FAST MS: Software for the Automated Analysis of Top-Down Mass Spectra of Polymeric Molecules Including RNA, DNA, and Proteins.

Abstract

Top-down mass spectrometry (MS) enables comprehensive characterization of modified proteins and nucleic acids and, when native electrospray ionization (ESI) is used, binding site mapping of their complexes with native or therapeutic ligands. However, the high complexity of top-down MS spectra poses a serious challenge to both manual and automated data interpretation, even when the protein, RNA, or DNA sequence and the type of modification or the ligand are known. Here, we introduce FAST MS, a user-friendly software that identifies, assigns and relatively quantifies signals of molecular and fragment ions in MS and MS/MS spectra of biopolymers with known sequence and provides a toolbox for statistical analysis. FAST MS searches mass spectra for ion signals by comparing all signals in the spectrum with isotopic profiles calculated from known sequences, resulting in superior sensitivity and an increased number of assigned fragment ions compared to algorithms that rely on artificial monomer units while maintaining the false positive rate on a moderate level (<5%). FAST MS is an open-source, cross-platform software for the accurate identification, localization and relative quantification of modifications, even in complex mixtures of positional isomers of proteins, oligonucleotides, or any other user-defined linear polymer.