Gas-phase Bio-conjugation in the Tandem Mass Spectrometry of Peptides, Proteins,

描述（由申请人提供）：本研究计划的总体目标是开发新的和改进的多肽离子结构表征方法，包括由酶或化学消化、全蛋白和蛋白质复合物形成的肽。近期目标是开发串联质谱的新方向，这将使处理蛋白质鉴定/表征问题的广泛新策略成为可能。绝大多数串联质谱实验依赖于裂解反应作为关键的结构信息反应。在这项工作中，我们试图在MSn实验的背景下增加气态离子选择性共价或非共价衍生化的能力。特定气相化学的发展从根本上促进了广泛应用的发展。正如溶液中的生物偶联化学已被分子生物学研究界广泛应用于各种测量策略中一样，影响气相生物偶联的能力将使许多结构表征的新方法成为可能。气相法有趣的特点之一是速度快（即几十毫秒的时间）

DESCRIPTION (provided by applicant): The overall thrust of this research initiative is to develop new and improved means for the structural characterization of polypeptide ions, including those formed from peptides derived from enzymatic or chemical digestion, whole proteins, and protein complexes. The near-term objective is to develop a new direction in tandem mass spectrometry that will enable a wide range of novel strategies to deal with protein identification/characterization problems. The vast majority of tandem mass spectrometry experiments rely on fragmentation as the key structurally informative reaction. In this work, we seek to add the capability for selective covalent or non-covalent derivatization of gaseous ions within the context of an MSn experiment. The development of specific gas-phase chemistries fundamentally enables the development of a very wide range of applications. Just as bio-conjugation chemistries in solution have become broadly used by the molecular biology research community in a wide range of measurement strategies, the ability to effect gas-phase bio- conjugation will enable many novel approaches to structural characterization. Among the interesting features of the gas-phase approach are that it is fast (i.e., tens of milliseconds time scale) and it completely avoids sample manipulation issues. The work will involve the development of novel selective reactions, many of which will be inspired by the rich bio-conjugation literature, the development of peptide ion-based applications, and the development of whole protein/protein complex ion-based applications, as reflected in the specific aims: Specific Aim 1: Discover/develop functional group specific ion/ion reactions for reactive amino acid side chains, including, but not limited to, lysine (and N-termini), residues with acidic side-chains (i.e., glutamic and aspartic acids) and C-termini, arginine, and cysteine, as well as common post-translational modifications, such as phosphate and sulfate. Specific Aim 2: Develop applications for the use of selective ion/ion reactions in the MSn of peptide ions in support of peptide-based (i.e., bottom-up) approaches in protein identification/characterization. Specific Aim 3: Develop applications for the use of selective ion/ion reactions in the MSn of intact protein and protein complex ions in support of protein-based (i.e., top-down) approaches in protein identification/characterization. The development of novel chemistries and methodologies based on selective gas-phase reactions will be relevant to many areas of bio-medical research by virtue of the role that tandem mass spectrometry already plays across the spectrum of health related science. The work in this proposal must necessarily be focused, and will be directed to protein analysis. However, functional group-specific gas-phase reactions can also lead to new applications in other areas, such as metabolomics, glycomics, lipidomics, etc. The potential bio- medical impact for this work, therefore, is unusually broad.