BET CELLS & MASS SPECTROMETERS

赌注细胞

• 批准号: 6486810
• 负责人: FF HSU
• 金额: $ 15.75万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-08-01 至 2002-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6486810
• 关键词: biomedical resource; genetics; hematology; human subject; male; proteins; spectrometry

Both types I and II diabetes mellitus result from abnormal function or loss of insulin-secreting beta cells in pancreatic islets. Beta cell membranes govern ionic conductances, exocytosis, and antigenic properties, and complex lipid components affect membrane functions. Characterization of islet lipids is difficult because islets are obtainable in only small quantities. Conventional methods of complex lipid analysis involve multiple chromatographic, digestion, and derivatization steps that each involve analyte losses, and FAB/MS is insufficiently sensitive to permit analysis of complex lipids in islet subcellular membranes. The great sensitivity of ESI/MS combined with structural information provided by collisionally activated dissociation and tandem mass spectrometry circumvents these difficulties. We have developed ESI/MS/MS methods that facilitate identification of small quantities of glycerophosphocholine lipids and of sulfatide species. ESI/MS has been used to identify a ll major phospholipid species in islet secretory granules and plasma membranes, to identify islet sulfatide species, and to examine stimulus-induced changes in amounts of islet phospholipid species. Characterization of islet lipids has led to the discovery of a new lipid metabolizing enzyme, which is now designated iPLA2. We have overexpressed this enzyme in Sf9 cells and are characterizing its post-translational modifications by HPLC/ESI/MS/MS analyses of tryptic digetsts of the recombinant protein.

I型和II型糖尿病都是由异常 胰岛中分泌胰岛素的β细胞的功能或丧失。 β细胞膜支配离子电导、胞吐作用和 抗原性，和复杂的脂类成分影响膜 功能。胰岛脂类的特征很难确定，因为 只有少量的小岛可以买到。常规方法 复杂的脂肪分析涉及多层析，消化， 和衍生化步骤，每个步骤都涉及分析物损失和FAB/MS 不够灵敏，不能分析复杂的脂类 胰岛亚细胞膜。电喷雾质谱联用的高灵敏度 通过碰撞激活提供的结构信息 解离和串联质谱学绕过了这些 困难。我们已经开发了ESI/MS/MS方法，以促进 微量甘油磷胆碱类脂的鉴定和鉴定 硫脂的种类。ESI/MS已被用来识别L1大调 胰岛分泌颗粒和质膜中的磷脂类物质， 鉴定胰岛硫脂的种类，并检测刺激诱导 胰岛磷脂种类的数量变化。特征描述 胰岛脂类导致了一种新的脂类代谢的发现 酶，现在命名为iPLA2。我们已经过度表达了这一点 Sf9细胞中的酶及其翻译后的特征 高效液相色谱/电喷雾质谱/电喷雾串联质谱法分析胰酶脱脂产物的研究 重组蛋白。