TCP3: QUALITATIVE & QUANTITATIVE PROTEOMIC ANALYSIS OF LYSINE MODIFICATIONS

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。 技术核心项目3：蛋白质修饰的定量质谱学和检测复杂混合物中的蛋白质修饰-AkHilesh Pandey，系助理教授。生物化学和免疫球蛋白。其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，他的方法可以通过预分馏(泥坑)方法常规地应用于蛋白质混合物。潘迪博士还领导了蛋白质生物信息学方面的一项重大工作。 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。 1.建立细胞培养中稳定同位素标记(SILAC)方法，一次从多个样品(最多四种不同状态)获得蛋白质和多肽水平的定量信息。这将允许同时相对定量i)所有4个状态的蛋白质丰度，以及ii)所有4个状态的翻译后修饰。 2.建立改进乙酰化、甲基化和泛素化多肽检测的质谱学方法。我们将开发前体离子扫描和中性损失扫描方法，从复杂混合物中选择性地识别携带乙酰基、甲基或泛素部分的多肽。 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储存库。以标准化格式生成涉及赖氨酸修饰的途径，并以XML文件格式生成蛋白质序列上的PTM映射，以便能够使用质谱学更好地搜索蛋白质数据库-派生

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