MASS SPECTRAL STUDIES OF DEPROTONATED PEPTIDES

本研究的目的是开发分析程序 酸性肽的负离子质谱分析。许多生物 肽和用于疾病治疗的肽具有许多谷氨酸 或天冬氨酸残基;其他含有酸性磷酸盐或硫酸盐 组 在过去的15年里，质谱法的进步 使其成为测定分子量的首选方法， 肽的序列。数百份报告已经出现在串联 带正电荷的质子化肽的质谱（MS/MS） 离子。 相比之下，相对较少的工作集中在消极的- 带电荷的去质子化离子，这些研究中的大多数都涉及 其中单电荷离子含有四个或更少的残基。然而，在这方面， 酸性肽通常比正离子更容易形成负离子 离子。 该项目将大量使用两种先进的方法 光谱法：电喷雾电离傅立叶变换离子回旋加速器 共振（ESI/FT-ICR）和基质辅助激光解吸电离 飞行时间（MALDI/TOF）。 ESI/FT-ICR研究将涉及低能 多重去质子化离子的碰撞诱导解离（CID）， 而MALDI/TOF工作将采用源后衰减（PSD）对单个 去质子化离子 主要目的是阐明解离 机制等各种氨基酸诱导的片段化 残基和它们的序列将被探索。 结构特征 包括谷氨酸和天冬氨酸残基，非 酸性残基，如脯氨酸和赖氨酸，C-末端端基（-OH 或NH 2）、高度酸性的磷酸盐和硫酸盐基团以及相互作用 在酸性和碱性残基之间。 对于多重去质子化离子 由ESI产生的离子电荷的数量和位置（因此 库仑排斥的大小）必须被考虑。 分子 将进行动力学计算，以深入了解库仑 能量和构象，这可能会影响解离。 的影响 将通过改变母体来评估碎片的内能 在FT-ICR和TOF中，通过增加PSD 在离子离开源之后发生碰撞。 负离子 解离将与相应的阳性对照进行比较。 离子。此外，影响负离子的结构因素 将对生产进行调查。 为了提高去质子化离子产率， MALDI，将为肽开发基本基质。

The objective of this research is to develop procedures for the analysis of acidic peptides by negative ion mass spectrometry. Many biological peptides and peptides used in disease treatment have numerous glutamic or aspartic acid residues; others contain acidic phosphate or sulfate groups. In the past fifteen years, advances in mass spectrometry have made it a method of choice for determining the molecular masses and sequences of peptides. Hundreds of reports have appeared on the tandem mass spectrometry (MS/MS) of positively-charged, protonated peptide ions. In contrast, relatively little work has focused on negatively- charged, deprotonated ions and the majority of these studies have dealt with singly charged ions containing four or fewer residues. However, acidic peptides often form negative ions more readily than positive ions. This project will utilize two advancing methodologies in mass spectrometry: electrospray ionization Fourier transform ion cyclotron resonance (ESI/FT-ICR) and matrix-assisted laser desorption ionization time-of-flight (MALDI/TOF). ESI/FT-ICR studies will involve low-energy collision-induced dissociation (CID) on multiply deprotonated ions, while MALDI/TOF work will employ post-source decay (PSD) on singly deprotonated ions. The major aim is to elucidate dissociation mechanisms. Fragmentations induced by various types of amino acid residues and by their sequences will be explored. Structural features under investigation include glutamic and aspartic acid residue, non- acidic residues such a proline and lysine, the C-terminal endgroup (-OH or NH2), highly acidic phosphate and sulfate groups, and interactions between acidic and basic residues. For multiply deprotonated ions generated by ESI, the number and locations of ionic charges (and thus the magnitude of Coulomb repulsion) must be considered. Molecular dynamics calculations will be performed to gain insight into Coulomb energy and conformations, which can affect dissociation. The impact of internal energy on fragmentation will be assessed by varying the parent ion excitation energy in the FT-ICR and, in the TOF, by augmenting PSD with collisions occurring after ions leave the source. Negative ion dissociation will be compared to that of the corresponding positive ions. In addition, structural factors that influence negative ion production will be investigated. To enhance deprotonated ion yields by MALDI, basic matrices will be developed for peptides.