Biozentrum, University of Basel, Spitalstrasse 41, Basel CH-4056, Switzerland.
Anal Chem. 2024 Nov 12;96(45):18061-18069. doi: 10.1021/acs.analchem.4c03622. Epub 2024 Oct 28.
Bacterial infections are a major threat to human health worldwide. A better understanding of the properties and physiology of bacterial pathogens in human tissues is required to develop urgently needed novel control strategies. Mass spectrometry-based proteomics could yield such data, but identifying and quantifying scarce bacterial proteins against a preponderance of human proteins is challenging. Here, we explored the recently introduced SureQuant method for highly sensitive targeted mass spectrometry. Using a major human pathogen, the Gram-positive bacteria , as an example, we evaluated several parameters, including the number of targets and intensity thresholds, for optimal qualitative and quantitative protein analysis. By comparison, we found that SureQuant achieved the same quantitative performance as standard parallel reaction monitoring while allowing accurate and precise quantification of up to 400 targets. SureQuant also surpassed the sensitivity and quantification capabilities of global data-independent acquisition methods. Finally, to facilitate method development, we provide optimized MS parameters for the sensitive quantification of different peptide panel sizes. This study provides a foundation for the broader application of SureQuant in the analysis of clinical specimens containing trace amounts of bacterial proteins as well as other studies requiring ultrasensitive detection of low-abundant proteins.
细菌感染是全球范围内对人类健康的重大威胁。为了开发急需的新型控制策略，需要更好地了解人类组织中细菌病原体的特性和生理学。基于质谱的蛋白质组学可以提供此类数据，但鉴定和定量丰度与人蛋白相比处于劣势的稀有细菌蛋白具有挑战性。在这里，我们探索了最近引入的 SureQuant 方法，用于高度敏感的靶向质谱分析。使用主要的人类病原体革兰氏阳性菌作为示例，我们评估了多个参数，包括目标数量和强度阈值，以实现最佳的定性和定量蛋白质分析。相比之下，我们发现 SureQuant 实现了与标准平行反应监测相同的定量性能，同时允许对多达 400 个目标进行准确和精确的定量。SureQuant 还超越了全局数据非依赖性采集方法的灵敏度和定量能力。最后，为了便于方法开发，我们提供了优化的 MS 参数，用于对不同肽面板大小进行灵敏定量。本研究为 SureQuant 在分析含有痕量细菌蛋白的临床标本以及其他需要超灵敏检测低丰度蛋白质的研究中的更广泛应用奠定了基础。
