们所连接的。然而，将观察到的光谱的特征与潜在的分子运动和相互作用明确地联系起来是具有挑战性的。 为了对ESR谱进行严格的解释，我们对完全溶剂化的T4溶菌酶进行了广泛的分子动力学（MD）模拟，在位置72和131处标记了自旋标记。这两个位点已经成为许多实验研究的对象，并且已经确立了它们自身作为α-螺旋表面上的原型溶剂暴露位点。 为了延长由模拟提供的时间窗口，随机马尔可夫模型，反映自旋标签侧链的动态方面，他们的旋转异构体的状态，从轨迹构建。 在没有可调参数的情况下，在三个不同磁场强度下计算的位置72和131的多频ESR谱与实验定量一致。 在动力学过程中，自旋标签具有相当大数量的构象，并显示出与蛋白质残基形成相互作用的显著倾向，而不是与螺旋沿着最近的相邻残基形成相互作用。从MD模拟中出现的自旋标记的详细图片统一了不同的光谱和晶体学数据，并提供了前所未有的深入了解它们的分子起源。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Multifrequency electron spin resonance (ESR) spectra provides a wealth of structural and dynamics information about the local environment of the spin label, and indirectly about the proteins to which they are attached. Unambiguously relating the features of the observed spectra to the underlying molecular motions and interactions is, however, challenging. To progress toward a rigorous interpretation of ESR spectra, we performed extensive molecular dynamics (MD) simulations of fully solvated T4 Lysozyme, labeled with spin labeled at positions 72 and 131. These two sites have been the object of numerous experimental studies, and have established themselves as prototypical solvent-exposed sites on the surface of alpha-helices. To extend the time window afforded by the simulations, stochastic Markov models, reflecting the dynamics of spin label side chains in terms of their rotameric states, are constructed from the trajectories. Without adjustable parameters, the multifrequency ESR spectra calculated at three different magnetic field strengths for positions 72 and 131 were found to be in quantitative agreement with experiment. During dynamics the spin label has access to a fairly large number of conformations and displays a significant propensity to form interactions with protein residues other than the nearest neighbors along the helix. The detailed picture of the spin label emerging from the MD simulations unifies the diverse spectroscopic and crystallographic data and provides unprecedented insight into their molecular origins.