Salivary Proteome Identification by MS

We identified salivary proteins using mass spectrometry-based proteomics. Examining 500 ng of peptides following tryptic digestion and peptide separation mass spectrometry data were obtained using a nanoElute LC system coupled to a timsTOF Pro mass spectrometer (Bruker Daltonics). In accordance with previous procedures [21 (link)], all MS/MS samples were examined using PEAKS Studio X+ (v.10.5, Bioinformatics Solutions). A protein sequence database of reviewed human proteins with 74,823 entries from UniProt was utilized for all searches, assuming the presence of the digestive enzyme trypsin. The InteractiVenn tool [22 (link)] was used to create a Venn diagram that displays the unique and shared proteins from each comparison. The Human Salivary Proteome Wiki (https://www.salivaryproteome.org/), a saliva proteome-focused, open-access database, was used to identify the origin of distinctive proteins [23 (link)]. Protein biological function was determined in FunRich software (v. 3.1.4) (accessed 21 June 2020) [24 (link)] using hypergeometric analysis where significance level was 0.01. In Funrich we add the list of all identified proteins with the “add dataset” function. In the “gene enrichment” option, we selected “compare” to identify the biological processes that presented the greatest differences between the groups. We chose the top 3 of those comparisons.