Browsing by Subject "top-down proteomics"

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    Large-Scale Qualitative and Quantitative Top-Down Proteomics Using Capillary Zone Electrophoresis-Electrospray Ionization-Tandem Mass Spectrometry with Nanograms of Proteome Samples
    (Springer, 2019-04-09) Lubeckyj, Rachele A.; Basharat, Abdul Rehman; Shen, Xiaojing; Liu, Xiaowen; Sun, Liangliang; BioHealth Informatics, School of Informatics and Computing
    Capillary zone electrophoresis-electrospray ionization-tandem mass spectrometry (CZE-ESI-MS/MS) has attracted attention recently for top-down proteomics because it can achieve highly efficient separation and very sensitive detection of proteins. However, separation window and sample loading volume of CZE need to be boosted for a better proteome coverage using CZE-MS/MS. Here, we present an improved CZE-MS/MS system that achieved a 180-min separation window and a 2-μL sample loading volume for top-down characterization of protein mixtures. The system obtained highly efficient separation of proteins with nearly one million theoretical plates for myoglobin and enabled baseline separation of three different proteoforms of myoglobin. The CZE-MS/MS system identified 797±21 proteoforms and 258±7 proteins (n=2) from an Escherichia coli (E. coli) proteome sample in a single run with only 250 ng of proteins injected. The system still identified 449±40 proteoforms and 173±6 proteins (n=2) from the E. coli sample when only 25 ng of proteins were injected per run. Single-shot CZE-MS/MS analyses of zebrafish brain cerebellum (Cb) and optic tectum (Teo) regions identified 1 730±196 proteoforms (n=3) and 2 024±255 proteoforms (n=3), respectively, with only 500-ng proteins loaded per run. Label-free quantitative top-down proteomics of zebrafish brain Cb and Teo regions revealed significant differences between Cb and Teo regarding the proteoform abundance. Over 700 proteoforms from 131 proteins had significantly higher abundance in Cb compared to Teo, and these proteins were highly enriched in several biological processes, including muscle contraction, glycolytic process, and mesenchyme migration.
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    MASH Explorer: A Universal Software Environment for Top-Down Proteomics
    (ACS, 2020-08) Wu, Zhijie; Roberts, David S.; Melby, Jake A.; Wenger, Kent; Wetzel, Molly; Gu, Yiwen; Ramanathan, Sudharshanan Govindaraj; Bayne, Elizabeth F.; Liu, Xiaowen; Sun, Ruixiang; Ong, Irene M.; McIlwain, Sean J.; Ge, Ying; BioHealth Informatics, School of Informatics and Computing
    Top-down mass spectrometry (MS)-based proteomics enable a comprehensive analysis of proteoforms with molecular specificity to achieve a proteome-wide understanding of protein functions. However, the lack of a universal software for top-down proteomics is becoming increasingly recognized as a major barrier, especially for newcomers. Here, we have developed MASH Explorer, a universal, comprehensive, and user-friendly software environment for top-down proteomics. MASH Explorer integrates multiple spectral deconvolution and database search algorithms into a single, universal platform which can process top-down proteomics data from various vendor formats, for the first time. It addresses the urgent need in the rapidly growing top-down proteomics community and is freely available to all users worldwide. With the critical need and tremendous support from the community, we envision that this MASH Explorer software package will play an integral role in advancing top-down proteomics to realize its full potential for biomedical research.