Quantitative Mapping of Protein Structure by Hydroxyl Radical Footprinting-Mediated Structural Mass Spectrometry: A Protection Factor Analysis
Quantitative Mapping of Protein Structure by Hydroxyl Radical Footprinting-Mediated Structural Mass Spectrometry: A Protection Factor Analysis
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DOI：
10.1016/j.bpj.2014.11.013
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发表时间：
2015-01-06
影响因子：
3.4
通讯作者：
Yang, Sichun
中科院分区：
文献类型：
--
作者：
Huang, Wei;Ravikumar, Krishnakumar M.;Yang, Sichun
羟基自由基足迹（HRF）的测量提供了丰富的信息，溶剂的氨基酸侧链的蛋白质的可及性。传统的HRF数据分析侧重于比较特定位点的修饰/足迹率的差异，以推断两种蛋白质状态的结构变化，例如，在自由态和配体结合态之间。然而，速率信息本身并不完全用于在绝对尺度上比较蛋白质内不同蛋白质位点的目的。为了提供这样的跨站点比较，我们提出了一种新的，据我们所知，数据分析算法，考虑到已知的固有反应性的氨基酸侧链的保护因子（PF）的测量足迹速率常数转换。要检查PF可用于结构解释的程度，PF分析适用于三个模型系统，其中文献中报道了辐解足迹数据。通过可视化的结构与PF值着色的个别肽，一个合理的看法，各种蛋白质位点的结构特征，关于其溶剂的可及性被揭示，其中高PF区域被掩埋和低PF区域更多地暴露于溶剂。此外，一个详细的分析相关的溶剂的可及性和局部结构接触凝溶胶蛋白显示PF值和各种结构措施之间的统计学显着的协议，表明PF来自这个PF分析很容易解释基本的HRF率测量。我们还对替代的、基于化学的HRF数据测试了这种PF分析，显示与单独检查HRF速率数据相比，模型蛋白Barstar的结构特性的相关性得到了改善。总的来说，这种PF分析不仅允许一种新的，据我们所知，通过使用足迹数据映射蛋白质结构的方法，而且还提升了HRF测量的使用，从定性的，跨状态比较到定量的，跨位点评估蛋白质结构的背景下，个人的构象状态的兴趣。
Measurements from hydroxyl radical footprinting (HRF) provide rich information about the solvent accessibility of amino acid side chains of a protein. Traditional HRF data analyses focus on comparing the difference in the modification/footprinting rate of a specific site to infer structural changes across two protein states, e.g., between a free and ligand-bound state. However, the rate information itself is not fully used for the purpose of comparing different protein sites within a protein on an absolute scale. To provide such a cross-site comparison, we present a new, to our knowledge, data analysis algorithm to convert the measured footprinting rate constant to a protection factor (PF) by taking into account the known intrinsic reactivity of amino acid side chain. To examine the extent to which PFs can be used for structural interpretation, this PF analysis is applied to three model systems where radiolytic footprinting data are reported in the literature. By visualizing structures colored with the PF values for individual peptides, a rational view of the structural features of various protein sites regarding their solvent accessibility is revealed, where high-PF regions are buried and low-PF regions are more exposed to the solvent. Furthermore, a detailed analysis correlating solvent accessibility and local structural contacts for gelsolin shows a statistically significant agreement between PF values and various structure measures, demonstrating that the PFs derived from this PF analysis readily explain fundamental HRF rate measurements. We also tested this PF analysis on alternative, chemical-based HRF data, showing improved correlations of structural properties of a model protein barstar compared to examining HRF rate data alone. Together, this PF analysis not only permits a novel, to our knowledge, approach of mapping protein structures by using footprinting data, but also elevates the use of HRF measurements from a qualitative, cross-state comparison to a quantitative, cross-site assessment of protein structures in the context of individual conformational states of interest.