负离子模式蛋白质组学的潜力:一种仅使用MS1的方法
Pelayo A Penanes1, Vladimir Gorshkov1, Mark V Ivanov2
1Department of Biochemistry and Molecular Biology, University of Southern Denmark, DK-5230 Odense M, Denmark.
Journal of proteome research
|July 3, 2023
概括
这项研究证明了使用负电离模式与DirectMS1方法的高效蛋白质识别,在10分钟内识别了1000多种蛋白质. 结合正和负离子模式,进一步提高了蛋白质识别产量.
科学领域:
- 蛋白质组学是指蛋白质组学.
- 质谱测量质量谱测量
- 分析化学 分析化学
背景情况:
- 目前的蛋白质组学主要使用正离子化,这对于酸性来说是低效的.
- 负电离模式为更广泛的检测提供了潜力,但尚未得到充分探索.
研究的目的:
- 使用DirectMS1方法研究和优化负电离模式下蛋白质识别效率.
- 评估负电离数据与正电离数据的互补价值.
- 评估不同蛋白酶在负离子模式下对蛋白质识别的影响.
主要方法:
- 使用了基于精确的质量和预测的保留时间的DirectMS1超快数据采集方法.
- 优化的分离和实验条件使用移动缓冲器与伊米达和异醇.
- 使用各种蛋白酶 (LysC,GluC,AspN,素) 进行蛋白质消化分析.
主要成果:
- 迄今为止,在负离子模式下实现了最高的蛋白质识别率,在1%的FDR中识别了人类细胞系中的1000多种蛋白质,在10分钟的梯度内.
- 结合正和负离子模式数据,已识别的蛋白质增加到1774.
- 素和LysC蛋白酶产生了最高的蛋白质识别率,与标准的正态蛋白质组消化相兼容.
结论:
- 与负电离相结合的DirectMS1方法显著提高了蛋白质识别效率.
- 负电离模式提供了补充蛋白质组数据,增加了总体覆盖范围.
- 在正态蛋白质组学中使用的标准蛋白酶对于负态分析是有效的,简化了工作流程.
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