| Autor: |
Ivanov MV; V. L. Talrose Institute for Energy Problems of Chemical Physics, N. N. Semenov Federal Research Center for Chemical Physics, Russian Academy of Sciences, 119334 Moscow, Russia., Bubis JA; V. L. Talrose Institute for Energy Problems of Chemical Physics, N. N. Semenov Federal Research Center for Chemical Physics, Russian Academy of Sciences, 119334 Moscow, Russia., Gorshkov V; Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense M DK-5230, Denmark., Tarasova IA; V. L. Talrose Institute for Energy Problems of Chemical Physics, N. N. Semenov Federal Research Center for Chemical Physics, Russian Academy of Sciences, 119334 Moscow, Russia., Levitsky LI; V. L. Talrose Institute for Energy Problems of Chemical Physics, N. N. Semenov Federal Research Center for Chemical Physics, Russian Academy of Sciences, 119334 Moscow, Russia., Lobas AA; V. L. Talrose Institute for Energy Problems of Chemical Physics, N. N. Semenov Federal Research Center for Chemical Physics, Russian Academy of Sciences, 119334 Moscow, Russia., Solovyeva EM; V. L. Talrose Institute for Energy Problems of Chemical Physics, N. N. Semenov Federal Research Center for Chemical Physics, Russian Academy of Sciences, 119334 Moscow, Russia., Pridatchenko ML; V. L. Talrose Institute for Energy Problems of Chemical Physics, N. N. Semenov Federal Research Center for Chemical Physics, Russian Academy of Sciences, 119334 Moscow, Russia., Kjeldsen F; Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense M DK-5230, Denmark., Gorshkov MV; V. L. Talrose Institute for Energy Problems of Chemical Physics, N. N. Semenov Federal Research Center for Chemical Physics, Russian Academy of Sciences, 119334 Moscow, Russia.; Moscow Institute of Physics and Technology (State University), 141700 Dolgoprudny, Russia. |
| Abstrakt: |
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. |
