Mustor Emilee M, Wohlfahrt Jessica, Guergues Jennifer, Stevens Stanley M, Shaw Lindsey Neil
Department of Molecular Biosciences, University of South Florida, Tampa, FL, USA.
Center for Antimicrobial Resistance, University of South Florida, Tampa, FL, USA.
bioRxiv. 2024 Aug 7:2024.08.07.607079. doi: 10.1101/2024.08.07.607079.
is a major human pathogen causing myriad infections in both community and healthcare settings. Although well studied, a comprehensive exploration of its dynamic and adaptive proteome is still somewhat lacking. Herein, we employed streamlined liquid- and gas-phase fractionation with PASEF analysis on a TIMS-TOF instrument to expand coverage and explore the dark proteome. In so doing, we captured the most comprehensive proteome to date, totaling 2,231 proteins (85.6% coverage), using a significantly simplified process that demonstrated high reproducibility with minimal input material. We then showcase application of this library for differential expression profiling by investigating temporal dynamics of the proteome. This revealed alterations in metabolic processes, ATP production, RNA processing, and stress-response proteins as cultures progressed to stationary growth. Notably, a significant portion of the library (94%) and proteome (80.5%) was identified by this single-shot, DIA-based analysis. Overall, our study shines new light on the hidden proteome, generating a valuable new resource to facilitate further study of this dangerous pathogen.
是一种主要的人类病原体，在社区和医疗环境中都会引发无数感染。尽管已得到充分研究，但对其动态和适应性蛋白质组的全面探索仍有所欠缺。在此，我们在TIMS-TOF仪器上采用简化的液相和气相分级分离结合PASEF分析，以扩大覆盖范围并探索暗蛋白质组。通过这样做，我们使用一个显著简化的过程捕获了迄今为止最全面的蛋白质组，共2231种蛋白质（覆盖率85.6%），该过程显示出高重现性且所需输入材料极少。然后，我们通过研究蛋白质组的时间动态，展示了该文库在差异表达谱分析中的应用。这揭示了随着培养进入稳定期生长，代谢过程、ATP产生、RNA加工和应激反应蛋白的变化。值得注意的是，通过这种基于DIA的单次分析鉴定出了文库的很大一部分（94%）和蛋白质组的80.5%。总体而言，我们的研究为隐藏的蛋白质组带来了新的认识，生成了一个有价值的新资源，以促进对这种危险病原体的进一步研究。
