Abstract
Top-down mass spectrometry (MS)-based proteomics is arguably a disruptive technology for the comprehensive analysis of all proteoforms arising from genetic variation, alternative splicing, and posttranslational modifications (PTMs). However, the complexity of top-down high-resolution mass spectra presents a significant challenge for data analysis. In contrast to the well-developed software packages available for data analysis in bottom-up proteomics, the data analysis tools in top-down proteomics remain underdeveloped. Moreover, despite recent efforts to develop algorithms and tools for the deconvolution of top-down high-resolution mass spectra and the identification of proteins from complex mixtures, a multifunctional software platform, which allows for the identification, quantitation, and characterization of proteoforms with visual validation, is still lacking. Herein, we have developed MASH Suite Pro, a comprehensive software tool for top-down proteomics with multifaceted functionality. MASH Suite Pro is capable of processing high-resolution MS and tandem MS (MS/MS) data using two deconvolution algorithms to optimize protein identification results. In addition, MASH Suite Pro allows for the characterization of PTMs and sequence variations, as well as the relative quantitation of multiple proteoforms in different experimental conditions. The program also provides visualization components for validation and correction of the computational outputs. Furthermore, MASH Suite Pro facilitates data reporting and presentation via direct output of the graphics. Thus, MASH Suite Pro significantly simplifies and speeds up the interpretation of high-resolution top-down proteomics data by integrating tools for protein identification, quantitation, characterization, and visual validation into a customizable and user-friendly interface. We envision that MASH Suite Pro will play an integral role in advancing the burgeoning field of top-down proteomics.
Highlights
Lution top-down proteomics data by integrating tools for protein identification, quantitation, characterization, and visual validation into a customizable and user-friendly interface
UniDec, which employs a Bayesian approach to separate mass and charge dimensions [27], can be applied to the deconvolution of high-resolution spectra. These algorithms assist in data processing, the deconvolution results often contain a considerable amount of misassigned peaks as a consequence of the complexity of the high-resolution mass spectrometry (MS) and MS and tandem MS (MS/MS) data generated in top-down proteomics experiments
Protein identification using Mascot Top Down is fundamentally similar to that used in bottom-up proteomics [34], and, it is somewhat limited in terms of identifying unexpected posttranslational modifications (PTMs)
Summary
We report the development of MASH Suite Pro, an integrated software platform, designed to incorporate tools for protein identification, quantitation, and characterization into a single comprehensive package for the analysis of topdown proteomics data. This program contains a user-friendly customizable interface similar to the previously developed MASH Suite [36] and has a number of new capabilities, including the ability to handle complex proteomics datasets from LC-MS and LC-MS/MS experiments, as well as the ability to identify unknown proteins and PTMs using MSAlignϩ [32]. MASH Suite Pro provides visualization components for the validation and correction of the computational outputs, which ensures accurate and reliable deconvolution of the spectra and localization of PTMs and sequence variations
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