PROTEIN IDENTIFICATION CORE

在生物体内表达的细胞内信号分子的全部成分 调查是一个极其复杂的分析挑战，有大量的 组件在大小和浓度上变化很大，并以高度 这些生物的中枢神经系统内的异质方式。应对这些挑战，并 为我们的用户提供一系列最先进的蛋白质识别技术，这个核心 提出了分子检测技术在三个方面的应用和发展 不同的区域。我们将(1)通过各种“自下而上”的方法识别信号分子 包括质量指纹图谱、自动化MS/MS测序和准确的质量测量，(2)通过完整的蛋白质分析来确定识别的细胞间信号分子的特征，以确定细胞间信号分子中常见的翻译后修饰，并纠正由于信号肽切割等原因而可能导致的基因组数据库中的错误，以及(3)通过各种MS成像技术定位细胞间信号分子，包括目前最先进的MS成像技术和建议的“拉伸样本”和“毛细管阵列”方法，这些方法有可能将成像方法的分辨率和灵敏度提高到用户所需的水平。我们用户对细胞间信号分子鉴定的需求在样本大小和复杂性上与单细胞生物的“鸟枪式”蛋白质组学有很大不同。有了与拟建中心相关的小组中的综合专业知识和设施，就有了在来源上识别和表征细胞间信号分子的独特机会。

The entire constituent of intracellular signaling molecules expressed in the organisms under investigation represents an extremely complex analytical challenge with a vast number of components varying dramatically in size and concentration and expressed in a highly heterogeneous manner within the CNS of these organisms. To address these challenges, and to provide our users with a battery of state-of-the-art protein identification techniques, this core proposes the use and development of techniques capable of molecular interrogation in three separate areas. We will (1) identify signaling molecules by a variety of "Bottom Up" approaches including mass fingerprinting, automated MS/MS sequencing, and accurate mass measurement, (2) characterize identified intercellular signaling molecules by intact protein analysis to identify post-translational modifications that are common among intercellular signaling molecules and correct for possible errors in the genomic databases due to signal peptide cleavage etc., and (3) localize intercellular signaling molecules by a variety of MS imaging techniques including current state-of-the-art and development of proposed "stretched sample" and "capillary array" methods that have the potential to enhance resolution and sensitivity of imaging methods to a level needed by our users. The needs for intercellular signaling molecule identification of our users are quite different from "shotgun" proteomics of unicellular organisms in sample size and complexity. With the combined expertise and facilities in the groups associated with the proposed center, a unique opportunity exists to identify and characterize intercellular signaling molecules at their sources.