TCP3: QUALITATIVE & QUANTITATIVE PROTEOMIC ANALYSIS OF LYSINE MODIFICATIONS

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。技术核心项目3：蛋白质修饰的定量质谱法和复杂混合物中蛋白质修饰的检测- Akhilesh Pandey，生物化学和IGM系助理教授。目的是开发定量方法来定位/检测蛋白质中的乙酰化、甲基化、泛素化和sumo化位点。此外，他的方法可以常规应用于蛋白质混合物的预分馏（MUDPIT）方法。潘迪博士还在蛋白质生物信息学方面做出了重大努力。这涉及到与印度班加罗尔生物信息学研究所的一项关键的数据库工作。建立细胞培养稳定同位素标记（SILAC）方法，在一次实验中从多个样品（多达四种不同状态）中获得蛋白质和肽水平的定量信息。这将允许同时相对定量i)所有4种状态下的蛋白质丰度，以及ii)所有4种状态下的翻译后修饰。2. 发展质谱方法以改进乙酰化、甲基化和泛素化肽的检测。我们将开发前体离子扫描和中性损失扫描方法，以选择性地从复杂混合物中识别携带乙酰基、甲基或泛素部分的肽。3. 开发用于定量分析的软件工具，并创建酵母和人类所有赖氨酸修饰的存储库。以标准化格式生成涉及赖氨酸修饰的通路，并以XML文件格式生成ptm在蛋白质序列上的映射，以便使用质谱-推导技术更好地搜索蛋白质数据库

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. Technology Core Projects 3: Quantitative mass spectrometry of protein modifications and detecting protein modifications in complex mixtures - Akhilesh Pandey, Assistant Professor, Dept. of Biological Chemistry and IGM. The aim is to develop quantitative methods for mapping/detecting sites of acetylation, methylation, ubiquitylation and SUMOylation in proteins. In addition, his methods can be routinely applied to protein mixtures by prefractionation (MUDPIT) methodologies. Dr. Pandey also heads a major effort in protein bioinformatics. This involves a critical databasing effort with the Institute of Bioinformatics in Bangalore, India. 1. To develop stable isotope labeling in cell culture (SILAC) method to obtain quantitative information at the protein and peptide level from multiple samples (up to four different states) in a single experiment. This will allow for simultaneous relative quantitation of i) protein abundance in all 4 states, and, ii) post-translational modifications in all 4 states. 2. To develop mass spectrometric methods for improved detection of acetylated, methylated and ubiquitylated peptides. We will develop precursor ion scanning as well as neutral loss scanning methods to selectively identify peptides carrying an acetyl, methyl or ubiquitin moieties from complex mixtures. 3. To develop software tools for analysis of quantitation and to create a repository of all lysine modifications in yeast and humans. To generate pathways involving lysine modifications in a standardized format and to generate mapping of PTMs on the protein sequences in XML file format to enable better searching of protein databases using mass spectrometry-derive