DOUBLE RESONANCE ECD

双共振ECD

• 批准号: 7955943
• 负责人: PETER B. O'CONNOR
• 金额: $ 0.7万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-06-01 至 2010-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7955943
• 关键词: American; Biology; Birds; Cations; Charge; Complex; Computer Retrieval of Information on Scientific Projects Database; Cyclic Peptides; Cyclosporins; Data; Dissociation; Electrons; Funding; Gramicidin; Grant; Hand; Heating; Hydrogen Bonding; Institution; Ions; Journals; Kinetics; Lasers; Manuscripts; Mass Spectrum Analysis; Measures; Medicine; Modeling; Molecular; Ornithine; Paper; Peptides; Publishing; Research; Research Personnel; Resources; Societies; Source; Techniques; Thermodynamics; United States National Institutes of Health; interest; millisecond; research study

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. The Double Resonance (DR) technique was adapted to electron capture dissociation (ECD) and applied to the analysis of the fragmentation timeframe of peptides. DR-ECD involves continuously ejecting an ion as it is being formed durine ECD. This ion is usually, but not necessarily, the charge reduced molecular radical cation. Since the ejection timeframe for on-resonant ejection in a 7T FTMS of our geometry is rougly 100 microseconds - 1 millisecond, any ions that are formed on a timeframe greater than 1 ms will be ejected. Thus, one can classify ions as "fast" or "slow" in terms of their ECD formation rate. The DR-ECD experiment on linear peptides showed that approximately half of all fragment ions are generated on a >1 millisecond timeframe. These are presumably fragments that are held together by hydrogen bonds - the dissociation of which determines the formation rate. This model does correlate with the DR-ECD data as reduction in the number of residues that can H-bond reduces the number of long-duration fragments ions. However, the cyclic peptide data is more interesting. Cyclosporin, which has no residues with sidechains that can hydrogen bond, generates many secondary fragments, but ALL of them are formed faster than 100 microseconds. On the other hand, gramicidin S, which has several ornithine residues, shows some slow-forming fragments. Furthermore, some of the fragments ions show "survival ratios" > 1, but this is correlated with double electron capture. The manuscript was published in the Journal of the American Society for Mass Spectrometry. Implementation of the infrared laser heating with the DR-ECD experiment allowed us to determine several important kinetic parameters for a set of peptides. We could measure the peptide refolding rate (by varying the delay between IR and ECD), we could measure the H-bond dissociation rate in a similar fashion, and we could measure the rate of H-atom abstraction in the radical cation complex, and it was measured to be ~5-10x higher than the H-bond dissociation rate. Finally, by calibrating the laser heating using a BIRD experiment, we were able to build Van 't Hoff plots from which we can extract thermodynamic parameters. We published one paper (Lin et al. 2008) in the Journal of the American Society for Mass Spectrometry (2008, 19, 870-879) regarding the kinetic data and have a second one in revision regarding the thermodynamic data.

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 将双共振（DR）技术应用于电子捕获解离（ECD），并应用于肽段片段化时间框架的分析。 DR-ECD涉及在ECD期间形成离子时连续喷射离子。 该离子通常但不一定是电荷减少的分子自由基阳离子。 由于在我们的几何形状的7 T FTMS中的共振喷射的喷射时间帧大约是100微秒-1毫秒，因此在大于1毫秒的时间帧上形成的任何离子都将被喷射。 因此，可以根据离子的ECD形成速率将离子分类为“快”或“慢”。 对线性肽的DR-ECD实验显示，所有碎片离子中的大约一半是在>1毫秒的时间范围内产生的。 这些可能是通过氢键结合在一起的碎片-氢键的解离决定了形成速率。 该模型确实与DR-ECD数据相关，因为可以氢键键合的残基数量减少会减少长持续时间碎片离子的数量。 然而，环肽数据更有趣。 环孢菌素没有带有可以氢键结合的侧链的残基，产生许多次级片段，但所有这些片段的形成速度都超过100微秒。 另一方面，短杆菌肽S，其中有几个鸟氨酸残基，显示出一些缓慢形成的片段。 此外，一些碎片离子显示“存活率”> 1，但这与双电子捕获相关。 该手稿发表在《美国质谱学会杂志》上。 红外激光加热与DR-ECD实验的实施使我们能够确定一组肽的几个重要的动力学参数。 我们可以测量肽重折叠速率（通过改变IR和ECD之间的延迟），我们可以以类似的方式测量H-键解离速率，并且我们可以测量自由基阳离子复合物中H-原子提取的速率，并且测得其比H-键解离速率高约5- 10倍。 最后，通过使用BIRD实验校准激光加热，我们能够建立货车't霍夫图，从中我们可以提取热力学参数。 我们在Journal of the American Society for Mass Spectrometry（2008，19，870-879）上发表了一篇关于动力学数据的论文（Lin等人，2008），并在修订中发表了第二篇关于热力学数据的论文。