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DirectMS1: MS/MS-Free Identification of 1000 Proteins of Cellular Proteomes in 5 Minutes

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posted on 05.03.2020, 13:09 by Mark V. Ivanov, Julia A. Bubis, Vladimir Gorshkov, Irina A. Tarasova, Lev I. Levitsky, Anna A. Lobas, Elizaveta M. Solovyeva, Marina L. Pridatchenko, Frank Kjeldsen, Mikhail V. Gorshkov
Proteome characterization relies heavily on tandem mass spectrometry (MS/MS) and is thus associated with instrumentation complexity, lengthy analysis time, and limited duty cycle. It was always tempting to implement approaches that do not require MS/MS, yet they were constantly failing to achieve a meaningful depth of quantitative proteome coverage within short experimental times, which is particularly important for clinical or biomarker-discovery applications. Here, we report on the first successful attempt to develop a truly MS/MS-free method, DirectMS1, for bottom-up proteomics. The method is compared with the standard MS/MS-based data-dependent acquisition approach for proteome-wide analysis using 5 min LC gradients. Specifically, we demonstrate identification of 1 000 protein groups for a standard HeLa cell line digest. The amount of loaded sample was varied in a range from 1 to 500 ng, and the method demonstrated 10-fold higher sensitivity. Combined with the recently introduced Diffacto approach for relative protein quantification, DirectMS1 outperforms most popular MS/MS-based label-free quantitation approaches because of significantly higher protein sequence coverage.