ELECTRON CAPTURE DISSOCIATION FOR DETERMINING H/D EXCHANGE RATES IN PEPTIDES

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 在分析和解释用于蛋白质结构分析的多维核磁共振数据时，限制因素之一是共振峰的分配。共鸣分配的标准策略既费力又昂贵。最近在使用稀疏标记策略进行多维核磁共振方面取得了进展，其中只标记和分析了有限数量的残基。为了利用更简单的 稀疏标记核磁共振产生的光谱，用于共振指认的替代技术正在进行中。 一种被研究的技术是使用酰胺氢交换动力学来指定共振。酰胺交换动力学可以很容易地通过核磁共振来跟踪；然而，共振峰的分配不能仅仅由核磁共振来确定。这个子项目试图利用快速胃酶消化，然后非遍历电子捕获 解离(ECD)质谱仪，以测量不同氨基酸残基的酰胺交换率。一旦可以测量每个标记残基的酰胺交换速率，就可以根据在核磁共振谱中测量的酰胺交换速率来分配稀疏标记核磁共振中的共振。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. One of the limiting factors in analysis and interpretation of multidimensional NMR data for protein structural analysis is the assignment of resonances. Standard strategies for resonance assignment are laborious and expensive. Recent progress has been made in the use of sparse labeling strategies for multidimensional NMR, where only a limited number of residues are labeled and analyzed. In order to take advantage of the simpler spectra resulting from sparse labeling NMR, alternative technology for resonance assignment are being pursued. One technology studied was the use of amide deuterium exchange kinetics to assign resonances. Amide exchange kinetics can be followed readily by NMR; however, the assignment of the resonances cannot be determined solely by NMR. This subproject attempts to utilize rapid pepsin digestion followed by non-ergodic electron capture dissociation (ECD) mass spectrometry in order to measure rates of amide exchange for different amino acid residues. Once the rate of amide exchange for each of the labeled residues can be measured, resonances in sparse labeling NMR can be assigned based on the rate of amide exchange measured in the NMR spectrum.