Computational protocol: The Histone Acetyltransferase Mst2 Protects Active Chromatin from Epigenetic Silencing by Acetylating the Ubiquitin Ligase Brl1

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Protocol publication

[…] Thermo RAW files were processed using Proteome Discoverer 2.1 software (Thermo Fisher), as described in the manufacturer’s instruction. Briefly, the Sequest search engine was used to search the MS2 spectra against the Schizosaccharomyces pombe UniProt database (downloaded on 30/01/2015) supplemented with common contaminating proteins. For total proteome analysis, cysteine carbamidomethylation and TMT tags on lysine and peptide N-termini were set as static modifications, whereas oxidation of methionine residues and acetylation protein N-termini were set as variable modifications. For acetyl-lysine enriched sample analysis, lysine acetylation and lysine TMT tags were set as variable modifications, while other modifications were set the same as for the proteome analysis. The assignments of the MS2 scans were filtered to allow 1% FDR. For reporter quantification, the S/N values were corrected for isotopic impurities of the TMT reagent using the values provided by the manufacturer. The sums across all TMT reporter channels were normalized assuming equal total protein content in each sample for proteome analysis, whereas for acetylome analysis normalization was based on total amount of acetylated peptides. All identified peptides from the proteome and acetylome experiments in this study are listed in . […]

Pipeline specifications

Software tools Proteome Discoverer, Comet
Application MS-based untargeted proteomics
Diseases Neoplasms