PTM-Shepherd: Analysis and Summarization of Post-Translational and Chemical Modifications From Open Search Results.

Open searching has proven to be an effective strategy for identifying both known and unknown modifications in shotgun proteomics experiments. Rather than being limited to a small set of user-specified modifications, open searches identify peptides with any mass shift that may correspond to a single modification or a combination of several modifications. Here we present PTM-Shepherd, a bioinformatics tool that automates characterization of post-translational modification profiles detected in open searches based on attributes, such as amino acid localization, fragmentation spectra similarity, retention time shifts, and relative modification rates.

Identification of modified peptides using localization-aware open search.

Identification of post-translationally or chemically modified peptides in mass spectrometry-based proteomics experiments is a crucial yet challenging task. We have recently introduced a fragment ion indexing method and the MSFragger search engine to empower an open search strategy for comprehensive analysis of modified peptides. However, this strategy does not consider fragment ions shifted by unknown modifications, preventing modification localization and limiting the sensitivity of the search.

Fast Quantitative Analysis of timsTOF PASEF Data with MSFragger and IonQuant.

Ion mobility brings an additional dimension of separation to LC-MS, improving identification of peptides and proteins in complex mixtures. A recently introduced timsTOF mass spectrometer (Bruker) couples trapped ion mobility separation to TOF mass analysis. With the parallel accumulation serial fragmentation (PASEF) method, the timsTOF platform achieves promising results, yet analysis of the data generated on this platform represents a major bottleneck.

Crystal-C: A Computational Tool for Refinement of Open Search Results.

Shotgun proteomics using liquid chromatography coupled to mass spectrometry (LC-MS) is commonly used to identify peptides containing post-translational modifications. With the emergence of fast database search tools such as MSFragger, the approach of enlarging precursor mass tolerances during the search (termed "open search") has been increasingly used for comprehensive characterization of post-translational and chemical modifications of protein samples. However, not all mass shifts detected using the open search strategy represent true modifications, as artifacts exist from sources such as unaccounted missed cleavages or peptide co-fragmentation (chimeric MS/MS spectra).

Integrated Proteogenomic Characterization of Clear Cell Renal Cell Carcinoma.

To elucidate the deregulated functional modules that drive clear cell renal cell carcinoma (ccRCC), we performed comprehensive genomic, epigenomic, transcriptomic, proteomic, and phosphoproteomic characterization of treatment-naive ccRCC and paired normal adjacent tissue samples. Genomic analyses identified a distinct molecular subgroup associated with genomic instability. Integration of proteogenomic measurements uniquely identified protein dysregulation of cellular mechanisms impacted by genomic alterations, including oxidative phosphorylation-related metabolism, protein translation processes, and phospho-signaling modules.

DeltaMass: Automated Detection and Visualization of Mass Shifts in Proteomic Open-Search Results.

Routine identification of thousands of proteins in a single LC-MS experiment has long become the norm. With these vast amounts of data, more rigorous treatment of modified forms of peptides becomes possible. "Open search", a protein database search with a large precursor ion mass tolerance window, is becoming a popular method to evaluate possible sets of post-translational and chemical modifications in samples.

IMTBX and Grppr: Software for Top-Down Proteomics Utilizing Ion Mobility-Mass Spectrometry.

Top-down proteomics has emerged as a transformative method for the analysis of protein sequence and post-translational modifications (PTMs). Top-down experiments have historically been performed primarily on ultrahigh resolution mass spectrometers due to the complexity of spectra resulting from fragmentation of intact proteins, but recent advances in coupling ion mobility separations to faster, lower resolution mass analyzers now offer a viable alternative. However, software capable of interpreting the highly complex two-dimensional spectra that result from coupling ion mobility separation to top-down experiments is currently lacking.

MSFragger: ultrafast and comprehensive peptide identification in mass spectrometry-based proteomics.

There is a need to better understand and handle the 'dark matter' of proteomics-the vast diversity of post-translational and chemical modifications that are unaccounted in a typical mass spectrometry-based analysis and thus remain unidentified. We present a fragment-ion indexing method, and its implementation in peptide identification tool MSFragger, that enables a more than 100-fold improvement in speed over most existing proteome database search tools. Using several large proteomic data sets, we demonstrate how MSFragger empowers the open database search concept for comprehensive identification of peptides and all their modified forms, uncovering dramatic differences in modification rates across experimental samples and conditions.

BatMass: a Java Software Platform for LC-MS Data Visualization in Proteomics and Metabolomics.

Mass spectrometry (MS) coupled to liquid chromatography (LC) is a commonly used technique in metabolomic and proteomic research. As the size and complexity of LC-MS-based experiments grow, it becomes increasingly more difficult to perform quality control of both raw data and processing results. In a practical setting, quality control steps for raw LC-MS data are often overlooked, and assessment of an experiment's success is based on some derived metrics such as "the number of identified compounds".