Database, online items and scripts from ‘InvitroSPI and a large database of proteasome-generated spliced and non-spliced peptides’

Manuscript abstract Noncanonical epitopes presented by Human Leucocyte Antigen class I (HLA-I) complexes to CD8+ T cells attracted the spotlight in the research of novel immunotherapies against cancer, infection and autoimmunity. Proteasomes, which are the main producers of HLA-I-bound antigenic peptides, can catalyze both peptide hydrolysis and peptide splicing. The prediction of proteasome-generated spliced peptides is an objective that still requires a reliable (and large) database of non-spliced and spliced peptides produced by these proteases. Here, we present an extended database of proteasome-generated spliced and non-spliced peptides, which was obtained by analyzing <em>in vitro</em> digestions of 80 unique synthetic polypeptide substrates, measured by different mass spectrometers. Peptides were identified through invitroSPI method, which was validated through <em>in silico</em> and <em>in vitro</em> strategies. The peptide product database contains 16,631 unique peptide products (5,493 non-spliced, 6,453 <em>cis</em>-spliced and 4,685 <em>trans</em>-spliced peptide products), and a substrate sequence variety that is a valuable source for predictors of proteasome-catalyzed peptide hydrolysis and splicing. Potential artefacts and skewed results due to different identification and analysis strategies are discussed. <br> File description <br> PBdataset-search-results.zip Mascot search results for the PB dataset, used for invitroSPI analysis of PB dataset. Contains a full list of all PSMs suggested by Mascot. <br> ProteasomeDB.csv Large database of proteasome-generated spliced and non-spliced peptides, generated by invitroSPI analysis of 80 synthetic polypeptide substrates. <br> databases.zip Custom databases used for search of in vitro digestion MS measurements. They contain all theoretically possible spliced and non-spliced peptides that could be derived from a given polypeptide. <br> analysis-scripts-figshare.zip Scripts used for the analysis presented in the corresponding manuscript. <br> invitroPB_scripts.zip Scripts for the invitroPB method. <br> Online-Table 1 Technical details of all 80 synthetic substrates and their measurement. <br> Online-Table 2 PSMs identified in the gp100 Fusion dataset <br> Online-Table 3 PSMs identified in the PB dataset <br> Online-Figure 1 Comparison of measured MS2 spectra of a representative pool of identified non-spliced and spliced peptide products and synthetic peptides derived from the gp100 dataset <br> Online-Figure 2 Comparison of measured and predicted MS2 spectra of non-spliced and spliced peptides identified by invitroSPI method in the PB dataset <br> Online-Figure 3 Comparison of measured and predicted MS2 spectra of non-spliced and spliced peptides identified by invitroPB method in the PB dataset