Top-Down Deep Sequencing of Ubiquitin Using Two-Dimensional Mass Spectrometry. Analytical Chemistry, 90(12):7302-7309, 2018. cited By 4
Top-Down Deep Sequencing of Ubiquitin Using Two-Dimensional Mass Spectrometry [link]Paper  doi  abstract   bibtex   
Two-dimensional mass spectrometry (2DMS) allows data independent fragmentation of all ions in a sample and correlation of fragment ions to their precursors without isolation prior to fragmentation. Developments in computer capabilities and implementations in Fourier transform ion cyclotron resonance (FTICR) MS over the past decade have allowed the technique to become a useful analytical tool for bottom-up proteomics (BUP) and, more recently, in top-down protein analysis (TDP). In this work, a new method of TDP is developed using 2D FTICR MS, called MS/2D FTICR MS or MS/2DMS. In MS/2DMS, an entire protein is initially fragmented in a hexapole collision cell, e.g., with collisionally activated dissociation (CAD). The primary fragments are then sent to the ICR cell, where 2DMS is performed with infrared multiphoton dissociation (IRMPD) or electron-capture dissociation (ECD). The resulting 2D mass spectra retain information equivalent to a set of TDP MS 3 experiments on the selected protein. Up to n - 1 fragmentation steps can be added to the process, as long as an ion of interest can be unambiguously fragmented before the ICR cell, leading to an MS n /2DMS experiment whose output is a 2D mass spectrum retaining information equivalent to MS n . MS/2DMS and MS/MS/2DMS are used in this work for the structural analysis of ubiquitin (Ubi), noting several unique features which aid fragment identification. The use of CAD-MS/IRMPD-2DMS, CAD-MS/ECD-2DMS, and MS 2 /2DMS using, respectively, in-source dissociation (ISD), CAD, and ECD-2DMS led to 97% cleavage coverage for Ubi. © 2018 American Chemical Society.

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