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bioRxiv · 10.1101/2024.11.08.622616

A Full Window Data Independent Acquisition Method for DeeperTop-down Proteomics

Sun, C.·Zhang, W.·Zhou, M.·Myu, M.·Xu, W.
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Abstract

Top-down proteomics (TDP) is emerging as a vital tool for the comprehensive characterization of proteoforms. However, as its core technology, top-down mass spectrometry (TDMS) still faces significant analytical challenges. While data-independent acquisition (DIA) has revolutionized bottom-up proteomics and metabolomics, they are rarely employed in TDP. The unique feature of protein ions in an electrospray mass spectrum, as well as the data complexity require the development of new DIA strategies. This study introduces a machine learning assisted Full Window DIA (FW-DIA) method that eliminates precursor ion isolation, making it compatible with a wide range of commercial mass spectrometers. Moreover, FW-DIA leverages all precursor protein ions to generate high-quality tandem mass spectra, enhancing signal intensities by [~]50-fold and protein sequence coverage by threefold in a modular protein analysis. The method was successfully applied to the analysis of a five-protein mixture under native conditions and Escherichia coli ribosomal proteoform characterization.

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Sun, C., Zhang, W., Zhou, M., Myu, M., Xu, W.. 2024-11-11. A Full Window Data Independent Acquisition Method for DeeperTop-down Proteomics. https://doi.org/10.1101/2024.11.08.622616

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